Food Aroma Evolution

During Food Processing,

Cooking, and Aging
 Food Analysis & Properties
Series Editor
Leo M.L. Nollet
University College Ghent, Belgium
This CRC series Food Analysis and Properties is designed to provide state-of-
the-art coverage on topics related to the understanding of the physical, chemi-
cal, and functional properties of food: including (1) recent analysis techniques
on the choice of food components; (2) developments and evolutions in analysis
techniques related to food; (3) recent trends in analysis techniques of specific
food components and/or a group of related food components.
Flow Injection Analysis of Food Additives
Edited by Claudia Ruiz-Capillas and Leo M.L. Nollet

Marine Microorganisms
Extraction and Analysis of Bioactive Compounds
Edited by Leo M.L. Nollet

Multiresidue Methods for the Analysis of Pesticide Residues in Food

Edited by Horacio Heinzen, Leo M.L. Nollet, and Amadeo R. Fernandez-Alba

Spectroscopic Methods in Food Analysis

Edited by Adriana S. Franca and Leo M.L. Nollet

Phenolic Compounds in Food

Characterization and Analysis
Edited by Leo M.L. Nollet and Janet Alejandra Gutierrez-Uribe

Testing and Analysis of GMO-Containing Foods and Feed

Edited by Salah E.O. Mahgoub and Leo M.L. Nollet

Fingerprinting Techniques in Food Authenticity and Traceability

Edited by K.S. Siddiqi and Leo M.L. Nollet

Hyperspectral Imaging Analysis and Applications for Food Quality

Edited by Nrusingha Charan Basantia, Leo M.L. Nollet, and Mohammed Kamruzzaman

Ambient Mass Spectroscopy Techniques in Food and the Environment

Edited by Leo M.L. Nollet and Basil K. Munjanja

Food Aroma Evolution: During Food Processing, Cooking and Aging

Edited by Matteo Bordiga and Leo M.L. Nollet
For more information, please visit the Series Page: https​://ww​w.crc​press​.com/​Food-​A naly​sis-​
Prope​rties​/ book​-seri​es/CR​C FOOD​A NPRO​
Food Aroma Evolution
During Food Processing,
Cooking, and Aging

Edited by
Matteo Bordiga
Dipartimento Di Scienze Del Farmaco
Leo M.L. Nollet
University College Ghent (Retired)
CRC Press
Taylor & Francis Group
6000 Broken Sound Parkway NW, Suite 300
Boca Raton, FL 33487-2742

© 2020 by Taylor & Francis Group, LLC

CRC Press is an imprint of Taylor & Francis Group, an Informa business

No claim to original U.S. Government works

International Standard Book Number-13: 978-1-138-33824-1 (Hardback)

This book contains information obtained from authentic and highly regarded sources. Reasonable efforts have been made to
publish reliable data and information, but the author and publisher cannot assume responsibility for the validity of all materi-
als or the consequences of their use. The authors and publishers have attempted to trace the copyright holders of all material
reproduced in this publication and apologize to copyright holders if permission to publish in this form has not been obtained.
If any copyright material has not been acknowledged please write and let us know so we may rectify in any future reprint.
Except as permitted under U.S. Copyright Law, no part of this book may be reprinted, reproduced, transmitted, or utilized in
any form by any electronic, mechanical, or other means, now known or hereafter invented, including photocopying, microfilm-
ing, and recording, or in any information storage or retrieval system, without written permission from the publishers.

For permission to photocopy or use material electronically from this work, please access www.copyright.com (http://www.
copyright.com/) or contact the Copyright Clearance Center, Inc. (CCC), 222 Rosewood Drive, Danvers, MA 01923, 978-750-
8400. CCC is a not-for-profit organization that provides licenses and registration for a variety of users. For organizations that
have been granted a photocopy license by the CCC, a separate system of payment has been arranged.

Trademark Notice: Product or corporate names may be trademarks or registered trademarks, and are used only for identifica-
tion and explanation without intent to infringe.

Library of Congress Cataloging‑in‑Publication Data

Names: Bordiga, Matteo, editor. | Nollet, Leo M.L., 1948- editor.

Title: Food aroma evolution : during food processing, cooking, and aging /
by Matteo Bordiga and Leo M.L. Nollet.
Description: 1st edition. | Boca Raton : CRC Press, 2019. | Series: Food
analysis & properties, 2475-7551 | Includes bibliographical references.
| Summary: “Of the five senses, smell is the most direct and food aromas
are the key drivers of our flavor experience. They are crucial for the
synergy of food and drinks. Up to 80% of what we call taste is actually
aroma. This book deals with how food aromas are developed and affected
during food processing, cooking, and aging.”-- Provided by publisher.
Identifiers: LCCN 2019026347 (print) | LCCN 2019026348 (ebook) | ISBN
9781138338241 (hardback) | ISBN 9780429441837 (pdf)
Subjects: LCSH: Food--Biotechnology. | Food--Odor. | Food--Sensory
evaluation. | Flavor.
Classification: LCC TP248.65.F66 F645 2019 (print) | LCC TP248.65.F66
(ebook) | DDC 664/.024--dc23
LC record available at https://lccn.loc.gov/2019026347
LC ebook record available at https://lccn.loc.gov/2019026348

Visit the Taylor & Francis Web site at

http://www.taylorandfrancis.com

and the CRC Press Web site at

http://www.crcpress.com
 Contents

Preface ix
Editors xi
Contributors xiii

Section I  AROMA, TASTE, AND FLAVOR

Chapter 1 Aroma and Odor 3
Christophe B.Y. Cordella

Chapter 2 Flavors and Taste 15

Christophe B.Y. Cordella

Chapter 3 Chemical Senses and Flavor Perception 23

Han-Seok Seo

Chapter 4 Aroma Compounds (Description, Biosynthesis,

and Regulation) 57
Mónika Valdenegro Espinoza, María Fernanda Flores Echeverría,
and Lida Fuentes Viveros

Chapter 5 Orthonasal and Retronasal Olfaction 99

Pengfei Han and Thomas Hummel

Section II  ANALYTICAL TECHNIQUES

Chapter 6 Extraction Methods of Volatile Compounds from Food
Matrices 123
Arthur Luiz Baião Dias, Francisco Manuel Barrales, and Philipe
dos Santos

Chapter 7 The Role of Gas Chromatography-Based Methodologies for

the Understanding of Food Aromas 141
Cátia Martins, Ângelo C. Salvador, and Sílvia M. Rocha

v
vi Contents

Chapter 8 Monitoring Food Aroma during Processing and Storage by

Rapid Analytical Methods: A Focus on Electronic Noses and
Mass Spectrometry-Based Systems 159
Aoife Power, Vi Khanh Truong, James Chapman, and
Daniel Cozzolino

Chapter 9 Hyphenated Electronic Nose Technique for Aroma Analysis

of Foods and Beverages 177
Adriana Marcia Graboski, Sandra Cristina Ballen,
Juliana Steffens, and Clarice Steffens

Chapter 10 Food Aroma Compounds by Capillary Electrophoresis 193

Raffaella Colombo and Adele Papetti

Chapter 11 Proton-Transfer-Reaction–Mass Spectrometry 217

Iuliia Khomenko and Brian Farneti

Chapter 12 Stable Isotope Dilution Assay 241

Hans-Georg Schmarr

Section III  PRINCIPLES OF PROCESSING,

EVOLUTION, AND MODIFICATION
Chapter 13 Food Processing, Cooking, and Aging: A Practical Case Study 261
Emmanuel Bertrand

Chapter 14 The Maillard Reaction 281

Joseph Provost

Chapter 15 Production of Food Aroma Compounds (Microbial and

Enzymatic Methodologies) 293
Lorena de Oliveira Felipe, Bruno Nicolau Paulino, Adones Sales,
Gustavo Molina, and Juliano Lemos Bicas

Chapter 16 Novel and Emerging Technologies (Benefits and Limitations) 307

Mohammad Hassan Kamani, Hanieh Amani,
Amir Yeganehshakib, and Amin Mousavi Khaneghah

Section IV AROMA COMPOUNDS IN FOOD MATRICES

Chapter 17 Distillates 337
Paul S. Hughes

Chapter 18 Evolution of Beer Aroma 353

Iztok Jože Košir and Miha Ocvirk

Chapter 19 Coffee Flavor 365

Sergio Pérez-Burillo, Matteo Bordiga, Silvia Pastoriza, and
José A. Rufián-Henares
 Contents vii

Chapter 20 Aroma Evolution in Chocolate Production 383

Roberta Ascrizzi, Luisa Pistelli, and Guido Flamini

Chapter 21 Bakery Products 415

Joana Pico and Juan Carlos Diego

Chapter 22 Recent Advances in the Study of Grape and Wine Volatile

Composition: Varietal, Fermentative, and Aging Aroma
Compounds 439
Pilar Rubio-Bretón, Maria Rosario Salinas, Ignacio Nevares,
Eva Pilar Pérez-Álvarez, Maria del Álamo-Sanza, Sandra Marín-San
Román, Gonzalo Luis Alonso, and Teresa Garde-Cerdán

Chapter 23 Milk/Dairy 465

Kieran Kilcawley

Chapter 24 Meat 487

Mónica Bueno, Thais Devincenzi, and Virginia Celia Resconi

Chapter 25 Fish 519

Asghar Amanpour, Gamze Guclu, and Serkan Selli

Chapter 26 Fruits and Vegetables 543

Rajnibhas Sukeaw Samakradhamrongthai

Chapter 27 Spices and Herbs 569

Alejandro Hernández, Emilio Aranda, Rocío Casquete, Cristina
Pereira, and Alberto Martín

Chapter 28 Off-Flavors in Alcoholic Beverages: An Overview 595

Rosa Perestrelo, Catarina Silva, and José S. Câmara

Section V INFLUENCES ON FLAVOR PERCEPTION

Chapter 29 Interactions between the Food Matrix and Aroma
Compounds in Relation to Perception 625
Elisabeth Guichard

Chapter 30 Food Emulsions as Flavor Delivery Systems 651

Like Mao, Yrjö H. Roos, Costas G. Biliaderis, and Song Miao

Chapter 31 Relationship between Structure and Odor 679

Valentina Villalobos Coa, Vito Lubes, Johannes Polster, Maria
Monteiro de Araújo Silva, and Giuseppe Lubes

Chapter 32 Bioactive Potential of Sesquiterpenes 695

Iramaia Néri-Numa, Kele A.C. Vespermann, Carlos H. Carvalho,
Bruno Nicolau Paulino, Maria J. Macedo, Gláucia M. Pastore,
Juliano Lemos Bicas, and Gustavo Molina
Index 719
 Preface

This book focuses on the description of aroma evolution in several food matrices. Of the five
senses, smell is the most “direct.” When we smell the aroma of a delicious dish, the odorous
molecules reach the nasal cavity and are captured by the mucous, which contains olfactory
receptors. These transmit a message to a specific area of our brain, through the olfactory
nerve. This direct contact between the nose and brain explains how a simple smell can trigger
an emotion. Furthermore, the olfactory experience influences the taste of food. This is due to
two processes, called “orthonasal olfaction” and “retronasal olfaction,” which transform the
olfactory signal into one of taste, thus enhancing our perception of flavor. For example, the
Maillard reaction helps create smells that are particularly welcome to the palate, on condition
that the food contains as little water as possible before being cooked. Actually, boiled foods
create some of the least appetizing cooking smells. Conversely, grilled or oven-baked foods
are characterized by “good smells”: high temperatures in fact diffuse odorous molecules more
effectively. Not just cooking, but also processing (such as fermentation) and aging are respon-
sible for food aroma evolution. A comprehensive evaluation of food requires that analytical
techniques keep pace with the available technology. As a result, a major objective in the chem-
istry of food aroma is concerned with the application and continual development of analytical
methods. This aspect, appearing particularly important, is discussed in depth in a dedicated
section of the book. The book results in a good mix of referenced research with practical
applications, also reporting case studies of the various applications of novel technologies.
In the last few years, there have been numerous publications focused on food processing
technology. Furthermore, numerous texts and reference books are available on fermented
foods such as wine, beer, and cheese. However, none of those sources deal with food aroma
evolution during different treatments (such as food processing, cooking, and aging) in a broad
spectrum, including the proper analytical methods. This text represents a comprehensive
reference book for students, educators, researchers, food processors, and food industry per-
sonals providing up-to-date insight. The range of techniques and materials covered provides
engineers and scientists working in the food industry with a valuable resource for their work.
The editors are very happy to thank all contributors for their appreciated contribu-
tions. They value all the time and energy spent to write the different chapters.

Matteo Bordiga, PhD

Leo M.L. Nollet, PhD

I’m honored to give a special thanks to Matteo for giving me the opportunity to work
with him on this project. This work will be a very respected volume in my book series
Food Analysis & Properties.

Leo M.L. Nollet

ix
 Editors

Matteo Bordiga, PhD, received a PhD in Food Science from the Università del Piemonte
Orientale, Novara, Italy in 2010. He received his MS in Chemistry and Pharmaceutical
Technologies from the same university. The main research activity of Dr. Bordiga con-
cerns food chemistry and investigating the different classes of polyphenols from an
analytical, technological, and nutritional point of view. More recently, he moved his
research interests toward wine chemistry, focusing his attention on the entire produc-
tion processss—from vine to glass. He has published more than 40 research papers in
peer-reviewed international journals. Since 2013, he has been an editorial board member
of the International Journal of Food Science & Technology. He has edited the books
Valorization of Wine Making By-Products (CRC Press, 2016) and Post-Fermentation
and -Distillation Technology: Stabilization, Aging, and Spoilage (CRC Press, 2018). All
these research activities have also been developed through important collaborations with
foreign institutions, including the Foods Science & Technology Department and Foods
for Health Institute, University of California, Davis, United States; Fundación Parque
Científico y Tecnológico de Albacete, and Instituto Regional de Investigación Científica
Aplicada, Universidad de Castilla-La Mancha, Ciudad Real, Spain.

Leo M.L. Nollet, PhD, earned an MS (1973) and PhD (1978) in Biology from the Katholieke
Universiteit Leuven, Belgium. He is an editor and associate editor of numerous books. He
edited for Marcel Dekker, New York—now CRC Press of the Taylor & Francis Group—
the first, second, and third editions of Food Analysis by HPLC and Handbook of Food
Analysis. The last edition is a two-volume book. Dr. Nollet also edited the Handbook
of Water Analysis (first, second, and third editions) and Chromatographic Analysis of
the Environment, third and fourth editions (CRC Press). With F. Toldrá, he coedited two
books published in 2006, 2007, and 2017: Advanced Technologies for Meat Processing
(CRC Press) and Advances in Food Diagnostics (Blackwell Publishing—now Wiley).
With M. Poschl, he coedited the book Radionuclide Concentrations in Food and the
Environment, also published in 2006 (CRC Press). Dr. Nollet has also coedited with Y.H.
Hui and other colleagues on several books: Handbook of Food Product Manufacturing
(Wiley, 2007), Handbook of Food Science, Technology, and Engineering (CRC Press,
2005), Food Biochemistry and Food Processing (first and second editions; Blackwell
Publishing—now Wiley—2006 and 2012), and the Handbook of Fruits and Vegetable
Flavors (Wiley, 2010). In addition, he edited the Handbook of Meat, Poultry, and Seafood
Quality, first and second editions (Blackwell Publishing—now Wiley—2007 and 2012).
From 2008 to 2011, he published five volumes on animal product-related books along
with F. Toldrá: Handbook of Muscle Foods Analysis, Handbook of Processed Meats
and Poultry Analysis, Handbook of Seafood and Seafood Products Analysis, Handbook
of Dairy Foods Analysis, and Handbook of Analysis of Edible Animal By-Products.
Also, in 2011, with F. Toldrá, he coedited two volumes for CRC Press: Safety Analysis of

xi
xii Editors

Foods of Animal Origin and Sensory Analysis of Foods of Animal Origin. In 2012, they
published the Handbook of Analysis of Active Compounds in Functional Foods. In a
coedition with Hamir Rathore, Handbook of Pesticides: Methods of Pesticides Residues
Analysis was marketed in 2009; Pesticides: Evaluation of Environmental Pollution in
2012; Biopesticides Handbook in 2015; and Green Pesticides Handbook: Essential Oils
for Pest Control in 2017. Other finished book projects include Food Allergens: Analysis,
Instrumentation, and Methods (with A. van Hengel; CRC Press, 2011) and Analysis
of Endocrine Compounds in Food (Wiley-Blackwell, 2011). Dr. Nollet’s recent proj-
ects include Proteomics in Foods with F. Toldrá (Springer, 2013) and Transformation
Products of Emerging Contaminants in the Environment: Analysis, Processes,
Occurrence, Effects, and Risks with D. Lambropoulou (Wiley, 2014). In the series Food
Analysis & Properties, he edited (with C. Ruiz-Capillas) Flow Injection Analysis of Food
Additives (CRC Press, 2015) and Marine Microorganisms: Extraction and Analysis of
Bioactive Compounds (CRC Press, 2016). With A.S. Franca, he edited Spectroscopic
Methods in Food Analysis (CRC Press, 2017) and with Horacio Heinzen and Amadeo R.
Fernandez-Alba, he edited Multiresidue Methods for the Analysis of Pesticide Residues
in Food (CRC Press, 2017). With J.A. Gutierrez-Uribe, he edited Phenolic Compounds
in Food: Characterization and Analysis (CRC Press, 2018); with Salah Mahgoub, he
edited Testing and Analysis of GMO-Containing Foods and Feed (CRC Press, 2018);
with K.S. Siddiqi, he edited Fingerprinting Techniques in Food Authentication and
Traceability (CRC Press, 2018); with N.C. Basantia and Mohammed Kamruzzaman, he
edited Hyperspectral Imaging Analysis and Applications for Food Quality (CRC Press,
2018); and with Basil K. Munjanja, he edited Ambient Mass Spectroscopy Techniques in
Food and the Environment (CRC Press, 2019).
 Contributors

Maria del Álamo-Sanza Sandra Cristina Ballen

Grupo UVaMOX, E.T.S. Ingenierías Department of Food Engineering
Agrarias URI—Campus of Erechim
Universidad de Valladolid Erechim, Rio Grande do Sul, Brazil
Palencia, Spain
Francisco Manuel Barrales
Gonzalo Luis Alonso Department of Food Engineering
Cátedra de Química Agrícola, E.T.S.I. School of Food Engineering
Agrónomos y Montes University of Campinas
Universidad de Castilla–La Mancha Campinas, São Paulo, Brazil
Albacete, Spain
Emmanuel Bertrand
Hanieh Amani INRA, UMR1163 Biodiversité et
Department of Grain and Industrial Plant Biotechnologie Fongiques
Technology Marseille, France
Faculty of Food Science
Szent István University Juliano Lemos Bicas
Budapest, Hungary Department of Food Science
School of Food Engineering
Asghar Amanpour University of Campinas
Department of Food Engineering Campinas, São Paulo, Brazil
Faculty of Agriculture
Cukurova University Costas G. Biliaderis
Adana, Turkey Department of Food Science
Faculty of Agriculture, Forestry and
Emilio Aranda Natural Environment
School of Agronomics Engineering Aristotle University of Thessaloniki
University Institute of Agronomics Thessaloniki, Greece
Resources (INURA)
University of Extremadura Mónica Bueno
Badajoz, Spain Laboratory of Foodomics
Institute of Food Science Research
Roberta Ascrizzi (CIAL, CSIC-UAM)
Department of Pharmacy Madrid, Spain
University of Pisa
Pisa, Italy

xiii
xiv Contributors

José S. Câmara Daniel Cozzolino

CQM—Centro de Química da Madeira School of Science
RMIT University
and
Melbourne, Victoria, Australia
Departamento de Química
Faculdade de Ciências Exatas e Thais Devincenzi
Engenharia Programa Carne y Lana
Universidade da Madeira Instituto Nacional de Investigación
Campus da Penteada Agropecuaria (INIA)
Funchal, Portugal Tacuarembó, Uruguay

Carlos H. Carvalho Arthur Luiz Baião Dias

Department of Food Science Department of Food Engineering
School of Food Engineering School of Food Engineering
University of Campinas University of Campinas
Campinas, São Paulo, Brazil Campinas, São Paulo, Brazil

Rocío Casquete Juan Carlos Diego

School of Agronomics Engineering Instrumental Techniques Laboratory
University Institute of Agronomics University of Valladolid
Resources (INURA) Valladolid, Spain
University of Extremadura
Badajoz, Spain María Fernanda Flores Echeverría
Sociedad Agroadvance Ltda
James Chapman Peñaflor, Chile
School of Medical and Applied Sciences
CQUniversity Mónika Valdenegro Espinoza
Rockhampton, Queensland, Australia Escuela de Agronomía
Facultad de Ciencias Agronómicas y de los
Valentina Villalobos Coa Alimentos
Laboratorio de Equilibrios en Solución Pontificia Universidad Católica de
Universidad Simón Bolívar (USB) Valparaíso
Caracas, Venezuela Quillota, Chile

Raffaella Colombo Brian Farneti

Department of Drug Sciences Research and Innovation Centre
University of Pavia Fondazione Edmund Mach
Pavia, Italy Trento, Italy

Christophe B.Y. Cordella Lorena de Oliveira Felipe

INRA, UMR 0914 Physiologie de Graduate School of Life and
la Nutrition et du Comportement Environmental Sciences
Alimentaire University of Tsukuba
Groupe Chimiométrie pour Tsukuba, Ibaraki, Japan
la Caractérisation de
Biomarqueurs—C2B Guido Flamini
Paris, France Department of Pharmacy
University of Pisa
Pisa, Italy
 Contributors xv

Teresa Garde-Cerdán Thomas Hummel

Grupo VIENAP Smell and Taste Clinic
Instituto de Ciencias de la Vid y del Department of Otorhinolaryngology
Vino (CSIC, Universidad de La Rioja, TU Dresden
Gobierno de La Rioja) Dresden, Germany
Logroño, Spain
Mohammad Hassan Kamani
Adriana Marcia Graboski Young Researchers and Elite Club
Department of Food Engineering Sabzevar Branch
Universidade Regional Integrada do Alto Islamic Azad University
Uruguai e das Missões Sabzevar, Iran
Erechim Campus
Erechim, Rio Grande do Sul, Brazil Amin Mousavi Khaneghah
Department of Food Science
Gamze Guclu Faculty of Food Engineering
Department of Food Engineering University of Campinas
Faculty of Agriculture Campinas, São Paulo, Brazil
Cukurova University
Adana, Turkey Iuliia Khomenko
Research and Innovation Centre
Elisabeth Guichard Fondazione Edmund Mach
Centre des Sciences du Goût et de Trento, Italy
l’Alimentation, AgroSup Dijon,
CNRS, INRA Kieran Kilcawley
Université Bourgogne Franche-Comté Food Quality & Sensory Science
Dijon, France Teagasc Food Research Centre
Moorepark
Pengfei Han Fermoy, Ireland
Key Laboratory of Cognition and
Personality Iztok Jože Košir
Ministry of Education Department for Agrochemistry and
Faculty of Psychology Brewing
Southwest University Slovenian Institute of Hop Research
Chongqing, China and Brewing
Žalec, Slovenia
Alejandro Hernández
School of Agronomics Engineering Giuseppe Lubes
University Institute of Agronomics Laboratorio de Equilibrios en Solución
Resources (INURA) Universidad Simón Bolívar (USB)
University of Extremadura Caracas, Venezuela
Badajoz, Spain
Vito Lubes
Paul S. Hughes Laboratorio de Equilibrios en Solución
Department of Food Science and Universidad Simón Bolívar (USB)
Technology Caracas, Venezuela
Oregon State University
Corvallis, Oregon
xvi Contributors

Maria J. Macedo Ignacio Nevares

Department of Food Science Grupo UVaMOX
School of Food Engineering E.T.S. Ingenierías Agrarias
University of Campinas Universidad de Valladolid
Campinas, São Paulo, Brazil Palencia, Spain

Like Mao Miha Ocvirk

Teagasc Food Research Centre Department for Agrochemistry and Brewing
Moorepark Slovenian Institute of Hop Research and
Fermoy, Ireland Brewing
Žalec, Slovenia
and
School of Food and Nutritional Sciences Adele Papetti
University College Cork Department of Drug Sciences
Cork, Ireland University of Pavia
Pavia, Italy
Alberto Martín
School of Agronomics Engineering Gláucia M. Pastore
University Institute of Agronomics Laboratory of Food Biotechnology
Resources (INURA) School of Food Engineering
University of Extremadura Institute of Science and Technology
Badajoz, Spain Universidade Federal dos Vales do
Jequitinhonha e Mucuri
Cátia Martins Diamantina, Minas Gerais, Brazil
QOPNA & LAQV-REQUIMTE
Department of Chemistry Silvia Pastoriza
University of Aveiro Facultad de Farmacia
Campus Universitário de Santiago Campus Universitario de Cartuja
Aveiro, Portugal Granada, Spain

Song Miao Bruno Nicolau Paulino

Teagasc Food Research Centre Faculty of Pharmaceutical Sciences
Moorepark Federal University of Amazonas
Fermoy, Ireland Manaus, Amazonas, Brazil

Gustavo Molina Cristina Pereira

Laboratory of Food Biotechnology School of Agronomics Engineering
Institute of Science and University Institute of Agronomics
Technology—UFVJM Resources (INURA)
Diamantina, Minas Gerais, Brazil University of Extremadura
Badajoz, Spain
Iramaia Néri-Numa
Laboratory of Food Biotechnology Rosa Perestrelo
School of Food Engineering CQM—Centro de Química
Institute of Science and Technology da Madeira
Universidade Federal dos Vales do Universidade da Madeira
Jequitinhonha e Mucuri Campus da Penteada
Diamantina, Minas Gerais, Brazil Funchal, Portugal
 Contributors xvii

Eva Pilar Pérez-Álvarez Sílvia M. Rocha

Grupo VIENAP QOPNA & LAQV-REQUIMTE
Instituto de Ciencias de la Vid y del Vino Department of Chemistry
Universidad de La Rioja University of Aveiro
Logroño, Spain Campus Universitário de Santiago
Aveiro, Portugal
and
Centro de Edafología y Biología Aplicada Sandra Marín-San Román
del Segura (CEBAS-CSIC) Grupo VIENAP
Departamento de Riego Instituto de Ciencias de la Vid y
Campus Universitario de Espinardo del Vino
Murcia, Spain Universidad de La Rioja
Logroño, Spain
Sergio Pérez-Burillo
Facultad de Farmacia Yrjö H. Roos
Campus Universitario de Cartuja School of Food and Nutritional
Granada, Spain Sciences
University College Cork
Joana Pico Cork, Ireland
Food Innovation, Structure and Health Lab
School of Engineering Pilar Rubio-Bretón
University of Guelph Grupo VIENAP
Guelph, Canada Instituto de Ciencias de la Vid y del Vino
Universidad de La Rioja
Luisa Pistelli Logroño, Spain
Department of Pharmacy
University of Pisa José A. Rufián-Henares
Pisa, Italy Facultad de Farmacia
Campus Universitario de Cartuja
Johannes Polster Granada, Spain
Nestlé Product Technology Centre
Orbe, Switzerland Adones Sales
Department of Food Science
Aoife Power School of Food Engineering
School of Medical and Applied Sciences University of Campinas
CQUniversity Campinas, São Paulo, Brazil
Rockhampton, Queensland, Australia
Maria Rosario Salinas
Joseph Provost Cátedra de Química Agrícola
University of San Diego E.T.S.I. Agrónomos y Montes
San Diego, California Universidad de Castilla-La Mancha
Albacete, Spain
Virginia Celia Resconi
Dep. Producción Animal y Ciencia de los Ângelo C. Salvador
Alimentos QOPNA & LAQV-REQUIMTE,
Facultad de Veterinaria Department of Chemistry
Universidad de Zaragoza University of Aveiro, Campus
Instituto Agroalimentario de Aragón Universitário de Santiago
IA2—CITA Aveiro, Portugal
Zaragoza, Spain
xviii Contributors

Rajnibhas Sukeaw Samakradhamrongthai Maria Monteiro de Araújo Silva

Department of Food Technology Nestlé Product Technology Centre Food
Faculty of Agro-Industry Singen, Germany
Prince of Songkla University
Songkhla, Thailand Juliana Steffens
Department of Food Engineering
Philipe dos Santos Universidade Regional Integrada do Alto
Department of Food Engineering Uruguai e das Missões
School of Food Engineering Erechim Campus
University of Campinas Erechim, Rio Grande do Sul, Brazil
Campinas, São Paulo, Brazil
Clarice Steffens
Hans-Georg Schmarr Department of Food Engineering
Dienstleistungszentrum Ländlicher Raum Universidade Regional Integrada do Alto
(DLR) Rheinpfalz Uruguai e das Missões
Institute for Viticulture and Oenology Erechim Campus
Neustadt an der Weinstraße, Germany Erechim, Rio Grande do Sul, Brazil
and
Vi Khanh Truong
Faculty for Chemistry School of Medical and Applied Sciences
University Duisburg-Essen CQUniversity
Essen, Germany Rockhampton, Queensland, Australia

Serkan Selli Kele. A.C. Vespermann

Department of Food Engineering Department of Food Science
Faculty of Agriculture School of Food Engineering
Cukurova University University of Campinas
Adana, Turkey Campinas, São Paulo, Brazil

Han-Seok Seo Lida Fuentes Viveros

Department of Food Science Centro Regional de Estudios en Alimentos
University of Arkansas Saludables
Fayetteville, Arkansas Valparaíso, Chile

Catarina Silva Amir Yeganehshakib

CQM—Centro de Química da Madeira School of Biomedical Sciences
Universidade da Madeira Symbiosis International University
Campus da Penteada Pune, India
Funchal, Portugal
 Section I
Aroma, Taste, and Flavor
 Chapter 1
Aroma and Odor
Christophe B.Y. Cordella

CONTENTS

1.1 Smell and Taste: Historical Aspects 3

1.2 Odor of Perfume: Some Social Aspects of Smell 4
1.3 The Scent of Molecules, Vectors of Smell, and Taste: A Molecular Point of View 5
1.4 Perception of Smells—The Sense of Smell 9
References 11

1.1 SMELL AND TASTE: HISTORICAL ASPECTS

Smell and flavor (more commonly known as taste) are two popular words in science,
particularly since the award of the Nobel Prize in Physiology or Medicine in 2004 to
Linda Buck and Richard Axel for their work that led to the discovery of the gene coding
for the synthesis of olfactory receptors. But this has not always been the case, quite the
contrary. Of the five human senses, taste and smell have long been considered as having
minor effects compared to vision or hearing, probably because of their apparent useless-
ness. Basically, humans need sight to move and see where they are going, and hearing to
communicate and protect themselves. In humans, few critical situations involve taste and
smell. In the Middle Ages, and the Renaissance, we paid more attention to masking odors
than studying them, and therefore there was no interest in understanding the mechanisms
that underlie their perception. This state of thinking has long prevailed and led to a total
misunderstanding of how smell and taste work. It was not until the early 20th century, in
the 1930s, that a scientific interest in odor and its perception was born. Dyson (Malcolm
Dyson, 1938) was one of the first to try to formalize a theoretical framework of smell.
Curiosity and scientific work on smell and the mechanisms of olfaction also developed
from ideas proposed by Moncrieff (1949, 1954), who created models of steric interaction
to explain the perception of odors. This was just the beginnings of a molecular under-
standing of smell and its molecular receptors.
Scientists became interested in the olfactory membrane, in odor receptors, the stereo-
chemical aspects of olfaction (two molecules that perfectly reflect one another in a mirror
can result from a different perception of smell), and in the physiological mechanisms of
the translation of smell, called transduction, which transforms chemical information into
electrical information interpreted by our brain. We now understand how much effort
and investigation is necessary to open the door to a clearer understanding of olfaction.
Thanks to a better understanding of olfaction, we know that these senses are intimately
linked to each other and cannot fully express themselves independently. By highlight-
ing the genetic mechanisms of olfaction, and by identifying the gene set coding for the

3
4 Food Aroma Evolution

synthesis of receptors (proteins) present in the neurons of the olfactory epithelium of the
same name, L. Buck and R. Axel further revisited knowledge on how olfaction works
(Buck and Axel, 1991). Olfaction is currently believed to be based on the interaction
between volatile molecules forming odor and olfactory receptors. The work of Buck and
Axel blows away older representations of odor and perception such as the vibrational or
wave vision of smell (Zhu et al., 2016), or the theory of odotopes and olfactophores (Mori
and Shepherd, 1994). The vibrational theory of smell considered the olfactory receptors
as a vibrational spectroscope. Molecules should be easily recognized by their vibrational
spectrum. The odotope theory was based on the shape of the molecular fragments imply-
ing a purely geometric recognition of the olfactory receptors.
Today, it is understood that an odorant must possess certain molecular features in
order to provide sensory properties. It must have some water solubility, a sufficiently high
vapor pressure, low polarity, some ability to dissolve in fat (lipophilicity), a surface activ-
ity, and a molecular weight lower than 300 Da. It does not need to have particular func-
tional chemical groups or be chemically active (Fernandez and Chemat, 2012). Odorant
compounds can have all the major functions of organic chemistry: alcohols, carbonyl
compounds (mainly aldehydes and ketones), esters, phenols, and sulfur or nitrogen deriv-
atives. Terpenes (C10 or C15 hydrocarbons) and terpenoids (functionalized terpenes) are
nevertheless the most widespread and the most abundant. The olfactory sense is able to
distinguish between a practically infinite number of chemical compounds at very low
concentrations. This number has been estimated at 400,000 (Mori and Yoshihara, 1995).
Recently, Bushdid et al. (2014) claimed that humans can discriminate more than one
trillion odors, but this assumption, based on the results of psychophysical testing, is
still debated in the scientific community. The actual capacity of humans to discriminate
between odor mixtures and also if any other animal can discriminate between them is
still unknown (Grabe and Sachse, 2018).

1.2 ODOR OF PERFUME: SOME SOCIAL ASPECTS OF SMELL

Even if we understand more about what a smell is and how we perceive it, the need to mask
some odors and especially body odors is not new. Since antiquity, we have used many dif-
ferent scented waters, extracts, and balms, often derived from natural plant extracts (e.g.,
geranium and vetiver on the island of Reunion or ylang-ylang in Mayotte) or of animal
origin (e.g., castoreum and civette, substances from the glandular secretions of small mam-
mals). Ancient civilizations, such as those of Egypt, Greece, Persia, and Rome, are rich in
examples of how perfumes and spices have been intricately woven into the fabric of vari-
ous societies. The Enuma Elish, a cuneiform text dating from thousands of years before
Christ, indicated that fragrant oils were widely used throughout the Middle East to provide
skincare and protection from the hot and dry environment, and spices and fragrances were
added to wine (Heidel, 1949). The development of tanneries in France (especially in Grasse,
France) in the Middle Ages, and then their decline, paved the way for modern perfumes.
Thanks to a microclimate favorable to the growth of scented plants, tanners evolved in the
18th century, initially becoming glovers and perfumers, and then perfumers exclusively.
Gloves and leather garments were often associated with fragrance. The idea was thus to
present to our taste and olfactory papillae another image that Mother Nature had sent us
from these skins. In public, we do not accept smells more now than we did before, but we
have learned to study them. Odors are recorded, analyzed, represented, and artificially
synthesized to mimic the originals found in nature. With a better understanding of the
 Aroma and Odor 5

molecular aspects of odors, we now know better how to satisfy our sensory organs and
how to show what we prefer. Odor control requires knowledge for measuring, differentiat-
ing, quantifying, and comparing them, which is the main purpose of the electronic nose.
Meanwhile, considerable interest has also developed in the study and understanding of the
mechanisms of the perception of taste sensations and, in this area, the electronic tongue is
the complementary tool of the e-nose.

1.3 THE SCENT OF MOLECULES, VECTORS OF SMELL,

AND TASTE: A MOLECULAR POINT OF VIEW

Leaning on a flower, a rose for example, to experience its scent or smell in places that we
pass every day is so familiar that we do not imagine living otherwise. One rarely won-
ders how this is possible and how the perception of smell helps us to live and understand
our environment. Yet behind this simple act lie extraordinary biochemical processes jug-
gling with nuances and diversity. In this area, many theories (Ohloff, 1994; Sell, 1999)
have been developed in order to explain the chemical and biochemical mechanisms that
enable a fragrant compound to generate a particular signal interpretable by the brain as an
odor. However, none of these theories have been able to report any experimental evidence.
Indeed, we have known since the early 20th century that some compounds, such as carvone
or menthol, have similar organoleptic properties even though they have a different chemical
formula. Carvone (Figure 1.1) is a very important monoterpene ketone for the flavor indus-
try. S-(+)-carvone is the main component of caraway oil and dill, with an odor resembling
these herbs. The other isomer (R-(−)-carvone) occurs at high concentrations (70–80%) in
spearmint oil and is also the major component responsible for its aroma. The aroma per-
ception threshold for carvone is between 6.7–820 ppb, for S-(+)-carvone, and 2.7–600
ppb for the R-(−)-isomer (Paula Dionísio et al., 2012). Conversely, we know of many mol-
ecules whose enantiomers cause different organoleptic sensations even though they have
the same chemical formula. Ohloff (Rienäcker and Ohloff, 1961) was the first to publish
results on the enantioselective perception of chiral odorous compounds: (+)-β-citronellol
was described as having a typical smell of lemon while (–)-β- citronellol produced an odor
of geraniums. Since then, many optically active odorous compounds have been identified
and are commonly used in perfumes and food flavorings (see Table 1.1) .

FIGURE 1.1 Carvone (2-me​thyl-​5-(1-​methy​lethe​nyl)-​2-cyc​lohex​en-1-​one).​ Member of a

family chemicals called terpenoids. Carvone has two mirror image forms or enantiomers
R(–)-carvone and S(+)-carvone.
6 Food Aroma Evolution

TABLE 1.1 Classical Example of Enantiomers Showing Different Odor Properties

Compounds
(See chemical structures in Figure 1.2) Odor description
7-Hydroxy-6,7-dihydro-citronellal (4) (+): Lily of the valley with green minty notes
(−): Sweet lily of the valley note
Linalool (5) (+): Sweet, petitgrain
(−): Woody, lavender
Carvone (6) (+): Caraway
(−): Spearmint
Nootkatone (7) (+): Grapefruit
(−): Woody, spicy
Nerol oxide (8) (+): Green, floral
(−): Green, spicy, geranium
Androstenone (9) (+): Odorless
(−): Sweaty, urine, strong, musky
Menthol (10) (−): Sweet, fresh, minty, strong cooling effect
(+): Dusty, vegetable, less minty, less cooling
Limonene (11) (+): Orange
(−): Turpentine
Source: Extracted from Brenna et al., 2003.

FIGURE 1.2 Classical enantiomers showing different odors properties. (Extracted from
Brenna et al., 2003.)
 Aroma and Odor 7

Thus, enantiomers differ either in the nature and quality of the odor (difference
in the sensations of smell) or in the intensity of the smell. Certain structural features
of molecules tend to increase the strength of odorants. Moncrieff listed the clearest of
these: (1) polar functional groups (OH, C=O, CN, SH, –O–, etc.) increase intensity,
(2) unsaturation generally increases intensity, (3) steric shielding of a functional group
decreases intensity, and (4) when two hydrogen bond acceptors are present, the odorant
is stronger when they are close to each other (Ohloff’s bifunctional rule (Ikan, 1995)).
The perception of smell is physiologically limited, and this limit is called the odor percep-
tion threshold. This can be expressed as the mass of odorant per unit volume of air and
is measured with various techniques such as olfactometry, in solution using Guadagni’s
method (Schimmer and Guadagni, 1962), or with the triangle test.
Usually, a scent is classified into two categories, depending on its use: fragrances and
flavorings. As reported by Brenna et al. (2003), in a significant review, fragrances are
chemical compounds used in functional and fine perfumery and flavorings are contained
in or added to foods and beverages. These can be natural or identical to natural to comply
with the regulations. Note that the enantiomeric excess of a compound, that is, the ratio
between the two optically active forms, plays an important role in the perception of odor.
The absolute configuration of chiral centers seems to have a particular influence on the
perception of odor. This involves a wide range of olfactory receptors, leading to a highly
generic form of the adaptability of protein receptors with multiple configurations of enan-
tiomeric odorants (see Figure 1.3 and Table 1.2). Finally, the concentration of an odorant
is also a key parameter that significantly changes the perception of the smell. A classic
example is thioterpineol which has a scent of passion fruit at low concentrations, a smell
of grapes at moderate concentrations, and produces a totally unpleasant or unsustainable
odor at high concentrations (Brenna et al., 2003; Malnic et al., 1999).
As indicated by Zampini and Spence (2012), smell and taste are part of a group of
perceptions that follow the natural order, and in the case of food we first find the visual

FIGURE 1.3 Irones 104–106 are the odoriferous principles of natural Orris root
oil. They were first isolated from the iris rhizome in 1893. (Extracted from Brenna
et al., 2003.)
8 Food Aroma Evolution

TABLE 1.2 Odor Descriptions of Irones Samples According to Brenna et al., 2003
Compound Odor
(see Figure 1.3) Odor description threshold
(+) 104a Violet-like, with woody, methylionone undertones 100 ppma
(–) 104a Slightly stronger with a distinct “orris-butter” character 10 ppma
(+) 104b It was described as the weakest of the α-isomers
(–) 104b It shows a weak violet/wood/red berry character. Neither (+)- nor
(−)-trans- α -irone possesses the characteristic “orris” odor
(+) 106a It shows a floral, fatty, sweet, and woody odor character, an
ionone-type odor with slightly sweet aspects
(–) 106a It shows a β-ionone-type odor of warm floral-woody tonality. 0.75 ng/lb
Green aspects are present, too. It shows some fruity nuances,
reminiscent of pineapples. The odor is linear, and it can be
considered a dry-down note
(+) 106b It is very weak, of a woody odor tonality 113.5 ng/lb
(–) 106b It is not very powerful, but it possesses a soft “orris-butter”-type 26.35 ng/lb
of odor
(+) 105 It possesses a β-ionone-type odor of warm floral-woody tonality
with green and anisic aspects. The odor is linear, and the tenacity
of the note is good. It can be considered a dry-down note
(–) 105 It has a woody odor with a distinct honey note, that is quite sweet.
Furthermore, it shows floral ionone-type facets, and a fruity
tonality, but also an unpleasant smoky character. It belongs to the
β-ionone-type family, without being very close to β-ionone
a Odor threshold by triangular test.
b Odor threshold by GC olfactometry.

properties, tactile and kinesthetic perceptions (via hands and cutlery), the smell, sen-
sations in the mouth: flavor, texture, aroma, and finally the residual perception (after
swallowing). Generally, this succession of perceptions often leads novice consumers to
confuse aroma (information from the olfactory epithelium) with flavor (information from
gustatory bulbs present mainly under the tongue). Having the feeling that perception is
localized in the mouth, one refers to this perception by using the generic term “taste,” a
term that can encompass information on texture. In fact, the combination of aroma and
taste is the flavor of food. We see here that flavor and smell are closely linked and play an
essential role in the sensory representation of objects around us. The odorous compounds
that form flavors are organic molecules of low molecular weight; they have a partial
­pressure of vapors that is sufficiently high at atmospheric pressure and ambient tem-
perature to allow a fraction to be released into the headspace surrounding their original
media (e.g., food). During breathing, the molecules reach the olfactory mucosa and cause
a stimulus. In a food, natural aroma represents a very small percentage (<2% by mass)
and consists of hundreds of volatile molecules (over 800 in coffee). Depending on the
nature of the food, these molecules do not play the same role in the formation of aroma.
Sometimes, a single molecule may carry the main aromatic note of the food (as in the case
of 1-octen-3-ol from fungi), or an illusion of the food can be given by a combination of
molecules (2-ethylbutyrate of methyl, trans-2-hexenal and hexenal in apple aroma) where
all the molecules are involved in the aroma but none, alone, is characteristic of the odor
(as in the case of pepper and chocolate).
 Aroma and Odor 9

It is still impossible today to predict the odor of a molecule from its chemical struc-
ture with a high degree of reliability. The establishment of structure–odor relationships
is intrinsically more complex than structure–activity relationships. This is because the
“answer” is not a physical property or a measurable biological activity, but a percep-
tion that can only be described qualitatively by the human subject. The question could
also be approached from two other points of view: on the one hand, the physiological
effects induced by odorants, and on the other hand the cognitive and linguistic aspects
related to the naming of odors. The former can be observed and measured in experimen-
tal situations by many techniques; the latter in the human sciences. In the area of ​​odor
description, mention must be made of the well-known deficiencies of our vocabulary
when it comes to describing odors (Rouby et al., 2002; Zampini and Spence, 2012).
Despite standardization efforts to arrive at a more objective description of these (Jaubert
et al., 1995), the problem of classifying responses into stable and perfectly differentiated
categories remains unresolved. Indeed, it appears that the olfactory space is a complex
multidimensional space in which categories do not organize hierarchically (Chastrette,
2002). Therefore effective structure–odor relationships only remain achievable for the
best-defined odor classes with a broad consensus.

1.4 PERCEPTION OF SMELLS—THE SENSE OF SMELL

The process begins in the nasal cavity where odorants bind to specific receptors (olfac-
tory receptors present in the olfactory epithelium). In humans, about 1% of the genome
is devoted to genetic information coding for the olfactory receptors. Thanks in part to
the work of Linda Buck and Richard Axel, we know now that these receptors belong to
a broader family of proteins called GPCRs (for “G protein-coupled receptors”). These
proteins have structural characteristics allowing them to adopt a large number of spatial
arrangements for the reception of odorant molecules. This variability in the form of the
molecular container that is the olfactory receptor is the major reason for the wide range
of odorants that can be detected and discriminated. Olfactory receptors constitute the
terminal portion of neurons dedicated to the sense of smell. Their ends called dendrites
are clustered in the olfactory bulb. Each olfactory neuron carries only one type of recep-
tor and several neurons expressing the same type of receptors are assembled in structures
called glomeruli. With an ordered projection of their axons, neurons having the same
receiver, scattered in the nasal mucosa, come together in one or two olfactory glomeruli.
Olfactory information is pooled before the cerebral step and leads to the formation of
a map of odors in the olfactory bulb. Therefore, molecules with different sizes or chemical
properties activate different areas of the olfactory bulb. These patterns function as virtual
“odor images,” and it has been hypothesized that these odor images are the basic build-
ing blocks of the odor discrimination process, analogous to the retinal images that are the
basis for the discrimination of visual pattern stimuli (Xu et al., 2003). The image produced
is different because the smell is also different (see Figure 1.4) (Zou et al., 2008). In fact,
the spatial organization of the olfactory system is much more dispersed and complex than
that exposed here. It has been reported not only that the sensory pathways of olfaction are
composed of a set of structures (see Figure 1.5) contributing to the detection, recording,
and processing of the olfactory stimulus, but it is also admitted that the sense of olfaction
exchanges multiple interactions with the other human senses (Auvray and Spence, 2008).
This multiplicity of interactions between taste, smell, touch, and the trigeminal system
(which will be discussed in the next chapter) has led many authors to define flavor as a
10 Food Aroma Evolution

FIGURE 1.4 Schematic overview of the basic steps of the central processing of odorous
stimuli. Odorants are first detected by receptors at the top of the nasal cavity, and from
there the signal travels to the olfactory bulb (1). This signal is then routed to the piriform
cortex (2) and subsequently to the orbitofrontal cortex (3), among other structures. Note
the dual route that odorants can take to reach the receptors at the top of the nasal cav-
ity. The route via the nostrils is known as orthonasal olfaction, whereas the route via
the back of the throat is known as retronasal olfaction. See the text for further details.
(Extracted from Lundström et al., 2010.)

synthesis of these systems, unified by the act of eating (Abdi, 2002; Auvray and Spence,
2008; Small and Prescott, 2005).
This operating mode and treatment of the olfactory stimulus involves a larger part of
the brain in humans than in other species. As reported by Lundtröm et al. (2010), in an
excellent review, this is a form of biological compensation that would increase the cogni-
tive processing of our brain while remaining independent of the number of peripheral
receptors, a mechanism that other non-primate species would not have.
The physiological aspects of smell and a description of the biochemical mechanisms
involved will be further described in Chapter 3: “Chemical Senses and Flavor Perception.”
Today, the field of neuroscience has made considerable progress, and brain imaging
techniques such as functional magnetic resonance imaging (fMRI) or positron emission
tomography (PET) allow us to go further in understanding the neuronal treatment of the
three chemical senses. Only in the last 25 years have significant advances been made in
our understanding of the cerebral location of the treatment of odorous stimuli, tastants,
and trigeminal stimuli (Gottfried, 2010; Savic et al., 2000; Savic, 2002a,b; Wiesmann
 Aroma and Odor 11

FIGURE 1.5 Olfactory sensory pathway—olfactory bulb projections. The outward-fac-

ing arrows represent inputs sent to other series of structures such as the obitofrontal cor-
tex (OFC), the agranular insula, the hippocampus, and so on. (Adapted from Lundström
et al., 2010.)

et al., 2004; Zatorre et al., 1992). Brain imaging is a field of research in very strong
development that involves important technical capacities that require a certain interdis-
ciplinary approach (neuroscience, magnetic resonance imaging, computer science, and
chemometrics) (Popovych et al., 2019).

REFERENCES

Abdi, H. (2002). What can cognitive psychology and sensory evaluation learn from each
other? Food Quality and Preference 13, 445–51.
Auvray, M. and Spence, C. (2008). The multisensory perception of flavor. Consciousness
and Cognition 17, 1016–31.
Brenna, E., Fuganti, C., and Serra, S. (2003). Enantioselective perception of chiral odor-
ants. Tetrahedron: Asymmetry 14, 1–42.
Buck, L. and Axel, R. (1991). A novel multigene family may encode odorant receptors: A
molecular basis for odor recognition. Cell 65, 175–87.
Bushdid, C., Magnasco, M. O., Vosshall, L. B., and Keller, A. (2014). Humans can dis-
criminate more than 1 trillion olfactory stimuli. Science 343, 1370–2.
Chastrette, M. (2002). Classification of odors and structure–odor relationships. In
Olfaction, Taste, and Cognition (A. Holley, B. Schaal, C. Rouby, D. Dubois, and R.
Gervais, eds.), pp. 100–16. Cambridge University Press, Cambridge.
Fernandez, X. and Chemat, F. (2012). La chimie des huiles essentielles: Tradition et
innovation, Vuibert, Paris.
12 Food Aroma Evolution

Gottfried, J. A. (2010). Central mechanisms of odour object perception. Nature Reviews

Neuroscience 11, 628.
Grabe, V. and Sachse, S. (2018). Fundamental principles of the olfactory code. Biosystems
164, 94–101.
Heidel, A. (1949). The Gilgamesh Epic and Old Testament Parallels, 2nd ed., University
of Chicago Press, Chicago, IL.
Ikan, R. (1995). Scent and fragrances: The fascination of odors and their chemical
perspenives, Günther Ohloff, translated from the original German by Wilhelm
Pickenhagen and Brian M. Lawrence, Springer-Verlag, Berlin, 1994. No of pages:
xvi + 238, price DM 128.00. ISBN 3-540-57108-6. Flavour and Fragrance Journal
10, 339 .
Jaubert, J. N., Tapiero, C., and Dore, J. C. (1995). The field of odors: Toward a universal
language for odor relationships. Perfumer & Flavorist 20, 1.
Lundström, J. N., Boesveldt, S., and Albrecht, J. (2010). Central processing of the chemi-
cal senses: An overview. ACS Chemical Neuroscience 2, 5–16.
Malcolm Dyson, G. (1938). The scientific basis of odour. Journal of the Society of
Chemical Industry 57, 647–51.
Malnic, B., Hirono, J., Sato, T., and Buck, L. B. (1999). Combinatorial receptor codes for
odors. Cell 96, 713–23.
Moncrieff, R. W. (1949). What is odor. A new theory. American Perfumer 54, 453.
Moncrieff, R. W. (1954). The characterization of odours. The Journal of Physiology 125,
453–65.
Mori, K. and Shepherd, G. M. (1994). Emerging principles of molecular signal process-
ing by mitral/tufted cells in the olfactory bulb. Seminars in Cell Biology 5, 65–74.
Mori, K. and Yoshihara, Y. (1995). Molecular recognition and olfactory processing in the
mammalian olfactory system. Progress in Neurobiology 45, 585–619.
Ohloff, G. (1994). Scent and Fragrances, Springer, Berlin Heidelberg.
Paula Dionísio, A., Molina, G., Souza de Carvalho, D., dos Santos, R., Bicas, J. L., and
Pastore, G. M. (2012). 11—Natural flavourings from biotechnology for foods and
beverages. In Natural Food Additives, Ingredients and Flavourings (D. Baines and
R. Seal, eds.), pp. 231–59. Woodhead Publishing, Sawston, United Kingdom.
Popovych, O. V., Manos, T., Hoffstaedter, F., and Eickhoff, S. B. (2019). What can com-
putational models contribute to neuroimaging data analytics? Frontiers in Systems
Neuroscience 12, 68.
Rienecker, R. and Ohloff, G. (1961). Optisch aktives β-Citronellol aus (+)- oder (–)-Pinan.
Angewandte Chemie 73, 240.
Rouby, C., Schaal, B., Dubois, D., Gervais, R., and Holley, A. (2002). Olfaction, Taste,
and Cognition, Cambridge University Press, Cambridge.
Savic, I. (2002a). Brain imaging studies of the functional organization of human olfac-
tion. The Neuroscientist 8, 204–11.
Savic, I. (2002b). Imaging of brain activation by odorants in humans. Current Opinion
in Neurobiology 12, 455–61.
Savic, I., Gulyas, B., Larsson, M., and Roland, P. (2000). Olfactory functions are medi-
ated by parallel and hierarchical processing. Neuron 26, 735–45.
Schimmer, S. and Guadagni, D. G. (1962). Relation between olfactory threshold concen-
tration and pyruvic acid content of onion juice. Journal of Food Science 27, 94–7.
Sell, C. (1999). Chemoreception. In The Chemistry of Fragrances (D. H. Pybus and
C. Sell, eds.), pp. 216–26. Royal Society of Chemistry, London.
 Aroma and Odor 13

Small, D. M. and Prescott, J. (2005). Odor/taste integration and the perception of flavor.
Experimental Brain Research 166, 345–57.
Wiesmann, M., Kettenmann, B., and Kobal, G. (2004). Functional magnetic resonance
imaging of human olfaction. Flavor Perception, 203–27.
Xu, F., Liu, N., Kida, I., Rothman, D. L., Hyder, F., and Shepherd, G. M. (2003). Odor
maps of aldehydes and esters revealed by functional MRI in the glomerular layer of
the mouse olfactory bulb. Proceedings of the National Academy of Sciences 100,
11029–34.
Zampini, M. and Spence, C. (2012). Assessing the role of visual and auditory cues in
multisensory perception of flavor. In The Neural Bases of Multisensory Processes
(M. M. Murray and M. T. Wallace, eds.), Chapter 37, CRC Press/Taylor & Francis.
Available from: https​://ww​w.ncb​i.nlm​.nih.​gov/b​ooks/​N BK92​852/.​
Zatorre, R. J., Jones-Gotman, M., Evans, A. C., and Meyer, E. (1992). Functional local-
ization and lateralization of human olfactory cortex. Nature 360, 339–40.
Zhu, C. C., Sun, Y. L., Luo, D. H., Bai, G., Zhou, J. L., Li, H., Yin, Y., and GholamHosseini,
H. (2016). The study of multimodal gas recognition algorithm based on machine
olfaction. 2016 ICCE International Conference on Consumer Electronics-China
(ICCE-China).
Zou, Z., Horowitz, L. F., Montmayeur, J. P., Snapper, S., and Buck, L. B. (2008). Genetic
tracing reveals a stereotyped sensory map in the olfactory cortex. Nature 452, 120.
 Chapter 2
Flavors and Taste
Christophe B.Y. Cordella

CONTENTS

2.1 Historical Aspects of the Taste Sense 15

2.2 Current Knowledge of the Taste Sense: What Is Flavor? 17
Glossary 21
References 21

2.1 HISTORICAL ASPECTS OF THE TASTE SENSE

Similar to the olfactory sense, written traces of the taste sense go back to ancient civiliza-
tion, especially the ancient texts of Greek scholars. Alcmaeon, Pythagoras’ first disciple (born
around 516 bc) (Vicq D’Azyr, 1787) describes the tongue as constituted by a multitude of tiny
pores admitting tastants to the sensorium: the brain. Democritus (460–370 bc), an atomic
theory supporter, defends the idea according that the basic sensations can be attributed to
specific geometric properties. Consequently, sourness is attributed to angular shapes, sweet-
ness to spherical shapes, and bitterness to small spherical shapes with hooks attached. The
first list of basic taste qualities appears with Aristotle (384–322 bc). Parmenides, Empedocles,
Anaxagoras, Democritus, Epicurus, and Heraclides held that the particular sensations are
produced in us by the symmetrical relations between the pores of the sense-organ and the
object of sense; that is, each sense has its own proper object of perception that exactly fits
into its pores (Beare, 1906). It is interesting to note that the conception of sense perception
in which some objects (molecules) should have a good correspondence with some anatomi-
cal parts of the sense-organ and can trigger actions/reactions, has probably inspired Emile
Fisher, 1500 years later, who will propose its key-lock model (1894) that explain the enzyme–
substrate interaction. The contribution of the Romans to physiology and medicine has been
fundamental and their knowledge of the meaning of taste has come to us through the work
and observations of Galen (ad 180–200) who listed, like Aristotle, seven basic taste sensa-
tions, but unfortunately his descriptions for some of them remained unclear, especially for
the qualifiers harsh, pungent, and astringent. Galen thought that taste was more reliable than
smell because odoriferous substances often secreted insufficient vapor into the air to stimu-
late the olfactory organ, while the tongue was directly stimulated by elementary particles.
Moreover, he suggested that the organ of taste was stimulated by even more coarse particles.
Clearly, Galen had not adopted the non-corpuscular concept of taste perception of Aristotle,
according to which taste is conveyed by the transfer of qualities of the object to the tongue.
Aristotle’s perception of taste was based on the observation that water that passes through
other substances is “charged” with certain qualities that communicate the sensation of the
aromas. Galen had not only proved irrefutably that the brain was the exclusive seat of all

15
16 Food Aroma Evolution

sensory perceptions, but he had been able, through detailed studies on the innervation of the
tongue, to correctly describe the different functions of the three main nerves supplying the
tongue and demonstrate their origin at the base of the brain (Siegel, 1970). The Middle Ages
is especially marked by the contribution of Arabic scholars who from the Greek writings
developed their own sciences; and in the field of physiology and medicine we note the writings
of Avicenna (980–1037), which lists five taste sensations: sweet, salty, bitter, sour, and insipid
(Gruner, 1973). Over 500 years later, in 1542, Western science returned to the forefront of
the international scene with the works of Jean Fernel (1497–1558, French physician and phi-
losopher). Fernel lists nine taste sensations of which seven are identical to those of Aristotle
and Galen. The two new ones are “fatty” and “insipid.” He defines the latter as probably not
a flavor but an absence of flavor, a kind of qualitative zero, that is, without any capacity to
trigger a stimulus in the mouth (tasteless). We now know that the taste sensation produced
by distilled water is mediated by fibers responding to the removal of the NaCl ions in saliva.
During the Age of Enlightenment, some scientists discovered new sensations that were first
taken to be fundamental sensations, such as “spiritual,” “acrid,” “aromatic,” “rough” pro-
posed by von Haller (von Haller and Cullen, 1803), or “alkaline” proposed by Horn, or
“metallic” proposed by Wundt. Von Haller listed 11 new sensations; however, he did not
understand the origin of these sensations, but he did observe that a stimulus too intense could
produce pain in the mouth. Subsequently, the more exotic qualities such as urinous, putrid,
or spirituous were gradually abandoned as the knowledge of the anatomy of the sense of taste
progressed (see Figure 2.1). The 19th century saw the emergence of two opposing scientific
views of taste: Kiesow’s and Öhrwall’s, students of W. Wundt and F. Holmgren, respectively.
Wundt and Holmgren were students of H. von Helmoltz, who proposed a distinction between
quality and modality. Briefly, Kiesow rejected Öhrwall’s assertion that the four basic tastes

FIGURE 2.1 Selected lists of taste quality names. (Adapted from Carterette and Friedman,
1978.)
 Flavors and Taste 17

were different modalities and instead insisted that chemical nature of the adequate stimuli for
these four tastes was sufficient to make them qualities within one modality. Kiesow claimed
to have demonstrated taste contrast and compensation, contrary to Öhrwall’s assertion that
these phenomena did not exist (Carterette and Friedman, 1978).
In the 20th century, the four basic sensations of taste were finally adopted and formalized
by Henning (1915) using a tetrahedron in which the primary sensations were placed in each
of the corners. Henning was not the first to make proposals of order systems for the sense
of taste; Linnaeus was the first to propose a list of seven categories of smell in the middle of
1700. And later in the 19th century, Zwaardemaker reformulated Linnaeus’ classificatory
system and enlarged it to nine categories. Finally, Henning transformed the list into a three-
dimensional system where odors vary in an incremental continuum and are related to one
another. The model is based on a three-dimensional representation (tetrahedron). The six
fundamental or primary odors were placed in each corner: flowery, foul, fruity, spicy, burned,
resinous. In this continuity, Henning also applied the same concept to represent the order and
variation of taste sensations (Schiffman and Erickson, 1971). But Henning believed that no
substance could create new sensations resulting from a combination of four primary sensa-
tions simultaneously. Therefore, the tetrahedron should be hollow. Henning’s model was a
useful tool for guiding research in the field but suffered from limitations, making it incom-
plete; for example, the fact that it is a closed-ended model. Thus, any stimuli which may not
be phenomenologically describable in terms of the four primary qualities cannot be ordered
by the model. Second, the model is only a general order of stimuli and the exact distances
between stimuli have not been determined. Thus, in addition to being qualitatively incom-
plete, Henning’s model is quantitatively imprecise. It will take a few more years for the scien-
tific community to recognize that the formation of taste sensations is expressed in a complex
multidimensional space, that is, one molecule can stimulate several sensory receptors and not
just one receptor cell. A combination of molecules produces a set of sensory stimuli which is
different from another combination of the same molecules in different proportions or concen-
trations. These facts are the basis of the multidimensional space of the odors’ perception. The
20th century witnessed interest in various issues such as cross-adaptation between substances
(that is related to the mechanism by which olfactory transductions are mediated by inde-
pendent pathways: InsP3- and cAMP-producing odorants) or the effect of adaptation on the
threshold of detection of the compounds. The advent of electrophysiology techniques revolu-
tionized the field and allowed further investigations into the measurement of neuronal activ-
ity associated with the sensation of taste. This is the appearance of the so-called “cross-fiber
patterning theory” of taste quality. The new psychophysics also helped dispel the confusion
surrounding tasteless and watery tastes. Because of the worldwide interest in the underlying
biological mechanisms, the search for taste (and olfaction) is now a truly international disci-
pline. Today, many research teams are working on this research object in the United States,
Japan, Germany, France, Chile, and other countries around the world.

2.2 CURRENT KNOWLEDGE OF THE TASTE SENSE: WHAT IS FLAVOR?

It is admitted that the perception of flavor is the most multisensory of our everyday expe-
riences, and among all our senses the perception of taste is 90% due to smell, especially
because smell interacts on two levels. On the one hand, the nose perceives aromas and on the
other hand, aromas are released when food is chewed and disintegrated in the mouth lead-
ing to complementary sensory stimuli. Therefore, under the effect of temperature increase,
new volatile molecules are released and reach the olfactory receptors, but this time through
18 Food Aroma Evolution

FIGURE 2.2 Temporal and spatial determinants of odor/taste integration. (Extracted

from Small and Prescott, 2005.)
the retronasal route (Figure 2.2). The combination of these two modes of perception with
sensory information from the tongue and oral cavity contributes to the taste formation.
Smell, therefore, plays a leading role in the taste sensation. But what is the mechanism of
this sense of taste? In the chemistry of the mouth, the decisive factor is the saliva that allows
the release of palatable molecules that partially react with these chemical receptors on the
tongue. The tongue possesses 10,000 sensory receptors, called taste buds. These organs are
also present on the soft palate, pharynx, larynx, epiglottis, uvula, the upper third of the
esophagus, and the lips and cheeks (Schiffman and Gatlin, 1993). Taste buds are located in
areas of the lingual mucosa called papillae, giving the tongue its rough texture. Three types
of papillae line the surface of the tongue: filiform (or foliate), fungiform, and circumvallate
papillae. The last two types contain the most taste buds. These three types of papillae cre-
ate the collection of five types of taste sensations caused by the mixture of sensations called
primary qualities (flavor sensations): sweet, sour, salty, bitter, and umami* or savory, which
was shown to be the fifth flavor only in the last 20 years. These flavors can be caused by
many different organic molecules. A sweet flavor is caused specifically by sugars, alcohols,
some amino acids, and some minerals. A sour flavor will naturally be caused by acid with
the release in the mouth of hydrogen ions, H+. In contrast, the ions of inorganic salts are
the cause of a salty taste. A well-known representative of inorganic salt is sodium chloride
which causes the most powerful salty sensation. Last, a bitter flavor will be induced by
organic molecules of larger size and molecular weight that belong to the family of alka-
loids, such as quinine, caffeine, nicotine, strychnine, and morphine. We find, however, that
some non-alkaloid molecules are capable of triggering a bitter taste, such as acetylsalicylic
acid (the active ingredient in aspirin). Although the sensation of taste may be triggered by
an odor or a trigeminal stimulus, it is always perceived as a taste (Lundstrom et al., 2011).
All tongues are sensitive to various tastes, but there are tongue-regional differences that
will define areas with a sensory preference. This preference is related to the nature of taste

* The umami taste is triggered by monosodium glutamate (E621-NaC5H8NO4 exhauster [Deppenweiler,

2014]) and recalls the taste of broth or meat. MSG is the sodium salt of glutamic acid. Glutamate is a natu-
rally occurring amino acid found in almost all foods, especially those rich in protein such as dairy, meat or
fish, and in many vegetables. Foods often used for their taste, such as mushrooms and tomatoes have high
levels of naturally occurring glutamate. The human body also produces glutamate, whose role is essential in
the normal functioning of the body.
 Flavors and Taste 19

papillae present in this region of the tongue. We know in particular that fungiform papillae
are more sensitive to sodium salts while the filiform papillae are preferentially sensitive to
acids, and circumvallate to bitter compounds (Schiffman and Gatlin, 1993). Moreover, the
receptors of the taste buds have very different sensibilities. Indeed, the bitter receptors can
detect traces of bitter compounds, while acid receptors are much less sensitive and react to
higher levels of concentration. There is a kind of order of sensitivity of taste receptors: bitter
>>> acid >> sweet > salty. The taste receptors operate on the principle of cell membranes
and exploit the concentration gradients phenomena between the internal environment (gus-
tatory neurons directly connected to brain areas of taste through three cranial nerves: the
facial nerve, the glossopharyngeal nerve, and the trigeminal nerve) and the external envi-
ronment (the external surface of the tongue and especially the cells forming the taste buds).
The concentration difference between the two media varies over time and in terms of the
substances that are placed in the mouth. These changes in concentration will trigger the
release of neurotransmitters in the sensory dendrites of taste cells, causing a depolarization
of tastes neurons’ extremity. This electrochemical disturbance is the starting point of the
chain of taste perception. Neurobiologists have discovered that our taste buds are not lim-
ited to transmitting five flavors: sweet, salty, sour, bitter, and umami. We actually perceive
a taste continuum that results from many flavors, but we have few words to express their
diversity. Language does not allow us to describe all the different sensations from one per-
son to another; the sensitivity to taste varies considerably.
The rapid growth of research on the topic of flavor perception in recent years (e.g.,
Verhagen, 2007; Verhagen and Engelen, 2006) is linked to the idea that gaining a better
understanding of how the multisensory integration taking place in the context of food per-
ception might impact the theories of multisensory integration in general (e.g., Simons and
Noble, 2003). On the other hand, it is also widely believed that the study of the multisen-
sory processes involved in flavor perception will have a number of important consequences
for the food and beverage industries, such as, for example, a better understanding of the
processes used by people to assess the acceptability and flavor of new products. One robust
finding to have emerged from recent psychophysical research on flavor perception is that
odors can elicit changes in the perceived sweetness of foodstuffs (e.g., Stevenson et al.,
1999). One of the main consequences of a better understanding of the multisensory pro-
cesses involved in the flavor perception is the potential benefits to an international flavoring
company, as it can reduce the concentration of the typically more expensive aroma added
to a flavor by changing the amount of tastant (typically much cheaper) that is added, while
still keeping the flavor profile delivered to their customer constant.
Another area of intense commercial interest currently revolves around seeing whether
the consumer’s brain can, in some sense, be tricked into perceiving tastes/flavors with-
out the need to include all the unhealthy ingredients that so many of us seem to crave.
There are also some interesting commercial opportunities here around exploiting genetic
differences in taste perception. Some people have 16 times more gustatory receptors on
their tongues than others. In a very real sense, then, we may well live in different taste
worlds. While some of the most profound differences in taste perception involve certain
bitter-tasting compounds, recent research has demonstrated that “supertasters” are also
more sensitive to the oral-somatosensory attributes of foodstuffs (for example, to the
fat in a salad dressing), and possibly also to certain olfactory stimuli, while at the same
time being less influenced by visual cues when judging taste/flavor. Interestingly, Gary
Pickering and colleagues have just published a paper suggesting that wine experts, if not
“foodies,” tend to be more sensitive to certain bitter-tasting compounds than the rest of
the population (Pickering et al., 2013).
20 Food Aroma Evolution

The link between the perception of taste and behavior is also a topic of interest and
an important area of research. A fundamental question in this area is what the important
functions of taste for animals and particularly for humans are. Research has shown that
taste is involved at many levels in the development of mammals and humans. For food
intake, taste sensations influence our thinking, our deciding, and our behavior toward
food both consciously and unconsciously. The sensory experience of food is a determin-
ing factor in the control of food intake, often attributed to the positive hedonic response
associated with certain sensory signals. Sensory cues based on the sight, smell, taste,
and texture of a food are operational before, during, and after a meal (McCrickerd and
Forde, 2016). A number of studies on consumer preferences have demonstrated that taste
has a social sense. S. Højlund related in an interesting review that

By moving the attention of taste as a physiological stimulus-response of individuals to

tasting as a shared cultural activity, it is possible to recognize the taster as a reflexive
actor that communicates, performs, manipulates, senses, changes and embodies taste—
rather than passively perceives a certain experience of food.
(Højlund, 2015)

There is evidence that taste plays a role in social communication in humans, but is it a general
characteristic in mammals and vertebrates? Probably it is. For many vertebrates, physical,
social contacts, including the licking of nonvolatile social chemical compounds from the geni-
tals of a related animal, urine, sweat, or saliva, help to route compounds to the vomeronasal
organ of many species of vertebrates that respond to conspecific compounds of social com-
munication (Chamero et al., 2012). In the case of invertebrates, the social function of taste
has been demonstrated in male drosophila which use taste to differentiate males and females
as well as to recognize the maturity status of females for mating (Bray and Amrein, 2003;
Koganezawa et al., 2010). The honeybee is another example of an invertebrate where it is
known that smell and taste are of great importance in the social organization of the species
(Robinson, 1996). The world of the honeybee is populated by pheromones, and taste plays
a key role because their survival depends on the collection and consumption of nectar and
pollen, as well as other natural products (de Brito Sanchez et al., 2014). The honeybee and its
chemical means of communication is an active field of research because the insect is a good
model for the study of interactions between individuals via volatile (pheromones produced by
the queen) or nonvolatile (liquid substances secreted or deposited by the honeybees) chemical
compounds. Studies have shown that the honeybee is sensitive to a large number of organic
substances such as glucose, fructose, maltose, sucrose, sodium chloride, potassium chloride,
or lithium chloride. Concerning taste perception more specifically, it has been demonstrated
that the honeybee has gustatory molecular receptors not only at the level of the head but
also at the level of the antennae, and it is now known that these taste receptors are useful for
identification of the chemical signature of the colony (de Brito Sanchez, 2011; Erickson, 1982;
Whitehead and Larsen, 1976; Wright, 2009). To conclude this introductory chapter on taste
and its molecular perception (flavors) and the intimate link between the perception of taste
and the perception of smell (see Chapter 1), one can highlight the capital role played by the
olfaction and taste senses in the evolution of man and hominids at all levels (Breslin, 2013;
Breslin and Spector, 2008). Indeed, the role of these two types of sensory perception (in com-
bination with others like sight, hearing, and touch) is major and determining in the behaviors
of living species and therefore in their survival and adaptation to the environments in which
they evolve. Therefore, smell and taste perception also depend on social behaviors (ingestion
or non-ingestion of food—food safety), choice of food according to the associated metabolic
 Flavors and Taste 21

consequences (the search for the nutritional optimum), research and recognition of sexual
partners, organization, and management of the group (in insects such as the bee or in group-
living animals such as wolves, lions etc.). The multimodal nature of taste perception requires
more studies implying a combinatorial approach of our senses. Only when true multimodal
flavor experiments are explored will we begin to understand how the human brain forms the
supramodal sensation of flavor (Lundstrom et al., 2011). At the molecular level, fundamental
questions are about to be solved, such as how the odor, taste, and trigeminal perception are
formed and how are they collated to create the perception of the flavor. Others are not yet
answered, such as those relating to social taste assessments occurring during human interac-
tions; kissing, for example (Breslin, 2008).

GLOSSARY

Flavor: The sensory impression of food or other substances, determined primarily

by the chemical senses of taste and smell. The “trigeminal senses,” which
detect chemical irritants in the mouth and throat, as well as temperature and
texture, are also important to the overall gestalt of flavor perception.
Gestalt: The psychology of form or gestaltism is a psychological, philosophical, and
biological theory, according to which the processes of perception and mental
representation spontaneously treat phenomena as global forms, structured or
not, rather than as addition or juxtaposition of simple elements.
Perception: T he conscious or unconscious awareness of things through the physical senses
(smell, taste, touch, sight, hearing) that give rise to experience.
Taste: A perception that results from stimulation of a gustatory nerve. Taste belongs
to the chemical sensing system. Tasting begins when molecules stimulate
special cells in the mouth or throat. These special cells transmit messages
through nerves to the brain, where specific tastes are identified.

REFERENCES

Beare, J. I. (1906). Greek Theories of Elementary Cognition from Alcmaeon to Aristotle,

Clarendon Press, Oxford University.
Bray, S. and Amrein, H. (2003). A putative Drosophila pheromone receptor expressed in
male-specific taste neurons is required for efficient courtship. Neuron 39, 1019–29.
Breslin, P. A. S. (2008). Multi-modal sensory integration: Evaluating foods and mates.
Chemosensory Perception 1, 92.
Breslin, P. A. S. (2013). An evolutionary perspective on food and human taste. Current
Biology 23, R409–18.
Breslin, P. A. S. and Spector, A. C. (2008). Mammalian taste perception. Current Biology
18, R148–55.
Carterette, E. C. and Friedman, M. P. (1978). History of taste research. In Handbook
of Perception (E. C. Carterette and M. P. Friedman, eds.), pp. xi. Academic Press,
London.
Chamero, P., Leinders-Zufall, T., and Zufall, F. (2012). From genes to social communi-
cation: Molecular sensing by the vomeronasal organ. Trends in Neurosciences 35,
597–606.
de Brito Sanchez, M. G. (2011). Taste perception in honey bees. Chemical Senses 36,
675–92.
22 Food Aroma Evolution

de Brito Sanchez, M. G., Lorenzo, E., Su, S., Liu, F., Zhan, Y., and Giurfa, M. (2014). The
tarsal taste of honey bees: Behavioral and electrophysiological analyses. Frontiers in
Behavioral Neuroscience 8, 25.
Deppenweiler, A. (2014). Le glutamate monosodique comme ehauster de goût: Confiance
ou méfiance? Sciences pharmaceutiques. Available at: https://dumas.ccsd.cnrs.fr/
dumas-01011277/.
Erickson, E. H., Jr. (1982). Evidence for electrostatic enhancement of odor receptor func-
tion by worker honeybee antennae. Bioelectromagnetics 3, 413–20.
Gruner, O. C. (1973). A Treatise on the Canon of Medicine of Avicenna: Incorporating
a Translation of the First Book, AMS Press, New York, NY.
von Haller, A. and Cullen, W. (1803). First Lines of Physiology, pp. 215–21. O. Penniman
& Co.
Henning, H. (1915). Der Geruch I. Zeitschrift fur Psychologie und Physiologie der
Sinnesorgane 78, 161–267.
Højlund, S. (2015). Taste as a social sense: Rethinking taste as a cultural activity. Flavour
4, 6.
Koganezawa, M., Haba, D., Matsuo, T., and Yamamoto, D. (2010). The shaping of male
courtship posture by lateralized gustatory inputs to male-specific interneurons.
Current Biology 20, 1–8.
Lundstrom, J. N., Boesveldt, S., and Albrecht, J. (2011). Central processing of the chemi-
cal senses: An overview. ACS Chemical Neuroscience 2, 5–16.
McCrickerd, K. and Forde, C. G. (2016). Sensory influences on food intake control:
Moving beyond palatability. Obesity Reviews 17, 18–29.
Pickering, G. J., Jain, A. K., and Bezawada, R. (2013). Super-tasting gastronomes?
Taste phenotype characterization of foodies and wine experts. Food Quality and
Preference 28, 85–91.
Robinson, G. E. (1996). Chemical communication in honeybees. Science 271, 1824–5.
Schiffman, S. S. and Erickson, R. P. (1971). A psychophysical model for gustatory quality.
Physiology & Behavior 7, 617–33.
Schiffman, S. S. and Gatlin, C. A. (1993). Clinical physiology of taste and smell. Annual
Review of Nutrition 13, 405–36.
Siegel, R. E. (1970). IV. Organ and perception of taste. In Galen on Sense Perception. His
Doctrines, Observations and Experiments on Vision, Hearing, Smell, Touch and
Pain, and Their Historical Sources, pp. 158–73. Karger Publishers, Basel.
Simons, C. T. and Nobel, A. C. (2003). Challenges for the sensory sciences from food and
wine industries. Nature Reviews Neuroscience 4, 599–605.
Small, D. M. and Prescott, J. (2005). Odor/taste integration and the perception of flavor.
Experimental Brain Research 166, 345–57.
Stevenson, R. J., Prescott, J., and Boakes, R. A. (1999). Confusing tastes and smells: How
odors can influence the perception of sweet and sour tastes. Chemical Senses 24, 627–35.
Vicq D’Azyr, F. (1787). Encyclopédie Méthodique, Médecine (Panckoucke, ed.), Vol. 1,
p. 931. F. Vicq D’Azyr, Liège.
Verhagen, J. V. and Engelen, L. (2006). The neurocognitive bases of human food percep-
tion: Sensory integration. Neuroscience and Biobehavioral Reviews 30, 613–50.
Whitehead, A. T. and Larsen, J. R. (1976). Electrophysiological responses of galeal con-
tact chemoreceptors of Apis mellifera to selected sugars and electrolytes. Journal of
Insect Physiology 22, 1609–16.
Wright, G. A. (2009). The “sweet tooth” of the honeybee: The perception of nectar and
its influence on honeybee behaviour. SEB Experimental Biology Series 63, 183–204.
 Chapter 3
Chemical Senses and
Flavor Perception
Han-Seok Seo

CONTENTS

3.1 Introduction 23
3.2 Chemosensory System Associated with Flavor Perception 25
3.2.1 Gustatory System 25
3.2.1.1 The Sense of Taste 25
3.2.1.2 Anatomy and Physiology 27
3.2.2 Olfactory System 29
3.2.2.1 The Sense of Smell 29
3.2.2.2 Orthonasal and Retronasal Olfaction 30
3.2.2.3 Anatomy and Physiology 30
3.2.3 Oral Somatosensory System 32
3.2.3.1 Somesthesis/Chemesthesis 32
3.2.3.2 Anatomy and Physiology 32
3.3 Multisensory Flavor Perception 34
3.3.1 Interactions between Chemosensory Cues in Flavor Perception 34
3.3.1.1 Crossmodal Correspondence 34
3.3.1.2 Lateralization/Localization 34
3.3.1.3 Intensity and Pleasantness 35
3.3.2 Effects of Tactile or Temperature Cues of a Stimulus Medium
on Flavor Perception 37
3.3.3 Effects of Visual or Auditory Cues on Flavor Perception 40
3.4 Conclusion 41
References 41

3.1 INTRODUCTION

Consider this scenario: Hannah and two friends, James and Olivia, have dinner at a jazz
club restaurant in New Orleans, Louisiana. They experience perceptions and acceptances
that may vary dynamically over a span of consumption that starts at the point of ordering
meal items, perhaps even from just entering the restaurant, up to the point of completing
their entire meal. Ingredient descriptions and images of menu meal items lead Hannah,
James, and Olivia to all order the same item: spicy crispy chicken tenders. Subsequently,
when the chicken tenders are served, Hannah, James, and Olivia experience a variety of

23
24 Food Aroma Evolution

sensory cues that include appearance, aroma, taste, aromatics, irritations, mouthfeel,
and sounds, all originating from the spicy chicken tenders. While Hannah likes her spicy
crispy chicken tenders, James and Olivia do not enjoy their portions. Because Olivia suf-
fers from a stuffy nose caused by a bad cold, the chicken tenders taste too bland to her.
James is quite sensitive to spicy components, so he is unable to continue eating the spicy
chicken tenders because of pain and irritation in his mouth. Hannah suggests that James
try drinking a milk beverage because she had previously read an article suggesting that
such a beverage could be effective in decreasing residual spiciness (Samant et al., 2016).
When James tries eating the spicy chicken tenders with a milk beverage, he feels better
because it aids in reducing residual burning. After consuming their main dishes, they
decide to order strawberry mousse cake desserts served on white plates in the presence
of jazz music. These desserts taste sweeter and more flavorful than they had expected,
increasing their acceptance (see Spence and Shankar, 2010; Piqueras-Fiszman et al.,
2012; Fiegel et al., 2014; Spence, 2015a). This scenario illustrates that the perception of
flavor during meal consumption can be dynamic and complex. The perception of flavor
can be influenced by sensory characteristics of the meal items, as well as non-sensory fac-
tors such as socio-demographics, personality traits, emotional states, and surrounding
contexts (Delwiche, 2004; Spence, 2015a,b).
The term “flavor” has been conceptualized and used differently by various authors
depending on their areas of expertise (e.g., flavor chemists, food scientists, and culinary
experts) and their socio-demographic profiles (e.g., gender, age group, language, and
culture) (Delwiche, 2003). For example, in a survey where 140 professionals working in
various areas (agriculture, food science, sensory evaluation, and chemical senses) were
questioned about their concept of flavor, they differed with respect to which attributes
they rated as essential in contributing to the flavor of food. However, there was a consen-
sus that smell and taste are considered to be the most essential sensations involved in the
perception of flavor (Delwiche, 2003).
Amerine et al. (1965) defined flavor as “the sum of perceptions resulting from stimu-
lation of the sense ends that are grouped together at the entrance of the alimentary and
respiratory tracts.” Hall (1968) further specified flavor as “the sensation produced by
a material taken in the mouth, perceived principally by the senses of taste and smell,
and also by the general pain, tactile and temperature receptors in the mouth. Flavor
denotes the sum of the characteristics of the material which produce that sensation.”
More recently, the International Standards Organization (2008; ISO 5492) characterized
flavor as a “complex combination of the olfactory, gustatory and trigeminal sensations
perceived during tasting. The flavor may be influenced by tactile, thermal, painful and/or
kinesthetic effects.” Some researchers, however, have suggested that sensations derived
from other sensory modules (e.g., the senses of vision and hearing) should also be added
to the definition of flavor (Spence, 2015a). Furthermore, others have proposed that all
sensory inputs related to a particular food should be considered as flavor components
(Verhagen and Engelen, 2006; Spence, 2015a). In this chapter, only sensory modules
evoked from ingested substances in the mouth, including gustatory, olfactory, and oral
somatosensory (focusing on trigeminal) sensations, will be considered as primary compo-
nents of flavor. This chapter introduces concepts, roles in daily life, anatomy and physiol-
ogy, and other influential factors related to olfactory, gustatory, and oral somatosensory
(in particular, trigeminal) systems predominantly associated with a perception of flavor.
This chapter will also address multisensory flavor perception, with an emphasis on both
(1) interactions of chemosensory cues and (2) effects of non-chemosensory cues on che-
mosensory perception.
 Chemical Senses and Flavor Perception 25

3.2 CHEMOSENSORY SYSTEM ASSOCIATED

WITH FLAVOR PERCEPTION

3.2.1 Gustatory System

3.2.1.1 The Sense of Taste

Taste is composed of sensations elicited by tasting compounds through an anatomi-
cally and physiologically defined gustatory system (Bachmanov and Beauchamp, 2007).
Because other sensations such as olfaction and somatosensation, along with taste sensa-
tions, may have been evoked in the mouth during eating and drinking, many people use
the word “taste” to describe sensations arising from the oral cavity (Bachmanov and
Beauchamp, 2007; Seo and Hummel, 2011a). In this sense, people with olfactory disor-
ders often report that they have lost their sense of taste.
The sense of taste plays an important role in judging whether something may be ben-
eficial or dangerous for humans to eat or drink (Massler, 1980; Seo and Hummel, 2011a).
Generally speaking, a bitter taste is associated with sensing of diverse natural toxins,
and abnormal bitterness is likely to be equated with dietary danger (Drewnowski and
Gomez-Carneros, 2000). For example, aversive bitterness can be detected from rancid
fats, hydrolyzed proteins, microbial fermentation, and plant-derived alkaloids, although
all toxic species do not necessarily produce humanly detectable bitter-taste signals (e.g.,
Dioscorea dumetorum, also known as the bitter yam and found in Africa, is almost
tasteless) (Hladik and Simmen, 1996). Even though there is no general rule relating con-
centrations of bitter-tasting compounds and their toxicities, bitter-tasting compounds are
detected by humans at much lower levels than are other tasting compounds (Hladik and
Simmen, 1996; Drewnowski and Gomez-Carneros, 2000).
The current consensus on taste quality is that human taste sensation can be clas-
sified into five basic taste qualities: sweet, salty, sour, bitter, and umami (or savory).
Human neonates exhibit differential facial expressions in response to qualities such as
sweet, salty, sour, and bitter tastes (Steiner, 1974; Rosenstein and Oster, 1988; Forestell
and Mennella, 2017). The sweet taste is related to signals reflecting the presence of
energy and nutrients in foods and beverages. Salty taste, mainly detected from sodium
salts, is thought to be attractive to people’s diets for electrolyte balance (Bachmanov
and Beauchamp, 2007). Sour tastes, along with bitter tastes, may be associated with sig-
nals related to spoiled substances (Bachmanov and Beauchamp, 2007). Like an infant’s
dislike for sour-tasting items, initial aversive responses may tend to change to positive
responses over time (Bossfeld et al., 2007; Forestell and Mennella, 2017). Finally, the
umami taste commonly detected from l-glutamate is associated with signals reflecting
the presence of protein (Bachmanov and Beauchamp, 2007). As shown in Figure 3.1,
basic taste qualities can interact with one another, leading to enhancement, suppres-
sion, or no effect on perceived intensity (Keast and Breslin, 2003). It should be noted
that binary taste interactions are compound-specific as well as intensity/concentration
specific (Keast and Breslin, 2003).
There is also growing interest in other taste qualities (i.e., the existence of a sixth
basic taste). These include a “fat taste” (Fukuwatari et al., 1997; Khan and Besnard,
2009; Mattes, 2009a,b; Keast and Costanzo, 2015) and a “starchy taste” (Sclafani,
2004; Lapis et al., 2014, 2016). CD-36 and G protein-coupled receptor 120 are consid-
ered to potentially allow humans to be able to detect free fatty acids varying in chain
length (C6 –C18) and saturation at low concentrations (Mattes, 2009a,b). However, it still
26 Food Aroma Evolution

FIGURE 3.1 Schematic review of binary taste interactions at (a) low intensity/concentra-
tion, (b) medium intensity/concentration, and (c) high intensity/concentration levels of taste
qualities. (Reprinted from Food Quality and Preference, 14(2), Keast and Breslin, “An over-
view of binary taste-taste interactions,” pp. 111–24, 2003, with permission from Elsevier.)

remains unclear as to whether there is a recognizable perception of fat independent of

other taste qualities (Keast and Costanzo, 2015). Recent studies have also demonstrated
that humans can detect glucose oligomers (average degree of polymerization: 7 and 14),
but not glucose polymers (average degree of polymerization: 44) (Lapis et al., 2016). The
detection of glucose oligomers seems to be independent of the sweet taste receptor, that
is, hT1R2/hT1R3 (Lapis et al., 2014, 2016). If there is an independent mechanism apart
from the sweet taste for glucose oligomers, “starchy taste” would be the most likely to be
associated with identifying a source of energy (Lapis et al., 2016). Hartley et al. (2019)
suggested the following comprehensive list of criteria for determining the appropriateness
of other new tastes:

1. A unique class of effective substances must exist.

2. Detection of effective substances must have an evolutionary perspective (e.g., the
body’s electrolyte balance for salty taste).
3. Unique receptors and neural transmission of the effective substances must exist.
4. Neurotransmission of electrical signals to taste processing regions of the brain
must occur.
5. The perceptual quality of the effective substances must be independent from
other taste qualities.
 Chemical Senses and Flavor Perception 27

6. Hedonic responses to the effective substances must exist.

7. Physiological and/or behavioral responses to the effective substances must exist.

It has been found that 4 to 10% of patients consulting a specialized smell and taste clinic
in the United States and Germany report impairment in their sense of taste (Deems et al.,
1991). Among the general population, the percentage of people with “hypogeusia” (i.e.,
gustatory hyposensitivity compared to young, healthy subjects) is estimated to be about
5%. People with “ageusia” (i.e., complete or pronounced reduction of the sense of taste
or a loss of sensitivity in a single taste quality) are considered very rare (Welge-Lüssen et
al., 2011; Fark et al., 2013). With respect to qualitative taste disorder, “parageusia” (i.e.,
inadequate or wrong taste sensation elicited by a taste stimulus) and “phantogeusia” (i.e.,
perception of taste in the absence of a taste stimulus) have been reported and often occur
together (Fark et al., 2013). Although the main reasons for taste disorders include cra-
niocerebral injury, infection of the upper respiratory tract, iatrogenic causes, side effects
of medication, exposure to toxic substances, and burning mouth syndrome (Hummel et
al., 2011; Fark et al., 2013), approximately 30% of taste disorder cases state no specific
cause (Fark et al., 2013).

3.2.1.2 Anatomy and Physiology

Each of the five basic taste qualities is mediated by distinct transduction pathways
expressed in specific subsets of taste receptor cells (Bachmanov and Beauchamp, 2007).
Sweet and umami tastes are specifically mediated by a family of three class-C G protein-
coupled receptors (GPCRs): T1R1, T1R2, and T1R3. Although T1R3 alone serves as a
taste receptor for sucrose and other sugars (Nelson et al., 2001; Zhao et al., 2003), the
heterodimers of T1R2 and T1R3 serve as sweet taste receptors for a variety of sweet-
tasting substances (Breslin and Huang, 2006; Temussi, 2009). A heterodimeric complex
of T1R1 and T1R3 also serves as a taste receptor for umami-tasting substances (Zhao
et al., 2003). Full-length mGluR1 and mGluR4 (Toyono et al., 2002) and a variant of
mGluR1 (i.e., taste-mGluR1, San Gabriel et al., 2009) are also considered candidates
for umami taste receptors (Yasumatsu et al., 2012). Bitter-tasting substances are bound
to the T2R family, a large family of class A GPCRs (Adler et al., 2000; Chandrashekar
et al., 2006). Salty-tasting (sodium) and sour-tasting (protons) substances are mediated
by specific ion channels (DeSimone and Lyall, 2006). With respect to the salty taste,
the selective epithelial amiloride-sensitive sodium channel (ENaC) is considered to be
involved, at least in rodents (Bachmanov and Beauchamp, 2007). Finally, multiple can-
didates have been found to mediate responses to sour-tasting substances. These include
acid-sensing ion channels (ASICS; Ugawa et al., 1998), neuronal amiloride-sensitive cat-
ion channel 1 (ACCN1), HCN1, and HCN4 from a family of hyperpolarization-activated
cyclic nucleotide-gated channels (HCNs; Stevens et al., 2001), transient receptor poten-
tial channels PKD1L3 and PKD2L1 (Huang et al., 2006; Ishimaru et al., 2006), and
TASK-1, and Na+-H+-exchanger isoform 1 (NHE-1) (Bachmanov and Beauchamp, 2007).
Taste receptor cells are mainly found in garlic- or rosebud-shaped multicellular clus-
ters called “taste buds.” Humans normally have between 5000 and 10,000 taste buds
(Loper et al., 2015). Taste buds have been observed not only on the tongue’s surface, but
also on the soft palate, the pharyngeal and laryngeal regions of the throat, the stom-
ach, and the gastrointestinal tract (Breslin and Huang, 2006; Rozengurt, 2006; Trivedi,
2012a; Rober and Chaudhari, 2017). Each taste bud contains a small opening, a “taste
pore,” on the upper surface of the taste papillae (Just et al., 2005). Through this taste
pore, substances to be tasted come into contact with taste receptor cells (Breslin and
28 Food Aroma Evolution

Huang, 2006; Seo and Hummel, 2011a). There are four principal types of cells, types
I, II, III, and IV, within a taste bud (Feng et al., 2014). They can be classified by dis-
tinct ultrastructural features and immuno-histochemical characteristics (Kataoka et al.,
2008). Type I (dark) cells with several long microvilli seem to have supportive or glia-like
functions in taste buds; they express enzymes and transporters required to remove extra-
cellular neurotransmitters (e.g., the neurotransmitter adenosine 5´-triphosphate (ATP))
(Bartel et al., 2006; Roper and Chaudhari, 2017). A subset of type I cells is considered
to be involved in salty-taste reception (Vandenbeuch et al., 2008; Feng et al., 2014). Type
II (light) cells, polarized cells with short microvilli, function as taste receptors for sweet,
bitter, umami, and possibly salty-tasting substances (Trivedi, 2012b). Most type II cells
express one class of taste GPCRs, that is, T1R or T2R, that respond to only one taste
quality (Roper and Chaudhari, 2017). However, since T1R1, T1R2, and T1R3 are often
co-expressed in taste bud cells, each taste bud responds to multiple tasting substances
(Roper and Chaudhari, 2017). Type III (dark) cells, thin spindle-shaped cells with single
microvillus, form synapses with gustatory nerve endings (Chaudhari and Roper, 2010;
Feng et al., 2014). Their subsets are considered to be involved in salty taste (Kataoka
et al., 2008; Oka et al., 2013; Feng et al., 2014) and/or sour taste perception (Huang
et al., 2006; Trivedi, 2012b). Type III cells are also likely to serve in integrating and
transmitting signals from type II cells to gustatory nerves (Tomchik et al., 2007; Feng
et al., 2014). Finally, type IV cells (also called basal cells), located near taste bud bases,
are newly generated taste precursor cells differentiating into mature taste cells (Trivedi,
2012b; Feng et al., 2014).
There are four types of taste papillae: fungiform, circumvallate, foliate, and filiform.
Fungiform (“mushroom-like”) papillae are distributed both on the anterior two-thirds
(Konstantindis, 2009) and on the side (Kullaa-Mikkonen et al., 1987) of the tongue.
Their densities are related to taste-intensity perception (Miller and Reedy, 1990) or
6-n-propylthiouracil (PROP) status (Eldeghaidy et al., 2018). For example, people with a
higher density of fungiform papillae may perceive some tastes as more intense than those
with a lower density of fungiform papillae (Miller and Reedy, 1990). However, little or
no association between fungiform papillae density and perceived taste intensity has been
observed (Feeney and Hayes, 2014; see also Dinnella et al., 2018). Women have been
found to possess more fungiform papillae than men (Bartoshuk et al., 1994; Dinnella
et al., 2018). Fungiform papillae density increases from childhood to adulthood (Correa
et al., 2013) then decreases with age (Dinnella et al., 2018). While circumvallate (“wall-
like”) papillae are present at the posterior third of the tongue, foliate (“leaf-like”) papillae
are positioned along its posterior lateral edges (Konstantindis, 2009). Finally, filiform
(“thread-like”) papillae with no taste buds are found on the tongue surface. They seem to
be involved in a somatosensory function, manipulation of food bolus, and/or saliva dis-
tribution on the tongue and food bolus (Konstantindis, 2009; Seo and Hummel, 2011a).
Interestingly, salivary secretion, flow, and profiles have been found to affect taste func-
tions, such as taste sensitivity (Christensen et al., 1987; Mese and Matsuo, 2007; Stolle
et al., 2017).
Three primary cranial nerves (CNs), facial (CN VII), glossopharyngeal (CN IX), and
vagus (CN X), are involved in the transmission and processing of taste signals to the gus-
tatory cortex. The facial nerve delivers taste signals via the chorda tympani nerve and the
greater superficial petrosal nerve (Spector, 2000; Breslin and Huang, 2006); the chorda
tympani nerve innervates the anterior two-thirds of the tongue and the greater superficial
petrosal nerve innervates taste buds on the soft palate (Konstantindis, 2009). The glos-
sopharyngeal nerve innervates the majority of the circumvallate papillae of the posterior
 Chemical Senses and Flavor Perception 29

tongue via the lingual-tonsillar branch (Spector, 2000; Konstantindis, 2009). Finally, the
vagus nerve innervates taste buds located on the laryngeal surface of the epiglottis, the
larynx, and the proximal part of the esophagus via the superior laryngeal nerve (Spector,
2000; Konstantindis, 2009).
The axons of gustatory neurons terminate in the rostral part of the nucleus of the sol-
itary tract in the medulla (Beckstead and Norgren, 1979). These neurons project into the
parvicelluar part of the posteromedial ventral thalamus through the central tegmental
tract (Huart et al., 2009; Seo and Hummel, 2011a; Iannilli and Gudziol, 2019). Neurons
from the thalamus project into the primary taste cortex in the frontal operculum and
adjoining insula areas (Huart et al., 2009) involved in taste intensity, identification, and
memory (Small, 2006; Iannilli and Gudziol, 2019). Each half of the human tongue was
found to be innervated by ipsilaterally ascending cranial nerves. However, the pathway
of taste laterality from the oral cavity to the primary taste cortex in humans remains
unclear (for a review, Iannilli and Gudziol, 2019). The primary taste cortex projects into
the secondary taste cortex. It includes the caudolateral orbitofrontal cortex, the cingulate
gyrus, the amygdala, the hypothalamus, and the basal ganglia (Sewards, 2004; Breslin
and Huang, 2006; Small, 2006; Huart et al., 2009; Seo and Hummel, 2011a). Detection
and suprathreshold intensity-related perception of gustatory stimuli are associated with
processing in the operculum/insula, while affective response is likely to be related to pro-
cessing in the orbitofrontal cortex (Small, 2006).

3.2.2 Olfactory System

3.2.2.1 The Sense of Smell

The sense of smell plays a more important role than commonly thought in a broader
range of daily tasks. First, the sense of smell serves as a detector of hazardous substances
(e.g., spoiled foods, poisonous fumes, gas leaks, etc.). Second, the sense of smell helps us
detect (Porter et al., 2007) and identify edible food sources (Fallon and Rozin, 1983). It
also influences food acceptability (Aschenbrenner et al., 2008), food preparation (Seo
and Hummel, 2009), eating behavior (Aschenbrenner et al., 2008), and nutritional status
(Duffy et al., 1995), although such effects have not been observed in all studies (Mattes,
2002). Finally, the sense of smell can serve as a social communicator (Stevenson, 2010)
by influencing reproductive behavior such as inbreeding avoidance, fitness detection in
prospective mates (Stevenson, 2010; Pause, 2016), and communication of emotion via
body odors (Zhou and Chen, 2009).
Prevalence of olfactory disorder in the general population is common, but most indi-
viduals are unaware of olfactory loss (Nordin et al., 2004; Croy et al., 2014). About 20%
of the general population have reported at least one type of olfactory disorder (Croy et al.,
2014), although the prevalence rate of such disorders varies in earlier studies (Nordin et
al., 2004; Croy et al., 2014; Noel et al., 2017). Olfactory disorders can be classified into
two major groups, quantitative and qualitative. A quantitative olfactory disorder may
include (1) “anosmia” (lack of ability to perceive odors), (2) “congenital anosmia” (condi-
tion in which people are born with an inability to perceive odors), (3) “specific anosmia”
(inability to perceive specific odor(s) due to the lack of certain olfactory receptors), (4)
“functional anosmia” (significantly reduced ability to perceive odors), (5) “hyposmia”
(reduced ability to perceive odors), and (6) “hyperosmia” (heightened ability to perceive
odors), based on the level of olfactory performance that can be quantified through clini-
cal testing (Hummel et al., 2016). It is estimated that the prevalence rate of congenital
30 Food Aroma Evolution

anosmia is one individual out of every 5000 to 10,000 (Croy et al., 2012). A qualitative
olfactory disorder includes (1) “parosmia” (also called “troposmia”; inadequate or wrong
perception of odor stimulus) and (2) “phantosmia” (perception of smell in the absence of
an odor stimulus) (Hummel et al., 2016). Both parosmic and phantosmic sensations are
typically described as distorted or unpleasant. For example, odors are often described
as “burned,” “rotten,” “chemical,” or “fecal” smells (Leopold, 2002; Frasnelli et al.,
2004; Hummel et al., 2016). In the general population, the prevalence rates of parosmia
and phantosmia have been reported as less than 4% (Nordin et al., 2007) and 0.8–2.1%
(Landis et al., 2004), respectively. The most common etiologies of olfactory disorder
are post-viral upper respiratory infection (18–45% of the clinical population) and nasal/
sinus disease (7–56%), followed by head trauma (8–20%) and exposure to toxins/drugs
(2–6%) (Nordin and Bramerson, 2008; Croy et al., 2014).
Olfactory disorders can significantly affect daily living activities and quality of life
(Miwa et al., 2001; Santos et al., 2004; Seo et al., 2009; Nordin et al., 2011; Croy et al.,
2014). Miwa et al. (2001) reported (1) safety-related activities such as reduced ability
to detect spoiled food (75% of 1407 clinical patients with olfactory disorder), gas leaks
(61%), or smoke (50%), and (2) eating-related activities such as eating (53%) or cook-
ing (49%) as the most frequently cited daily activities impaired by olfactory disorder.
Similarly, Santos et al. (2004) reported that cooking-related incidents (45% of 445 clini-
cal patients with olfactory disorder) and ingestion of spoiled food (25%) are the most
common hazardous events associated with patients’ olfactory disorders. The incidence
of hazardous events increases with the degree of olfactory disorder (Pence et al., 2014).

3.2.2.2 Orthonasal and Retronasal Olfaction

People perceive volatile aromatic compounds via two different pathways. First, volatile
aromatic compounds can be perceived via the external nares during sniffing or nasal
inhalation. This is referred to as “orthonasal olfaction,” providing external information
associated with a food source, edibility, toxicity/danger, or social communication (Rozin,
1982; Shepherd, 2006; Seo and Hummel, 2011a). The other pathway is the mouth (oral
cavity or nasopharynx) via which volatile aromatic compounds can be perceived dur-
ing eating, drinking, or exhalation, a condition referred to as “retronasal olfaction.”
It provides information about what is consumed in the mouth (Rozin, 1982; Shepherd,
2006; Seo and Hummel, 2011a; Goldberg et al., 2018). Retronasal odors from the food
or beverage matrix are typically released from the mouth during consumption (Shepherd,
2006). Since they are localized to the mouth (Lim and Johnson, 2012), people appear to
frequently confuse retronasal stimulation with taste (Seo and Hummel, 2011a); that is,
they experience “taste–smell confusion” (Murphy et al., 1977; Murphy and Cain, 1980;
Rozin, 1982).

3.2.2.3 Anatomy and Physiology

Through either orthonasal or retronasal pathways, volatile aromatic compounds of sub-
stances reach the olfactory epithelium, a layered structure residing in the upper part of
the nasal cavity. More specifically, it resides bilaterally within the olfactory cleft and
extends into the superior turbinate and superior part of the middle turbinate (Rawson et
al., 1997; Rawon and Yee, 2006; Seo and Hummel, 2011a). The olfactory epithelium is
composed of different cell types, including Bowman’s gland cells, horizontal basal cells,
globose basal cells, both immature and mature olfactory neurons, and sustentacular cells
(Lavoie et al., 2017). Basal cells are capable of proliferating and differentiating into either
neural or non-neural cells throughout adulthood. Both mature and immature olfactory
 Chemical Senses and Flavor Perception 31

neurons are positioned in the intermediate layer of the olfactory epithelium (Lavoie et
al., 2017). The apical layer also contains sustentacular cells and sensory cilia that can be
projected from the dendrites of olfactory neurons (Lavoie et al., 2017).
When volatile aromatic compounds bind to the olfactory receptors on the sensory
cilia, the membrane-bound protein structure changes, allowing extracellular calcium
ions to enter a cell (Hornung, 2006). It then creates a generator potential that produces
an electronic signal flowing ipsilaterally from the axons of the olfactory neurons to the
olfactory bulb located in the anterior cranial fossa, above the cribriform plate of the eth-
moid (Hornung, 2006; Huart et al., 2009).
The olfactory bulb is the first relay station in the olfactory system (Freiherr, 2016). It
exhibits relatively large individual size variation, ranging from 37 to 98 mm3 with respect
to its volume in healthy adults (Buschhüter et al., 2008). The volume of the olfactory
bulb, larger in males than in females (Buschhüter et al., 2008), has been found to be cor-
related with olfactory function both in healthy people (Buschhüter et al., 2008; Hummel
et al., 2013) and people with olfactory disorder (Haehner et al., 2008; Rombaux et al.,
2010). The olfactory bulb has sphere-shaped glomeruli that contain synapses between
olfactory receptor neurons and dendrites of the mitral cells (Huart et al., 2009), relaying
olfactory information to second-order neurons, mitral and tufted cells (Imai, 2014). The
mitral and tufted cells are modulated by intrabulbar circuits and centrifugal inputs, with
the neurons generating a unique odor code (Imai, 2014).
From the olfactory bulb, olfactory information is transmitted via the lateral olfactory
tract to other cortical olfactory structures. These include the piriform cortex, the entorhi-
nal cortex, the amygdala, and periamygdaloid cortex, the olfactory tubercle, and the ante-
rior olfactory nucleus (Freiherr, 2016). Although the anterior and posterior sub-regions of
the piriform cortex appear to be histologically identical, they provide different functions
(Gottfried, 2006). The anterior part of the piriform cortex is associated with initial neural
representation of an odorant and encoding of its molecular features (Davison and Ehlers,
2011). The posterior or temporal part of the piriform cortex is related to perceptual infor-
mation related to odor quality and categorization (Gottfried et al., 2006; Howard et al.,
2009; Freiherr, 2016). The entorhinal cortex is considered a gateway to the hippocampus
responsible for memory processes (Insausti et al., 2002; Freiherr, 2016). The amygdala
and periamygdaloid cortex are considered responsible for cognitive evaluation of olfac-
tory information (Freiherr, 2016) and intensity coding of odors with emotional salience
(Winston et al., 2005; Grabenhorst et al., 2007; Freiherr, 2016).
Olfactory information further projects into brain areas responsible for cognitive odor
processing and perception. These include the orbitofrontal cortex, the insular, the hippo-
campus, the thalamus, the hypothalamus, the cingulate cortex, the ventral striatum, and
the cerebellum (Freiherr, 2016). While olfactory bulb volume is related to odor identifica-
tion performance (Buschhüter et al., 2008), the gray matter volume of the orbitofrontal
cortex (OFC) is associated with odor threshold and odor discrimination performances
(Seubert et al., 2013; Freiherr, 2016). The insular has been found to be involved in an inte-
gration of flavor perception, a perception of unpleasant olfactory or intranasal trigemi-
nal stimuli (Albrecht et al., 2010), and a multisensory integration of negatively valenced
stimuli (e.g., unpleasant odor and disgusted face) (Seubert et al., 2010; Freiherr, 2016).
The hippocampus is responsible for emotional and memory-related processing of olfac-
tory stimuli and multisensory integration of olfactory inputs (Freiherr, 2016). Olfactory
information is transmitted to cortical areas without a thalamic relay, but there is an indi-
rect pathway of olfactory processing through the mediodorsal thalamus to the neocortex
(Freiherr, 2016). The mediodorsal thalamic nucleus receives direct inputs from primary
32 Food Aroma Evolution

olfactory areas that include the piriform cortex, the enthorhinal cortex, and the amyg-
dala, and has reciprocal connections with the OFC (Courtiol and Wilson, 2014, 2015). It
is considered to be a higher-order olfactory thalamus (Freiherr, 2016). The mediodorsal
thalamic nucleus appears to be related to olfactory functions, including odor perception
and discrimination, attention to odors, and coding of the hedonic valence of odor stimuli
(Courtiol and Wilson, 2015).

3.2.3 Oral Somatosensory System

3.2.3.1 Somesthesis/Chemesthesis
During eating or drinking, people often perceive various kinds of physical or chemical sen-
sations such as “softness,” “roughness,” “stickiness,” “irritation,” “burning,” “stinging,”
“cooling,” or “tickling,” along with olfactory and gustatory sensations in the mouth. These
other sensations, neither olfactory nor gustatory, are characterized as “oral somesthesis.”
They provide information about the physicochemical constituents and characteristics of
substances taken into the mouth (Lim, 2016). In particular, chemosensory sensations,
neither olfactory nor gustatory, were described as “common chemical senses” by George
H. Parker in 1912. He reported that animals have three distinct types of chemosensory
receptors: olfactory, gustatory, and common chemical (Slack, 2016). However, the term
itself and the concept of “common chemical senses” were actively debated (Green, 2016).
The term “chemesthesis” was introduced to define the chemical sensibility of the skin and
mucous membranes as senses other than chemical (Green et al., 1990; Green and Lawless,
1991; Green, 2016). The term chemesthesis refers to any somatosensory response to an
irritant or a noxious chemical (Slack, 2016). Chemesthetic sensations have been found to
be mediated via thermoreceptors, nociceptors, and mechanoreceptors of the somatosen-
sory nerves (Green, 1996, 2016; Lim, 2016). Those evoked within the nose and the mouth
are primarily mediated via the trigeminal nerve (CN V) that is innervated in the nasal
mucosa and the anterior regions of the oral cavity. Thus, chemicals that evoke sensations
other than olfactory or gustatory sensations are often described as “trigeminal stimuli”
(Green, 2016). The glossopharyngeal and vagal nerves of the somatosensory nerves are
involved in chemesthetic sensations in response to irritants present in the posterior regions
of the oral cavity (Green, 1996; Rentmeister-Bryant and Green, 1997; Slack, 2016).

3.2.3.2 Anatomy and Physiology

Somatosensory nerves contain three types of somesthesis-related receptors: mechanore-
ceptors, thermoreceptors, and nociceptors (Vallbo et al., 1979). Oral mechanoreceptors
responsible for sensory perceptions of pressure, slip, vibration, and movements occurring
in the mouth play an important role in the safe manipulation of food bolus (Engelen,
2012). More specifically, orofacial mechanoreceptors are involved in multiple activities
that include (1) position and movement of the tongue, (2) manipulation of food bolus
suitable for chewing and swallowing, (3) prevention of biting of the tongue and cheek
during oral processing, and (4) swallowing without chocking (Engelen, 2012). In orofa-
cial areas, there are three types of receptors: (1) slow-adapting (SA) type I receptors (SA
I) that end in Merkel cells, (2) slow-adapting type II receptors (SA II) that end in Ruffling
corpuscles, and (3) fast-adapting (FA) type I afferents (FA I) that end in Meissner cor-
puscles (Engelen, 2012); however, no fast-adapting type II afferents (FA II) that end in
the Pacinian corpuscles have been found in orofacial areas (Trulsson and Essick, 2010;
Engelen, 2012). In the mouth, SA I receptors with a high degree of spatial resolution are
 Chemical Senses and Flavor Perception 33

associated with texture and shape perceptions of food bolus in the processes of chewing
and manipulation (Engelen, 2012). SA II receptors in the mouth are associated with size
perception of food bolus as well as shape and position perceptions of the tongue (Engelen,
2012). While FA I afferents seem to be involved in the perception of vibratory sensation in
the mouth (Engelen, 2012), further research is needed to clarify their roles in the mouth.
Proprioceptors in the face and periodontal mechanoreceptors have also been found to
play a crucial role in the perception of mechanical characteristics of food bolus during
eating (Engelen, 2012; Higaki et al., 2014).
Nociception, the sensation of pain, is mediated by free nerve endings (nocicep-
tors). Myelinated Aδ mechanical and thermal fibers mediate fast and sharp pain stimuli.
Unmyelinated C-fibers transmit slow and dull pain (Silberstein, 2003; Engelen, 2012).
Because nociceptors are polymodal neurons, they are involved in a variety of mechani-
cal, chemical, and thermal stimuli (Engelen, 2012) as well as auditory and visual stimuli
(Mickle et al., 2016).
Cutaneous thermal information is processed via multiple classes of afferent nerve
fibers (Schepers and Ringkamp, 2010). While warm stimuli are processed by warm
fibers (unmyelinated C-fibers) at an average rate of 1.0 m/s (LaMotte and Campbell,
1978; Darian-Smith et al., 1979), cold stimuli are thought to be processed by either
thinly-myelinated Aδ fiber conducting at 9 to 15 m/s or unmyelinated C-fibers (Schepers
and Ringkamp, 2010; Engelen, 2012). The activity of several transient receptor poten-
tial (TRP) ion channels has been found to depend on surrounding temperatures ranging
from noxious cold to noxious heat (Voets et al., 2004). In particular, among 28 different
TRP channels in humans, three TRP families, that is, TRP vanilloid channels (TRPV),
ankyrin transmembrane protein channels (TRPA), and melastatin or long TRP channels
(TRPM), seem to be temperature-sensitive (Schepers and Ringkamp, 2010). The TRPV1
channel is activated not only by noxious heat above 43°C, but also by multiple chemical
stimuli such as capsaicin (hot peppers), eugenol (cloves), gingerol (ginger), allicin (gar-
lic), and acids, evoking perceptions of tingling, sharpness, and burning (Caterina et al.,
1997; Frasnelli and Manescu, 2016). The TRPV3 channel is activated by noxious warm
stimuli above 39°C and chemical stimuli such as monoterpenoid phenol present in high
concentrations in the essential oils of oregano, thymol (thyme), eugenol (cloves), and car-
veol (Viana, 2011; Frasnelli and Manescu, 2016). In contrast, the TRPA1 and TRPM8
channels are activated by cold stimuli. The TRPA1 channels are activated by noxious cold
below 17°C, as well as allyl thioisocyanate (mustard), cinnamaldehyde, allicin, and gin-
gerol (Viana, 2011; Frasnelli and Manescu, 2016). Volatile organic compounds present in
cigarette smoke and smog, triggering eye irritation, coughing, and mucous secretion, are
also TRPA1 agonists (Viana, 2011). The TRPM8 channels are activated by cold stimuli
between 8 and 25°C, as well as natural and synthetic cooling stimuli such as menthol and
eucalyptol (Viana, 2011; Engelen, 2012).
The trigeminal nerve (CN V), the largest cranial nerve, has three major branches:
the ophthalmic nerve (CN V1), the maxillary nerve (CN V2), and the mandibular nerve
(CN V3) (Doty and Cometto-Muñiz, 2003). The ophthalmic nerve branch is involved in
sensory inputs from the upper part of the head, the forehead, the upper eyelid, the nasal
mucosa, and the tip of the nose (Huart et al., 2009; Bathla and Hegde, 2013). The maxil-
lary nerve branch is associated with sensory inputs originating from the middle third of
the head, the lower eyelid, the cheek, the nares, the upper lip, and the upper teeth (Huart
et al., 2009; Bathla and Hegde, 2013). Finally, the mandibular nerve carries both sensory
and motor information coming from the lower third of the head, the chin, the lower lip,
the lower teeth, and the jaw (Huart et al., 2009; Seo and Hummel, 2011a; Bathla and
34 Food Aroma Evolution

Hegde, 2013). These three branches converge on the trigeminal ganglion (also referred to
as Gasserian Ganglion) located within the Meckel’s cave (Huart et al., 2009; Bathla and
Hegde, 2013). The axons of the first-order neurons from the trigeminal ganglion enter
the brainstem at the level of the pons. Within the brainstem, fibers segregate into three
different sensory nuclei: the principal sensory nucleus, the mesencephalic nucleus, and the
spinal trigeminal nucleus (Walker, 1990; Bathla and Hegde, 2013). The principal sensory
nucleus, positioned in the pontine tegmentum, mediates tactile sensations, in particular
pressure and light touch from V1–V3, while the mesencephalic nucleus, located at the
junction of pons and mid-brain, mediates proprioceptive sensations from V3 (e.g., mas-
ticatory muscles, teeth, hard palate, periodontium, and the temporomandibular joint)
(Walker, 1990; Bathla and Hegde, 2013). The spinal trigeminal nucleus, extended from
the pontomedullary junction to the upper cervical cord, mediates pain and temperature
sensations from V1–V3 (Walker, 1990; Doty and Cometto-Muñiz, 2003; Huart et al.,
2009; Bathla and Hegde, 2013). The second-order neurons transmit to the contralateral
side and ascend through the trigemino-thalamic tract toward the ventro-postero-medial
nucleus of the thalamus (Huart et al., 2009). Via the posterior arm of the internal cap-
sule, the third-order neurons project from the thalamus to the primary somatosensory
cortex, the brain area responsible for somatosensation (Huart et al., 2009; Frasnelli and
Manescu, 2016). Other important processing areas include the secondary somatosen-
sory cortex, the amygdala, and the hippocampus (Huart et al., 2009; Seo and Hummel,
2011a; Han et al., 2018).

3.3 MULTISENSORY FLAVOR PERCEPTION

3.3.1 Interactions between Chemosensory Cues in Flavor Perception

3.3.1.1 Crossmodal Correspondence
Crossmodal correspondence, characterized as “a compatibility effect between attributes
or dimensions of a stimulus (i.e., an object or event) in different sensory modalities (be
they redundant or not)” (Spence, 2011), is innate and developed by perceptual learn-
ing (Gilbert et al., 1996; Schifferstein and Tanudjaja, 2004; Spence, 2011). Crossmodal
correspondence has been found to exist between chemosensory cues. People often con-
fuse retronasal stimulation with taste (Murphy et al., 1977; Murphy and Cain, 1980;
Rozin, 1982) because taste cues are often perceived with retronasal odors during eating
and drinking. Based on frequent experiences of smell and taste co-occurrence in the
mouth, people are accustomed to matching certain orthonasal or retronasal odors with
specific taste qualities (Stevenson and Boakes, 2004). For example, while aromas/flavors
of strawberry, vanilla, or caramel are likely to be matched with a sweet taste, aromas/
flavors of lemon or citrus fruits are often paired with a sour taste. Crossmodal correspon-
dences between olfactory and trigeminal stimuli have also been reported. Bensafi et al.
(2013), for example, showed that participants matched intranasal carbon dioxide (CO2)
with orange odor, but not with rose odor, probably due to their previous experience with
a mixture of orange odor and carbon dioxide in soft drinks.

3.3.1.2 Lateralization/Localization
Humans can lateralize trigeminal stimuli, such as carbon dioxide with a high degree of
accuracy (Kobal et al., 1989; Kleemann et al., 2009). However, it seems to be difficult
for humans to lateralize pure odor stimuli without accompanying trigeminal stimulation
 Chemical Senses and Flavor Perception 35

when they are presented only to the left or the right nostril (Schneider and Schmidt, 1967;
Kobal et al., 1989; Radil and Wysocki, 1998; Frasnelli et al., 2009; Kleemann et al., 2009;
Tremblay and Frasnelli, 2018). Although contradictory findings have also been reported
(Negoias et al., 2013), olfactory lateralization seems to occur only when an odor stimulus
triggers the trigeminal somatosensory system. In a study conducted by Kleemann et al.
(2009), participants were unable to localize a pure odor stimulus (hydrogen sulfide, H 2S),
stimulating the olfactory system only at low concentrations. They were able, however,
to localize the odor stimulus accompanying trigeminal stimulation (isoamyl acetate) or
the trigeminal stimulus (carbon dioxide, CO2). It should also be noted that a majority
of odorants (also tasting substances) typically produce trigeminal stimulation at a high
concentration level (Doty et al., 1978; Hummel and Livermore, 2002).
Von Békésy (1964) suggested that olfactory stimuli could be localized by their dif-
ferences with respect to concentration and time to reach the nostril. He showed that the
perceived location of orthonasal odors, depending on the time interval between the odor
and taste stimulation, changes from the nose to the mouth. However, it should be noted
that, in his study, only odorants capable of evoking a trigeminal sensation (e.g., benzol,
cloves, lavender, and eucalyptus) were used and participants (N = 3) had some previ-
ous experiences with localization tasks (Kleemann et al., 2009). In addition, in a study
by Negoias et al. (2013), olfactory stimuli presented to only one nostril could not be
correctly lateralized by untrained participants, while their ability to lateralize olfactory
stimuli was improved after completing four training sessions in olfactory lateralization.
Several studies have demonstrated that humans can determine whether an olfactory
stimulus comes either from the tip of the nose or from the back of the mouth (Heilmann
and Hummel, 2004; Small et al., 2005). In other words, nasal airflow direction to the
olfactory epithelium (i.e., anterior delivery of orthonasal stimulation versus posterior
delivery of retronasal stimulation) is likely to help humans in localizing odor stimulation
(Mozell, 1970; Seo and Hummel, 2016). It is unlikely, however, that determining nasal
airflow direction is necessary for localizing the olfactory stimulation when a gustatory
stimulus is also present in the mouth (Seo and Hummel, 2016). For example, combined
olfactory and gustatory stimuli (e.g., clove odor and acid solution) were perceived as a
single sensation occurring in the mouth when they were presented “simultaneously” (Von
Békésy, 1964). Such a single perception was also found even when the odor stimulus was
presented via an orthonasal route (Stevenson et al., 2011). Since a gustatory stimulation
may induce participants’ selective attention toward tasting substances, participants are
likely to be less attentive to placing an odor stimulus in an orthonasal odor referral to the
nose (Stevenson et al., 2011; Seo and Hummel, 2016). While oral somatosensory stimu-
lation of taste stimuli may also be involved in taste-smell confusion (Murphy and Cain,
1980), such stimulation alone seems to be relatively ineffective in eliciting retronasal odor
localization (referral) to the mouth because it constantly presents in the mouth during
involuntary swallowing, breathing, and talking (Lim and Johnson, 2011; Stevenson et
al., 2011; Seo and Hummel, 2016). For retronasal odor localization to the mouth, the
presence of congruent taste in the mouth has been found to be crucial (Lim and Johnson,
2011, 2012; Lim et al., 2014; Fondberg et al., 2018), while oral tactile stimulation alone
contributed no retronasal odor referral to the mouth (Lim and Johnson, 2011).

3.3.1.3 Intensity and Pleasantness

Bimodal congruency between chemosensory stimuli plays a vital role in modulating per-
ceived intensity and pleasantness of either single or mixed stimuli (Small and Prescott,
2005; Seo and Hummel, 2011a). More specifically, congruent odor stimuli are likely
36 Food Aroma Evolution

to enhance the perceived intensity and/or pleasantness of corresponding taste stimuli

(Frank and Byram, 1988; Bingham et al., 1990; Small and Prescott, 2005). However,
the enhancement of perceived taste intensity was not observed in some earlier studies
(Bingham et al., 1990; Frank and Van der Klaauw, 1992; Frank et al., 1993). For exam-
ple, Frank and Byram (1988) showed that strawberry retronasal odor but not peanut
butter retronasal odor was found to increase the perceived sweetness of whipped cream,
while the strawberry odor did not enhance the saltiness of sodium chloride solution.
Congruent gustatory cues can also modulate olfactory perception (Dalton et al.,
2000; Green et al., 2012; Fujimaru and Lim, 2013; Lim et al., 2014). As shown in Figure
3.2, Dalton et al. (2000) showed that participants were more sensitive to cherry/almond-
like odor (benzaldehyde) in the presence of a subthreshold concentration of congruent
taste (saccharin) in the mouth, but not in the presence of an incongruent taste (monoso-
dium glutamate) or of deionized water. Using olfactory event-related potentials (ERPs),
Welge-Lüssen et al. (2005) also showed that a congruent combination (e.g., vanillin
odor and sweet taste) could produce higher amplitudes and shorter latencies of N1 and
P2 peaks than an incongruent combination (e.g., vanillin odor and sour taste). This
suggests that gustatory–olfactory interaction occurs at relatively early levels of neural
processing. Neuroimaging studies have also found that the insula/operculum, the cau-
dal orbital frontal cortex, and the anterior cingulate cortex are involved in interactions
between gustatory and olfactory cues for flavor perception (Small et al., 1997; Cerf-
Ducastel and Murphy, 2001; Small et al., 2004, 2005; Small, 2006; Seo and Hummel,
2011a; Seo et al., 2013).

FIGURE 3.2 Congruent taste-enhanced odor sensitivity. Participants were more sensitive
(positive numbers represent increases in sensitivity) to cherry/almond-like odor (benzal-
dehyde) in the presence of a subthreshold concentration of congruent taste (saccharin) in
the mouth. This was not observed in the presence of an incongruent taste (monosodium
glutamate, MSG) or of deionized water. (Reprinted from Nature Neuroscience, 3(5),
Dalton, Doolittle, Nagata, and Breslin, “The merging of the senses: integration of sub-
threshold taste and smell,” pp. 431–432, 2000, with permission from Springer Nature.)
 Chemical Senses and Flavor Perception 37

Congruency-enhanced pleasantness of chemosensory stimuli between olfactory and

trigeminal stimuli has also been observed. Specifically, a congruent mixture of olfactory
and trigeminal stimuli (e.g., orange odor and carbon dioxide) was rated as more pleasant
than an incongruent mixture (e.g., rose odor and carbon dioxide) (Bensafi et al., 2013).
The congruency-enhanced pleasantness in the mixture of olfactory and trigeminal stim-
uli was found to be related to increased neural activities in the hippocampus and anterior
cingulate gyrus (Bensafi et al., 2013). Apart from a congruency between olfactory and
trigeminal stimuli, it has been found that olfactory and trigeminal stimuli interact by
mutually enhancing or suppressing one another at peripheral, central, or perceptual levels
(Cain and Murphy, 1980; Hummel and Livermore, 2002; Frasnelli et al., 2004; Brand,
2006; Pellegrino et al., 2017; Tremblay and Frasnelli, 2018).

3.3.2 Effects of Tactile or Temperature Cues of a

Stimulus Medium on Flavor Perception

Tactile cues of a stimulus medium have been found to influence chemosensory or flavor
perception (Delwiche, 2004; Verhagen and Engelen, 2006). Typically, as the viscosity of a
flavor medium increases, both sensitivity to (Mackey and Valassi, 1956; Stone and Oliver,
1966) and perceived intensity of taste, odor, or flavor stimulus (Moskowitz and Arabie,
1970; Pangborn et al., 1973, 1978; Pangborn and Szczesniak, 1974; Christensen, 1980;
Bult et al., 2007) are likely to diminish. The mechanism for suppression of flavor percep-
tion by enhanced viscosity remains unclear. It could be that the viscous characteristic of
a flavor medium evokes trigeminal stimulation, reducing flavor perception (Bayarri et
al., 2006) or inducing selective attention to the medium (Seo and Hummel, 2016). The
viscous characteristics of the medium may also modulate the pathways of releasing and/
or transporting flavor components from the medium matrix to receptors (Bayarri et al.,
2006; Mao et al., 2017). Hardness (or firmness) of a flavor medium has been found to
affect perceived flavor intensity in semi-solid and solid matrices (Seo and Hummel, 2016).
In general, perceived flavor intensity decreases with increasing hardness (or firmness) of
semi-solid and solid matrices (Weel et al., 2002; Visschers et al., 2006). Weel et al. (2002)
showed that the hardness of a whey protein gel system could change the perceived flavor
intensity, probably because of a change in mouthfeel, without affecting the in-nose fla-
vor concentration. The crispness level of a food matrix has also been found to modulate
flavor perception. For example, as shown in Figure 3.3, Luckett et al. (2016) showed
that flavors were rated as more intense and maximum flavor perception occurred sooner
with an increase in crispness level of potato chips. Interestingly, the impact of crispness
level on flavor perception was more pronounced for older participants than for younger
or middle-aged groups, probably due to an association with mastication patterns, such
as the number of chews. In fact, the textural properties of semi-solid or solid foods can
change consumers’ mastication patterns. These include chewing rate, chewing duration,
and the number of chews, all of which can affect temporal flavor dynamics as well as
general flavor perception (Blissett et al., 2006; Tarrega et al., 2008; Luckett et al., 2016;
Luckett and Seo, 2017).
The temperature of a stimulus medium is another important factor influencing taste,
odor, or flavor perception (Olson et al., 1980; Kähkönen et al., 1995; Cruz and Green,
2000; Engelen et al., 2003; Ventanas et al., 2010). Although there were conflicting results
among them, previous psychophysical studies using basic taste solutions have shown
effects of solution temperature on taste intensity (Hahn and Günther, 1932; McBurney
38 Food Aroma Evolution

FIGURE 3.3 Mean time-intensity curves of potato chips as a function of crispness level
(low, medium, and high), flavor type (plain, cheese, and spicy), and age group [younger
(20–25 years old), middle-aged (40–45 years old), and older (65+ years old) groups].
Flavors were rated as more intense and maximum flavor perception occurred quicker
with an increase in crispness level of potato chips. The effect of crispness level on fla-
vor perception was more clearly observed in the older participants than in the younger
or middle-aged groups. (Reprinted from Food Quality and Preference, 51, Luckett,
Meullenet, and Seo, “Crispness level of potato chips affects temporal dynamics of flavor
perception and mastication patterns in adults of different age groups,” pp. 8–19, 2016,
with permission from Elsevier.)

et al., 1973; Moskowitz, 1973; Bartoshuk et al., 1982; Prescott et al., 1984; Calviño,
1986; Green and Frankmann, 1987, 1988; Schiffman et al., 2000; Green and Andrew,
2017; for a review, Lemon, 2017). One plausible explanation for such inconsistency is
that tongue temperature had not been carefully controlled over the testing interval (Green
and Frankmann, 1987; Delwiche, 2004). In other words, variations in oral temperature
could have possibly altered the perceived intensity of basic taste solutions (Cruz and
Green, 2000; Green, 2002).
The effects of oral temperature on taste, odor, or flavor perception have also been
studied using more complex matrices such as food and beverage models (Drake et al.,
2005; Ross and Weller, 2008; Mony et al., 2013; Kim et al., 2015; Stokes et al., 2016;
Steen et al., 2017; Adhikari et al., 2018; Pramudya and Seo, 2018a,b; Chapko and Seo,
2019), in which the effects of oral or product temperature were dependent on the type of
food (Mela et al., 1994; Engelen et al., 2003; Mony et al., 2013; Kim et al., 2015) and
dietary behaviors (Kim et al., 2015). For example, Kim et al. (2015) asked both trained
panelists and untrained consumers to rate saltiness intensities in sodium chloride (NaCl)
solution, chicken broth, and miso soup at five different temperatures: 80, 70, 60, 50, and
 Chemical Senses and Flavor Perception 39

40°C. Neither trained nor untrained consumer panelists were able to find a significant
difference in saltiness intensity among the NaCl solutions. However, untrained consum-
ers, not trained panelists, rated chicken broth and miso soup as less salty when served at
70 and/or 80°C compared to when there were served at 40 to 60°C. Consumers accus-
tomed to frequently consuming hot dishes also rated the soup samples served at 60°C sig-
nificantly saltier than those less accustomed to consuming hot dishes. Recent studies have
also demonstrated the effect of serving/consumption temperature on flavor perception of
brewed coffee (Stokes et al., 2016; Steen et al., 2017; Adhikari et al., 2018; Chapko and
Seo, 2019). Chapko and Seo (2019) asked six trained panelists to rate intensities of 32
sensory attributes (3 appearance, 12 aromas, 13 flavors, 2 tastes, and 2 mouthfeels) with
respect to three varieties (Colombian, Ethiopian, and Kenyan) of brewed coffee presented
at four different serving temperatures: 75, 55, 40, and 25°C, respectively. Interestingly, as
shown in Figure 3.4, a principal component analysis (PCA) showed that serving tempera-
ture could account for the greater amount of data variation (63.28% of total variation)
than coffee variety (21.24%). For example, aromas and flavors of “roasted” and “cof-
fee impression” were more associated with brewed coffee served at higher temperatures
(70 and 55°C), while “stale” flavor was more likely to be associated with brewed coffee
served at lower temperatures (40 and 25°C). These results emphasize that sensory attri-
butes of brewed coffee should be evaluated at multiple s­ erving temperatures, for example,
both higher (70 to 55°C) and lower (40 to 25°C) ones (Chapko and Seo, 2019). Taken
together, the effects of oral or product temperature on taste, odor, or flavor perception
must be interpreted from multiple perspectives.

FIGURE 3.4 Two plots of principal component analysis with respect to three cof-
fee varieties (Colombian, Ethiopian, and Kenyan) of brewed coffee samples evaluated
at four serving temperatures: 75, 55, 40, and 25°C. (A) Represents 24 sensory attri-
butes used for data analysis and (B) represents 12 brewed coffee samples (three variet-
ies × four serving temperatures). “A,” “T,” and “F” represent aroma, taste, and flavor,
respectively. (Reprinted from Food Research International, In Press, Chapko and Seo,
“Characterizing product temperature-dependent sensory perception of brewed coffee
beverages: Descriptive sensory analysis.” https​://do​i.org ​/10.1​016/j​.food​res.2​018.1​2 .026,
2019, with permission from Elsevier.)
40 Food Aroma Evolution

3.3.3 Effects of Visual or Auditory Cues on Flavor Perception

Some researchers have proposed that all sensory inputs related to substances con-
sumed during eating or drinking could be interactive. This will affect flavor perception
(Verhagen and Engelen, 2006; Spence, 2015a), although chemosensory cues, that is, gus-
tatory, olfactory, and trigeminal sensations, are considered to be major components of
flavor. Earlier studies provided empirical evidence that certain gustatory or olfactory
cues could be matched with specific attributes of non-chemosensory cues. These would
include color hue (Gilbert et al., 1996; Zellner et al., 2008; for a review, Spence et al.,
2015), color brightness (Von Hornbostel, 1931; Fiore, 1993), shapes (Hanson-Vaux et al.,
2013), symbols (Seo et al., 2010), pitch (Crisinel and Spence, 2010; Wang et al., 2016),
and timbre (Crisinel and Spence, 2011, 2012).
Bimodal congruency between chemosensory and non-chemosensory cues (e.g.,
visual or auditory) has been found to play a crucial role in modulating chemosensory or
flavor perception (Delwiche, 2004; Zampini et al., 2007, 2008; Seo et al., 2010; Shankar
et al., 2010; Seo and Hummel, 2014). More specifically, congruent colors are more likely
than incongruent colors to increase flavor discrimination (Zampini et al., 2007) or iden-
tification (DuBose et al., 1980; Stillman, 1993; Philipsen et al., 1995). Interestingly,
when white wines are colored red using an odorless dye, individuals describe them using
odor-related terms more associated with red wines than with white wines (Morrot et
al., 2001). Also, when the color of a tasting substance is more intense, individuals tend
to rate taste, odor, or flavor cues as more intense (DuBose et al., 1980; Johnson et al.,
1982; Johnson and Clydesdale, 1982; also see Frank et al., 1989; Lavin and Lawless,
1998; Delwiche, 2004). Such effects of color cues on taste/odor/flavor perceptions are
influenced by many other factors, including participant age (Philipsen et al., 1995; Lavin
and Lawless, 1998), gender, cultural background (Shankar et al., 2010), odor stimulus
delivery route (Koza et al., 2005), and experimental contexts (Seo and Hummel, 2011a).
For example, the effect of darker red colors on sweetness intensity was found to be
present in adults, but not in children aged between 5 and 10 (Lavin and Lawless, 1998).
Koza et al. (2005) also demonstrated that when colored odorous solutions were sniffed,
the colors increased the perceived intensities of the odor stimuli, but an opposite effect
of color on odor intensity was found when solutions were smelled retronasally (from the
mouth). Moreover, in a study conducted by Shankar et al. (2010), 70% of British par-
ticipants, when presented with a brown colored drink, matched the brown color with a
cola flavor, while 49% of Taiwanese participants associated the same color with a grape
flavor. Such examples illustrate that the effect of a visual cue such as color on chemo-
sensory or flavor perception should be carefully approached because there may be many
influential factors (Spence et al., 2010).
Although auditory cues have been considered to represent a “forgotten” flavor sense
(Delwiche, 2004; Spence, 2012), there is a rapidly growing body of empirical evidence
that auditory cues can be matched with gustatory, olfactory, or flavor stimuli (for a
review, Spence, 2012). For example, Crisinel and Spence (2010) showed that 12 different
taste and flavor stimuli were matched with different levels of pitch. Low-pitched sounds
generated by brass instruments were frequently matched with the bitter taste of a caffeine
solution, while high-pitched sounds produced by a piano were often matched with the
sweet taste of a sucrose solution.
Auditory cues can also affect perceived intensities and likings (Ferber and Cabanac,
1987; Woods et al., 2011; Stafford et al., 2012; Fiegel et al., 2014; Yan and Dando, 2015;
Kantono et al., 2016, 2018; Reinoso Carvalho et al., 2016, 2017) as well as discrimination
 Chemical Senses and Flavor Perception 41

(Pellegrino et al., 2015) of chemosensory or flavor stimuli, although these effects were not
consistently reported in earlier studies. Woods et al. (2011) showed that loud background
noise (75–85 dB) more than quiet background noise (45–55 dB) could decrease perceived
intensities of sweetness and saltiness (see also Stafford et al., 2012). Similarly, cabin noise
presented at a level of 80 to 85 dB was found to decrease sweetness intensity of a sweet
sucrose solution, while it increased the umami intensity of an umami monosodium glu-
tamate (MSG) solution (Yan and Dando, 2015). A hedonic tone of auditory stimuli is
also likely to be transformed into hedonic ratings of odor, flavor, and food/drink stimuli
(Seo and Hummel, 2011b; Fiegel et al., 2014, 2019; Kantono et al., 2016, 2018; Reinoso
Carvalho et al., 2016).

3.4 CONCLUSION

The flavor is a complex and dynamic combination of gustatory, olfactory, and oral
somatosensory (in particular trigeminal) sensations evoked by ingested substances during
eating and drinking. Although gustatory, olfactory, and oral somatosensory (trigeminal)
systems are different at the peripheral level, each system has its own role in daily life.
They often interact at the peripheral and central nervous levels. In addition, flavor per-
ception is influenced by many other factors that include non-chemosensory cues, demo-
graphic profiles, and environmental eating contexts. Flavor perception should therefore
be considered to be a Gestalt concept.

REFERENCES

Adhikari, J., Chambers, E., IV, and Koppel, K. 2018. Impact of consumption tempera-
ture on sensory properties of hot brewed coffee. Food Research International, 115,
95–104.
Adler, E., Hoon, M. A., Mueller, K. L., Chandrashekar, J., Ryba, N. J. P., and Zuker, C.
S. 2000. A novel family of mammalian taste receptors. Cell, 100, 693–702.
Albrecht, J., Kopietz, R., Frasnelli, J., Wiesmann, M., Hummel, T., and Lundström, J.
N. 2010. The neuronal correlates of intranasal trigeminal function—An ALE meta-
analysis of human functional brain imaging data. Brain Research Reviews, 62,
183–96.
Amerine, M. A., Pangborn, R. M., and Roessler, E. B. 1965. Principles of Sensory
Evaluation of Food. New York, NY: Academic Press.
Aschenbrenner, K., Hummel, C., Teszmer, K., Krone, F., Ishimaru, T., Seo, H.-S., and
Hummel, T. 2008. The influence of olfactory loss on dietary behaviors. Laryngoscope
118, 135–44.
Bachmanov, A. A. and Beauchamp, G. K. 2007. Taste receptor genes. Annual Review of
Nutrition, 27, 389–414.
Bartel, D. L., Sullivan, S. L., Lavoie, E. G., Sévigny, J., and Finger, T. E. 2006. Nucleoside
triphosphate diphosphohydrolase-2 is the ecto-ATPase of type I cells in taste buds.
Journal of Computational Neurology, 497, 1–12.
Bartoshuk, L. M., Duffy, V. B., and Miller, I. J. 1994. PTC/PROP tasting: Anatomy,
psychophysics, and sex effects. Physiology & Behaviors, 56, 1165–71.
Bartoshuk, L. M., Rennert, K., Rodin, J., and Stevens, J. C. 1982. Effects of temperature
on the perceived sweetness of sucrose. Physiology & Behavior, 28, 905–10.
42 Food Aroma Evolution

Bathla, G. and Hegde, A. N. 2013. The trigeminal nerve: An illustrated review of its
imaging anatomy and pathology. Clinical Radiology, 68, 203–13.
Bayarri, S., Taylor, A. J., and Hort, J. 2006. The role of fat in flavor perception: Effect
of partition and viscosity in model emulsions. Journal of Agricultural and Food
Chemistry, 54, 8862–8.
Beckstead, R. M. and Norgren, R. 1979. An autoradiographic examination of the central
distribution of the trigeminal, facial, glosspharyngeal, and vagal nerves in the mon-
key. Journal of Computational Neurology, 184, 455–72.
Bensafi, M., Iannilli, E., Schriever, V. A., Poncelet, J., Seo, H.-S., Gerber, J., Rouby, C.,
and Hummel, T. 2013. Cross-modal integration of emotions in the chemical senses.
Frontiers in Human Neuroscience, 7, 883.
Bingham, A. F., Birch, G. G., de Graaf, C., Behan, J. M., and Perring, K. D. 1990.
Sensory studies with sucrose-maltol mixtures. Chemical Senses, 15, 447–56.
Blissett, A., Hort, J., and Taylor, A. J. 2006. Influence of chewing and swallowing behav-
ior on volatile release in two confectionery systems. Journal of Texture Studies, 37,
476–96.
Blossfeld, I., Collins, A., Boland, S., Baixauli, R., Kiely, M., and Delahunty, C. 2007.
Relationships between acceptance of sour taste and fruit intakes in 18-month-old
infants. British Journal of Nutrition, 98, 1084–91.
Brand, G. 2006. Olfactory/trigeminal interactions in nasal chemoreception. Neuroscience
& Behavioral Reviews, 30, 908–17.
Breslin, P. A. S. and Huang, L. 2006. Human taste: Peripheral anatomy, taste trans-
duction, and coding. In Taste and Smell. An Update. Advances in Oto-Rhino-
Laryngology, eds. T. Hummel and A. Welge-Lüssen, pp. 152–90. Basel: Karger.
Bult, J. H., de Wijk, R. A., and Hummel, T. 2007. Investigations on multimodal sensory
integration: texture, taste, and ortho- and retronasal olfactory stimuli in concert.
Neuroscience Letters, 411, 6–10.
Buschhüter, D., Smitka, M., Puschmann, S., Gerber, J. C., Witt, M., Abolmaali, N. D.,
and Hummel, T. 2008. Correlation between olfactory bulb volume and olfactory
function. Neuroimage, 42, 498–502.
Cain, W. S. and Murphy, C. L. 1980. Interaction between chemoreceptive modalities of
odour and irritation. Nature, 284, 255–7.
Calviño, A. M. 1986. Perception of sweetness: The effects of concentration and tempera-
ture. Physiology & Behavior, 36, 1021–8.
Caterina, M. J., Schumacher, M. A., Tomiaga, M., Rosen, T. A., Levine, J. D., and Julius,
D. 1997. The capsaicin receptor: A heat-activated ion channel in the pain pathway.
Nature, 389, 816–24.
Cerf-Ducastel, B. and Murphy, C. 2001. fMRI activation in response to odorants orally
delivered in aqueous solutions. Chemical Senses, 26, 625–37.
Chandrashekar, J., Hoon, M. A., Ryba, N. J., and Zuker, C. S. 2006. The receptors and
cells for mammalian taste. Nature, 444, 288–94.
Chapko, M. J. and Seo, H.-S. 2019. Characterizing product temperature-dependent
sensory perception of brewed coffee beverages: Descriptive sensory analysis.
Food Research International, In press. https​: //do​i.org ​/10/1​016/j​.food​res.2​018.1​
2.026​.
Chaudhari, N. And Roper, S. D. 2010. The cell biology of taste. Journal of Cell Biology,
190, 285–96.
Christensen, C. M. 1980. Effects of solution viscosity on perceived saltiness and sweet-
ness. Perception & Psychophysics, 28, 347–53.
 Chemical Senses and Flavor Perception 43

Christensen, C. M., Brand, J. G., and Malamud, D. 1987. Salivary changes in solu-
tion pH: A source of individual differences in sour taste perception. Physiology &
Behavior, 40, 221–7.
Correa, M., Hutchinson, I., Laing, D. G., and Jinks, A. L. 2013. Changes in fungiform
papillae density during development in humans. Chemical Senses, 38, 519–27.
Courtiol, E. and Wilson, D. A. 2014. Thalamic olfaction: Characterizing odor processing
in the mediodorsal thalamus of the rat. Journal of Neurophysiology, 111, 1274–85.
Courtiol, E. and Wilson, D. A. 2015. The olfactory thalamus: Unanswered questions
about the role of the mediodorsal thalamic nucleus in olfaction. Frontiers in Neural
Circuits, 9, 49.
Crisinel, A.-S. and Spence, C. 2010. A sweet sound? Food names reveal implicit associa-
tions between taste and pitch. Perception, 39, 417–25.
Crisinel, A.-S. and Spence, C. 2011. Crossmodal associations between flavoured milk
solutions and musical notes. Acta Psychologica, 138, 155–61.
Crisinel, A.-S. and Spence, C. 2012. The impact of pleasantness ratings on crossmodal
associations between food samples and musical notes. Food Quality and Preference,
24, 136–40.
Croy, I., Negoias, S., Novakova, L., Landis, B., and Hummel, T. 2012. Learning about
the functions of the olfactory system from people without a sense of smell. PLoS
ONE, 7, e33365.
Croy, I., Nordin, S., and Hummel, T. 2014. Olfactory disorders and quality of life—An
updated review. Chemical Senses, 39, 185–94.
Cruz, A. and Green, B. G. 2000. Thermal stimulation of taste. Nature, 403, 889–92.
Dalton, P., Doolittele, N., Nagata, H., and Breslin, P. A. 2000. The merging of the senses:
Integration of subthreshold taste and smell. Nature Neuroscience, 3, 431–2.
Darian-Smith, I., Johnson, K. O., and LaMotte, C. 1979. Warm fibers innervating
palmar and digital skin of the monkey: Responses to thermal stimuli. Journal of
Neurophysiology, 42, 1297–315.
Davison, I. G. and Ehlers, M. D. 2011. Neural circuit mechanisms for pattern detection
and feature combination in olfactory cortex, Neuron, 70, 82–94.
Deems, D. A., Doty, R. L., Settle, R. G., Moore-Gillon, V., Shaman, P., Mester, A. F.,
Kimmelman, C. P., Brightman, V. J., and Snow, J. B., Jr. 1991. Smell and taste dis-
orders, a study of 750 patients from the University of Pennsylvania Smell and Taste
Center. Archives of Otolaryngology—Head & Neck Surgery, 117, 519–28.
Delwiche, J. 2003. Attributes believed to impact flavor: An opinion survey. Journal of
Sensory Studies, 18, 347–52.
Delwiche, J. 2004. The impact of perceptual interactions on perceived flavor. Food
Quality and Preference, 15, 137–46.
DeSimone, J. A. and Lyall, V. 2006. Taste receptors in the gastrointestinal tract. III. Salty
and sour taste: Sensing of sodium and protons by the tongue. American Journal of
Physiology-Gastrointestinal and Liver Physiology, 291, G1005–10.
Dinnella, C., Monteleone, E., Piochi, M., Spinelli, S., Prescott, J., Pierguidi, L., Gasperi,
F., Laureati, M., Pagliarini, E., Predieri, S., Torri, L., Barbieri, S., Valli, E., Bianchi,
P., Braghieri, A., Del Caro, A., Di Monaco, R., Favotto, S., and Moneta, E. 2018.
Individual variation in PROP status, fungiform papillae density, and responsiveness
to taste stimuli in a large population sample. Chemical Senses, 43, 697–710.
Doty, R. L., Brugger, W. P. E., Jurs, P. C., Orndorff, M. A., Snyder, P. J., and Lowry, L.
D. 1978. Intranasal trigeminal stimulation from odorous volatiles: Psychometric
responses from anosmic and normal humans. Physiology & Behavior, 20, 175–85.
44 Food Aroma Evolution

Doty, R. L. and Cometto-Muñiz, I. E. 2003. Trigeminal chemosensation. In Handbook

of Olfaction and Gustation, 2nd ed., ed. R. D. Tory, pp. 981–99. New York, NY:
Marcel Dekker.
Drake, M. A., Yates, M. D., and Gerard, P. D. 2005. Impact of serving temperature on
trained panel perception of cheddar cheese flavor attributes. Journal of Sensory
Studies, 20, 147–55.
Drewnowski, A. and Gomez-Carneros, C. 2000. Bitter taste, phytonutrients, and the
consumer: A review. American Journal of Clinical Nutrition, 72, 1424–35.
DuBose, C. N., Cardello, A. V., and Maller, O. 1980. Effects of colorants and flavorants
on identification, perceived flavor intensity, and hedonic quality of fruit-flavored
beverages and cake. Journal of Food Science, 45, 1393–9.
Duffy, V. B., Backstrand, J. R., and Ferris, A. M. 1995. Olfactory dysfunction and related
nutritional risk in free-living, elderly women. Journal of the American Dietetic
Association, 95, 879–84.
Eldeghaidy, S., Thomas, D., Skinner, M., Ford, R., Giesbrecht, T., Thomas, A., Hort, J.,
and Francis, S. 2018. An automated method to detect and quantify fungiform papil-
lae in the human tongue: Validation and relationship to phenotypical differences in
taste perception. Physiology & Behavior, 184, 226–34.
Engelen, L. 2012. Chapter 2. Oral receptors. In Food Oral Processing: Fundamentals of
Eating and Sensory Perception, eds. J. Chen and L. Engelen, pp. 15–43. Oxford:
Blackwell Publishing.
Engelen, L., de Wijk, R. A., Prinz, J. F., Janssen, A. M., Weenen, H., and Bosman, F.
2003. The effect of oral and product temperature on the perception of flavor and
texture attributes of semi-solids. Appetite, 41, 273–81.
Fallon, A. and Rozin, P. 1983. The psychological bases of food rejection by humans.
Ecology of Food and Nutrition, 13, 15–26.
Fark, T., Hummel, C., Hähner, A., Nin, T., and Hummel, T. 2013. Characteristics of
taste disorders. European Archives of Otorhinolaryngology, 270, 1855–60.
Feeney, E. L. and Hayes, J. E. 2014. Regional differences in suprathreshold intensity for
bitter and umami taste. Chemosensory Perception, 7, 147–57.
Feng, P., Huang, L., and Wang, H. 2014. Taste bud homeostasis in health, disease, and
aging. Chemical Senses, 39, 3–16.
Ferber, C. and Cabanac, M. 1987. Influence of noise on gustatory affective ratings and
preference for sweet or salt. Appetite, 8, 229–35.
Fiegel, A., Childress, A., Beekman, T. L., and Seo, H.-S. 2019. Variations in food
acceptability with respect to pitch, tempo, and volume levels of background music.
Multisensory Research, 32, 319–46, In Press.
Fiegel, A., Meullenet, J.-F., Harrington, R. J., Humble, R., and Seo, H.-S. 2014.
Background music genre can modulate flavor pleasantness and overall impression of
food stimuli. Appetite, 76, 144–52.
Fiore, A. M. 1993. Multisensory integration of visual, tactile, and olfactory aesthetic
cues of appearance. Clothing and Textiles Research Journal, 11, 45–52.
Fondberg, R., Lundström, J. N., Blöchl, M., Olsson, M. J., and Seubert, J. 2018.
Multisensory flavor perception: The relationship between congruency, pleasantness,
and odor referral to the mouth. Appetite, 125, 244–52.
Forestell, C. A. and Mennella, J. A. 2017. The relationship between infant facial expres-
sions and food acceptance. Current Nutrition Reports, 6, 141–7.
Frank, R. A. and Byram, J. 1988. Taste-smell interactions are tastant and odorant depen-
dent. Chemical Senses, 13, 445–55.
 Chemical Senses and Flavor Perception 45

Frank, R. A., Ducheny, K., and Mize, S. J. S. 1989. Strawberry odor, but not red color,
enhances the sweetness of sucrose solutions. Chemical Senses, 14, 371–7.
Frank, R. A. and Van der Klaauw, N. J. 1992. The influence of stimulus context and
instructional set on odor-induced enhancement of taste. Chemical Senses, 17, 625.
Frank, R. A., Van der Klaauw, N. J., and Schifferstein, H. N. 1993. Both perceptual and
conceptual factors influence taste-odor and taste-taste interactions. Perception &
Psychophysics, 54, 343–54.
Frasnelli, J., Charbonneau, G., Collignon, O., and Lepore, F. 2009. Odor localization
and sniffing. Chemical Senses, 34, 139–44.
Frasnelli, J., Landis, B. N., Heilmann, S., Hauswald, B., Hüttenbrink, K. B., Lacroix,
J. S., Leopold, D. A., and Hummel, T. 2004. Clinical presentation of qualita-
tive olfactory dysfunction. European Archives of Oto-Rhino-Laryngology, 261,
411–5.
Frasnelli, J. and Manescu, S. 2016. Chapter 46. The intranasal trigeminal system. In
Springer Handbook of Odor, ed. A. Buettner, pp. 881–95. New York, NY: Springer.
Freiherr, J. 2016. Chapter 38. Cortical olfactory processing. In Springer Handbook of
Odor, ed. A. Buettner, pp. 759–67. New York, NY: Springer.
Fujimaru, T. and Lim, J. 2013. Effects of stimulus intensity on odor enhancement by
taste. Chemosensory Perception, 6, 1–7.
Fukuwatari, T., Kawada, T., Tsuruta, M., Hiraoka, T., Iwanaga, T., Sugimoto, E., and
Fushiki, T. 1997. Expression of the putative membrane fatty acid transporter (FAT)
in taste buds of the circumvallate papillae in rats. FEBS Letters, 414, 461–4.
Gilbert, A. N., Martin, R., and Kemp, S. E. 1996. Cross-modal correspondence between
vision and olfaction: The color of smells. American Journal of Psychology, 109,
335–51.
Goldberg, E. M., Wang, K., Goldberg, J., and Aliani, M. 2018. Factors affecting the
ortho- and retronasal perception of flavors: A review. Critical Reviews in Food
Science and Nutrition, 58, 913–23.
Gottfried, J. A. 2006. Smell: Central nervous processing. In Taste and Smell. An Update.
Advances in Oto-Rhino-Laryngology, eds. T. Hummel and A. Welge-Lüssen, pp.
44–69. Basel: Karger.
Gottfried, J. A., Winston, J. S., and Dolan, R. J. 2006. Dissociable codes of odor quality
and odorant structure in human piriform cortex. Neuron, 49, 467–79.
Grabenhorst, F., Rolls, E. T., Margot, C., da Silva, M. A. A. P., and Velazco, M. I. 2007.
How pleasant and unpleasant stimuli combine in different brain regions: Odor mix-
tures. Journal of Neuroscience, 27, 13532–40.
Green, B. G. 1996. Chemesthesis: Pungency as a component of flavor. Trends in Food
Science & Technology, 7, 415–20.
Green, B. G. 2002. Studying taste as a cutaneous sense. Food Quality and Preference,
14, 99–109.
Green, B. G. 2016. Chapter 1. Introduction. What is chemesthesis? In Chemesthesis:
Chemical Tough in Food and Eating, eds. S. T. McDonald, D. A. Bolliet, and J. E.
Hayes, pp. 1–7. Oxford: John Wiley & Sons.
Green, B. G. and Andrew, K. 2017. Stimulus-dependent effects of temperature on bitter
taste in humans. Chemical Senses, 2017, 153–60.
Green, B. G. and Frankmann, S. P. 1987. The effect of cooling the tongue on the per-
ceived intensity of taste. Chemical Senses, 12, 609–19.
Green, B. G. and Frankmann, S. P. 1988. The effect of cooling on the perception of car-
bohydrate and intensive sweeteners. Physiology & Behavior, 43, 515–9.
46 Food Aroma Evolution

Green, B. G. and Lawless. H. T. 1991. The psychophysics of somatosensory chemorecep-

tion in the nose and mouth. In Smell and Taste in Health and Disease, eds. T. V.
Getchell, R. L. Doty, L. M. Bartoshuk, and J. B. Snow Jr., pp. 235–54. New York,
NY: Raven Press.
Green, B. G., Mason, J. R., and Kare, M. R. 1990. Preface. In Chemical Senses, Irritation,
eds. B. G. Green, J. R. Mason, and M. R. Kare, pp. v–vii. New York, NY: Marcel
Dekker.
Green, B. G., Nachtigal, D., Hammond, S., and Lim, J. 2012. Enhancement of retronasal
odors by taste. Chemical Senses, 37, 77–86.
Haehner, A., Rodewald, A., Gerber, J. C., and Hummel, T. 2008. Correlation of olfac-
tory function with changes in the volume of the human olfactory bulb. Archives of
Otolaryngology—Head & Neck Surgery, 134, 621–4.
Hahn, H. and Günther, H. 1932. Über die Reize und die Reizbedingungen des
Geschmackssinnes. Pflügers Archiv: European Journal of Physiology, 231, 48–67.
Hall, R. L. 1968. Food flavors: Benefits and problems. Food Technology, 22, 1388–92.
Han, P., Mann, S., Raue, C., Warr, J., and Hummel, T. 2018. Pepper with and without a
sting: Brain processing of intranasal trigeminal and olfactory stimuli from the same
source. Brain Research, 1700, 41–6.
Hanson-Vaux, G., Crisinel, A.-S., and Spence, C. 2013. Smelling shapes: Crossmodal
correspondences between odors and shapes. Chemical Senses, 38, 161–6.
Hartley, I. E., Liem, D. G., and Keast, R. 2019. Umami as an “Alimentary” taste. A new
perspective on taste classification. Nutrients, 11, 182.
Heilmann, S. and Hummel, T. 2004. A new method for comparing orthonasal and retro-
nasal olfaction. Behavioral Neuroscience, 118, 412–9.
Higaki, N., Goto, T., Ishida, Y., Watanabe, M., Tomotake, Y., and Ichikawa, T. 2014.
Do sensation differences exist between dental implants and natural teeth? A meta-
analysis. Clinical Oral Implants Research, 25, 1307–10.
Hladik, C. M. and Simmen, B. 1996. Taste perception and feeding behavior in nonhu-
man primates and human populations. Evolutionary Anthropology, 5, 58–71.
Hornung, D. E. 2006. Nasal anatomy and the sense of smell. In Taste and Smell. An
Update. Advances in Oto-Rhino-Laryngology, eds. T. Hummel and A. Welge-
Lüssen, pp. 1–22. Basel: Karger.
Howard, J. D., Plailly, J., Grueschow, M., Haynes, J. D., and Gottfried, J. A. 2009.
Odor quality coding and categorization in human posterior piriform cortex. Nature
Neuroscience, 12, 932–38.
Huang, A. L., Chen, X., Hoon, M. A., Chandrashekar, J., Guo, W., Tränkner, D., Ryba,
N. J. P., and Zuker, C. S. 2006. The cells and logic for mammalian sour taste detec-
tion. Nature, 442, 934–8.
Huart, C., Collet, S., and Rombaux, P. 2009. Chemosensory pathways: From periphery
to cortex. B-ENT, 5(Suppl. 13), 3–9.
Hummel, T., Croy, I., and Haehner, A. 2016. Chapter 16, Olfactory disorders and conse-
quences. In Flavor from Food to Behaviors, Wellbeing and Health, eds. P. Etiévant, E.
Guichard, C. Salles, and A. Voilley, pp. 361–77. New York, NY: Woodhead Publishing.
Hummel, T., Haehner, A., Hummel, C., Croy, I., and Iannilli, E. 2013. Lateralized differ-
ences in olfactory bulb volume relate to lateralized differences in olfactory function.
Neuroscience, 237, 51–5.
Hummel, T., Landis, B. N., and Hüttenbrink, K. B. 2011. Dysfunction of the chemical
senses smell and taste. Laryngorhinootologie, 90(Suppl. 1), S44–55.
 Chemical Senses and Flavor Perception 47

Hummel, T. and Livermore, A. 2002. Intranasal chemosensory function of the trigeminal

nerve and aspects of its relation to olfaction. International Archives of Occupational
and Environmental Health, 75, 305–13.
Iannilli, E. and Gudziol, V. 2019. Gustatory pathway in humans: A review of mod-
els of taste perception and their potential lateralization. Journal of Neuroscience
Research, 97, 230–40.
Imai, T. 2014. Construction of functional neuronal circuitry in the olfactory bulb.
Seminars in Cell & Developmental Biology, 35, 180–8.
Insausti, R., Marcos, P., Arroyo-Jiménez, M. M., Blaizot, X., and Martínez-Marcos, A.
2002. Comparative aspects of the olfactory portion of the entorhinal cortex and
its projection to the hippocampus in rodents, nonhuman primates, and the human
brain. Brain Research Bulletin, 57, 557–60.
International Standards Organization. 2008. Standard 5492: Terms Relating to Sensory
Analysis. International Organization for Standardization. Vienna, Austria: Austrian
Standards Institute.
Ishimaru, Y., Inada, H., Kubota, M., Zhuang, H., Tominaga, M., and Matusnami, H.
2006. Transient receptor potential family members PKD1L3 and PKD2L1 form a
candidate sour taste receptor. Proceeding of the National Academy of Sciences of
the United States of America, 103, 12569–74.
Johnson, J. and Clydesdale, F. M. 1982. Perceived sweetness and redness in colored
sucrose solutions. Journal of Food Science, 47, 747–52.
Johnson, J. L., Dzendolet, E., Damon, R., Sawyer, M., and Clydesdale, F. M. 1982.
Psychophysical relationships between perceived sweetness and color in cheery-fla-
vored beverages. Journal of Food Protection, 45, 601–6.
Just, T., Stave, J., Pau, H. W., and Guthoff, R. 2005. In vivo observation of papillae of the
human tongue using confocal laser scanning microscopy. ORL: Journal for Oto-
Rhino-Laryngology and Its Related Specialties, 67, 207–12.
Kähkönen, P., Tuorila, H., and Hyvönen, L. 1995. Dairy fat content and serving tempera-
ture as determinants of sensory and hedonic characteristics in cheese soup. Food
Quality and Preference, 6, 127–33.
Kantono, K., Hamid, N., Shepherd, D., Lin, Y. H. T., Brad, C., Grazioli, G., and Carr,
T. 2018. The effect of music on gelato perception in different eating contexts. Food
Research International, 113, 43–56.
Kantono, K., Hamid, N., Shepherd, D., Yoo, M. J. Y., Grazioli, G., and Carr, B. T. 2016.
Listening to music can influence hedonic and sensory perceptions of gelati. Appetite,
100, 244–55.
Kataoka, S., Yang, R., Ishimaru, Y., Matsunami, H., Sévigny, J., Kinnamon, J. C., and
Finger, T. E. 2008. The candidate sour taste receptor, PKD2L1, is expressed by type
III taste cells in the mouse. Chemical Senses, 33, 243–54.
Keast, R. S. J. and Breslin, P. A. S. 2003. An overview of binary taste-taste interactions.
Food Quality and Preference, 14, 111–24.
Keast, R. S. J. and Costanzo, A. 2015. Is fat the sixth taste primary? Evidence and impli-
cations. Flavour, 4, 5.
Khan, N. A. and Besnard, P. 2009. Oro-sensory perception of dietary lipids: New insights
into the fat taste transduction. Biochemical Biophysical Acta, 1791, 149–55.
Kim, J.-W., Samant, S. S., Seo, Y., and Seo, H.-S. 2015. Variation in saltiness percep-
tion of soup with respect to soup serving temperature and consumer dietary habits.
Appetite, 84, 73–8.
48 Food Aroma Evolution

Kleemann, A. M., Albrecht, J., Schöpf, V., Haegler, K., Kopietz, R., Hempel, J. M., Linn,
J., Flanagin, V. L., Fesl, G., and Wiesmann, M. 2009. Trigeminal perception is nec-
essary to localize odors. Physiology & Behavior, 97, 401–5.
Kobal, G., Van Toller, S., and Hummel, T. 1989. Is there directional smelling? Experientia,
45, 130–2.
Konstantindis, I. 2009. The taste peripheral system. B-ENT, 5 (Suppl. 13), 115–21.
Koza, B. J., Cilmi, A., Dolese, M., and Zellner, D. A. 2005. Color enhances orthonasal
olfactory intensity and reduces retronasal olfactory intensity. Chemical Senses, 30,
643–9.
Kullaa-Mikkonen, A., Koponen, A., and Seilonen, A. 1987. Quantitative study of human
papillae and taste buds: Variation with aging and in different morphological forms
of the tongue. Gerodontics, 3, 131–5.
LaMotte, R. H. and Campbell, J. N. (1978). Comparison of responses of warm and noci-
ceptive C-fiber afferents in monkey with human judgments of thermal pain. Journal
of Neurophysiology, 41, 509–28.
Landis, B. N., Konnerth, C. G., and Hummel, T. 2004. A study on the frequency of olfac-
tory function. Laryngoscope, 114, 1764–9.
Lapis, T. J., Penner, M. H., and Lim, J. 2014. Evidence that humans can taste glucose
polymers. Chemical Senses, 39, 737–47.
Lapis, T. J., Penner, M. H., and Lim, J. 2016. Humans can taste glucose oligomers inde-
pendent of the hT1R2/hT1R3 sweet taste receptors. Chemical Senses, 41, 755–62.
Lavin, J. and Lawless, H. T. 1998. Effects of color and odor on judgments of sweetness
among children and adults. Food Quality and Preference, 9, 283–9.
Lavoie, J., Gassó Astorga, P., Segal-Gavish, H., Wu, Y. C., Chung, Y., Cascella, N. G.,
Sawa, A., and Ishizuka, K. 2017. The olfactory neural epithelium as a tool in neuro-
science. Trends in Molecular Medicine, 32, 100–3.
Lemon, C. H. 2017. Modulation of taste processing by temperature. American Journal of
Physiology, Regulatory, Integrative and Comparative Physiology, 313, R305–21.
Leopold, D. 2002. Distortion of olfactory perception: Diagnosis and treatment. Chemical
Senses, 27, 611–5.
Lim, J. 2016. Chapter 3. Oral referral. In Multisensory Flavor Perception. From
Fundamental Neuroscience Through to the Marketplace, eds. B. Piqueras-Fiszman
and C. Spence, pp. 37–58. New York, NY: Woodhead Publishing.
Lim, J., Fujimaru, T., and Linscott, T. D. 2014. The role of congruency in taste-odor
interactions. Food Quality and Preference, 34, 5–13.
Lim, J. and Johnson, M. B. 2011. Potential mechanisms of retronasal odor referral to the
mouth. Chemical Senses, 36, 283–9.
Lim, J. and Johnson, M. B. 2012. The role of congruency in retronasal odor referral to
the mouth. Chemical Senses, 37, 515–22.
Loper, H. B., La Sala, M., Dotson, C., and Steinle, N. 2015. Taste perception, associated
hormonal modulation, and nutrient intake. Nutrition Reviews, 73, 83–91.
Luckett, C. R., Meullenet, J.-F., and Seo, H.-S. 2016. Crispness level of potato chips
affects temporal dynamics of flavor perception and mastication patterns in adults of
different age groups. Food Quality and Preference, 51, 8–19.
Luckett, C. R. and Seo, H.-S. 2017. The effects of both chewing rate and chewing dura-
tion on temporal flavor perception. Chemosensory Perception, 10, 13–22.
Mackey, A. O. and Valassi, K. 1956. The discernment of primary tastes in the presence
of different food textures. Food Technology, 10, 238–40.
 Chemical Senses and Flavor Perception 49

Mao, L., Roos, Y. H., Biliaderis, C. G., and Miao, S. 2017. Food emulsions as deliv-
ery systems for flavor compounds: A review. Critical Reviews in Food Science and
Nutrition, 57, 3173–87.
Massler, M. 1980. Geriatric nutrition: The role of taste and smell in appetite. Journal of
Prosthetic Dentistry, 43, 247–50.
Mattes, R. D. 2002. The chemical senses and nutrition in aging: Challenging old assump-
tions. Journal of the American Dietetic Association, 102, 192–6.
Mattes, R. D. 2009a. Is there a fatty acid taste? Annual Review of Nutrition, 29, 305–27.
Mattes, R. D. 2009b. Oral detection of short-, medium-, and long-chain free fatty acids
in humans. Chemical Senses, 34, 145–50.
McBurney, D. H., Collings, V. B., and Glanz, L. M. 1973. Temperature dependence of
human taste responses. Physiology & Behavior, 11, 89–94.
Mela, D. J., Langley, K. R., and Martin, A. 1994. No effect of oral or sample temperature
on sensory assessment of fat content. Physiology & Behavior, 56, 655–8.
Mese, H. and Matsuo, R. (2007). Salivary secretion, taste and hyposalivation. Journal of
Oral Rehabilitation, 34, 711–23.
Mickle, A. D., Shepherd, A. J., and Mohapatra, D. P. 2016. Nociceptive TRP channels:
Sensory detectors and transducers in multiple pain pathologies. Pharmaceuticals, 9, 72.
Miller, I. J. and Reedy, F. E. 1990. Variations in human taste bud density and taste inten-
sity perception. Physiology & Behavior, 47, 1213–9.
Miwa, T., Furukawa, M., Tsukatani, T., Costanzo, R. M., DiNardo, L. J., and Reiter, E.
R. 2001. Impact of olfactory impairment on quality of life and disability. Archives
of Otolaryngology—Head & Neck Surgery, 127, 497–503.
Mony, P., Tokar, T., Pang, P., Fiegel, A., Meullenet, J.-F., and Seo, H.-S. 2013. Temperature
of served water can modulate sensory perception and acceptance of food. Food
Quality and Preference, 28, 449–55.
Morrot, G., Brochet, F., and Dubourdieu, D. 2001. The color of odors. Brain and
Language, 79, 309–20.
Moskowitz, H. R. 1973. Effect of solution temperature on taste intensity in humans.
Physiology & Behavior, 10, 289–92.
Moskowitz, H. R. and Arabie, P. 1970. Taste intensity as a function of stimulus concen-
tration and solvent viscosity. Journal of Texture Studies, 1, 502–10.
Mozell, M. M. 1970. Evidence for a chromatographic model of olfaction. Journal of
General Physiology, 56, 46–63.
Murphy, C. and Cain, W. S. 1980. Taste and olfaction: Independence vs interaction.
Physiology & Behavior, 24, 601–5.
Murphy, C., Cain, W. S., and Bartoshuk, L. M. 1977. Mutual action of taste and olfac-
tion. Sensory Processes, 1, 204–11.
Negoias, S., Aszmann, O., Croy, I., and Hummel, T. 2013. Localization of odors can be
learned. Chemical Senses, 38, 553–62.
Nelson, G., Hoon, M. A., Chandrashekar, J., Zhang, Y., Ryba, N. J., and Zuker, C. S.
2001. Mammalian sweet taste receptors. Cell, 106, 381–90.
Noel, J., Habib, A.-R. R., Thamboo, A., and Patel, Z. M. 2017. Variables associated with
olfactory disorders in adults: A U.S. population-based analysis. World Journal of
Otorhinolaryngology—Head and Neck Surgery, 3, 9–16.
Nordin, S., Blomqvist, E. H., Olsson, P., Stjarne, P., and Ehnhage, A. 2011. Effects of
smell loss on daily life and adopted coping strategies in patients with nasal polyposis
with asthma. Acta Oto-Laryngologica, 131, 826–32.
50 Food Aroma Evolution

Nordin, S. and Bramerson, A. 2008. Complaints of olfactory disorders: Epidemiology,

assessment and clinical implications. Current Opinion in Allergy and Clinical
Immunology, 8, 10–5.
Nordin, S., Bramerson, A., and Bende, M. 2004. Prevalence of self-reported poor odor
detection sensitivity: The Skovde population-based study. Acta Oto-Laryngologica,
124, 1171–3.
Nordin, S., Bramerson, A., Millqvist, E., and Bende, M. 2007. Prevalence of parosmia:
The Skovde population-based studies. Rhinology, 45, 50–3.
Oka, Y., Butnaru, M., von Buchholtz, L., Ryba, N. J., and Zuker, C. S. 2013. High salt
recruits aversive taste pathways. Nature, 494, 472–5.
Olson, D. G., Caporaso, F., and Mandigo, R. W. 1980. Effects of serving temperature
on sensory evaluation of beef steaks from different muscles and carcass maturities.
Journal of Food Science, 45, 627–31.
Pangborn, R. M., Gibbs, Z. M., and Tassan, C. 1978. Effect of hydrocolloids on apparent
viscosity and sensory properties of selected beverages. Journal of Texture Studies,
9, 415–36.
Pangborn, R. M. and Szczensniak, A. S. 1974. Effect of hydrocolloids and viscosity
on flavor and odor intensities of aromatic flavor compounds. Journal of Texture
Studies, 4, 467–82.
Pangborn, R. M., Trabue, I. M., and Szczesniak, A. S. 1973. Effect of hydrocolloids on
oral viscosity and basic taste intensities. Journal of Texture Studies, 4, 224–41.
Parker, G. H. 1912. The relation of smell, taste and the common chemical sense in ver-
tebrates. Journal of the Academy of Natural Sciences of Philadelphia, 15, 221–34.
Pause, B. 2016. Chapter 52. Human chemosensory communication. In Springer
Handbook of Odor, ed. A. Buettner, pp. 987–1010, New York, NY: Springer.
Pellegrino, R., Drechsler, E., Hummel, C., Warr, J., and Hummel, T. 2017. Bimodal
odor processing with a trigeminal component at sub- and suprathreshold levels.
Neuroscience, 363, 43–9.
Pellegrino, R., Luckett, C. R., Shinn, S. E., Mayfield, S., Gude, K., Rhea, A., and Seo, H.-
S. 2015. Effects of background sound on consumers’ sensory discriminatory ability
among foods. Food Quality and Preference, 43, 71–8.
Pence, T. S., Reiter, E. R., DiNardo, L. J., and Costanzo, R. M. 2014. Risk factors for
hazardous events in olfactory-impaired patients. JAMA Otolaryngology—Head &
Neck Surgery, 140, 951–5.
Philipsen, D. H., Clydesdale, F. M., Griffin, R. W., and Stern, P. 1995. Consumer age
affects response to sensory characteristics of a cherry flavored beverage. Journal of
Food Science, 60, 364–8.
Piqueras-Fiszman, B., Alcaide, J., Roura, E., and Spence, C. 2012. Is it the plate or is it
the food? Assessing the influence of the color (black or white) and shape of the plate
on the perception of the food placed on it. Food Quality and Preference, 24, 205–8.
Porter, J., Craven, B., Khan, R., Chang, S.-J., Kang, I., Judkewitz, B., Volpe, J., Settles, G.,
and Sobel, N. 2007. Mechanisms of scent tracking in humans. Nature Neuroscience,
10, 27–9.
Pramudya, R. C. and Seo, H.-S. 2018a. Using Check-All-That-Apply (CATA) method
for determining product temperature-dependent sensory-attribute variations: A case
study of cooked rice. Food Research International, 105, 724–32.
Pramudya, R. C. and Seo, H.-S. 2018b. Influences of product temperature on emotional
responses to, and sensory attributes of, coffee and green tea beverages. Frontiers in
Psychology, 8, 2264.
 Chemical Senses and Flavor Perception 51

Prescott, J., Allen, S., and Stephens, L. 1984. Interactions between oral chemical irrita-
tion, taste, and temperature. Chemical Senses, 18, 389–404.
Radil, T. and Wysocki, C. J. 1998. Spatiotemporal masking in pure olfaction. Annals of
the New York Academy of Sciences, 855, 641–4.
Rawson, N. E., Gomez, G., Cowart, B., Brand, J. G., Lowry, L. D., Pribitkin, E. A., and
Restrepo, D. 1997. Selectivity and response characteristics of human olfactory neu-
rons. Journal of Neurophysiology, 77, 1606–13.
Rawson, N. E. and Yee, K. K. 2006. Transduction and coding. In Taste and Smell.
An Update. Advances in Oto-Rhino-Laryngology, eds. T. Hummel and A. Welge-
Lüssen, pp. 23–43. Basel: Karger.
Reinoso Carvalho, F., Wang, Q. (J.), Van Ee, R., Persoone, D., and Spence, C. 2017.
“Smooth operator”: Music modulates the perceived creaminess, sweetness, and bit-
terness of chocolate. Appetite, 108, 383–90.
Reinoso Carvalho, F., Wang, Q. (J.), Van Ee, R., and Spence, C. 2016. The influence
of soundscapes on the perception and evaluation of beers. Food Quality and
Preference, 52, 32–41.
Rentmeister-Bryant, H. and Green, B. G. 1997. Perceived irritation during ingestion of
capsaicin or piperine: Comparison of trigeminal and non-trigeminal areas. Chemical
Senses, 22, 257–66.
Rombaux, P., Potier, H., Markessis, E., Duprez, T., and Hummel, T. 2010. Olfactory
bulb volume and depth of olfactory sulcus in patients with idiopathic olfactory loss.
European Archives of Oto-Rhino-Laryngology, 267, 1551–6.
Roper, S. D. and Chaudhari, N. 2017. Taste buds: Cells, signals and synapses. Nature
Reviews Neuroscience, 18, 485–97.
Rosenstein, D. and Oster, H. 1988. Differential facial responses to four basic tastes in
newborns. Children Development, 59, 1555–68.
Ross, C. F. and Weller, K. 2008. Effect of serving temperature on the sensory attributes
of red and white wines. Journal of Sensory Studies, 23, 398–416.
Rozengurt, E. 2006. Taste receptors in the gastrointestinal tract. I. Bitter taste recep-
tors and alpha-gustducin n the mammalian gut. American Journal of Physiology-
Gastrointestinal and Liver Physiology, 291, G171–7.
Rozin, P. 1982. “Taste-smell confusions” and the duality of the olfactory sense. Perception
& Psychophysics, 31, 397–401.
Samant, S. S., Cho, S., Whitmore, A. D., Oliveira, S. B. S., Mariz, T. B., and Seo, H.-
S. 2016. The influence of beverages on residual spiciness elicited by eating spicy
chicken meat: Time-intensity analysis. International Journal of Food Science and
Technology, 51, 2406–15.
San Gabriel, A., Maekawa, T., Uneyama, H., and Toril, K. 2009. Metabotropic glutamate
receptor type 1 in taste tissue. American Journal of Clinical Nutrition, 90, 7435–65.
Santos, D. V., Reiter, E. R., DiNardo, L. J., and Costanzo, R. M. 2004. Hazardous events
associated with impaired olfactory function. Archives of Otolaryngology—Head &
Neck Surgery, 130, 317–9.
Schepers, R. J. and Ringkamp, M. 2010. Thermoreceptors and thermosensitive afferents.
Neuroscience and Biobehavioral Reviews, 34, 177–84.
Schifferstein, H. N. J. and Tanudjaja, I. 2004. Visualising fragrances through colours:
The mediating role of emotions. Perception, 33, 1249–66.
Schiffman, S. S., Sattely-Miller, E. A., Graham, B. G., Bennett, J. L., Booth, B. J.,
Desai, N., and Bishay, I. 2000. Effect of temperature, pH, and ions on sweet taste.
Physiology & Behavior, 68, 469–81.
52 Food Aroma Evolution

Schneider, R. A. and Schmidt, C. E. 1967. Dependency of olfactory localization on non-

olfactory cues. Physiology & Behavior, 2, 305–09.
Sclafani, A. 2004. The sixth taste? Appetite, 43, 1–3.
Seo, H.-S., Arshamian, A., Schemmer, K., Scheer, I., Sander, T., Ritter, G., and Hummel,
T. 2010. Cross-modal integration between odors and abstract symbols. Neuroscience
Letters, 478, 175–8.
Seo, H.-S. and Hummel, T. 2009. Effects of olfactory dysfunction on sensory evaluation
and preparation of foods. Appetite, 53, 314–21.
Seo, H.-S. and Hummel, T. 2011a. Chapter 3. Smell, taste, and flavor. In Food Flavors
Chemical, Sensory and Technological Properties, ed. H. Jeleń, pp. 35–63. New
York, NY: CRC Press.
Seo, H.-S. and Hummel, T. 2011b. Auditory-olfactory integration: Congruent or pleasant
sounds amplify odor pleasantness. Chemical Senses, 36, 301–9.
Seo, H.-S. and Hummel, T. 2014. Congruent sound can modulate odor pleasantness.
Chemical Senses, 39, 215–28.
Seo, H.-S. and Hummel, T. 2016. Chapter 47. Cross-modal integration in olfactory per-
ception. In Springer Handbook of Odor, ed. A. Buettner, pp. 897–918. New York,
NY: Springer.
Seo, H.-S., Iannilli, E., Hummel, C., Okazaki, Y., Buschhüter, D., Gerber, J., Krammer, G.
E., Van Lengerich, B., and Hummel, T. 2013. A salty-congruent odor enhances salti-
ness: Functional magnetic resonance imaging study. Human Brain Mapping, 34, 62–76.
Seo, H.-S., Jeon, K. J., Hummel, T., and Hummel, T. 2009. Influences of olfactory impair-
ment on depression, cognitive performance, and quality of life in Korean elderly.
European Archives of Oto-Rhino-Laryngology, 266, 1739–45.
Seubert, J., Freiherr, J., Frasnelli, J., Hummel, T., and Lundström, J. N. 2013.
Orbitofrontal cortex and olfactory bulb volume predict distinct of olfactory perfor-
mance in healthy subjects. Cerebral Cortex, 23, 2448–56.
Seubert, J., Kellermann, T., Loughead, J., Boers, F., Brensinger, C., Schneider, F., and
Habel, U. 2010. Processing of disgusted faces is facilitated by odor primes: A func-
tional MRI study. Neuroimage, 53, 746–56.
Sewards, T. V. 2004. Dual separate pathways for sensory and hedonic aspects of taste.
Brain Research Bulletin, 62, 271–83.
Shankar, M. U., Levitan, C. A., and Spence, C. 2010. Grape expectations: The role of cog-
nitive influence on color-flavor interactions. Conscious and Cognition, 19, 380–90.
Shepherd, G. M. 2006. Smell images and the flavour system in the human brain. Nature,
444, 316–21.
Silberstein, S. D. 2003. Neurotoxins in the neurobiology of pain. Journal of Head and
Face Pain, 43, 2–8.
Slack, J. P. 2016. Chapter 21. Molecular pharmacology of chemesthesis. In Chemosensory
Transduction. The Detection of Odors, Tastes, and Other Chemostimuli, eds. F.
Zufall and S. D. Munger, pp. 375–91. New York, NY: Academic Press.
Small, D. M. 2006. Central gustatory processing in humans. In Taste and Smell. An
Update. Advances in Oto-Rhino-Laryngology, eds. T. Hummel, and A. Welge-
Lüssen, pp. 152–90. Basel: Karger.
Small, D. M., Gerber, J. C., Mak, Y. E., and Hummel, T. 2005. Differential neural
responses evoked by orthonasal versus retronasal odorant perception in humans.
Neuron, 48, 593–605.
Small, D. M., Jones-Gotman, M., Zatorre, R. J., Petrides, M., and Evans, A. C. 1997.
Flavor processing: More than the sum of its parts. Neuroreport, 8, 3913–7.
 Chemical Senses and Flavor Perception 53

Small, D. M. and Prescott, J. 2005. Odor/taste integration and the perception of flavor.
Experimental Brain Research, 166, 345–57.
Small, D. M., Voss, J., Mak, Y. E., Simmons, K. B., Parrish, T., and Gitelman, D. 2004.
Experience-dependent neural integration of taste and smell in the human brain.
Journal of Neurophysiology, 92, 1892–903.
Spector, A. C. 2000. Linking gustatory neurobiology to behavior in vertebrates.
Neuroscience & Biobehavioral Reviews, 24, 391–416.
Spence, C. 2011. Crossmodal correspondences: A tutorial review. Attention, Perception,
& Psychophysics, 73, 971–5.
Spence, C. 2012. Auditory contributions to flavour perception and feeding behaviour.
Physiology & Behavior, 107, 505–15.
Spence, C. 2015a. Eating with our ears: Assessing the importance of the sounds of con-
sumption on our perception and enjoyment of multisensory flavour experiences.
Flavour, 4, 3.
Spence, C. 2015b. Multisensory flavor perception. Cell, 161, 24–35.
Spence, C., Levitan, C. A., Shankar, M. U., and Zampini, M. 2010. Does food color
influence taste and flavor perceptions in humans? Chemosensory Perception, 3,
68–84.
Spence, C. and Shankar, M. U. 2010. The influence of auditory cues on the perception of,
and responses to, food and drink. Journal of Sensory Studies, 25, 406–30.
Spence, C., Wan, X., Woods, A., Velasco, C., Deng, J., Youssef, J., and Deroy, O. 2015.
On tasty colours and colourful tastes? Assessing, explaining, and utilizing crossmo-
dal correspondences between colours and basic tastes. Flavour, 4, 23.
Stafford, L. D., Fernandes, M., and Agobiani, E. 2012. Effects of noise and distraction
on alcohol perception. Food Quality and Preference, 24, 218–24.
Steen, I., Waehrens, S. S., Petersen, M. A., Münchow, M., and Bredie, W. L. P. 2017.
Influence of serving temperature on flavour perception and release of Bourbon
Caturra coffee. Food Chemistry, 219, 61–8.
Steiner, J. E. 1974. Discussion paper: Innate, discriminative human facial expressions
to taste and smell stimulation. Annals of the New York Academy of Sciences, 237,
229–33.
Stevens, D. R., Seifert, R., Bufe, B., Müller, F., Kremmer, E., Gauss, R., Meyerhof, W.,
Kaupp, U. B., and Lindermann, B. 2001. Hyperpolarization-activated channels
HCN1 and HCN4 mediate responses to sour stimuli. Nature, 413, 631–5.
Stevenson, R. J. 2010. An initial evaluation of the function of human olfaction. Chemical
Senses, 35, 3–20.
Stevenson, R. J. and Boakes, R. A. 2004. Sweet and sour smells: Learned synaesthesia
between the senses of taste and smell. In The Handbook of Multisensory Processing,
eds. G. A. Calvert, C. Spence, and B. E. Stein, pp. 69–83. Cambridge, MA: MIT
Press.
Stevenson, R. J., Oaten, M. J., and Mahmut, M. K. 2011. The role of taste and oral
somatosensation in olfactory localization. Quarterly Journal of Experimental
Psychology, 64, 224–40.
Stillman, J. A. 1993. Color influences flavor identification in fruit-flavored beverages.
Journal of Food Science, 58, 810–2.
Stokes, C. N., O’Sullivan, M. G., and Kerry, J. P. 2016. Assessment of black coffee
temperature profiles consumed from paper-based cups and effect on affective and
descriptive product sensory attributes. International Journal of Food Science &
Technology, 51, 2041–8.
54 Food Aroma Evolution

Stolle, T., Grondinger, F., Dunkel, A., Meng, C., Médard, G., Kuster, B., and Hofmann,
T. 2017. Salivary proteome patterns affecting human salt taste sensitivity. Journal of
Agricultural and Food Chemistry, 65, 9275–86.
Stone, H. and Oliver, S. 1966. Effect of viscosity on the detection of relative sweetness
intensity of sucrose solutions. Journal of Food Science, 31, 129–34.
Tarrega, A., Yven, C., Semon, E., and Salles, C. 2008. Aroma release and chewing activ-
ity during eating different model cheeses. International Dairy Journal, 18, 849–57.
Temussi, P. A. 2009. Sweet, bitter and umami receptors: A complex relationship. Cell,
34, 296–302,
Tomchik, S. M., Berg, S., Kim, J. W., Chaudhari, N., and Roper, S. D. 2007. Breadth
of tuning and taste coding in mammalian taste buds. Journal of Neuroscience, 27,
10840–8.
Toyono, T., Seta, Y., Kataoka, S., Harada, H., Morotomi, T., Kawano, S., Shigemoto,
R., and Toyoshima K. 2002. Expression of the metabotropic glutamate receptor,
mGluR4a, in the taste hairs of taste buds in rat gustatory papillae. Archives of
Histology and Cytology, 65, 91–6.
Tremblay, C. and Frasnelli, J. 2018. Olfactory and trigeminal systems interact in the
periphery. Chemical Senses, 43, 611–6.
Trivedi, B. P. 2012a. Neuroscience: Hardwired for taste. Nature, 486, S7–S9.
Trivedi, B. P. 2012b. Gustatory system: The finer points of taste. Nature, 486, S2–S3.
Trulsson, M. and Essick, G. K. 2010. Sensations evoked by microstimulation of sin-
gle mechanoreceptive afferents innervating the human face and mouth. Journal of
Neurophysiology, 103, 1741–7.
Ugawa, S., Minami Y., Guo, W., Saishin, Y., Takatuji, K., Yamamoto, T., Tohyama, M.,
and Shimada, S. 1998. Receptor that leaves a sour taste in the mouth. Nature, 395,
555–6.
Vallbo, A. B., Hagbarth, K.-E., Torebjörk, H. E., and Wallin, B. G. 1979. Somatosensory,
proprioceptive, and sympathetic activity in human peripheral nerves. Physiological
Reviews, 59, 919–57.
Vandenbeuch, A., Clapp, T. R., and Kinnamon, S. C. 2008. Amiloride-sensitive channels
in type 1 fungiform taste cells in mouse. BMC Neuroscience, 9, 1.
Ventanas, S., Mustonen, S., Puolanne, E., and Tuorila, H. 2010. Odour and flavour
perception in flavoured model systems: Influence of sodium chloride, umami com-
pounds and serving temperature. Food Quality and Preference, 21, 453–62.
Verhagen, J. V. and Engelen, L. 2006. The neurocognitive bases of human multimodal
food perception: Sensory integration. Neuroscience and Biobehavioral Reviews, 30,
613–50.
Viana, F. 2011. Chemosensory properties of the trigeminal system. ACS Chemical
Neuroscience, 19, 38–50.
Visschers, R. W., Jacobs, M. A., Frasnelli, J., Hummel, T., Burgering, M., and Boelrijk,
A. E. M. 2006. Cross-modality of texture and aroma perception is independent of
orthonasal or retronasal stimulation. Journal of Agricultural and Food Chemistry,
54, 5509–15.
Voets, T., Droogmans, G., Wissenbach, U., Janssens, A., Flockerzi, V., and Nilius, B.
2004. The principle of temperature-dependent gating in cold- and heat-sensitive
TRP channels. Nature, 430, 748–54.
Von Békésy, G. 1964. Olfactory analogue to directional hearing. Journal of Applied
Physiology, 19, 369–73.
 Chemical Senses and Flavor Perception 55

Von Hornbostel, E. M. 1931. Über Geruchshelligkeit. Pflügers Archiv für die gesamte
Physiologie des Menschen und der Tiere. European Journal of Physiology, 227, 517–38.
Walker, H. K. 1990. Chapter 61. Cranial nerve V: The trigeminal nerve. In Clinical
Methods: The History, Physical, and Laboratory Examinations, 3rd ed., eds. H. K.
Walker, W. D. Hall, and J. W. Hurst, pp. 318–21, Boston, MA: Butterworths.
Wang, Q. J., Wang, S., and Spence, C. 2016. “Turn up the taste”: Assessing the role of
taste intensity and emotion in mediating crossmodal correspondences between basic
tastes and pitch. Chemical Senses, 41, 345–56.
Weel, K. G. C., Boelrijk, A. E. M., Alting, A. C., van Mil, P. J. J. M., Burger, J. J.,
Gruppen, H., Voragen, A. G. J., and Smith, G. 2002. Flavor release and perception
of flavored whey protein gels: Perception is determined by texture rather than by
release. Journal of Agricultural and Food Chemistry, 50, 5149–55.
Welge-Lüssen, A., Dorig, P., Wolfensberger, M., Krone, F., and Hummel, T. 2011. A
study about the frequency of taste disorders. Journal of Neurology, 258, 386–92.
Welge-Lüssen, A., Drago, J., Wolfensberger, M., and Hummel, T. 2005. Gustatory stimu-
lation influences the processing of intranasal stimuli. Brain Research, 1038, 69–75.
Winston, J. S., Gottfried, J. A., Kilner, J. M., and Dolan, R. J. 2005. Integrated neural
representations of odor intensity and affective valence in human amygdala. Journal
of Neuroscience, 25, 8903–7.
Woods, A. T., Poliakoff, E., Lloyd, D. M., Kuenzel, J., Hodson, R., Gonda, H., Batchelor,
J., Dijksterhuis, G. B., and Thomas, A. 2011. Effect of background noise on food
perception. Food Quality and Preference, 22, 42–7.
Yan, K. S. and Dando, R. 2015. A crossmodal role for audition in taste perception. Journal
of Experimental Psychology: Human Perception and Performance, 41, 590.
Yasumatsu, K., Ogiwara, Y., Takai, S., Yoshida, R., Iwatsuki, K., Torii, K., Margolskee,
R. F., and Ninomiya, Y. 2012. Umami taste in mice uses multiple receptors and
transduction pathways. Journal of Physiology, 590, 1155–70.
Zampini, M., Sanabria, D., Phillips, N., and Spence, C. 2007. The multisensory per-
ception of flavor: Assessing the influence of color cues on flavor discrimination
responses. Food Quality and Preference, 18, 975–84.
Zampini, M., Wantling, E., Phillips, N., and Spence, C. 2008. Multisensory flavor per-
ception: Assessing the influence of fruit acids and color cues on the perception of
fruit-flavored beverages. Food Quality and Preference, 19, 335–43.
Zellner, D. A., McGarry, A. M., Mattern-McClory, R., and Abreu, D. 2008. Masculinity/
femininity of fine fragrances affects color-odor correspondences: A case for cogni-
tions influencing cross-modal correspondences. Chemical Senses, 33, 211–22.
Zhao, G. Q., Zhang, Y., Hoon, M. A., Chandrashekar, J., Erlenbach, I., Ryba, N. J., and
Zuker, C. S. 2003. The receptors for mammalian sweet and umami taste. Cell, 115,
255–66.
Zhou, W. and Chen, D. 2009. Sociochemosensory and emotional functions. Psychological
Science, 20, 1118–24.
 Chapter 4
Aroma Compounds (Description,
Biosynthesis, and Regulation)
Mónika Valdenegro Espinoza, María Fernanda
Flores Echeverría, and Lida Fuentes Viveros

CONTENTS

4.1 Introduction 58
4.1.1 Concept of Flavor: Taste and Aroma 58
4.1.2 Impact Compounds of Natural Aromas: Odor Threshold 59
4.2 Chemistry and Organoleptic Properties of Volatile Aroma Compounds 61
4.2.1 Volatile Compounds 61
4.2.2 Esters 62
4.2.3 Lactones 63
4.2.4 Terpenoids 65
4.2.5 Aldehydes and Ketones 66
4.2.5.1 Aldehydes 66
4.2.5.2 Ketones 67
4.2.6 Phenylpropenes and Other Aromatic Derivatives 67
4.2.7 Alcohols 68
4.2.8 Furans and Its Derivatives 68
4.2.9 Pyrazines 69
4.2.10 Sulfur Compounds 69
4.3 Biosynthesis 70
4.3.1 Fatty Acid-Derived and Lipophilic Compounds 70
4.3.1.1 Lipoxygenase Pathway (in Chain Oxidation) 71
4.3.1.2 α and β-Oxidation 73
4.3.2 Amino Acid Derivates 75
4.3.2.1 Acids, Alcohols, Aldehydes, Esters, Lactones, and N- and
S-Containing Flavor Molecules 75
4.3.2.2 Phenylpropenes and Other Aromatic Derivatives 76
4.3.3 Carbohydrate-Derived Flavor Compounds 78
4.3.3.1 Furanones and Pyrones 78
4.3.3.2 Terpenoids 79
4.3.3.3 Apocarotenoids 80
References 81

57
58 Food Aroma Evolution

4.1 INTRODUCTION

4.1.1 Concept of Flavor: Taste and Aroma

The acceptance of a food depends on many factors, which highlight their sensory proper-
ties such as color, appearance, taste, aroma, texture, and even the sound that is generated
during chewing (Ponce, 2006; Singh et al., 2013; Parker, 2015; Forney and Song, 2018).
When food is consumed, the interaction of taste, odor, and textural feeling provide an
overall sensation which is best defined by the word “flavor.” The taste is the result of two
classes of compounds: some of them are nonvolatile compounds associated and respon-
sible for the taste (El Hadi et al., 2013) and others are aroma substances responsible for
odors. However, there are compounds which provide both sensations. When chemical
compounds interact with taste receptors, located in the taste buds of the tongue, four
important basic taste perceptions are provided: sour, sweet, bitter, and salty; the fifth
basic taste is stimulated by glutamate substances. Aroma substances are volatile com-
pounds, which are perceived by the odor receptor sites of the smell organ, that is, the
olfactory tissue of the nasal cavity. They reach the receptors when drawn in through the
nose (orthonasal detection) and via the throat after being released by chewing (retronasal
detection) (Taylor et al., 2000).
The concept of aroma substances, like the concept of taste substances, should be used
loosely, since a compound might contribute to the typical odor or taste of one food, while
in another food it might cause a faulty odor or taste, or both, resulting in an off-flavor.
The aroma of a food can provide various functions, among them can be linked emotion-
ally with past experiences, stimulate our appetite, or can alert us about the safety of such
foods, in the presence of a rancid product for example (Parker, 2015). The aroma of food
has several functions, not only conveying the essential character of the food and provid-
ing variety and interest to what we consume but also alerting us to rancid and unsafe
food, stimulating the appetite as well as providing an emotional link to past experiences.
The compounds that are responsible for aroma are highly volatile, low-molecular weight
compounds that are present in foods at low levels (Parker, 2015). The food habits of
people are determined to a large extent by the aroma and flavor of the products that they
consume and that allow their development and survival (Ponce, 2006). Innovations in the
knowledge of the generation of aromas and flavors have made the development of new
foods possible. These new food products will be accepted or rejected by the consumers
primarily based on their characteristics, aroma, and flavor, regardless of nutritional qual-
ity, toxicology, or their advantages.
Fruit quality includes both its preharvest development, such as changes in color,
flavor, and texture as fruits develop, grow, and ripen, as well as its maintenance
following harvest as the perishable tissues senesce (Ogundiwin et al., 2009). Flavor
consists both of the perception in the mouth (sweetness, acidity or bitterness) and of
the odor, produced by several volatile compounds. All plants are able to emit volatile
organic compounds (VOCs), and the content and composition of these molecules
show both genotypic variation and phenotypic plasticity (Maffei, 2010, Vrhovsek et
al., 2014). As aroma is one of the most appreciated fruit characteristics, volatile flavor
compounds are likely to play a key role in determining the perception and accept-
ability of products by consumers. Identification of key volatile flavor metabolites that
carry the unique character of the natural fruit is essential, as it provides the principal
sensory identity and characteristic flavor of the fruit (Cheong et al., 2010; El Hadi
et al., 2013).
 Aroma Compounds (Description, Biosynthesis, and Regulation) 59

4.1.2 Impact Compounds of Natural Aromas: Odor Threshold

Compounds defined as aromas are highly volatile substances, with low molecular weight
and present in low concentrations in food (ca. 10–15 mg/kg), and they have a funda-
mental effect on their quality and acceptance (Belitz et al., 2008; Parker, 2015; Forney
and Song, 2018, Pernolle and Briand, 2004). In general, however, a great variety of com-
pounds is often present in fruits and vegetables as well (Belitz et al., 2008, Rodríguez
et al., 2013). In processed foods this variety can be high, comprising a large number of
compounds, especially when foods are made by thermal processes alone (e.g., coffee) or
in combination with fermentation (e.g., bread, beer, cocoa, or tea) (Gasser and Grosch,
1988). Only a limited number are important for aroma (Belitz et al., 2008). A threshold
concentration is defined as the concentration at which an individual first perceives the
stimulus. For aroma, this can be either a detection threshold—the point at which the indi-
vidual can sense an aroma, or a recognition threshold—the point at which an individual
can recognize the aroma (Parker, 2015). The threshold value corresponds to the lowest
concentration of a compound that is just enough for the recognition of its odor, and is
called the odor threshold (recognition threshold) (Table 4.1). The volatile profile of most
foods contains many odor-active compounds, but very few of these actually give charac-
ter to the food. For example, cooked meat contains hundreds of odor-active compounds
(Cerny, 2012); many of which impart generic savory notes when roasted, toasted, or

TABLE 4.1 Odor Threshold Values in Water of Some Aroma

Compounds (20°C)
Compound Threshold Value (mL−1)
Ethanol 100
Maltol 9
Furfural 3.0
Hexanol 2.5
Benzaldehyde 0.35
Vanillin 0.02
Raspberry ketone 0.01
Limonene 0.01
Linalool 0.006
Hexanal 0.0045
2-Phenylethanal 0.004
Methylpropanal 0.001
Ethyl Butyrate 0.001
(+)-Nootkatone 0.001
(−)-Nootkatone 1.0
Filbertone 0.00005
Methylthiol 0.00002
2-Isobutyl-3-methoxypyrazine 0.000002
1-p-Menthene-8-thiol 0.0000002
Source: Data from Belitz et al. (2008). Aroma compounds. In: Food
Chemistry. Springer. Belitz H., Grosch W., Schieberle, P. (Eds.)
339–402, doi: 10.1007/978-3-540-69934-7_6.
60 Food Aroma Evolution

fried, but they are also present in snacks, fries, nuts, and so on. Others impart seemingly
unrelated aromas such as herbaceous or vegetal, rose, mushroom, and candy floss (cot-
ton candy). However, there are only a few compounds that impart a characteristic meaty
aroma, and the most common examples are 2-methyl-3-furanthiol and bis-(2-methyl-
3-furan) disulfide, called the “character impact compounds” of meat because, without
these, the food would be unrecognizable (Parker, 2015).
The threshold values are frequently determined by smelling (orthonasal value) and
tasting the sample (retronasal value). The threshold concentrations (values) for aroma
compounds are dependent on their vapor pressure, which is affected by both tempera-
ture and medium. The “aroma value” of a compound is calculated considering the ratio
between the concentration of the compound in the food and its odor threshold. The
evaluation of volatile compounds on the basis of the aroma value provides only a rough
pattern at first, and once the odor threshold of a compound and its concentration in the
extract have been determined, the odor-activity value (OAV) can be calculated. The OAV
is defined as the concentration of the aroma compound divided by its odor threshold
(Doty, 1991).
Flavor and aroma are important characteristics that contribute to the quality of dif-
ferent foods, especially fresh fruits and vegetables (Tomiyama et al., 2012; Pérez et al.,
1992; 1996a; 1996b). Forney and Song (2018) point out that “It is estimated that more
than 480 species of fruits and 600 species of vegetables are consumed worldwide.”
However, in Western markets, about 25 fruit species and 40 vegetable species domi-
nate, with an additional 80 or so “specialty” fruits and vegetables available seasonally
(CPMA, 2012). Each of these fruits and vegetables has a unique flavor and aroma pro-
file, with a physical and metabolic process that occurs as fruits and vegetables mature
both prior to and following harvest, as constituents responsible for flavor and aroma
continually alter.
The flavor and aroma of foods are physiological phenomena that are closely related;
however, the compounds responsible in each case have different physical and chemical
properties. In the first case, they are substances of higher molecular weight, nonvolatile,
soluble in water and in smaller numbers than those related to the aroma, which must nec-
essarily be volatile for reaching the olfactory centers. Both flavor and aroma are a human
perception of a complex combination of volatile, nonvolatile, and structural components
contributing to aroma and taste, as well as appearance and texture (Drewnowski, 1997).
Volatile compounds are responsible for the aroma of fruits and vegetables and provide the
unique flavor characteristics that distinguish different commodities. A desirable comple-
ment of volatile compounds is required to ensure consumer acceptance. Volatile aroma
compounds are perceived through the human sense of smell when these compounds are
detected by receptors on the olfactory epithelium, located in the nasal cavity. These nerve
receptors have a wide range of sensitivity depending on the compound and it is estimated
that there are about 1000 different olfactory receptors (Pernollet and Briand, 2004).
Signals from these receptors are interpreted by the brain, resulting in a perception of
aroma and flavor. Smelling volatile compounds through the nose (orthonasal) provides a
different perception of the aroma than when volatiles are perceived through the back of
the throat (retronasal), which occurs during chewing and swallowing (Voirol and Daget,
1987). Another important characteristic is the chiral nature of these compounds because
the aroma and taste chemoreceptors have the ability to distinguish between the various
enantiomeric forms (Ponce, 2006).
The chemical constituents of fruits and vegetables that provide the stimulus that
results in the perception of flavor and aroma are diverse. Volatile compounds are typically
 Aroma Compounds (Description, Biosynthesis, and Regulation) 61

analyzed by gas chromatography, which has identified over 8000 different volatile com-
pounds in foods (VCF Online, 2016; Song and Forney, 2008) and each commodity has
a unique volatile profile, which varies within a species depending on cultivar, maturity,
and other factors (Forney, 2001; Baldwin et al., 2007). In addition to primary volatile
compounds that are present in fruits and vegetables, secondary volatile compounds are
formed when plant tissues are disrupted by processes such as cutting or cooking (Kays
and Wang, 2000). Physical actions including bruising, cutting, chewing, freezing, and
heating result in cell rupture, the mixing of enzymes and substrates, and chemical and
physiological responses that initiate the production of a variety of compounds, many of
which contribute to flavor (Beaulieu and Baldwin, 2002).

4.2 CHEMISTRY AND ORGANOLEPTIC PROPERTIES

OF VOLATILE AROMA COMPOUNDS

Volatile aroma compounds are responsible for odor and contribute to the overall flavor
of the fresh and processed fruit (Brackmann et al., 1993). In the case of fruits and veg-
etables, the volatile compounds that determine the aroma and contribute to the flavor
are found in low concentrations (rarely exceeding 30 ppm), and the concentration of
individual volatile compounds can vary widely (Forney and Song, 2018). However, the
most abundant volatile compounds present in a product may not be the most important
contributors to the flavor. The odor threshold properties of individual compounds are
highly variable and this aspect is the reason that the most abundant volatile compounds
present in a product may not be the most important contributors to the flavor. Among
volatile compounds typically found in fresh fruits and vegetables, the odor threshold may
differ by 108-fold or more (Forney, 2001; Forney and Song, 2018). Although different
fruits often share many aromatic characteristics, each fruit has a distinctive aroma that
depends upon the combination of volatiles, the concentration and the perception thresh-
old of individual volatile compounds (Tucker, 1993; Misry et al., 1997).

4.2.1 Volatile Compounds

Volatile compounds present in fruit and vegetables are mainly comprised of diverse
classes of chemicals, including esters, alcohols, aldehydes, ketones, lactones, and ter-
penoids (Forney and Song, 2018). However, some sulfur compounds, such as S-methyl
thiobutanoate, 3-(methylthio) propanal, 2-(methylthio) ethyl acetate, 3-(methylthio)
ethyl propanoate, and 3-(methylthio) propyl acetate, also contribute to the flavor of fruit
such as melons (Cucumis melo L.) (Song and Forney, 2008). The contribution of each
compound to the specific aroma profile of each cultivar depends on the activity and
substrate specificity of the relevant enzymes in the biosynthetic pathway, the substrate
availability, the odor threshold above which the compound can be detected by smell, and
the presence of other compounds (Rizzolo et al., 2006). For example, more than 285
volatile aroma compounds have been reported from various mango cultivars, including
monoterpenes, sesquiterpenes, esters, aldehydes, ketones, alcohols, carboxylic acids, ali-
phatic hydrocarbons, and aromatics (Preethi et al., 2014, Uji et al., 2015). Hydrocarbon
monoterpenes and sesquiterpenes contribute 70–90% of the total volatile aroma com-
pounds (Winterhalter, 1991). Examples of different groups of volatile compounds can be
observed in Figures 4.1 through 4.4.
62 Food Aroma Evolution

FIGURE 4.1 Examples of odor-active compounds containing carbon, hydrogen, and

oxygen. (Reprinted and modified with permission from Parker J.K. (2015). Introduction
to aroma compounds in foods. In: J.K. Parker, S. Elmore, and L. Methven (Eds.), Flavour
Development, Analysis and Perception in Food and Beverages. Woodhead Publishing:
Cambridge.)

4.2.2 Esters

Esters are formed by the esterification of carboxylic acid derivatives and alcohols. Esters
are the predominant volatile compound in many fruits (for example melons, apples, pine-
apples, and strawberries) and typically contribute fruity aromas and flavors. Esters also
contribute to the more delicate aromas found in cured ham (Theron et al., 2010) and some
cheeses. Ethyl butanoate and ethyl hexanoate are key odorants in Parmigiano Reggiano
(Qian and Reineccius, 2003) and blue cheese (Qian et al., 2002). Fruits and vegetables are
able to utilize a wide variety of these substrates, resulting in a vast array of esters having
a variety of distinctive aromas. In apples, alkyl esters comprise about 90% of the volatile
compounds (Dixon and Hewett, 2000). The most abundant compound is ethyl acetate,
present in most ripe or ripening fruits. Gonzalez et al. (2009) showed in strawberries a
marked increase in the abundance of acetates, promoted by an increase in the alcohol
acyl transferase that brings about the esterification of acyl-CoAs and alcohols. In this
sense, ethyl esters are major components of fruit aroma, particularly in ripe fruit where
the production of ethanol has boosted their formation. Esters are the most abundant
volatile compounds emitted by apple and, together with α-farnesene, have been proposed
for cultivar classification (Young et al., 2005). Ethyl 2-methyl butanoate, 2-methyl butyl
acetate, and hexyl acetate contribute mostly to the characteristic aroma of “Fuji” apples,
while ethyl butanoate and ethyl 2-methyl butanoate are the active odor compounds in
“Elstar” apples, and ethyl butanoate, acetaldehyde, 2-methyl butanol, and ethyl methyl
propanoate in “Cox Orange” (Berger, 2007). The longer chain ethyl esters become soapy,
 Aroma Compounds (Description, Biosynthesis, and Regulation) 63

FIGURE 4.2 Examples of odor-active compounds, oxygen heterocycles, and phenols.

(Reprinted with permission from Parker J. K. (2015). Introduction to aroma compounds
in foods. In: J.K. Parker, S. Elmore, and L. Methven (Eds.), Flavour Development,
Analysis and Perception in Food and Beverages. Woodhead Publishing: Cambridge.)

cheesy, and waxy. Some esters can be quite characteristic of specific fruits: 3-methylbutyl
acetate is characteristic of pear or pear drops, allyl hexanoate, is typically in pineapple,
cis-3-hexenyl butanoate imparts the green leafy aroma of the parent alcohol, and the C9-
esters are important for melon aroma.

4.2.3 Lactones

Lactones are cyclic organic (or intramolecular) esters that are potent aroma compounds
formed from the corresponding hydroxy acid and contribute to the flavor of some
fruits and vegetables. Those based on a furan ring are γ-lactones (e.g., γ-octalactone
(or 4-octanolide) and γ-decalactone (4-decanolide)) and tend to impart peachy, creamy,
and coconut aromas. Consequently, they are very popular in tropical flavors; for exam-
ple, γ-decalactone is the major lactone in both peaches and nectarines (Engel et al.,
1988a) and has a threshold of 11 μg/kg in water. The δ-lactones, which are based on a
pyran ring, are less odor-active than their furanyl isomers (Parker, 2015). Both γ- and
δ-lactones contribute to the aroma of peach. Eduardo et al. (2010) identified six lac-
tones that are important to peach aroma: γ-decalactone, γ-dodecalactone, γ-octalactone,
δ-decalactone, γ-nonalactone, and δ-dodecalactone. γ-Decalactone is described as hav-
ing a “creamy, fruity, peach-like” odor. In celery, the three most potent odorants are the
phthalides 3-butylphthalide, sedanenolide, and sedanolide, the former being described
as “green spicy” and the latter two as “sweet spicy” (Kurobayashi et al., 2006). When
64 Food Aroma Evolution

FIGURE 4.3 Examples of odor-active nitrogen compounds. (Reprinted with permission

from Parker J. K. (2015). Introduction to aroma compounds in foods. In: J.K. Parker,
S. Elmore, and L. Methven (Eds.), Flavour Development, Analysis and Perception in
Food and Beverages. Woodhead Publishing: Cambridge.)

FIGURE 4.4 Examples of odor-active sulfur compounds. (Reprinted with permission

from Parker J. K. (2015). Introduction to aroma compounds in foods. In: J.K. Parker,
S. Elmore, and L. Methven (Eds.), Flavour Development, Analysis and Perception in
Food and Beverages. Woodhead Publishing: Cambridge.)
 Aroma Compounds (Description, Biosynthesis, and Regulation) 65

these compounds were added to chicken broth, they were found to enhance the per-
ceived intensities of “umami” and “sweet” flavor (Kurobayashi et al., 2008). Several lac-
tones were identified in an extract of sweet cream butter, of which δ-decalactone had the
highest OAV and was believed to contribute to the sweet cream aroma. γ-Nonalactone,
δ-decalactone, and the two unsaturated lactones (5-hydroxyoct-2-enoic acid lactone
and 5-hydroxydec-2-enoic acid lactone) were found to have relatively high OAVs in milk
chocolate (Schnermann and Schieberle, 1997). All of them were also found in cocoa, but
the 5-hydroxydec-2-enoic acid lactone had been used in the production of the chocolate.
Jasmine lactone provides a floral petal-like aroma to green tea (Katsuno et al., 2014), and
lactones also make a significant contribution to the volatile profile of Bourbon whiskey,
with δ-nonalactone having a FD factor of 2048, and cis-3-methyl-4-octanolide (which is
also known as whiskey lactone) and γ-decalactone also contributing to the aroma. The
odor thresholds of lactones decrease significantly as the number of constituent carbons
increases (Parker, 2015; Forney and Song, 2018). Sensory important lactones usually pos-
sess 8–12 carbon atoms and some of them are very potent flavor components for a variety
of fruits (Basear and Demirci, 2007). All lactones originate from their corresponding 4‐
or 5‐hydroxy carboxylic acids, which are formed by either (i) reduction of oxo acids by
nicotinamide adenine dinucleotide (NAD)‐linked reductase, (ii) hydration of unsaturated
fatty acids, (iii) epoxidation and hydrolysis of unsaturated fatty acids, or (iv) reduction of
hydroperoxides (Schöttler and Boland, 1996; Schwab et al., 2008).

4.2.4 Terpenoids

Terpenoids are the largest and most diverse group of volatile compounds that contrib-
ute to fruit and vegetable flavor, constituting one of the most diverse families of natural
products, with over 40,000 different molecular structures (Schwab et al., 2008; Forney
and Song, 2018). Some nonvolatile terpenoids are recognized as phytohormones—gib-
berellins, abscisic acid, brassinosteroids—involved in important plant processes, such
as membrane structure, photosynthesis, redox chemistry, and growth regulation, that
regulate fruit development and have relevant importance in non-climacteric fruit ripen-
ing (Croteau et al., 2000; Cherian et al., 2014). The volatile terpenoids—monoterpenoids
(C10) and sesquiterpenoids (C15)—have been associated with the flavor profiles of most
fruits and the scent of flowers at varying levels. Citrus fruit aroma consists mostly of
these terpenes, which accumulate in specialized oil glands in the flavedo (the external
part of the peel) and oil bodies in the juice sacs. The monoterpene R-limonene normally
accounts for over 90% of the essential oils of the citrus fruit (Weiss, 1997). The sesquiter-
penes valencene and α‐ and β-sinensal, although present in minor quantities in oranges,
play an important role in the characteristic flavor and aroma of an orange fruit (Vora
et al., 1983; Weiss, 1997; Maccarone et al., 1998). Nootkatone, a putative derivative of
valencene, is a small fraction of the essential oils, but has a dominant role in the flavor
and aroma of grapefruit (Shaw and Wilson, 1981), while the monoterpene S-linalool was
found to be an important general strawberry aroma compound (Larsen and Poll, 1992;
Aharoni et al., 2004, Van de Poel et al., 2014) and is found in many other fruits including
peaches, guavas, nectarines, papayas, mangoes, passion fruits, tomatoes, litchi, oranges,
prickly pears, and koubos (Flath and Takahashi, 1978; Idstein et al., 1985; Bernreuther
and Schreier, 1991; Visai and Vanoli, 1997; Ong and Acree, 1998; Baldwin et al., 2000;
Ninio et al., 2003). The combination of the monoterpenes geraniol, citronellol, and rose
oxide is a key component of the characteristic aroma of aromatic Muscat grapes as well
66 Food Aroma Evolution

as the special scent of roses (Dunphy and Allcock, 1972; Bayrak, 1994; Luan et al.,
2005). C13 norisoprenoids, which are derived from carotenoids, are the most abundant
volatiles in raspberry fruit and comprised 64% to 94% of the total volatile content among
nine raspberry genotypes (Forney et al., 2015; Forney and Song, 2018).
Primary terpenes contributing significantly to aroma as determined by gas chro-
matography–olfactory detection (GCO) include myrcene, terpinolene, sabinene, and
1,3,8-p-menthatriene (Fukuda et al., 2013), which have “herbaceous and woody,” “sweet
and piney,” “woody and spicy,” and “camphoraceous and herbal” aroma notes, respec-
tively (Forney and Song, 2018). The terpenoid geosmin is responsible for the “earthy”
aroma found in red beet (Acree et al., 1976). In tomato, the terpenoids geranial, β-ionone,
β-damascenone, and 6-methyl-5-heptene-2-one contribute “fruity” and “floral” aromas
(Klee and Tieman, 2013).

4.2.5 Aldehydes and Ketones

4.2.5.1 Aldehydes
Aldehydes and ketones are organic compounds which incorporate a carbonyl functional
group, C=O. The carbon atom of this group has two remaining bonds that may be occu-
pied by hydrogen or alkyl or aryl substituents. If at least one of these substituents is
hydrogen, the compound is an aldehyde; if neither is hydrogen, the compound is a ketone
(Azzara and Campbell, 1992; Schwab et al., 2008; El Hadi et al., 2013; Forney and Song,
2018). Aldehydes are common components of many foods or flavorings, having an inter-
esting low odor threshold. The straight-chain unbranched aldehydes are present in differ-
ent food and plant tissues. Therefore, acetaldehydes impart fruity ester notes and are one
of the main components of flavoring of fruit, whereas propanal, butanal, and pentanal
(C3 –C5 aldehydes) tend to have a rather chemical/malty/green note of hard define (Foney
and Song, 2018). Many volatile aldehydes have remarkable odor properties (Van Gemert,
2000). This kind of compound can be formed from alcohols and other precursors by oxi-
dation processes, so the changes in aroma properties linked to oxidation are very often
related to the formation of aldehydes (Azzara and Campbell, 1992).
Many aldehydes with two double bonds have low odor thresholds, for example, the
2,4-alkadienals have great importance in fried aromas and have some characteristic fried
notes, for example, spicy touches could be present in fried chicken (Gasser and Grosch,
1990) or in cooked beef Gasser and Grosch, 1988). Also, the aroma threshold in water
is 0.2 µg k−1 (Belitz et al., 2004). Another close series of aldehydes, trans-4,5-epoxy-(E)-
2-alkenals are frequently founded in different foods. They all have a metallic odor being
the most potent trans-4,5-epoxy-(E)-2-decenal in soy milk (Kaneko et al., 2011), potato
chips (Kasuga et al., 2008), and in black tea (Kumazawa et al., 2008).
Saturated and unsaturated volatile C6 and C9 aldehydes and alcohols are key con-
tributors to the characteristic flavors of fruits, vegetables, and green leaves (Schwab et al.,
2008; El Hadi et al., 2013), being widely used as food additives that grants a characteris-
tic “fresh green” odor. Aldehydes and alcohols are found ubiquitously in the plant king-
dom at high concentrations, and they are derived from the degradation of branched-chain
and aromatic amino acids or methionine constitute (Schwab et al., 2008). The C6 alde-
hydes hexanal, (Z)-3-hexenal, and (E)-2-hexenal contribute “green” aroma notes, which
are emitted when tissues are disrupted by cutting or chewing in both vegetables and
fruits. These aldehydes are important flavor components of many fruits and vegetables
including apples (Defilippi et al., 2005; Villatoro et al., 2008), oranges (Gonzalez-Mas
 Aroma Compounds (Description, Biosynthesis, and Regulation) 67

et al., 2011), raspberries (Klesk et al., 2004), peaches (Engel et al., 1988; Aubert et al.,
2003; Aubert and Milhet, 2007; Wang et al., 2009), strawberries (Jetti et al., 2007),
cherries, tomato, cucumber, and spinach (Belitz et al., 2009; Forney and Song, 2018).
In some cases, hexanal and related C6 aldehydes decreased as fruit matures on the plant
(Engel et al., 1988). The C9 aldehydes contribute to green-bean and cut-grass impres-
sions in foods when assessed alone and to fresh green aromas; (E,Z)-2,6-nonadienal and
(E)-2-nonenal are key aroma components of cucumber, contributing “cucumber-like”
and “green, fatty” aromas (Schieberle et al., 1990), and (Z)-3-nonenal contributes to
the “green apple” note in apples (Belitz et al., 2009). Benzaldehyde is also an impor-
tant aroma compound in cherry and other stone fruit (Belitz et al., 2009; Forney and
Song, 2018). Aldehydes containing an aromatic ring, such as benzaldehyde, are pres-
ent in cherries and almonds; phenylacetaldehyde (rose and honey) and cinnamaldehyde
are important components of foods and flavorings. One example of these is vanillin
(4-hydroxy-3-metoxybenzaldehyde).

4.2.5.2 Ketones
This type of compound contributes characteristic aromas to different types of fruits and
vegetables, especially C5, C7, and C8 ketones (Forney and Song, 2018). The straight-chain
methylketones, that contains one carbonyl group in the 2-position, like 2-heptanone,
impart both a blue cheese and fruit pear aroma, while 3-octanone has earthy, mushroom
notes (Cho et al., 2006). Another type of compound in this group are the α-dicarbonyl
compounds; 2,3 pentanedione and 2,3-butanedione or diacetyl have lower fat thresholds
and bring butter and creamy notes to many cooked foods. In fruits, this type of com-
pound contributes to floral aromas, like in grapefruit, where they bring a “geranium-like”
odor (Buettner and Schieberle, 2001). In raspberry, 1-(p-hydroxyphenyl)-3-butanone is a
great contributor to fruit flavor, but Klesk et al. (2004) showed in aroma extract dilution
analysis of “Meeker” raspberries that this type of compound was a minor contributor to
the aroma.

4.2.6 Phenylpropenes and Other Aromatic Derivatives

Phenylpropanoid and benzenoid volatile compounds contribute to the aromas and scents
of many plant species and play important roles in plant communication with defense
pathways (Dudareva and Pichersky, 2006; Knudsen and Gershenzon, 2006; Pichersky
et al., 2006; Naoumkina et al., 2010.). Benzyl alcohol derivative, 1,3,5-trimethoxyben-
zene, has been identified as a key component of the odor of Chinese rose (Yomogida,
1992). This volatile is an effective sedative and has been used as a cosmetic additive
(Shoji et al., 2000). The biosynthesis pathway is thought to begin with phloroglucinol
and includes three methylation steps. Many modern rose varieties synthesize a related
compound, 3,5-dimethoxytoluene (Flament et al., 1993), from orcinol (3,5-dihydroxy-
toluene) by two successive methylations. Vanillin (4-hydroxy-3-methoxybenzaldehyde) is
the most widely used flavor compound in the world. It is the principal flavor component
of the vanilla extract obtained from cured pods (beans) of the orchid Vanilla planifolia
Andrews. Vanillin accumulates in the secretion around the seeds in the mature fruits. A
unique secretory tissue composed of closely packed unicellular hairs is located in three
gaps between the placentas along the central fruit cavity. These cells seem to be respon-
sible for vanillin secretion (Joel et al., 2003). Vanilla extract is valued as a natural flavor,
but, because of its cost and limited availability, less than 1% of the annual world demand
68 Food Aroma Evolution

for vanillin is isolated from its natural source (Walton et al., 2003). Most of the vanillin
used by the flavor industry originates from chemical methods that use guaiacol, eugenol,
or lignin as starting materials (Rao and Ravishankar, 2000).

4.2.7 Alcohols

With respect to alcohol volatile compounds, this type of compound is important in alco-
holic beverages and in most fruits and vegetables but tends to play a minor role in aroma
and flavor. The release of volatile flavor materials from alcoholic beverages depends not
only on the concentration of volatiles in the solution but also by interactions between vol-
atiles through the presence of various nonvolatile materials and ethanol concentration.
Ethanol is the most abundant of the volatile compounds in wine and it can modify both
the sensory perception of aromatic attributes as well as the detection of volatile com-
pounds (Goldner et al., 2009). In another food, the presence of butanol has been reported
to contribute to an apple aroma (Young et al., 1996). In watermelon, the C9 compound
alcohol (Z,Z)-3,6-nonadienol is one of the compounds with major contribution to flavor,
being described as “watermelon, fruity, fresh, cucumber” aroma (Xisto et al., 2012). In
other fruits and vegetables, C6 compound alcohols such as hexanol, (Z)-3-hexenol and
(E)-2-hexenol bring “herbaceous” aromas (Belitz et al., 2009).

4.2.8 Furans and Its Derivatives

Furan and its derivatives are naturally occurring compounds found at very low levels in
many foods and drinks, and they are associated with the flavor of foods. These include
commercially prepared foods as well as homemade foods. Furans are a major class of
compounds formed during the Maillard reaction and their presence in foods is well docu-
mented (Maga, 1979). Literature data indicate multiple sources of furan formation origi-
nating from (i) thermal degradation/Maillard reaction reducing sugars, alone or in the
presence of amino acids, (ii) thermal degradation of certain amino acids, and the ther-
mal oxidation of (iii) ascorbic acid, (iv) polyunsaturated fatty acids, and (v) carotenoids
(Yaylayan, 2006, Pérez and Yaylayan, 2004). The primary source of furan in food is the
thermal degradation of carbohydrates, such as glucose, lactose, and fructose (Vranová
and Ciesarová, 2009). A wide range of compounds can be formed during thermal pro-
cessing of food, some of which are relevant for aroma (e.g., furfural), while others are of
great health concern (e.g., furan). The formation of furan under pyrolytic conditions has
been studied in simple model systems revealing more precursor classes, that is, (i) ascor-
bic acid and related compounds; (ii) Maillard type systems containing amino acids and
reducing sugars; (iii) lipid oxidation of unsaturated fatty acids or triglycerides; and (iv)
carotenoids (Pérez and Yaylayan, 2004; Becalski and Seaman, 2005; Yaylayan, 2006).
Furthermore, the effect of ionizing radiation on furan formation in real (apple and orange
juice) and model systems has been studied (Fan, 2005a,b). Furan and its derivatives were
identified in a small number of heat-treated foods back in the 1960s and 1970s. Furans
are important compounds contributing to fruit aroma. Those present in fresh produce
are mainly furanoid terpenes. One example is linalool oxide, which brings a floral herby
note, but in this type of product it is a signal of an oxidation process and loss of qual-
ity. In other foods like grapes, wine, and tea, some compounds like theaspirane, another
bicyclic furanoid derived from carotenoids, exist as four diastereoisomers (Collin et al.,
 Aroma Compounds (Description, Biosynthesis, and Regulation) 69

2011). Each of them has different aroma properties: camphoraceous notes (the 2R and
the 2S,5S isomers), blackcurrant (the 2R,5S isomer) or smells like naphthalene (the 2S,5R
isomer). Furaneol brings a “sweet, caramel, floral, strawberry-like” aroma and contrib-
utes to the flavor of fruits including strawberry, blackberry, raspberry, pineapple, mango,
and tomato (Du and Qian, 2008; Forney and Song, 2018). In strawberry, both furaneol
and mesifuran (2,5-dimethyl-4-methoxy-2H-furan-3-one) have been shown to increase
during fruit ripening (Pérez et al., 1996a, 1996b). Most of the furans are formed during
the thermal processing of ingredients. Those found in fresh produce tend to be furanoid
terpenes, such as linalool oxide, which brings a floral herby note but tends to appear dur-
ing storage and is indicative of oxidation.

4.2.9 Pyrazines

Pyrazines are heterocyclic nitrogen-containing compounds that contribute to flavor in

several vegetables and specifically in some processed products like wine (Ryona et al.,
2008), contributing “green bell peppers” notes (Forney and Song, 2018). Most pyrazines
are generated during the thermal processing of foods at temperatures >100°C. The simple
unsubstituted or monosubstituted pyrazines have a roasted, biscuity aroma and rela-
tively high aroma thresholds, but as substitution increases, the odor threshold decreases
(Parker, 2015). They have very strong odors. One example is 3-isobutyl-2-methoxypyr-
azine, reported to have an odor threshold of only 2 parts per 1012 parts of water (Buttery
et al. (1969) cited by Forney and Song (2018)). Hinterholzer et al. (1998) analyzed volatile
profiles in different raw vegetables, and they detected 3-isobutyl 2-methoxypyrazine in
green and red bell peppers and French beans. They also quantified the presence of 3-sec-
butyl 2-methoxypyrazyne in carrots, parsnips, and beets, and 3-isopropil 2- methoxypyr-
azine in peas, broad beans, asparagus, and cucumbers. The pyrazines found in uncooked
potatoes and vegetables are methoxy-substituted, and powerful odorants. For example,
2-methoxy-3-isobutylpyrazine is the character impact compound in green bell pepper and
is identified as the most potent odorant in raw French beans (Hinterholzer et al., 1998).
The homologous 2-isopropyl-3-methoxypyrazine is known as bean pyrazine because it
imparts earthy, pea, and beany notes to soy milk (Kaneko et al., 2011), earthy notes to
potato (Buttery and Ling, 1973), and was also found to be odor-active in parsley leaves
(Jung et al., 1992) and gravies containing vegetables (Christlbauer and Schieberle, 2009).

4.2.10 Sulfur Compounds

Volatile organic compounds (VOCs) comprise a wide diversity of low-molecular weight

secondary metabolites, with an appreciable vapor pressure under ambient conditions
(McGorrin, 2011). Although some VOCs are probably common to almost all plants,
others are specific to only one or a few related taxa. To the first type belong the so-called
“green leaf” volatiles (GLVs) because of their “fresh green” odor. This group comprises
short-chain (C6) acyclic aldehydes, alcohols, and their esters, produced by plants from
most taxa as a wound response via the enzymatic metabolism of polyunsaturated fatty
acids. However, species- or genus-specific VOCs have been described in some species, such
as the sulfur-containing VOCs of Alliaceae and Brassicaceae (Qualley and Dudareva,
2001). In these cases, many strong aromas are the response of sulfur-containing com-
pounds. Glucosinolates are sulfur-rich, anionic natural products that upon hydrolysis
70 Food Aroma Evolution

by endogenous thioglucosidases called myrosinases produce several different products

(e.g., isothiocyanates, thiocyanates, and nitriles) (Halkier and Gershenzon, 2006). They
contain sulfur groups and are present in numerous species belonging to the Brassicaceae
family (Verkerk et al., 2009, Agerbirk and Olsen, 2012). Chemically, glucosinolates are
composed of thiohydroximate-O-sulfonate group linked to glucose, and an alkyl, aral-
kyl, or indolyl side chain (R) (Barba et al., 2016). It is known that the enantiomers of chi-
ral flavor compounds often have different sensory properties (McGorrin, 2011). In some
cases, one chiral form may exhibit a lower flavor threshold relative to its epimer. In other
situations, the aroma may change flavor character between the two enantiomeric forms,
or shift in character from odor to odorless. The sensory properties of enantiomers of
sulfur volatile compounds have been described and compared (Bernreuther et al., 1997;
Engel et al., 2001).

4.3 BIOSYNTHESIS

Many plant flavor compounds are accumulated and biosynthesized in specialized ana-
tomical structures (Bagchi, 2000; Croteau et al., 2000; Schwab et al., 2008). Therefore,
these tissues contain many of the biosynthetic enzymes, showing a high expression of
genes involved in the production of such metabolites (Schwab et al., 2008). This section
describes the biogenetic origin of plant-derived flavor molecules, with emphasis on bio-
synthesis process described on fruit.

4.3.1 Fatty Acid-Derived and Lipophilic Compounds

The main part of plant volatiles is synthesized from saturated and unsaturated fatty acids.
Fatty acid-derived straight-chain alcohols, aldehydes, acids, ketones, esters, and lactones
(C1–C20) are found extensively in the plant kingdom at high concentrations, being the
major building blocks of many straight-chain volatile flavor compounds of fresh fruit
flavors (Sanz et al., 1997; Schwab et al., 2008; Aragüez and Valpuesta, 2013; El Hadi et
al., 2013; Forney and Song, 2018). These compounds are fundamentally formed by three
biosynthesis processes: α-oxidation, β-oxidation, and the lipoxygenase (LOX) pathway
(Schwab and Schreier, 2002; Conde et al., 2008; Schwab et al., 2008; Espino-Díaz et
al., 2016). Aroma volatiles in intact fruit are formed via the β-oxidation biosynthetic
pathway, whereas when fruit tissue is disrupted, volatiles are formed via the lipoxygenase
pathway (Lea, 1995; Vogt et al., 2013; 160Wu et al., 2018). However, identification of
a number of oxylipin containing phosphatidylglycerols, monogalactosyldiacylglycerols,
and digalactosyldiacylglycerols reveal that direct oxidation of the fatty acid side chain in
acylglycerides is possible (Buseman et al., 2006; Schwab et al., 2008). Therefore, some
studies suggest that increased availability of fatty acid, along with higher membrane per-
meability, during fruit ripening might allow the LOX pathway to become active in intact
plant tissue and to function as an alternative to β-oxidation (Guadagni et al., 1971).
Many of the aliphatic esters, alcohols, acids, and carbonyls found in fruits are derived
from the oxidative degradation of linoleic and linolenic acids (Reineccius, 2006). In addi-
tion, some of the volatile compounds derived from the enzyme-catalyzed oxidative break-
down of unsaturated fatty acids may also be produced by autoxidation (Chan, 1987).
Autoxidation of linoleic acid produces 9,13-hydroperoxides, whereas linolenic acid also
produces 12,16-hydroperoxides (Berger, 2007). Hexanal and 2,4-decadienal are the
 Aroma Compounds (Description, Biosynthesis, and Regulation) 71

primary oxidation products of linoleic acid, while autoxidation of linolenic acid produces
2,4-heptadienal as the major product. Further autoxidation of these aldehydes leads to
the formation of other volatile products (Chan, 1987). It was reported that the constitu-
tive over-expression of a yeast (Saccharomyces cerevisiae) Δ9-desaturase, to catalyze the
conversion of linoleic acid to linolenic acid, in tomato produced a modified oxidation
pattern of lipids, with the result that the concentration of some short-chain alcohols and
aldehydes increased derived from fatty acids, such as (Z)-3-hexenol, 1-hexanol, hexanal,
and (Z)-3-hexenal (Wang et al., 1996). The over-expression of ω‐3 fatty acid desaturases
(FAD) from Brassica napus (BnFAD3) and potato (StFAD7) produced an increase in
linolenic/linoleic acid ratio in tomato leaves and fruits. In addition, the transgenic lines
presented an increase of the (Z)-3-hexenal/hexanal ratio and were more tolerant to chill-
ing (Domínguez et al., 2010).

4.3.1.1 Lipoxygenase Pathway (in Chain Oxidation)

The metabolism of polyunsaturated fatty acids, via the first LOX catalyzed step and the
subsequent reactions, is commonly known as the LOX pathway (Schwab et al., 2008; El
Hadi et al., 2013). Four enzymes have been described in this biosynthetic pathway: lipox-
ygenase (LOX), hydroperoxide lyase (HPL), 3Z,2E-enal isomerase, and alcohol dehydro-
genase (ADH) (Schwab et al., 2008; El Hadi et al., 2013). In intact fruit, enzymes in the
LOX pathway and their substrates have different subcellular locations, preventing the
formation of volatile compounds (El Hadi et al., 2013). During ripening, cell walls and
membranes may become more permeable, allowing the LOX pathway to become active
without tissue disruption (El Hadi et al., 2013), providing substrates for ester production
(De Pooter et al., 1983), and acting as an alternative to β-oxidation of fatty acids during
LOX biosynthetic activation by ripening. Conversely, when the fruit is homogenized, lin-
oleic and linolenic acid are oxidized to various C6 and C9 aldehydes by the effect of free
enzymes (Lea, 1995). Therefore, it is important that the studies of volatile compounds
by destructive techniques take into account the biochemical changes involved during the
extraction process.
LOX is a non-heme, iron-containing dioxygenase that catalyzes the regio- and enan-
tioselective dioxygenation of unsaturated fatty acids such as linoleic and α-linolenic acid
containing one or more 1Z,4Z-pentadienoic moieties (Liavonchanka and Feussner, 2006;
Schwab et al., 2008). Several LOX have been characterized in plants, being essential
components of the oxylipin pathway, converting fatty acids into hydroperoxides for the
generation of flavors such as 3Z-hexenol, 2E-hexenal, and 2E,6Z-nonadienal. The LOX
enzyme classification is on base to their specificity of the hydrocarbon backbone of fatty
acid oxygenation, that is, oxygenation at C9 (9-LOX) or at C13 (13-LOX) leads to the (9S)
and (13S)-hydroperoxy derivatives, respectively. Plant LOX have been grouped according
to their sequence similarity into two gene sub-families. Enzymes carrying no plastidic
transit peptide (type-1 LOX) show a high sequence similarity (>75%) between them,
while the type-2 LOX have only a moderate sequence similarity (approximately 35%)
(Liavonchanka and Feussner, 2006; Schwab et al., 2008).
The action of the HPL enzyme involves the cleaving of fatty acid hydroperoxides
generated from the LOX reaction, resulting in the formation of ω-oxo acids and volatile
C6 and C9 aldehydes. Similar to LOX, HPL can be classified into two groups according
to substrate specificity (Noordermeer et al., 2001). HPL is a member of the cytochrome
P450 family CYP74B/C that acts on a hydroperoxy functionality in a lipid peroxide with-
out any co-factor, being responsible for the emission of small aldehydes that act as green
leaf volatiles (Lee et al., 2008). The study of HPL role in LOX pathway, shows that
72 Food Aroma Evolution

down-regulation of HPL in potato plants (Salas et al., 2005) induced an increase in LOX
activity but a decrease of most of the C6 volatile compounds. On the other hand, regard-
ing compartmentalization of these enzymes, the heterologous expression of grapevine
(Vitis vinifera) hydroperoxide lyase (VvHPL1) merged to green fluorescent protein (GFP)
in tobacco BY-2 cells show that the potential location of HPL is in plastids (Akaberi et
al., 2018).
The ADH enzyme catalyzed the metabolism of aldehydes (C6 and C9) to form the
corresponding alcohols. ADH genes related to aroma production are expressed and reg-
ulated during fruit ripening (Manríquez et al., 2006). Over-expression of the tomato
SlADH2 gene led to the improved flavor of the fruit by modifying the levels of short-
chain aldehydes and alcohols, particularly 3Z-hexenol (Speirs et al., 1998; Prestage et
al., 1999). On the other hand, when a tomato short-chain ADH (SlscADH1) was silenced
specifically in fruit, these tomatoes accumulated higher concentrations of C5 and C6 vola-
tile compounds of the LOX pathway (Moummou et al., 2012). In contrast, the grapevine
ADH (VvADH2), over-expressed and silenced in grape berries, didn’t affect the con-
tent of free or bound volatile compounds, except benzyl alcohol and 2-phenylethanol
(Torregrosa et al., 2008).
The higher contents of fatty acids and fatty acid-derived volatiles present in tomato
fruit, in parallel with the induced expression of TomloxC, HPL, ADH2, and increased
activities of LOX, ADH, and HPL, were observed in abscisic acid (ABA)-treated tomato
fruit. Promoter region analysis shows that the cis-acting elements involved in ABA
responsiveness (ABREs) can be found in the 2000 bp region upstream of TomloxC and
HPL, suggesting that the accumulation of fatty acid volatiles is regulated by ABA, among
other factors (Wu et al., 2018).
Some genetic efforts have been made at an aroma improvement breeding program for
apples (Dunemann et al., 2009; Marconi et al., 2018). Although, putatively, a quantita-
tive trait locus (QTLs) for a LOX candidate gene was mapped (Dunemann et al., 2009),
the authors suggest that it would need to be validated using more apple cultivars and spe-
cies to better understand the genetics of apple aroma and to develop molecular markers
involved in volatile biosynthesis for the development of a breeding program. Thus, big
data analysis correlating quality parameters and molecular markers has been useful to
differentiate a particular locality gene of apple varieties from other European locations
(Marconi et al., 2018).
In grapes, the RNA sequencing (RNA-Seq) analysis of berries treated with CPPU
[forchlorfenuron N-(2-chloro-4-pyridyl)-N-phenylurea], a synthetic cytokine-like plant
regulator that promotes grape berry set and development, shows that differentially
expressed genes (DEGs) were associated with the formation of volatile compounds. These
DEGs were associated with fatty acid degradation and biosynthesis, phenylpropanoid
metabolism and biosynthesis, and carotenoid biosynthesis genes, such as CCDs (carot-
enoid cleavage dioxygenase), LOX, GGDP reductase (geranylgeranyl diphosphate reduc-
tase), PAL (phenylalanine ammonia-lyase) (Wang et al., 2017).
With regard to volatile compound biosynthesis in bananas (Musa sp), genes that
encode enzymes for the important steps of aroma production include BanLOX, BanPDC,
BanHPL, BanBCAT, BanADH, and BanAAT (Beekwilder et al., 2004; Yang et al.,
2011a) A recent study has revealed genes involved in ester formation from amino acids,
saturated fatty acids, and unsaturated fatty acids such as lipoxygenases, transferases,
and acetyltransferases (Asif et al., 2014). During the storage of bananas, the expres-
sions of a subset of aroma biosynthetic genes including MaOMT1, MaMT1, MaGT1,
MaBCAT1, MaACY1, MaAGT1, and BanAAT were down-regulated when pre-stored
 Aroma Compounds (Description, Biosynthesis, and Regulation) 73

at 7°C compared to those at 22°C during the ripening process of bananas. Furthermore,
two transcriptional factors—MabZIP4 and MabZIP5—were found to be regulators of
gene expression from these aroma biosynthetic genes (Guo et al., 2018).

4.3.1.2 α and β-Oxidation

Although the degradation of straight-chain fatty acids by α and β-oxidation is an impor-
tant process for the formation of flavor molecules, many aspects of plant biosynthesis
must be understood (Schwab et al., 2008; El Hadi et al., 2013). The α-oxidation mecha-
nism involves enzymatic degradation of free fatty acids (C12 –C18) via one or two inter-
mediates producing C(n−1) long-chain fatty aldehydes and CO2 (Hamberg et al., 1999;
Schwab et al., 2008), described in the plant catalysis by an enzyme with dual function:
α-dioxygenase/peroxidase and NAD+ oxidoreductase (Saffert et al., 2000). On the other
hand, the β-oxidation results in the successive removal of C2 units (acetyl-CoA) from the
parent fatty acid (Goepfert and Poirier, 2007), being the primary biosynthetic process
providing alcohols and acyl coenzyme A (CoAs) for ester formation (Sanz et al., 1997).
Fatty acid acyl-CoA derivatives are converted to shorter chain acyl-CoAs by losing two
carbons in every round of the β-oxidation cycle, a process that requires NAD, flavinade-
nine dinucleotide (FAD) and free CoA. Acyl-CoAs are reduced by the action of acyl-CoA
reductase to aldehyde that in turn is reduced by ADH to alcohol for use by alcohol acyl
transferases (AAT) to produce esters (Bartley et al., 1985). Pear and apple aromas are two
classic examples of volatile formation through the β-oxidation pathway (Paillard, 1990).
The biosynthesis of lactones, key aroma components in fruits such as peach and nectar-
ine (γ-decalactone and γ-dodecalactone), pineapple (δ-octalactone), or coconut (Cocos
nucifera L.) (γ-octalatcone), is also associated with the β-oxidation pathway (Tressl and
Albrecht, 1986), without ruling out its association with the LOX pathway.
Aliphatic short and medium-chain aldehydes and alcohols have been described in
different plant parts, being probably formed by enzymatic reduction of the parent acyl-
CoAs (Flamini et al., 2007). Particularly, alcohols are less important as flavor mole-
cules due to their high odor thresholds in comparison with their aldehyde homologous.
Alternatively, alcohols can also be formed by ADH. This enzyme-mediated hydroge-
nation of aldehydes, and medium-chain aldehydes are intermediates of the α-oxidation
cycle, starting with common fatty acids (Hamberg et al., 1999). An ADH with specific
substrate preference has been isolated from melons (Manríquez et al., 2006). Specifically,
flavor ester production relies upon the supply of acyl-CoAs formed during β-oxidation
and alcohols. Alcohol acyl transferases (AAT) are capable of combining various alcohols
and acyl-CoAs, resulting in the synthesis of a wide range of esters, thus accounting for
the diversity of esters. Several AAT genes have been isolated and characterized in fruit
and vegetables (Aharoni et al., 2000; Beekwilder et al., 2004; El-Sharkawy et al., 2005;
Gonzalez et al., 2009; Balbontin et al., 2010). Aliphatic esters are emitted by vegetative
tissues and contribute to the aroma of nearly all fruits, being some of them responsible
for a particular fruit or floral aroma.
AAT enzymes catalyze the last step in ester formation by transacylation from an acyl-
CoA to an alcohol, for example, the biosynthesis of volatile butanoates and hexanoates
in strawberry fruit (Aharoni et al., 2000; Cumplido-Laso et al., 2012). Combinations
between different alcohols and acyl-CoAs will result in the formation of a range of esters
in different fruit species. The most likely precursors for the esters are lipids and amino
acids (Beekwilder et al., 2004). For example, in strawberry, the amino acid Ala has been
implicated in the formation of ethyl esters during ripening (Pérez et al., 1992). However,
it has been suggested that selectivity of enzymes preceding AATs in the biosynthetic
74 Food Aroma Evolution

pathway also determined the ester profile of fruits (Wyllie and Fellman, 2000). In addi-
tion, the oxidative degradation of linoleic and linolenic acids results in the production of
volatile aldehydes, which in turn are utilized by alcohol dehydrogenases to form alcohols.
The latter is subsequently converted to hexyl and hexenyl esters (Pérez et al., 1996; Shalit
et al., 2001).
The activity of AAT enzymes and gene expression pattern has been subject of several
investigations on extracts of various fruit species, including banana, strawberry, melon
(Cucumis melo), apple, grape, apricot, and mountain papaya (Vasconcellea pubescens)
(Aharoni et al., 2000; Wyllie and Fellman, 2000; Shalit et al., 2001; Pérez et al., 2002;
Beekwilder et al., 2004; Wang and De Luca, 2005; Gónzalez et al., 2009; González-
Aguero et al., 2009; Balbontin et al., 2010). In strawberry and banana, the AAT activity
showed a broad substrate specificity (alcohols and acyl-CoAs) and an activity depen-
dent on maturation (Beekwilder et al., 2004). In the last decades, fruit expressed genes
encoding enzymes with AAT activity have been isolated and characterized from straw-
berry, melon, banana, and mountain papaya (Aharoni et al., 2000; Yahyaoui et al., 2002;
Beekwilder et al., 2004; Balbontin et al., 2010). Aharoni et al. (2000) utilized cDNA
microarrays for gene expression profiling during strawberry (Fragaria ananassa) fruit
development and identified the SAAT gene, which showed a strong induction upon rip-
ening. Moreover, the recombinant SAAT enzyme could catalyze the formation of esters
typically found in strawberry, using aliphatic, medium-chain alcohols (e.g., octanol)
in combination with various chain length (up to C10 tested) acyl-CoAs as substrates.
The activity of SAAT was much lower for aromatic substrates, and no activity could be
detected with the monoterpene alcohol, linalool. Yahyaoui et al. (2002) reported on the
molecular and biochemical characterization of a ripening-induced and ethylene-regulated
AAT gene (CM-AAT1) from ripe melon fruit. As in the case of SAAT, the recombinant
CM-AAT1 enzyme was capable of producing esters from a wide range of alcohols and
acyl-CoAs. While CM-AAT1 and the SAAT proteins share only 21% sequence iden-
tity, they show a similar preference toward alcohols. The comparison of recombinant
AAT isolated from fruit of wild strawberry (Fragaria vesca) and banana (Musa sapien-
tum) with SAAT enzyme, showed that the substrate preference of recombinant enzymes
was not necessarily reflected in the representation of esters in the corresponding fruit
volatile profiles. The above suggests that the specific profile of a given fruit species is to
a significant extent determined by the supply of precursors (Beekwilder et al., 2004).
Furthermore, the obtention of transgenic petunia (Petunia hybrida) plants overexpress-
ing the SAAT gene showed that the volatile profile was found to be unaltered while the
expression of SAAT and the activity of AAT was detected in transgenic plants. Feeding of
isoamyl alcohol to explants of transgenic lines resulted in the emission of the correspond-
ing acetyl ester and confirmed that the availability of alcohol substrates is an important
parameter to consider for engineering volatile ester formation in plants (Beekwilder et al.,
2004). Esters have been described as the main volatile compounds produced during fruit
ripening of mountain papaya (Vasconcellea pubescens), and most of them are dependent
on ethylene. The transcript accumulation pattern and increase in AAT activity showed
that the VpAAT1 gene is expressed exclusively in fruit tissues and that a high level of
transcripts is accumulated during ripening, it is induced by ethylene and it is avoided by
1-methylcyclopropene (1-MCP) treatment (Balbontin et al., 2010). The data indicate that
VpAAT1 is involved in aroma formation and that ethylene plays a major role in regulat-
ing its expression. On other hands, the over-expression of VpAAT1 gene in yeasts and
in vitro assay with different precursors suggest that this enzyme has a preference for the
formation of benzyl acetate (Balbontin et al., 2010).
 Aroma Compounds (Description, Biosynthesis, and Regulation) 75

4.3.2 Amino Acid Derivates

Aldehydes and alcohols derived from the degradation of branched-chain and aromatic
amino acids or methionine are abundant plant volatiles. However, many aspects of their
biosynthesis pathways are still unknown. Many branched-chain esters give their charac-
teristic to many fruits; for example, 2-methyl-butyl acetate has a strong apple scent and is
associated with apple varieties that are rich in aroma such as “Gala,” “Fuji,” and “Golden
Delicious” (Dixon and Hewett, 2000; Holland et al., 2005); isoamyl acetate is one of the
key constituents of banana flavor (Surburg and Panten, 2005) and grants a strong fruity
odor described as similar to banana or pear; methyl 2-methyl butanoate determines the
characteristic aroma of prickly pear (Weckerle et al., 2001), while a combination of isoamy-
lacetate and 2-methyl-butyl acetate and other volatiles imparts the unique aroma of melons
(Beaulieu and Grimm, 2001; Jordan et al., 2001; Shalit et al., 2001). Branched amino acid
metabolism plays an important role not only in flavor and volatile biosynthesis but also as
an important nutrient for human health (Klee and Tieman, 2013).

4.3.2.1 Acids, Alcohols, Aldehydes, Esters, Lactones, and

N- and S-Containing Flavor Molecules
The knowledge of amino acids catabolism in microorganisms proposes three biochemical
reactions: (i) formation of 2-ketoacids by aminotransferases, (ii) decarboxylation to alde-
hydes, and (iii) reduction to 2-hydroxyacids (Marilley and Casey, 2004; Schwab et al.,
2008). Compounds derived from leucine such as 3-methylbutanal, 3-methylbutanol, and
3-methylbutanoic acid, as well as phenylacetaldehyde and 2-phenylethanol formed from
phenylalanine, are abundant in tomato, strawberry, and grape varieties, among other
fruits (Aubert et al., 2005). Alcohols and acids derived from amino acids can also be
esterified to compounds with a large impact on fruit odor, such as 3-methylbutyl acetate
and 3-methylbutyl butanoate in banana (Nogueira et al., 2003).
The isolation of gene encoding enzymes responsible for the direct decarboxylation of
phenylalanine from tomato, petunia, and rose was one of the first signs that alternative
catabolic pathways exist in plants (Kaminaga et al., 2006; Tieman et al., 2006; Schwab
et al., 2008). Studies of amino acid catabolism performed in tomato (Solanum lycoper-
sicum L., Solanaceae) fruit indicated that the catabolism of l-phenylalanine into aroma
volatiles is initially mediated by decarboxylation, followed by deamination (Tieman et
al., 2006). In contrast, in petunia (Petunia hybrida, Solanaceae) and rose (Rosa hybrida,
Rosaceae) petals, it has been observed that one enzyme is able both to decarboxylate and
to deaminate l-phenylalanine to release phenylacetaldehyde (Kaminaga et al., 2006),
which indicates that different biosynthetic routes can be used. Although the enzymes
display subtle differences in sequences and enzymatic properties, their down-regulation
led to reduced emission of 2-phenylacetaldehyde and 2-phenylethanol. Transgenic tomato
lines have shown that the over-expression of the amino acid decarboxylase resulted in
fruits with up to 10-fold increased levels of 2-phenylacetaldehyde, 2-phenylethanol, and
1-nitro-2-phenylethane (Tieman et al., 2006). The modulation of the emission of these
compounds can have different effects, observing that at low concentrations 2-phenyletha-
nol and 2-phenylacetaldehyde are associated with pleasant, sweet, flowery notes, while at
high levels, 2-phenylacetaldehyde is unpleasant (Tadmor et al., 2002).
The essential amino acids, l-phenylalanine, l-methionine, l-leucine, l-isoleucine,
and l-valine, are metabolized into aroma compounds in melon fruit (Gonda et al., 2010;
Gonda et al., 2013; Gonda et al., 2018), and the action of aminotransferases (AT) has
been suggested as a key step for generating the respective α-keto acids. In apple, BCAT,
76 Food Aroma Evolution

ArAT, and amino acid decarboxylases (AADC) were up-regulated during ripening and
further enhanced by ethylene treatment (Yang et al., 2011b). The ethylene treatment at the
pre-climacteric stage of apple growth showed an early burst of branched-chain volatiles,
including 2-methylbutanol, 2-methylbutylacetate, and butyl-2-methylbutanaote increase
(Song, 1994). In oriental sweet melon, it has been observed that ethylene regulated the
metabolism of branched-chain amino acids, including valine, leucine, and isoleucine,
as well as phenylalanine and cysteine, and enhanced the formation of aroma volatiles,
especially methyl-branched and aromatic esters (Li et al., 2016). The activities of amino-
transaminase BCAT, ArAT, and pyruvate dehydrogenase (PDH), as well as the expression
patterns of CmBCAT1 and CmArAT1, showed a clear ethylene regulation (Li et al., 2016).
On the other hand, a diverse catabolic pathway of amino acids to generate differ-
ent aromatic compounds has been described in onion (Allium cepa), garlic (A. sativum)
(Jones et al., 2004; Lanzotti, 2006), maize (Frey et al., 2000), cruciferous vegetables
such as mustard, broccoli, cauliflower, kale, turnips, collards, brussels sprouts, cab-
bage, radish, and watercress (Chen and Andreasson, 2001; Bak et al., 2006; Bones and
Rossiter, 2006).

4.3.2.2 Phenylpropenes and Other Aromatic Derivatives

Phenylpropanoid and benzenoid volatile compounds, primarily derived from phenylala-
nine, contribute to the aromas and scents of many plant species and play important roles
in plant communication with defense pathways (Dudareva and Pichersky, 2006; Knudsen
and Gershenzon, 2006; Pichersky et al., 2006; Naoumkina et al., 2010.). Several enzymes
that catalyze the final steps in the biosynthesis of these compounds have been isolated and
characterized; however, there is still much to understand about the early steps leading
to the formation of the benzenoid backbone (Beuerle and Pichersky, 2002; Schnepp and
Dudareva, 2006; Wildermuth, 2006; Schwab et al., 2008).
The biosynthesis of benzenoids from phenylalanine requires shortening of the carbon
skeleton side chain by a C2 unit, which can be carried out by β-oxidative or non-oxi-
dative pathway (Boatright et al., 2004). The existence of both routes has been dem-
onstrated by experiments with stable isotope-labeled precursors in different plants. In
tobacco (Nicotiana tabacum) leaves (Ribnicky et al., 1998), the studies suggested that
benzoic acid is produced from phenylalanine derived cinnamic acid via the β-oxidative
pathway, first yielding benzoyl-CoA, which can then be hydrolyzed by a thioesterase to
free benzoic acid. Conversely, the same experiments in Hypericum androsaemum cell
cultures (Ahmed et al., 2002), together with initial enzyme characterization, supported
the existence of a pathway for non-oxidative conversion of cinnamic acid to benzaldehyde
to obtain benzoic acid, which can be further converted to benzoyl-CoA (Beuerle and
Pichersky, 2002). On the other hand, in vivo isotope labeling and metabolic flux analysis
of the benzenoid network in petunia flowers revealed that both pathways yield benze-
noid compounds and that benzyl benzoate is an intermediate between l‐phenylalanine
and benzoic acid (Boatright et al., 2004). The generation of transgenic petunia plants
with reduced or eliminated expression of benzoyl-CoA—phenylethanol/benzyl alcohol
benzoyltransferase (BPBT) enzyme that uses benzoyl-CoA and benzyl alcohol to make
benzyl benzoate—showed that the decrease or elimination of benzyl benzoate forma-
tion decreased the endogenous pool of benzyl acid and methyl benzoate emission but
increased the emission of benzyl alcohol and benzylaldehyde, confirming the contribu-
tion of benzyl benzoate to benzoic acid formation (Orlova et al., 2006). The dilution
of 2H-5-phenylalanine suggests an alternative pathway from a different precursor than
phenylalanine, possibly phenylpyruvate (Schwab et al., 2008).
 Aroma Compounds (Description, Biosynthesis, and Regulation) 77

Phenylpropenes such as trans-anethole, eugenol, and isoeugenol are produced by

plants as defense compounds against animals and microorganisms and as floral attrac-
tants of pollinators (Schwab et al., 2008). Moreover, humans have used phenylpropenes
since antiquity as medicinal agents and for food preservation and flavoring (Gross et al.,
2002). Sweet basil (Ocimum basilicum) contains an enzyme that can use coniferyl ace-
tate and NADPH to form eugenol (Koeduka et al., 2006) and petunia (Petunia hybrida
cv. Mitchell) flowers possess an enzyme homologous to basil that also uses the same
substrates but catalyzes the formation of isoeugenol. These enzymes are phenylpropene
forming enzymes that belong to a structural family of NADPH dependent reductases that
includes pinoresinol/lariciresinol reductase, isoflavone reductase, and phenylcoumaran
benzylic ether reductase (Koeduka et al., 2006).
Phenylpropenes and benzoids are further subjected to methylation catalyzed by plant
O-methyltransferases (OMTs) (Ibrahim et al., 1998). Various OMTs involved in the bio-
synthetic pathways of floral scent components have been identified and characterized. For
example, S-adenosyl-l-methionine (iso)eugenol OMT, which catalyzes the methylation
of eugenol and isoeugenol to form the volatiles methyleugenol and isomethyleugenol,
has been isolated from Clarkia breweri (Wang et al., 1997). Eugenol OMT and chavi-
col OMT, which convert eugenol and chavicol to methyleugenol and methylchavicol,
respectively, have been identified in Ocimum basilicum (Lewinsohn et al., 2000; Gang
et al., 2002). Similarly, enzymatic activities able to convert trans-anole to trans-anethole
and chavicol to estagole have been demonstrated in Foeniculum vulgare tissues (Gross et
al., 2002, 2006). Transient over-expression in tobacco showed that OMT gene isolated
from apple (MdOMT1) utilized a range of phenylpropene substrates and catalyzed the
conversion of chavicol to estragole (Yauk et al., 2015). The same study showed that mul-
tiple transgenic apple lines (cv. Royal Gala) with reduced MdOMT1 expression produced
lower levels of methylated phenylpropenes, including estragole and methyleugenol indi-
cate that MdOMT1 is required for the production of methylated phenylpropenes in apple
and that phenylpropenes including estragole may contribute to ripe apple fruit aroma
(Yauk et al., 2015). A phloroglucinol O-methyltransferase that methylates the first step
to produce the intermediate 3,5-dihydroxyanisole has been isolated from rose petals, and
the two previously described orcinol O-methyltransferases catalyze the subsequent steps
(Wu et al., 2004).
As studies of the enzymes responsible for the synthesis of aromatic compounds prog-
ress, the possibilities of joining different substrates increase. Therefore, recently, it has
been shown that AATs from ripe strawberry (SAAT1) and tomato (SlAAT1) fruit can
also utilize p-coumaryl and coniferyl alcohols, indicating that ripening-related AATs are
likely to link volatile ester and phenylpropene production in many different fruits (Yauk
et al., 2017).
Methyl salicylate and methyl benzoate are common components of floral scent and
are believed to be important attractants of insect pollinators (Dobson, 1994; Dudareva
et al., 1998, 2000; Dudareva and Pichersky, 2000). Enzymes that catalyze the forma-
tion of methyl salicylate and methyl benzoate from salicylic acid (SA) and benzoic acid
(BA), respectively, have been characterized from flowers of Clarkia breweri, snapdragon
(Antirrhinum majus), petunia, Arabidopsis thaliana, and Stephanotis floribunda (Ross
et al., 1999; Murfitt et al., 2000; Negre et al., 2002; Pott et al., 2002; Chen et al.,
2003). While these enzymes use S-adenosyl-l-methionine as the methyl donor, as do
many previously characterized methyltransferases that act on a variety of substrates
(e.g., DNA, protein, phenylpropanoids), these SA and BA carboxyl methyltransferases
have primary amino acid sequences that show no significant sequence identity to other
78 Food Aroma Evolution

methyltransferases. Interestingly, a group of N-methyltransferases involved in the biosyn-

thesis of the alkaloid caffeine, including theobromine synthase, share sequence similarity
with the SA and BA carboxyl methyltransferases (D’Auria et al., 2003). These enzymes
were therefore grouped into a new class of methyltransferases designated the SABATH
methyltransferases, and this family now also includes jasmonic acid methyltransfer-
ase (Seo et al., 2001), indole-acetic acid methyltransferase (Zubieta et al., 2003), and
cinnamic/p-coumaric acid methyltransferase (Kapteyn et al., 2007). The three dimen-
sional structure of C. breweri SA carboxyl methyltransferase (Zubieta et al., 2003), com-
bined with in silico modeling of the active site pocket in the Nicotiana suaveolens and S.
floribunda enzymes (Pott et al., 2004), also indicates that these enzymes have a unique
structure that is distinct from those of unrelated methyltransferases found in plants (Noel
et al., 2003).
Different reports suggest that vanillin could be synthesized from phenylpropanoid
precursors (Schwab et al., 2008), and a three-step pathway for vanillin biosynthesis
from 4-coumaric acid has been proposed based on precursor accumulation and feed-
ing cell cultures of V. planifolia with the proposed precursors (Havkin‐Frenkel et al.,
1999). In this pathway, 4-coumaric acid is first converted to 4-hydroxybenzaldehyde
via 4-hydroxybenzaldehyde synthase through a chain shortening step (Podstolski et al.,
2002). Then, 4-hydroxybenzaldehyde synthase carries out the hydroxylation of position
3 on the ring of p-hydroxybenzyl alcohol converting it to 3,4-dihydroxybenzyl alcohol
or aldehyde. The final enzymatic step was shown to be catalyzed by a multifunctional V.
planifolia OMT that had a broad substrate range, including 3,4-dihydroxybenzaldehyde
(Pak et al., 2004).

4.3.3 Carbohydrate-Derived Flavor Compounds

4.3.3.1 Furanones and Pyrones

Only a limited number of natural volatiles originate directly from carbohydrates with-
out prior degradation of the carbon skeleton; such compounds include the furanones
and pyrones (Bood and Zabetakis, 2002), which are important fruit constituents and
have been isolated from the bark and leaves of several tree species (Schwab and Roscher,
1997). Pyrone maltol and substituted 4-hydroxy-3-(2H)-furanones constitute flavor
molecules with exceptional low odor thresholds. Furanones are emitted only by the
fruits; conversely, maltol has been isolated from the bark and leaves of Larix deciduas,
Evodiopanax innovans, Cercidiphyllum japonicum, and Pinaceae plants (Tiefel and
Berger, 1993; Schwab et al., 2008). Regarding the precursor of furaneol, experiments
using the labeled precursors revealed that d-fructose-1,6-diphosphate is an efficient bio-
genetic precursor of furaneol. In strawberry (Fragaria ananassa) and tomato (Solanum
lycopersicum), the hexose diphosphate is converted to 4-hydroxy-5-methyl-2-methylene-
3-(2,H)-furanone, which serves as the substrate for an enone oxidoreductase isolated
from ripe fruit (Raab et al., 2006; Klein et al., 2007).
Furaneol is one of the key flavor compounds in the attractive aroma of fruits (Farine
et al., 1994). In strawberry, furaneol is further metabolized by an O-methyltransferase
(FaOMT) to methoxyfuraneol (Wein et al., 2002). The in vivo methylation of furaneol
by FaOM has been demonstrated by the genetic transformation of strawberry with the
FaOMT sequence in the antisense orientation, under the control of a constitutive pro-
moter, resulting in a near-total loss of the methoxyfuraneol volatile (Lunkenbein et al.,
2006). However, the reduced level of methoxyfuraneol was only perceived by one-third
 Aroma Compounds (Description, Biosynthesis, and Regulation) 79

of the volunteer panelists, consistent with results obtained by aroma extract dilution
assays. Norfuraneol and homofuraneol have been identified in tomato and melon fruits,
respectively, but their biogenetic pathways and that of maltol remain unknown (Schwab
and Roscher, 1997; Schwab et al., 2008). However, studies in tomato and yeast have
identified phosphorylated carbohydrates as potential precursors of the furanones (Sasaki
et al., 1991; Hauck et al., 2003).

4.3.3.2 Terpenoids
These molecules are enzymatically synthesized de novo from acetyl-CoA and pyruvate
available from the carbohydrate present in plastids and the cytoplasm, without an impor-
tant contribution of acetyl-CoA from fatty acid oxidation in peroxisomes (Schwab et al.,
2008). Despite their diversity, all terpenoids derive from the common building units iso-
pentenyl diphosphate (IDP) and its isomer dimethylallyl diphosphate (DMADP) (Croteau
and Karp, 1991; McGarvey and Croteau, 1995; Croteau et al., 2000). Both IDP and
DMADP are synthesized via two parallel pathways in plants: (i) the mevalonate (MVA)
pathway, which is active in the cytosol; and (ii) the methylerythritol 4‐phosphate (MEP)
pathway, which is active in the plastids (Lichtenthaler, 1999; Rodriguez‐Concepción and
Boronat, 2002; Rohdich et al., 2002; Rohmer, 2003). In recent years, there has been
significant progress in understanding the subcellular distribution of substrates with dif-
fering chain length and cross-talk between the two pathways for substrate formation
(Gutensohn et al., 2013; Rasulov et al., 2015; Dong et al., 2016; Pazouki and Niinemets,
2016). The cytosolic pathway is responsible for the synthesis of sesquiterpenes, phytos-
terols, and ubiquinone, whereas monoterpenes, gibberellins, abscisic acid, carotenoids
and the prenyl moiety of chlorophylls, plastoquinone and tocopherol are produced in
plastids (Lichtenthaler, 1999; Rodriguez‐Concepción and Boronat, 2002; Rohdich et al.,
2002; Rohmer, 2003), but indications of cross‐talk between the plastidic and cytosolic
pathways have been found in tobacco, Arabidopsis, and snapdragon petals (Ohara et al.,
2003; Aharoni et al., 2004; Dudareva et al., 2005). The direct precursors of terpenoids,
linear geranyl diphosphate (GDP, C10), farnesyl diphosphate (FDP, C15), and geranyl-
geranyl diphosphate (GGDP, C20), are produced by the activities of three prenyl trans-
ferases (PTs). Terpene synthases are the primary enzymes responsible for catalyzing the
formation of hemiterpenes (C5), monoterpenes (C10), sesquiterpenes (C15) or diterpenes
(C20) from the substrates DMADP, GDP, FDP, or GGDP, respectively.
Prenyl transferases catalyze the addition of IDP units to prenyl diphosphates with
allylic double bonds to the diphosphate moiety. Most of the PTs accept DMADP as the
initial substrate, but they also bind GDP or FDP depending on the particular prenyl-
transferase (Tarshis et al., 1994, 1996; Greenhagen and Chappell, 2001; Withers and
Keasling, 2007). The availability of GDP and FDP are often the key factor in the pro-
duction of monoterpenes and sesquiterpenes in plants. In fact, multi-substrate enzymes
can form monoterpenes with GDP as the substrate and sesquiterpenes with FDP as the
substrate (Davidovich-Rikanati et al., 2008; Gutensohn et al., 2013; Pazouki et al., 2015;
Pazouki and Niinemets, 2016). This problem was elegantly overcome in metabolic engi-
neering experiments by the co‐expression of GDP and FDP synthases with appropriate
monoterpene and sesquiterpene synthases over-expressed in tobacco (Wu et al., 2006).
This strategy, together with the targeting of the over-expression to the plastid compart-
ment, resulted in increased synthesis of the sesquiterpenes amorpha‐4,11‐diene and
patchoulol and the monoterpene S‐limonene (Wu et al., 2006). The third phase of terpene
volatile biosynthesis involves the conversion of the various prenyl diphosphates DMADP,
GDP, FDP, and GGDP to hemiterpenes, monoterpenes, sesquiterpenes, and diterpenes,
80 Food Aroma Evolution

respectively, by the large family of the terpene synthases (TPSs) (Pazouki and Niinemets,
2016). Triterpenes (and sterols) and tetraterpenes (such as carotenoids) are derived from
the condensation of two molecules of FDP or GGDP, respectively. Evolutionarily, plant
hemiterpene, monoterpene, sesquiterpene, and diterpene synthases are related to each
other and are structurally distinct from triterpene or tetraterpene synthases (Bohlmann
and Keeling, 2008). Many TPSs have been isolated and characterized from various plant
species (Bohlmann et al., 1998; Tholl, 2006; Davidovich-Rikanati et al., 2008; Gutensohn
et al., 2013; Pazouki et al., 2015). In tomato and other plants, there is evidence that
several TPSs are multi-substrate enzymes, capable of synthesizing terpenes of different
chain length depending on corresponding substrate availability (Davidovich-Rikanati et
al., 2008; Gutensohn et al., 2013; Pazouki et al., 2015). Among such multi-substrate
enzymes, some can form monoterpenes with GDP as the substrate and sesquiterpenes
with FDP as the substrate (Davidovich-Rikanati et al., 2008; Gutensohn et al., 2013;
Pazouki et al., 2015; Pazouki and Niinemets, 2016). It has also been reported that the
cultivated apple genome (Malus domestica) contains 55 putative terpene synthase TPS
genes but only 10 are functional (Nieuwenhuizen et al., 2013). While many terpene vola-
tiles are direct products of TPSs, many others are formed through transformation of the
initial products by oxidation, dehydrogenation, acylation, and other reactions (Croteau
and Karp, 1991; Croteau et al., 2000; Dudareva et al., 2004; Pichersky et al., 2006). For
example, (–)-(1R,2S,5R)-menthol, the principal monoterpene of commercial peppermint
essential oil is formed by eight enzymatic steps involving monoterpene synthases, isom-
erases, and reductases (Turner and Croteau, 2004; Ringer et al., 2005). The biosynthe-
sis starts with the formation of 4S-limonene from GPP and ends with the reduction of
(–)-menthone to (°)-menthol.

4.3.3.3 Apocarotenoids
Apocarotenoids are commonly found in the flowers, fruits, and the leaves of many plants
(Winterhalter and Rouseff, 2001), and possess flavor aroma properties together with low
aroma thresholds. They are found among the potent flavor compounds in wines and con-
tribute to floral and fruity attributes (Winterhalter and Schreier, 1994, 2002). Carotenoid
cleavage dioxygenases catalyze the oxidative cleavage of carotenoids, resulting in the
production of apocarotenoids (Schmidt et al., 2006). CCDs often exhibit a preference
for several substrates, contributing to the diversity of apocarotenoids found in nature.
The synthesis of β-ionone, geranyl acetone and 6-methyl-5-hepten-2-one in tomato fruits
increase 10–20‐fold during fruit ripening, and these compounds were shown to be pro-
duced by the activity of the genes LeCCD1A and LeCCD1B that were isolated from
tomato fruits (Simkin et al., 2004). In tomato fruit, β-ionone is present at very low con-
centrations (4 nLL −1), but due to its low odor threshold (0.007 nLL −1), it is the second
most important volatile contributing to fruit flavor (Baldwin et al., 2000). Silencing of
LeCCD1A and LeCCD1B resulted in a significant decrease in the β-ionone content of
ripe fruits, implying a role for these genes in C13 norisoprenoid synthesis in vivo (Simkin
et al., 2004). Reduction of Petunia hybrida CCD1 transcript levels in transgenic plants
led to a 58–76% decrease in β‐ionone synthesis in the corollas of selected petunia lines,
indicating a significant role for this enzyme in volatile synthesis (Simkin et al., 2004).
Also, a potential CCD gene was identified among a Vitis vinifera L. EST collection, and
recombinant expression of VvCCD1 confirmed that the gene encodes a functional CCD
that cleaves zeaxanthin symmetrically yielding 3-hydroxy-β-ionone and a C14 aldehyde
(Mathieu et al., 2005). CCDs were also found to be involved in the formation of impor-
tant aroma compounds in melon (Cucumis melo). The product of the CmCCD1 gene,
 Aroma Compounds (Description, Biosynthesis, and Regulation) 81

whose expression is up-regulated upon fruit development, was shown to cleave carot-
enoids, generating geranylacetone from phytoene, pseudoionone from lycopene, β-ionone
from β-carotene, and α-ionone and pseudoionone from δ-carotene (Ibdah et al., 2006).
A recent study of carotenoid biosynthesis in bilberry or European blueberry showed
an increase of expression of the gene encoding for carotenoid cleavage dioxygenase
(VmCCD1) and 9-cis-epoxycarotenoid dioxygenase (VmNCED), indicating enzymatic
carotenoid cleavage and degradation during ripening (Karppinen et al., 2016). Instead,
mature bilberry fruits responded specifically to red/far-red light wavelengths by induc-
ing the expression of both the carotenoid biosynthetic and the cleavage genes indicating
tissue and developmental stage-specific regulation of apocarotenoid formation by light
quality (Karppinen et al., 2016).

REFERENCES

Acree, T.E., Lee, C.Y., Butts, R.M., and Barnard, J. 1976. Geosmin, the earthy compo-
nent of table beet odor. J. Agric. Food Chem. 24, 430–431.
Agerbirk, N. and Olsen, C.E. 2012. Glucosinolate structures in evolution. Phytochemistry
77, 16–45.
Aharoni, A., Giri, A.P., Verstappen, F.W.A., Bertea, C.M., Sevenier, R., Sun, Z., Jongsma,
M.A., Schwab, W., and Bouwmeester, H.J. 2004. Gain and loss of fruit flavor com-
pounds produced by wild and cultivated strawberry species. Plant Cell 16, 3110–31.
Aharoni, A., Keizer, L.C.P., Bouwmeester, H.J., Sun, Z., Alvarez-Huerta, M., Verhoeven,
H.A., Blaas, J., van Houwelingen, A.M.M.L., De Vos, R.C.H., van Der Voet, H.,
Jansen, R.C., Guis, M., Mol, J., Davis, R.W., Schena, M., Van Tunen, A.J., and
O’Connell, A.P. 2000. Identification of the SAAT gene involved in strawberry flavor
biogenesis by use of DNA microarrays. Plant Cell 12, 647–62.
Ahmed, M.A., El–Mawla, A., and Beerhues, L. 2002. Benzoic acid biosynthesis in cell
cultures of Hypericum androsaemum. Planta 214, 727–33.
Akaberi, S., Wang, H., Claudel, P., Riemann, M., Hause, B., Hugueney, P., and Nick, P.
2018. Grapevine fatty acid hydroperoxide lyase generates actin-disrupting volatiles
and promotes defence-related cell death. J. Exp. Bot. 25, 69(12), 2883–96.
Aragüez, I. and Valpuesta, V. 2013. Metabolic engineering of aroma components in
fruits. Biotechnol. J. 8, 1144–1158.
Asif, M.H., Lakhwani, D., Pathak, S., Gupta, P., Bag, S.K., Nath, P., and Trivedi, P.K.
2014. Transcriptome analysis of ripe and unripe fruit tissue of banana identifies
major metabolic networks involved in fruit ripening process. BMC Plant Biol.
14, 316.
Aubert, C. and Milhet, C. 2007. Distribution of the volatile compounds in the different
parts of a white-fleshed peach (Prunus persica L. Batsch). Food Chem. 102, 375–84.
Aubert, C., Baumann, S., and Arguel, H. 2005. Optimization of the analysis of flavor
volatile compounds by liquid-liquid microextraction (LLME): Application to the
aroma analysis of melons, peaches, grapes, strawberries, and tomatoes. J. Agric.
Food Chem. 53, 8881–8895.
Aubert, C., Gunata, Z., Ambid, C., and Baumes, R. 2003. Changes in physicochemical
characteristics and volatile constituents of yellow- and white-fleshed nectarines dur-
ing maturation and artificial ripening. J. Agric. Food Chem. 51, 3083–91.
Azzara, C.D. and Campbell, L.B. 1992. Off-flavors of dairy products. In G. Charalambous
(Ed.), Off-Flavors in Foods and Beverages. Elsevier: Amsterdam, pp. 329–74.
82 Food Aroma Evolution

Bagchi, G.D. 2000. Essential oil producing glands in some important aromatic plants:
Structure and process of secretion. J. Med. Aromat. Plant Sci. 22, 605–15.
Bak, S., Paquette, S.M., and Morant, M. 2006. Cyanogenic glycosides: A case study for
evolution and application of cytochromes P450. Phytochem. Rev. 5, 309–29.
Balbontin, C., Gaete-Eastman, C., Fuentes, L., Figueroa, C.R., Herrera, R., Manriquez,
D., and Moya-Leon, M.A. 2010. VpAAT1, a gene encoding an alcohol acyltrans-
ferase, is involved in ester biosynthesis during ripening of mountain papaya fruit. J.
Agric. Food Chem. 58, 5114–5121.
Baldwin, E.A., Plotto, A., and Goodner, K. 2007. Shelf-life versus flavour-life for fruits
and vegetables: How to evaluate this complex trait. Stewart Postharvest Rev. 3(1),
1–10.
Baldwin, E.A., Scott, J.W., Shewmaker, C.K., and Schuch, W. 2000. Flavor trivia and
tomato aroma: Biochemistry and possible mechanisms for control of important
aroma components. HortScience 35, 1013–22.
Barba, F., Nikmaram, N., Roohinejad, S., Khelfa, A., Zhu, Z., and Koubaa, M. 2016.
Bioavailability of glucosinolates and their breakdown products: Impact of process-
ing. Front. Nutr. 3, 24.
Bartley, I.M., Stoker, P.G., Martin, A.D.E., Hatfield, S.G.S., and Knee, M. 1985.
Synthesis of aroma compounds by apples supplied with alcohols and methyl esters
of fatty acids. J. Sci. Food Agric. 36, 567–74.
Basear, K.H.C. and Demirci, F. 2007. Chemistry of essential oil. In: R.G. Berger (Eds.),
Flavors and Fragrances. Heidelberg: Springer, pp. 43–86.
Bayrak, A. 1994. Volatile oil composition of Turkish rose (Rosa damascena). J. Agric.
Food. Chem. 64, 441–8.
Beaulieu, J.C. and Baldwin, E. 2002. Flavor and aroma of fresh-cut fruits and vegetables.
In: O. Lamikanra (Ed.), Fresh-Cut Fruits and Vegetables: Science, Technology and
Market. CRC Press: Boca Raton, FL.
Beaulieu, J.C. and Grimm, C.C. 2001. Identification of volatile compounds in cantaloupe
at various developmental stages using solid phase microextraction. J. Agric. Food
Chem. 49, 1345–1352.
Becalski, A. and Seaman, S. 2005. Furan precursors in food: A model study and develop-
ment of a simple headspace method for determination of furan. J. AOAC Int. 88,
102–6.
Beekwilder, J., Alvarez-Huerta, M., Neef, E., Verstappen, F.W., Bouwmeester, H.J., and
Aharoni, A. 2004. Functional characterization of enzymes forming volatile esters
from strawberry and banana. Plant Physiol. 135(4), 1865–78.
Belitz, H., Grosch, W., and Schieberle, P. 2008. Aroma compounds. In: H. Belitz, W.
Grosch, and P. Schieberle (Eds.), Food Chem. Springer: Garching, pp. 339–402.
Belitz, H.-D., Grosch, W., and Schieberle, P. 2004. Food Chemistry. Springer: Garching.
Belitz, H.D., Grosch, W., and Schieberle, P. 2009. Aroma Compounds in Food Chemistry,
4th ed. Springer: Berlin, Germany, pp. 340–400.
Berger, R.G. 2007. Flavours and Fragrances-Chemistry, Bioprocessing and Sustainability.
Springer-Verlag: Berlin, Germany.
Bernreuther, A. and Schreier, P. 1991. Multidimensional gas chromatography/mass spec-
trometry: A powerful tool for the direct chiral evaluation of aroma compounds in
plant tissues. 2. Linalool in essential oils and fruits. Phytochem. Anal. 2, 167–70.
Bernreuther, A., Epperlein, U., and Koppenhoefer, B. 1997. In: R. Marsili (Ed.), Techniques
for Analyzing Food Aroma. Marcel Dekker: New York, NY, pp. 143–207.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 83

Beuerle, T. and Pichersky, E. 2002. Purification and characterization of benzoate:

Coenzyme A ligase from Clarkia breweri. Arch. Biochem. Biophys. 400, 258–64.
Boatright, J., Negre, F., Chen, X., Kish, C.M., Wood, B., Peel, G., Orlova, I., Gang, D.,
Rhodes, D., and Dudareva, N. 2004. Understanding in vivo benzenoid metabolism
in petunia petal tissue. Plant Physiol. 135, 1993–2011.
Bohlmann, J. and Keeling, C. 2008. Terpenoid biomaterials. Plant J. 54, 656–69.
Bohlmann, J., Meyer-Gauen, G., and Croteau, R. 1998. Plant terpenoid synthases:
Molecular biology and phylogenetic analysis. Proc. Natl Acad. Sci. USA, 95, 4126–33.
Bones, A.M. and Rossiter, J.T. 2006. The enzymic and chemically induced decomposi-
tion of glucosinolates. Phytochem. 67, 1053–67.
Bood, J. and Zabetakis, I. 2002. The biosynthesis of strawberry flavor (II): Biosynthetic
and molecular biology studies. J. Food Sci. 67(1), 2–8.
Brackmann, A., Streif, J., and Bangerth, F. 1993. Relationship between a reduced aroma
production and lipid metabolism of apples after long-term controlled-atmosphere
storage. J. Am. Soc. Horticult. Sci. 118, 243–7.
Buettner, A. and Schieberle, P. 2001. Application of a comparative aroma extract dilu-
tion analysis to monitor changes in orange juice aroma compounds during pro-
cessing. In: J. V. Leland, P. Schieberle, A. Buettner, and T.E. Acree (Eds.), Gas
Chromatography – Olfactometry. American Chemical Society: Washington, DC,
pp. 33–45.
Buseman, C.M., Tamura, P., and Sparks, A.A. 2006. Wounding stimulates the accumu-
lation of glycerolipids containing oxophytodienoic acid and dinoroxophytodienoic
acid in Arabidopsis leaves. Plant Physiol. 142, 28–39.
Buttery, R.G. and Ling, L.C. 1973. Earthy aroma of potatoes. J. Agric. Food Chem. 21,
745–746.
Buttery, R. G., Seifert, R. M., Guadagni, D. G., and Ling, L. C. 1969. Characterization of
some volatile constituents of bell peppers. J. Agric. Food Chem. 17, 1322–7.
Cerny, C. 2012. Savory flavours. In: L.M.L. Nollet (Ed.), Handbook of Meat. Poultry
and Seafood Quality, 2nd ed. Wiley-Blackwell: Chichester.
Chan, H.W.S. 1987. Autoxidation of Unsaturated Lipids. Academic Press: London, UK.
Chen, F., D’Auria, J.C., Tholl, D., Ross, J.R., Gershenzon, J., Noel, J.P., and Pichersky, E.
2003. An Arabidopsis thaliana gene for methylsalicylate biosynthesis, identified by
a biochemical genomics approach, has a role in defense. Plant J. 36, 577–88.
Chen, S. and Andreasson, E. 2001. Update on glucosinolate metabolism and transport.
Plant Physiol. Biochem. 39, 743–58.
Cheong, K.W., Tan, C.P., Mirhosseini, H., Hamid, N.S.A., Osman, A., and Basri, M.
2010. Equilibrium headspace analysis of volatile flavour compounds extracted from
soursop (Anoan muricata) using solid phase microextraction. Food Res. Int. 43,
1267–76.
Cherian, S., Figueroa, C.R., and Nair, H. 2014. Movers and shakers’ in the regulation of
fruit ripening: A cross-dissection of climacteric versus non-climacteric fruit. J. Exp.
Bot. 65, 4705–22.
Christlbauer, M. and Schieberle, P. 2009. Characterization of the key aroma compounds
in beef and pork vegetable gravies á la chef by application of the aroma extract dilu-
tion analysis. J. Agric. Food Chem. 57, 9114–9122.
Cho, I, Kim, S.Y., Choi, H.K., and Kim, Y.S. 2006. Characterization of aroma-active
compounds in raw and cooked pine-mushrooms (Tricholoma matsutake Sing). J.
Agric. Food Chem. 54, 6332–5.
84 Food Aroma Evolution

Collin, S., Nizet, S., Claeys Bouuaert, T., and Despatures, P.-M. 2011. Main odorants in
Jura florsherry wines. Relative contributions of sotolon, abhexon, and theaspirane-
derived compounds. J. Agric. Food Chem. 60, 380–7.
Conde, C., Delrot, S., and Gerós, H. 2008. Physiological, biochemical and molecular
changes occurring during olive development and ripening. J. Plant Physiol. 165(15),
1545–62.
CPMA. 2012. CPMA Fruit and Vegetable Availability Guide. Canada Produce Marketing
Association. http:​//www​.cpma​.ca/p​df/He​althN​utrit​ion/C​PMA_F​ruit_​Veget​able_​
Avail​abili​t y_Gu​ide_2​012_e​n.pdf​, accessed March 05, 2019.
Croteau, R. and Karp, F. 1991. Origin of natural odorants. In: P.M. Muller and D.
Lamparsky (Eds.), Perfumes: Art, Science and Technology. London: Elsevier
Applied Science, pp. 101–26.
Croteau, R., Kutchan, T. M., and Lewis, N.G. 2000. Natural products (secondary
metabolites). In B. Buchanan, W. Gruissem, and R. Jones (Eds.), Biochemistry and
Molecular Biology of Plants. American Society of Plant Physiologists: Rockville,
MD, pp. 1250–318.
Cumplido-Laso, G., Medina-Puche, L., Moyano, E., Hoffmann, T., Sinz, Q., Ring,
L., Studart-Wittkowski, C., Caballero, J. L., Schwab, W., Muñoz-Blanco, J., and
Blanco-Portales, R. 2012. The fruit ripening-related gene FaAAT2 encodes an
acyl transferase involved in strawberry aroma biogenesis. J. Exp. Bot. 63(11),
4275–90.
D’Auria, J.C., Chen, F. and Pichersky, E. 2003. The SABATH family of MTs in
Arabidopsis thaliana and other plant species. Recent Adv. Phytochem. 37, 95–125.
Davidovich-Rikanati, R., Lewinsohn, E., Bar, E., Iijima, Y., Pichersky, E., and Sitrit, Y.
2008. Over-expression of the lemon basil alpha-zingiberene synthase gene increases
both mono- and sesquiterpene contents in tomato fruit. Plant J. 56(2), 228–38.
Defilippi, B., Kader, A. and Dandekar A.M. 2005. Apple aroma: Alcohol acyltransferase,
a rate limiting step for ester biosynthesis, is regulated by ethylene. Plant Sci. 168,
1199–1210.
De Pooter, H.L., Montens, J.P., Dirinck, J., Willaert, G.A., and Schamp, N.M. 1983.
Treatment of Golden Delicious apple with aldehydes and carboxylic acids: Effect on
the headspace composition. J. Agric. Food Chem. 37, 813–18.
Dixon, J. and Hewett, E.W. 2000. Factors affecting apple aroma/flavour volatile concen-
tration: A review. New Zeal. J. Crop Horticult. Sci. 28, 155–73.
Dobson, H. 1994. Floral Volatiles in Insect Biology. In: E. A. Bernays (Ed.), Insect–Plant
Interactions. Boca Raton, FL: CRC Press, pp. 47–81.
Domínguez, T., Hernández, M.L., Pennycooke, J.C., Jiménez, P., Martínez-Rivas, J.M.,
Sanz, C., Stockinger, E.J., Sánchez-Serrano, J.J., and Sanmartín, M. 2010. Increasing
ω-3 desaturase expression in tomato results in altered aroma profile and enhanced
resistance to cold stress. Plant Physiol. 153, 655–65.
Dong, L., Jongedijk, E., Bouwmeester, H., and Van Der Krol, A. 2016. Monoterpene byo-
sinthesis potential of plant subcellular compartments. New Phytol. 209, 679–690.
Doty, R.L. 1991. Psychophysical measurement of odor perception in humans. In: D.
Laing, R. Doty, and W. Breipohl (Eds.), The Human Sense of Smell. Springer-Verlag:
Berlin Heidelberg.
Drewnowski, A. 1997. Taste preferences and food intake. Ann. Rev. Nutr. 17, 237–53.
Du, X. and Qian, M. 2008. Quantification of 2,5-dimethyl-4 hydroxy-3(2H)-furanone
using solid-phase extraction and direct microvial insert thermal desorption gas
chromatography-mass spectrometry. J. Chromatogr. A 1208, 197–201.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 85

Dudareva, N. and Pichersky, E. 2000. Biochemical and molecular genetic aspects of flo-
ral scents. Plant Physiol. 122, 627–33.
Dudareva, N. and Pichersky, E. 2006. Floral scent metabolic pathways: Their regulation
and evolution. In: N. Dudareva and E. Pichersky (Eds.), Biology of Floral Scent.
Boca Raton, FL: CRC Press, pp. 55–78.
Dudareva, N., Andersson, S., Orlova, I., Gatto, N., Reichelt, M.,Rhodes, D., Boland, W.,
and Gershenzon, J. 2005. The nonmevalonate pathway supports both monoterpene
and sesquiterpene formation in snapdragon flowers. Proc. Natl Acad. Sci. USA,
102, 933–8.
Dudareva, N., Murfitt, L.M., Mann, C.J., Gorenstein, N., Kolosova, N., Kish, C.M.,
Bonham, C., and Wood, K. 2000. Developmental regulation of methyl benzoate
biosynthesis and emission in snapdragon flowers. Plant Cell 12, 949–61.
Dudareva, N., Pichersky, E., and Gershenzon, J. 2004. Biochemistry of plant volatiles.
Plant Physiol. 135, 1893–902.
Dudareva, N., Raguso, R.A., Wang, J., Ross, J.R., and Pichersky, E. 1998 Floral scent
production in Clarkia breweri. III. Enzymatic synthesis and emission of benzenoid
esters. Plant Physiol. 116, 599–604.
Dunemann, F., Ulrich, D., Boudichevskaia, A., Grafe, C., and Weber, W. E. 2009. QTL
mapping of aroma compounds analysed by headspace solid-phase microextraction
gas chromatography in the apple progeny ‘Discovery’ × ‘Prima’. Mol. Breed. 23,
501–21.
Dunphy, P. J. and Allcock, C. 1972. Isolation and properties of a monoterpene reductase
from rose petals. Phytochemistry 11, 1887–91.
Eduardo, I., Chietera, G., Bassi, D., Rossini, L., and Vecchietti, A. 2010. Identification
of key odor volatile compounds in the essential oil of nine peach accessions. J. Sci.
Food Agric. 90, 1146–54.
El Hadi, M.A., Zhang, F.J., Wu, F.F., Zhou, C.H., and Tao, J. 2013. Advances in fruit
aroma volatile research. Molecules 11, 18(7), 8200–29.
El-Sharkawy, I., Manríquez, D., Flores, F.B., Regad, F., Bouzayen, M., Latché, A., and
Pech, J. C. 2005. Functional characterization of a melon alcohol acyl-transferase
gene family involved in the biosynthesis of ester volatiles. Identification of the cru-
cial role of a threonine residue for enzyme activity. Plant Mol. Biol. 59, 345–62.
Engel, K.H., Flath, R.A., Buttery, R.G., Mon, T.R., Ramming, D.W., and Teranishi, R.
1988. Investigation of volatile constituents in nectarines. 1. Analytical and sensory
characterization of aroma components in some nectarine cultivars. J. Agric. Food
Chem. 36, 549–53.
Engel, K.-H., Schellenberg, A., and Schmarr, H.-G. 2001. In: G.R. Takeoka, M. Guntert,
and K.-H. Engel (Eds.), Aroma Active Compounds in Foods. ACS Symposium Series
794. American Chemical Society: Washington, DC, pp. 138–48.
Espino-Díaz, M., Sepúlveda, D.R., González-Aguilar, G., and Olivas, G.I. 2016.
Biochemistry of apple aroma: A review. Food Technol. Biotechnol. 54(4), 375–97.
Fan, X. 2005a. Impact of ionizing radiation and thermal treatments on furan levels in
fruit juice. J. Food Sci. 70, E409–14.
Fan, X. 2005b. Formation of furan from carbohydrates and ascorbic acid following
exposure to ionizing radiation and thermal processing. J. Agric. Food Chem. 53,
7826–31.
Farine, J.-P., Le Quere, J.-L., Duffy, J., Everaerts, C., and Brossut, R. 1994. Male sex pher-
omone of cockroach Eurycotis floridana (Walker) (Blattidae, Polyzosteriinae): Role
and composition of tergites 2 and 8 secretions. J. Biol. Chem. Ecol. 20, 2291–306.
86 Food Aroma Evolution

Flath, R.A. and Takahashi, J.M. 1978. Volatile constituents of prickly pear (Opuntia
ficus indica Mill., de Castilla variety). Institut za Primenu Nuklearne Energije u
Poljoprivredi, Veterinarstvu i Sumarstvu, Zemun (Yugoslavia).
Flament, I., Debonneville, C., and Furrer, A. 1993. Volatile constituents of roses:
Characterization of cultivars based on the headspace analysis of living flower emis-
sions. In: R. Teranishi, R.G. Buttery, and H. Sugisawa (Eds.), Bioactive Volatile
Compounds from Plants. American Chemical Society: Washington, DC, pp. 269– 281.
Flamini, G., Tebano, M., and Cioni, P.L. 2007. Volatiles emission patterns of different
plant organs and pollen of Citrus limon. Anal. Chim. Acta 589, 120–124.
Forney, C. and Song, J. 2018. Flavor and aromas: Biological functions, Chapter 24. In:
Elhadi M. Yahía (Ed.), Fruit and Vegetable Phytochemical: Chemistry and Human
Health, vol. I, 2nd ed. John Wiley & Sons, pp. 515–39.
Forney, C.F. 2001. Horticultural and other factors affecting aroma volatile composition
of small fruit. HortTechnology 11, 529–38.
Forney, C.F., Jamieson, A.R., Munro Pennell, K.D., Jordan, M.A., and Fillmore, S.A.E.
2015. Relationships between fruit composition and storage life in air or controlled
atmosphere of red raspberry. Post. Biol. and Tech. 10, 121–30.
Frey, M., Stettner, C., Pare, P.W., Schmelz, E.A., Tumlinson, J.H., and Gierl, A. 2000. An
herbivore elicitor activates the gene for indole emission in maize. Proc. Natl Acad.
Sci. USA 97, 14801–6.
Fukuda, T., Tanaka, H., Ihori, H., Okazaki, K., Shinano, T., and Fukumori, Y. 2013.
Identification of important volatiles in fresh and processing carrot varieties: Using
kuroda and flakee types. Food Sci. Technol. Res. 19, 497–504.
Gang, D.R., Lavid, N., Zubieta, C., Chen, F., Beuerle, T., Lewinsohn, E., Noel, J.P., and
Pichersky, E. 2002. Characterization of phenylpropene O–methytransferases from
sweet basil: Facile change of substrate specificity and convergent evolution within a
plant OMT family. Plant Cell 14, 505–19.
Gasser, U. and Grosch, W. 1988. Identification of volatile flavor compounds with high
aroma values from cooked beef. Z. Lebensm. Unters. Forsch. 186, 489–94.
Gasser, U. and Grosch, W. 1990. Primary odorants of chicken broth. A comparative
study with meat broths from cow and ox. Z. Lebensm. Unters. Forsch. 190, 3–8.
Goepfert, S. and Poirier, Y. 2007. β-Oxidation in fatty acid degradation and beyond.
Curr. Opin. Plant Biol. 10, 245–51.
Goldner, M.C., Zamora, M.C., Di Leo, P., Gianninoto, H., and Bandoni, A. 2009. Effect
of ethanol level in the perception of aroma attributes and the detection of volatile
compounds in red wine. J. Sens. Stud. 24(2), 243–57.
Gonda, I., Bar, E., Portnoy, V., Lev, S., Burger, J., Schaffer, A.A., Tadmor, Y., Gepstein,
S., Giovannoni, J.J., Katzir, N., and Lewinsohn, E. 2010. Branched-chain and aro-
matic amino acid catabolism into aroma volatiles in Cucumis melo L. fruit. J. Exp.
Bot. 61(4), 1111–23.
Gonda, I., Davidovich-Rikanati, R., Bar, E., Lev, S., Jhirad, P., Meshulam, Y.,
Wissotsky, G., Portnoy, V., Burger, J., Schaffer, A.A., Tadmor, Y., Giovannoni, J.J.,
Fei, Z., Fait, A., Katzir, N., and Lewinsohn, E. 2018. Differential metabolism of
­l-phenylalanine in the formation of aromatic volatiles in melon (Cucumis melo L.)
fruit. Phytochemistry 148, 122–31.
Gonda, I., Lev, S., Bar, E., Sikron, N., Portnoy, V., Davidovich-Rikanati, R., Burger, J.,
Schaffer, A.A., Tadmor, Y., Giovannonni, J.J., Huang, M., Fei, Z., Katzir, N., Fait,
A., and Lewinsohn, E. 2013. Catabolism of l-methionine in the formation of sulfur
and other volatiles in melon (Cucumis melo L.) fruit. Plant J. 74, 458–72.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 87

Gónzalez, M., Gaete-Eastman, C., Valdenegro, M., Figueroa, C.R., Fuentes, L., Herrera,
R., and Moya-León, M.A. 2009. Aroma development during ripening of F. chiloen-
sis fruit and participation of an alcohol acyltransferase (FcAAT1) gene. J. Agric.
Food Chem. 57, 9123–32.
González-Aguero, M., Troncoso, S., Gudenschwager, O., Campos-Vargas, R., Moya-
León, M.A., and Defilippi, B.G. 2009. Differential expression levels of aroma-related
genes during ripening of apricot (Prunus armeniaca L.). Plant Physiol. Biochem. 47,
435–40.
González-Mas, M.C., Rambla, J.L., Alamar, M.C., Gutiérrez, A., and Granell, A. 2011.
Comparative analysis of the volatile fraction of fruit juice from different citrus spe-
cies. PLoS ONE 6, e22016.
Greenhagen, B. and Chappell, J. 2001. Molecular scaffolds for chemical wizardry: Learning
nature’s rules for terpene cyclases. Proc. Natl Acad. Sci. USA 98, 13479–81.
Gross, M., Friedman, J., Dudai, N., Larkov, O., Cohen, Y., Bar, E., Ravid, U., Putievsky,
E., and Lewinsohn, E. 2002. Biosynthesis of estragole and t-anethole in bitter fennel
(Foeniculum vulgare Mill. var. vulgare) chemotypes. Changes in SAM:phenylpropene
O-methyltransferase activities during development. Plant Sci. 163, 1047–53.
Gross, M., Joel, D.M., Cohen, Y., Bar, E., Friedman, J., and Lewinsohn, E. 2006
Ontogenesis of mericarps of bitter fennel (Foeniculum vulgare Mill. var. vulgare) as
related to t-anethole accumulation. Israel J. Plant Sci. 54, 309–16.
Guadagni, D.G., Bomben, J.L., and Hudson, J.S. 1971. Factors influencing the develop-
ment of aroma in apple peel. J. Sci. Food Agric. 22, 110–15.
Guo, Y.F., Zhang, Y.L., Shan, W., Cai, Y.J., Liang, S.M., Chen, J.Y., Lu, W.J., and Kuang, J.F.
2018. Identification of two transcriptional activators MabZIP4/5 in controlling aroma
biosynthetic genes during banana ripening. J. Agric. Food Chem. 20, 66(24), 6142–50.
Gutensohn, M., Orlova, I., Nguyen, T.T., Davidovich-Rikanati, R., Ferruzzi, M.G.,
Sitrit, Y., Lewinsohn, E., Pichersky, E., and Dudareva, N. 2013. Cytosolic monoter-
pene biosynthesis is supported by plastid-generated geranyl diphosphate substrate in
transgenic tomato fruits. Plant J. 75(3), 351–63.
Halkier, B. Gershenzon. 2006. Biology and biochemistry of glucosinolates. Annu. Rev.
Plant Biol. 57, 303–33.
Hamberg, M., Sanz, A., and Castresana, C. 1999. α-Oxidation of fatty acids in higher
plants. Identification of a pathogen‐inducible oxygenase (piox) as an α‐dioxygenase
and biosynthesis of 2‐hydroperoxylinolenic acid. J. Biol. Chem. 274, 24503–13.
Hauck, T., Hubner, Y., Bruhlmann, F., and Schwab, W. 2003. Alternative pathway
for the formation of 4,5-dihydroxy-2,3-pentanedione, the proposed precursor of
4-hydroxy-5-methyl- 3(2H)-furanone as well as autoinducer-2, and its detection as
natural constituent of tomato fruit. Biochem. Biophys. Acta 1623, 109–19.
Havkin-Frenkel, D., Podstolski, A., Witkowska, E., Molecki, P., and Mlkolajczyk, M.
1999. Vanillin biosynthetic pathways: An overview. In: T.J. Fu, G. Singh, and W.R.
Curtis (Eds.), Plant Cell and Tissue Culture for the Production of Food Ingredients.
New York: Kluwer/Plenum, pp. 35–43.
Hinterholzer, A., Lemos, T., and Schieberle, P. 1998. Identification of key odorants in
raw French beans and changes during cooking. Z. Lebensm. Unters. Forsch. A 207,
219–22.
Holland, D., Larkov, O., Bar-Yaákov, I., Bar, E., Zax, A., and Brandeis, E. 2005.
Developmental and varietal differences in volatile ester formation and acetyl-CoA:
Alcohol acetyl transferase activities in apple (Malus domestica Borkh.) fruit. J.
Agric. Food Chem. 53, 7198–203.
88 Food Aroma Evolution

Ibdah, M., Azulay, Y., Portnoy, V., Wasserman, B., Bar, E., Meir, A., Burger, Y.,
Hirchberg, J., Schaffer, A.A., Katzir, N., Tadmor, Y., and Lewinsohn, E. 2006.
Functional characterization of CmCDD1, a carotenoid cleavage dioxygenase from
melon. Phytochemistry 67, 1579–89.
Ibrahim, R.K., Bruneau, A., and Bantignies, B. 1998. Plant O-methyltransferases:
Molecular analysis, common signature and classification. Plant Mol. Biol. 36, 1–10.
Idstein, H., Keller, T., and Schreier, P. 1985. Volatile constituents of mountain papaya
(Carica candamarcensis, Syn. C. pubescens Lenne et Koch) fruit. J. Agric. Food
Chem. 33, 663–6.
Jetti, R.R., Yang, E., Kurnianta, A., Finn, C., and Qian, M.C. 2007. Quantification of
selected aroma-active compounds in strawberries by headspace solid-phase micro-
extraction gas chromatography and correlation with sensory descriptive analysis.
J. Food Sci. 72, S487–96.
Joel, D.M., French, J.C., Graft, N., Kourteva, G., Dixon, R.A., and Havkin-Frenkel, D.
2003. A hairy tissue produces vanillin. Isr. J. Plant Sci. 51, 157–159.
Jones, M.G., Hughes, J., Tregova, A., Milne, J., Tomsett, A.B., and Collin, H.A. 2004.
Biosynthesis of the flavor precursors of onion and garlic. J. Exp. Bot. 55, 1903–18.
Jordan, M.J., Shaw, P.E., and Goodner, K.L. 2001. Volatile components in aqueous
essence and fresh fruit of Cucumis melo cv. Athena (muskmelon) by GC–MS and
GC–O. J. Agric. Food Chem. 49, 5929– 5933.
Jung, H.P., Sen, A., and Grosch, W. 1992. Evaluation of potent odorants in parsley leaves
[Petroselinum crispum (Mill) Nym. ssp. crispum] by aroma extract dilution analysis
Lebensm.-Wiss. u.-Technol. 25, 55–60.
Kaminaga, Y., Schnepp, J., Peel, G., Kish, C.M, Ben-Nissan, G., Weiss, D., Orlova, I.,
Lavie, O., Rhodes, D., Wood, K., Porterfield, D.M., Cooper, A.J., Schloss, J.V.,
Pichersky, E., Vainstein, A., and Dudareva, N. 2006. Plant phenylacetaldehyde syn-
thase is a bifunctional homotetrameric enzyme that catalyzes phenylalanine decar-
boxylation and oxidation. J. Biol. Chem. 281, 23357–66.
Kaneko, S., Kumazawa, K., and Nishimura, O. 2011. Studies on the key aroma com-
pounds in soy milk made from three different soybean cultivars. J. Agric. Food
Chem. 59, 12204–9.
Kapteyn, J., Qualley, A.V., Xie, Z., Fridman, E., Dudareva, N., and Gang, D.R. 2007.
Evolution of cinnamate/p–coumarate carboxyl methyltransferases and their role in
the biosynthesis of methylcinnamate. Plant Cell 19, 3212–29.
Karppinen, K., Zoratti, L., Sarala, M., Carvalho, E., Hirsimäki, J., Mentula, H.,
Martens, S., Häggman, H., and Jaakola, L. 2016. Carotenoid metabolism during
bilberry (Vaccinium myrtillus L.) fruit development under different light conditions
is regulated by biosynthesis and degradation. BMC Plant Biol. 16, 95. Published Apr
21, 2016. doi: 10.1186/s12870-016-0785-5.
Kasuga, H., Asada, S., Iiyoshi, T., and Honda, S. 2008. Aroma components of potato
chips. Koryo 239, 35–41.
Katsuno, T., Kasuga, H., Kusano, Y., Yaguchi, Y., Tomomura, M., Cui, J., Yang., Z,
Baldermann, S., Nakamura, Y., Ohnishi, T., Mase, N., and Watanabe, N. 2014.
Characterisation of odorant compounds and their biochemical formation in green
tea with a low temperature storage process. Food Chem. 148, 388–95.
Kays, S.J. and Wang, Y. 2000. Thermally induced flavor compounds. HortSc. 35,
1002–12.
Klee, H.J. and Tieman, D.M. 2013. Genetic challenges of flavor improvement in tomato.
Trends Genet. 29, 257–62.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 89

Klein, D., Fink, B., Arold, B., Eisenreich, W., and Schwab, W. 2007. Functional char-
acterization of enone oxidoreductases from strawberry and tomato fruit. J. Agric.
Food Chem. 55, 6705–11.
Klesk, K., Qian, M., and Martin, R. 2004. Aroma extract dilution analysis of cv. Meeker
(Rubus idaeus L.) red raspberries from Oregon and Washington. J. Agric. Food
Chem. 52, 5155–61.
Knudsen, J.T. and Gershenzon, J. 2006 The chemical diversity of floral scent. In: N.
Dudareva and E. Pichersky (Eds.), Biology of Floral Scent. Boca Raton, FL: CRC
Press, pp. 27–52.
Koeduka, T., Fridman, E., and Gang, D.R. 2006. Eugenol and isoeugenol, characteristic
aromatic constituents of spices, are biosynthesized via reduction of a coniferyl alco-
hol ester. Proc. Natl Acad. Sci. USA 103, 10128–33.
Kumazawa, K., Wada, Y., and Masuda, H. 2008. Flavor contribution and formation of
epoxydecenal isomers in black tea. ACS Symp. Ser. 988, 136–46.
Kurobayashi, Y., Katsumi, Y., Fujita, A., Morimitsu,Y., and Kubota, K. 2008. Flavor
enhancement of chicken broth from boiled celery constituents. J. Agric. Food.
Chem. 56, 512–16.
Kurobayashi, Y., Kouno, E., Fujita, A., Morimitsu, Y., and Kubota, K. 2006. Potent
odorants characterize the aroma quality of leaves and stalks in raw and boiled cel-
ery. Biosci. Biotechnol. Biochem. 70, 958–65.
Lanzotti, V. 2006. The analysis of onion and garlic. J. Chromatogr. A 1112, 3–22.
Larsen, M. and Poll, L. 1992. Odour thresholds of some important aroma compounds in
strawberries. Z. Lebensm. Unters. Forsch. 195, 120–3.
Lea, A. G. H. 1995. Apple juice. In: P.R. Ashurts (Ed.), Production and Packaging
of Non-Carbonated Fruit Juices and Fruit Beverages, 3rd ed. Springer: Berlin,
Germany, pp. 153–96.
Lee, D. S., Nioche, P., Hamberg, M., and Raman, C. S. 2008. Structural insights into the
evolutionary paths of oxylipin biosynthetic enzymes. Nature 18, 455(7211), 363–8.
Lewinsohn, E., Ziv-Raz, I., Dudai, N., Tadmor, Y., Lastochkin, E., Larkov, O.,
Chaimovitsh, D., Ravid, U., Putievsky, E., and Pichersky, E. 2000. Biosynthesis of
estragole and methyl-eugenol in sweet basil (Ocimum basilicum L). Developmental
and chemotypic association of allylphenyl O-methyltransferase activities. Plant Sci.
160, 27–35.
Liavonchanka, A. and Feussner, I. 2006. Lipoxygenases: Occurrence, functions and
catalysis. J. Plant Physiol. 163, 348–357.
Li, Y., Qi, H., Jin, Y., Tian, X., Sui, L., and Qiu, Y. 2016. Role of ethylene in biosynthetic
pathway of related-aroma volatiles derived from amino acids in oriental sweet mel-
ons (Cucumis melo var. makuwa Makino). Sci. Hortic. 201, 24–35.
Lichtenthaler, H.K 1999. The 1-deoxy-d-xylulose-5-phosphate pathway of isoprenoid
biosynthesis in plants. Annu. Rev. Plant Physiol. Plant Mol. Biol. 50, 47–65.
Luan, F., Mosandl, A., Münch, A., and Wüst, M. 2005. Metabolism of geraniol in grape
berry mesocarp of Vitis vinifera L. cv. Scheurebe: Demonstration of stereoselective
reduction, E/Z isomerization, oxidation. Phytochemistry 66, 295–303.
Lunkenbein, S., Salentijn, E.M.J., Coiner, H.A., Boone, M.J., Krens, F.A., and Schwab,
W. 2006. Up- and down-regulation of Fragaria ·ananassa O-methyltransferase:
Impacts on furanone and phenylpropanoid metabolism. J. Exp. Bot. 57,
2445–53.
Maccarone, E., Campisi, S., Fallico, B., Rapisarda, P., and Sgarlata, R. 1998. Flavor com-
ponents of Italian orange juices. J. Agric. Food Chem. 46, 2293–8.
90 Food Aroma Evolution

Maffei, M.E. 2010. Changes in biosynthesis of aroma volatile compounds during on-tree
maturation of “Pink Lady” apples. South Afr. J. Bot. 76, 612–31.
Maga, J.A. 1989. Flavor formation and retention during extrusion. In: C. Mercier, P.
Linko, and J. Harper (Eds.), Extrusion Cooking. American Association of Cereal
Chemists: Saint Paul, MN, pp. 387–398.
Marconi, G., Ferradini, N., Russi, L., Concezzi, L., Veronesi, F., and Albertini, E. 2018.
Genetic characterization of the apple Germplasm collection in central Italy: The
value of local varieties. Front. Plant Sci. 10, 9, 1460.
Marilley, L. and Casey, M.G. 2004. Flavors of cheese products: Metabolic pathways,
analytical tools and identification of producing strains. Int. J. Food Microbiol. 90,
139–159.
Mathieu, S., Terrier, N., Procureur, J., Bigey, F., and Gunata, Z. 2005. A carotenoid
cleavage dioxygenase from Vitis vinifera L.: Functional characterization and expres-
sion during grape berry development in relation to C13 -norisoprenoid accumulation.
J. Exp. Bot. 56, 2721–31.
McGarvey, D.J. and Croteau, R. 1995. Terpenoid metabolism. Plant Cell 7, 1015–1026.
McGorrin, R. 2011. The significance of volatile sulfur compounds in food flavors, an
overview, Chapter 1. In: Qian et al. (Eds.), Volatile Sulfur Compounds in Food. ACS
Symposium Series. American Chemical Society: Washington, DC, pp. 1–29.
Misry, B.S., Reineccius, T., and Olson, L.K. 1997. Gaschromotography–olfactometry
for the determination of key odorants in foods. In: R. Marsili (Ed.), Techniques for
Analyzing Food Aroma. Marcel Dekker: New York, NY.
Moummou, H., Tonfack, L.B., Chervin, C., Benichou, M., Youmbi, E., Ginies, C., Latché,
A., Pech, J.C., and van der Rest, B. 2012. Functional characterization of SlscADH1,
a fruit-ripening‐associated short-chain alcohol dehydrogenase of tomato. J. Plant
Physiol. 169, 1435–44.
Murfitt, L.M., Kolosova, N., Mann, C.J., and Dudareva, N. 2000. Purification and char-
acterization of S-adenosyl-l-methionine: Benzoic acid carboxyl methyltransferase,
the enzyme responsible for biosynthesis of the volatile ester methyl benzoate in flow-
ers of Anthirrinum majus. Arch. Biochem. Biophys. 382, 145–51.
Naoumkina, M.A., Zhao, Q., Gallego-Giraldo, L., Dai, X., Zhao, P.X., and Dixon, R.A.
2010. Genome wide analysis of phenylpropanoid defence pathways. Mol. Plant
Pathol. 11(6), 829–46.
Negre, F., Kolosova, N., Knoll, J., Kish, C.M., and Dudareva, N. 2002 Novel S-adenosyl-
l-methionine:salicylic acid carboxyl methyltransferase, an enzyme responsible for
biosynthesis of methyl salicylate and methyl benzoate, is not involved in floral scent
production in snapdragon flowers. Arch. Biochem. Biophys. 406, 261–270.
Nieuwenhuizen, N.J., Green, S.A., and Chen, X. 2013. Functional genomics reveals that
a compact terpene synthase gene family can account for terpene volatile production
in apple. Plant Physiol 161, 787–804.
Ninio, R., Lewinsohn, E., Mizrahi, Y., and Sitrit, Y. 2003. Changes in sugars, acids and
volatiles during ripening of koubo (Cereus peruvianus [L.] Miller) fruits. J. Agric.
Food Chem. 51, 797–801.
Noel, J.P., Dixon, R.A., Pichersky, E., Zubieta, C., and Ferrer, J.L. 2003. Structural,
functional, and evolutionary basis for methylation of plant small molecules. Recent
Adv. Phytochem. 37, 37–58.
Nogueira, J.M.F., Fernandes, P.J.P., and Nascimento, A.M.D. 2003. Composition of
volatiles of banana cultivars from Madeira island. Phytochem. Anal. 14, 87–90.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 91

Noordermeer, M.A., Veldink, G.A., and Vliegenthart, J.F.G. 2001 .Fatty acid hydroper-
oxide lyase: a plant cytochrome P450 enzyme involved in wound healing and pest
resistance. Chembiochem 2, 494–504.
Ogundiwin, E.A., Peace, C.P., Gradziel, T.M., Parfitt, D.E., Bliss, F.A.I., and Crisosto,
C. H. A. 2009. Fruit quality gene map of Prunus. BMC Genom. 10, 587.
Ohara, K., Ujihara, T., Endo, T., Sato, F., and Yazaki, K. 2003. Limonene production in
tobacco with Perilla limonene synthase cDNA. J. Exp. Bot. 54, 2635–42.
Ong, P.K.C. and Acree, T.E. 1998. Gas chromatography olfactory analysis of lychee
(Litchi chinesis Sonn.). J. Agric. Food Chem. 46, 2282–6.
Orlova, I., Marshall-Colón, A., Schnepp, J., Wood, B., Varbanova, M., Fridman, E.,
Blakeslee, J.J., Peer, W.A., Murphy, A.S., Rhodes, D., Pichersky, E., and Dudareva, N.
2006. Reduction of benzenoid synthesis in petunia flowers reveals multiple pathways
to benzoic acid and enhancement in auxin transport. Plant Cell 18, 3458–75.
Paillard, N.M.M. 1990. The flavour of apples, pears and quinces. In: L.D. Morton and
A.J. MacLeod (Eds.), Food Flavours, Part C: The Flavour of Fruits. Elsevier Science
Publishing Company Inc.: Amsterdam, the Netherlands, pp. 1–41.
Pak, F.E., Gropper, S., Dai, W.D., Havkin-Frenkel, D., and Belanger, F.C. 2004.
Characterization of a multifunctional methyltransferase from the orchid Vanilla
planifolia. Plant Cell Rep. 22, 959–66.
Parker, J.K. 2015. Introduction to aroma compounds in foods. In: J.K. Parker, S. Elmore,
and L. Methven (Eds.), Flavour Development, Analysis and Perception in Food and
Beverages. Woodhead Publishing: Cambridge.
Pazouki, L. and Niinemets, Ü. 2016. Multi-substrate Terpene synthases: Their occur-
rence and physiological significance. Front. Plant Sci. 7, 1019.
Pazouki, L., Memari, H.R., Kännaste, A., Bichele, R., and Niinemets Ü. 2015. Germacrene
A synthase in yarrow (Achillea millefolium) is an enzyme with mixed substrate spec-
ificity: Gene cloning, functional characterization and expression analysis. Front.
Plant Sci. 6, 111.
Pérez, A.G., Olías, R., Sanz, C., Olías J.M. 1996. Furanones in strawberries: Evolution
during ripening and postharvest shelf life. J. Agric. Food Chem. 44, 3620–4.
Pérez, A.G., Ríos, J.J., Sanz, C., and Olías, J.M. 1992. Aroma components and free
amino acids in strawberry variety Chandler during ripening. J. Agric. Food Chem.
40, 2232–5.
Pérez, A.G., Olías, R., Luaces, P., and Sanz, C. 2002. Biosynthesis of strawberry aroma
compounds through amino acid metabolism. J. Agric. Food Chem. 50, 4037–4042.
Pérez, A.G., Sanz, C., Olías, R., Ríos, J.J., and Olías, J.M. 1996. Evolution of strawberry
alcohol acyltransferase activity during fruit development and storage. J. Agric. Food
Chem. 44, 3286–90.
Pérez, L.C. and Yaylayan, V.A. 2004. Origin and mechanistic pathways of formation of
the parent furan—A food toxicant. J. Agric. Food Chem. 52, 6830–6.
Pernolle, J.C. and Briand, L. 2004. Structural recognition between odorants, olfactory-
binding proteins and olfactory receptors: First events on odour coding. In: A. Taylor
and D. Roberts (Eds.), Flavor Perception. Blackwell: Oxford.
Pichersky, E., Noel, J.P., and Dudareva, N. 2006. Biosynthesis of plant volatiles: Nature’s
diversity and ingenuity. Science 311, 808–811.
Podstolski, A., Havin-Frenkel, D., Malinowski, J., Blount, J.W., Kourteva, G., and Dixon,
R.A. 2002. Unusual 4–hydroxybenzaldehyde synthase activity from tissue cultures
of vanilla orchid Vanilla planifolia. Phytochem. 61, 611–20.
92 Food Aroma Evolution

Ponce, E. 2006. Aroma y Sabor, Chapter 4. In: Salvador Badui Dergal (Ed.), Química de
los Alimentos, 4th ed. Pearson Education: Mexico.
Pott, M.B., Pichersky, E., and Piechulla, B. 2002. Evening-specific oscillations of scent
emission, SAMT enzyme activity, and SAMT mRNA in flowers of Stephanotis flo-
ribunda. J. Plant Physiol. 159, 925–34.
Preethi, P., Soorianathasundaram, K., and Subramanian, K.S. 2014. Aroma volatile com-
pounds of mango cultivars neelum and banganapalli. Biochem. Cell. Arch. 14(2),
2–6.
Prestage, S., Linforth, R.S.T., Taylor, A.J., Lee, E., Speirs, J., and Schuch, W. 1999.
Volatile production in tomato fruit with modified alcohol dehydrogenase activity. J.
Sci. Food Agric. 79, 131–6.
Qian, M. and Reineccius, G. 2003. Static headspace and aroma extract dilution analysis
of Parmigiano Reggiano cheese. J. Food Sci. 68, 794–8.
Qian, M., Nelson, C., and Bloomer, S. 2002. Evaluation of fat-derived aroma compounds
in blue cheese by dynamic headspace GC/olfactometry-MS. J. Am. Oil Chem. Soc.
79, 663–7.
Qualley, A. and Dudareva, N. 2001. Plant volatiles. In: Encyclopedia of Life Sciences.
John Wiley & Sons: Chichester, UK.
Raab, T., López-Ráez, J.A., Klein, D., Caballero, J.L., Moyano, E., Schwab, W., and
Muñoz-Blanco, J. 2006. FaQR, required for the biosynthesis of the strawberry fla-
vor compound 4-hydroxy-2,5- dimethyl-3(2H)-furanone, encodes an enone oxido-
reductase. Plant Cell 18, 1023–37.
Rao, S.R. and Ravishankar, G.A. 2000. Vanilla flavour: Production by conventional and
biotechnological routes. J. Sci. Food. Agric. 80, 289–304.
Rasulov, B., Talts, E., Kaennaste, A., and Niinemets, U. 2015. Bisphosphonate inhibi-
tors reveal a large elasticity of plastidic isoprenoid synthesis pathway in isoprene-
emitting hybrid aspen. Plant Physiol. 168, 532–548.
Reineccius, G. 2006. Flavor Chemistry and Technology, 2nd ed. CRC Press: Boca
Raton, FL.
Ribnicky, D.M., Shulaev, V., and Raskin, I. 1998. Intermediates of salicylic acid biosyn-
thesis in tobacco. Plant Physiol. 118, 565–72.
Ringer, K.L., Davis, E.M., and Croteau, R. 2005. Monoterpene metabolism: Cloning,
expression, and characterization of (−)-isopiperitenol/(−)-carveol dehydrogenase of
peppermint and spearmint. Plant Physiol. 137, 863–872.
Rizzolo, A., Grassi, M., and Zerbini, P.E. 2006. Influence of harvest date on ripening
and volatile compounds in the scab-resistant apple cultivar “Golden Orange.” J.
Horticult. Sci. Biotechnol. 81, 681–90.
Rodríguez, A., Alquézar, B., and Peña, L. 2013. Fruit aromas in mature fleshy fruits as
signals of readiness for predation and seed dispersal. New Phytol. 197, 36–48.
Rodriguez-Concepción, M. and Boronat, A. 2002. Elucidation of the methylerythritol
phosphate pathway for isoprenoid biosynthesis in bacteria and plastids. A metabolic
milestone achieved through genomics. Plant Physiol. 130, 1079–89.
Rohdich, F., Hecht, S., Gartner, K., Adam, P., Krieger, C., Amslinger, S., Arigoni, D.,
Bacher, A., and Eisenreich, W. 2002. Studies on the nonmevalonate terpene biosyn-
thetic pathway: Metabolic role of IspH (LytB) protein. Proc. Natl Acad. Sci. USA
99, 1158–63.
Rohmer, M. 2003. Mevalonate-independent methylerythritol phosphate pathway for iso-
prenoid biosynthesis: Elucidation and distribution. Pure Appl. Chem. 75, 375–388.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 93

Ross, J.R., Nam, K.H., D’Auria, J.C., and Pichersky, E. 1999 S-adenosyl-
l-methionine:salicylic acid carboxyl methyltransferase, an enzyme involved in floral
scent production and plant defense, represents a new class of plant methyltransfer-
ases. Arch. Biochem. Biophys. 367, 9–16.
Ryona, I., Pan, B.S., Intrigliolo, D.S., Lakso, A.N., and Sacks, G.L. 2008. Effects of
cluster light exposure on 3-isobutyl 2-methoxypyrazine accumulation and degrada-
tion patterns in red wine grapes (Vitis vinifera L. cv. Cabernet Franc). J. Agric. Food
Chem. 56, 10838–46.
Saffert, A., Hartmann-Schreier, J., Schön, A., and Schreier, P. 2000. A dual function
α‐dioxygenase‐peroxidase and NAD+ oxidoreductase active enzyme from germinating
pea rationalizing α-oxidation of fatty acids in plants. Plant Physiol. 123, 1545–1551.
Salas, J.J., Sanchez, C., Garcia-Gonzalez, D.L., and Aparicio, R. 2005. Impact of the sup-
pression of lipoxygenase and hydroperoxide lyase on the quality of the green odor in
green leaves. J. Agric. Food Chem. 53, 1648–1655.
Sanz, C., Olías, J.M., and Peéez, A.G. 1997. Phytochemistry of fruit and vegetables. In:
Aroma Biochemistry of Fruits and Vegetables. Oxford University Press Inc.: New
York pp. 125–55.
Sasaki, M., Nunomura, N., and Matsudo, T. 1991. Biosynthesis of 4-hydroxy-2(or
5)-ethyl-5(or 2)-methyl-3(2H)-furanone by yeasts. J. Agric. Food Chem. 39, 934–8.
Schieberle, P., Ofner, S., and Grosch, W. 1990. Evaluation of potent odorants in cucum-
bers (Cucumis sativus) and muskmelons (Cucumis melo) by aroma extract dilution
analysis. J. Food Sci. 55, 193–195.
Schmidt, H., Kurtzer, R., Eisenreich, W., and Schwab, W. 2006. The carotenase AtCCD1
from Arabidopsis thaliana is a dioxygenase. J. Biol. Chem. 281, 9845–51.
Schnepp, J. and Dudareva, N. 2006 Floral scent: Biosynthesis, regulation, and genetic
modifications. In: C. Ainsworth (Ed.), Flowering and its Manipulation. Oxford:
Blackwell Publishing, pp. 240–57.
Schnermann, P. and Schieberle, P. 1997. Evaluation of key odorants in milk chocolate and
cocoa mass by aroma extract dilution analyses. J. Agric. Food Chem. 45, 867–872.
Schwab, W. and Roscher, R. 1997. 4-Hydroxy-3(2H)-furanones: Natural and Maillard
product. Res. Dev. Phytochem. 1, 643–673.
Schwab, W. and Schreier, P. 2002. Enzymic formation of flavor volatiles from lipids. In:
T.M. Kuo and H.W. Gardner (Eds.), Lipid Biotechnology. Marcel Dekker: New
York, pp. 293–318.
Schwab, W., Davidovich-Rikanati, R., and Lewinsohn, E. 2008. Biosynthesis of plant-
derived flavor compounds. Plant J. 54(4), 712–32.
Schwab, W., Schaart, J. G., and Rosati, C. 2009. Molecular biology research in Fragaria.
In: K.M. Folta and S.E. Gardiner (Eds.), Genetics and Genomics of Rosaceae, vol.
6. Springer: New York, pp. 457–86.
Seo, H.S., Song, J.T., Cheong, J.J., Lee, Y.W., Hwang, I., Lee, J.S. and Choi, Y.D. 2001.
Jasmonic acid carboxyl methyltransferase: A key enzyme for jasmonate-regulated
plant responses. Proc. Natl Acad. Sci. USA 98, 4788–93.
Shalit, M., Katzir, N., Tadmor, Y., Larkov, O., Burger, Y., Schalechet, F., Lastochkin, E.,
Ravid, U., Amar, O., Edelstein, M., Karchi, Z., and Lewinsohn, E. 2001. Acetyl-
CoA: Alcohol acetyl transferase activity and aroma formation in ripening melon
fruits. J. Agric. Food Chem. 49, 794–9.
Shaw, P.E. and Wilson, C.W. 1981. Importance of nootkatone to the aroma of grapefruit
oil and the flavor of grapefruit juice. J. Agric. Food Chem. 29, 677–9.
94 Food Aroma Evolution

Shoji, K., Takeshi, M., Joichi, A., and Yomogida, K. 2000. Sedative for vaporizing inha-
lation and sedative perfume composition containing the same as active ingredient.
Japan Patent Gazette, JP2000-086478. Tokyo: Japan Patent Office.
Simkin, A., Schwartz, S., Auldridge, M., Taylor, M., and Klee, H.J. 2004. The tomato
carotenoid cleavage dioxygenase 1 genes contribute to the formation of the flavor
volatiles b-ionone, pseudoionone, and geranylacetone. Plant J. 40, 882–892.
Singh, Z., Singh, R., Sane, V., and Nath, P. 2013. Mango: Postharvest biology and bio-
technology. Crit. Rev. Plant Sci. 32, 217–236.
Song, J. 1994. Enfluss verschiedener Erntzeitpunkte auf die Fruchtreife unter besonderer
Beruecksichtigung der Aromabildung bei Aepfeln, Tomaten und Erdbeeren. Ulrich
E. Grauer: Stuttgart, Germany.
Song, J. and Forney, C.F. 2008. Flavour volatile production and regulation in fruit. Can.
J. Plant Sci. 88, 537–50.
Speirs, J., Lee, E., Holt, K., Yong‐Duk, K., Steele Scott, N., Loveys, B., and Schuch,
W. 1998. Genetic manipulation of alcohol dehydrogenase levels in ripening tomato
fruit affects the balance of some flavor aldehydes and alcohols. Plant Physiol. 117,
1047–58.
Surburg, H. and Panten, J. (Eds.). 2005. Common Fragrance and Flavor Materials:
Preparation, Properties and Uses. Wiley-VCH: Weinheim, Germany.
Tadmor, Y., Fridman, E., Gur, A., Larkov, O., Lastochkin, E., Ravid, U., Zamir, D., and
Lewinsohn, E. 2002. Identification of malodorous, a wild species allele affecting
tomato aroma that was selected against during domestication. J. Agric. Food Chem.
50, 2005–2009.
Tarshis, L.C., Proteau, P.J., Kellogg, B.A., Sacchettini, J.C., and Poulter, C.D. 1996.
Regulation of product chain length by isoprenyl diphosphate synthases. Proc. Natl.
Acad. Sci. USA 93, 15018–23.
Tarshis, L.C., Yan, M., Poulter, C.D., and Sacchettini, J.C. 1994. Crystal structure
of recombinant farnesyl diphosphate synthase at 2.6 A° resolution. Biochem. 33,
10871–10877.
Taylor, A.J., Linforth, R.S.T., Baek, I., Brauss, M.S., Davidson, J.M., and Gray, D.A.
2000. Flavor release and flavor perception. In: S.J. Risch and C.-T. Ho (Eds.),
Flavor Chemistry: Industrial and Academic Research. American Chemical Society:
Washington, DC, pp. 151–65.
Theron, L., Tournayre, P., Kondjoyan, N., Abouelkaram, S., Sante-Lhoutellier, V., and
Berdague, J.-L. 2010. Analysis of the volatile profile and identification of odour-
active compounds in Bayonne ham. Meat Sci. 85, 453–60.
Tholl, D. 2006. Terpene synthases and the regulation, diversity and biological roles of
terpene metabolism. Curr. Opin. Plant Biol. 9, 297–304.
Tiefel, P. and Berger, R.G. 1993. Seasonal variation of the concentrations of maltol and
maltol glucoside in leaves of Cercidiphyllum japonicum. J. Sci. Food Agric. 63,
59–61.
Tieman, D., Taylor, M., Schauer, N., Fernie, A.R., Hanson, A.D., and Klee, H.J. 2006.
Tomato aromatic amino acid decarboxylases participate in synthesis of the flavor
volatiles 2-phenylethanol and 2-phenylacetaldehyde. Proc. Natl Acad. Sci. USA
103, 8287–92.
Tomiyama, K., Aoki, H., Oikawa, T., Sakurai, K., Kashara, Y., and Kawakami, Y. 2012.
Characteristic volatile components of Japanese sour citrus fruits: Yuzu, Sudachi and
Kabosu. Flavour Fragr. J. 27, 341–55.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 95

Torregrosa, L., Pradal, M., Souquet, J.M., and Rambert, M. 2008. Manipulation of
VvAdh to investigate its function in grape berry development. Plant Sci. 174, 149–55.
Tressl, R. and Albrecht, W. 1986. Biogenesis of aroma compounds through acyl path-
ways. In: T.H. Parliment and R. Croteau (Eds.), Biogeneration of Aromas. ACS:
Washington, DC, pp. 114–33.
Tucker, G.A. 1993. Introduction. In: G.B. Seymour, J.E. Taylor, Gregory A. Tucker
(Eds.), Biochemistry of Fruit Ripening. Chapman & Hall, Springer: Dordrecht.
Turner, G.W. and Croteau, R. 2004. Organization of monoterpene biosynthesis in
Mentha. Immunocytochemical localizations of geranyl diphosphate synthase, lim-
onene-6-hydroxylase, isopiperitenol dehydrogenase, and pulegone reductase. Plant
Physiol. 136, 4215–4227.
Uji, Y., Ozawa, R., Shishido, H., Taniguchi, S., Takabayashi, J., Akimitsu, K., and Gomi,
K. 2015. Isolation of a sesquiterpene synthase expressing in specialized epithelial
cells surrounding the secretory cavities in rough lemon (Citrus jambhiri). J. Plant
Physiol. 15, 180, 67–71.
Van de Poel, B, Vandendriessche, T., Hertog, M., Nicolai, B., and Geeraerd, A. 2014.
Detached ripening of non-climacteric strawberry impairs aroma profile and fruit
quality. Postharvest Biol. Technol. 95, 70–80.
Van Gemert, L.J. 2000. Compilations of Odour Threshold Values in Air and Water.
Boelens Aroma Chemical Information Service.
VCF Online. 2016. Volatile compounds in food 16.3. Triskelion. http:​//www​.vcf-​onlin​
e.nl/​VcfCo​mpoun​ds.cf​m, accessed March 4, 2019.
Verkerk, R., Schreiner, M., Krumbein, A., Ciska, E., Holst, B., Rowland, I., De Schrijver,
R., Hansen, M., Gerhäuser, C., Mithen, R., and Dekker, M. 2009. Glucosinolates in
Brassica vegetables: The influence of the food supply chain on intake, bioavailability
and human health. Mol. Nutr. Food Res. 53(Suppl. 2), S219.
Villatoro, C., Altisent, R., Echeverría, G., Graell, J., López, M. L., and Lara, I. 2008.
Changes in biosynthesis of aroma volatile compounds during on-tree maturation of
“Pink Lady” apples. Postharvest Biol. Technol. 47, 286–95.
Visai, C. and Vanoli, M. 1997. Volatile compound production during growth and ripen-
ing of peaches and nectarines. Sci. Horticult. 70, 15–24.
Vogt, J., Schiller, D., Ulrich, D., Schwab, W., and Dunemann, F. 2013. Identification of
lipoxygenase (LOX) genes putatively involved in fruit flavour formation in apple
(Malus × domestica). Tree Genet. Genomes 9, 1493–511.
Voirol, E. and Daget, N. 1987. Nasal and retronasal olfactory perception of a meat
aroma. In: M. Martens, G.A. Dalen, and H. Russwurm (Eds.), Flavor Science and
Technology. Wiley: New York, NY.
Vora, J.D., Matthews, R.F., Crandall, P.G., and Cook, R. 1983. Preparation and chemi-
cal composition of orange oil concentrates. J. Food Sci. 48, 1197–9.
Vranová, J. and Ciesarová, Z. 2009. Furan in food—A review. Food Research Institute,
Bratislava, Slovak Republic. Czech J. Food Sci. 27(1), 1–10.
Vrhovsek, U., Lotti, C., Masuero, D., Carlin, S., Weingart, G., and Mattivi, F. 2014.
Quantitative metabolic profiling of grape, apple and raspberry volatile compounds
(VOCs) using a GC/MS/MS method. J. Chromatogr. B. 966, 132–9.
Walton, N.J., Mayer, M.J., and Narbad, A. 2003 Vanillin. Phytochem. 63, 505–515.
Wang, C., Chin, C.-K., Ho, C.-T., Hwang, C.-F., Polashock, J., and Martin, C. 1996.
Changes of fatty acids and fatty acid‐derived flavor compounds by expressing the
yeast Δ‐9 desaturase gene in tomato. J. Agric. Food Chem. 44, 3399–402.
96 Food Aroma Evolution

Wang, J. and De Luca, V. 2005. The biosynthesis and regulation of biosynthesis of

Concord grape fruit esters, including “foxy” methyl anthranilate. Plant J. 44,
606–19.
Wang, J., Dudareva, N., Bhakta, S., Raguso, R.A., and Pichersky, E. 1997. Floral scent
production in Clarkia breweri (Onagraceae). II. Localization and developmental
modulation of the enzyme SAM:(iso)eugenol O-methyltransferase and phenylpro-
panoid emission. Plant Physiol. 114, 213–221.
Wang, W., Khalil-Ur-Rehman, M., Feng, J., and Tao, J. 2017. RNA-seq based transcrip-
tomic analysis of CPPU treated grape berries and emission of volatile compounds. J.
Plant Physiol. 218, 155–66.
Wang, Y.J., Yang, C.X., Li, S.H., Yang, L., Wang, Y.N., Zhao, J.B., and Jiang, Q. 2009.
Volatile characteristics of 50 peaches and nectarines evaluated by HP-SPME with
GC-MS. Food Chem. 116, 356–64.
Weckerle, B., Bastl-Borrmann, R., Richling, E., Hör, K., Ruff, C., and Schreier, P. 2001.
Cactus pear (Opuntia ficus indica) flavor constituents: Chiral evaluation (MDGC–
MS) and isotope ratio (HRGC–IRMS) analysis. Flav. Frag. J. 16, 360–363.
Wein, M., Lavid, N., Lunkenbein, S., Lewinsohn, E., Schwab, W., and Kaldenhoff, R.
2002. Isolation, cloning and expression of a multifunctional O-methyltransferase
capable of forming 2,5- dimethyl-4-methoxy-3(2H)-furanone, one of the key aroma
compounds in strawberry fruits. Plant J. 31, 755–765.
Weiss, E.A. 1997. Essential Oil Crops. CAB International: Wallingford, UK.
Wildermuth, M.C. 2006. Variations on a theme: Synthesis and modification of plant
benzoic acids. Curr. Opin. Plant Biol. 9, 288–296.
Winterhalter, P. 1991. Fruits IV. In: H. Maarse (Ed.), Volatile Compounds in Foods and
Beverage. New York: Marcel Dekker Inc., pp. 389–409.
Winterhalter, P. and Rouseff, R. 2001. Carotenoid-derived aroma compounds: An intro-
duction. In: P. Winterhalter and R. Rouseff (Eds.), Carotenoid-Derived Aroma
Compounds. American Chemical Society, pp.1–17.
Winterhalter, P. and Rouseff, R.L. (Eds.). 2002. Carotenoid derived aroma compounds:
An introduction. In: Carotenoid-Derived Aroma Compounds, ACS Symposium
Series 802. American Chemical Society: Washington, DC, pp. 1–17.
Winterhalter, P. and Schreier, P. 1994. C13-Norisoprenoid glycosides in plant tissues:
An overview on their occurrence, composition and role as flavour precursors. Flav.
Frag. J. 9, 281–7.
Withers, S.T. and Keasling, J.D. 2007. Biosynthesis and engineering of isoprenoid small
molecules. Appl. Microbiol. Biotechnol. 73, 980–990.
Wu, Q., Tao, X., Ai, X., Luo, Z., Mao, L., Ying, T., and Li, L. 2018. Contribution of
abscisic acid to aromatic volatiles in cherry tomato (Solanum lycopersicum L.) fruit
during postharvest ripening. Plant Physiol. Biochem. 130, 205–14.
Wu, S., Schalk, M., Clark, A., Miles, R.B., Coates, R., and Chappell, J. 2006. Redirection
of cytosolic or plastidic isoprenoid precursors elevates terpene production in plants.
Nat. Biotechnol. 24, 1441– 7.
Wu, S.Q., Watanabe, N., Mita, S., Dohra, H., Ueda, Y., Shibuya, M., and Ebizuka, Y.
2004. The key role of phloroglucinol O–methyltransferase in the biosynthesis of
Rosa chinensis volatile 1,3,5- trimethoxybenzene. Plant Physiol. 135, 95–102.
Wyllie, S.G. and Fellman, J.K. 2000. Formation of volatile branched chain esters in
bananas (Musa sapientum L.). J. Agric. Food Chem. 48, 3493–6.
 Aroma Compounds (Description, Biosynthesis, and Regulation) 97

Xisto, A.L., Vilas Boas, E., Nunes, E., Montero, B., and Guerreiro, M.C. 2012. Volatile
profile and physical, chemical, and biochemical changes in fresh cut watermelon
during storage. Ciênc. Tecnol. Aliment. 32, 173–178.
Yahyaoui, F., Wongs-Aree, C., Latché, A., Hackett, R., Grierson, D., and Pech, J.C.
2002. Molecular and biochemical characteristics of a gene encoding an alcohol acyl-
transferase involved in the generation of aroma volatile esters during melon ripen-
ing. Eur. J. Biochem. 269, 2359–2366.
Yang, C.X., Wang, Y.J., Wu, B.H., Fang, J.B., and Li, S.H. 2011a. Volatile compounds
evolution of three table grapes with different flavor during and after maturation.
Food Chem. 128, 823–30.
Yang, X.T., Song, J., Fillmore, S., Pang, X.Q., and Zhang, Z.Q. 2011b. Effect of high
temperature on color, chlorophyll fuorescence and volatile biosynthesis in green-ripe
banana fruit. Postharvest Biol. Technol. 62, 246–57.
Yauk, Y.K., Chagné, D., Tomes, S., Matich, A.J., Wang, M.Y., Chen, X., Maddumage,
R., Hunt, M.B., Rowan, D.D., and Atkinson, R.G. 2015. The O-methyltransferase
gene MdoOMT1 is required for biosynthesis of methylated phenylpropenes in ripe
apple fruit. Plant J. 82, 937–950.
Yauk, Y.K., Souleyre, E.J.F., Matich, A.J., Chen, X., Wang, M.Y., Plunkett, B., Dare,
A.P., Espley, R.V., Tomes, S., Chagné, D., and Atkinson, R.G. 2017. Alcohol acyl
transferase 1 links two distinct volatile pathways that produce esters and phenylpro-
penes in apple fruit. Plant J. 91(2), 292–305.
Yaylayan, V.A. 2006. Precursors, formation and determination of furan in food. J.
Verbrauch. Lebensm. 1, 5–9.
Yomogida, K. 1992. Scent of modern roses. Koryo 175, 65–89.
Young, J.C., Chu, C.L.G., Lu, X.W., and Zhu, H.H. 2004 Ester variability in apple vari-
eties as determined by solid-phase microextraction and gas chromatography-mass
spectrometry. J. Agric. Food Chem. 52, 8086–8093.
Young, J.C., George Chu, C.L., Lu, X., and Zhu, H. 2005. Ester variability in apple vari-
eties as determined by solid-phase microextraction and gas chromatography–mass
spectrometry. J. Agric. Food Chem. 52(26), 8086–93.
Zubieta, C., Ross, J.R., Koscheski, P., Yang, Y., Pichersky, E., and Noel, J.P. 2003.
Structural basis for substrate recognition in the salicylic acid carboxyl methyltrans-
ferase family. Plant Cell 15, 1704–16.
 Chapter 5
Orthonasal and
Retronasal Olfaction
Pengfei Han and Thomas Hummel

CONTENTS

5.1 Basics of Human Olfaction 99

5.2 Orthonasal and Retronasal Olfaction 100
5.3 Methods to Investigate Orthonasal Olfaction 101
5.4 Method to Study Retronasal Olfaction 102
5.5 Differences between Orthonasal and Retronasal Olfaction 102
5.5.1 Differences at the Psychophysical and Peripheral Levels 103
5.5.2 Differentiations at the Neural Level 103
5.6 The Role of Ortho- and Retronasal Olfaction in Food Perception 104
5.6.1 Orthonasal Food Odor Perception 105
5.6.2 Retronasal Food Odor Perception 105
5.7 Factors Influencing Olfactory Perception 106
5.7.1 Gender 106
5.7.2 Age 107
5.7.3 Genetic Factors 108
5.7.4 Oral Environment (Saliva and Microbiota) 109
5.7.5 Hormonal Levels 110
5.7.6 Olfactory Disorders 110
5.8 Conclusions 111
Acknowledgment 111
References 111

5.1 BASICS OF HUMAN OLFACTION

Olfaction is phylogenetically considered the oldest sense, yet remains the least well under-
stood (Philpott et al., 2008). For humans, the sense of smell is more important than is
generally realized (McGann, 2017), as suggested by the important role it plays in the evo-
lution of human diet, food ingestion, threat detection, and social behaviors (Shepherd,
2004; Stevenson, 2010).
Olfaction comprises the chemosensory modality dedicated to detecting low concen-
trations of airborne, volatile chemical substances (Ache and Young, 2005). The ascending
pathway for odor perception begins in the olfactory epithelium, where odor molecules
make contact with sensory endings of olfactory receptor neurons. The olfactory epithe-
lium comprises 6–30 million neurons. They express hundreds of receptor proteins which
enable detection and discrimination of thousands of odorants (Menashe and Lancet,

99
100 Food Aroma Evolution

2006). The axons of sensory neurons in the olfactory epithelium project to the olfactory
bulb—the first olfactory brain structure, in which different odor molecules are repre-
sented by different patterns of spatial activities. Axonal projections from these cells are
conveyed via the lateral olfactory tract onto the primary and secondary olfactory cortex
(Gottfried, 2010). Thus, an odor perception (e.g., banana or bacon) is the brain’s inter-
pretation of the activation pattern of many peripheral sensory neurons that are differen-
tially sensitive to a wide variety of odor molecules (Firestein, 2001).
There are different dimensions commonly used to define odor perception, which
include odor quality and intensity, as well as the affective (hedonic) dimension, or the
familiarity with an odor. For humans, the most common feature of olfactory function
to be quantified is the ability to identify an odor (odor identification), but there are also
other aspects of olfactory functions, such as the ability to discriminate between odors
(odor discrimination) or to detect odors at low concentrations (odor threshold).

5.2 ORTHONASAL AND RETRONASAL OLFACTION

In humans and other similarly behaving animals, there are two ways to sample and trans-
port odor molecules from the outside atmosphere to the olfactory epithelium. Orthonasal
olfaction occurs when odor molecules are sampled via nasal inhalation or sniff; odors are
delivered with inhaled air to the olfactory receptors through the nose. Retronasal olfac-
tion is naturally initiated by the chewing and swallowing of foods. Volatile molecules
are released into the back of the oral cavity and travel through the nasopharynx during
exhalation and subsequently stimulate receptors on the olfactory epithelium in the nose
(Figure 5.1). Because retronasal sensations are typically perceived through the mouth (Lim
and Johnson, 2011), retronasal sensations are commonly referred to as “taste.” In addi-
tion, smell–taste confusions are so profound that there are even languages (e.g., Swiss
German) that do not have a proper word for “smell” but largely refer to it as “taste.”

FIGURE 5.1 The orthonasal (dark gray) and the retronasal (light gray) route for olfaction.
 Orthonasal and Retronasal Olfaction 101

In this chapter, the methods used to investigate orthonasal and retronasal olfaction
will be first introduced. Although the same set of olfactory receptor neurons are being
activated, olfactory perceptions through the orthonasal and retronasal routes are differ-
ent at both the psychophysical and neural levels. In addition, this chapter will compare
and discuss the roles the two olfactory routes play in food perception and nutrition.
Finally, factors that influence olfaction in humans will be summarized.

5.3 METHODS TO INVESTIGATE ORTHONASAL OLFACTION

Among the most widely used orthonasal olfactory tests is the “Sniffin’ Sticks” battery
(Burghart Messtechnik GmbH, Wedel, Germany) (Figure 5.2) (Kobal et al., 1996). The
Sniffin’ Sticks test is based on felt-tip pen-like devices containing common odors selected
specifically to be applicable in the general European population (Hummel et al., 1997).
The classic test consists of three parts: odorant threshold, discrimination, and identifica-
tion (TDI) abilities (Hummel et al., 1997), which allows a comprehensive description
of the overall olfactory function. The Sniffin’ Sticks test was also modified into other
versions; for example, the “Sniffin’ Kids” test for children between 6 to 17 years old
(Schriever et al., 2014). Another version is the extended test for odor identification with
32 items, which was developed in order to create more precise tools enabling repeated,
longitudinal testing of small changes of olfactory subfunction (Haehner et al., 2009;
Sorokowska et al., 2015a). A major advantage of the “Sniffin’ Sticks” is the availability of
normative data generated in large groups of healthy subjects which can be applied to both
healthy people and patients with olfactory dysfunctions (Hummel et al., 2007).
Another popular test for olfactory identification is the University of Pennsylvania Smell
Identification Test (UPSIT), which contains 40 microencapsulated odorants on the paper
of the test kits (Doty et al., 1984b). The identification scores of the UPSIT test have been

FIGURE 5.2 The “Sniffin’ Sticks” smell test with its three parts: Odor identification
(foreground tube rack, with list of verbal and graphical descriptors of odors); the odor
discrimination test (middle tube rack); and the phenyl ethyl alcohol odor threshold test
(back ground tube rack).
102 Food Aroma Evolution

shown to be comparable with the Sniffin’ Sticks identification test (Hugh et al., 2015). The
UPSIT offers the advantage of being able to be self-administered by most participants. By
microencapsulating odorants, olfactory testing can reach large amounts of subjects, for
example, through mailing. In addition, normative data based on results from nearly 4000
people are available, and an individual’s percentile rank can be determined from tables
based upon data from healthy people of the same age and gender (Doty, 1997).

5.4 METHOD TO STUDY RETRONASAL OLFACTION

The key issue for the investigation of retronasal olfaction is the delivery of odorous stim-
uli to the back of the throat. Psychophysical tests to explore retronasal olfactory function
include flavor identification tests such as the “taste powders test” (Heilmann et al., 2002;
for an international version see Croy et al. [2014]), or the “candy smell test” (Renner
et al., 2009). Taste powders are a test kit with 20 grocery-available food powders. The
powders are administered to the middle of the tongue, and subjects are asked to identify
the “taste” from a list of four items using a multiple forced choice procedure (Heilmann
et al., 2002). Similarly, the candy smell test consists of 23 hard candies, each containing
one unique aroma. The aromas are congruent to sweet taste. Both tests show good test–
retest reliability and are commonly used in clinical measurement of retronasal olfaction.
Although these tests address the retronasal application of odors, they cannot allow
direct comparisons to stimuli that are presented in front of the nose, simply because the
oral administration of odors may produce gustatory, thermal, and mechanical sensa-
tions which may interact with olfactory mediated sensations (Spence, 2013). To avoid
this situation, some researchers have placed odors in containers and then placed them
on the tongue to deliver odor retronasally (Sun and Halpern, 2005), while others asked
subjects to sniff the headspace of an odorous liquid or inhale the same headspace through
the mouth followed by nasal exhalation (Burdach et al., 1984; Voiol and Daget, 1986).
However, these methods have limitations since the odor concentration in the oral cavity
is non-predictable and the mechanical stimulation of intraoral surfaces is not fully sup-
pressed (Bojanowski and Hummel, 2012).
A more defined retronasal stimulation is possible when using a computer-controlled
air-dilution olfactometer (Kobal, 1981), which allows researchers to control the concentra-
tion and flow rate of an odor stimulus. Under endoscopic control, Heilmann and Hummel
inserted a tube below the lower turbinate of the nasal cavity and placed its opening in
the epipharynx (Heilmann and Hummel, 2004). In this way, the odor stimuli could be
presented to the olfactory epithelium retronasally without additional activation of sensors
in the oral cavity. Using the computer-controlled olfactometer, this form of stimulation
provides excellent control over both odor concentration and stimulus onset/offset.

5.5 DIFFERENCES BETWEEN ORTHONASAL

AND RETRONASAL OLFACTION

Although both routes deliver the odor molecules to the same receptor fields in the olfac-
tory epithelium, sensation and perception from orthonasal or retronasal olfaction have been
reported differently. Traditionally, retronasal smell has been considered equivalent to ortho-
nasal smell, just coming from a different direction. However, the validity of such an assump-
tion has come under question. Odors presented via the retronasal route appear to evoke
 Orthonasal and Retronasal Olfaction 103

different sensations compared to orthonasal presentation (Frasnelli et al., 2008). Orthonasal

and retronasal olfaction differ in terms of odor threshold, odor intensity, ability to localize
an odor, and the neuronal processing. Differences in airflow patterns, apart from cognitive
factors, play an important role in perceptual differences between ortho- and retronasal pre-
sentations of odors (Mozell, 1964; Rebello et al., 2015; Scott et al., 2007, 2014). The direc-
tion of the odorized airstream (e.g., forward or backward) and the consequential differences
in absorption patterns of odor molecules to the olfactory epithelium across the olfactory
epithelium probably leads to differences in the processing of odorous information.

5.5.1 Differences at the Psychophysical and Peripheral Levels

A number of studies have reported psychophysical differences between ortho- and retro-
nasal olfaction. Thresholds to odor stimuli are typically lower for the orthonasal com-
pared to the retronasal route; in other words, people are more sensitive to orthonasal
than to retronasal stimuli (Duffy et al., 1999; Heilmann and Hummel, 2004; Melzner et
al., 2011; Voiol and Daget, 1986). The reason for such differences has been postulated
to be due to different stimulation patterns of the olfactory epithelium, or due to different
temporal patterns of activation (Engen, 1982).
On the suprathreshold level, Pierce and Halpern compared ortho- and retronasal
odor identification abilities (Pierce and Halpern, 1996). They found odor identification
to be significantly better when odors were presented orthonasally. In addition, intensity
ratings were higher for orthonasal than for retronasal stimuli (Heilmann and Hummel,
2004). It has also been reported that the odor identification is more efficient when stimuli
are presented orthonasally (Burdach et al., 1984). Thus, retronasal olfactory stimula-
tion appears to lead to a lower degree of activation of the olfactory system than ortho-
nasal stimulation. At the same time, measures of odor concentrations in the olfactory
cleft appeared to indicate that both absolute odor concentrations and time course of the
stimuli were similar, regardless of whether odors were presented in the back or in the
front of the nose. Intranasal air flow exhibited subtle differences in relation to stimulus
presentation. For example, numerous studies indicate that pure olfactory stimuli cannot
be lateralized when the stimulus is applied to one of the two nostrils (Kobal et al., 1989;
von Skramlik, 1925); however, subjects are able to differentiate between ortho- and ret-
ronasal presentation of an odorant (Hummel, 2008).
Differences between ortho- and retronasal olfactory functions also affect the sali-
vary response. Bender et al. observed recovery from salivary habituation (e.g., reduced
saliva secretion upon repeated exposure to a certain odor stimulus) upon presentation
of the same odor via a novel route (e.g., changed from orthonasal to retronasal or vice
versa) (Bender et al., 2009). A similar finding was reported from a recent study where
adaptation was observed to odors delivered orthonasally but not retronasally (Pierce and
Simons, 2018). This lack of cross-adaptation demonstrates that presenting an odor via
different pathways represents a distinct sensory signal (Bender et al., 2009).

5.5.2 Differentiations at the Neural Level

In humans, when recording electrical potentials at the olfactory epithelium (electro-

olfactograms, EOGs) in response to orthonasal or retronasal stimulation with phenyl-
ethyl alcohol, smaller electro-olfactogram signal amplitudes were observed for retronasal
104 Food Aroma Evolution

as compared to orthonasal stimulation, indicating that the perceptual difference starts

at the level of the olfactory epithelium (Hummel et al., 2017a). Gautam and Verhagen
(Gautam and Verhagen, 2012) showed that in the olfactory bulb, retronasally presented
odors induced not only smaller response amplitude but also longer response onset laten-
cies when compared to orthonasally presented odors in a rat model.
Electroencephalogram-derived olfactory event-related potentials (OERPs) also indi-
cated differences between orthonasal and retronasal odor perception. For example, a
prolonged OERP latency and smaller amplitude were shown when a fruity odor stimulus
was delivered retronasally as compared to orthonasally, which was in accordance to a
stronger perceived intensity through the orthonasal route (Ishii et al., 2008). In addition,
the OERP seemed to be odor- and context-dependent. When a non-food odorant (e.g.,
lavender) was presented in a contextually unusual site, that is, retronasally, the peak
amplitude of OERP was larger compared to the presentation of the same odorant at
an orthonasal site. This was the other way around for a food-related odor (e.g., choco-
late). These findings clearly indicate the contextual differences in information processing
depending on the route of odor presentation (Hummel and Heilmann, 2008).
Human brain imaging studies demonstrate similar yet different patterns of brain acti-
vation in response to orthonasal and retronasal odor stimulation. By applying odorants in
aqueous solution, previous studies have found that multiple brain regions, including the
piriform cortex, amygdala, orbitofrontal cortex (OFC), insular cortex, and hippocampus,
known as the regions activated by orthonasal stimulation, were also activated in response
to retronasal odor stimulation (Cerf-Ducastel and Murphy, 2001; de Araujo et al., 2003).
However, activation by retronasal odor stimulation was found at the base of the central
sulcus, corresponding to the primary representation of the oral cavity-related stimulation,
possibly reflecting that retronasal odors are referred to the mouth (Boling et al., 2002;
Pardo et al., 1997; Yamashita et al., 1999). When the same set of odors were perceived
through either orthonasal and retronasal routes, larger brain activation in the insula, oper-
cula, thalamus, hippocampus, amygdala, piriform, and caudolateral OFC was found in
response to orthonasal odor stimuli, whereas the pregenual cingulate, posterior cingulate,
medial OFC, and superior temporal gyrus extending into the temporal operculum show
greater activation after retronasal olfactory stimulation (Small et al., 2005). Importantly,
this difference was only found for food-related odors (Small et al., 2005).
Taken together, these findings indicate that orthonasal and retronasal olfaction rep-
resent two qualitatively distinct systems. This finding supports one of the first theories
of orthonasal versus retronasal differences coined by Rozin (1982), stating that there
are different behavioral consequences depending on the two types of information. In
short, orthonasal stimulation represents information from odor sources in the environ-
ment ranging from animals over plants to fire. Retronasal stimulation signals informa-
tion from odor sources in the oral cavity, which is generally an object that has been
previously selected as food.

5.6 THE ROLE OF ORTHO- AND RETRONASAL

OLFACTION IN FOOD PERCEPTION

For food odors, orthonasal perception represents information from the food sources in
the environment, while retronasal stimulation signals information from odor sources in
the oral cavity. An increasing body of literature describes how the perceptions of ortho-
nasal and retronasal food odor presentation routes differ.
 Orthonasal and Retronasal Olfaction 105

5.6.1 Orthonasal Food Odor Perception

A major role of orthonasal odor perception is to identify and spatially localize the food
sources. Human neonates tended to move toward a pad scented with their mother’s
breast odor in contrast to a clean control pad (Varendi and Porter, 2001), indicating the
early development of orthonasal olfaction to guide food- and nutrient-seeking behav-
ior (Adam-Darque et al., 2017; Varendi et al., 1994). In addition, orthonasal odors are
needed to identify a food’s suitability for ingestion, reflecting prior learning about the
food’s immediate and delayed consequences.
Food-related odors can be categorized according to their “taste” attributes, which
may be utilized to estimate the dominant macronutrients (e.g., carbohydrate, protein or
fat) in certain foods (Denzer-Lippmann et al., 2017). For example, sweet food odors usu-
ally emanate from food high in sugar content. In addition, through orthonasal olfaction,
humans can discriminate high concentrations of long-chain fatty acids in vapor phase
(Bolton and Halpern, 2010) and were able to discriminate between skimmed, medium,
and full fat milk samples with an overall accuracy of 40–55% correct trials in three
consecutive experiments, a value that is significantly above the expected chance level
(33.3%) (Boesveldt and Lundstrom, 2014). Thus, the orthonasal olfaction may function
as a detection system for nutrients content within natural food sources.
The presence of food odor typically triggers the “cephalic phase response” (CRR)—a
series of anticipatory physiological regulations related to feeding, including salivation
and secretion of hormones and digestive enzymes. CRS is to prepare the body for inges-
tion and digestion of specific foods or macronutrients (Smeets et al., 2010). In addition,
food odors may stimulate appetite for food consumption (Ramaekers et al., 2014; Zoon
et al., 2016). The odor-induced appetite seems to follow a sensory-specific manner; for
example, savory food odor increased the appetite for savory food, but decreased appetite
for sweet foods, and vice versa after exposure to sweet food odors. In addition, exposure
to food odors can direct choice toward the food that is signaled by the odor specifically.
For example, it has been reported that sub-threshold exposure to fruit odors prior to a
meal led participants to choose more fruit and vegetable-based foods at a subsequent meal
(Gaillet et al., 2013; Gaillet-Torrent et al., 2014; Marty et al., 2017). However, Zoon et
al. (2014) found that exposure to ambient odors that signaled high or low energy-dense
sweet and savory foods had no effect on the consumption of these foods, and this did not
vary with hunger state. In contrast to the appetizing effect, another study reported that
orthonasal exposure to dark chocolate odor suppressed hunger and reduced the ghrelin
concentration level (Massolt et al., 2010).
Taken together, orthonasal food odor can direct people toward food sources; how-
ever, the extent to which these odors are utilized to regulate appetite (including both
its stimulation and inhibition) or subsequent food choices remains unclear and requires
more research.

5.6.2 Retronasal Food Odor Perception

In real life, retronasal perception of food odors occurs when food is chewed in the mouth.
Retronasal odor perception often, if not always, interacts with signals from other sen-
sory modalities during food consumption, such as taste, somatosensation, or audition,
and is considered a fundamental factor in the formation of food flavor (Shepherd, 2006).
Retronasal olfaction is often referred to the mouth, which is augmented in the presence of
106 Food Aroma Evolution

a congruent taste (Lim and Johnson, 2011,2012). When retronasal olfaction connects to
other sensations in the mouth, for example, taste or somatosensation, the unified percept
is projected to the brain’s mouth area, eventually leading to the well-known sensory confu-
sion between taste and retronasal olfaction (Murphy and Cain, 1980; Rozin, 1982). Loss of
retronasal olfaction reduces the food-induced sensations related to gustation, texture, and
chemesthesis. As a result, the ability to identify flavors is severely limited (Running, 2018).
As compared to orthonasal odor perception, which is related to food or nutrient
anticipation, retronasal odor perception is suggested to be related to food reward (Small
et al., 2005). Possibly, retronasal odors are linked to nutrient intake and become satiat-
ing. In addition, oral exposure to odors may also affect sensory-specific satiety because
cephalic phase responses are generally stronger during oral than during odor stimula-
tion. A series of studies suggested that retronasal odor stimulation may be used to induce
satiety during consumption and ultimately may contribute to a decrease in food intake
(Ruijschop et al., 2009a). Indeed, Ruijschop et al. (2008) demonstrated that participants
felt significantly more satiated and had less desire to eat congruent (sweet) foods if they
were stimulated with a longer retronasal odor profile during (sweet) milk consumption.
However, in that study, there was no effect of retronasal odor exposure on the subsequent
milk intake, while a later study established a 9% lower food intake when participants
were exposed to a high retronasal condition versus other conditions with lower concen-
trations or shorter exposure duration (Ramaekers et al., 2013). In another study, a nega-
tive correlation was observed between the extent of retronasal aroma release and the ad
libitum food consumption (Ruijschop et al., 2009b). Collectively, these results indicated
that changing the release of retronasal aroma through modulation of concentration and
exposure time may affect perceived satiety and food intake, but perhaps not in all situa-
tions or for all participants. In addition, the composition of the odor seems to affect the
satiating effects of retronasal perception, that is, the more complex an aroma, the higher
the satiation (Ruijschop et al., 2010).

5.7 FACTORS INFLUENCING OLFACTORY PERCEPTION

Human olfactory perception differs enormously between individuals, with large reported
perceptual variations in the intensity and pleasantness of a given odor. Like in other
senses, humans vary widely in their olfactory sensitivity and quality perception. This
variability includes differences in general olfactory acuity and in the sensitivity toward
particular odorants.

5.7.1 Gender

In terms of odor detection, there is no conclusive empirical evidence for gender difference
(Doty and Cameron, 2009; Larsson et al., 2000). Nevertheless, where gender differences
exist, women are usually more sensitive to odors compared to men (Dalton et al., 2002;
Kobal et al., 2001; Koelega, 1994). Females are better at odor discrimination and identi-
fication (Cain, 1982; Ferdenzi et al., 2013), which could be related to gender differences
in verbal proficiency (Larsson et al., 2003). In neuroimaging measurement with odor
perception, females exhibited stronger responses in the frontal brain cortex, including the
OFC (Royet et al., 2003; Yousem et al., 1999), a brain structure that is involved in odor
identification (Kjelvik et al., 2012). For retronasal olfaction, women also outperformed
 Orthonasal and Retronasal Olfaction 107

men (Heilmann et al., 2002). Overall, gender differences in olfactory ability appear to be
due to aspects of olfactory processing that require higher-level cognition, such as odor
identification or odor memory.
There is also evidence suggesting gender differences in odor intensity or pleasant-
ness. For example, with a large cross-cultural sample size (n = 772), Ferdenzi et al.
(2013) showed that females rated odors more intense compared to males. In another
study, Olofsson and Nordin (2004) found that females, as compared to males, provided
higher intensity and unpleasantness ratings for olfactory/trigeminal stimuli, which was
reflected as more identifiable early components, larger amplitudes and shorter laten-
cies for the relatively exogenous late OERP component. During repeated exposure to
an odor, women showed smaller decrease in liking as compared to men (Triscoli et al.,
2014).

5.7.2 Age

The human olfactory system is well-formed during fetal development and olfactory func-
tions are already present at birth (Adam-Darque et al., 2017; Dominguez, 2011). Overall
olfactory performance increases with age and peaks at the age of 20 to 30 years, and then
declines with aging (Figure 5.3). In adults, aging often involves various transformations
of the olfactory system, which leads to a decreased olfactory function (for review see
Doty and Kamath, 2014).
The overwhelming evidence indicates a decline of odor sensitivity with aging, pro-
nouncedly so in older people (Cain et al., 1990; Griep et al., 1995; Larsson et al., 2009,
2000; Stuck et al., 2006). The decline of sensitivity toward retronasal odors has also
been described (Duffy et al., 1999; Stevens and Cain, 1986). Some authors suggested that

FIGURE 5.3 Age-related changes of the “Sniffin’ Sticks” test TDI scores in healthy sub-
jects (n = 479). With the exception of the youngest subjects, TDI scores decreased as a
function of age. (Adapted from Kobal et al., 2000.)
108 Food Aroma Evolution

aging impairs detection thresholds of different odors in a similar way (Cain and Gent,
1991; Cowart, 1989; Stevens et al., 1988). Others, however, referred to the stimulus
specificity in olfactory aging, suggesting that some flavors might fade faster than others
(Cain et al., 1990). For example, Flaherty and Lim showed decreased retronasal respon-
siveness among older participants for vanilla and soy sauce odors but not for strawberry
or chicken odors (Flaherty and Lim, 2016).
Odor discrimination and recognition, which engage more cognitive load (Hedner et
al., 2010) are developed through learning and cognitive development. Children showed
an age-related increase in their olfactory abilities (Sorokowska et al., 2015b), and they
were poorer in discriminating or recognizing odors as compared to adults, which may
result from the lesser experiences with odors, relative to adults (Stevenson et al., 2007).
For adults, significant age-related alterations are generally observed for odor identifica-
tion (Doty et al., 1984a; Evans et al., 1995; Larsson et al., 2000; Sorokowska et al.,
2015b) or discrimination (Cain et al., 1990).
Age-related changes in olfaction are also indicated by ERP measures. For adults,
the decreased olfactory performance with aging was associated with longer latencies or
smaller amplitudes for the OERP components (Evans et al., 1995; Hummel et al., 1998;
Stuck et al., 2006). Evans et al. (1995) also reported a significant life-span prolongation
of the latency for late OERP components. In addition, one fMRI study found reduced
activation in the olfactory brain areas in response to odors among older participants
(Suzuki et al., 2001). Aging is also related to decreased odor pleasantness (Joussain et al.,
2013). Taken together, the age-related changes in olfaction are observed at all levels of
information processing.

5.7.3 Genetic Factors

Individual variation of olfactory perception is largely due to genetic polymorphisms; these

include olfactory receptor genes, signal transduction genes, as well as genes involved in
propagation and processing of the olfactory input, such as those underlying the develop-
ment of olfactory epithelium and olfactory bulb (Hasin-Brumshtein et al., 2009).
Qualitatively, the genetic variations of the olfactory receptor genes explain much of
the differences in sensitivity for odors, and, consecutively, flavor experience of foods. For
example, the rs6591536 single-nucleotide polymorphism (SNP) variation of the olfactory
receptor OR5A1 explains perceptual sensitivity to the odorant β-ionone (Jaeger et al.,
2013). Several other food odors have been associated with genetic variation in odorant
receptor genes. Sensitivity to odors associated with SNPs in specific genes includes isova-
leric acid (cheesy, sweaty), β-ionone (floral), cis-3-hexen-1-ol (green, grassy), and guaiacol
(smoky) (Jaeger et al., 2013; Mainland et al., 2014; Menashe et al., 2007). Sensitivity to
other food-related odors, such as the isobutyraldehyde (malty), β-damascenone (floral),
and 2-heptanone (banana), has only been associated with regions known to encode odor
receptors (McRae et al., 2013).
Genetic variations also contribute to qualitative odor perception. The smell of the
steroid androstenone, which can be found in the skin and adipose tissue of male pigs,
varies among individuals with some experiencing no sensation, others a sweet, floral-
like sensation, and others an unpleasant sweaty, urinous sensation. Gene association
studies in humans indicate that SNPs in the OR7D4 olfactory receptor gene are at least
partially responsible for perceptual differences for the odor of androstenone (Keller et
al., 2007). The unpleasant perception (e.g., “soapiness”) for cilantro is also suggested to
 Orthonasal and Retronasal Olfaction 109

be genetically determined (Eriksson et al., 2012; Mauer, 2011). An SNP in the OR4N5
OR6A2 variation has been implicated in cilantro soapiness and disliking, as well as one
near the bitter taste receptor gene TAS2R1, may contribute to the pleasantness of cilantro
odor, as individuals homozygous for both minor alleles at these locations disliked cilan-
tro (Mauer, 2011). However, assessing the genetic variability is difficult to separate from
cultural exposure to the herb, as this varies widely among ethnic groups.
Additionally, a well-documented individual odor perceptual difference due to muta-
tions in odor receptor genes is the inability to smell a particular chemical compound,
called “specific anosmia” (Amoore, 1977). Such specific anosmias have been reported
for many food-related odors, for example, isobutyric acid (Amoore et al., 1968) or tri-
methylamine (Amoore and Forrester, 1976), among others. It is also assumed that these
idiosyncrasies are based on specific genetic differences between individuals (Croy et al.,
2015; Keller et al., 2007).

5.7.4 Oral Environment (Saliva and Microbiota)

The perception of quality and intensity for an odor are mainly linked to the chemi-
cal structure (Stevens, 1960) and the concentration of the molecule (Chastrette, 1997),
respectively. In addition, salivary proteins and microbiota have been shown to influence
the release of odor molecules in the oral cavity (Canon et al., 2018; Muñoz-González
et al., 2018).
The salivary proteins (e.g., mucins and enzymes) have been shown to interact with
aroma compounds, consequently affecting aroma concentration and retronasal percep-
tion (Buettner, 2002a,b; Friel and Taylor, 2001; Muñoz-González et al., 2018; Pagès-
Hélary et al., 2014). The addition of artificial saliva to reconstituted red wines decreased
the release of the more hydrophilic molecules but enhanced that of hydrophobic mol-
ecules (Mitropoulou et al., 2011). Compared to artificial saliva, human saliva affects the
release of ketones and esters differently (Pagès-Hélary et al., 2014), which is possibly due
to different types and amounts of protein found in natural human saliva. Furthermore,
one study showed how the individual differences in saliva composition (such as protein
content and total antioxidant capacity) could be responsible for differences in aroma
release (Muñoz-González et al., 2018). They found that centrifugation tends to reduce the
effect of saliva and its interindividual variability (Muñoz-González et al., 2018). Another
study showed that saliva from obese individuals presented a diminished aroma release
compared with normal-weight subjects, and this fact was again related to the total pro-
tein content and the total antioxidant capacity determined in saliva (Piombino et al.,
2014). Moreover, enzymes present in saliva can metabolize aroma compounds (Muñoz-
González et al., 2018) or glycosidic aroma precursors (Muñoz-González et al., 2015)
which could modify the temporal aroma perception (Muñoz-González et al., 2015).
The microbiological composition in saliva also has an impact on odor perception
(Muñoz-González et al., 2015; Piombino et al., 2014). For example, Muñoz-González et
al. found obese people to have more Firmicutes and Actinobacteria, while they had less
Proteobacteria and Fusobacteria, as compared to the normal-weight participants, which
also contributed to the suppressed aroma release from the wine sample (Piombino et al.,
2014). However, until now most of the studies have been carried out in well-controlled in
vitro situations that could not have represented the complexity and dynamics occurring at
the human mouth level. Future studies are needed with sensory evaluation by human partici-
pants in order to better understand the impact of the oral environment on odor perceptions.
110 Food Aroma Evolution

5.7.5 Hormonal Levels

The olfactory system is considered to be tightly intertwined with the endocrine system in
the regulation of chemical state and nutritional needs (Palouzier-Paulignan et al., 2012).
Olfaction is modulated by both orexigenic (appetite-stimulating) and anorexigenic (appe-
tite-suppressing) signals, of which the production and site of action are located in the
periphery (e.g., gastrointestinal tract, adipose tissue, and others) and/or in the central
nervous system (e.g., hypothalamus, reward circuits) (Soria-Gomez et al., 2014a). It has
been shown that ghrelin increases exploratory sniffing and enhances odor detection in
both humans (Tong et al., 2011) and rodents (Loch et al., 2015; Tong et al., 2011), possi-
bly by acting in the olfactory cortex. The endocannabinoids and exogenous cannabinoids
increased odor detection in fasted mice, which linking the feeling of hunger to stronger/
more effective odor processing (Soria-Gomez et al., 2014b). A lower olfactory capacity is
related to higher circulating concentrations of endocannabinoid 2-arachidonoylglycerol
and higher body mass index in women (Pastor et al., 2016). In humans, administration
of the opioid remifentanil raised olfactory thresholds while having little or no influence
on odor discrimination and odor identification performance (Lötsch et al., 2001), and
pharmacological blocking of opioid receptors was found to attenuate olfactory hedonic
responses (Yeomans and Wright, 1991). Thus, there is abundant evidence that orexigenic
factors enhance olfactory performance.
Conversely, satiety and hypophagic molecules induce a decrease in olfactory sensitiv-
ity. Two of the main hormones involved in satiety processes are leptin and insulin (Coll
et al., 2007; Morton et al., 2006). Importantly, both hormones and their receptors are
also locally expressed in olfactory circuits, mainly in the olfactory mucosa and olfac-
tory bulb. In rodents, a decreased performance in olfactory detection was observed upon
the central administration of insulin (Aimé et al., 2012) or leptin (Julliard et al., 2007),
mimicking the satiation state. In humans, reduced olfactory sensitivity was observed with
elevated insulin levels (Brünner et al., 2013; Ketterer et al., 2011), or with increased
visceral fat, which is linked to enhanced leptin concentrations (Fernandez-Garcia et al.,
2017). In addition, elevated leptinemia is reflected by low ratings of black pepper-odor
pleasantness (Trellakis et al., 2011). However, a recent study showed intranasal insulin
application significantly decreased the sensitivity to n-butanol but not to peanut odor,
and was observed only in females, which suggested odor- and gender-dependent effects
on odor thresholds (Rodriguez-Raecke et al., 2018). In addition, blood leptin concentra-
tion is correlated, in a gender-specific way, to odor identification (Karlsson et al., 2002).
Altogether, these data show that the nutritional status and its hormonal components
modulate olfactory circuits, consequently changing the olfactory behaviors related to
food consumption.

5.7.6 Olfactory Disorders

Olfactory dysfunction includes qualitative and quantitative disorders (Hong et al., 2012;
Hummel et al., 2017b). Quantitative olfactory dysfunction refers to a decreased, weaker
odor perception, and includes hyposmia (the sense of smell is partially reduced) and anosmia
(the sense of smell is lost completely or reduced to an extent that is not useful in daily life)
(Hummel et al., 2017b). Qualitative olfactory dysfunction refers to distorted perceptions of
odor quality such as parosmia (distorted odor perceptions in the presence of an odor source)
and ­phantosmia (reported odor sensation in the absence of an odor) (Frasnelli et al., 2004).
 Orthonasal and Retronasal Olfaction 111

The incidence of anosmia in the general population is about 5%, and is more frequent among
older people (Bramerson et al., 2004; Landis et al., 2004; Murphy et al., 2002; Vennemann et
al., 2008). The major causes of olfactory dysfunctions include chronic rhinosinusitis with or
without nasal polyps, acute infections of the upper respiratory tract, head trauma, and idio-
pathic olfactory loss (Temmel et al., 2002). Other causes include neurodegenerative disorders
(such as Parkinson’s disease and Alzheimer’s disease).
Olfactory loss can influence both orthonasal and retronasal olfaction. The presence of
nasal polyposis influences orthonasal but typically to a lesser degree retronasal olfactory
function (Landis et al., 2003). In addition, olfactory loss has a clear influence on food odor
perception and eating behaviors (Aschenbrenner et al., 2008). Despite the colloquial meaning
of the word “taste,” much of a food’s overall flavor actually comes from odor: specifically, ret-
ronasal olfaction. When nasal passages are inflamed or otherwise blocked, the movement of
air is restricted and results in a lack of sensation. Loss of retronasal olfaction leads to reduced
eating-related sensations, including gustation, texture, and chemesthesis. As a result, the abil-
ity to identify flavors is severely limited. For example, people with olfactory loss rated food
odors as less intense (Seo and Hummel, 2009). Olfactory loss could further result in difficul-
ties related to eating, preparing food/cooking, and reduced appetite. Interestingly, however,
these eating problems do not lead to a general pattern of reduced food intake (Aschenbrenner
et al., 2008; Temmel et al., 2002). In a study by Ferris and Duffy, 18% of the patients with
olfactory loss described an increased food consumption while 20% showed a decrease, and
the majority reported no change in food consumption (Ferris and Duffy, 1989).

5.8 CONCLUSIONS

Orthonasal and retronasal olfaction are linked but also separated, particularly in food
odor perception, suggesting a dual sensory process (Rozin, 1982). The difference not only
lies in the perceptual level, but also in the neural level. In addition, individual differences
in terms of olfactory perception are modulated by age, sex, genetic, and hormonal fac-
tors. More research is necessary to better understand the mechanism of orthonasal and
retronasal olfactory perception and their differential roles in health and diseases.

ACKNOWLEDGMENT

Special thanks go to Cornelia Hummel for preparing Figure 5.1.

REFERENCES

Ache B. W. and Young J. M. 2005. Olfaction: Diverse species, conserved principles.

Neuron 48: 417–30.
Adam-Darque A., Grouiller F., Vasung L., Ha-Vinh Leuchter R., Pollien P., Lazeyras F.,
and Huppi P. S. 2017. fMRI-based neuronal response to new odorants in the new-
born brain. Cereb Cortex 28: 1–7.
Aimé P., Hegoburu C., Jaillard T., Degletagne C., Garcia S., Messaoudi B., Thevenet
M., Lorsignol A., Duchamp C., and Mouly A.-M. 2012. A physiological increase
of insulin in the olfactory bulb decreases detection of a learned aversive odor and
abolishes food odor-induced sniffing behavior in rats. PLoS One 7: e51227.
112 Food Aroma Evolution

Amoore J. E. 1977. Specific anosmia and the concept of primary odors. Chem Senses 2:
267–81.
Amoore J. E and Forrester L. J. 1976. Specific anosmia to trimethylamine: The fishy pri-
mary odor. J Chem Ecol 2: 49–56.
Amoore J. E., Venstrom D., and Davis A. R. 1968. Measurement of specific anosmia.
Percept Mot Skills 26: 143–64.
Aschenbrenner K., Hummel C., Teszmer K., Krone F., Ishimaru T., Seo H. S., and Hummel T.
2008. The influence of olfactory loss on dietary behaviors. Laryngoscope 118: 135–44.
Bender G., Hummel T., Negoias S., and Small D. M. 2009. Separate signals for orthonasal
vs. retronasal perception of food but not nonfood odors. Behav Neurosci 123: 481–9.
Boesveldt S. and Lundstrom J. N. 2014. Detecting fat content of food from a distance:
Olfactory-based fat discrimination in humans. PLoS One 9: e85977.
Bojanowski V. and Hummel T. 2012. Retronasal perception of odors. Physiol Behav 107:
484–87.
Boling W., Reutens D. C., and Olivier A. 2002. Functional topography of the low post-
central area. J Neurosurg 97: 388–95.
Bolton B. and Halpern B. P. 2010. Orthonasal and retronasal but not oral-cavity-only
discrimination of vapor-phase fatty acids. Chem Senses 35: 229–38.
Bramerson A., Johansson L., Ek L., Nordin S., and Bende M. 2004. Prevalence of olfac-
tory dysfunction: The skovde population-based study. Laryngoscope 114: 733–7.
Brünner Y. F., Benedict C., and Freiherr J. 2013. Intranasal insulin reduces olfactory sen-
sitivity in normosmic humans. J Clin Endocrinol Metab 98: E1626–30.
Buettner A. 2002a. Influence of human saliva on odorant concentrations. 2. Aldehydes,
alcohols, 3-alkyl-2-methoxypyrazines, methoxyphenols, and 3-hydroxy-4,5-di-
methyl-2(5H)-furanone. J Agric Food Chem 50: 7105–10.
Buettner A. 2002b. Influence of human salivary enzymes on odorant concentration
changes occurring in vivo. 1. Esters and thiols. J Agric Food Chem 50: 3283–89.
Burdach K. J., Kroeze J. H., and Koster E. P. 1984. Nasal, retronasal, and gustatory per-
ception: An experimental comparison. Percept Psychophys 36: 205–8.
Cain W. S. 1982. Odor identification by males and females: Predictions vs performance.
Chem Senses 7: 129–42.
Cain W. S. and Gent J. F. 1991. Olfactory sensitivity: Reliability, generality, and associa-
tion with aging. J Exp Psychol Hum Percept Perform 17: 382–91.
Cain W. S., Reid F., and Stevens J. C. 1990. Missing ingredients: Aging and the discrimi-
nation of flavor. J Nutr Elder 9: 3–15.
Canon F., Neiers F., and Guichard E. 2018. Saliva and flavor perception: perspectives.
J Agric Food Chem 66: 7873–9.
Cerf-Ducastel B. and Murphy C. 2001. fMRI activation in response to odorants orally
delivered in aqueous solutions. Chem Senses 26: 625–37.
Chastrette M. 1997. Trends in structure-odor relationship. SAR QSAR Environ Res 6:
215–54.
Coll A. P., Farooqi I. S., and O’Rahilly S. 2007. The hormonal control of food intake.
Cell 129: 251–62.
Cowart B. J. 1989. Relationships between taste and smell across the adult life span a. Ann
N Y Acad Sci 561: 39–55.
Croy I., Hoffmann H., Philpott C., Rombaux P., Welge-Luessen A., Vodicka J.,
Konstantinidis I., Morera E., and Hummel T. 2014. Retronasal testing of olfac-
tory function: An investigation and comparison in seven countries. Eur Arch
Otorhinolaryngol 271: 1087–95.
 Orthonasal and Retronasal Olfaction 113

Croy I., Olgun S., Mueller L., Schmidt A., Muench M., Hummel C., Gisselmann G., Hatt
H., and Hummel T. 2015. Peripheral adaptive filtering in human olfaction? Three
studies on prevalence and effects of olfactory training in specific anosmia in more
than 1600 participants. Cortex 73: 180–87.
Dalton P., Doolittle N., and Breslin P. A. 2002. Gender-specific induction of enhanced
sensitivity to odors. Nat Neurosci 5: 199–200.
de Araujo I. E., Rolls E. T., Kringelbach M. L., McGlone F., and Phillips N. 2003. Taste-
olfactory convergence, and the representation of the pleasantness of flavour, in the
human brain. Eur J Neurosci 18: 2059–68.
Denzer-Lippmann M. Y., Beauchamp J., Freiherr J., Thuerauf N., Kornhuber J., and
Buettner A. 2017. Development and validation of a Food-Associated Olfactory Test
(FAOT). Chem Senses 42: 47–57.
Dominguez P. R. 2011. The study of postnatal and later development of the taste and
olfactory systems using the human brain mapping approach: An update. Brain Res
Bull 84: 118–24.
Doty R. L. 1997. Studies of human olfaction from the University of Pennsylvania Smell
and Taste Center. Chem Senses 22: 565–86.
Doty R. L. and Cameron E. L. 2009. Sex differences and reproductive hormone influ-
ences on human odor perception. Physiol Behav 97: 213–28.
Doty R. L. and Kamath V. 2014. The influences of age on olfaction: A review. Front
Psychol 5: 20.
Doty R.L., Shaman P., Applebaum S. L., Giberson R., Siksorski L., and Rosenberg L.
1984a. Smell identification ability: Changes with age. Science 226: 1441–3.
Doty R. L., Shaman P., and Dann M. 1984b. Development of the University of
Pennsylvania Smell Identification Test: A standardized microencapsulated test of
olfactory function. Physiol Behav 32: 489–502.
Duffy V. B., Cain W. S., and Ferris A. M. 1999. Measurement of sensitivity to olfactory
flavor: Application in a study of aging and dentures. Chem Senses 24: 671–7.
Engen T. 1982. The Perception of Odors. Toronto, ON: Academic Press.
Eriksson N., Wu S., Do C. B., Kiefer A. K., Tung J. Y., Mountain J. L., Hinds D. A., and
Francke U. 2012. A genetic variant near olfactory receptor genes influences cilantro
preference. Flavour 1: 22.
Evans W. J., Cui L., and Starr A. 1995. Olfactory event-related potentials in normal
human subjects: Effects of age and gender. Electroencephalogr Clin Neurophysiol
95: 293–301.
Ferdenzi C., Roberts S. C., Schirmer A., Delplanque S., Cekic S., Porcherot C., Cayeux
I., Sander D., and Grandjean D. 2013. Variability of affective responses to odors:
Culture, gender, and olfactory knowledge. Chem Senses 38: 175–86.
Fernandez-Garcia J. C., Alcaide J., Santiago-Fernandez C., Roca-Rodriguez M. M.,
Aguera Z., Banos R., Botella C., de la Torre R., Fernandez-Real J. M., Fruhbeck G.,
Gomez-Ambrosi J., Jimenez-Murcia S., Menchon J. M., Casanueva F. F., Fernandez-
Aranda F., Tinahones F. J., and Garrido-Sanchez L. 2017. An increase in visceral
fat is associated with a decrease in the taste and olfactory capacity. PLoS One 12:
e0171204.
Ferris A. M. and Duffy V. B. 1989. Effect of olfactory deficits on nutritional status: Does
age predict persons at risk? Ann N Y Acad Sci 561: 113–23.
Firestein S. 2001. How the olfactory system makes sense of scents. Nature 413:
211–18.
114 Food Aroma Evolution

Flaherty T. J. and Lim J. 2016. Individual differences in retronasal odor responsiveness:

Effects of aging and concurrent taste. Chemosens Percept 10: 91–103.
Frasnelli J., Landis B. N., Heilmann S., Hauswald B., Hüttenbrink K. B., Lacroix J. S.,
Leopold D. A., and Hummel T. 2004. Clinical presentation of qualitative olfactory
dysfunction. Eur Arch Otorhinolaryngol 261: 411–5.
Frasnelli J., Ungermann M., and Hummel T. 2008. Ortho- and retronasal presentation of
olfactory stimuli modulates odor percepts. Chemosens Percept 1: 9–15.
Friel E. N. and Taylor A. J. 2001. Effect of salivary components on volatile partitioning
from solutions. J Agric Food Chem 49: 3898–905.
Gaillet M., Sulmont-Rossé C., Issanchou S., Chabanet C., and Chambaron S. 2013.
Priming effects of an olfactory food cue on subsequent food-related behaviour. Food
Qual Prefer 30: 274–81.
Gaillet-Torrent M., Sulmont-Rossé C., Issanchou S., Chabanet C., and Chambaron S.
2014. Impact of a non-attentively perceived odour on subsequent food choices.
Appetite 76: 17–22.
Gautam S. H. and Verhagen J. V. 2012. Retronasal odor representations in the dorsal
olfactory bulb of rats. J Neurosci 32: 7949–59.
Gottfried J. A. 2010. Central mechanisms of odour object perception. Nat Rev Neurosci
11: 628–41.
Griep M. I., Mets T. F., Vercruysse A., Cromphout I., Ponjaert I., Toft J., and Massart
D. L. 1995. Food odor thresholds in relation to age, nutritional, and health status. J
Gerontol A Biol Sci Med Sci 50: B407–14.
Haehner A., Mayer A. M., Landis B. N., Pournaras I., Lill K., Gudziol V., and Hummel
T. 2009. High test–retest reliability of the extended version of the “Sniffin’ Sticks”
test. Chem Senses 34: 705–11.
Hasin-Brumshtein Y., Lancet D., and Olender T. 2009. Human olfaction: From genomic
variation to phenotypic diversity. Trends Genet 25: 178–84.
Hedner M., Larsson M., Arnold N., Zucco G. M., and Hummel T. 2010. Cognitive fac-
tors in odor detection, odor discrimination, and odor identification tasks. J Clin
Exp Neuropsychol 32: 1062–7.
Heilmann S. and Hummel T. 2004. A new method for comparing orthonasal and retro-
nasal olfaction. Behav Neurosci 118: 412–9.
Heilmann S., Strehle G., Rosenheim K., Damm M., and Hummel T. 2002. Clinical
assessment of retronasal olfactory function. Arch Otolaryngol Head Neck Surg
128: 414–8.
Hong S. C., Holbrook E. H., Leopold D. A., and Hummel T. 2012. Distorted olfactory
perception: A systematic review. Acta Otolaryngol 132 Suppl 1: S27–31.
Hugh S. C., Siu J., Hummel T., Forte V., Campisi P., Papsin B. C., and Propst E. J. 2015.
Olfactory testing in children using objective tools: Comparison of Sniffin’Sticks and
University of Pennsylvania Smell Identification Test (UPSIT). J Otolaryngol Head
Neck Surg 44: 10.
Hummel T. 2008. Retronasal perception of odors. Chem Biodivers 5: 853–61.
Hummel T., Barz S., Pauli E., and Kobal G. 1998. Chemosensory event-related potentials
change with age. Electroencephalogr Clin Neurophysiol 108: 208–17.
Hummel T. and Heilmann S. 2008. Olfactory event-related potentials in response to
ortho- and retronasal stimulation with odors related or unrelated to foods. Int
Dairy J 18: 874–8.
 Orthonasal and Retronasal Olfaction 115

Hummel T., Kobal G., Gudziol H., and Mackay-Sim A. 2007. Normative data for the
“Sniffin’ Sticks” including tests of odor identification, odor discrimination, and
olfactory thresholds: An upgrade based on a group of more than 3,000 subjects.
Eur Arch Otorhinolaryngol 264: 237–43.
Hummel T., Sekinger B., Wolf S. R., Pauli E., and Kobal G. 1997. “Sniffin’ Sticks”:
Olfactory performance assessed by the combined testing of odor identification, odor
discrimination and olfactory threshold. Chem Senses 22: 39–52.
Hummel T., Seo H. S., Pellegrino R., and Heilmann S. 2017a. Electro-olfactograms in
humans in response to ortho-and retronasal chemosensory stimulation. Chemosens
Percept 10: 114–8.
Hummel T., Whitcroft K. L., Andrews P., Altundag A., Cinghi C., Costanzo R. M.,
Damm M., Frasnelli J., Gudziol H., Gupta N., Haehner A., Holbrook E., Hong S.
C., Hornung D., Hüttenbrink K. B., Kamel R., Kobayashi M., Konstantinidis I.,
Landis B. N., Leopold D. A., Macchi A., Miwa T., Moesges R., Mullol J., Mueller
C. A., Ottaviano G., Passali G. C., Philpott C., Pinto J. M., Ramakrishnan V. J.,
Rombaux P., Roth Y., Schlosser R. A., Shu B., Soler G., Stjarne P., Stuck B. A.,
Vodicka J., and Welge-Luessen A. 2017b. Position paper on olfactory dysfunction.
Rhinology doi:10.4193/Rhin16.248.
Ishii A., Roudnitzky N., Beno N., Bensafi M., Hummel T., Rouby C., and Thomas-
Danguin T. 2008. Synergy and masking in odor mixtures: An electrophysiological
study of orthonasal vs. retronasal perception. Chem Senses 33: 553–61.
Jaeger S. R., McRae J. F., Bava C. M., Beresford M. K., Hunter D., Jia Y., Chheang S. L.,
Jin D., Peng M., Gamble J. C., Atkinson K. R., Axten L. G., Paisley A. G., Tooman
L., Pineau B., Rouse S. A., and Newcomb R. D. 2013. A Mendelian trait for olfac-
tory sensitivity affects odor experience and food selection. Curr Biol 23: 1601–5.
Joussain P., Thevenet M., Rouby C., and Bensafi M. 2013. Effect of aging on hedonic
appreciation of pleasant and unpleasant odors. PLoS One 8: e61376.
Julliard A. K., Chaput M. A., Apelbaum A., Aimé P., Mahfouz M., and Duchamp-Viret
P. 2007. Changes in rat olfactory detection performance induced by orexin and
leptin mimicking fasting and satiation. Behav Brain Res 183: 123–9.
Karlsson A., Lindroos A., Lissner L., Torgerson J., Carlsson B., Carlsson L., and Sjöström
L. 2002. Evidence for gender-specific associations between leptin and olfaction. J
Gender-Specific Med 5: 25–32.
Keller A., Zhuang H., Chi Q., Vosshall L. B., and Matsunami H. 2007. Genetic variation
in a human odorant receptor alters odour perception. Nature 449: 468–72.
Ketterer C., Heni M., Thamer C., Herzberg-Schafer S. A., Haring H. U., and Fritsche A.
2011. Acute, short-term hyperinsulinemia increases olfactory threshold in healthy
subjects. Int J Obes (Lond) 35: 1135–8.
Kjelvik G., Evensmoen H. R., Brezova V., and Haberg A. K. 2012. The human brain
representation of odor identification. J Neurophysiol 108: 645–57.
Kobal G. 1981. Elektrophysiologische untersuchungen des menschlichen geruchssinns.
Stuttgart: Thieme Verlag.
Kobal G., Hummel T., Sekinger B., Barz S., Roscher S., and Wolf S. 1996. “Sniffin’sticks”:
Screening of olfactory performance. Rhinology 34: 222–6.
Kobal G., Klimek L., Wolfensberger M., Gudziol H., Temmel A., Owen C. M., Seeber
H., Pauli E., and Hummel T. 2000. Multicenter investigation of 1,036 subjects
using a standardized method for the assessment of olfactory function combining
tests of odor identification, odor discrimination, and olfactory thresholds. Eur Arch
Otorhinolaryngol 257: 205–11.
116 Food Aroma Evolution

Kobal G., Palisch K., Wolf S. R., Meyer E. D., Hüttenbrink K. B., Roscher S., Wagner
R., and Hummel T. 2001. A threshold-like measure for the assessment of olfactory
sensitivity: The “random” procedure. Eur Arch Otorhinolaryngol 258: 168–72.
Kobal G., Van Toller S., and Hummel T. 1989. Is there directional smelling? Experientia
45: 130–2.
Koelega H. S. 1994. Sex differences in olfactory sensitivity and the problem of the gener-
ality of smell acuity. Percept Mot Skills 78: 203–13.
Landis B. N., Giger R., Ricchetti A., Leuchter I., Hugentobler M., Hummel T., and
Lacroix J. S. 2003. Retronasal olfactory function in nasal polyposis. Laryngoscope
113: 1993–7.
Landis B. N., Konnerth C. G., and Hummel T. 2004. A study on the frequency of olfac-
tory dysfunction. Laryngoscope 114: 1764–9.
Larsson M., Farde L., Hummel T., Witt M., Lindroth N. E., and Backman L. 2009. Age-
related loss of olfactory sensitivity: Association to dopamine transporter binding in
putamen. Neuroscience 161: 422–6.
Larsson M., Finkel D., and Pedersen N. L. 2000. Odor identification: Influences of age,
gender, cognition, and personality. J Gerontol B Psychol Sci Soc Sci 55: P304–10.
Larsson M., Lovden M., and Nilsson L. G. 2003. Sex differences in recollective experi-
ence for olfactory and verbal information. Acta Psychol (Amst) 112: 89–103.
Lim J. and Johnson M. B. 2011. Potential mechanisms of retronasal odor referral to the
mouth. Chem Senses 36: 283–9.
Lim J. and Johnson M. B. 2012. The role of congruency in retronasal odor referral to the
mouth. Chem Senses 37: 515–22.
Loch D., Breer H., and Strotmann J. 2015. Endocrine modulation of olfactory respon-
siveness: Effects of the orexigenic hormone ghrelin. Chem Senses 40: 469–79.
Lötsch J., Darimont J., Skarke C., Zimmermann M., Hummel T., and Geisslinger G.
2001. Effects of the opioid remifentanil on olfactory function in healthy volunteers.
Life Sci 69: 2279–85.
Mainland J. D., Keller A., Li Y. R., Zhou T., Trimmer C., Snyder L. L., Moberly A. H.,
Adipietro K. A., Liu W. L., Zhuang H., Zhan S., Lee S. S., Lin A., and Matsunami
H. 2014. The missense of smell: Functional variability in the human odorant recep-
tor repertoire. Nat Neurosci 17: 114–20.
Marty L., Bentivegna H., Nicklaus S., Monnery-Patris S., and Chambaron S. 2017. Non-
conscious effect of food odors on children’s food choices varies by weight status.
Front Nutr 4: 16.
Massolt E. T., van Haard P. M., Rehfeld J. F., Posthuma E. F., van der Veer E., and
Schweitzer DH. 2010. Appetite suppression through smelling of dark chocolate cor-
relates with changes in ghrelin in young women. Regul Pept 161: 81–6.
Mauer L. 2011. Genetic Determinants of Cilantro Preference. University of Toronto.
McGann J. P. 2017. Poor human olfaction is a 19th-century myth. Science 356: 597+.
McRae J. F., Jaeger S. R., Bava C. M., Beresford M. K., Hunter D., Jia Y., Chheang S. L.,
Jin D., Peng M., Gamble J. C., Atkinson K. R., Axten L. G., Paisley A. G., Williams
L., Tooman L., Pineau B., Rouse S. A., and Newcomb R. D. 2013. Identification of
regions associated with variation in sensitivity to food-related odors in the human
genome. Curr Biol 23: 1596–600.
Melzner J., Bitter T., Guntinas-Lichius O., Gottschall R., Walther M., and Gudziol H.
2011. Comparison of the orthonasal and retronasal detection thresholds for carbon
dioxide in humans. Chem Senses 36: 435–41.
 Orthonasal and Retronasal Olfaction 117

Menashe I., Abaffy T., Hasin Y., Goshen S., Yahalom V,. Luetje C. W., and Lancet D.
2007. Genetic elucidation of human hyperosmia to isovaleric acid. PLoS Biol 5: e284.
Menashe I. and Lancet D. 2006. Variations in the human olfactory receptor pathway.
Cell Mol Life Sci 63: 1485–93.
Mitropoulou A., Hatzidimitriou E., and Paraskevopoulou A. 2011. Aroma release of
a model wine solution as influenced by the presence of non-volatile components.
Effect of commercial tannin extracts, polysaccharides and artificial saliva. Food Res
Int 44: 1561–70.
Morton G. J., Cummings D. E., Baskin D. G., Barsh G. S., and Schwartz M. W. 2006.
Central nervous system control of food intake and body weight. Nature 443: 289–95.
Mozell M. M. 1964. Evidence for sorption as a mechanism of the olfactory analysis of
vapours. Nature 203: 1181.
Muñoz-González C., Cueva C., Pozo-Bayón M. A., and Moreno-Arribas M. V. 2015.
Ability of human oral microbiota to produce wine odorant aglycones from odour-
less grape glycosidic aroma precursors. Food Chem 187: 112–9.
Muñoz-González C., Feron G., Brule M., and Canon F. 2018. Understanding the release
and metabolism of aroma compounds using micro-volume saliva samples by ex vivo
approaches. Food Chem 240: 275–85.
Muñoz-González C., Feron G., and Canon F. 2018. Main effects of human saliva on
flavour perception and the potential contribution to food consumption. Proc Nutr
Soc 77: 1–9.
Murphy C. and Cain W. S. 1980. Taste and olfaction: Idependence vs interaction. Physio
Behav 24: 601–5.
Murphy C., Schubert C. R., Cruickshanks K. J., Klein B. E., Klein R., and Nondahl D.
M. 2002. Prevalence of olfactory impairment in older adults. JAMA 288: 2307–12.
Olofsson J. K. and Nordin S. 2004. Gender differences in chemosensory perception and
event-related potentials. Chem Senses 29: 629–37.
Pagès-Hélary S., Andriot I., Guichard E., and Canon F. 2014. Retention effect of human
saliva on aroma release and respective contribution of salivary mucin and α-amylase.
Food Res Int 64: 424–31.
Palouzier-Paulignan B., Lacroix M. C., Aimé P., Baly C., Caillol M., Congar P., Julliard
A. K., Tucker K., and Fadool D. A. 2012. Olfaction under metabolic influences.
Chem Senses 37: 769–97.
Pardo J. V., Wood T. D., Costello P. A., Pardo P. J., and Lee J. T. 1997. PET study of the
localization and laterality of lingual somatosensory processing in humans. Neurosci
Lett 234: 23–6.
Pastor A., Fernandez-Aranda F., Fito M., Jimenez-Murcia S., Botella C., Fernandez-
Real J. M., Fruhbeck G., Tinahones F. J., Fagundo A. B., Rodriguez J., Aguera Z.,
Langohr K., Casanueva F. F., and de la Torre R. 2016. A lower olfactory capacity
is related to higher circulating concentrations of endocannabinoid 2-arachidonoylg-
lycerol and higher body mass index in women. PLoS One 11: e0148734.
Philpott C M., Bennett A., and Murty G. E. 2008. A brief history of olfaction and olfac-
tometry. J Laryngol Otol 122: 657–62.
Pierce A. M. and Simons C. .T. 2018. Olfactory adaptation is dependent on route of deliv-
ery. Chem Senses 43: 197–203.
Pierce J. and Halpern B. P. 1996. Orthonasal and retronasal odorant identification based
upon vapor phase input from common substances. Chem Senses 21: 529–43.
118 Food Aroma Evolution

Piombino P., Genovese A., Esposito S., Moio L., Cutolo P. P., Chambery A., Severino V.,
Moneta E., Smith D. P., Owens S. M., Gilbert J. A., and Ercolini D. 2014. Saliva
from obese individuals suppresses the release of aroma compounds from wine. PLoS
One 9: e85611.
Ramaekers M. G., Boesveldt S., Lakemond C. M., van Boekel M. A., and Luning P. A.
2014. Odors: Appetizing or satiating? Development of appetite during odor expo-
sure over time. Int J Obes (Lond) 38: 650–6.
Ramaekers M. G., Luning P. A., Ruijschop R. M., Lakemond C. M., Bult J. H., Gort G.,
and van Boekel M. A. 2013. Aroma exposure time and aroma concentration in rela-
tion to satiation. Br J Nutr 111: 1–9.
Rebello M. R., Kandukuru P., and Verhagen J. V. 2015. Direct behavioral and neuro-
physiological evidence for retronasal olfaction in mice. PLoS One 10: e0117218.
Renner B., Mueller C. A., Dreier J., Faulhaber S., Rascher W., and Kobal G. 2009. The
candy smell test: A new test for retronasal olfactory performance. Laryngoscope
119: 487–95.
Rodriguez-Raecke R., Brünner Y. F., Kofoet A., Mutic S., Benedict C., and Freiherr J.
2018. Odor sensitivity after intranasal insulin application is modulated by gender.
Front Endocrinol 9: 580.
Royet J. P., Plailly J., Delon-Martin C., Kareken D. A., and Segebarth C. 2003. fMRI of
emotional responses to odors: Influence of hedonic valence and judgment, handed-
ness, and gender. Neuroimage 20: 713–28.
Rozin P. 1982. “Taste-smell confusions” and the duality of the olfactory sense. Atten
Percept Psychophys 31: 397–401.
Ruijschop R. M., Boelrijk A. E., Burgering M. J., de Graaf C., and Westerterp-Plantenga
M. S. 2010. Acute effects of complexity in aroma composition on satiation and food
intake. Chem Senses 35: 91–100.
Ruijschop R. M., Boelrijk A. E., de Graaf C., and Westerterp-Plantenga M. S. 2009a.
Retronasal aroma release and satiation: A review. J Agric Food Chem 57: 9888–94.
Ruijschop R. M., Boelrijk A. E., de Ru J. A., de Graaf C., and Westerterp-Plantenga M.
S. 2008. Effects of retro-nasal aroma release on satiation. Br J Nutr 99: 1140–8.
Ruijschop R. M., Burgering M. J., Jacobs M. A., and Boelrijk A. E. 2009b. Retro-nasal
aroma release depends on both subject and product differences: A link to food
intake regulation? Chem Senses 34: 395–403.
Running C. A. 2018. Oral sensations and secretions. Physiol Behav 193: 234–7.
Schriever V. A., Mori E., Petters W., Boerner C., Smitka M., and Hummel T. 2014. The
“Sniffin’ Kids” test-a 14-item odor identification test for children. PLoS One 9:
e101086.
Scott J. W., Acevedo H. P., Sherrill L., and Phan M. 2007. Responses of the rat olfactory
epithelium to retronasal air flow. J Neurophysiol 97: 1941–50.
Scott J W., Sherrill L., Jiang J., and Zhao K. 2014. Tuning to odor solubility and sorption
pattern in olfactory epithelial responses. J Neurosci 34: 2025–36.
Seo H. S. and Hummel T. 2009. Effects of olfactory dysfunction on sensory evaluation
and preparation of foods. Appetite 53: 314–21.
Shepherd G. M. 2004. The human sense of smell: Are we better than we think? PLoS
Biol 2: E146.
Shepherd G. M. 2006. Smell images and the flavour system in the human brain. Nature
444: 316–21.
 Orthonasal and Retronasal Olfaction 119

Small D. M., Gerber J. C., Mak Y. E., and Hummel T. 2005. Differential neural responses
evoked by orthonasal versus retronasal odorant perception in humans. Neuron 47:
593–605.
Smeets P. A., Erkner A., and de Graaf C. 2010. Cephalic phase responses and appetite.
Nutr Rev 68: 643–55.
Soria-Gomez E., Bellocchio L., and Marsicano G. 2014a. New insights on food intake
control by olfactory processes: The emerging role of the endocannabinoid system.
Mol Cell Endocrinol 397: 59–66.
Soria-Gomez E., Bellocchio L., Reguero L., Lepousez G., Martin C., Bendahmane M.,
Ruehle S., Remmers F., Desprez T., Matias I., Wiesner T., Cannich A., Nissant A.,
Wadleigh A/, Pape H. C., Chiarlone A. P., Quarta C., Verrier D., Vincent P., Massa
F., Lutz B., Guzman M., Gurden H., Ferreira G., Lledo P. M., Grandes P., and
Marsicano G. 2014b. The endocannabinoid system controls food intake via olfac-
tory processes. Nat Neurosci 17: 407–15.
Sorokowska A., Albrecht E., Haehner A., and Hummel T. 2015a. Extended version of the
“Sniffin’ Sticks” identification test: Test–retest reliability and validity. J Neurosci
Methods 243: 111–4.
Sorokowska A., Schriever V. A., Gudziol V., Hummel C., Hahner A., Iannilli E., Sinding
C., Aziz M., Seo H. S., Negoias S., and Hummel T. 2015b. Changes of olfactory
abilities in relation to age: Odor identification in more than 1400 people aged 4 to
80 years. Eur Arch Otorhinolaryngol 272: 1937–44.
Spence C. 2013. Multisensory flavour perception. Curr Biol 23: R365–9.
Stevens J. C. and Cain W. S. 1986. Smelling via the mouth: Effect of aging. Percept
Psychophys 40: 142–6.
Stevens J. C., Cain W. S., and Burke R. J. 1988. Variability of olfactory thresholds. Chem
Senses 13: 643–53.
Stevens S. S. 1960. The psychophysics of sensory function. Am Scientist 48: 226–53.
Stevenson R. J. 2010. An initial evaluation of the functions of human olfaction. Chem
Senses 35: 3–20.
Stevenson R. J., Mahmut M., and Sundqvist N. 2007. Age-related changes in odor dis-
crimination. Dev Psychol 43: 253–60.
Stuck B. A., Frey S., Freiburg C,. Hormann K., Zahnert T., and Hummel T. 2006.
Chemosensory event-related potentials in relation to side of stimulation, age, sex,
and stimulus concentration. Clin Neurophysiol 117: 1367–75.
Sun B. C. and Halpern B. P. 2005. Identification of air phase retronasal and orthonasal
odorant pairs. Chem Senses 30: 693–706.
Suzuki Y., Critchley H. D., Suckling J., Fukuda R., Williams S. C., Andrew C., Howard
R., Ouldred E., Bryant C., Swift C. G., and Jackson S. H. 2001. Functional mag-
netic resonance imaging of odor identification: The effect of aging. J Gerontol A
Biol Sci Med Sci 56: M756–60.
Temmel A. F., Quint C., Schickinger-Fischer B., Klimek L., Stoller E., and Hummel T.
2002. Characteristics of olfactory disorders in relation to major causes of olfactory
loss. Arch Otolaryngol Head Neck Surg 128: 635–41.
Tong J., Mannea E, Aimé P., Pfluger P. T., Yi C. X., Castaneda T. R., Davis H. W.,
Ren X., Pixley S., Benoit S., Julliard K., Woods S. C., Horvath T. L., Sleeman M.
M., D’Alessio D., Obici S., Frank R., and Tschop M. H. 2011. Ghrelin enhances
olfactory sensitivity and exploratory sniffing in rodents and humans. J Neurosci 31:
5841–6.
120 Food Aroma Evolution

Trellakis S., Tagay S., Fischer C., Rydleuskaya A., Scherag A., Bruderek K., Schlegl S.,
Greve J., Canbay A. E., Lang S., and Brandau S. 2011. Ghrelin, leptin and adipo-
nectin as possible predictors of the hedonic value of odors. Regul Pept 167: 112–7.
Triscoli C., Croy I., Olausson H., and Sailer U. 2014. Liking and wanting pleasant odors:
Different effects of repetitive exposure in men and women. Front Psychol 5: 526.
Varendi H. and Porter R. H. 2001. Breast odour as the only maternal stimulus elicits
crawling towards the odour source. Acta Paediatr 90: 372–5.
Varendi H., Porter R. H., and Winberg J. 1994. Does the newborn baby find the nipple
by smell? Lancet 344: 989–90.
Vennemann M. M., Hummel T., and Berger K. 2008. The association between smoking
and smell and taste impairment in the general population. J Neurol 255: 1121–6.
Voiol E. and Daget N. 1986. Comparative study of nasal and retronasal olfactory percep-
tion. Lebens Wiss Technol 19: 316–9.
von Skramlik E. 1925. Über die Lokalisation der Empfindungen bei den niederen Sinnen.
Zeitschr Sinnesphysiol 56: 69.
Yamashita H., Kumamoto Y., Nakashima T., Yamamoto T., Inokuchi A., and Komiyama
S. 1999. Magnetic sensory cortical responses evoked by tactile stimulations
of the human face, oral cavity and flap reconstructions of the tongue. Eur Arch
Otorhinolaryngol 256 Suppl 1: S42–6.
Yeomans M. R. and Wright P. 1991. Lower pleasantness of palatable foods in nalmefene-
treated human volunteers. Appetite 16: 249–59.
Yousem D. M., Maldjian J. A., Siddiqi F., Hummel T., Alsop D. C., Geckle R. J., Bilker
W. B., and Doty R. L. 1999. Gender effects on odor-stimulated functional magnetic
resonance imaging. Brain Res 818: 480–7.
Zoon H. F., de Graaf C., and Boesveldt S. 2016. Food odours direct specific appetite.
Foods 5: 12.
Zoon H. F., He W., de Wijk R. A., de Graaf C., and Boesveldt S. 2014. Food preference
and intake in response to ambient odours in overweight and normal-weight females.
Physiol Behav 133: 190–6.
 Section II
Analytical Techniques
 Chapter 6
Extraction Methods of Volatile
Compounds from Food Matrices
Arthur Luiz Baião Dias, Francisco Manuel
Barrales, and Philipe dos Santos

CONTENTS

6.1 Introduction 123

6.2 Extraction Methods for Liquid Samples 124
6.2.1 Liquid–Liquid Extraction 124
6.2.2 Liquid–Solid Extraction 125
6.2.3 Solid-Phase Extraction 125
6.3 Extraction Methods for Liquid or Solid Samples 126
6.3.1 Static Headspace Extraction 126
6.3.2 Dynamic Headspace Extraction (Purge and Trap) 127
6.3.3 Solid-Phase Microextraction 129
6.4 Extraction Methods for Solid Samples 129
6.4.1 Conventional Methods 130
6.4.2 Ultrasound-Assisted Extraction 130
6.4.3 Microwave-Assisted Extraction 132
6.4.4 Supercritical and Pressurized Fluid Extraction 133
References 134

6.1 INTRODUCTION

Volatile compound fractions in food matrices are associated with the aromas and flavors
that are constructed during the preparation, cooking, and consumption of the food. A
multifaceted series of factors can affect the perception, quantification, and analysis of
the food’s flavor. Generally, the analysis involves the identification and quantification of
an analyte or a group of analytes. According to Sides, Robards, and Helliwell (2000),
odor analysis involves the determination of a physiological concept via physicochemical
measurements of a complex system aimed to the detection of a wide range of compounds
and this introduces some unique analytical constraints. Generally, the food flavor is
composed of volatile organic compounds (VOCs), as well as their derivatives, and other
nonvolatile compounds. Several factors can complicate the analysis of the food’s flavor.
First, a recurrent problem is identifying the compounds contributing to the food aroma,
which is generally represented by many classes of compounds and may be present in
low or trace concentrations (Sides, Robards, and Helliwell, 2000). Another group of

123
124 Food Aroma Evolution

factors that can influence an analysis are the differences in the physical proprieties of
those compounds, since each class of compound or each compound has specific thermo-
dynamic proprieties and can interact with the matrix and solvents in many ways, which
affects the extraction and separation. However, the use of an adequate technique can
facilitate the separation of a specific compound or a group. Thus, the objective of this
chapter is to provide an overview of the different methodologies to extract, isolate, and
purify volatile compounds from food matrices. The methods are arranged according to
the sample’s physical state.

6.2 EXTRACTION METHODS FOR LIQUID SAMPLES

6.2.1 Liquid–Liquid Extraction

Liquid–liquid extraction (LLE) is based on the relative solubility of two different immis-
cible liquids, usually a polar and a nonpolar solvent; therefore, this technique has also
been referred to as immiscible solvent extraction. Usually, one phase is aqueous, and the
other is an organic solvent, where the analyte is transferred from one phase to the other
liquid immiscible phase, due to the chemical potential of the solutes. The immiscibility
of the solvents allows for an easy separation of the phases in a separatory funnel, where
the lower layer can be collected through the bottom, and the upper layer can be removed
through the top, since the position of each phase in the funnel depends on the densities
of the liquids (Wells, 2003). The LLE method has disadvantages compared to other food
flavor extraction methods, such as the exposure to large volumes of organic solvents and
the formation of emulsions. Moreover, this method has a limitation when extracting and
isolating flavor compounds from food matrices containing a significant content of lipids,
because this method also extracts lipids. Therefore, in these cases, further purification
steps are required for the separation and isolation of the flavor compounds. Despite the
simplicity of the method, there is a tendency to replace LLE with other techniques, due
to, on the one hand, trace analysis requiring expensive high-purity solvents, and on
the other hand the elevated use of environmentally damaging and unhealthy solvents
associated with this technique (Augusto, Lopes, and Zini, 2003; Wells, 2003). To over-
come these drawbacks, other sample preparation techniques have been developed and
implemented; for example, solid-phase extraction (SPE) and solid-phase microextrac-
tion (SPME). Meanwhile, there are several papers that have used this technique for food
aroma analysis, such as for wine and grape varieties (López and Gómez, 2000; Rocha
et al., 2000; Mestres, Busto, and Guasch, 2000), and for the isolation of volatile com-
pounds in fruits and vegetables, for example cherry tomatoes (Selli et al., 2014), and
oranges (Kelebek and Selli, 2011), and for the extraction of rose water (Canbay, 2017).
The wide applications of this technique is evidence of its success for food aroma analy-
ses; however, according to Varlet, Prost, and Serot (2007), LLE is not recommended for
recovering the volatile aldehydes in smoked fish. Moreover, according to Marsili (2016),
researchers identified 80 neutral volatiles in raw milk from different species using vac-
uum distillation and liquid–liquid extraction followed by high-resolution gas chroma-
tography (HRGC). Also, despite the significant number of volatiles identified, the results
of their work did not identify which compounds were the most important to good milk
flavor. Therefore, this method has some restrictions and limitations. However, it is an
excellent preliminary test for research of food flavoring compounds due to the simplicity
and low cost.
 Extraction Methods of Volatile Compounds  125

6.2.2 Liquid–Solid Extraction

Liquid–solid extractions (LSEs) were used to concentrate semi-volatile compounds from liq-
uids into a solid. In summary, the liquid sample is placed in contact with the bulk solid
extracting phase, and after a determined period an equilibrium is established between the two
phases. Next, the physical separation of the solid and liquid phases is performed, by decant-
ing or filtering, and finally the extraction from the solid is conducted with a suitable solvent
to isolate the volatile compounds. Different modifications to the LSE of volatile organic com-
pounds have developed several extraction techniques; for example, solid-phase extraction,
solid-phase microextraction (SPME), and stir bar sorptive extraction (SBSE) (Wells, 2003).

6.2.3 Solid-Phase Extraction

Solid-phase extraction was a technique developed in the mid-1970s and was introduced
in the market in the 1980s (Panighel and Flamini, 2015). This methodology of sample
preparation is based on the sorption of the analyte onto a cartridge and its recovery by
elution using a suitable solvent, resulting in a concentration and purification/isolation of
the target compound or a class of compounds. The capability of the cartridge to extract
some target compound depends on the bed sorbent, sample volume loaded, and the char-
acteristics and volume of the solvents and eluents used in the analysis. The performance
of the method can also be affected by the breakthrough volume, which is defined as
the maximum volume of sample that can be introduced into the sorbent (Panighel and
Flamini, 2015; Wells, 2003). The SPE technique is composed of four stages: (i) column
preparation; (ii) sample loading; (iii) column post-wash; and (iv) sample desorption, as
shown in Figure 6.1. However, recent advances in sorbent technology removed the column
preparation step. The objective of the pre-wash or column preparation and post-wash

FIGURE 6.1 Four basic steps for solid-phase extraction: (1) Conditioning the sorbent prior to
sample application ensures reproducible retention of the compound of interest; (2) Retention
of the adsorbed isolate, undesired matrix, and other undesired matrix compounds; (3) Rinse
the column(s) to remove undesired components; (4) Elution of desired components remain.
126 Food Aroma Evolution

stage is a condition of the stationary phase and removal of undesirable contaminants,

respectively. Generally, the target compounds are retained on the sorbent, the interfer-
ers go through the stationary phase, and an elution solvent with an appropriate solvent
recovers the adsorbed analytes. Usually, in analytical procedures, SPE is carried out using
small columns or cartridges containing the solid stationary phase (sorbent). In volatile
compounds analysis, the most common sorbent used is octadecyl (C18); this material
allows a reversed-phase extraction of mid-polar to nonpolar analytes (Wells, 2003). SPE
has been widely used in grape and wine volatiles analysis (Campone et al., 2018; Picard
et al., 2018; Weldegergis et al., 2011; Williams et al., 1982). SPE has also been applied to
characterize butter flavor (Vreuls et al., 1999), to aromatic compounds from fruit pulps
(Boulanger and Crouzet, 2000), to the analysis of flavor-related to alkylbenzenes in ciga-
rette smoke (Stanfill and Ashley, 2000), and to other nonvolatile compounds present in
food matrices. For more details and examples, the papers published by Andrade-Eiroa
and collaborators (Andrade-Eiroa et al., 2016a,b) give an excellent review of SPEs.

6.3 EXTRACTION METHODS FOR LIQUID OR SOLID SAMPLES

6.3.1 Static Headspace Extraction

This technique is over 30 years old and has been applied to a variety of food matrices for
the extraction of volatile organic compounds, such as fish products (Fukami et al., 2002;
Girard and Nakai, 1991; Duflos et al., 2006; Li et al., 2013) and vegetables (Colina-Coca
et al., 2013; Molina-Calle, Capote and de Castro, 2007), among others. The versatility
of this method allows the sample to be solid or liquid. Meanwhile, its low sensitivity does
not allow for the analysis of trace or high boiling point compounds (Pico et al., 2016). The
most significant advantage of the static headspace extraction technique is the simplicity of
the liquid samples preparation. The preparation involves only transferring the sample into
the vial, typically of 10 to 20 mL, and sealing it immediately. Meanwhile, for solids, the
sample must be ground to increase the superficial area available for the analyte volatiliza-
tion to the headspace, and occasionally the solid sample might be dissolved or suspended
in a liquid to attain equilibrium inside the vial faster. Once the sample is inside the vial and
hermetically sealed, it is incubated at a controlled temperature. The volatile analytes diffuse
to the headspace of the vial. When the equilibrium between the gas and liquid (or solid)
phase is achieved, an aliquot of the headspace gas is injected into a gas chromatograph (GC)
for analysis, as illustrated in Figure 6.2a. The last step may be manual or automatic (Slack,
Snow, and Kou, 2003). The automatic system consists of three stages: (i) Equilibrium; (ii)
Pressurization; and (iii) Sampling, as illustrated in Figure 6.2b.

(i) Equilibrium: This is the most critical stage. Special attention must be given to
equilibrium temperature and time when developing a method. The equilibrium
time is considered once the sample is inserted into the vial and sealed, until the
insertion of the sample needle into the vial. Each compound migrates from the
sample matrix to the gas phase at its own rate, according to the temperature.
Therefore, the minimum equilibrium time depends on the slowest diffusion com-
pound. High volatility compounds might start the migration to the gas phase dur-
ing the vial preparation, occasioning the loss of some volatile compounds before
they arrive in the vial. In these cases, it is recommended to keep the preparation
of the samples at a low temperature. In addition, there are different migration
 Extraction Methods of Volatile Compounds  127

FIGURE 6.2 Schematic diagram of headspace extraction autosampler (a) and the steps
for balanced pressure sampling in GC headspace analysis (b). (Adapted from Kolb 1999.)

rates from the sample matrix to the headspace, since each compound has its
own solubility in the sample matrix, which also depends on the temperature.
Usually, the rise of temperature diminishes the solubility of the VOCs, increas-
ing the concentration of the analyte in the headspace at higher temperatures. It
is recommended to use a temperature of at least 15°C over room temperature to
ensure the correct temperature control. Special attention must be given to higher
temperatures, which may cause degradation of thermolabile compounds.
(ii) Pressurization: Once equilibrium is attained, the headspace gas is ready to be
transferred into a GC. The most common transference mechanism involves the
pressurization of the vial headspace with an inert gas, using a heated hollow
needle, followed by a pressure release into the pneumatic sampler, through the
same needle. Another strategy is to use a system with a sampler loop. In this case,
the pressure inside the vial will transfer the sample into an internal sampler loop,
from where the sample accesses the GC inlet by actioning the sampler valve.
(iii) Sample transference: After pressurization, the gas sample into the vial flows to
the pneumatic sampler through the needle, moved by the pressure gradient estab-
lished during pressurization.

6.3.2 Dynamic Headspace Extraction (Purge and Trap)

Dynamic headspace extraction (DHE) may be used for liquid as well as for solid samples.
It is mainly used when there is a small quantity of the analyte in the sample, at trace
level, or when an exhaustive extraction is needed, and it has been applied for a variety
of matrices, such as sponge cake (Pozo-Bayón et al., 2007), fruit (Mamede and Pastore,
2006), and meat (Madruga et al., 2009), among others. However, it must be consid-
ered that DHE only allows for measurement of the ratio of specific volatile compounds
128 Food Aroma Evolution

FIGURE 6.3 Schematic diagram of a typical purge and trap–GC system (a), and the
needle sparger for purge and trap (b). (Adapted from Zang et al. 2017.)

concentrated over the sorbent surface. Meanwhile, static headspace extraction (SHS)
allows the measurement of the ratio of all of the volatile compounds contained in the
gaseous phase inside the vial (Pico et al., 2016). DHS is similar to SHS; however, the
VOCs are removed continuously from the sample by a continuous inert gas flow. In this
way, a concentration gradient that favors the process exists. The system is composed of a
purge vessel, a sorbent trap (usually Tenax-TA®), a six-way valve, and transference lines,
as illustrated in Figure 6.3a. The sample is inserted into the purge vessel, and a purge
gas (usual helium) passes through the sample continuously carrying the VOCs into the
trap, where the sorbents retain them. When the purge ends, the trap is heated to desorb
the analytes into the GC (Soria, García-Sarrió, and Sanz, 2015). There are three kinds of
purge vessels: (i) frit spargers; (ii) fritless spargers; and (iii) needle spargers. Frit spargers
(Figure 6.3a) are used for liquids that are relatively clear, that are not prone to foam, and
that have solid particles that may clog the system. Fritless spargers and needle spargers
(Figure 6.3b) are recommended for particulate systems and liquids with proteins that are
prone to foam. The trap is usually a stainless-steel tube of 3 mm (ID) and 25 mm long,
packed with different layers of sorbents. The trap must retain the analyte and not intro-
duce impurities. The system operation consists of a series of steps: (i) Purge, a period of
10 to 15 minutes, wherein the carrier gas extracts the VOCs and leads them to the trap,
the samples might be heated by an electrical heater or by microwaves to accelerate the
mass transference from the matrix to the gas phase; (ii) Dry purge, wherein the carrier
gas only passes through the trap, and not through the sample in order to eliminate the
water that might be carried out into the trap along with the analyte; (iii) Desorption pre-
heating, wherein the carrier gas is turned off and the trap is heated to 5 to 10°C below
the desorption temperature, to accelerate desorption; (iv) Desorption, wherein the trap is
heated to 180 to 250°C and the GC gas carrier is turned on in a reversed flow, for about
1 to 4 min. The gas carrying the VOCs is conducted into a GC inlet; (v) Trap bake, the
gas flow returns to the initial direction and the trap temperature rises to 15°C above the
desorption temperature to eliminate possible contamination in the trap and gets the sys-
tem ready for the next sample (Slack, Snow, and Kou, 2003).
 Extraction Methods of Volatile Compounds  129

6.3.3 Solid-Phase Microextraction

This technique was first developed by Belardi and Pawaliszyn (1989) while looking to
reduce the costs and time employed to perform liquid–liquid extraction and solid-phase
extraction. This method has been commercially available since 1993 and has been widely
applied since then on several matrices, such as fish products (Jiménez-Martín et al., 2015),
fruit (Chen et al., 2018; Sdiri et al., 2017), off-flavors (Marsili, 1999; Matsushita et al.,
2017), and spices (Korkmaz, Hayaloglu, and Atasoy, 2017), among others. Solid-phase
microextraction (SPME) is a microscale approach to solid-phase extraction for the extrac-
tion and preconcentration of analytes. The SPME methodology is based on the partition-
ing of analytes between a reusable coated fiber (a stationary phase) and a sample. In SPME,
the analyte molecules must migrate from the sample and penetrate into the fiber coating,
so the mass transfer resistance must be overcome to reach the equilibrium or adsorption
equilibrium, and then the fiber is inserted into the GC inlet to desorb and analyze the
analytes (Merkle, Kleeberg, and Fritsche, 2015). There are two possible approaches for
the analyte adsorption process into the fiber, direct and headspace (HS-SPME). In direct
adsorption, the fiber is immersed into the sample matrix or in a solution containing the
sample. Thus, the fiber might be exposed to nonvolatile compounds that will contaminate
the sample and may affect the chromatography and reduce the number of times that the
fiber might be reused. On the other hand, in HS-SPME, the fiber is inserted into the vial’s
headspace, which contains the sample, the vial is heated, and the volatile compounds are
transferred from the liquid phase to the gaseous phase, and then they are absorbed into
the fiber. Therefore, only volatile compounds reach for the fiber, avoiding undesirable
compounds (Slack, Snow, and Kou, 2003). There are several fiber coatings on the market,
which may be arranged in three groups, polar, semi-polar, and nonpolar. To choose a fiber
coating, one has to consider the nature of the analytes. A fiber coating with similar polar-
ity to that of the analyte will favor its adsorption. Thus, the extraction will be selective,
reducing the chance of extracting contaminant compounds (Valente and Augusto, 2000).
SPME exhibits several advantages over traditional extraction methods, such as to
be a rapid, simple, sensitive, and solvent-free method, and have linear results for a wide
range of concentrations and analytes (Nerín et al., 2009). However, the disadvantages
of SPME are the limited number of commercially available stationary phases (fiber coat-
ings), low recommended operating temperature (240–80°C), the instability and swelling
in organic solvents, breakage of the fiber, stripping of coatings, bending of the needle,
and the cost. Other disadvantages are the limited lifetime of the fiber and the low extrac-
tion efficiencies (Nerín et al., 2009; Merkle, Kleeberg, and Fritsche, 2015). SPME has
been extensively applied to the sampling and analysis of aroma in several raw materials
and food products. Optimization processes for this method involve the selection of the
fiber coating, as well as the fiber diameter, and the extraction conditions, stirring, tem-
perature, direct immersion or headspace, vial volume, sample volume, equilibrium time,
fiber exposure time, and fiber preparation and conservation.

6.4 EXTRACTION METHODS FOR SOLID SAMPLES

In general, analysis of aromas or volatile compounds from plants or foods involves two
steps: extraction and analytical methods. Extraction of analytes aims to separate the
target compounds from the matrix and increase their concentration level. The sample
preparation is an important step which determines the quality of the analysis, and it is
130 Food Aroma Evolution

also the primary source of systematic errors and the lack of precision of analytical meth-
ods (Armenta, Garrigues, and de la Guardia, 2015). A pre-treatment is required before
extraction, aiming at the reduction of the particle size and an increase in the diffusion
of analytes from the sample to the solvent. The choice of appropriate parameters (e.g.,
solvent, sample size, pressure, temperature, number of cycles, and extraction time) is nec-
essary to optimize the process. Moreover, extraction efficiency is mainly influenced by
three factors: the solubility of an analyte on a solvent, the mass transfer properties, and
the matrix effects (Kou and Mitra, 2003).

6.4.1 Conventional Methods

Conventional extraction methods, such as hydro-distillation (HD), simultaneous distilla-

tion extraction (SDE), Soxhlet extraction, and ultrasound-assisted extraction (UAE) are
operated under atmospheric pressure and under heating. These methods consume a large
amount of solvent and may have a long extraction time. On the other hand, there are
other “green” and “innovative” extraction methods, such as supercritical fluid extraction
(SFE), pressurized fluid extraction (PLE), and microwave-assisted extraction (MAE) that
are faster, demand less consumption of solvent, and require less energy to operate (Kou
and Mitra, 2003). Soxhlet is a benchmark method used for the extraction of semi-volatile
organics from solid samples. The main advantages are the independence of the matrix,
the low cost of the equipment, and that further filtration is not required. However, the
disadvantages are the long extraction time and the relatively large amount of solvent
consumed. Study of this methodology to quantify aroma compounds of promising appli-
cation in food industries has been conducted over the years. Recent works have identified
volatile substances from wheat breadcrumb and gluten-free flours (Pico et al., 2016) and
cornstarch (Pico et al., 2018). Soxhlet extraction was also used to identify volatile com-
pounds from grape seed oils (Al Juhaimi and Özcan, 2018).

6.4.2 Ultrasound-Assisted Extraction

UAE can enhance the extraction yield since it increases the mass transfer between the
solvent and plant matrix. The cavitation bubbles lead to a cell disruption near the solid
surface, which improves the solvent penetration and can also break the cell walls. Among
the advantages of UAE are less dependency of the solvent, better solvent penetration,
extraction at lower temperatures, reduced extraction time, a higher yield of extracted
compounds, and faster start-up. Disadvantages are the amount of solvent required and
possible extraction evaporation. A study compared the extraction of volatile compounds
from tea leaves using Soxhlet, ultrasound-assisted extraction, and simultaneous distilla-
tion extraction. It was observed that Soxhlet obtained the highest extraction yield (Gao
et al., 2017). In another study, the quantification of volatile compounds from Schinus
terebinthifolius Raddi fruits was performed by UAC. The authors noticed that high yields
of the extracts might be due to the extraction of high-molecular weight compounds (e.g.,
triterpenes and carotenoids) (Silva et al., 2017). Ultrasonic waves are mechanical vibra-
tions applied to solids, liquids, or gases with frequencies exceeding 20 kHz. Such waves
are different from electromagnetic waves since they need matter to propagate. Their
frequency and wavelength characterize them, and the mathematical product of these
parameters results in the wave velocity through the medium (Wang and Weller, 2006).
 Extraction Methods of Volatile Compounds  131

Ultrasonic waves cause effects of expansion and compression on the matter. The expan-
sion can create bubbles in a liquid and produce negative pressure, while the collapse of
the formed bubbles can cause cavitation. The collapse of bubbles near the cellular walls
produces cellular disruption, and as a result, there is better penetration of the solvent into
the cells, and consequently an increase in the mass transfer (Esclapez et al., 2011). Figure
6.4 is a schematic representation of the effects of the ultrasound process on the interface
of a plant matrix.
In Figure 6.4, the formation of bubbles can be seen in the first stage, (a). Such bubbles
undergo expansion and compression (b), which will cause their collapse or implosion
(c). Eventually, should this collapse occur near the array interface, it can generate shock
waves that will disturb the wall of the matrix, consequently releasing the intraparticular
material into the solvent (d) (Esclapez et al., 2011). An extremely important factor in
ultrasonic-assisted extraction is the extractor configuration, which may have the trans-
ducer coupled to the extraction vessel or have an ultrasonic probe immersed in the sol-
vent/matrix medium. Figure 6.5 shows some extractor geometries schematically with
ultrasonic waves.
Currently, the most widely used laboratory scale extraction systems are ultrasonic
baths or direct sonification. Ultrasonic baths consist of a transducer coupled to a tank con-
taining a liquid responsible for transferring the waves to a container containing the extrac-
tive matrix, according to Figure 6.5a. The disadvantages of this geometry are the lack of
uniformity of ultrasound wave distribution. Thus, direct sonification emerged as an alter-
native to improve the distribution of the ultrasonic waves through the extraction medium.
This system consists of inserting an ultrasonic probe directly into the solvent/matrix mix-
ture, as shown in Figure 6.5b and c (Esclapez et al., 2011). The use of ultrasound was con-
ducted in the extraction of aroma compounds from aged brandies, tea, wine, and garlic,
in the extraction of antioxidants from pomegranate peel, and carotenoids from tomato
waste (Chemat et al., 2017). Moreover, some studies extracted lycopene from tomatoes,

FIGURE 6.4 Schematic representation of the effects of the ultrasound process on the
matrix vegetable. (Adapted from Esclapez et al., 2011; and Capote and de Castro,
2007.)
132 Food Aroma Evolution

FIGURE 6.5 Typical extractor configurations: (a) transducer coupled to a vessel; trans-
ducer (probe) immersed in solvent/plant matrix medium, (b) in batch and in continuous
(considering the fluid/solvent) (c). (Adapted from Esclapez et al., 2011.)

phenolics from strawberries, citrus peel, and coconut shell powder, anthocyanins from red
raspberries, and capsaicinoids from peppers (Chemat and Khan, 2011).

6.4.3 Microwave-Assisted Extraction

Microwave-assisted extraction works with the dissipation of the electromagnetic

waves in the irradiated medium through heat. Among the main advantages of this
technique are the reduced costs, easy equipment manipulation, higher purity of the
final products, reduced extraction time, and energy consumption. However, MAE
requires a post-extraction step (e.g., cooling and filtration) which can extend the
process and it is quite an exhaustive process including interfering species that need
cleanup before the analysis. For food production, it is worth mentioning that this
method allows the reduction of the equipment size, a faster response to heating
processes, and increases of the production and elimination of post-treatment steps
(Chemat et al., 2017). Solvent-free microwave extraction (SFME) is an adaption of
MAE that uses a combination of microwave heating and dry distillation under atmo-
spheric pressure without the use of solvent or water. This process has been used to
extract aroma compounds from citrus fruit (e.g., orange [Ferhat et al., 2006] and
lemon [Ferhat et al., 2007a]), aromatic herbs (e.g., basil, mint, and thyme) and spices
(e.g., cumin and anise [Lucchesi, Chemat, and Smadja, 2004]). In a recent study com-
paring SFME with a conventional hydro-distillation extraction to obtain essential oil
from Origanum vulgare L, it was observed that SFME was more efficient with a higher
extraction yield, and in a shorter extraction time (Bayramoglu, Sahin, and Sumnu,
2008). Another study compared microwave-assisted simultaneous distillation-solvent
extraction (MW-SDE) with conventional SDE in the extraction of volatile compounds
from fresh aromatic herb (Zygophyllum album L.). It was noticed that MW-SDE was
faster, required less solvent amounts and less energy in comparison to conventional
SDE extraction (Ferhat et al., 2007b).
 Extraction Methods of Volatile Compounds  133

6.4.4 Supercritical and Pressurized Fluid Extraction

Among the innovative extraction processes, SFE appears as an alternative to the use of
conventional methods. SFE is considered a green methodology that aims to obtain com-
pounds with reduced energy requirements, shorter extraction times, and using solvents
generally recognized as safe (GRAS). Supercritical fluids enhance transport properties,
due to their high diffusivity and low viscosity, being able to readily diffuse through solid
materials with faster extraction rates. Supercritical carbon dioxide (SC-CO2) is the most
common solvent used since it is a non-toxic, non-flammable, non-polluting, and low-cost
solvent; it is relatively inert and can be recovered (Brunner, 2005). Moreover, it works
at moderate critical conditions (Tc = 31.05°C and Pc = 7.38 MPa) that are crucial in the
preservation of aroma compounds from extracts. However, the equipment costs can be
a disadvantage of this technique when scale-up is aimed for. Another SC-CO2 limita-
tion is its low polarity making the process more appropriate for extracting nonpolar
compounds.
The process of extraction with a supercritical fluid, specifically using CO2 , occurs in
two stages: the extraction of the solutes and the separation of the solute from the solvent.
The first step is to manipulate the carbon dioxide in the binomial pressure/temperature
in order to obtain the highest solvation of the target solutes. The solvent flows into the
extractor and through the entire plant matrix, solubilizing the solutes. The solvent/solute
then mixture goes to the second step, in which the pressure is reduced below the criti-
cal point value. In this way, the solvent changes its state of supercritical aggregation to
gas, reducing its power of solvation, and consequently precipitation of the solute occurs.
Thus, the solute is recovered, and the gas is redirected into a recycle (Raventós, Duarte,
and Alarcón, 2002; Martínez, 2007). Figure 6.6 shows the supercritical fluid extraction
process. The recycle conducts the gaseous carbon dioxide into a condenser, where the gas

FIGURE 6.6 Flow diagram of a supercritical extraction process from solid matrices.
(Adapted from Rosa et al., 2008.)
134 Food Aroma Evolution

is liquefied. Thereafter, the CO2 has its pressure increased above the pressure of the criti-
cal point, as a result of the work of a pump, and its elevated temperature up to the desired
operating temperature by a heater (Rosa et al., 2008).
The use of SFE to obtain extracts enriched in a specific compound of interest for the
food industries has been carried out throughout the years, such as the extraction of caf-
feine from green coffee beans, free fatty acids from rice bran oil, tocopherol from wheat
germ oil, and beta-carotene from crude palm oil. SFE has been used in the fractionation
of fatty acid ethyl esters from fish oil, extraction of essential oils from seeds of a pome-
granate, walnut oil, carrot fruit, and carotenoids from pumpkins. The application of
SFE was also studied before analysis of volatile compounds in beverages and sugar cane
(Herrero et al., 2010). The use of SFE assisted by ultrasound (SFE-US) has emerged as an
alternative for extracting and quantifying bioactive compounds from food samples, since
the ultrasound waves can increase the rupture of the cell walls of the matrix enhancing
solvent penetration and extraction yield. SFE-US was studied for the extraction of aroma
compounds from ginger (Balachandran et al., 2006), essential oil from almond (Riera
et al., 2004), antioxidants from blackberry (Pasquel Reátegui et al., 2014), tocopherol
and tocotrienols from passion fruit (Barrales, Rezende, and Martínez, 2015), and capsa-
icinoids from peppers (Santos et al., 2015; Dias et al., 2016). Pressurized liquid extraction
(PLE) is an alternative technique that uses liquid, like ethanol or/and water, as solvents at
elevated pressure and temperature, enhancing the solubility and mass transfer properties
of an analyte in a solvent. Moreover, the higher temperature diminishes the solvent’s vis-
cosity and surface tension, improving the solvent penetration in the matrix sample. PLE
works with a reduced extraction time and has less solvent consumption, and it does not
require a filtration step when compared to conventional processes, which is important
taking into account the use of automated and online systems. Furthermore, the use of
polar solvents, such as water and ethanol, covers the extraction of compounds of high
and intermediate polarity. The main drawback of this technique is the cost of equip-
ment. The use of PLE to obtain food aroma compounds was recently performed in the
quantification of anthocyanins, phenolics, antioxidants, isoflavonoids, carotenoids, and
capsaicinoids (Mustafa and Turner, 2011).

REFERENCES

Al Juhaimi, Fahad and Mehmet Musa Özcan. 2018. Effect of cold press and soxhlet
extraction systems on fatty acid, tocopherol contents, and phenolic compounds of
various grape seed oils. Journal of Food Processing and Preservation 42 (1):e13417.
Andrade-Eiroa, Auréa, Moisés Canle, Valérie Leroy-Cancellieri, and Víctor Cerdà.
2016a. Solid-phase extraction of organic compounds: A critical review: Part I. TrAC
Trends in Analytical Chemistry 80:641–54.
Andrade-Eiroa, Auréa, Moisés Canle, Valérie Leroy-Cancellieri, and Víctor Cerdà.
2016b. Solid-phase extraction of organic compounds: A critical review: Part II.
TrAC Trends in Analytical Chemistry 80:655–67.
Armenta, Sergio, Salvador Garrigues, and Miguel de la Guardia. 2015. The role of green
extraction techniques in Green Analytical Chemistry. TrAC Trends in Analytical
Chemistry 71:2–8.
Augusto, Fabio, Alexandre Leite e Lopes, and Claudia Alcaraz Zini. 2003. Sampling
and sample preparation for analysis of aromas and fragrances. TrAC Trends in
Analytical Chemistry 22 (3):160–9.
 Extraction Methods of Volatile Compounds  135

Balachandran, S, SE Kentish, R Mawson, and M Ashokkumar. 2006. Ultrasonic

enhancement of the supercritical extraction from ginger. Ultrasonics Sonochemistry
13 (6):471–9.
Barrales, Francisco Manuel, Camila Alves Rezende, and Julian Martínez. 2015.
Supercritical CO2 extraction of passion fruit (Passiflora edulis sp.) seed oil assisted
by ultrasound. The Journal of Supercritical Fluids 104:183–92.
Bayramoglu, Beste, Serpil Sahin, and Gulum Sumnu. 2008. Solvent-free microwave
extraction of essential oil from oregano. Journal of Food Engineering 88 (4):535–40.
Belardi, Robert P and Janusz B Pawliszyn. 1989. The application of chemically modified fused
silica fibers in the extraction of organics from water matrix samples and their rapid trans-
fer to capillary columns. Water Quality Research Journal of Canada 24 (1):179–91.
Boulanger, Renaud and Jean Crouzet. 2000. Free and bound flavour components of
Amazonian fruits: 2. Cupuacu volatile compounds. Flavour and Fragrance Journal
15 (4):251–7.
Brunner, Gerd. 2005. Supercritical fluids: Technology and application to food processing.
Journal of Food Engineering 67 (1):21–33.
Campone, Luca, Anna Lisa Piccinelli, Rita Celano, Imma Pagano, Mariateresa Russo,
and Luca Rastrelli. 2018. Rapid and automated on-line solid-phase extraction
HPLC–MS/MS with peak focusing for the determination of ochratoxin A in wine
samples. Food Chemistry 244:128–35.
Canbay, Hale Seçilmiş. 2017. Effectiveness of liquid-liquid extraction, solid-phase extrac-
tion, and headspace technique for determination of some volatile water-soluble
compounds of rose aromatic water. International Journal of Analytical Chemistry
2017:4870671.
Capote, F Priego and MD Luque de Castro. 2007. Analytical Applications of Ultrasound.
Vol. 26. Elsevier.
Chemat, Farid, Natacha Rombaut, Alice Meullemiestre, Mohammad Turk, Sandrine
Perino, Anne-Sylvie Fabiano-Tixier, and Maryline Abert-Vian. 2017. Review of
Green Food Processing techniques. Preservation, transformation, and extraction.
Innovative Food Science & Emerging Technologies 41:357–77.
Chemat, Farid, Natacha Rombaut, Anne-Gaëlle Sicaire, Alice Meullemiestre, Anne-Sylvie
Fabiano-Tixier, and Maryline Abert-Vian. 2017. Ultrasound assisted extraction of
food and natural products. Mechanisms, techniques, combinations, protocols and
applications. A review. Ultrasonics Sonochemistry 34:540–60.
Chemat, Farid and Muhammed Kamran Khan. 2011. Applications of ultrasound in food
technology: Processing, preservation and extraction. Ultrasonics Sonochemistry 18
(4):813–35.
Chen, Yangyang, Hao Yin, Xiao Wu, Xinjie Shi, Kaijie Qi, and Shaoling Zhang. 2018.
Comparative analysis of the volatile organic compounds in mature fruits of 12
Occidental pear (Pyrus communis L.) cultivars. Scientia Horticulturae 240:239–48.
Colina-Coca, Clara, Diana González-Peña, Estela Vega, Begoña de Ancos, and
Concepción Sánchez-Moreno. 2013. Novel approach for the determination of vola-
tile compounds in processed onion by headspace gas chromatography–mass spec-
trometry (HS GC–MS). Talanta 103:137–44.
Dias, Arthur Luiz Baião, Camilla Scarelli Arroio Sergio, Philipe Santos, Gerardo
Fernandéz Barbero, Camila Alves Rezende, and Julian Martínez. 2016. Effect of
ultrasound on the supercritical CO2 extraction of bioactive compounds from dedo
de moça pepper (Capsicum baccatum L. var. pendulum). Ultrasonics Sonochemistry
31:284–94.
136 Food Aroma Evolution

Duflos, Guillaume, Vincent Marcel Coin, Marie Cornu, Jean‐Francois Antinelli, and Pierre
Malle. 2006. Determination of volatile compounds to characterize fish spoilage using
headspace/mass spectrometry and solid-phase microextraction/gas chromatography/
mass spectrometry. Journal of the Science of Food and Agriculture 86 (4):600–11.
Esclapez, MD, JV García-Pérez, A Mulet, and JA Cárcel. 2011. Ultrasound-assisted
extraction of natural products. Food Engineering Reviews 3 (2):108.
Ferhat, Mohamed A, Brahim Y Meklati, and Farid Chemat. 2007a. Comparison of dif-
ferent isolation methods of essential oil from Citrus fruits: Cold pressing, hydro-
distillation and microwave “dry”distillation. Flavour and Fragrance Journal 22
(6):494–504.
Ferhat, Mohamed A, Brahim Y Meklati, Jacqueline Smadja, and Farid Chemat. 2006.
An improved microwave Clevenger apparatus for distillation of essential oils from
orange peel. Journal of Chromatography A 1112 (1):121–6.
Ferhat, Mohamed A, Nacéra Tigrine-Kordjani, Smain Chemat, BrahimY Meklati, and
Farid Chemat. 2007b. Rapid extraction of volatile compounds using a new simul-
taneous microwave distillation: Solvent extraction device. Chromatographia 65
(3–4):217–22.
Fukami, Katsuya, Sachiyo Ishiyama, Hitoshi Yaguramaki, Takuya Masuzawa, Yoshimitu
Nabeta, Kenichi Endo, and Mitsuya Shimoda. 2002. Identification of distinctive
volatile compounds in fish sauce. Journal of Agricultural and Food Chemistry 50
(19):5412–6.
Gao, Xuemei, Shidong Lv, Yuanshuang Wu, Jiangbing Li, Wenrui Zhang, Wen Meng,
Chen Wang, and Qingxiong Meng. 2017. Volatile components of essential oils
extracted from Pu-erh ripe tea by different extraction methods. International
Journal of Food Properties 20 (Supp. 1):S240–53.
Girard, B and S Nakai. 1991. Static headspace gas chromatographic method for volatiles
in canned salmon. Journal of Food Science 56 (5):1271–4.
Herrero, Miguel, José A Mendiola, Alejandro Cifuentes, and Elena Ibáñez. 2010.
Supercritical fluid extraction: Recent advances and applications. Journal of
Chromatography A 1217 (16):2495–511.
Jiménez-Martín, Estefanía, Adem Gharsallaoui, Trinidad Pérez-Palacios, Jorge Ruiz
Carrascal, and Teresa Antequera Rojas. 2015. Volatile compounds and physico-
chemical characteristics during storage of microcapsules from different fish oil
emulsions. Food and Bioproducts Processing 96:52–64.
Kelebek, Hasim and Serkan Selli. 2011. Determination of volatile, phenolic, organic acid
and sugar components in a Turkish cv. Dortyol (Citrus sinensis L. Osbeck) orange
juice. Journal of the Science of Food and Agriculture 91 (10):1855–62.
Kolb, Bruno. 1999. Headspace sampling with capillary columns. Journal of
Chromatography A 842 (1):163–205.
Korkmaz, Aziz, Ali Adnan Hayaloglu, and Ahmet Ferit Atasoy. 2017. Evaluation of the
volatile compounds of fresh ripened Capsicum annuum and its spice pepper (dried
red pepper flakes and isot). LWT—Food Science and Technology 84:842–50.
Kou, Dawen and Somenath Mitra. 2003. Extraction of semi-volatile organic compounds
from solid matrices. Sample Preparation Techniques in Analytical Chemistry
162:139–78.
Li, Chunping, Jiajia Wu, Yan Li, and Zhiyuan Dai. 2013. Identification of the aroma
compounds in stinky mandarin fish (S iniperca chuatsi) and comparison of vol-
atiles during fermentation and storage. International Journal of Food Science &
Technology 48 (11):2429–37.
 Extraction Methods of Volatile Compounds  137

López, E Falqué and E Fernández Gómez. 2000. Comparison of solvents for determina-
tion of monoterpenes in wine using liquid-liquid extraction. Chromatographia 52
(11):798–802.
Lucchesi, Marie E, Farid Chemat, and Jacqueline Smadja. 2004. Solvent-free microwave
extraction: An innovative tool for rapid extraction of essential oil from aromatic
herbs and spices. Journal of Microwave Power and Electromagnetic Energy 39
(3–4):135–9.
Madruga, Marta S, J Stephen Elmore, Andrew T Dodson, and Donald S Mottram. 2009.
Volatile flavour profile of goat meat extracted by three widely used techniques. Food
Chemistry 115 (3):1081–7.
Mamede, Maria EO and Gláucia M Pastore. 2006. Study of methods for the extraction
of volatile compounds from fermented grape must. Food Chemistry 96 (4):586–90.
Marsili, R. 2016. Flavors and off-flavors in dairy foods. In Reference Module in Food
Science. Elsevier.
Marsili, RT. 1999. SPME–MS–MVA as an electronic nose for the study of off-flavors in
milk. Journal of Agricultural and Food Chemistry 47 (2):648–54.
Martínez, José L. 2007. Supercritical Fluid Extraction of Nutraceuticals and Bioactive
Compounds. CRC Press.
Matsushita, Taku, Miki Sakuma, Shiori Tazawa, Taiki Hatase, Nobutaka Shirasaki, and
Yoshihiko Matsui. 2017. Use of gas chromatography–mass spectrometry–olfactom-
etry and a conventional flask test to identify off-flavor compounds generated from
phenylalanine during chlorination of drinking water. Water Research 125:332–40.
Merkle, Sybille, Kim Karen Kleeberg, and Jan Fritsche. 2015. Recent developments and
applications of solid-phase microextraction (SPME) in food and environmental
analysis—A review. Chromatography 2 (3):293–381.
Mestres, M, O Busto, and J Guasch. 2000. Analysis of organic sulfur compounds in wine
aroma. Journal of Chromatography A 881 (1–2):569–81.
Molina-Calle, María, Feliciano Priego-Capote, and María D Luque de Castro. 2017.
Headspace–GC–MS volatile profile of black garlic vs fresh garlic: Evolution along
fermentation and behavior under heating. LWT-Food Science and Technology
80:98–105.
Mustafa, Arwa and Charlotta Turner. 2011. Pressurized liquid extraction as a green
approach in food and herbal plants extraction: A review. Analytica Chimica Acta
703 (1):8–18.
Nerín, C, J Salafranca, M Aznar, and R Batlle. 2009. Critical review on recent devel-
opments in solventless techniques for extraction of analytes. Analytical and
Bioanalytical Chemistry 393 (3):809.
Panighel, Annarita and Riccardo Flamini. 2015. Solid-phase extraction and solid-phase
microextraction in grape and wine volatile compounds analysis. Sample Preparation
2:55–65.
Pasquel Reátegui, José Luis, Ana Paula da Fonseca Machado, Gerardo F Barbero, Camila
A Rezende, and Julian Martínez. 2014. Extraction of antioxidant compounds from
blackberry (Rubus sp.) bagasse using supercritical CO2 assisted by ultrasound. The
Journal of Supercritical Fluids 94:223–33.
Picard, Magali, Céline Franc, Gilles de Revel, and Stéphanie Marchand. 2018. Dual
solid-phase and stir bar sorptive extraction combined with gas chromatogra-
phy–mass spectrometry analysis provides a suitable tool for assaying limonene-
derived mint aroma compounds in red wine. Analytica Chimica Acta 1001:
168–78.
138 Food Aroma Evolution

Pico, Joana, Jesús Tapia, José Bernal, and Manuel Gómez. 2018. Comparison of different
extraction methodologies for the analysis of volatile compounds in gluten-free flours
and corn starch by GC/QTOF. Food Chemistry 267:303–12.
Pico, Joana, Manuel Gómez, José Bernal, and José Luis Bernal. 2016. Analytical methods
for volatile compounds in wheat bread. Journal of Chromatography A 1428:55–71.
Pico, Joana, María Jesús Nozal, Manuel Gómez, and José Luis Bernal. 2016. An alterna-
tive method based on enzymatic fat hydrolysis to quantify volatile compounds in
wheat bread crumb. Food Chemistry 206:110–8.
Pozo-Bayón, Maria Angeles, Alejandro Ruíz-Rodríguez, Karine Pernin, and Nathalie
Cayot. 2007. Influence of eggs on the aroma composition of a sponge cake and on
the aroma release in model studies on flavored sponge cakes. Journal of Agricultural
and Food Chemistry 55 (4):1418–26.
Raventós, M, S Duarte, and R Alarcón. 2002. Application and possibilities of super-
critical CO2 extraction in food processing industry: An overview. Food Science and
Technology International 8 (5):269–84.
Riera, E, Y Golás, A Blanco, JA Gallego, M Blasco, and A Mulet. 2004. Mass trans-
fer enhancement in supercritical fluids extraction by means of power ultrasound.
Ultrasonics Sonochemistry 11 (3):241–4.
Rocha, S, P Coutinho, A Barros, MA Coimbra, I Delgadillo, and A Dias Cardoso. 2000.
Aroma potential of two bairrada white grape varieties: Maria Gomes and Bical.
Journal of Agricultural and Food Chemistry 48 (10):4802–7.
Rosa, Paulo TV, Juan Carlos Parajó, Herminia Domínguez, Andrés Moure, Beatriz
Díaz-Reinoso, RL Smith, Masaaki Toyomizu, Louw J Florusse, Cor J Peters, and
Motonobu Goto. 2008. Supercritical and pressurized fluid extraction applied to the
food industry. In Extracting Bioactive Compounds for Food Products. Theory and
Applications 272-400. CRC Press. ISBN 978-1-4200-6237-3.
Santos, Philipe, Ana C Aguiar, Gerardo F Barbero, Camila A Rezende, and Julian
Martínez. 2015. Supercritical carbon dioxide extraction of capsaicinoids from
malagueta pepper (Capsicum frutescens L.) assisted by ultrasound. Ultrasonics
Sonochemistry 22:78–88.
Sdiri, Sawsen, José L Rambla, Cristina Besada, Antonio Granell, and Alejandra Salvador.
2017. Changes in the volatile profile of citrus fruit submitted to postharvest degreen-
ing treatment. Postharvest Biology and Technology 133:48–56.
Selli, S, H Kelebek, MT Ayseli, and H Tokbas. 2014. Characterization of the most aroma-
active compounds in cherry tomato by application of the aroma extract dilution
analysis. Food Chemistry 165:540–6.
Sides, Alasdair, Kevin Robards, and Stuart Helliwell. 2000. Developments in extraction
techniques and their application to analysis of volatiles in foods. TrAC Trends in
Analytical Chemistry 19 (5):322–9.
Silva, Bruno Guzzo da, Ana Maria Frattini Fileti, Mary Ann Foglio, Ana Lúcia Tasca
Gois Ruiz, and Paulo de Tarso Vieira e Rosa. 2017. Supercritical carbon dioxide
extraction of compounds from Schinus terebinthifolius Raddi fruits: Effects of oper-
ating conditions on global yield, volatile compounds, and antiproliferative activity
against human tumor cell lines. The Journal of Supercritical Fluids 130:10–6.
Slack, Gregory C, Nicholas H Snow, and Dawen Kou. 2003. Extraction of Volatile
Organic Compounds from Solids and Liquids. Vol. 162. Wiley-Interscience,
Hoboken, NJ.
Soria, Ana C, MJ García-Sarrió, and M Luz Sanz. 2015. Volatile sampling by headspace
techniques. TrAC Trends in Analytical Chemistry 71:85–99.
 Extraction Methods of Volatile Compounds  139

Stanfill, Stephen B and David L Ashley. 2000. Quantitation of flavor-related alkenylben-

zenes in tobacco smoke particulate by selected ion monitoring gas chromatography–
mass spectrometry. Journal of Agricultural and Food Chemistry 48 (4):1298–306.
Valente, Antonio Luiz Pires and Fabio Augusto. 2000. Microextração por fase sólida.
Química Nova 23 (4): 523–530.
Varlet, Vincent, Carole Prost, and Thierry Serot. 2007. Volatile aldehydes in smoked fish:
Analysis methods, occurence and mechanisms of formation. Food Chemistry 105
(4):1536–56.
Vreuls, René JJ, Arnold van der Heijden, A Udo, and Mohamed Adahchour. 1999. Trace-
level determination of polar flavour compounds in butter by solid-phase extraction
and gas chromatography–mass spectrometry. Journal of Chromatography A 844
(1–2):295–305.
Wang, Lijun and Curtis L Weller. 2006. Recent advances in extraction of nutraceuticals
from plants. Trends in Food Science & Technology 17 (6):300–12.
Weldegergis, Berhane T, Andrew M Crouch, Tadeusz Górecki, and André de Villiers.
2011. Solid-phase extraction in combination with comprehensive two-dimensional
gas chromatography coupled to time-of-flight mass spectrometry for the detailed
investigation of volatiles in South African red wines. Analytica Chimica Acta 701
(1):98–111.
Wells, Martha JM. 2003. Principles of extraction and the extraction of semi-volatile
organics from liquids. Sample Preparation Techniques in Analytical Chemistry
162:37.
Williams, PJ, CR Strauss, B Wilson, and RA Massy-Westropp. 1982. Use of C18 reversed-
phase liquid chromatography for the isolation of monoterpene glycosides and nor-
isoprenoid precursors from grape juice and wines. Journal of Chromatography A
235 (2):471–80.
Zang, Xiaohuan, Weiqian Liang, Qingyun Chang, Tong Wu, Chun Wang, and Zhi Wang.
2017. Determination of volatile organic compounds in pen inks by a dynamic head-
space needle trap device combined with gas chromatography–mass spectrometry.
Journal of Chromatography A 1513:27–34.
 Chapter 7
The Role of Gas
Chromatography-Based
Methodologies for the
Understanding of Food Aromas
Cátia Martins, Ângelo C. Salvador, and Sílvia M. Rocha

CONTENTS

7.1 Introduction 141

7.2 Sample Preparation toward Gas Chromatographic Analysis 142
7.3 The Role of Gas Chromatographic-Based Tools: Principles and
Potentialities 143
7.3.1 One-Dimensional Gas Chromatography (1D-GC) 144
7.3.2 Comprehensive Two-Dimensional Gas Chromatography (GC×GC) 145
7.3.3 Olfactometry (GC-O) 150
7.4 Concluding Remarks and Future Trends 152
Acknowledgments 153
References 153

7.1 INTRODUCTION

Food aroma is of major concern to food researchers and industrial teams as it represents
a significant factor influencing the public’s food-buying decisions, as well as being associ-
ated with food quality and safety. Odorous compounds are typically volatile or semi-vola-
tile in nature and have a low molecular weight (the majority below 300 amu). Despite this
apparently limited range, odorous stimuli belong to a broad variety of s­ ubstance classes
that comprise diverse structural moieties such as ester, alcohol, aldehyde, or ketone func-
tions, among others, having aromatic or aliphatic forms, or comprising of thio and other
heteroatomic groups (Baldovini, 2017).
A detailed analysis of the literature allows us to infer that there is some misconcep-
tion between characterizing volatile food composition and understanding its aroma, as
the study of a food aroma is much more than the analysis of its volatile composition.
Indeed, the human nose’s perception of aromas depends, among other factors, on the
extension of the volatiles released from the matrix and the odor properties of the com-
pounds, in which these molecules do not contribute equally to the overall flavor profile of
a sample; thus, a higher chromatographic area generated by a chemical detector does not

141
142 Food Aroma Evolution

necessarily correspond to higher odor intensities (Zellner et al., 2008). The aroma percep-
tion also depends on the characteristics of the nasal and oral physiology of mucosa and
microbiota. The experiences and memories of the individual and their sensory acuity are
other significant parameters critical to aroma recognition. Aroma characterization has
been an issue of concern for a long time and new devices and innovative approaches have
been developed, such as gas chromatography-olfactometry (GC-O) that combines senso-
rial and instrumental data (Giungato et al., 2018; Song and Liu, 2018), or highly sensitive
equipment, for instance, comprehensive two-dimensional gas chromatograph coupled to
a mass spectrometer with a time-of-flight analyzer (GC×GC-ToF-MS). The determination
of aroma molecules, as well as the knowledge of their origin and possible evolution, for
instance, during ripening, processing, storage, or aging, are crucial for understanding and
modulating the aroma properties of foods and their perception by the consumer. These
challenges are intimately connected with the use of gas chromatographic methodologies.
Thus, this chapter aims to highlight the role of gas chromatographic-based techniques to
understand the aroma of foods. The 1-D, GC×GC, and GC-O will be presented, as well as
the concepts and main potentialities and challenges related to the use of these tools. First,
the phases preceding chromatographic analysis will be briefly presented.

7.2 SAMPLE PREPARATION TOWARD GAS

CHROMATOGRAPHIC ANALYSIS

Food volatile characterization should be carefully planned, and the construction of a

workflow that includes the sequence of all steps, that is, sampling, extraction, instrumen-
tal analysis and data processing and interpretation, is recommended (Figure 7.1).
This chapter will focus on instrumental analysis using gas chromatographic-based
techniques; nevertheless, there are several considerations with sampling and extraction
that the authors want to emphasize as they can contribute to misinterpretations of the
data. In fact, food aroma scientists face enormous challenges regarding this topic, and,
as extraction techniques were discussed in detail in a previous chapter, this section only
highlights some concerns that are antecedent to aroma chromatographic analysis per se.
The sampling and extraction steps should be carefully established in order to generate
volatile-related data that may be useful for food item aroma comprehension. Therefore,

FIGURE 7.1 Workflow that summarizes the main steps that may be used for food vola-
tile characterization, including the odorant molecules, here illustrated by beer as a case
study: (1) sampling, (2) extraction, (3) instrumental analysis, and (4) data processing and
interpretation.
 The Role of Gas Chromatography-Based Methodologies  143

it is extremely important to have a representative aliquot of the food item (even in size/
amount), whose analysis may reflect the active odor volatiles released from the food item,
as close as possible. Also, another crucial factor is the number of samples, which should
be representative and statistically significant of the overall food population, and indepen-
dent sample aliquots may be preferred over analytical replicates (Giungato et al., 2018;
Schieberle and Molyneux, 2012). The perceived aroma of food items usually reflects the
free volatile compounds. However, an in-depth study may also comprise the analysis of
aroma precursor compounds, such as in the form of glycones, or high-molecular weight
molecules. For instance, monoterpenic compounds may be present in fruits linked to sugar
molecules (glycones); thus, these compounds, as nonvolatile, do not contribute to aroma
perception. However, during ripening or technological processes, the glycosidic-linkage
may be broken, and the free released compounds may consequently contribute for aroma
(Rocha et al., 2010). Thus, to estimate the potential impact of the compounds present in
the form of glycones, and to mimic the effect of physiologic or technological processes
that may contribute to their release, the sampling step may include an acid and/or enzy-
matic hydrolysis before the extraction step. Food aroma compounds are generally present
in low concentration, being a common practice to concentrate volatiles extracted from
foods before their analysis. Many extraction techniques are often used to extract and/or
to simultaneously extract and concentrate volatile compounds from foods, such as head-
space, solvent, or sorptive based techniques. For instance, after extraction with solvents,
and before chromatographic analysis, the solvent should be removed to concentrate the
volatile fraction. Otherwise, sorptive techniques, such as solid-phase microextraction
(SPME), promote the simultaneous extraction and concentration of volatiles. A food’s
volatile composition can be significantly changed particularly during pre-treatment tech-
niques if an adequate procedure is not performed. For instance, compounds may be lost or
modified during the extraction steps. Factors such as concentration of the solvent extracts
before analysis, storage before analysis, or cross-contamination with storage and/or analy-
sis materials may influence the volatile composition (Nongonierma et al., 2007). Hence,
the main requirements of an extraction protocol are to ensure that volatiles are extracted
in a representative way; their variability, as a result of the extraction step, is minimized;
and compounds are not modified due to the extraction method (Rodrigues et al., 2016).

7.3 THE ROLE OF GAS CHROMATOGRAPHIC-BASED

TOOLS: PRINCIPLES AND POTENTIALITIES

The volatile characterization of food items is usually performed using gas chromato-
graphic techniques. One-dimensional gas chromatography (1D-GC) is the most common
and widely used analytical method, not only by food researchers but also by industrial
teams. Nevertheless, the complexity of the volatile composition of foods usually exceeds
the capacity of one-dimensional separation. Co-elutions may compromise reliable identifi-
cations. Thus, there is a need to constantly seek for more sensitive and selective analytical
tools that are able to study targeted and untargeted compounds from these complex matri-
ces, with short instrumental analysis and data processing times. Noteworthy improve-
ments have been occurring such as the development of new GC column stationary phases
(namely ionic liquids), enhancement of instrumental technologies (e.g., new detectors, such
as high resolution mass spectrometers), and more powerful data analysis through new
software and/or algorithms (Prebihalo et al., 2018; Wong, Hayward, and Zhang, 2013).
Also, instrumentation has evolved from 1D-GC to multidimensional gas chromatography
144 Food Aroma Evolution

(MDGC), namely comprehensive two-dimensional gas chromatography (GC×GC) that

has been applied to determine the volatile profile from a wide variety of foods and bever-
ages, such as beer (Martins et al., 2015, 2018), clams (Rocha et al., 2013), elderflowers
(Salvador, Silvestre, and Rocha, 2017), elderberries (Salvador et al., 2016), grapes (Rocha
et al., 2007), aromatic plants (Petronilho et al., 2013, 2011; Jalali et al., 2012, 2013), sea
salt (Silva et al., 2015, 2010), and wine (Santos et al., 2015, 2013; Perestrelo et al., 2010,
2011). Furthermore, instrumental devices that combine the chromatographic and senso-
rial evaluation of food items, such as gas chromatography-olfactometry (GC-O), are very
powerful tools for unveiling food aromas. This technique combines both olfactometry and
mass spectrometry fields, where the obtained data complement the information of each
detection mode, used for rapid mapping of aroma-active compounds, identification of key
aroma-active compounds, cluster analysis based on the aroma-active compounds, relation-
ship between odorants and sensory properties, and elucidation of formation mechanism of
important odorants. In the following sections, the main gas chromatographic-based tools
used for the understanding of food aromas will be presented.

7.3.1 One-Dimensional Gas Chromatography (1D-GC)

Gas chromatographic separation processes foresee that the compounds separate ­according
to partitioning (dispersion and specific interactions) between two immiscible phases,
the mobile phase (e.g., helium, hydrogen, or nitrogen) and the stationary phase (Grob
and Barry, 2004). 1D-GC schematic representation is shown in Figure 7.2, where the
GC column is connected in between an injector and a detector, within a temperature
programmable oven. As food volatile components exhibit a high diversity of chemical
structures, there are a wide range of stationary phases for GC columns, such as polyethyl-
ene glycol (polar) and 5% of phenylmethylpolysiloxane or equivalents (nonpolar), which
are frequently used for foods volatile profile determination. Different types of detectors
may also be connected with the gas chromatograph, the flame ionization detector (FID)
is commonly used, which is cheaper and more used for target analysis than the mass
spectrometry detector (MS). This last type of detector allows for powerful and effective
compound identification, based on a mass spectrum fragmentation pattern, and possible
comparison with MS databases, also promoting high sensitivity and sensibility.
GC-MS equipment has robust software that facilitates data processing, either for
quantitative or qualitative purposes (Dettmer, Aronov, and Hammock, 2007; Milman,
2015). Indeed, different approaches may be used to acquire and process the MS data: full-
scan (scanning in a m/z range), single or multiple ion monitoring (SIM or MIM—data

FIGURE 7.2 Schematic illustration of the one-dimensional gas chromatographic system

(1D-GC).
 The Role of Gas Chromatography-Based Methodologies  145

acquisition using specific m/z as diagnostic ions), and ion extraction mode (IEC—use of
specific m/z as diagnostic ions to process MS data). As peaks co-elution may occur in the
full-scan data acquisition mode, SIM and IEC modes may be used as strategies to over-
come this problem, since they can increase the specificity and sensitivity, particularly in
target analysis (Tranchida et al., 2004; Petronilho, Coimbra, and Rocha, 2014; Martins,
Almeida, and Rocha, 2017). IEC allows the minimization of the contribution of co-eluted
peaks, contributing to increasing the target compound’s peak area. Thus, the combina-
tion of the data obtained through full-scan mode acquisition (which gives the food’s
global volatile profile) and the data achieved by IEC processing (the use of specific m/z
as diagnostic ions that highlights a particular chemical family) allows in-depth informa-
tion about a food’s volatile composition, particularly in the improvement of the target
compound’s data. For quantitative or semiquantitative purposes, the chromatographic
area is used to estimate the amount of each analyte. Raw data may be normalized versus
an internal (or external) standard, and analyte amount can be estimated through calibra-
tion curves or the standard addition method. Different types of internal standards may
be used for quantification purposes, depending on the applied methodologies and the
target compounds. The use of specific isotope-labeled internal standards is recommended
once they assure an equal response factor for the analytes; however, they can be quite
expensive. Accurate analyte identification is only confirmed through the co-injection of
authentic standards; however, they can be expensive or not commercially available. For
that reason, the use of the commercial/open-access mass spectra databases can be deci-
sive tools for compound putative identification (Dettmer, Aronov, and Hammock, 2007;
Milman, 2015). Confidence in the identification of the compound may be improved using
other strategies, namely through the use of retention indices (RI) values. Basically, a
n-alkanes series (typically ranging from C6 to C20) is injected in the same GC program
of the sample, and retention time of each n-alkane is recorded. An RI value of 100 times
the number of carbon is attributed to each n-alkane. Then, a compound’s retention time
is normalized using the retention times of the adjacent eluting n-alkanes, through the
van Den Dool and Kratz equation (van Den Dool and Dec. Kratz, 1963). The calcu-
lated RI can be compared with RIs available in the literature or open sources and that
were achieved with a similar GC column to the column used experimentally. This previ-
ous fact is possible because RIs are not dependent on several GC instrumental features
(e.g., carrier gas type and velocity, column’ diameter and length, film thickness, among
others), which allows for comparison with other analytical laboratories (Babushok,
­
2015; Grob and Barry, 2004). Although 1D-GC is widely used in the qualitative and
quantitative analysis of a wide range of foods, providing high-quality analytical data,
sometimes its complexity exceeds the separation capacity of a single chromatographic
column, producing overloaded chromatograms. In such cases, co-elution peaks might
occur, which makes compound identification and quantification difficult. Therefore, in
order to increase chromatographic resolution, independent techniques have been devel-
oped, namely comprehensive two-dimensional gas chromatography (GC×GC) that is a
powerful and high throughput solution (Mondello et al., 2008; Tranchida et al., 2004).

7.3.2 Comprehensive Two-Dimensional Gas Chromatography (GC×GC)

Multidimensional gas chromatography (MDGC) plays a key role in the determination of

a sample’s volatile profile, namely in the comprehensive mode (GC×GC). The separation
of chromatographic peaks in complex matrices (Marriott et al., 2012; Mondello et al.,
146 Food Aroma Evolution

FIGURE 7.3 Schematic illustration of a comprehensive two-dimensional gas chro-

matographic system coupled with mass spectrometry and a time-of-flight analyzer
(GC×GC-ToF-MS).

2008), as well as possible enantiomeric recognition (Cao et al., 2011), are important fea-
tures of MDGC.
In-depth and detailed characterization of a food’s volatile profile can be achieved by
GC×GC (Figure 7.3), comprising two orthogonal separation mechanisms that combine
the use of two GC columns, which are coated with specific stationary phases ruled by
different properties, namely volatility, chirality, or polarity. The column combination
most commonly used is a nonpolar 1D column (separation ruled by volatility, generally
30 m) and a polar 2D column (separation ruled by polarity, usually 1–2 m) (Cordero et
al., 2018). A specific interface, called a modulator, makes the connection between the
two GC columns. A modulator is responsible for the transfer of small portions of the elu-
ate (2–8 s) from the primary 1D column, and then for re-injecting it into the secondary
2 D column, preserving the integrity of the 1D separation, once each peak is modulated

several times. Different categories of modulators are available, namely valve-based, flow,
and thermal. The collection of the eluate is made by a loop or channel, either valve-based
or through flow modulation (often called the same); while in thermal modulation, the
eluate is trapped using temperature. This last modulator type is the most used, particu-
larly the cryomodulator, where a fast cooling of the eluate occurs using a cryogenic jet
(for instance with liquid nitrogen) followed by a fast heating applied using a hot-gas jet,
this allows the immobilization and then remobilization of the compounds (Mondello et
al., 2008; Marriott et al., 2012; Prebihalo et al., 2018). Narrow peaks are produced once
the collected fractions are no bigger than ¼ of the width peak; thus, they are “flash”
separated before the end of the modulation time. However, the wraparound phenomenon
might occur when the separation of an analyte exceeds the modulation time, and it is
not finished before the next modulation (Figure 7.4). This phenomenon may promote
co-elutions, interfering in the precise quantification of the compounds (Marriott, Massil,
and Hügel, 2004, 2012; Mondello et al., 2008), being necessary to formerly optimize the
modulation time to avoid wraparound.
The obtained GC×GC peaks are very narrow, with a typical width at half-height of
0.1 s or less, and several data points are needed to record these peaks, which imply higher
data acquisition rates (ca. 100 full mass-range spectra per second) that are only possible
with a time-of–flight mass spectrometer (ToF-MS) (Dallüge, Beens, and Brinkman, 2003;
Mondello et al., 2008). Furthermore, reliable spectra deconvolution of overlapped peaks
is possible due to full mass spectra acquisition and continuity (the same ion abundance
ratios are observed in all the point of the chromatographic peak for the different masses
in the spectrum) of the ToF-MS, even at trace levels (Górecki, Panić, and Oldridge, 2006;
 The Role of Gas Chromatography-Based Methodologies  147

FIGURE 7.4 Schematic representation of the wraparound phenomenon: the separa-

tion of an analyte exceeds the modulation time, and it is not finished before the next
modulation.

Grob and Barry, 2004). This data complexity can be handled through specific soft-
ware that allows GC×GC-ToF-MS data processing. One example is ChromaTOF® that
includes different algorithms (True Signal Deconvolution® and automated peak finding),
able to perform the acquisition, processing, and data report. This software also cre-
ates and allows the visualization of the GC×GC chromatograms, in a contour plot or a
three-dimensional plot (Dallüge, Beens, and Brinkman, 2003). Regarding analytes quan-
tification, the same strategies reported for 1D-GC analysis, could also be implemented
for GC×GC data. For identification purposes, the same criteria described previously for
1D-GC, namely the co-injection of standards (when it is possible); MS spectral similar-
ity, calculated RI and comparison with literature are other possibilities for identification.
Furthermore, GC×GC provides an extra powerful tool for analyte identification, namely
the formation of structured chromatograms. The orthogonal separation mechanism of
GC×GC (promoted by the combination of a nonpolar with a polar column) lets the two-
dimensional spatial distribution of the compounds be ruled by their chemical proper-
ties, that is, chemically related compounds are organized in the same two-dimensional
chromatographic space, determining a two-dimensional “chemical map.” This feature is
particularly helpful in the achievement of reliable identifications, especially in the analy-
sis of complex matrices like foods, and when chemical standards are absent; it can also
simplify the data analysis, and consequently decrease the analysis time (Cordero et al.,
2015; Marriott et al., 2012). Figure 7.5 shows an example of a structured chromatogram
that highlights one of the most important tools of GC×GC compared to 1D-GC. For
instance, hydrocarbons present a lower retention time for the second dimension (2tR) once
they are nonpolar compounds, contrary to organic acids that present higher polarity, and
therefore exhibit the highest 2tR.
In recent years, GC×GC-ToF-MS has been widely applied in food research
(Nolvachai, Kulsing, and Marriott, 2017; Dymerski, 2018). For instance, several stud-
ies have been performed to determine specific volatile components or chemical families
of different foods, namely beer (Martins et al., 2018), elderflowers (Salvador, Silvestre,
148 Food Aroma Evolution

FIGURE 7.5 Three-dimensional GC×GC-ToF-MS total ion chromatogram plot obtained

from a lager beer analysis. A structured chromatogram is shown though the bands and
clusters that are formed by structurally related compounds of each beer volatile chemical
family.

and Rocha, 2017), elderberries (Salvador et al., 2016), grape (Rocha et al., 2007), or
aromatic plants (Jalali et al., 2013, 2012; Petronilho et al., 2013, 2011). Figure 7.6
shows a practical example of the use of GC×GC-ToF-MS to access the evolution of
elderberries’ volatile composition (Salvador et al., 2016), namely through the volatile
profile of the Sambucus nigra variety during the ripening process (before and after a
ripe state). The clusters delineated on the contour plots (Figure 7.6) allow for elucidation
of the volatile profile of the chemical families (monoterpenic and sesquiterpenic com-
pounds, and norisoprenoids) for each type of elderberry (unripe and ripe), and reveals
the utility of the compound’s relative spatial position within the same chemical family.
The achieved GC×GC-ToF-MS contour plots are snapshots of the actual ripening stage
of the elderberry under study, and their visual analysis allows for observation of the
differences between the two stages of ripening. A visual chromatogram inspection of
Figure 7.6 reveals an increase in the diversity of detected chemical compounds during
the elderberries’ ripening, and a decrease of their content, particularly of monoterpe-
nic compounds. These differences in elderberry plots allow for the conclusion that the
analysis of chromatogram contour plots demonstrate the potentiality for rapid assess-
ment of the volatile varietal profiles of elderberries, which may have a further impact
on volatile composition of elderberry-based products, and eventually, on their aroma.
 The Role of Gas Chromatography-Based Methodologies  149

FIGURE 7.6 A blowup of a part of a GC×GC-ToF-MS chromatogram contour plot

obtained for a Sambucus nigra variety in IEC mode (m/z 93, 169, 204) for unripe and
ripe elderberry. The chromatographic spaces corresponding to monoterpenic compounds
(C10), norisoprenoids (C13), and sesquiterpenic compounds (C15) are highlighted.

The advantages of GC×GC application in the compound’s separation that have the
same volatility can be shown through the practical example shown in Figure 7.7a, in
which β-ocimene and 1,1,3,5-tetramethylcyclohexane present similar volatility (same
retention time for the first dimension—1t R : 528 s); therefore, co-eluting on the 1D col-
umn (Equity-5). Nevertheless, they can be separated by the 2D column (DB-FFAP),
within only around half of a second, once they present different polarities (2t R of 0.530
and 0.590 s, respectively). Also, the lower detection limits and spectral quality of
GC×GC-ToF-MS gives a huge advantage for the determination of trace compounds,
which remains to be one of the main challenges for food volatile composition. In fact,
the narrow peak (ca. 39 ms) achieved for β-ocimene in beer volatile determination
(Figure 7.7b) had an appropriate spectral quality of the mass spectrum (similarity value
926/1000) compared with a commercial database. Hence, β-ocimene can be putatively
identified based on the combination of several parameters: its retention times (1t R and
2t ), RI calculation, comparison with RIs, its mass spectrum pattern, and comparison
R
with commercial databases.
GC×GC-ToF-MS overcomes several drawbacks of the conventional 1D-GC, namely
shorter run times, enhanced resolution and peak capacity, lower detection limits, and
150 Food Aroma Evolution

FIGURE 7.7 (a) Blowup of a part of a GC×GC-ToF-MS total ion chromatogram contour
plot obtained from a lager beer analysis, illustrating the separation of (1) 1,1,3,5-tet-
ramethylcyclohexane, (2) β-ocimene, and (3) γ-terpinene; (b) β-ocimene, a beer trace
component, presenting a peak of 39 ms-wide, is easily identified with a very high mass
spectrum similarity (926 of 1000), based on the comparison with Wiley MS database.

higher mass sensitivity and selectivity as a result of the peak focusing in the modula-
tor (Seeley and Seeley, 2013; Tranchida et al., 2016; Mondello et al., 2008; Marriott,
Massil, and Hügel, 2004). Also, signal-to-noise ratio is enhanced for GC×GC, compared
to 1D-GC (Dallüge, Beens, and Brinkman, 2003). However, GC×GC has some disad-
vantages, namely the high costs related to the equipment acquisition, and its operation,
and maintenance. Also, the instrumentation is complex; thus, an expert technician is
required, the data generated are quite complex, and their alignment, integration, and
processing are a time-consuming task (Mondello et al., 2008).

7.3.3 Olfactometry (GC-O)

For many decades, the in-depth characterization of a food’s volatile compounds was
accomplished through extraction and gas chromatographic techniques, which revealed
the identification of more than 10,000 volatile compounds. Although only a few of them
 The Role of Gas Chromatography-Based Methodologies  151

FIGURE 7.8 Schematic illustration of a gas chromatography-olfactometry system (GC-O).

are aroma-active and affect the overall aroma profile of foods (Song and Liu, 2018). This
illustrates one of the main drawbacks of GC-MS-based techniques, that is, the chemical
structures of the volatile compounds present in foods may be elucidated; however, these
techniques are incapable of determining the odor properties and contribution to the food
samples (Chen and Ho, 2006; Song and Liu, 2018).
Gas chromatography-olfactometry (Figure 7.8) complements the aforementioned
drawback of MS-based techniques using the combination of compound separation
through the GC capillary column and the olfactometer detector (Zellner et al., 2008).
When coupled with analytical techniques, GC-O becomes a precise, descriptive approach
to characterize stimuli, evaluating and measuring impressions, which enables the compre-
hension and quantification of a sensorial characteristic (Zellner et al., 2008). GC-O was
invented in 1964 (Fuller, Steltenkamp, and Tisseiand, 1964) and has undergone several
improvements over the years. Even so, the concept was maintained in which the GC efflu-
ent is mixed with humid air or inert gas and sent to the olfactometer for sniffing (Figure
7.8). The GC system is equipped with a common detector, such as FID or MS, plus a
non-destructive thermal conductivity detection system with the outlet connected to a
sniffing port (Song and Liu, 2018). The identification of aroma-active compounds is con-
ducted by the same criteria previously reported for 1D-GC and GC×GC. Additionally,
the sensorial detector gives the aroma descriptor of the detected analyte, as well as its
intensity, which should be compared with the odor descriptions of reference compounds
(Song and Liu, 2018). GC-O data may be collected and processed through different olfac-
tometric methods: dilution, time-intensity, detection frequency, and posterior intensity
methods. The most common method is dilution analysis, in which the aroma extract is
successively diluted until there is no odor perception by panelists, and Charm (combined
hedonic aroma response method) or AEDA (aroma extraction dilution analysis) represent
the most common strategies for this (Zellner et al., 2008). More recently, Osme (time-
intensity method) has been used as a popular method for measuring the potency of flavor
components (Chen and Ho, 2006). Despite the huge utility of the GC-O analysis, the
implementation of this technique represents several challenges. The chemical complexity
of food flavor implies the occurrence of co-elutions that may occur both in nonpolar and
polar stationary phases, leading to the inaccurate identification of odor-active compounds
(Zellner et al., 2008). Key odor-active compounds may be present in the matrix at a trace
level, and the co-elution of compounds may easily occur making the correlation between
the chromatographic peaks and the perceived aroma difficult to assess (Giungato et al.,
2018). Theoretically, this drawback may be overcome by using multidimensional GC
combined with olfactometry (MDGC-O), being, for instance, already carried out, on a
152 Food Aroma Evolution

fresh puree of kiwi (Jordán et al., 2002) which allowed for the identification and report of
several components for the first time in this matrix. Nevertheless, GC×GC-O remains to
be a quite a challenging technique as the human breathing cycle is slower than the sepa-
ration, that is, the formation of very narrow chromatographic peaks (e.g., 100–400 ms)
during one modulation time period (Giungato et al., 2018). Another drawback of GC-O
systems is due to the fact that the chromatographic separation implies that volatile com-
ponents are assessed individually and their impact on the food versus the isolated compo-
nent is not linear (due to the masking and synergistic effects that might occur between the
volatile components), thus the overall perceived aroma cannot be fully elucidated (Hallier
et al., 2004). The human factor is another limitation of the GC-O system, as olfactory
capacity and odor thresholds may vary significantly both within and between people, and
also cases of specific anosmia may occur, that is, some people with an otherwise normal
sense of smell are unable to detect families of similar smelling compounds (Delahunty,
Eyres, and Dufour, 2006). Additionally, the lack of concentration, breathing cycle, health
status of the individual, and the natural variation of the olfactory response over the time
are factors of biased results (Plutowska and Wardencki, 2008; Delahunty, Eyres, and
Dufour, 2006). Thus, the number of assessors (panelists that will sniff at the olfactomet-
ric port) should be significant and appropriate, in order to guarantee the representative-
ness of sensory evaluation and to preserve the reliability of the results (Giungato et al.,
2018). GC-O appears as a powerful tool for investigations in the food industry, through
the detection and analysis of key aroma contributing components that are used in a wide
range of applications in flavor analysis (Giungato et al., 2018). For instance, Grosch
summarized the use of GC-O and the character-impact odorants identified in olive oil,
butter, cheese, meat, bread, beer, green tea, spices, and also the study of off-flavors from
several food items (Grosch, 1994). Other food items analyzed by GC-O involving either
Charm analysis or the AEDA techniques included fruit and fruit products, namely grape
juice (Baek et al., 1997), alcoholic beverages, such as wine (Campo et al., 2011), chocolate
and cocoa (Schnermann and Schieberle, 1997), roasted coffee beans and brewed coffee
(Semmelroch et al., 1995), dairy products (Curioni and Bosset, 2002), among numerous
other food and food products (Chen and Ho, 2006; Zellner et al., 2008).

7.4 CONCLUDING REMARKS AND FUTURE TRENDS

The aroma of a food can be studied through different techniques, choosing the most
suitable equipment is crucial for the achievement of reliable data due to the food’s com-
plexity and the analytes’ concentration (which may be in different orders of magnitude).
Nevertheless, the authors would like to reinforce the idea that food aroma is much more
than its volatile components, since most of them are not aroma-active, and may not con-
tribute to the overall aroma properties. Gas chromatography is extremely important for
the determination of a food’s volatile composition and for monitoring the food aroma
evolution, 1D-GC being the most inexpensive and widely used equipment. However, the
co-elution frequency in 1D-GC, mainly in the study of food samples, is high, which
may cause misidentifications and respective incorrect quantifications, particularly for
trace compounds. Therefore, MDGC techniques, for instance, GC×GC, assume a key
role in the study and reliable, in-depth characterization of the food’s volatile composi-
tion, particularly due to the presence of structured chromatograms. The instrumental
improvement of gas chromatographic techniques has led to long lists of volatiles that
may or may not have a sensory relevance to food. Olfactometry techniques, specifically
 The Role of Gas Chromatography-Based Methodologies  153

GC-O, have been challenged to enlighten and complement this avalanche of data by
understanding the role of odor in the volatile components present in foods. MDGC tech-
niques are essential: for the in-depth characterization of a food’s volatile composition; for
the detection of key odorants and elucidation of their formation; for the discrimination
of enantiomers and co-eluting compounds that can be trace elements with peculiar odor
characteristics (Cordero et al., 2018). Nevertheless, the relation between a food’s vola-
tile composition and flavor perception is still challenging, and requires other techniques
as a complement, such as olfactometry. In recent years, significant improvements have
occurred in GC systems, in order to increase resolution and sensitivity. The development
of new stationary phases of GC columns have been growing (e.g., ionic liquid station-
ary phases) (Prebihalo et al., 2018), as well as the development of simpler and effective
modulators (Prebihalo et al., 2018; Cordero et al., 2018). The huge amount of data gener-
ated by GC×GC-ToF-MS brings challenging problems, that can be overcome through the
­development of new, effective, and powerful GC×GC-ToF-MS data processing methods,
as well as new data analysis methods (Cordero et al., 2015, 2018; Prebihalo et al., 2018).
Moreover, the development of new mass spectra libraries is required (Dymerski, 2018). In
the near future, developments will occur on faster and miniaturized solutions (even por-
table devices), with reduced costs for the production and performance of the equipment
(Dymerski, 2018) that will allow real-time monitoring of food aroma evolution, and that
can be routinely used in labs. Several challenges still remain in food aroma analysis, such
as the development of faster and more powerful instrumental techniques, which will
access the flavor compounds with higher accuracy and sensitivity. Also, the development
of specific flavor-isolation techniques will allow more reliable and accurate identification
of specific groups of flavor compounds. Furthermore, suitable techniques that mimic the
interaction between humans and foods are still needed, which will provide knowledge
about which compounds have the greatest impact on how a food tastes. Altogether, when
analyzing a food product, it is crucial to plan and measure all the parameters that will
possibly have an impact on its odor, and the nature of the food product and the link of
sensory data should always be considered and required to fully understand the impact of
the analytical results.

ACKNOWLEDGMENTS

Thanks are due to FCT/MEC for the financial support to the QOPNA and LAQV-
REQUIMTE Research Unit (FCT UID/QUI/00062/2019), through national funds and
where applicable co-financed by the FEDER, within the PT2020 Partnership Agreement.

REFERENCES

Babushok, V. I. 2015. Chromatographic retention indices in identification of chemical

compounds. Trends in Analytical Chemistry 69: 98–104.
Baek, H. H., Cadwallader, K. R., Marroquin, E., and Silva, J. L. 1997. Identification of
predominant aroma compounds in Muscadine grape juice. Journal of Food Science
62 (2): 249–52.
Baldovini, N. 2017. Natural fragrant raw materials. In Handbook of Odor, edited by
Andrea Buettner, pp. 39–62. Cham, Switzerland: Springer International Publishing
AG.
154 Food Aroma Evolution

Campo, E., Malfeito-Ferreira, M., Loureiro, V., Cacho, J., and Ferreira, V. 2011.
Analytical and sensorial characterization of the aroma of wines produced with sour
rotten grapes using GC-O and GC-MS : Identification of key aroma compounds.
Journal of Agricultural and Food Chemistry 59: 2543–53.
Cao, G., Shan, Q., Li, X., Cong, X., Zhang, Y., Cai, H., and Cai, B. 2011. Analysis of
fresh Mentha haplocalyx volatile components by comprehensive two-dimensional
gas chromatography and high-resolution time-of-flight mass spectrometry. Analyst
136 (22): 4653–61.
Chen, Y. and Ho, C. 2006. Flavor analysis in food. In Encyclopedia of Analytical
Chemistry. Hoboken, NJ: Wiley.
Cordero, C., Kiefl, J., Schieberle, P., Reichenbach, S. E., and Bicchi, C. 2015.
Comprehensive two-dimensional gas chromatography and food sensory properties:
Potential and challenges. Analytical and Bioanalytical Chemistry 407 (1): 169–91.
Cordero, C., Schmarr, H. G., Reichenbach, S. E., and Bicchi, C. 2018. Current develop-
ments in analyzing food volatiles by multidimensional gas chromatographic tech-
niques. Journal of Agricultural and Food Chemistry 66 (10): 2226–36.
Curioni, P. M. G. and Bosset, J. O. 2002. Key odorants in various cheese types as
determined by gas chromatography-olfactometry. International Dairy Journal 12:
959–84.
Dallüge, J., Beens, J., and Brinkman, U. A. Th. 2003. Comprehensive two-dimen-
sional gas chromatography: A powerful and versatile analytical tool. Journal of
Chromatography A 1000 (1–2): 69–108.
Delahunty, C. M., Eyres, G., and Dufour, J. -P. 2006. Gas chromatography-olfactometry.
Journal of Separation Science 29: 2107–25.
Dettmer, K., Aronov, P. A., and Hammock, B. D. 2007. Mass spectrometry-based metab-
olomics. Mass Spectrometry Reviews 26: 51–78.
Dymerski, T. 2018. Two-dimensional gas chromatography coupled with mass spectrom-
etry in food analysis. Critical Reviews in Analytical Chemistry 48 (4): 252–78.
Fuller, G. H., Steltenkamp, R., and Tisseiand, G. A. 1964. The gas chromatograph with
human sensor: Perfumer model. Annals New York Academy of Sciences 116: 711–24.
Giungato, P., Gilio, A. D., Palmisani, J., Marzocca, A., Mazzone, A., Brattoli, M., Giua,
R., and Gennaro, G. de. 2018. Synergistic approaches for odor-active compounds
monitoring and identification: State of the art, integration, limits and potentialities
of analytical and sensorial techniques. Trends in Analytical Chemistry 107: 116–29.
Górecki, T., Panić, O., and Oldridge, N. 2006. Recent advances in comprehensive two-
dimensional gas chromatography (GC×GC). Journal of Liquid Chromatography &
Related Technologies 29 (7–8): 1077–104.
Grob, R. L. and Barry, E. F., eds. 2004. Modern Practice of Gas Chromatography, 4th
ed. NJ, United States of America: John Wiley & Sons, Inc.
Grosch, W. 1994. Determination of potent odourants in foods by Aroma Extract Dilution
Analysis (AEDA) and calculation of Odour Activity Values (OAVs). Flavour and
Fragrance Journal 9: 147–58.
Hallier, A., Courcoux, P., Thierry, S., and Prost, C. 2004. New gas chromatography–olfac-
tometric investigative method, and its application to cooked Silurus glanis (European
catfish) odor characterization. Journal of Chromatography A 1056: 201–8.
Jalali, H. T., Petronilho, S., José, J., Coimbra, M. A., Domingues, M. R. M., Ebrahimian,
Z. J., Silvestre, A. J. D., and Rocha, S. M. 2012. Deeper insight into the monoterpe-
nic composition of Ferula gummosa oleo-gum-resin from Iran. Industrial Crops &
Products 36 (1): 500–7.
 The Role of Gas Chromatography-Based Methodologies  155

Jalali, H. T., Petronilho, S., Villaverde, J. J., Coimbra, M. A., Domingues, M. R. M.,
Ebrahimian, Z. J., Silvestre, A. J. D., and Rocha, S. M. 2013. Assessment of the
sesquiterpenic profile of Ferula gummosa oleo-gum-resin (galbanum) from Iran.
Contributes to its valuation as a potential source of sesquiterpenic compounds.
Industrial Crops and Products 44: 185–91.
Jordán, M. J., Margaría, C. A., Shaw, P. E., and Goodner, K. L. 2002. Aroma active
components in aqueous kiwi fruit essence and kiwi fruit puree by GC-MS and
multidimensional GC/GC-O. Journal of Agricultural and Food Chemistry 50 (19):
5386–90.
Marriott, P. J., Chin, S. T., Maikhunthod, B., Schmarr, H. G., and Bieri, S. 2012.
Multidimensional gas chromatography. Trends in Analytical Chemistry 34:
1–20.
Marriott, P. J., Massil, T., and Hügel, H. 2004. Molecular structure retention relation-
ships in comprehensive two-dimensional gas chromatography. Journal of Separation
Science 27 (15–16): 1273–84.
Martins, C., Almeida, A., and Rocha, S. M. 2017. Recent advances and challenges
for beer volatile characterization based on gas chromatographic techniques.
In Recent Advances in Analytical Techniques, edited by Rida Ahmed Atta-ur-
Rahman and Sibel A. Ozkan, pp. 141–99. Sharjah, U. A. E.: Bentham Science
Publishers.
Martins, C., Brandão, T., Almeida, A., and Rocha, S. M. 2015. Insights on beer volatile
profile: Optimization of solid-phase microextraction procedure taking advantage
of the comprehensive two-dimensional gas chromatography structured separation.
Journal of Separation Science 38 (12): 2140–8.
Martins, C., Brandão, T., Almeida, A., and Rocha, S. M. 2018. Unveiling the lager beer
volatile terpenic compounds. Food Research International 114: 199–207.
Milman, B. L. 2015. General principles of identification by mass spectrometry. Trends in
Analytical Chemistry 69: 24–33.
Mondello, L., Tranchida, P., Dugo, P., and Dugo, G. 2008. Comprehensive two-dimen-
sional gas chromatography-mass spectrometry: A review. Mass Spectrometry
Reviews 27: 101–24.
Nolvachai, Y., Kulsing, C., and Marriott, P. J. 2017. Multidimensional gas chromatogra-
phy in food analysis. Trends in Analytical Chemistry 96: 124–37.
Nongonierma, A., Voilley, A., Cayot, P., Quéré, J. L., and Springett, M. 2007. Mechanisms
of extraction of aroma compounds from foods, using adsorbents. Effect of various
parameters. Food Reviews International 9129: 51–94.
Perestrelo, R., Barros, A. S., Câmara, J. S., and Rocha, S. M. 2011. In-depth search
focused on furans, lactones, volatile phenols, and acetals as potential age markers of
Madeira wines by comprehensive two-dimensional gas chromatography with time-
of-flight mass spectrometry combined with solid phase microextraction. Journal of
Agricultural and Food Chemistry 59 (7): 3186–204.
Perestrelo, R., Petronilho, S., Câmara, J. S., and Rocha, S. M. 2010. Comprehensive
two-dimensional gas chromatography with time-of-flight mass spectrometry com-
bined with solid phase microextraction as a powerful tool for quantification of
ethyl carbamate in fortified wines. The case study of Madeira wine. Journal of
Chromatography A 1217 (20): 3441–5.
Petronilho, S., Coimbra, M. A., and Rocha, S. M. 2014. A critical review on extraction
techniques and gas chromatography based determination of grapevine derived ses-
quiterpenes. Analytica Chimica Acta 846: 8–35.
156 Food Aroma Evolution

Petronilho, S., Maraschin, M., Delgadillo, I., Coimbra, M. A., and Rocha, S. M. 2011.
Sesquiterpenic composition of the inflorescences of Brazilian chamomile (Matricaria
recutita L.): Impact of the agricultural practices. Industrial Crops and Products 34
(3): 1482–90.
Petronilho, S., Rocha, S. M., Ramírez-Chávez, E., Molina-Torres, J., and Rios-Chavez, P.
2013. Assessment of the terpenic profile of Callistemon citrinus (Curtis) Skeels from
Mexico. Industrial Crops and Products 46: 369–79.
Plutowska, B. and Wardencki, W. 2008. Application of gas chromatography–olfactom-
etry (GC–O) in analysis and quality assessment of alcoholic beverages—A review.
Food Chemistry 107: 449–63.
Prebihalo, S. E., Berrier, K. L., Freye, C. E., Bahaghighat, H. D., Moore, N. R., Pinkerton,
D. K., and Synovec, R. E. 2018. Multidimensional gas chromatography: Advances
in instrumentation, chemometrics, and applications. Analytical Chemistry 90:
505–32.
Rocha, S. M., Coelho, E., Zrostlíková, J., Delgadillo, I., and Coimbra, M. A. 2007.
Comprehensive two-dimensional gas chromatography with time-of-flight mass
spectrometry of monoterpenoids as a powerful tool for grape origin traceability.
Journal of Chromatography A 1161 (1–2): 292–9.
Rocha, S. M., Coutinho, P., Coelho, E., Barros, A. S., Delgadillo, I., and Coimbra, M.
A. 2010. Relationships between the varietal volatile composition of the musts and
white wine aroma quality. A four year feasibility study. LWT—Food Science and
Technology 43 (10): 1508–16. Elsevier.
Rocha, S. M., Freitas, R., Cardoso, P., Santos, M., Martins, R., and Figueira, E. 2013.
Exploring the potentialities of comprehensive two-dimensional gas chromatog-
raphy coupled to time of flight mass spectrometry to distinguish bivalve species:
Comparison of two clam species (Venerupis decussata and Venerupis philippina-
rum). Journal of Chromatography A 1315: 152–61.
Rodrigues, A., Caldana, C., Jorge, T. F., Schmidt, R., Dongen, J. T. V., Thomas-Oates,
J., and Anto, C. 2016. Mass spectrometry-based plant metabolomics: Metabolite
responses to abiotic stress. Mass Spectrometry Reviews 35 (5): 620–49.
Salvador, Â. C., Rudnitskaya, A., Silvestre, A. J. D., and Rocha, S. M. 2016. Metabolomic-
based strategy for fingerprinting of Sambucus nigra L. berry volatile terpenoids and
norisoprenoids: Influence of ripening and cultivar. Journal of Agricultural and Food
Chemistry 64 (26): 5428–38.
Salvador, Â. C., Silvestre, A. J. D., and Rocha, S. M. 2017. Unveiling elderflowers
(Sambucus nigra L.) volatile terpenic and norisoprenoids profile: Effects of different
postharvest conditions. Food Chemistry 229: 276–85.
Santos, M. C., Nunes, C., Rocha, M. A. M., Rodrigues, A., Rocha, S. M., Saraiva, J.
A., and Coimbra, M. A. 2013. Impact of high pressure treatments on the physi-
cochemical properties of a sulphur dioxide-free white wine during bottle storage:
Evidence for Maillard reaction acceleration. Innovative Food Science & Emerging
Technologies 20: 51–8.
Santos, M. C., Nunes, C., Rocha, M. A. M., Rodrigues, A., Rocha, S. M., Saraiva, J. A.,
and Coimbra, M. A. 2015. High pressure treatments accelerate changes in volatile
composition of sulphur dioxide-free wine during bottle storage. Food Chemistry
188: 406–14.
Schieberle, P. and Molyneux, R. J. 2012. Quantitation of sensory-active and bioactive
constituents of food: A Journal of Agricultural and Food Chemistry perspective.
Journal of Agricultural and Food Chemistry 60 (10): 2404–8.
 The Role of Gas Chromatography-Based Methodologies  157

Schnermann, P. and Schieberle, P. 1997. Evaluation of key odorants in milk chocolate and
cocoa mass by Aroma Extract Dilution Analyses. Journal of Agricultural and Food
Chemistry 45: 867–72.
Seeley, J. V. and Seeley, S. K. 2013. Multidimensional gas chromatography: Fundamental
advances and new applications. Analytical Chemistry 85 (2): 557–78.
Semmelroch, P., Laskawy, G., Blank, I., and Grosch, W. 1995. Determination of potent
odourants in roasted coffee by stable isotope dilution assays. Flavour and Fragrance
Journal 10: 1–7.
Silva, I., Coimbra, M. A., Barros, A. S., Marriott, P. J., and Rocha, S. M. 2015. Can
volatile organic compounds be markers of sea salt? Food Chemistry 169: 102–13.
Elsevier.
Silva, I., Rocha, S. M., Coimbra, M. A., and Marriott, P. J. 2010. Headspace solid-phase
microextraction combined with comprehensive two-dimensional gas chromatogra-
phy time-of-flight mass spectrometry for the determination of volatile compounds
from marine salt. Journal of Chromatography A 1217 (34): 5511–21.
Song, H. and Liu, J. 2018. GC-O-MS technique and its applications in food flavor analy-
sis. Food Research International 114: 187–98. Elsevier.
Tranchida, P. Q., Dugo, P., Dugo, G., and Mondello, L. 2004. Comprehensive two-
dimensional chromatography in food analysis. Journal of Chromatography A 1054
(1–2): 3–16.
Tranchida, P. Q., Purcaro, G., Maimone, M., and Mondello, L. 2016. Impact of com-
prehensive two-dimensional gas chromatography with mass spectrometry on food
analysis. Journal of Separation Science 39 (1): 149–61.
van Den Dool, H. and Dec. Kratz, P. 1963. A generalization of the retention index system
including linear temperature programmed gas–liquid partition chromatography.
Journal of Chromatography A 11: 463–71.
Wong, J. W., Hayward, D. G., and Zhang, K. 2013. Gas chromatography-mass spec-
trometry techniques for multiresidue pesticide analysis in agricultural commodities.
In Comprehensive Analytical Chemistry, edited by Imma Ferrer and E. Michael
Thurman, 1st ed., Vol. 61, pp. 3–22. Oxford: Elsevier B.V.
Zellner, A., Dugo, P., Dugo, G., and Mondello, L. 2008. Gas chromatography–olfactom-
etry in food flavour analysis. Journal of Chromatography A 1186: 123–43.
 Chapter 8
Monitoring Food Aroma during
Processing and Storage by Rapid
Analytical Methods: A Focus
on Electronic Noses and Mass
Spectrometry-Based Systems
Aoife Power, Vi Khanh Truong, James
Chapman, and Daniel Cozzolino

CONTENTS

8.1 Introduction 159

8.2 Sensors 160
8.3 Data Integration and Analysis 163
8.4 Examples and Applications in Food Systems 163
8.4.1 Electronic Noses and Mass Spectrometry-Based Systems 163
8.4.2 Electronic Tongues 168
8.5 Conclusion 169
References 169

8.1 INTRODUCTION

Flavor perception is influenced by a multitude of sensory modalities which are in turn the
product of a complex combination of chemical compounds in the food matrix. For exam-
ple, in wine and other alcoholic beverages, it is their aroma (or smell), that contributes
significantly to their taste, similar trends can also be observed in other foods such as meat
and cheese. Consequently, the flavor and/or taste of a product is ultimately determined by
the presence and concentration of chemical volatile compounds (CVC) (Polášková et al.,
2008). Many of the analytical methods used to study aroma in foods typically involve the
preparation of an extract (or the collection of volatiles with traps) followed by chromato-
graphic separation and detection (Linforth, 2000; Taylor and Linforth, 2000; Kress-
Rogers and Brimelow, 2001; Polášková et al., 2008; Ebeler and Thorngate, 2009; Ross,
2009; Lesschaeve and Noble, 2010; Heaven and Nash, 2012; Sáenz-Navajas et al., 2012).
However, other aspects of the food process involved in the perception of aroma and taste,
such as the temporal dimension of the eating/drinking process, which has an influence

159
160 Food Aroma Evolution

on the release and transport of chemical compounds to the olfactory epithelium, are
not considered in such analyses, although the temporal dimension is a central feature of
the eating and/or drinking process (Linforth, 2000; Kress-Rogers and Brimelow, 2001;
Polášková et al., 2008; Ebeler and Thorngate, 2009; Ross, 2009; Lesschaeve and Noble,
2010; Heaven and Nash, 2012; Sáenz-Navajas et al., 2012). Typically, human senses do
not react to the absolute intensity of a stimulus, but they do respond to the rate of change
of that particular stimulus (Linforth, 2000; Kress-Rogers and Brimelow, 2001; Polášková
et al., 2008; Ebeler and Thorngate, 2009; Ross, 2009; Lesschaeve and Noble, 2010;
Heaven and Nash, 2012; Sáenz-Navajas et al., 2012). Therefore, the temporal dimension
associated with the sensorial response should be measured using methods that can relate
to time and the intensity of such compounds in the food. This approach has also led to the
development of mathematical models to describe how perception is affected by temporal
changes in breath aroma concentration (Linforth, 2000; Kress-Rogers and Brimelow,
2001; Polášková et al., 2008; Ebeler and Thorngate, 2009; Ross, 2009; Lesschaeve and
Noble, 2010; Heaven and Nash, 2012; Sáenz-Navajas et al., 2012).

8.2 SENSORS

When food is stored, it releases specific volatile compounds, such as hydrocarbons, ethanol,
aldehydes, acids, ethers, and esters (Gallo and Ferranti, 2016; Ghasemi-Varnamkhasti and
Lozano, 2016; Matindoust et al., 2016). The presence and concentration of these volatile
compounds are different and depend on other factors like the raw material, brand and type
of food, storage conditions (e.g., temperature, humidity), and issues associated with spoil-
age (e.g., presence of microorganisms) (Gallo and Ferranti, 2016; Ghasemi-Varnamkhasti
and Lozano, 2016; Matindoust et al., 2016). These volatile compounds have been proposed
as being like fingerprints of the factors affecting food during storage and processing (Gallo
and Ferranti, 2016; Ghasemi-Varnamkhasti and Lozano, 2016; Matindoust et al., 2016).
The ideal flavor or aroma analysis tool would be able to monitor changes in the tem-
poral dimension, and thus make objective measurements related to perception (Linforth,
2000; Polášková et al., 2008; Lesschaeve and Noble, 2010). To achieve this in a timely
manner and at low cost, sensitive analytical systems which have a selectivity and sen-
sitivity comparable with the human olfactory receptor are required (Linforth, 2000;
Polášková et al., 2008; Lesschaeve and Noble, 2010). Several limitations in the use of
existing sensors are evident; for example, while electroencephalography (EEG) and mag-
netic resonance imaging (MRI) can both be used to monitor a participants brain activity
directly when they are encountering a taste or aroma, the instrumentation itself is not
practical from an industrial point of view (e.g., excessive cost, lack of portability, too
time-consuming) (Linforth, 2000; Polášková et al., 2008; Lesschaeve and Noble, 2010).
Alternatively, the use of purely instrumental techniques to follow changes in breath
volatile concentration during eating or drinking, relating to the observed patterns associ-
ated with sensory perception, could be implemented by the food industry; the best rep-
resentation of such modern tools are the so-called electronic noses (ENs) (Moseley and
Norris, 1991; Baek et al., 1999; Hines et al., 1999; Hurst, 1999; Citterio and Suzuki,
2008; Lesschaeve and Noble, 2010; Brattoli et al., 2011; Croissant et al., 2011; Li-Chan
et al., 2011). The EN employs arrays of sensors (in much the same way as the olfactory
epithelium has arrays of receptors) which can be used to monitor volatile compounds
(Hurst, 1999; Linforth, 2000; Kress-Rogers and Brimelow, 2001; Polášková et al., 2008;
Ebeler and Thorngate, 2009; Ross, 2009; Lesschaeve and Noble, 2010; Heaven and Nash,
 Processing and Storage by Rapid Analytical Methods 161

2012; Sáenz-Navajas et al., 2012). All the sensors have a certain specificity, such that one
sensor responds more to esters than aldehydes, whereas the opposite might be true for
another type of sensor. To date, however, no sensor array can respond in a manner com-
parable with that of the human olfactory system. Moreover, they lack in both sensitivity
(ppm/ppb concentration) and in some cases selectivity. Coupled with their comparatively
slow response rate, it makes existing instrumentation incapable of appropriately monitor-
ing rapid changes in breath volatile concentration (Mielle, 1996; Lindinger and Jordan,
1998; Hines et al., 1999; Hurst, 1999; Wilson et al., 2001; Pearce et al., 2006; Citterio
and Suzuki, 2008; Röck et al., 2008; Brattoli et al., 2011; Heaven and Nash, 2012).
Several studies have reported the application of ENs for quality control in several
foods (e.g., grapes, wine, saffron, medicinal plants). However, a limited number of reports
can be found in the literature on the application of the EN technology for the online or
real-time monitoring of aroma during food processing and storage.
Electronic noses can also be based on conducting organic polymers (COP) (Suppes et
al., 2008; Kukla et al., 2009; Sanaeifar et al., 2017). These are a set of active mechanisms
which detect odors and convert chemical vapors into electrical signals. Conducting polymer
characteristics depend on doping levels, ion size of the dopant, water content, and proton-
ation levels. These sensors may be classified into a mode of transduction and application
(Suppes et al., 2008; Kukla et al., 2009; Sanaeifar et al., 2017). The other group of EN
instruments available on the market are those based on a metal oxide semiconductor, which
is a form of gas sensor that can detect the early signs of meat spoilage (Sanaeifar et al., 2017).
It has been demonstrated that the ideal instrumental method must be objective, cost
effective, and provide rapid, reproducible results, with continuous operation. However,
to date, instrumental methods for sensory analysis lack the ability to reliably perceive all
of these key sensory attributes, and depending on the food matrix being analyzed current
methods have been inconsistent in their predictive relationships between sensory and
instrumental measurements (Martens, 1999).
Real-time quantitative detection of volatiles can be achieved using an electron impact
source with a membrane separator between the source and the external environment coupled
with a mass spectrometer (Linforth, 2000; Kress-Rogers and Brimelow, 2001). The advan-
tages of the mass spectrometer (relative to the electronic nose) are its fast response rate and
greater sensitivity (Suppes et al., 2008; Kukla et al., 2009; Sanaeifar et al., 2017). However,
often the separator membrane reduces the technique’s overall sensitivity, while its selective
permeability reduces its application potential. In recent years, the further development of mass
spectrometer systems, such as atmospheric pressure chemical ionization–mass spectrometry
(APCI-MS) and proton transfer reaction–mass spectrometry are envisioned to be capable of
the real-time detection of compounds at ppb concentrations (Hurst, 1999; Linforth, 2000;
Kress-Rogers and Brimelow, 2001; Martı ́ et al., 2005; Sanaeifar et al., 2017).
As the demands of public health and consumer safety authorities increase, the devel-
opment of rapid screening techniques to determine the quality characteristics of foods
and beverages has become of great interest to the modern food industry. Ideally, these
techniques should be relatively inexpensive, easy to operate, and require little or no sam-
ple preparation allowing them to be used in-line or at-line. Electronic noses, electronic
tongues, and optical methods often based on vibrational spectroscopy have all demon-
strated some potential to characterize complex food or beverage samples rather than rely-
ing solely on sensory analysis using human subjects (Bartlett et al., 1997; Hurst, 1999;
Guadarrama et al., 2001; Deisingh et al., 2004; Gutiérrez et al., 2007; Röck et al., 2008).
Thus, an automatic real-time aroma tracking system is needed to complement existing
analytical systems (Sanaeifar et al., 2017).
162 Food Aroma Evolution

As reported by other authors, ENs are instruments designed to detect and identify
chemical volatile compounds released by the food. As described before, an EN uses an
array of semi-selective gas sensors which are subjected to the volatile components of
food products to generate specific fingerprints (aroma responses) (Ampuero and Bosset,
2003; Luykx and Van Ruth, 2008; Wilson and Baietto, 2009; Sanaeifar et al., 2017).
The fingerprints of conditions in the headspace of a food sample are utilized to construct
an aroma dataset (Cynkar et al., 2010; Sanaeifar et al., 2017). Gas chromatography,
mass spectrometry, and the combined technique gas chromatography–mass spectrom-
etry allow both qualitative and quantitative analyses to be performed; accordingly, these
techniques have been widely used to analyze the volatile compounds released by food.
However, these methods require complex and costly sample pre-treatments and are not
suitable for the online monitoring of volatile compounds (Gilar et al., 2003; Phan et al.,
2012; Castro-Puyana et al., 2017; Kuuliala et al., 2018). Rapid developments in materials
and electronic techniques have allowed for the creation of electronic nose systems based
on electrochemical reactions. Such systems allow cheap online measurement to be made
using small sample volumes and have therefore been used widely to measure volatile
compounds released from food. However, electronic sensors have short useful lifetimes
due to surface fouling and hysteresis and suffer from cross-sensitivity in their practical
application (Loutfi et al., 2015; Verma and Yadava, 2015; Gu et al., 2017; Kodogiannis,
2017; Sanaeifar et al., 2017; Wojnowski et al., 2017; Kuuliala et al., 2018; Majchrzak et
al., 2018). Table 8.1. summarizes the advantages and disadvantages of different types of
electronic noses used by the food industry (Sanaeifar et al., 2017).
Other sensors based on vibrational spectroscopy, infrared base sensors (near and
mid infrared), have shown promise as rapid methods for the real-time monitoring of
aroma and flavor in different food matrices. Consequently, infrared spectroscopy (IR)
has become one of the most attractive and used methods of analysis in recent years, as
it provides simultaneous, rapid, and non-destructive quantitation of the major chemical
components of many agriculture-related products and plant materials (Su et al., 2017).
Recently, new applications involving the determination of other minor compounds in

TABLE 8.1 Advantages and Disadvantages of Commonly Used Electronic Noses

Conducting
Metal Oxide Polymer Electrochemical Mass Spectrometry
Advantages Sensitive to a Sensitive to a Robust High sensitivity
wide range of wide range of instrumentation and stability,
chemical chemical ideal for on-site
compounds, fast compounds, fast analysis, quality
recovery, low recovery, diverse control, enables
cost, and high range of quantitative and
reproducibility polymer qualitative
coatings analysis
Disadvantages Sensors tend to High sensor drift Low sensitivity, High cost, large
drift, affected by and susceptible no adequate for size instruments
humidity (sensor to humidity foods with high (however, some
saturation) and/or high concentration of portable
moisture aromatic instrument
hydrocarbons in become available)
the matrix
Source: Adapted from Sanaeifar et al., 2017.
 Processing and Storage by Rapid Analytical Methods 163

plant materials have also been reported (Kačuráková and Wilson, 2001; Cozzolino, 2014;
Gordon et al., 2018; Roberts et al., 2018; Bureau et al., 2019).
It has been reported that most volatile compounds released by foodstuffs have spe-
cific infrared absorption characteristics (Kačuráková and Wilson, 2001; Cozzolino,
2014; Gordon et al., 2018; Roberts et al., 2018; Bureau et al., 2019). Volatile compounds
released by food can be analyzed qualitatively and quantitatively using these specific
infrared absorption peaks. In recent decades this has resulted in multiple research groups
studying infrared methods for detecting volatile compounds released by food during stor-
age, maturity, and spoilage processes (Kačuráková and Wilson, 2001; Cozzolino, 2014;
Gordon et al., 2018; Roberts et al., 2018; Bureau et al., 2019).
The use of mid infrared spectroscopy (MIR) can also be performed using an open-
path mode. Open-path MIR has been used extensively in environmental monitoring to
quantify the components of gas clouds from distances of several meters to kilometers (La
Spina et al., 2013; Akagi et al., 2014; Kira et al., 2016). The development of the focal
plane detector technique has allowed open-path MIR to be used for imaging detection
and to determine the spatial distributions of gas clouds directly. Hence, wide-area and
remote detection of volatile components released from food is possible with the use of
open-path MIR (Jiao et al., 2018).
Colorimetric sensor arrays use chemical reactions between sensitive dyes and volatile
compounds and have been used to monitor food spoilage. Although these methods allow
volatile compounds to be visualized, the methods have problems associated with cross-
sensitivity and environmental pollution (Huang et al., 2014; Morsy et al., 2016; Sachdev
et al., 2016; Schaude et al., 2017; Domínguez-Aragón et al., 2018).

8.3 DATA INTEGRATION AND ANALYSIS

During the application of electronic noses and tongues, large datasets are generated.
Therefore, in order to obtain the relevant information from the data generated by such
instruments, different techniques based on multivariate data analysis (e.g., principal com-
ponents analysis; partial least squares) and pattern recognition techniques (e.g., discrimi-
nant and cluster analysis) are used to detect and identify specific trends in the chemistry
or chemical changes of the food under analysis. The application of these data analysis
methods allows for the quantitative (e.g., calibration models) and qualitative analysis (e.g.,
cluster analysis, patterns) of the data generated. Specific details about multivariate data
analysis methods and techniques, including pattern recognition methods, can be found
elsewhere (Adams, 1995; Brereton, 2000, 2008; Esbensen, 2002; Gishen et al., 2005;
Geladi and Kowalski, 1986; Geladi, 2003; Granato et al., 2014; Martens and Martens,
2001; Naes et al., 2002). Overall, the examples presented in this chapter highlight the
need to combine chemometric and rapid analytical techniques to develop appropriate
applications. Figure 8.1 shows the integration of sensors and multivariate data analysis
methods to monitor and control the aroma of foods during processing and storage.

8.4 EXAMPLES AND APPLICATIONS IN FOOD SYSTEMS

8.4.1 Electronic Noses and Mass Spectrometry-Based Systems

Andrés and collaborators (2002) reported on the use of a solid-phase microextraction

(SPME) system coupled to a new direct-extraction device (DED) to monitor volatile
164 Food Aroma Evolution

FIGURE 8.1 Schematic representation of the use of electronic noses and chemometric
tools to control and monitor food processes.

compounds during dry-cured ham ripening. The authors indicated that a DED allows for
the utilization of the SPME fiber in the core of the sample with no damage to the fiber,
enabling extraction of volatiles from the solid sample while avoiding sample handling
(Andrés et al., 2002). According to the authors, major groups of volatile compounds
extracted with SPME-DED agreed with the available scientific literature on the volatiles
present in dry-cured ham, such as 3-methylbutanal or hexanal. In addition, observed
changes in the profile of those volatile compounds throughout processing followed a
typical pattern for the formation of volatile compounds (Andrés et al., 2002). Pionnier
and collaborators (2004) monitored the kinetics of aroma compounds released during
cheese consumption using an online atmospheric pressure ionization–mass spectrometry
(API MS) system and an off-line solid-phase micro extraction-gas chromatography–mass
spectrometry (SPME-GC-MS) instrument. The authors reported that ethyl hexanoate,
heptan-2-one, and heptan-2-ol were the main volatile compounds. However, only the
concentrations of ethyl hexanoate and heptan-2-one could be determined by the API-MS
system (Pionnier et al., 2004).
The combination of instrumental methods and sensory analysis is an excellent tool
for the determination of the duration of the storage, the quality, and the origin of foods
(Barié et al., 2015). Barié and collaborators (2015) reported on the use of polymer-coated
surface acoustic wave (SAW) sensor arrays combined with SPME microextraction as
a technique for a highly sensitive and selective organic gas detection system (Barié et
al., 2015). These authors reported that discrimination between apple varieties, ripe and
unripe pineapple, and finally the detection of off-flavor compounds was possible using
the proposed system. The main advantages of the described system included its short
analysis time (e.g., sample preparation, extraction, and measuring times) and low cost
(Barié et al., 2015). Since the device is highly portable, a broad range of applications
might be possible, including the online measurement of chemical and biochemical pro-
cesses and detection of microorganism contamination (Barié et al., 2015).
The rapid, objective monitoring of the ripening process of fermented food products
is of great interest to the food industry. The application of an EN instrument to monitor
this process was reported by Trihaas and Nielse (2005). In this study, headspace samples
from two types of Danish blue cheese (traditional cheese and cheese from pasteurized
milk) were analyzed (Trihaas and Nielsen, 2005). Data from the response of the chemical
 Processing and Storage by Rapid Analytical Methods 165

sensors were used to model the changes that occurred during shelf life (5, 8, 12, 20, and
33 weeks after brining) using multivariate analysis methods (Trihaas and Nielsen, 2005).
The authors pre-processed the signal generated by the EN instrument using multiplica-
tive scatter correction (MSC) (Trihaas and Nielsen, 2005). Differences, reflected in the
cheese smell, were greater at early ripening stages, as determined by the EN system used
(Trihaas and Nielsen, 2005).
Rega and co-workers (2009) reported the use of dynamic SPME to monitor the
release of volatile compounds generated during the baking process of cereal products
(sponge cake model) by directly sampling its baking vapors. The steam generated during
the process was analyzed using the dynamic SPME system, allowing for the monitoring
of several volatile compounds with very different volatility and hydrophobicity properties
(e.g., 5-hydroxymethylfurfural and 2-methyl-propanal) (Rega et al., 2009).
The capability of a sensor array EN to discriminate between different milk sam-
ples by sensing their aroma was reported by Brambilla and collaborators (2010). The
authors analyzed milk samples sourced from different production batches, brands, and
processing systems (ultra-high temperature [UHT], partly skimmed, and commercially
available). Principal component analysis (PCA) was used to put samples into different
groups according to their odor profile. Moreover, the analysis of the olfactory finger-
prints showed that 2 hours after the opening of the packaging, the flavor of anomalous
samples evolved in a different way from that of the normal ones (Brambilla et al., 2010).
Linear discriminant analysis (LDA) was also used to analyze the data generated by the
EN system, where more than 98% of the samples were correctly classified (Brambilla
et al., 2010).
Volatile compounds in wines arise from many sources, such as from grapes during
fermentation, yeast during fermentation, and/or during post-fermentation, storage, and
processing (Callejón et al., 2012). Factors influencing the extraction of volatiles from
grapes during fermentation have not been widely studied; an improved understanding of
the processes and mechanisms involved in the formation or release of wine aroma com-
pounds from grapes during fermentation could help winemakers to optimize or control
wine composition and aroma (Callejón et al., 2012). These authors evaluated the effects
of different skin contact times on changes in concentrations of volatile aroma compounds
during the fermentation of Cabernet Sauvignon grapes using a SPME system coupled to
GC-MS (Callejón et al., 2012). Results reported by these authors indicated that the dura-
tion and timing of the skin contact during fermentation showed a measurable effect on
volatile composition (Callejón et al., 2012).
Volatile compounds are important factors that affect the quality and flavor of dried
medicinal and aromatic plant products such as mint (Mentha spicata L.) leaves (Kiani
et al., 2018). Work by Kiani et al. (2018) evaluated the capability of an EN system as an
alternative tool for monitoring the dynamic changes of the CVC of mint leaves during
the hot air-drying process. The monitoring system includes a gas sensor array based on a
metal oxide semiconductor (MOS) sensor system (Kiani et al., 2018). During the drying
process, aroma parameters, humidity, and temperature of the outlet air, as well as sample
weight, were acquired at given dryer settings. These authors reported that PCA analysis
showed that the fingerprints of CVC for mint leaves are significantly different due to
the drying conditions. In addition, partial least squares (PLS), multiple linear regression
(MLR), and principal component regression (PCR) were applied to correlate the aroma
parameters to the mint leaves’ moisture content as well as to determine the end-point of
the drying process (Kiani et al., 2018). The reported models had high correlation coef-
ficients in cross-validation (R>0.90) with an acceptable error margin (root mean square
166 Food Aroma Evolution

standard error of prediction, RMSEP=5.79). The overall results demonstrated the effec-
tiveness and feasibility of monitoring mint leaf drying conditions using an electronic nose
system (Kiani et al., 2018).
Brazinha and collaborators (2010) experimentally demonstrated the use of mass
spectrometry (MS) for the online, quantitative monitoring of organophilic pervaporation
processes operated under variable operational parameters (different temperature param-
eters, condensation/downstream pressure) (Brazinha et al., 2010). The authors reported
that, due to its high sensitivity, mass spectrometry is particularly suitable for the online
monitoring of aromas in dilute streams under reduced pressure, which is similar to how
natural aromas permeate (Brazinha et al., 2010). The authors also demonstrated how
mass spectrometry is suitable for the online monitoring of solvent and co-solvent per-
meants (water and ethanol in the selected case study), enabling the technique for online
determination of selectivity of the solutes of interest (Brazinha et al., 2010). Moreover,
the MS system has proven to be a powerful tool for studying the fractionation of aro-
mas, recovered by integrated pervaporation–condensation processes, allowing the online
detection and monitoring of each solute vapor in the permeate stream. Additionally, the
described technique allowed for accurate transient studies as it was capable of acquiring
data points every 12–5 s in real time (Brazinha et al., 2010).
Aprea and co-workers (2006) reported the use of PTR-MS to characterize olive
oil headspace without concentration of the volatiles or pre-treatment of the samples.
Comparison of extra virgin and defective (rancid) samples, as described by a panel of
sensory judges, and the monitoring of thermo-oxidation processes were discussed (Aprea
et al., 2006). Overall, PTR-MS was able to monitor rancidity in olive oil samples (Aprea
et al., 2006).
Coffees from different origins were roasted to different roast degrees and vary-
ing times using different temperature roasting profiles (Gloess et al., 2014). The for-
mation of CVC during roasting was analyzed online using a PTR-ToF-MS instrument.
These authors analyzed Coffea arabica from Colombia, Guatemala (Antigua La Ceiba),
Ethiopia (Yirga Cheffe, Djimmah), and Coffea canephora var. robusta from Indonesia
(Malangsari); the roasting profiles ranged from high temperature short time (HTST) to
low temperature long time (LTLT) roasting (e.g., from medium to dark roast) (Gloess
et al., 2014). The dynamic release of the online monitored CVC differed for the different
coffees and showed a strong relation with the combination of time/temperature roasting
profile (Gloess et al., 2014). The authors reported that for Guatemalan coffee samples,
the formation of CVC started relatively early in the roasting process, while the CVC
formation started much later in the case of Yirga Cheffe and Malangsari (Gloess et al.,
2014). Differences were associated with the interactions between time and temperature
of roasting, as well as the degree of roasting, and were influenced by both the intensity
of volatiles and different chemical composition derived from the different coffee varieties
and origins analyzed (Gloess et al., 2014).
The hot water extraction process used to make an espresso coffee is affected by many
factors, and the proper understanding of how these factors impact the profile of the final
cup are important for the industry (Sanchez-Lopez et al., 2016). Sanchez-Lopez et al.
(2016) investigated the effect of water temperature and pressure on the extraction kinet-
ics of CVC in coffee using online monitoring of volatiles directly from the coffee flow
using PTR-ToF-MS. The application of hierarchical cluster analysis (HCA) to the data
allowed the grouping of the identified compounds into five families according to their
time-intensity profiles (Sanchez-Lopez et al., 2016). The volatile compounds that were
grouped into each family had similar physicochemical properties with polarity being
 Processing and Storage by Rapid Analytical Methods 167

determined as one of the main forces driving the CVC extraction kinetics; the effect
of temperature on extractability was more pronounced for the less polar compounds
(Sanchez-Lopez et al., 2016). It was observed by the authors that an increase in tempera-
ture produced a significant increase in the extraction of CVC, especially during the final
stages of the extraction (Sachez Lopez et al., 2016).
Gloess et al. (2014, 2018) used ion mobility spectrometry–mass spectrometry
(IMS-MS) with corona discharge ionization for the online analysis of coffee roasting.
The authors reported that this was the first time that the formation of CVC during coffee
roasting was monitored not only in a positive but also in a negative ion mode, and not
only with a MS system, but also with IMS (Gloess et al., 2018). The temporal evolution
of more than 150 CVC was monitored during the roasting of Brazilian Coffea arabica.
The use of IMS-MS allowed for the separation of isobaric and isomeric compounds. In a
positive ion mode, isomers of alkyl pyrazines were found to exhibit distinct time-intensity
profiles during roasting, providing a unique insight into the complex chemistry of this
important class of aroma active compounds (Gloess et al., 2018). A negative ion mode
gave access to species that were poorly detectable by other online methods, such as acids.
In this study, the release of fatty acids during coffee roasting was investigated in detail.
The author documents how the concentration of the fatty acids increases early in the
roasting process followed by a decrease at the same time as other CVC start to be formed
(Gloess et al., 2018).
Wei and co-workers (2019) analyzed a wide variety of volatile constituents in vari-
ous flaxseed oils (FSO) prepared from the roasting of the flaxseeds at different tempera-
tures using SPME coupled with GC-MS. The authors identified more than 60 volatile
compounds that were correlated with sensory analysis. The results indicated that all of
the FSO samples had similar intensities for “oily” and “herbaceous” aroma descriptors,
where “roasted” and “almond” aromas were statistically different (p < 0.05). The use of
PCA showed that the first principal component (PC1) and the second principal compo-
nent (PC2) explained 84% and 9%, respectively, of the variability associated with the
volatile compounds present in the set of samples analyzed and prepared from different
roasting conditions (Wei et al., 2019).
Mouret and collaborators (2014) investigated the production of major volatile com-
pounds associated with the predominant higher alcohols and esters formed during alco-
holic wine fermentation and monitored it using an online GC system. The accuracy and
frequency of the GC system measurements made it possible to calculate kinetic param-
eters, rates, and specific rates of production (Mouret et al., 2014). Results from this study
indicated that esters (substantial proportions of which are lost in the off-gas, rather than
of the total production (liquid content + losses) can lead to misinterpretation of the yeast
metabolism (Mouret et al., 2014). The authors also highlighted that the specific produc-
tion rate of individual higher alcohols reached their maximum values before the exhaus-
tion of the corresponding precursor amino acids (Mouret et al., 2014). Isobutanol and
isoamyl alcohol were formed from carbon metabolites and nitrogen metabolites and were
consequently produced continuously throughout the fermentation process. On the other
hand, propanol synthesis was strongly correlated with the presence of assimilable nitro-
gen, during both the growth and stationary phases (Mouret et al., 2014). Acetate ester
concentrations correlated linearly with the concentrations of the corresponding higher
alcohols, indicating that the availability of the precursors is the main limiting factor for
producing these esters. The authors theorized that these results opened up the possibility
for innovative approaches based on metabolic flux analysis taking such dynamics into
account (Mouret et al., 2014).
168 Food Aroma Evolution

FIGURE 8.2 The use of chemometric (principal component analysis) to monitor changes
during processing.

Xu and collaborators (2019) monitored changes in the volatile compounds of germi-

nated chickpea, lentil, and yellow pea flour samples over the course of a 6-day germina-
tion using SPME-GC-MS/Olfactometry. The authors identified more than 100 volatile
components involving 19 odor-active components utilizing GC-O. Both PCA and HCA
methods revealed that lentil and yellow pea flour samples had similar aromatic profiles,
while the decrease of bean flavor compounds along with the occurrence of unpleasant fla-
vors was detected in chickpea flours upon germination (Xu et al., 2019). Six bean flavor
markers, including hexanal, (E,E)-2,4-nonadienal, (E,E)-2,4-decadienal, 3-methyl-1-bu-
tanol, 1-hexanol, and 2-pentyl-furan, were employed to quantify bean flavor formation
in the flours over the course of germination (Xu et al., 2019).
The integration of sensors with multivariate data analysis or chemometrics to moni-
tor a given process is illustrated in Figure 8.2. Changes during the process can be easily
detected and/or monitored by the application of techniques such as PCA analysis.

8.4.2 Electronic Tongues

The perception of aroma during oral processing is a complex interplay of many factors,
partly related to the food matrix as well as to the oral processing conditions (Benjamin et
al., 2012). The role of the tongue in transporting the bolus through the mouth by pressing
it against the palate has been widely studied; however, the relationship between tongue
pressures generated and CVC release is not clear (Benjamin et al., 2012). Pressure pat-
terns during swallowing were found to be unique across subjects which may suggest per-
sonal flavor perception (Benjamin et al., 2012). These authors described an experiment
for the release of volatile compounds in vitro during the process of eating, using a model
mouth capable of reproducing actual human tongue pressure patterns with a computer-
controlled artificial tongue driven by an actuator. In this experiment, tongue functional-
ity was monitored by pressure and force sensors (Benjamin et al., 2012). The system was
designed to incorporate oral features and conditions such as temperature, saliva flow,
gas flow, and appropriate oral cavity volumes attached to an online CVC measurement
system using proton transfer reaction–mass spectrometry (Benjamin et al., 2012). The
authors concluded that the development of an innovative model mouth has a potential
industrial application as a product development evaluation tool for food and pharmaceu-
tical companies. Overall, the proposed system can provide fast and accurate feedback on
 Processing and Storage by Rapid Analytical Methods 169

the product’s flavor release and its physical properties under oral conditions (Benjamin et
al., 2012). The use of electronic tongues to monitor food spoilage, enzymatic activity, and
the production of toxic compounds derived from the storage of foods has been reviewed
by Ghasemi-Varnamkhasti and collaborators (2018).

8.5 CONCLUSION

The potential savings, in time and cost of analysis coupled with the environmentally
friendly nature of these technologies, has positioned the so-called electronic nose and
tongue systems as attractive techniques in the field of the quality analysis of foods (e.g.,
storage, process).
Relatively inexpensive ENs such as a metal oxide sensor are commercially available
and have been widely used by the food industry. However, volatile interference with the
sensors determines the use of sample pre-treatment steps, which compromises the speed
and the simplicity of the technique. However, a mass spectrometry-based electronic nose
allows for the collection of large amounts of information from a sample in a very short
time without the need for chromatographic separation or sample pre-processing, reduc-
ing the duration of the analysis. These technologies offer an exciting prospect of poten-
tially providing the means for the development of small scale, cheap, portable hand-held
instruments, which would be of great benefit to the whole food supply chain.
The combination of sensors and chemometrics is a relatively new approach in
the analysis of the aroma and taste of foods, and it can be used as a rapid technique
for monitoring changes during processing and storage. However, data pre-treatment
and care in the use of chemometric methods are required due to expected collinearity
issues among variables, differences in signal and possible instrument drift and base-
line shifts. Although these technologies are still relatively new, they will potentially
become more readily available to smaller companies and businesses, as tools to detect,
monitor, and prevent food spoilage and to monitor different steps during the produc-
tion of foods.
Despite multiple publications in the scientific literature regarding developments in
ENs, there is a clear gap between the robust real-world application of this technology
and the developments in both research and academia. This is a consequence of various
roadblocks that still hinder the growth and uptake of these applications, such as the hesi-
tancy of the industry to accept the integration of chemistry and mathematics (the benefits
of chemometrics are often ignored by those who prefer to employ classical statistics), the
lack of formal (academic) education in the use and application of instrumental methods
as high throughput tools, or the implementation of a holistic approach to complex sys-
tems analysis.

REFERENCES

Adams, M. J. (1995). “Chemometrics in analytical spectroscopy.” In RSC Spectroscopy

Monographs. Edited by N. W. Barnett. The Royal Society of Chemistry, UK. 216 p.
Akagi, S. K., I. R. Burling, A. Mendoza, T. J. Johnson, M. Cameron, D. W. Griffith, C.
Paton-Walsh, D. R. Weise, J. Reardon, R. J. Yokelson (2014). “Field measurements
of trace gases emitted by prescribed fires in southeastern US pine forests using an
open-path FTIR system.” Atmospheric Chemistry and Physics 14(1): 199–215.
170 Food Aroma Evolution

Ampuero, S. and J. Bosset (2003). “The electronic nose applied to dairy products: A
review.” Sensors and Actuators B: Chemical 94(1): 1–12.
Andrés, A., R. Cava, and J. Ruiz (2002). “Monitoring volatile compounds during dry-
cured ham ripening by solid-phase microextraction coupled to a new direct-extrac-
tion device.” Journal of Chromatography A 963(1–2): 83–8.
Aprea, E., F. Biasioli, G. Sani, C. Cantini, T. D. Märk, and F. Gasperi (2006). “Proton
transfer reaction–mass spectrometry (PTR-MS) headspace analysis for rapid detec-
tion of oxidative alteration of olive oil.” Journal of Agricultural and Food Chemistry
54(20): 7635–40.
Baek, I., R. S. Linforth, A. Blake, and A. Taylor (1999). “Sensory perception is related
to the rate of change of volatile concentration in-nose during eating of model gels.”
Chemical Senses 24(2): 155–60.
Barié, N., M. Bücking, U. Stahl, and M. Rap (2015). “Detection of coffee flavour age-
ing by solid-phase microextraction/surface acoustic wave sensor array technique
(SPME/SAW).” Food Chemistry 176: 212–8.
Bartlett, P. N., J. M. Elliott, and J. W. Gardner (1997). “Electronic noses and their appli-
cation in the food industry.” Food Technology 51: 44–8.
Benjamin, O., P. Silcock, J. Beauchamp, A. Buettner, and D. W. Everett (2012). “Tongue
pressure and oral conditions affect volatile release from liquid systems in a model
mouth.” Journal of Agricultural and Food Chemistry 60(39): 9918–27.
Brambilla, M., P. Navarotto, and V. la Sicurezza Alimentare (2010). “Application of
E-NOSE technology for ultra-high temperature processed partly skimmed milk pro-
duction batches monitoring.” Chemical Engineering Transactions 23: 171–6.
Brattoli, M., G. De Gennaro, V. De Pinto, A. Demarinis Loiotile, S. Lovascio, and M.
Penza (2011). “Odour detection methods: Olfactometry and chemical sensors.”
Sensors (Basel) 11(5): 5290–322.
Brazinha, C., A. Fonseca, O. Teodoro, and J. G. Crespo (2010). “On-line and real-
time monitoring of organophilic pervaporation by mass spectrometry.” Journal of
Membrane Science 347(1–2): 83–92.
Brereton, R. G. (2000). “Introduction to multivariate calibration in analytical chemis-
try.” The Analyst 125: 2125–54.
Brereton, R. G. (2008). Applied Chemometrics for Scientist. John Wiley & Sons,
Chichester, UK.
Bureau, S., D. Cozzolino, and C. J. Clark (2019). “Contributions of Fourier-transform
mid infrared (FT-MIR) spectroscopy to the study of fruit and vegetables: A review.”
Postharvest Biology Technology 148: 1–14.
Callejón, R. M., B. Margulies, G. D. Hirson, and S. E. Ebeler (2012). “Dynamic changes
in volatile compounds during fermentation of Cabernet Sauvignon grapes with and
without skins.” American Journal of Enology and Viticulture 63(3): 301–12.
Castro-Puyana, M., R. Pérez-Míguez, L. Montero, and M. Herrero (2017). “Reprint of:
Application of mass spectrometry-based metabolomics approaches for food safety,
quality and traceability.” TrAC Trends in Analytical Chemistry 96: 62–78.
Citterio, D. and K. Suzuki (2008). “Smart taste sensors.” Analytical Chemistry 80(11):
3965.
Cozzolino, D. (2014). “An overview of the use of infrared spectroscopy and chemometrics
in authenticity and traceability of cereals.” Food Research International 60: 262–5.
Croissant, A., D. Watson, and M. Drake (2011). “Application of sensory and instrumen-
tal volatile analyses to dairy products.” Annual Review of Food Science Technology
2: 395–421.
 Processing and Storage by Rapid Analytical Methods 171

Cynkar, W., R. Dambergs, P. Smith, and D. Cozzolino (2010). “Classification of

Tempranillo wines according to geographic origin: Combination of mass spectrom-
etry based electronic nose and chemometrics.” Analytica Chimica Acta 660(1–2):
227–31.
Deisingh, A. K., D. C. Stone, and M. Thompson (2004). “Applications of electronic noses
and tongues in food analysis.” International Journal of Food Science Technology
39(6): 587–604.
Domínguez-Aragón, A., J. A. Olmedo-Martínez, and E. A. Zaragoza-Contreras (2018).
“Colorimetric sensor based on a poly (ortho-phenylenediamine-co-aniline) copo-
lymer for the monitoring of tilapia (Orechromis niloticus) freshness.” Sensors &
Actuators B: Chemical 259: 170–6.
Ebeler, S. E. and J. H. Thorngate (2009). “Wine chemistry and flavor: Looking into the
crystal glass.” Journal of Agricultural Food Chemistry 57(18): 8098–108.
Esbensen, K. H. (2002). Multivariate Data Analysis in Practice. CAMO Process AS,
Oslo, Norway.
Gallo, M. and P. Ferranti (2016). “The evolution of analytical chemistry methods in
foodomics.” Journal of Chromatography A 1428: 3–15.
Geladi, P. (2003). “Chemometrics in spectroscopy. Part I. Classical chemometrics.”
Spectrochimica Acta Part B 58: 767–82.
Geladi, P. and B. R. Kowalski (1986). “Partial least-squares regression: A tutorial.”
Analytica Chimica Acta 185: 1–15.
Ghasemi-Varnamkhasti, M., C. Apetrei, L. Lozano, and A. Anyogu (2018). “Potential
use of electronic noses, electronic tongues and biosensors as multisensor systems for
spoilage examination in foods.” Trends in Food Science & Technology 80: 71–92.
Ghasemi-Varnamkhasti, M. and J. Lozano (2016). “Electronic nose as an innovative
measurement system for the quality assurance and control of bakery products: A
review.” Engineering in Agriculture, Environment Food 9(4): 365–74.
Gilar, M., K. J. Fountain, Y. Budman, J. L. Holyoke, H. Davoudi, and J. C. Gebler
(2003). “Characterization of therapeutic oligonucleotides using liquid chromatog-
raphy with on-line mass spectrometry detection.” Oligonucleotides 13(4): 229–43.
Gishen, M., R. G. Dambergs, and D. Cozzolino (2005). “Grape and wine analysis—
Enhancing the power of spectroscopy with chemometrics. A review of some appli-
cations in the Australian wine industry.” Australian Journal of Grape and Wine
Research 11: 296–305.
Gloess, Alexia N., C. Yeretzian, R. Knochenmuss, and M. Groessl (2018). “On-line anal-
ysis of coffee roasting with ion mobility spectrometry–mass spectrometry (IMS–
MS).” International Journal of Mass Spectroscopy 424: 49–57.
Gloess, Alexia N., A. Vietri, F. Wieland, S. Smrke, B. Schönbächler, J. A. López, S.
Petrozzi, S. Bongers, T. Koziorowski, and C. Yeretzian (2014). “Evidence of differ-
ent flavour formation dynamics by roasting coffee from different origins: On-line
analysis with PTR-ToF-MS.” International Journal of Mass Spectroscopy 365–66:
324–37.
Gordon, R., J. Chapman, A. Power, S. Chandra, J. Roberts, and D. Cozzolino (2018).
“Unfrazzled by fizziness: Identification of beers using attenuated total reflectance
mid-infrared spectroscopy and multivariate analysis.” Food Analytical Methods 11:
1–8.
Granato, D., V. M. A. Calado, and B. Jarvis (2014). “Observations on the use of statis-
tical methods in food science and technology.” Food Research International 55:
137–59.
172 Food Aroma Evolution

Gu, X., Y. Sun, K. Tu, and L. Pan (2017). “Evaluation of lipid oxidation of Chinese-style
sausage during processing and storage based on electronic nose.” Meat Science 133:
1–9.
Guadarrama, A., J. Fernandez, M. Iniguez, J. Souto, and J. De Saja (2001). “Discrimination
of wine aroma using an array of conducting polymer sensors in conjunction with
solid-phase micro-extraction (SPME) technique.” Sensors & Actuators B: Chemical
77(1–2): 401–8.
Gutiérrez, A., J. Burgos, C. Garcerá, M. Zarzo, M. Ruiz, and E. Moltó (2007).
“Optimization of an aroma sensor for assessing grape quality for wine making.”
Spanish Journal of Agricultural Research 5(2): 157–63.
Heaven, M. W. and D. Nash (2012). “Recent analyses using solid phase microextraction
in industries related to food made into or from liquids.” Food Control 27(1): 214–27.
Hines, E., E. Llobet, and J. Gardner (1999). “Electronic noses: A review of signal pro-
cessing techniques.” IEE Proceedings—Circuits, Devices Systems 146(6): 297–310.
Huang, X., X. Zou, J. Zhao, J. Shi, X. Zhang, Z. Li, and L. Shen (2014). “Sensing the
quality parameters of Chinese traditional Yao-meat by using a colorimetric sensor
combined with genetic algorithm partial least squares regression.” Meat Science
98(2): 203–10.
Hurst, W. J. (1999). “Electronic noses and sensor array based systems: Design and appli-
cations.” Proceedings of the 5th International Symposium on Olfaction and the
Electronic Nose, Baltimore, MD, September 27–30, 1998. Technomic Publishing
Company.
Jiao, L., Y. Guo, J. Chen, X. Zhao, and D. Dong (2018). “Detecting volatile compounds
in food by open-path Fourier transform infrared spectroscopy.” Food Research
International 119: 968–73.
Kačuráková, M. and R. Wilson (2001). “Developments in mid-infrared FT-IR spectros-
copy of selected carbohydrates.” Carbohydrate Polymers 44(4): 291–303.
Kiani, S., Saeid M., and M. Ghasemi-Varnamkhasti (2018). “Real-time aroma monitor-
ing of mint (Mentha spicata L.) leaves during the drying process using electronic
nose system.” Measurement 124: 447–52.
Kira, O., R. Linker, and Y. Dubowski (2016). “Detection and quantification of water-
based aerosols using active open-path FTIR.” Scientific Reports 6: 25110.
Kodogiannis, V. S. (2017). “Application of an electronic nose coupled with fuzzy-wavelet
network for the detection of meat spoilage.” Food Bioprocess Technology 10(4):
730–49.
Kress-Rogers, E. and C. J. Brimelow (2001). Instrumentation and Sensors for the Food
Industry. Woodhead Publishing.
Kukla, A., A. Pavluchenko, Y. M. Shirshov, N. Konoshchuk, and O. Y. Posudievsky
(2009). “Application of sensor arrays based on thin films of conducting polymers
for chemical recognition of volatile organic solvents.” Sensors and Actuators B:
Chemical 135(2): 541–51.
Kuuliala, L., Y. Al Hage, A.-G. Ioannidis, M. Sader, F.-M. Kerckhof, M. Vanderroost,
N. Boon, B. De Baets, B. De Meulenaer, and P. Ragaert (2018). “Microbiological,
chemical and sensory spoilage analysis of raw Atlantic cod (Gadus morhua) stored
under modified atmospheres.” Food Microbiology 70: 232–44.
La Spina, A., M. Burton, R. Harig, F. Mure, P. Rusch, M. Jordan, and T. Caltabiano
(2013). “New insights into volcanic processes at Stromboli from Cerberus, a remote-
controlled open-path FTIR scanner system.” Journal of Volcanology Geothermal
Research 249: 66–76.
 Processing and Storage by Rapid Analytical Methods 173

Lesschaeve, I. and A. Noble (2010). “Sensory analysis of wine.” In Managing Wine

Quality: Viticulture and Wine Quality. Elsevier, Woodhead Publishing, New Delhi,
India. 189–217 p.
Li-Chan, E., J. Chalmers, and P. Griffiths (2011). Applications of Vibrational Spectroscopy
in Food Science. John Wiley & Sons.
Lindinger, W. and A. Jordan (1998). “Proton-transfer-reaction mass spectrometry (PTR–
MS): On-line monitoring of volatile organic compounds at pptv levels.” Chemical
Society Reviews 27(5): 347–75.
Linforth, R. S. T. (2000). “Developments in instrumental techniques for food flavour
evaluation: Future prospects.” Journal of the Science of Food Agriculture 80(14):
2044–8.
Loutfi, A., S. Coradeschi, G. K. Mani, P. Shankar, and J. B. B. Rayappan (2015).
“Electronic noses for food quality: A review.” Journal of Food Engineering 144:
103–11.
Luykx, D. M. and S. M. Van Ruth (2008). “An overview of analytical methods for deter-
mining the geographical origin of food products.” Food Chemistry 107(2): 897–911.
Majchrzak, T., W. Wojnowski, T. Dymerski, J. Gębicki, and J. Namieśnik (2018).
“Electronic noses in classification and quality control of edible oils: A review.” Food
Chemistry 246: 192–201.
Martens, M. (1999). “A philosophy for sensory science.” Food Quality and Preference
10(4–5): 233–44.
Martens, H. and M. Martens (2001). Multivariate Analysis of Quality. An Introduction.
John Wiley & Sons, Chichester, UK.
Martı ́, M. P., O. Busto, J. Guasch, and R. Boqué (2005). “Electronic noses in the qual-
ity control of alcoholic beverages.” TrAC Trends in Analytical Chemistry 24(1):
57–66.
Matindoust, S., M. Baghaei-Nejad, Z. Zou, and L.-R. Zheng (2016). “Food quality and
safety monitoring using gas sensor array in intelligent packaging.” Sensor Review
36(2): 169–83.
Mielle, P. (1996). “‘Electronic noses’: Towards the objective instrumental characteriza-
tion of food aroma.” Trends in Food Science Technology 7(12): 432–38.
Morsy, M. K., K. Zór, N. Kostesha, T. S. Alstrøm, A. Heiskanen, H. El-Tanahi, A.
Sharoba, D. Papkovsky, J. Larsen, and H. Khalaf (2016). “Development and valida-
tion of a colorimetric sensor array for fish spoilage monitoring.” Food Control 60:
346–52.
Moseley, P. T. and J. O. Norris (1991). Techniques and Mechanisms in Gas Sensing.
Taylor & Francis.
Mouret, J. R., M. Perez, M. Angenieux, P. Nicolle, V. Fairnes, and J. M. Sablayrolles
(2014). “Online-based kinetic analysis of higher alcohol and ester synthesis during
winemaking fermentations.” Food and Bioprocess Technology 7(5): 1235–45.
Naes, T., T. Isaksson, T. Fearn, and T. Davies (2002). A User-friendly Guide to
Multivariate Calibration and Classification. NIR Publications, Chichester, UK. 420
p.
Pearce, T. C., S. S. Schiffman, H. T. Nagle, and J. W. Gardner (2006). Handbook of
Machine Olfaction: Electronic Nose Technology. Wiley-VCH Verlag GmbH & Co.
KGaA.
Phan, N.-T., K.-H. Kim, E.-C. Jeon, U.-H. Kim, J. R. Sohn, and S. K. Pandey (2012).
“Analysis of volatile organic compounds released during food decaying processes.”
Environmental Monitoring Assessment 184(3): 1683–92.
174 Food Aroma Evolution

Pionnier, E., C. Chabanet, L. Mioche, J.-L. Le Quéré, and C. Salles (2004). “1. In vivo
aroma release during eating of a model cheese: Relationships with oral parameters.”
Journal of Agricultural and Food Chemistry 52(3): 557–64.
Polášková, P., J. Herszage, and S. E. Ebeler (2008). “Wine flavor: Chemistry in a glass.”
Chemical Society Reviews 37(11): 2478–89.
Rega, B., A. Guerard, J. Delarue, M. Maire, and P. Giampaoli (2009). “On-line dynamic
HS-SPME for monitoring endogenous aroma compounds released during the bak-
ing of a model cake.” Food Chemistry 112(1): 9–17.
Roberts, J. J., A. Power, J. Chapman, S. Chandra, and D. Cozzolino (2018). “Vibrational
spectroscopy methods for agro-food product analysis.” Comprehensive Analytical
Chemistry 80: 51–68. Elsevier.
Röck, F., N. Barsan, and U. Weimar (2008). “Electronic nose: Current status and future
trends.” Chemical Reviews 108(2): 705–25.
Ross, C. F. (2009). “Sensory science at the human–machine interface.” Trends in Food
Science Technology 20(2): 63–72.
Sachdev, D., V. Kumar, P. H. Maheshwari, R. Pasricha, and N. Baghel (2016). “Silver
based nanomaterial, as a selective colorimetric sensor for visual detection of post
harvest spoilage in onion.” Sensors & Actuators B: Chemical 228: 471–9.
Sachez Lopez, Jose A., M. Wellinger, Alexia N. Gloess, Ralf Zimmermann, and Chahan
Yeretzian (2016). “Extraction kinetics of coffee aroma compounds using a semi-
automatic machine: On-line analysis by PTR-ToF-MS.” International Journal of
Mass Spectroscopy 401: 22–30.
Sáenz-Navajas, M.-P., P. Fernández-Zurbano, and V. Ferreira (2012). “Contribution of
nonvolatile composition to wine flavor.” Food Reviews International 28(4): 389–411.
Sanaeifar, A., H. ZakiDizaji, A. Jafari, and M. de la Guardia (2017). “Early detection of
contamination and defect in foodstuffs by electronic nose: A review.” TrAC Trends
in Analytical Chemistry 97: 257–71.
Schaude, C., C. Meindl, E. Fröhlich, J. Attard, and G. J. Mohr (2017). “Developing a
sensor layer for the optical detection of amines during food spoilage.” Talanta 170:
481–7.
Su, W.-H., H.-J. He, and D.-W. Sun (2017). “Non-destructive and rapid evaluation of
staple foods quality by using spectroscopic techniques: A review.” Critical Reviews
in Food Science Nutrition 57(5): 1039–51.
Suppes, G. M., B. A. Deore, and M. S. Freund (2008). “Porous conducting polymer/
heteropolyoxometalate hybrid material for electrochemical supercapacitor applica-
tions.” Langmuir 24(3): 1064–9.
Taylor, A. J. and R. S. Linforth (2000). “Techniques for measuring volatile release in vivo
during consumption of food.” ACS Symposium Series, American Chemical Society,
Washington, DC, 1999.
Trihaas, J. and P. V. Nielsen (2005). “Electronic nose technology in quality assessment:
Monitoring the ripening process of Danish blue cheese.” Journal of Food Science
70(1): E44–9.
Verma, P. and R. Yadava (2015). “Polymer selection for SAW sensor array based elec-
tronic noses by fuzzy c-means clustering of partition coefficients: Model studies
on detection of freshness and spoilage of milk and fish.” Sensors & Actuators B:
Chemical 209: 751–69.
Wei, C., Q. Zhou, B. Han, Z. Chen, and W. Liu (2019). “Changes occurring in the vola-
tile constituents of flaxseed oils (FSOs) prepared with diverse roasting conditions.”
European Journal of Lipid Science and Technology 121(1): 1–12.
 Processing and Storage by Rapid Analytical Methods 175

Wilson, A. and M. Baietto (2009). “Applications and advances in electronic-nose tech-

nologies.” Sensors 9(7): 5099–148.
Wilson, D. M., S. Hoyt, J. Janata, K. Booksh, and L. Obando (2001). “Chemical sensors
for portable, handheld field instruments.” IEEE Sensors Journal 1(4): 256–74.
Wojnowski, W., T. Majchrzak, T. Dymerski, J. Gębicki, and J. Namieśnik (2017).
“Electronic noses: Powerful tools in meat quality assessment.” Meat Science 131:
119–31.
Xu, M., Z. Jin, Y. Lan, J. Rao, and B. Chen (2019). “HS-SPME-GC–MS/olfactometry
combined with chemometrics to assess the impact of germination on flavor attri-
butes of chickpea, lentil, and yellow pea flours.” Food Chemistry 280: 83–95.
 Chapter 9
Hyphenated Electronic Nose
Technique for Aroma Analysis
of Foods and Beverages
Adriana Marcia Graboski, Sandra Cristina Ballen,
Juliana Steffens, and Clarice Steffens

CONTENTS

9.1 Introduction 177

9.2 Electronic Nose 178
9.2.1 Sampling System 179
9.2.2 Detection System (Sensor Array) 180
9.2.2.1 Gas Sensor Response Mechanism 182
9.2.3 Signal Processing System and Standard Recognition Methods 183
9.3 Electronic Nose Application 183
9.4 Conclusion and Future Perspectives 186
Conflict of Interest 187
Acknowledgment 187
References 187

9.1 INTRODUCTION

Human olfaction is still the main instrument for evaluating the aroma of many products,
as well as being used for identifying the deterioration of a wide range of foods (Ghasemi-
Varnamkhasti et al., 2018). Aroma is closely linked to the acceptance and quality of
food. The flavor of a food is strongly influenced by its aroma. The chemical sensation
caused by the taste of a food is due to the presence of the odors of small molecules that
are sufficiently volatile so as to reach the sensory receptors during eating. There a layer
of mucus that dissolves the molecules as soon as they reach these receptors; the molecules
are perceived by the nose’s olfactory epithelium, which is then recognized by the brain as
food characteristic (Firestein, 2001; Plutowska and Wardencki, 2007; Santonico et al.,
2008). The aromas are used to characterize, improve, standardize, and/or reconstitute
the aroma/flavor of the food, mask undesirable flavors and aromas, and improve sensory
quality (Fani, 2015).
Different aromas can be classified as either natural or synthetic. Natural flavors are
obtained by physical, microbiological, or enzymatic methods from natural raw materi-
als. They include essential oils, extracts (liquid and dry), balsams, and isolated natural

177
178 Food Aroma Evolution

flavoring substances. However, synthetic flavors are those obtained by chemical processes
and are comprised of flavors identical to natural and artificial flavors. There are also
mixtures of flavors, reaction/processing flavors, and smoke flavors (Agência Nacional de
Vigilância Sanitária [ANVISA], 2007).
Food flavors are the result of a balance between the concentrations of flavor com-
pounds, which have different chemical natures, in the vapor phase. A distortion of the
aroma profile may occur according to the modification of the food composition by favor-
ing or hindering the release of certain volatile compounds (Seuvre et al., 2007).
To evaluate the aroma quality in a food, sensory analysis, which uses human olfac-
tion, is one of the most commonly used techniques. However, there are many limitations
such as that the panel of testers is subjective and suffers incoherence and unpredictabil-
ity due to individual variability, and decreased sensitivity due to prolonged exposure,
fatigue, and variable mental states, and that it is time-consuming and limited to non-toxic
compounds (Banerjee et al., 2012).
Due to the deficiencies of sensory analysis methods, instrumental methods are used
as a complement in the evaluation of foods and allow for detailed qualitative analysis
(Wyllie, 2008). However, instrumental methods, such as gas chromatography/mass spec-
trometry (GC/MS), are also expensive and require trained people and considerable time
for the analysis (Pizzoni et al., 2015). These methods sometimes do not provide accurate
and reliable information (Ghasemi-Varnamkhasti et al., 2018).
The development of faster and more efficient identification methods remains the focus
of scientific research. In this sense, the electronic nose has attracted attention in many
branches of industry for its potential applicability in aroma analysis, such as for quality and
loss control, volatile release, and detection of aromas in food and beverages (Loutfi et al.,
2015). The electronic nose has several advantages over traditional methods of gas analysis.
Unlike other analytical instruments, it identifies the mixture of all the volatile compounds
present in the sample, without identification of individual chemical species. It is considered
a fast method, easy to operate, of small size, and it can perform the analysis in situ (Sun et
al., 2018). It is also low cost, simple to handle, and has good portability. No reagents are
needed for analysis and many applications are possible (Escuderos et al., 2013).

9.2 ELECTRONIC NOSE

The first electronic nose model was developed by Persaud and Dodd (1982) around 1980.
An electronic nose is an instrument that comprises an array of chemical sensors with
partial specificity and an adequate pattern recognition system capable of distinguishing
simple or complex odors (Gardner and Bartlett, 1994). It allows for an analysis of the
composition of the gas mixture as a whole without the separation and identification of
its specific components. It is an instrument that comprises a series of gas sensors (array)
with different selectivities toward analytes (cross response) and an appropriate pattern
recognition system.
Intrinsically, each sensor exhibits a different selectivity and sensitivity with respect
to particular components of the sample; however, they generate a chemical image char-
acteristic of the gas mixture, called a “fingerprint.” The samples will be evaluated and
classified (in terms of quality) based on these fingerprints or patterns (Gorji-Chakespari
et al., 2017; Gębicki and Szulczyński, 2018).
The operation of an electronic nose tries to mimic the biological process of human
olfaction in aroma identification. In the electronic nose, the volatile compounds must be
 Hyphenated Electronic Nose Technique  179

introduced through a sampling mechanism and transferred to the chamber containing the
gas sensors. The human nose olfactory system is replaced by the sensors, which, when in
contact with the aroma, interact with the sensitive layer. This layer undergoes reversible
physical and/or chemical alterations, causing changes in the electrical properties, such as
tension, for example, that will be transduced into electrical signals, pre-processed, and
assessed by software. The results are later identified by a pattern recognition system.
In the human nose, these electrical stimuli are transmitted to the brain which pro-
duces a recognition pattern in the memory. The brain can identify, classify, or perform
a hedonic analysis of the sample (Deshmukh et al., 2015; Lisboa, Page, and Guy, 2009;
Tudor Kalit et al., 2014; Santos, Lozano, and Aleixandre, 2017). A comparative illustra-
tion of the fundamental relations of the process of aroma recognition by a human and
electronic nose is presented in Figure 9.1 (Kiani, Minaei, and Ghasemi-Varnamkhasti,
2016; Vagin and Winquist, 2015; Ramgir, 2013).
The electronic nose (sensor array) mimics the behavior of the biological receptors in
the human olfactory epithelium. It is used to mimic the behavior of the human brain in
the task of recognizing and comparing odors or flavors. It can not only classify samples
but accurately determine the concentration of substances when combined with a suitable
multivariate calibration tool.
Figure 9.2 shows an electronic nose system used for aroma detection. These systems
generally consist of three main parts: (a) Sampling system in a chamber, (b) detection sys-
tem with an array of sensors, and (c) data processing and pattern recognition algorithms
in a computer (Kiani, Minaei, and Ghasemi-Varnamkhasti, 2016; Vagin and Winquist,
2015; Ramgir, 2013).

9.2.1 Sampling System

Sample handling is an important step in a detection system using an electronic nose,

where the quality of the analysis can be improved by using a suitable sampling system
(Sanaeifar et al., 2017). The proper sampling of the volatile fraction and its transfer to

FIGURE 9.1 Comparison of the components involved in the detection and identification
of aromas by human nose and electronic nose.

FIGURE 9.2 Electronic nose scheme for detection of aromas.

180 Food Aroma Evolution

the sensor chamber represents a significant challenge during the design of the analytical
methodology (Majchrzak et al., 2018).
The sampling chamber should be constructed of inert materials according to sample
size, not be reactive with the volatile compounds, and not release odors. A heating or
temperature control system must be used in order to increase the concentration of the
volatile components in the top of the chamber and maintain the same conditions in all
experiments (Sanaeifar et al., 2017).
Sampling can be performed in a variety of ways through the use of headspace (static or
dynamic) by using a diffusion method or sample enrichment (Cui et al., 2018; Estakhroyeh,
Rashedi, and Mehran, 2017; Sanaeifar et al., 2017). Headspace is usually performed in a
static or dynamic mode. In a static mode, the samples are incubated for a predetermined
time. An aliquot of the top volatiles is collected using a syringe and then injected into the
sensory chamber. The fact that only a relatively small fraction can be sampled is the main
limitation of this method (Majchrzak et al., 2018). However, it has the advantage of avoid-
ing the dilution of the volatiles in the free space, which increases the stability and sensitiv-
ity of the sensor and avoids disturbances within the sampling chamber (Wilson, 2012).
In the dynamic mode, an airflow is used to load the volatile compounds into the sensory
chamber (Ghasemi-Varnamkhasti et al., 2018), which is generally conducted using con-
trolled air pressure or nitrogen gas pumps (Triyana et al., 2015).

9.2.2 Detection System (Sensor Array)

The detection unit or multiple sensor array is an array of sensor elements capable of
transducing chemical changes or interactions in measurable signals. This unit is con-
sidered the most important part of the electronic nose system (Kalantar-zadeh and Fry,
2008; Ramgir, 2013; Rudnitskaya, 2018). The most common types of sensors, the opera-
tion principles, and the advantages and disadvantages are shown in Table 9.1 (Wojnowski
et al., 2017; Wardencki, Chmiel, and Dymerski, 2013; Deshmukh et al., 2015; Kiani,
Minaei, and Ghasemi-Varnamkhasti, 2016; Tiggemann et al., 2016).
Among the sensor types, metal oxide semiconductor (MOS) and conducting poly-
mer (CP) are the most commonly used in electronic nose systems (Kiani, Minaei, and
Ghasemi-Varnamkhasti, 2016). The principle of a sensor’s operation is based on the
interaction of the sensitive layer with the volatile compounds of the sample being ana-
lyzed (Di Rosa et al., 2017). For this, the sensors are exposed to an inert gas (reference
gas) for a baseline construction and to obtain a reference parameter, as can be seen in
Figure 9.3. Later the volatile compounds generated by the headspace are transported
through a tube to the chamber, in which they come in contact with the sensors. Then,
again, an inert gas is generated to remove the volatile compounds adhered to the surface
of the sensors and prepare them for a new measurement cycle (Nagle, Gutierrez-Osuna,
and Schiffman, 1998).
Gas sensors interact with the analyte to be detected by binding their molecules to
the surface of the sensitive layer using one or more mechanisms, including absorption,
adsorption, chemisorption, or chemical reactions. These generate physical and/or chemi-
cal changes, promoted by these processes, which are measured as an electrical signal
(Arshak et al., 2004).
An ideal gas sensor must have good reliability, robustness, high sensitivity, selectivity,
and reversibility. In order to have a system with selective and reversible responses, a series
of detection layers with different chemical properties must be used (James et al., 2005).
 Hyphenated Electronic Nose Technique  181

TABLE 9.1 Types of Sensors Most Used in Electronic Nose Systems, Operation Principles,
and the Advantages and Disadvantages
Sensor Type Operation Principle Advantages Disadvantages
Conducting When the conductive Operate at room High susceptibility to
polymer (CP) layer comes in contact temperature changes in relative
with the molecules of High stability humidity
the analyte, the Wide range of applications Short lifetime
conductivity of the Excellent reproducibility (typically 9–18
sensor changes High sensitivity months) due to
High selectivity oxidation of the
Short response time polymer
Easy synthesis
Metal oxide The detection Operate at 50–170°C, Operation under
semiconductor mechanism is based which reduces the impact ambient conditions
field-effect on the induction of of relative humidity on the must be under
transistor polarization on the output signal constant control,
(MOSFET) catalytically active Small size which excludes its
surface caused by an High selectivity and application in
intermediate reaction sensitivity portable and
Good sensitivity to toxic commercial
and flammable substances equipment
Metal oxide Show a change of Low response to humidity Vulnerable to
semiconductor resistance/tension on Good sensitivity, in the poisoning by volatile
(MOS) exposure to certain order of sub-ppm levels sulfur compounds
analytes caused by the Good discrimination and ethanol
reaction of volatile High response High power
chemical compounds High recovery time consumption
catalyzed by metal
oxides
Piezoelectric Alteration in the mass Fast response time Susceptibility to
Quartz crystal of the piezoelectric Low cost ambient temperature
microbalance sensor occurs during Potential for high changes
(QCM) its exposure to integration Complex fabrication
Surface odoriferous process
acoustic wave compounds
(SAW) sensors (adsorption/
Bulk acoustic absorption of the
wave (BAW) compound), which
causes a change in the
resonance frequency
Electrochemical Constructed from Very low power Low sensitivity
sensors electrodes immersed consumption
in an electrolyte. The Operate at room
analyte molecules are temperature
reduced or oxidized Not susceptible to humidity
causing a change in Robust
voltage or resistance
(Continued )
182 Food Aroma Evolution

TABLE 9.1 (Continued) Types of Sensors Most Used in Electronic Nose Systems,
Operation Principle, and the Advantages and Disadvantages
Sensor Type Operation Principle Advantages Disadvantages
Optical sensors Measure the Low energy consumption High cost
modulation of light High sensitivity Complex construction
properties or Ability to identify
characteristics individual compounds in
(changes in light mixtures
absorption, Short response time
polarization, Not susceptible to humidity
fluorescence,
wavelength, etc.) on
gaseous analytes
exposure
Mass Involves the Short response time High power
spectrometer- introduction of High sensitivity consumption
based volatile compounds in Qualitative and Complex construction
electronic the ionization quantitative analysis High cost
noses chamber of a mass Big size
spectrometer
instrument without
previous
chromatographic
separation

FIGURE 9.3 The typical response of the gas sensor when submitted to an analyte.

9.2.2.1 Gas Sensor Response Mechanism

Gas sensor characterization/performance in an electronic nose system usually involves
the study of parameters such as sensitivity, selectivity, response time, reversibility, repro-
ducibility, and the limit of detection, among others (Liu et al., 2012b; Skoog et al., 2005;
Dey, 2018). The following details the characterization of the response of the sensor.
 Hyphenated Electronic Nose Technique  183

(a) Sensitivity is the ratio of the slope in the calibration curve and the standard
deviation of the analytical signal at a given concentration.
(b) Selectivity is the ability of the sensor to identify a specific gas in a gas mixture.
(c) Response time is defined as the time spent by a sensor to generate a measurable
stable output signal corresponding to 63.2% of its stable maximum value after
the addition of the analyte.
(d) Reversibility is the ability of the sensor to return to its original state (baseline)
after analyte detection.
(e) Reproducibility means the sensors must have a stable and reproducible signal
over a period of time.
(f) Limit of detection is the lowest concentration that can be distinguished with a
certain level of confidence.

An ideal sensor must have high sensitivity, selectivity, and stability; fast response and
recovery times; and a low manufacturing cost (Dey, 2018).

9.2.3 Signal Processing System and Standard Recognition Methods

In most electronic noses, the processing system consists of a computer with software for
the acquisition and collection of the signals generated during volatile compounds detec-
tion (Majchrzak et al., 2018).
Pre-processing techniques are applied to the data in order to reduce the amount of
information being analyzed and to obtain the “olfactory pattern” of the samples and
to extract the static parameters of the measurements (Rodriguez-Gamboa, Albarracin-
Estrada, and Delgado-Trejos, 2011; Capelli et al., 2014). This procedure involves the
extraction of certain significant characteristics of the sensors’ response curves in order
to produce a set of numerical data that can be processed by the recognition system of the
electronic nose (Capelli et al., 2014).
Data analyses using algorithms are used to perform qualitative and quantitative clas-
sifications. Two classes of statistical methods are used most. Supervised methods are used to
classify unknown characteristics of a dataset that present common properties based on train-
ing samples, including artificial neural network (ANN), linear discriminant analysis (LDA),
and support vector machine (SVM) methods. Unsupervised methods such as principal com-
ponent analysis (PCA) and cluster analysis (CA) separate input data into different clusters
based on the similarity of sample characteristics (Cui et al., 2018; Majchrzak et al., 2018).

9.3 ELECTRONIC NOSE APPLICATION

The electronic nose can be applied in many areas of industrial production and human
activities, medical diagnostics, environmental monitoring, quality control of food prod-
ucts, or security systems. In the food industry, it is an important tool that can be used
during food and beverage processing, determination of shelf life, freshness assessment,
and authenticity or tamper assessment (Peris and Escuder-Gilabert, 2013). Some exam-
ples of the application of the hyphenated electronic nose technique in aroma analysis in
foods and beverages are presented below.
In food production processes that have chemical parameters that vary over time, it
is of fundamental importance to comply with the regulations related to quality control
184 Food Aroma Evolution

FIGURE 9.4 Electronic nose system applied to detect and discriminate volatile com-
pounds in gummy candies aromatized with apple, strawberry, and grape. The gas sensor
array is based on graphite interdigitated electrodes with conducting polymer layer.

(Peris and Escuder-Gilabert, 2013). Problems with food authenticity assessment (adul-
teration) can cause a serious risk to health in some instances. In this way, the use of
advanced sensor arrays have been reported as being used for authenticity/adulteration
assessment in oils, juices, canned of fruits, milk, meat, tea, wine, whisky, and spices
(Peris and Escuder-Gilabert, 2016).
The aroma is used as a quality parameter and product conformity indicator, intimately
related to the product’s acceptance by the consumer. For this purpose, Graboski et al.
(2018a, 2018b) applied an electronic nose composed of an array of gas sensors with differ-
ent polyaniline-based sensors for aroma volatile compound detection (apple, strawberry,
and grape) in gummy candy. The authors observed that this system was able to discriminate
between the aromas in the food matrix (gummy candies) using PCA (Figure 9.4).
The control of the degree of maturity of fruit is very important due to its susceptibil-
ity to disease and fast deterioration. Electronic nose systems can be used for minimizing
production losses and also preserving the unique features of each fruit. Manzoli et al.
(2011) used a low-cost sensor array system for monitoring banana ripeness (Figure 9.5).
The volatile compound released during banana ripening is ethylene; when the fruit is ripe
and full-ripe it releases 2-hexenal, eugenol, and isopentanol. In the experiment, unripe
bunches of bananas were used, and the maturation volatiles over the course of 5 days
were evaluated. The criterion of ripeness was the color of the banana peel. The sensor
array was able to produce a distinct pattern of signals that could be used to characterize
the bananas’ degree of ripeness.
To evaluate the preservation of the leaf aroma in the drying process of medicinal
and aromatic plants, Kiani, Minaei, and Ghasemi-Varnamkhasti (2018) identified vola-
tile compounds released by the foodstuff using an electronic nose. In the drying of mint
leaves, in which aroma volatilization may alter the aromatic quality, the electronic nose
system was capable of differentiating the mint aroma in different drying stages (fresh and
dried mint) and of determining the time to terminate the drying process.
In the beverage industry, for classification and quality-control purposes, different
fruit aromas (strawberry, lemon, cherry, and melon) were evaluated using an electronic
nose (Adak and Yumusak, 2016). By the use of neural networks, it was possible to train
the nose separately with backpropagation, for 60% of the sample’s values for all four of
the aroma types.
 Hyphenated Electronic Nose Technique  185

FIGURE 9.5 Experimental setup of an electronic nose based on a gas sensor with poly-
aniline films applied to detect the volatiles emitted during banana ripening. Numbers
indicate gas sensor (1), glass chamber (2), data acquisition (3), and computer (4).

Xiao et al. (2014) used an electronic nose to classify Chinese liquor (an alcoholic
drink made from starter cultures) aroma mixtures with different origins and liquor flavor
types. They identified 86 aromas, including acids, esters, alcohols, aldehydes, ketones,
phenols, nitrous, and sulfuric compounds; and the electronic nose could differentiate
between the liquor origins and flavor types.
Liu et al. (2012a) evaluated 20 Chinese spirits which belonged to eight different
flavors using a portable electronic nose. The authors found that a combination of an
appropriate sensor array with a pattern recognition method could evaluate the quality
and flavor of the samples.
In a study by Gupta, Variyar, and Sharma (2015) using aroma data, in food irradi-
ated by electromagnetic γ-rays or electron beams for improving safety and shelf life and
reducing microbial contamination, the electronic nose was able to identify a food that
was or not subjected to irradiation. First PCA and later LDA were used to classify grape
and apple fruits in a chemometric approach. A 100% success rate was obtained for dis-
criminating between irradiated samples at all doses (0.1, 0.25, 0.5, 1.0, 1.5, and 2.0 kGy).
The quality of black tea aroma of multiple tea gardens spread across north and
north-east India was evaluated by Tudu et al. (2009) using an electronic nose and a neu-
ral network for classification. It was possible to classify four tea gardens in north and
north-east India using this system. Black tea quality is quite complex due to the presence
of innumerable compounds. In this tea, a fermentation process can occur with biochemi-
cal reactions in green tea leaves. The transformation from a grassy to sweet floral aroma
was identified by the authors using an electronic nose.
Sharma et al. (2015) monitored the fermentation of black tea in real time using an
electronic nose (an array of quartz crystal microbalance sensors coated with glucose,
maltose, maltodextrin, β-cyclodextrin, d-glucose, and polyethylene glycols). The highest
peak of the sensor represented the optimum fermentation point, proving that the elec-
tronic nose was able to identify the fermentation profile.
The organoleptic requirements for evaluating honey odor included the analysis of
dominant pollen in honey deposits, type, and origin. A classification of Polish honey types
(acacia flower, linden flower, rape, buckwheat, and honeydew) was studied by Dymerski
et al. (2014) using an electronic nose with chemometric data analysis. With PCA and
186 Food Aroma Evolution

cluster analysis, it was possible to discriminate between three types of honey (acacia
flower, linden flower, and rape). Ampuero, Bogdanov, and Bosset (2004) employed an
electronic nose for an investigation of honey matrices, and a good correlation was found
between the data of the study and the classical method for the determination of the
botanical origin of honey.
Physiological changes from the minimal processing of vegetables can cause loss of
taste and production of off-flavors in these products. Torri, Sinelli, and Limbo (2010)
applied a commercial electronic nose to monitor the changes of volatile compounds in
minimally processed pineapples during their storage at different temperatures. According
to PCA, there was much more evident discrimination between the samples kept at higher
temperatures than at lower temperatures. By CA there was a separation into two groups,
“fresh pineapple” and “old pineapple,” making it possible to determine a stability time
(maximum acceptability time for the loss of freshness) of 5.3 days at 5.3°C, 2.7 days at
8.6°C, and 1.2 days at 15.8°C. Therefore, this approach can be applied for evaluating
loss of aroma quality, freshness, and influence on aroma and taste quality during storage.
In the meat and fish industry, it is fundamental to have a fast and accurate detection
system for microbial spoilage to avoid production losses. Food spoilage is associated
with microbial volatiles, organoleptic changes, and the development of off-odors. These
parameters can be used as indicators of bacterial presence and, subsequently, of meat
quality. The commercial electronic nose was used for the detection of these microbial vol-
atiles in beef food spoilage and was able to classify stored beef into two classes (“spoiled”
and “unspoiled”) (Balasubramanian et al., 2004). It was also employed to monitor the
spoilage of packaged beef fillet at different storage temperatures (0, 4, 8, 12, and 16°C).
Food additives are added in products to enhance taste and preserve flavor or appear-
ance during processing, packaging, and storage. But its presence in food products must be
considered by consumers because in some cases it can represent health risks. Electronic
nose application in citrus juices for monitoring food additives (benzoic acid and chitosan)
was reported by Qiu and Wang (2017). The authors used a linear discriminant analysis
for discrimination and classification and obtained 85.5 and 95.0% cross-validation for
benzoic acid and chitosan, respectively, as observed in Figure 9.6.
The addition, substitution, and removal of substances and the dilution of products
are considered food adulterations. Controls on adulteration are required to avoid harm to
human health. In this way, the hyphenated electronic nose can be applied for evaluating
common fraudulent procedures (partial or complete substitution of an authentic ingredi-
ent), considering a specific odor (pattern recognition system) or authentic or traditional
aroma of a product. For this purpose, many foods have been monitored such as milk,
cheese, several oils (olive, sesame, palm olein, virgin coconut, flaxseed), meat and meat
products, honey, juices, wine, vinegar, and alcoholic beverages (Gliszczyńska-Świgło and
Chmielewski, 2017).

9.4 CONCLUSION AND FUTURE PERSPECTIVES

The accuracy of an electronic nose’s performance may be limited by extrinsic factors

such as humidity and temperature, or by intrinsic factors such as sensor drift or instru-
mentation errors. To classify and authenticate different analytes or properties in complex
samples, a standard sample for evaluating the sensor’s response is required. To make this
possible, different kinds of electrodes and an array of sensors can be used with a high
sensitivity and even selectivity. The response obtained can be evaluated using a statistical
 Hyphenated Electronic Nose Technique  187

FIGURE 9.6 Schematic representation of the electronic nose applied to citrus juice to
discriminate benzoic acid and chitosan additives.

tool to discriminate between aroma samples. In future works, the use of statistical tools
and more artificial intelligence in these devices are expected to discriminate between
samples with very similar behavior.

CONFLICT OF INTEREST

The authors declare that they do not have a conflict of interest.

ACKNOWLEDGMENT

The authors acknowledge the National Council for Scientific and Technological
Development, Coordination for the Improvement of Higher Education Personnel,
Research Support Foundation of the State of Rio Grande, and Research and Projects
Financing for the support received.

REFERENCES

Adak, M., and Y. Nejat. 2016. “Classification of e-nose aroma data of four fruit types by
ABC-based neural network.” Sensors 16 (3): 304.
Agência Nacional de Vigilância Sanitária (ANVISA). 2007. Resolução de diretoria cole-
giada—RDC no 2, de 15 de janeiro de 2007. ANVISA. http:​//por​tal.a​nvisa​.gov.​
br/do​cumen​ts/10​181/2​71837​6/RDC​_ 02_2​0 07_C​OMP.p​df/c9​66caf​f-1c1​9-4a2​f-87a​
6-05f​7a09e​940b.​
Ampuero, S., S. Bogdanov, and J.-O. Bosset. 2004. “Classification of unifloral honeys
with an MS-based electronic nose using different sampling modes: SHS, SPME and
INDEX.” European Food Research and Technology 218 (2): 198–207.
188 Food Aroma Evolution

Arshak, K., E. Moore, G.M. Lyons, J. Harris, and S. Clifford. 2004. “A review of gas
sensors employed in electronic nose applications.” Sensor Review 24 (2): 181–98.
Balasubramanian, S., S. Panigrahi, C.M. Logue, M. Marchello, C. Doetkott, H. Gu, J.
Sherwood, and L. Nolan. 2004. “Spoilage identification of beef using an electronic
nose system.” Transactions of the ASAE 47 (5): 1625–33.
Banerjee, R., B. Tudu, L. Shaw, A. Jana, N. Bhattacharyya, and R. Bandyopadhyay.
2012. “Instrumental testing of tea by combining the responses of electronic nose
and tongue.” Journal of Food Engineering 110 (3): 356–63.
Capelli, L., S. Sironi, and R. Del Rosso. 2014. “Electronic noses for environmental moni-
toring applications.” Sensors 14 (11). Multidisciplinary Digital Publishing Institute:
19979–87.
Cui, S., P. Ling, H. Zhu, and H. Keener. 2018. “Plant pest detection using an artificial
nose system: A review.” Sensors 18 (2): 378.
Deshmukh, S., R. Bandyopadhyay, N. Bhattacharyya, R.A. Pandey, and A. Jana. 2015.
“Application of electronic nose for industrial odors and gaseous emissions measure-
ment and monitoring—An overview.” Talanta 144. Elsevier: 329–40.
Dey, A. 2018. “Semiconductor metal oxide gas sensors: A review.” Materials Science and
Engineering: B 229 (March). Elsevier: 206–17.
Dymerski, T., J. Gębicki, W. Wardencki, and J. Namieśnik. 2014. “Application of an
electronic nose instrument to fast classification of polish honey types.” Sensors 14
(6): 10709–24.
Escuderos, M.E., M. García, A. Jiménez, and M.C. Horrillo. 2013. “Edible and non-edi-
ble olive oils discrimination by the application of a sensory olfactory system based
on tin dioxide sensors.” Food Chemistry 136 (3–4). Elsevier: 1154–9.
Estakhroyeh, H.R., E. Rashedi, and M. Mehran. 2017. “Design and construction of
electronic nose for multi-purpose applications by sensor array arrangement using
IBGSA.” Journal of Intelligent & Robotic Systems 92 (2): 205–21.
Fani, M. 2015. “Dossie de aromas.” Food Ingredients Brasil (FBI) 17 (33): 30–53. http:​
//www​.revi​sta-f​i .com ​/edic​oes_m​ateri​as.ph​p?id_​edica​o=70.​
Firestein, S. 2001. “How the olfactory system makes sense of scents.” Nature 413 (6852): 211–8.
Gardner, J.W., and P.N. Bartlett. 1994. “A brief history of electronic noses.” Sensors and
Actuators B: Chemical 18 (1–3): 210–1.
Gębicki, J., and B. Szulczyński. 2018. “Discrimination of selected fungi species based on
their odor profile using prototypes of electronic nose instruments.” Measurement
116 (November, 2017): 307–13.
Ghasemi-Varnamkhasti, M., C. Apetrei, J. Lozano, and A. Anyogu. 2018. “Potential
use of electronic noses, electronic tongues and biosensors as multisensor systems
for spoilage examination in foods.” Trends in Food Science & Technology 80
(October): 71–92.
Gliszczyńska-Świgło, A., and J. Chmielewski. 2017. “Electronic nose as a tool for moni-
toring the authenticity of food: A review.” Food Analytical Methods 10 (6): 1800–16.
Gorji-Chakespari, A., A.M. Nikbakht, F. Sefidkon, M. Ghasemi-Varnamkhasti, and E.L.
Valero. 2017. “Classification of essential oil composition in rosa damascena mill.
genotypes using an electronic nose.” Journal of Applied Research on Medicinal and
Aromatic Plants 4 (March): 27–34.
Graboski, A.M., E. Galvagni, A. Manzoli, F.M. Shimizu, C.A. Zakrzevski, T.A.
Weschenfelder, J. Steffens, and C. Steffens. 2018a. “Lab-Made electronic-nose with
polyaniline sensor array used in classification of different aromas in gummy can-
dies.” Food Research International 113: 309–15.
 Hyphenated Electronic Nose Technique  189

Graboski, A.M., S.C. Ballen, A. Manzoli, F.M. Shimizu, C.A. Zakrzevski, J. Steffens,
and C. Steffens. 2018b. “Array of different polyaniline-based sensors for detec-
tion of volatile compounds in gummy candy.” Food Analytical Methods 11 (1):
77–87.
Gupta, S., P.S. Variyar, and A. Sharma. 2015. “Application of mass spectrometry based
electronic nose and chemometrics for fingerprinting radiation treatment.” Radiation
Physics and Chemistry 106 (January): 348–54.
James, D., S.M. Scott, Z. Ali, and W.T. O’Hare. 2005. “Chemical sensors for electronic
nose systems.” Microchimica Acta 149 (1–2): 1–17.
Kalantar-zadeh, K., and B. Fry. 2008. “Sensor characteristics and physical effects.” In
Nanotechnology-Enabled Sensors, pp. 13–62. Boston, MA: Springer US.
Kiani, S., S. Minaei, and M. Ghasemi-Varnamkhasti. 2016. “Application of electronic
nose systems for assessing quality of medicinal and aromatic plant products: A
review.” Journal of Applied Research on Medicinal and Aromatic Plants 78 (3):
195–8.
Kiani, S., S. Minaei, and M. Ghasemi-Varnamkhasti. 2018. “Real-time aroma monitor-
ing of mint (Mentha Spicata L.) leaves during the drying process using electronic
nose system.” Measurement 124 (August): 447–52.
Lisboa, H.M., T. Page, and C. Guy. 2009. “Gestão de odores : Fundamentos do nariz
eletrônico.” Engenharia Sanitária Ambiental 14 (1): 9–18.
Liu, M., X. Han, K. Tu, L. Pan, J. Tu, L. Tang, P. Liu, G. Zhan, Q. Zhong, and Z. Xiong.
2012a. “Application of electronic nose in Chinese spirits quality control and flavour
assessment.” Food Control 26 (2): 564–70.
Liu, X., S. Cheng, H. Liu, S. Hu, D. Zhang, and H. Ning. 2012b. “A survey on gas sens-
ing technology.” Sensors 12 (12): 9635–65.
Loutfi, A., S. Coradeschi, G.K. Mani, P. Shankar, and J.B.B. Rayappan. 2015. “Electronic
noses for food quality: A review.” Journal of Food Engineering 144. Elsevier:
103–11.
Majchrzak, T., W. Wojnowski, T. Dymerski, J. Gębicki, and J. Namieśnik. 2018.
“Electronic noses in classification and quality control of edible oils: A review.” Food
Chemistry 246 (April). Elsevier: 192–201.
Manzoli, A., C. Steffens, R.T. Paschoalin, A. Correa, W. Alves, F. Leite, and P. Herrmann.
2011. “Low-cost gas sensors produced by the graphite line-patterning technique
applied to monitoring banana ripeness.” Sensors 11 (12): 6425–34.
Nagle, H.T., R. Gutierrez-Osuna, and S.S. Schiffman. 1998. “The how and why of elec-
tronic noses.” IEEE Spectrum 35 (9): 22–31.
Peris, M., and L. Escuder-Gilabert. 2013. “On-line monitoring of food fermentation pro-
cesses using electronic noses and electronic tongues: A review.” Analytica Chimica
Acta 804 (December): 29–36.
Peris, M., and L. Escuder-Gilabert. 2016. “Electronic noses and tongues to assess
food authenticity and adulteration.” Trends in Food Science & Technology 58
(December). Elsevier: 40–54.
Persaud, K., and G. Dodd. 1982. “Analysis of discrimination mechanisms in the mam-
malian olfactory system using a model nose.” Nature 299 (5881): 352–5. http:​//
www​.ncbi​.nlm.​nih.g​ov/pu​bmed/​71103​56.
Pizzoni, D., D. Compagnone, C. Di Natale, N. D’Alessandro, and P. Pittia. 2015.
“Evaluation of aroma release of gummy candies added with strawberry flavours by
gas-chromatography/mass-spectrometry and gas sensors arrays.” Journal of Food
Engineering 167 (December). Elsevier: 77–86.
190 Food Aroma Evolution

Plutowska, B., and W. Wardencki. 2007. “Aromagrams—Aromatic profiles in the appre-

ciation of food quality.” Food Chemistry 101 (2): 845–72.
Qiu, S., and J. Wang. 2017. “The prediction of food additives in the fruit juice based
on electronic nose with chemometrics.” Food Chemistry 230 (September). Elsevier:
208–14.
Ramgir, N.S. 2013. “Electronic Nose based on nanomaterials: Issues, challenges, and
prospects.” ISRN Nanomaterials 2013: 1–21.
Rodriguez-Gamboa, J.C., E.S. Albarracin-Estrada, and E. Delgado-Trejos. 2011.
“Quality control through electronic nose system.” In Modern Approaches to
Quality Control, pp. 505–22. Janeza Trdine: Croatia. InTech.
Rosa, A.R.D., F. Leone, F. Cheli, and V. Chiofalo. 2017. “Fusion of electronic nose, elec-
tronic tongue and computer vision for animal source food authentication and qual-
ity assessment—A review.” Journal of Food Engineering 210 (October). Elsevier:
62–75.
Rudnitskaya, A. 2018. “Calibration update and drift correction for electronic noses and
tongues.” Frontiers in Chemistry 6 (September): 433.
Sanaeifar, A., H. ZakiDizaji, A. Jafari, and M. de la Guardia. 2017. “Early detection of
contamination and defect in foodstuffs by electronic nose: A review.” TrAC Trends
in Analytical Chemistry 97 (December). Elsevier B.V.: 257–71.
Santonico, M., P. Pittia, G. Pennazza, E. Martinelli, M. Bernabei, R. Paolesse, A.
D’Amico, D. Compagnone, and C. Di Natale. 2008. “Study of the aroma of arti-
ficially flavoured custards by chemical sensor array fingerprinting.” Sensors and
Actuators B: Chemical 133 (1): 345–51.
Santos, J.P., J. Lozano, and M. Aleixandre. 2017. “Electronic noses applications in beer
technology.” In Brewing Technology, pp. 177–200. InTech.
Seuvre, A.M., E. Philippe, S. Rochard, and A. Voilley. 2007. “Kinetic study of the release
of aroma compounds in different model food systems.” Food Research International
40 (4): 480–92.
Sharma, P., A. Ghosh, B. Tudu, S. Sabhapondit, B.D. Baruah, P. Tamuly, N. Bhattacharyya,
and R. Bandyopadhyay. 2015. “Monitoring the fermentation process of black tea
using QCM sensor based electronic nose.” Sensors and Actuators B: Chemical 219
(November): 146–57.
Skoog, D., D. West, J. Holler, and S. Crouch. 2005. Fundamentals of analytical chemis-
try. Analytical Chemistry 398: 27–8. Belmont: Thomson-Brooks/Cole.
Sun, F., Z. Wu, Y. Chen, J. Li, S. He, and R. Bai. 2018. “Analysis of odors from thermally
modified bamboo assessed by an electronic nose.” Building and Environment 144
(August): 386–91.
Tiggemann, L., S. Ballen, C. Bocalon, A.M. Graboski, A. Manzoli, P.S. De Paula
Herrmann, J. Steffens, E. Valduga, and C. Steffens. 2016. “Low-cost gas sensors
with polyaniline film for aroma detection.” Journal of Food Engineering 180: 16–21.
Torri, L., N. Sinelli, and S. Limbo. 2010. “Shelf life evaluation of fresh-cut pineapple by
using an electronic nose.” Postharvest Biology and Technology 56 (3): 239–45.
Triyana, K., M.T. Subekti, P. Aji, S.N. Hidayat, and A. Rohman. 2015. “Development of
electronic nose with low-cost dynamic headspace for classifying vegetable oils and
animal fats.” Applied Mechanics and Materials 771 (July): 50–4.
Tudor Kalit, M., K. Marković, S. Kalit, N. Vahčić, and J. Havranek. 2014. “Application
of electronic nose and electronic tongue in the dairy industry.” Mljekarstvo 64 (4).
Hrvatska mljekarska udruga: 228–44.
 Hyphenated Electronic Nose Technique  191

Tudu, B., A. Jana, A. Metla, D. Ghosh, N. Bhattacharyya, and R. Bandyopadhyay. 2009.

“Electronic nose for black tea quality evaluation by an incremental RBF network.”
Sensors and Actuators B: Chemical 138 (1): 90–95.
Vagin, M.Y., and F. Winquist. 2015. “Electronic noses and tongues in food safety assur-
ance.” In High Throughput Screening for Food Safety Assessment, pp. 265–83.
Elsevier.
Wardencki, W., T. Chmiel, and T. Dymerski. 2013. “Gas chromatography-olfactometry
(GC-O), electronic noses (e-noses) and electronic tongues (e-tongues) for in vivo
food flavour measurement.” In Instrumental Assessment of Food Sensory Quality,
pp. 195–229. Elsevier.
Wilson, A.D. 2012. “Advanced methods for teaching electronic-nose technologies to diag-
nosticians and clinical laboratory technicians.” Procedia—Social and Behavioral
Sciences 46: 4544–54.
Wojnowski, W., T. Majchrzak, T. Dymerski, J. Gębicki, and J. Namieśnik. 2017.
“Electronic noses: Powerful tools in meat quality assessment.” Meat Science 131
(May): 119–31.
Wyllie, S.G. 2008. “Flavour quality of fruit and vegetables: Are we on the brink of major
advances?” In Fruit and Vegetable Flavour, pp. 3–10. Woodhead Publishing.
Xiao, Z., D. Yu, Y. Niu, F. Chen, S. Song, J. Zhu, and G. Zhu. 2014. “Characterization
of aroma compounds of Chinese famous liquors by gas chromatography–mass
spectrometry and flash GC electronic-nose.” Journal of Chromatography B 945–6
(January): 92–100.
 Chapter 10
Food Aroma Compounds by
Capillary Electrophoresis
Raffaella Colombo and Adele Papetti

CONTENTS

10.1 Introduction 194

10.2 CE Instrumentation 194
10.2.1 System 194
10.2.2 Capillaries 194
10.2.3 Buffers 195
10.2.4 I njection and Separation 196
10.2.5 Detectors 196
10.3 Theoretical Principles 197
10.4 Separation Modes 197
10.4.1 Micellar Electrokinetic Chromatography (MEKC) 197
10.4.2 Chiral Capillary Electrophoresis (CCE) 198
10.4.3 Capillary Electrochromatography (CEC) 198
10.4.4 Non-Aqueous Capillary Electrophoresis (NACE) 198
10.5 Advantages and Limitations 199
10.6 Applications 199
10.6.1 Non-Enzymatic Reactions 199
10.6.1.1 Amines 200
10.6.1.2 Furanones 200
10.6.1.3 Phenols 200
10.6.1.4 Pyranones 201
10.6.1.5 Pyrazines 201
10.6.1.6 Pyridines 202
10.6.1.7 Thiols, Thioethers, and Di- and Trisulfides 202
10.6.1.8 Amino Acids as Precursors of Food Aroma Compounds 202
10.6.1.9 Other Substances as Precursors of Aroma Compounds 203
10.6.2 Enzymatic Reactions 203
10.6.2.1 Carbonyl Compounds 203
10.6.2.2 Terpenes 206
10.6.2.3 Other Substances as Precursors of Aroma Compounds 206
10.6.2.4 Phenols as Precursors of Food Aroma Compounds 206
References 210

193
194 Food Aroma Evolution

10.1 INTRODUCTION

Capillary electrophoresis (CE) is a microscale technique, and its principle consists in the
migration of ions/charged molecules in a buffer solution through an open, fused-silica
capillary under an applied electric field. The separation is based on a molecule mass-to-
charge ratio. From the early 1980s to date, CE has resulted in the application of different
CE separation modes, the progression of coupling CE with sensitive detection systems,
and the advances in microchip-CE technology. This improves the versatility of CE with
important applications in food quality and safety, in food processing and stability, and
also in foodomics (Pinero, Bauza, and Arce, 2011; Karlinsey, 2012; Acunha et al., 2016;
Papetti and Colombo, in press). In particular, miniaturized CE systems (microchip-CE
devices), which simultaneously allow sample preparation, separation, and quantification
in a chip, and ensure rapid and sensitive methods for detecting fraud or contamination
(Martín, Vilela, and Escarpa, 2012).

10.2 CE INSTRUMENTATION

10.2.1 System

CE separations occur in a bare fused-silica capillary (30 to 100 cm long), with an inner
diameter (i.d.) of 50–100 µm and an outer diameter (o.d.) of 150–360 μm. Both cap-
illary ends are immersed in buffer (background electrolyte, BGE) reservoirs, together
with platinum electrodes to keep the conductivity at the applied voltage (10–30 kV).
Capillaries are covered with a copolymer (polyimide) with heat resistance and flexibil-
ity, not transparent to the UV light. Therefore, it is necessary to remove it to create a
detection window, which is transparent to UV, providing online detection (Camilleri,
1997; Whatley, 2001). The voltage is applied to the total capillary length (L), but the
analytes only cross the capillary until they reach the effective capillary length (l), which
is the distance from the injection to the detection window. As a high electric field and
the electrophoretic principle can cause Joule heating and zone broadening; the capillary
temperature must be controlled with high-speed forced-air coolers or recirculating liq-
uid coolant systems. Temperature also becomes an important parameter for maintaining
a constant current and buffer viscosity and reproducible migration times (Weinberger,
2000; Whatley, 2001). Notwithstanding the presence of controlled temperature systems,
the temperature inside the capillary can only be estimated, and the generated Joule heat-
ing increases proportionally over the electric field (applied voltage/capillary length) and
the BGE conductivity and concentration (Whatley, 2001). Figure 10.1 illustrates a sche-
matic CE system.

10.2.2 Capillaries

CE uncoated capillary walls are made of silanol groups (Si–OH), which are weak acids
(pK~7), and have a negative charge in alkaline conditions and buffers. To have a stable
system, operative conditions must be setup in a pH range of 2–9 with buffers, respon-
sible for pH control, and analysis reproducibility. The application of a voltage gives an
electrical double layer (electric ζ potential) along the inner wall, in which buffer cations
are attracted to the negative electrode, creating the so-called electroosmotic flow (EOF)
 Food Aroma Compounds by Capillary Electrophoresis 195

FIGURE 10.1 Schematic representation of a CE system. (Reprinted with permission from

Karlinsey, J. M. (2012). Electrophoresis. In Y. Picò (Ed.), Chemical Analysis of Food:
Techniques and Applications (pp. 375–405). Waltham (MA): Academic Press Elsevier.)

(Slater, Tessier, and Kopecka, 2010). A new uncoated capillary must be conditioned
with an appropriate volume of strong bases and buffers to create an EOF. EOF velocity
depends on the applied electric field; it is directly proportional to buffer dielectric con-
stant and ζ potential, and indirectly proportional to buffer viscosity. It is often necessary
for suppressing EOF because of capillary–analyte interaction/absorption, which causes
EOF mobility variation and irreproducibility. In this case, the capillary inner wall is
chemically modified by covalent bonding (covalent coating) with neutral or hydrophilic
substituents or by adding BGE polymeric modifiers (dynamic or adsorptive coating)
(Horvath and Dolník, 2001; Whatley, 2001; Slater, Tessier, and Kopecka, 2010).
A new capillary has to be pre-conditioned at high-pressure values (≥1 bar). The
reagents for the conditioning procedure are different, based on uncoated or coated capil-
laries. An uncoated capillary is rinsed with 10 to 15 column volumes of strong bases,
followed by the same column volumes of the BGE, to generate EOF. For a coated capil-
lary, in which EOF must be suppressed, other solvents, such as ethanol or toluene, and
appropriate coated agents are used. In addition, a conditioning inter-run is necessary to
regenerate the capillary wall at the end of each analysis (Weinberger, 2000; Whatley,
2001; Slater, Tessier, and Kopecka, 2010).

10.2.3 Buffers

The BGE modulates pH and silanol dissociation and drives the migration of analytes,
which differ for migration times and mobilities. Buffers must be chosen based on a high-
salt purity, high-buffer capacity (pH = pKa±1), low-absorbance, low-conductivity, and
low-temperature coefficient. In the choice of a run buffer the parameters to take into
account are pH value, ionic strength, types of salts (inorganic or organic), and operating
temperature. Organic buffers have high ionic strength, which is correlated with a high
196 Food Aroma Evolution

buffer capacity; they exhibit low mobility/conductivity and less Joule heating, giving the
possibility of using higher voltages with an increase in peak efficiency. On the contrary,
organic buffers absorb UV–Vis light, and this causes an increase in baseline absorbance
(Reijenga et al., 1996; Camilleri, 1997; Weinberger, 2000). The ionic force of BGE can
be modulated in relation to the ionic force of the sample; in fact, a difference in the con-
centration/conductivity between a sample buffer and BGE allows for so-called “sample
stacking,” a procedure able to increase peak efficiency and sensitivity (Camilleri, 1997;
Whatley, 2001).

10.2.4 Injection and Separation

The injection plug amount must be of 1–50 nL to optimize efficiency and resolution. The
most used type of injection is by pressure or vacuum (hydrodynamic injection) by an on-
board air pump, in which two parameters (pressure value and time) can be setup. The
injected volume can be calculated with the Poiseuille equation; it is directly proportional
to pressure, capillary inner diameter and pressure application time, and indirectly pro-
portional to BGE viscosity and total capillary length (Whatley, 2001). Another type of
injection consists of the use of low voltage values over short time periods (electrokinetic
injection). It is rarely used because of a low reproducibility.
After a sample injection, the application of voltage allows for the migration of ions.

10.2.5 Detectors

CE detection occurs on-capillary with the advantage of having no void volumes and the
disadvantage of a short path length (i.e., i.d.), which is responsible for the low concentra-
tion sensitivity of this technique. Absorbance detectors are the most used and can be UV
detectors, which use only a portion of the available energy, or photodiode array detectors
(PDA), which exploit the entire spectrum of UV light, with the advantage of estimating
peak purity. For compounds, which have no or low UV absorption, such as inorganic
ions or some acids, respectively, indirect UV detection can be a solution. In indirect UV,
a molecule with a UV absorption and a mobility similar to the analyte, is added to BGE.
When analytes migrate in the BGE, a displacement of the absorbing molecule occurs, and
a decrease in absorbance is registered (Camilleri, 1997).
Electrochemical (EC) detectors consist mainly of amperometric detectors (AD), in
which current is measured. They are more sensitive in comparison with UV detection and
are based on oxidation/reduction reactions between analytes and electrodes. This type
of detection can be very useful in food analysis for compounds in trace levels (Karlinsey,
2012). Recently, the development of advanced EC detectors, such as CE contactless cou-
pled detection (CE-CCD) and CE capacitively coupled contactless conductivity detection
(CE‐C4D), has allowed for an increase in sensitivity, which is very useful in food analysis
(Elbashir, Schmitz, and Aboul-Enein, 2017).
Another detection system with increased sensitivity is represented by the laser-
induced fluorescence (LIF) detector, which requires a process of derivatization (on- or
off-capillary) for non-fluorescent molecules (Karlinsey, 2012). In the case of food aroma
compounds, it is mainly used for amino acid precursors.
Mass spectrometer (MS) detectors can add information on molecular weight and the
mode MS/MS (MS2) also provides structural information. Coupling with CE improves
 Food Aroma Compounds by Capillary Electrophoresis 197

velocity, resolving capacity, selectivity, and sensitivity. CE-MS is mostly applied to the
analysis of traces of contaminants, residues, and biomarkers to ensure quality and
authenticity in food samples (Ravelo-Pérez et al., 2009; Ibáñez et al., 2013).

10.3 THEORETICAL PRINCIPLES

The electrophoretic mobility (µe) represents the velocity (v) of an ion/analyte through the
capillary, in which an electric field (E) is applied (µe=v/E). EOF represents the velocity of
the bulk flow (vEOF) in this electric field and it is expressed as a mobility (µEOF or µ0).
In normal polarity and with injection at a node pole, cations and neutral species migrate
with EOF and anions migrate against it. In reverse polarity, the inner wall assumes a
positive charge, and the EOF moves in the opposite direction. The so-called apparent
mobility (µapp) refers to the observed mobility of an analyte, which takes into account
its effective mobility (µeff) and the EOF contribution (µapp= µEOF+µeff). The mobility
is calculated based on migration time, voltage, and capillary length (µapp=l/tE; E=V/L;
µapp=lL/Vt) (Camilleri, 1997). In ideal conditions (a small injection plug length and the
absence of an interaction between the analyte and the capillary), only longitudinal diffu-
sion is present, and as a consequence peak efficiency depends only on the solute molecular
diffusion coefficient.

10.4 SEPARATION MODES

When only BGE is used, the technique is called capillary zone electrophoresis (CZE)
or free-solution capillary electrophoresis (FSCE), and it can be applied only to charged
analytes. The simple addition of different additives to BGE produces different CE-modes
and separation principles. Here the main CE-modes applied in the analysis of food aroma
compounds were selected. Table 10.1 shows the principles and parameters of CE-modes
applied to food aroma compounds.

10.4.1 Micellar Electrokinetic Chromatography (MEKC)

The presence of ionic or non-ionic surfactants (sodium dodecyl sulfate, dodecyltrimeth-

ylammonium bromide, Triton X-100), which are added to BGE in concentrations useful
to constitute micelles, creates a sort of pseudophase, and for this reason the technique is

TABLE 10.1 CE Separation Modes Mainly Used in the Analysis of Food Aroma Compounds
Stationary
CE Mode Principle BGE Additives Phase Type of Molecule
CZE Mobility / / Charged
MEKC Hydrophobic/ionic interaction Surfactant / Neutral/charged
CCE Diastereoisomer formation Chiral selector / Chiral
CEC Mobility / Present Neutral/charged
Hydrophobic/ionic interaction
NACE ion-pairing effects Organic solvent / Low soluble
198 Food Aroma Evolution

named “chromatohraphy.” In fact, they act as a pseudo-stationary phase with a hydro-

phobic core and a hydrophilic outer surface able to create hydrophobic and/or ionic inter-
actions with the analytes. This technique also allows for the separation of neutral species
contrary to CZE. The use of a basic buffer is recommended to minimize interactions
between the surfactant and the capillary wall (Terabe, 2009).
The MEKC separation mode can also be applied in chiral conditions (see Section
10.4.2), allowing enatiomeric separation, such as for studying the stability of catechins in
tea infusions (Mirasoli et al., 2014).

10.4.2 Chiral Capillary Electrophoresis (CCE)

CCE allows for the performance of chiral separations, adding neutral or charged chi-
ral selectors to the BGE without the stationary phases of chiral liquid chromatography
(LC). In the food field, it is mainly used for the separation of racemic amino acids, as
potential precursors of food aroma compounds (Belitz, Grosch, and Schieberle, 2009;
Acunha et al., 2016). Chiral selectors have different structures (cyclodextrins (CDs),
crown ethers, proteins, and macrocyclic antibiotics). In CDs, which are cyclic oligosac-
charides, and crown ethers, the presence of a cavity favors the formation of a hydropho-
bic or hydrophilic complexation, respectively (Rizzi, 2001; Tsioupi, Stefan-Vanstaden,
and Kapnissi-Christodoulou, 2013). CDs and their derivatives are the most widely used
selectors because of high BGE solubility, no or low absorption in the UV range, and high
versatility. It is also important to setup the selector concentration. This parameter influ-
ences the effective mobilities and as a consequence the method’s selectivity (Rizzi, 2001;
Tsioupi, Stefan-Vanstaden, and Kapnissi-Christodoulou, 2013).

10.4.3 Capillary Electrochromatography (CEC)

CEC is a hybrid technique of CE and LC; in fact, the capillary contains a stationary phase
and consequently two principles, such as partition and mobility, drive the separation.
This technique finds application in the analysis of aromatic compounds (Svec, 2004).
The development of new coating polymers or materials has considerably increased CEC
specificity and versatility (Iacob, Bodoki, and Oprean, 2014; Tarongoy, Haddad, and
Quirino, 2018).

10.4.4 Non-Aqueous Capillary Electrophoresis (NACE)

NACE is a CE mode in which organic solvents, such as methanol, ethanol, and acetoni-
trile, are added to the BGE. It is useful for analytes with very low solubility in aqueous
solutions. This addition improves CZE selectivity. NACE can be used as an alternative to
MEKC (Pinero, Bauza, and Arce, 2011; Kenndler, 2014).
Other CE modes, such as capillary gel electrophoresis (CGE) or capillary isotacho-
phoresis (cITP) and capillary isoelectric focusing (cIEF), not treated here, are specific
for DNA or peptides and proteins, respectively. In these CE modes, running buffers are
replaced by other solutions (gel, particular buffers, or ampolytes) (Pinero, Bauza, and
Arce, 2011; Karlinsey, 2012; Acunha et al., 2016; Papetti and Colombo, in press). Table
10.1 illustrates different CE modes with principles, characteristics, and applications.
 Food Aroma Compounds by Capillary Electrophoresis 199

10.5 ADVANTAGES AND LIMITATIONS

On-capillary detection contributes to an increase in peak efficiency, which is also ensured

by the geometry of the capillary. In fact, this induces the creation of a uniform flow,
contrary to the laminar flow of LC. Analysis time in CE is short (10–20 min); the small
volumes (injection and capillary) allow for the low consumption of the sample and buffer,
and the free-solution system gives the benefit of environmental compatibility. In addition,
the high surface area to volume ratio minimizes the Joule heating effect and partially
resolves the problem of reproducibility. The problem of errors in quantitative analysis,
due to the different migration velocities of the solutes, can be resolved. In fact, the over-
estimation of the peak area of solutes with low mobility and the underestimation of the
peak area for those with high mobility are normalized considering the peak area divided
by the migration time (area normalization) (Camilleri, 1997).
The possibility of modulating many parameters, such as capillary type and length, buf-
fer (pH, type, and ionic force), injection type and values, voltage, temperature, conditioning
type and time, and the possibility of applying different separation modes, provides a great
potential for a wide range of applications (ions, drugs, proteins, natural products, and food-
related complex molecules from amino acids to lipids, carbohydrates, DNAs, and vitamins)
(Reijenga et al., 1996; Pinero, Bauza, and Arce, 2011; Papetti and Colombo, in press).
Poor sensitivity remains the only limit, which depends on the small light path (~30 µm
with an internal diameter of 50 µm), but it can be enhanced, not only by using appropriate
detectors, but also by using capillaries with an extended path length or offline or online
preconcentration procedures (Camilleri, 1997; Whatley, 2001; Breadmore et al., 2017).
Table 10.2 illustrates the specific advantages and limitations of each CE-mode treated here.

10.6 APPLICATIONS

10.6.1 Non-Enzymatic Reactions

These reactions, such as lipid peroxidation, the Maillard reaction (for example cara-
melization), and amino acid degradation, if occurring at room temperature, influence
aroma compounds only during long storage. On the contrary, heat treatment increases
kinetics and aroma diversity in the case of roasting and frying. This type of reaction
can be the source of a great number of volatile compounds, but they are usually in small

TABLE 10.2 Pros and Cons of Different CE Modes

CE Mode Pros Cons
CZE Simplicity Limited to charged molecules
MEKC Neutral molecules Interaction of the analyte with
Improved analyte solubility surfactant
CCE Efficiency Setup of separative conditions
Low consumption of chiral selectors
CEC Efficiency Setup of BGE composition
Easy coupling with MS detectors high voltages
NACE Improved analyte solubility Variations in pKa values and mobility
Efficiency
Selectivity
200 Food Aroma Evolution

concentrations, so only a few generated compounds will be aroma active. The parameters
which mainly influence non-enzymatic reactions consist of precursor type and quantity,
temperature, and time (Belitz, Grosch, and Schieberle, 2009).

10.6.1.1 Amines
Amines are Strecker-products, and their formation is pH-dependent (Belitz, Grosch, and
Schieberle, 2009). Among them, 2-phenylethylamine is a flavoring agent used as a food
additive, and a lot of its derivatives represent synthetic adulterants of dietary supplements
(Pawar and Grundel, 2017). He et al. (2016) demonstrated the application of a CZE-UV
method for monitoring biogenic amines, 2-phenylethylamine included, in biological sam-
ples, and this method could also be applied in food aroma analysis.

10.6.1.2 Furanones
Among furanones, which are secondary products of the Maillard reaction, 4-hydroxy-
2,5-dimethyl-3-(2H)-furanone, also known as strawberry furanone or furaneol, are
responsible for a caramel-like odor present in many food products, mainly in fruit (pine-
apple, strawberry), boiled beef, and medium roasted coffee. It is a racemic compound,
and CCE can be a powerful technique. A CCE method was set up to monitor the racemi-
zation rate in different pH storage conditions (Raab et al., 2003). Also, 3-hydroxy-4,5-di-
methyl-2-(5H)-furanone, named Sotolon, is a racemic furanone; its enantiomers do not
differ in aroma quality (caramel), and it is typical of coffee and sherry. Taga et al. (2012)
analyzed this molecule using CZE-UV, without considering enatiomers, but with the aim
of monitoring Sotolon as a food additive. Without any sample preparation, in comparison
with LC and gas chromatography (GC) techniques, it was possible to quantify ppm levels.

10.6.1.3 Phenols
These compounds can be derived from phenolic acids and lignin in the presence of heat
or microorganisms. They are present in food and food products, such as meat, coffee,
milk, beer, whiskey, cooked apple, and asparagus. They give a different aroma quality,
mainly smoky, depending on the type of phenol. For example, p-cresol, which is respon-
sible for the smoky aroma of coffee, milk, and roasted peanuts, can be analyzed using
CEC. Notwithstanding, the focus of this work was not to analyze aroma compounds but
to determine contaminated samples, this method can rapidly and efficiently obtain the soil
content of phenols, p-cresol included, after a sample preconcentration step with supercriti-
cal fluid extraction (Fung and Long, 2001). Among phenols in food, 4-ethylphenol, with a
woody aroma, can be detected by CZE. This study aimed to determine different bisphenols
and 4-ethylphenol as contaminants in plastic water bottles. A solid phase extraction pro-
cedure prior to analysis and an AD also allowed for the quantification of traces (Browne et
al., 2013). Vanillin with a vanilla aroma characterizes vanilla, butter, rum, coffee, cooked
asparagus, and can be analyzed using MEKC-UV. The setup method with an optimization
of buffer concentration, pH, and modifier has been useful for rapidly detecting and quanti-
fying (5–500µg/mL) four flavor compounds (vanillin, ethylvanillin 2-methoxyphenol, and
2-ethoxyphenol) in cocoa drinks (Ohashi et al., 2007). Also CZE methods without a sam-
ple pre-treatment (Panossian et al., 2001; Lahouidak et al., 2018) or with simple online pre-
concentration steps (Heller et al., 2011) rapidly allowed for the quantification of aldehydes,
vanillin included, in brandy, whiskey, and wine. This determination is very important for
recognizing aged or counterfeit products (these products contain only vanillin and no other
aldehydes) (Panossian et al., 2001). Lahouidak et al. (2018) recently set up a method able to
simultaneously quantify natural, synthetic, and prohibited flavoring compounds. It allowed
 Food Aroma Compounds by Capillary Electrophoresis 201

FIGURE 10.2 Comparison among CE profiles of a natural vanilla extract (A) and
non-natural vanilla products (B–D) to determine vanilla flavors for food authenticity
(COUM, coumarin; EVA, ethyl vanillin; VAN, vanillin; PHB, p-hydroxybenzaldehyde;
VANA, vanillic acid; PHBA, p-hydroxybenzoic acid). (Reprinted with permission from
Lahouidak, S., Salghi, R., Zougagh, M., and Ríos, A. (2018). Capillary electrophoresis
method for the discrimination between natural and artificial vanilla flavor for controlling
food frauds. Electrophoresis, 39, 1628–33. doi: 10.1002/elps.201700480.)

for the rapid gain of a complete fingerprint of vanilla markers, discriminating between van-
illin and artificial vanilla compounds (vanillic acids, ethyl vanillin), and adulterants, such
as coumarin, as illustrated in Figure 10.2. For the rapid determination of vanilla-related
flavors in food fraud, a CE microchip coupled with electrochemical detectors can also be
applied (Avila et al., 2007).
A chiral MEKC-PDA method was applied to the analysis of catechins in tea infusions
obtained using tea leaves stored in different conditions (Mirasoli et al., 2014). In this
work, BGE was added with sodium dodecyl sulfate and (2-hydroxypropyl)-β-cyclodextrin
as a chiral selector, in order to identify (+)-catechin, epicatechin, epigallocatechin, epi-
catechin gallate, epigallocatechingallate, and caffeine. In aged samples, catechin content
decreased or increased in relation to the storage time.

10.6.1.4 Pyranones
Maltol, with its vanilla-related flavor, represents a product obtained from carbohydrates,
which increases the sweet taste in food; it is found mainly in roasted coffee, heated butter,
and biscuits. Its synthetic products, which are sweeter than maltol, are used extensively
in food aromatization (Belitz, Grosch, and Schieberle, 2009). Avila et al. (2007) set up
a CE-microchip device with an ED for food authenticity, discriminating simultaneously
between maltol and ethyl maltol, together with vanillin, ethyl vanillin, and vanillic alcohol.

10.6.1.5 Pyrazines
These compounds are both synthesized and degraded by a few types of bacteria and
fungi and formed during the heating of food. They contribute to odors, characterizing
202 Food Aroma Evolution

roasted food, such as coffee, beef products, bread, corn, and fried potatoes, and for
this reason they are often used as flavoring additives in the modern-day food indus-
try (Müller and Rappert, 2010; Migita et al., 2017). In the literature there are no
CE-methods applied to volatile pyrazines, but some CE-modes, which have been setup
in different food matrices to analyze other aromatic eterocyclic compounds, such as for
example triazines, can be adapted for this purpose (Elbashir and Aboul-Enein, 2015).
CZE (Arribas et al., 2011) and MEKC (Fang et al., 2014) are the most powerful tech-
niques; in addition, CEC (Chang et al., 2006) and NACE (Carabias-Martínez et al.,
2006) can also be potential tools.
10.6.1.6 Pyridines
Pyridine derivatives are the source of a typical roasted/cracker-like odor, typical of bread,
cooked meat, and a particular type of rice (Mirasoli et al., 2014; Wei et al., 2017). Their
amount increases in a temperature range of 50–75°C over time (1 hour). Pyridines, such
as 2-acetylpyridine, have a low UV absorbance, but with a preconcentration step CZE
could also be a potential technique of analysis. Hattori et al. (2017) setup a method
in which CZE was used to analyze pyridines, after a preconcentration step, obtained
through transient isotachophoresis.

10.6.1.7 Thiols, Thioethers, and Di- and Trisulfides

These compounds can be generated following heat or long storage times, starting mainly
from amino acids (Cys and Met) or monosaccharides. They are responsible for a roasted,
toasted, and meat-like odor, but also a sulfurous and cabbage-like aroma. Among them,
2-methyl-3-furanthiol, which characterizes the typical meat odor of fermented soy sauce
(Meng et al., 2017) and wine (Tominaga and Dubourdieu, 2006), represents an interest-
ing sulfur compound, nowadays analyzed by GC; however, CE-MS could also be a poten-
tial technique for quantifying it, because of its very low odor threshold.

10.6.1.8 Amino Acids as Precursors of Food Aroma Compounds

In some foods, as for example grape, wine, and vinegar, amino acids represent important
precursors of aroma compounds, such as alcohols, aldehydes, esters, and ketonic acids,
which are responsible for important organoleptic properties (taste, aroma, and color).
In general, it is not particularly easy to analyze amino acids with proper sensitivity, but
the use of LIF and electrochemical detectors can overcome this disadvantage (Callejón,
Troncoso, and Morales, 2010).
Amino acids, such as glycine, alanine (Ala), valine (Val), leucine (Leu), isoleucine (Ile),
and phenylalanine (Phe), are precursors of carbonyl compounds and can undergo deg-
radation, giving origin to Strecker aldehydes, responsible for a malty aroma. A Strecker
reaction also involves methionine (Met), Val, Leu, Ile, and Phe, which become precursors
of amines, giving rise to a malty/fishy odor (Belitz, Grosch, and Schieberle, 2009).
In general, amino acids and racemic amino acids in food are an important marker
of authenticity, quality, and origin and can be analyzed by CZE, MEKC, and CCE to
monitor the formation of aroma substances (Karlinsey, 2012; Viglio et al., 2012; Tsioupi,
Stefan-Vanstaden, and Kapnissi-Christodoulou, 2013; Poinsot et al., 2018). In particular,
Val, the precursor of methylpropanal and Leu and Ile, precursors of 3-methylbutanal and
2-methylbutanal, respectively, can be rapidly quantified using a setup CCE method with
indirect UV detection (Qiu et al., 2017).
Cysteine (Cys) and Met are precursors of sulfurous compounds (Belitz, Grosch, and
Schieberle, 2009) and total Cys or Met content can be rapidly monitored by CZE-UV
 Food Aroma Compounds by Capillary Electrophoresis 203

with uncoated capillaries (Kubalczyk and Bald, 2009) or innovative coated capillaries
(Vitali et al., 2014). Ala under heating can also be a precursor of odorous pyrazines
(Belitz, Grosch, and Schieberle, 2009).

10.6.1.9 Other Substances as Precursors of Aroma Compounds

Thiamine, which is a precursor of sulfurous compounds (i.e., 2-methyl-3-furanthiol), is a
water-soluble vitamin, and a potential candidate for a technique in a free solution, such
as CZE (Belitz, Grosch, and Schieberle, 2009). It can be analyzed using CZE-UV (Vitali
et al., 2014) and CZE-ESI-MS for increasing method selectivity and sensitivity (Trenerry,
2001; Maráková et al., 2014).
Among other precursors of sulfurous molecules (thiols, thioethers, di-, and trisul-
fides), monosaccharides can also be analyzed by CE. Sugars, such as ribose, but also
rhamnose and glucose, in the presence of Cys and under a heating process, are the source
of 3-mercapto-2-pentanone, 2-methyl-3-furanthiol, and 2-furfurylthiol with a charac-
teristic meat aroma (Belitz, Grosch, and Schieberle, 2009). Saccharides (ribose and glu-
cose) are also food additives, mainly present in juices, soft and high-energy drinks, and
wines; they can be easily and rapidly detected and quantified by CE-UV (Rovio, Yli-
Kauhaluoma, and Sirén, 2007) or by CE-C4D, improving CE-sensitivity (Tůma et al.,
2011; Vochyánová et al., 2012). Recently, a CE-MS was set up to rapidly analyze the
profile of monosaccharides (ribose, rhamnose, and glucose included) as a signal in coffee
samples adulterated with soybean and corn (Daniel et al., 2018).

10.6.2 Enzymatic Reactions

Enzymes can be indirectly involved in food aroma formation and enhancement; in fact,
after tissue disruption, they could release precursor compounds (e.g., ortho-quinone
structures from phenolic compounds or amino acids from proteins or sugar from poly-
saccharides) which are then converted in aroma compounds by further non-enzymatic
reactions (Belitz, Grosch, and Schieberle, 2009).

10.6.2.1 Carbonyl Compounds

Ketones and aldehydes have a sensory relevance in foodstuff as they can both improve
food quality by generating a pleasant flavor and odor, and also indicate food deteriora-
tion, bacterial fermentation, and off-flavors. Therefore, their detection attracts a growing
relevance and CE with UV detection has emerged in the last few years due to its main
known advantages. A derivatization step is always needed to avoid losses of the carbonyl
compounds. Among aldehydes, cis and trans 2-hexenal, possessing the odor of freshly cut
grass and leaves, have been determined together with acetaldehyde in different yogurts,
juices, and yogurt–juice mixtures using a fully automated approach based on pervapora-
tion coupled online with MEKC using a flow injection manifold and the replenishment
system of a CE apparatus (Ruiz-Jiménez and Luque de Castro, 2006). Figure 10.3 illus-
trates this combined approach.
The integration of a pervaporation step in an analytical process other than sepa-
ration represents a good alternative to headspace static or dynamic systems (Bryce,
Izquierdo, and Luque de Castro, 1997). Furthermore, this approach is time-saving as
the removal of analytes from the matrix, their preconcentration, and derivatization take
place in the acceptor chamber of the evaporator and require about 10 min, and the sep-
aration of the analytes in CE can be simultaneous with the next sample pervaporation.
204 Food Aroma Evolution

FIGURE 10.3 Setup of a pervaporation and CE interface to determine volatile aldeydes

in slurry samples, as yogurt and juice (AS, airstream; WS, water stream; SS, sample
stream; PP, peristaltic pump; DR, derivatisation reagent; SV, selection valve; IV, injec-
tion valve; PH, pre-heater; M, membrane; PU, pervaporation unit; WB, water bath;. W,
waste). (Reprinted with permission from Ruiz-Jiménez, J., and Luque de Castro, M. D.
(2006). Online pervaporation-capillary electrophoresis for the determination of volatile
analytes in food slurries. Journal of Chromatography A, 1128, 251–8. doi:10.1016/j.
chroma.2006.06.031.)

The validated CZE-PDA method improves sensitivity by 10 times. Acetaldehyde can be

considered as an indicator of off-flavors and is classified as toxic by the International
Agency for Research on Cancer (1985). A novel home-made miniaturized CZE-AD
system was proposed for its fast detection in wine and waterlogged samples by D.
Zhang et al. (2011). 2-thiobarbituric acid was selected as a derivatization agent as it is
an electro-attractive species that easily originates adducts, thus facilitating the deter-
mination of acetaldehyde. This derivatization agent was also used for the detection
of other aldehydes, such as hexanal, 2,3-butanedione (diacetyl), and methyglyoxal, in
different food matrices, that is, olive and sunflower seed oils, white, red and glutinous
rice wines, and water-soaked products (sea cucumber, jelly fish, and tendons of beef)
using CZE-AD. The home-built three-electrode electrochemical cell which was used
consisted of a 300 mm diameter carbon disk working electrode, a platinum auxiliary
electrode, and a saturated calomel electrode as the reference electrode in combination
with a BAS LC-3D amperometric detector. The developed method was simple, accu-
rate, and useful for the analysis of aldehydes in many different food matrices. Figure
10.4 shows the detection of acetaldehyde and formaldehyde through the formation of
adducts with 2-thiobarbituric acid (J.-B. Zhang et al., 2011). Another derivatization
agent, 4-hydrazinobenzoic acid (the primary terminal amine in this hydrazine reacts
with the carbonyl group forming an imide), was recently used in the development of a
CZE-PDA method for the analysis of different wine samples. A gas-diffusion microex-
traction method was applied, thus obtaining a pre-concentrated and cleaned-up (only
volatile and semi-volatile compounds are extracted) sample. This methodology showed
good results regarding linearity and precision (de Lima et al., 2018). Benzhydrazide,
a water-soluble substance, was used as a derivatizing agent for the determination of
aldehydes in yogurt and vinegar using MEKC-PDA. The ­derativization reaction is fast,
simple, not strongly pH- and temperature-dependent, giving rise to only one derivative
for each aldehyde which is stable up to 3 hours (Donegatti, Moreira Gonçalves, and
Pereira, 2017). Methylgyoxal was also quantified, even at low concentrations, in tap
water and beer samples using an optimized CZE-PDA method; o-phenylendiamine was
 Food Aroma Compounds by Capillary Electrophoresis 205

FIGURE 10.4 CZE-ED profiles of the standard mixture (A), Chinese white liquor (B),
and white wine (C). 1, acetaldehyde-2-thiobarbituric acid adduct; 2, formaldehyde-
2-thiobarbituric acid adduct; 3, 2-thiobarbituric acid. (Adapted with permission from
Zhang, D., Zhang, J., Li, M., Li, W., Aimaiti, G., Tuersun, G., Ye, J., and Chu, Q. (2011).
A novel miniaturized electrophoretic method for determining formaldehyde and acetal-
dehyde in food using 2-thiobarbituric acid derivatization. Food Chemistry, 129, 206–12.
doi:10.1016/j.foodchem.2011.04.025.)

used as a derivatizing agent and a C18 SPE as a purification step prior to the analysis
(do Rosário et al., 2005). An environmentally friendly miniature CZE-AD method
developed for the trace quantification of four aliphatic aldehydes present in water sam-
ples, as disinfection by-products, also provided a good separation of glyoxal and meth-
ylglyoxal after derivatization with 2-thiobarbituric acid. This new method was based
on thee hollow-fiber liquid-phase microextraction of the analytes (Li et al., 2015).
206 Food Aroma Evolution

10.6.2.2 Terpenes
Terpenes are generally analyzed using gas chromatography. In the last two decades, only
a CEC-PDA method has been developed for the detection and quantification of 11 aro-
matic and terpenic compounds in extracts of the spices from Piper nigrum. A C18 sor-
bent was the best sorbent for packing the fused-silica capillary and a mixture of 50 mM
ammonium acetate solution (pH 6.0) and ACN, 10:90, v/v, the best mobile phase for the
separation of all the tested analytes (Musenga et al., 2007).

10.6.2.3 Other Substances as Precursors of Aroma Compounds

Allium genus vegetables are rich in S-alk(en)yl-l-cysteine-sulfoxides, the precursor of
the typical odor and flavor of these vegetables. Methyl-cysteine sulfoxide (methiin)
and 2-propenyl-cysteine sulfoxide (alliin) have a special flavor which is called kakumi.
As both of these compounds have no specific UV absorbance, an electrolyte for indi-
rect detection was selected, and the developed CE-UV method did not require any
derivatization (but only vegetable boiling, extraction, dilution, and filtration); it was
simple and required less than 25 min for each analysis (Horie and Yamashita, 2006).
Another method also applicable to the analysis of isoalliin, propiin (S-propylcysteine-
S-oxide), ethiin (S-ethylcysteine-S-oxide), and butiin (S-butylcysteine-S-oxide) was
setup by Kubec and Dadáková (2008). The methanolic extracts of Allium and Brassica
vegetables were derivatized with fluorenylmethyl chloroformate, and subsequently
separated by MECK-UV. Simplicity, sensitivity, high specificity, and very low run-
ning costs, make this method suitable for the routine analysis of large numbers of
samples. Glucosinolates (alkyl-N-hydroximine esters with a β-dthioglucopyranoside
group linked to the hydroximine carbon in Z configuration to the sulfate group) are
precursors of isothiocyanates (ITCs), generated by the action of enzyme myrosinase;
ITCs are responsible for the pungent sensory characteristics of Brassicaceae vegeta-
ble food, and they are easily recognized. These compounds are not easily detectable
in UV but can still be detected after derivatization, a process not compatible with
myrosinase measurements. Therefore, Bellostas and co-workers developed a micel-
lar electrokinetic capillary chromatography for monitoring the myrosinase catalyzed
hydrolysis of 2-hydroxy substituted glucosinolates (progoitrin, glucosibarin, gluco-
barbarin, glucotropaeolin, and gluconasturtiin), and the simultaneous formation of
the corresponding degradation products which are not ITCs at all (oxazolidine-2-thi-
ones and nitriles) (Bellostas, Sørensen, and Sørensen, 2006). Recently, a fast, robust,
and simple MECK method was developed and validated for the simultaneous quan-
tification of glucosinolates and ITCs by Gonda et al. (2016). The low detectability of
ITCs in UV was overcome using in-vial derivatizing with mercaptoacetic acid, with-
out inhibiting the enzyme activity. Different Brussel sprouts, horseradish, watercress,
and radish samples were successfully analyzed, and sinigrin, gluonasturtiin, and allyl
isothiocyanate quantified. This method can be used not only for glucosinolate deter-
mination, but also for myrosinase activity measurement, and isothiocyanate release
estimation.

10.6.2.4 Phenols as Precursors of Food Aroma Compounds

2,4,6-trichloroanisole, usually produced by naturally occurring airborne fungi and bacte-
ria (Aspergillus sp., Penicillium sp., Actinomycetes, Botrytis cinerea, and Streptomyces),
is the main component responsible for cork taint in wines; 2,4,6-trichlorophenol is its
 Food Aroma Compounds by Capillary Electrophoresis 207

TABLE 10.3 Food Aroma Compounds Detected by CE Techniques

Aroma Type of
Food Aroma Compound Description Food CE Mode
Furaneol Strawberry- Fruit CCE-PDA
like Beer (Raab et al., 2003)
Pineapple- Coffee
like (medium
Caramel roasted)
Boiled beef
White bread
Emmental
cheese
Sotolon Caramel Coffee CZE-PDA
Sherry (Taga et al., 2012)

p-Cresol Smoky Coffee CEC-PDA

Sherry (Fung and Long,
Milk 2001)
Roasted
peanuts
Asparagus

4-Ethylphenol Woody Coffee CZE-AD

Milk (Browne et al., 2013)
Tomatoes,
Roasted
peanuts
Soya sauce

(Continued )
208 Food Aroma Evolution

TABLE 10.3 (Continued) Food Aroma Compounds Detected by CE Techniques

Aroma Type of
Food Aroma Compound Description Food CE Mode
Vanillin Vanilla Vanilla MEKC-PDA (Ohashi
Coffee et al., 2007)
Rum CZE-PDA (Panossian
Whiskey et al., 2001; Heller et
Asparagus al., 2011; Lahouidak
Cooked et al., 2018)
butter CE-microchip-ED
(Avila et al., 2007)

Catechins Bitter Tea Chiral MEKC-PDA

*–H = (–)-Catechin (Mirasoli et al., 2014)
–OH = (–)-Gallocatechin

**–H=Epicatechin
–OH=Epigallocatechin

***–H=Epicatechin gallate
–OH=Epigallocatechin gallate

(Continued )
 Food Aroma Compounds by Capillary Electrophoresis 209

TABLE 10.3 (Continued) Food Aroma Compounds Detected by CE Techniques

Aroma Type of
Food Aroma Compound Description Food CE Mode
Maltol Caramel Roasted CE-microchip-ED
coffee (Avila et al., 2007)
Biscuit
Cooked
butter
Chocolate
Beer

2-Acetylpyridine Roasted White bread CZE-PDA (Hattori et

al., 2017)

2-Hexenal Freshly cut Yogurt MEKC-PDA (Ruiz-

grass/ Juice Jiménez and Luque
leaves Wine de Castro, 2006)
Fruity Sherry CZE-ED/AD (D.
Zhang et al., 2011;
J.-B. Zhang et al.,
Acetaldehyde 2011)

Methylglioxal Cheesy Wine CZE-PDA (do Rosário

Beer et al., 2005)
Biscuit
Bread

Terpenic compounds Woody Fruit CEC-PDA (Musenga

Vegetables et al., 2007)
Jam
Wine

precursor and it has been detected in wines using a headspace single drop microextrac-
tion automatically inline coupled with CE. This extraction becomes a very promising
sample pre-treatment technique when a single drop hanging at the inlet tip of a capillary
for CE is used as the acceptor phase; in fact, high enrichment factors were obtained in
a short time. Furthermore, a headspace single drop microextraction performed before
210 Food Aroma Evolution

the sample injection and a large volume sample stack using an electroosmotic flow
pump after the sample injection leads to the achievement of enrichment factors of sev-
eral thousand-fold, with LODs in the nanomolar range for chlorophenols in wine (Park
et al., 2012).
Table 10.3 illustrates a summary of published CE applications in food aroma
compounds.

REFERENCES

Acunha, T., Ibáñez, C., García-Cañas, V., Simó, C., and Cifuentes, A. (2016). Recent
advances in the application of capillary electromigration methods for food analysis
and foodomics. Electrophoresis, 37, 111–41. doi:10.1002/elps.201500291.
Arribas, A.S., Moreno, M., Bermejo, E., Zapardiel, A., and Chicharro, M. (2011).
CZE separation of amitrol and triazine herbicides in environmental water sam-
ples with acid-assisted on-column preconcentration. Electrophoresis, 32, 275–83.
doi:10.1002/elps.201000371.
Avila, M., González, M.C., Zougagh, M., Escarpa, A., and Ríos, A. (2007). Rapid sample
screening method for authenticity controlling vanilla flavors using a CE microchip
approach with electrochemical detection. Electrophoresis, 28, 4233–9. doi:10.1002/
elps.200700277.
Belitz, H.-D., Grosch, W., and Schieberle, P. (2009). Aroma compounds. In H.-D.
Belitz, W. Grosch, and P. Schieberle (Eds.), Food Chemistry (pp. 360–89). Berlin:
Springer-Verlag.
Bellostas, N., Sørensen, J.C., and Sørensen, H. (2006). Micellar electrokinetic capillary
chromatography—Synchronous monitoring of substrate and products in the myros-
inase catalysed hydrolysis of glucosinolates. Journal of Chromatography A, 1130,
246–52. doi:10.1016/j.chroma.2006.05.079.
Breadmore, M.C., Wuethrich, A., Li, F., Phung, S.C., Kalsoom, U., Cabot, J.M.,
Tehranirokh, M., Shallan, A.I., Abdul Keyon, A.S., See, H.H., Dawod, M., and
Quirino, J.P. (2017). Recent advances in enhancing the sensitivity of electropho-
resis and electrochromatography in capillaries and microchips (2014–2016).
Electrophoresis, 38, 33–59. doi:10.1002/elps.201600331.
Browne, D.J., Zhou, L., Luong, J.H., and Glennon, J.D. (2013). CE with a boron-doped
diamond electrode for trace detection of endocrine disruptors in water samples.
Electrophoresis, 34, 2025–32. doi:10.1002/elps.201200480.
Bryce, D.W., Izquierdo, A., and Luque de Castro, M.D. (1997). Pervaporation as an alter-
native to headspace. Analytical Chemistry, 69, 844–7. doi:10.1021/ac960456s.
Callejón, R.M., Troncoso, A.M., and Morales, M.L. (2010). Determination of amino
acids in grape-derived products: A review. Talanta, 81, 1143–52. doi:10.1016/j.
talanta.2010.02.040.
Camilleri, P. (1997). Capillary Electrophoresis: Theory and Practice. Boca Raton, FL:
CRC Press.
Carabias-Martínez, R., Rodríguez-Gonzalo, E., Miranda-Cruz, E., Domínguez Álvarez,
J., and Hernández-Méndez, J. (2006). Comparison of a non-aqueous capillary elec-
trophoresis method with high performance liquid chromatography for the determi-
nation of herbicides and metabolites in water samples. Journal of Chromatography
A, 1122, 194–201. doi:10.1016/j.chroma.2006.04.017.
 Food Aroma Compounds by Capillary Electrophoresis 211

Chang, C.H., Chen C.J., Chuang, Y.C., and Her, G.R. (2006). Analysis of triazines by
capillary electrochromatography/electrospray ionization–mass spectrometry using
a low-flow sheath liquid interface. Electrophoresis, 27, 4303–11. doi:10.1002/
elps.200600251.
Daniel, D., Lopes, F.S., Santos, V.B.D., and do Lago, C.L. (2018). Detection of coffee
adulteration with soybean and corn by capillary electrophoresis-tandem mass spec-
trometry. Food Chemistry, 243, 305–10. doi:10.1016/j.foodchem.2017.09.140.
de Lima, L.F., Brandão, P.F., Donegatti, T.A., Ramos, R.M., Moreira Gonçalves, L.,
Cardoso, A.A., Pereira, E.A., and Rodrigues, J.A. (2018). 4-hydrazinobenzoic acid
as a derivatizing agent for aldehyde analysis by HPLC-UV and CE-DAD. Talanta,
187, 113–9. doi:10.1016/j.talanta.2018.04.091.
do Rosário, P.M.Á., Cordeiro, C.A.A., Freire, A.P., and Nogueira, J.M.F. (2005). Analysis
of methylglyoxal in water and biological matrices by capillary zone electrophoresis
with diode array detection. Electrophoresis, 26, 1760–7.
Donegatti, T.A., Moreira Gonçalves, L., and Pereira, E.A. (2017). Derivatizing assay
for the determination of aldehydes using micellar electrokinetic chromatography.
Electrophoresis, 38, 1068–74. doi:10.1002/elps.201600483
Elbashir, A., Schmitz, O.J., and Aboul‐Enein, H.Y. (2017). Application of capillary elec-
trophoresis with capacitively coupled contactless conductivity detection (CE‐C4D):
An update. Biomedical Chromatography, 31, e3945. doi:10.1002/bmc.3945.
Elbashir, A.A. and Aboul-Enein, H.Y. (2015). Separation and analysis of triazine herbi-
cide residues by capillary electrophoresis. Biomedical Chromatography, 29, 835–
42. doi:10.1002/bmc.3381.
Fang, R., Chen, G.H., Yi, L.X., Shao, Y.X., Zhang, L., Cai, Q.H., and Xiao, J. (2014).
Determination of eight triazine herbicide residues in cereal and vegetable by micel-
lar electrokinetic capillary chromatography with online sweeping. Food Chemistry,
145, 41–8. doi:10.1016/j.foodchem.2013.08.028.
Fung, Y.S. and Long, Y.H. (2001). Determination of phenols in soil by supercritical fluid
extraction-capillary electrochromatography. Journal of Chromatography A, 907,
301–11. doi:10.1016/S0021-9673(00)01039-6.
Gonda, S., Kiss-Szikszai, A., Szűcs, Z., Nguyen, N.M., and Vasas, G. (2016). Myrosinase
compatible simultaneous determination of glucosinolates and allyl isothiocyanate
by Capillary Electrophoresis Micellar Electrokinetic Chromatography (CE-MEKC).
Phytochemical Analysis, 27, 191–8. doi:10.1002/pca.2615.
Hattori, T., Okamura, H., Asaoka, S., and Fukushi, K. (2017). Capillary zone electro-
phoresis determination of aniline and pyridine in sewage samples using transient
isotachophoresis with a system-induced terminator. Journal of Chromatography A,
1511, 132–7. doi:10.1016/j.chroma.2017.07.009.
He, L., Ren, J., Shi, Z., and Xu, Z. (2016). Separation of key biogenic amines by capillary elec-
trophoresis and determination of possible indicators of sport fatigue in athlete’s urine.
Journal of Chromatography Science, 54, 1428–34. doi:10.1093/chromsci/bmw065.
Heller, M., Vitali, L., Oliveira, M.A., Costa, A.C., and Micke, G.A. (2011). A rapid
sample screening method for authenticity control of whiskey using capillary electro-
phoresis with online preconcentration. Journal of Agricultural and Food Chemistry,
59, 6882–8. doi:10.1021/jf202218r.
Horie, H. and Yamashita, K. (2006). Non-derivatized analysis of methiin and alliin in
vegetables by capillary electrophoresis. Journal of Chromatography A, 1132, 337–
9. doi:10.1016/j.chroma.2006.09.018.
212 Food Aroma Evolution

Horvath, J. and Dolník, V. (2001). Polymer wall coatings for capillary electrophoresis.
Electrophoresis, 22, 644–55. doi:1​0.100​2 /152​2-268​3(200​102)2​2:4<6​44:AI ​D -ELP​
S644>​3.0.C​O;2-3​.
Iacob, B.C., Bodoki, E., and Oprean, R. (2014). Recent advances in capillary electrochro-
matography using molecularly imprinted polymers. Electrophoresis, 35, 2722–32.
doi:10.1002/elps.201400253.
Ibáñez, C., García-Cañas, V., Valdés, A., and Simó, C. (2013). Novel MS-based approaches
and applications in food metabolomics. Trends in Analytical Chemistry, 52, 100–
11. doi:10.1016/j.trac.2013.06.015.
International Agency for Research on Cancer (IARC). (1985). Allyl Compounds,
Aldehydes, Epoxides and Peroxides, IARC Monographs on the Evaluation of the
Carcinogenic Risk of Chemicals to Humans. Lyon, France. https​://mo​nogra​phs.i​
arc.f​r/iar​c-mon​ograp​hs-on​-the-​evalu​ation​-of-c​arcin​ogeni​c-ris​ks-to​-huma​ns-79​/.
Karlinsey, J.M. (2012). Electrophoresis. In Y. Picò (Ed.), Chemical Analysis of Food:
Techniques and Applications (pp. 375–405). Waltham, MA: Academic Press Elsevier.
Kenndler, E. (2014). A critical overview of non-aqueous capillary electrophoresis. Part
I: Mobility and separation selectivity. Electrophoresis, 1335, 16–30. doi:10.1016/j.
chroma.2014.01.010.
Kubalczyk, P. and Bald, E. (2009). Analysis of orange juice for total cysteine and gluta-
thione content by CZE with UV-absorption detection. Electrophoresis, 30, 2280–3.
doi:10.1002/elps.200800741.
Kubec, R. and Dadáková, E. (2008). Quantitative determination of S-alk(en)ylcys-
teine-S-oxides by micellar electrokinetic capillary chromatography. Journal of
Chromatography A, 1212, 154–7. doi:10.1016/j.chroma.2008.10.024.
Lahouidak, S., Salghi, R., Zougagh, M., and Ríos, A. (2018). Capillary electrophoresis
method for the discrimination between natural and artificial vanilla flavor for con-
trolling food frauds. Electrophoresis, 39, 1628–33. doi:10.1002/elps.201700480.
Li, Y., Yi, F., Zheng, Y., Wang, Y., Ye, J., and Chu, Q. (2015). Hollow-fiber liquid-phase
microextraction coupled with miniature capillary electrophoresis for the trace anal-
ysis of four aliphatic aldehydes in water samples. Journal of Separation Science, 38,
2873–9. doi:10.1002/jssc.201500323.
Maráková, K., Piestanský, J., Havránek, E., and Mikus, P. (2014). Simultaneous analysis
of vitamins B in pharmaceuticals and dietary supplements by capillary electropho-
resis hyphenated with triple quadrupole mass spectrometry. Pharmazie, 69, 663–8.
doi:10.1691/ph.2014.4020.
Martín, A., Vilela D., and Escarpa, D.A. (2012). Food analysis on microchip electrophore-
sis: An updated review. Electrophoresis, 33, 2212–27. doi:10.1002/elps.201200049.
Meng, Q., Kitagawa, R., Imamura, M., Katayama, H., Obata, A., and Sugawara, E.
(2017). Contribution of 2-methyl-3-furanthiol to the cooked meat-like aroma of
fermented soy sauce. Bioscience, Biotechnology and Biochemistry, 81, 168–72. doi
:10.1080/09168451.2016.1238295.
Migita, K., Iiduka, T., Tsukamoto, K., Sugiura, S., Tanaka, G., Sakamaki, G., Yamamoto,
Y., Takeshige, Y., Miyazawa, T., Kojima, A., Nakatake, T., Okitani, A., and
Matsuishi, M. (2017). Retort beef aroma that gives preferable properties to canned
beef products and its aroma components. Animal Science Journal, 88, 2050–6.
doi:10.1111/asj.12876.
Mirasoli, M., Gotti, R., Di Fusco, M., Leoni, A., Colliva, C., and Roda, A. (2014). Electronic
nose and chiral-capillary electrophoresis in evaluation of the quality changes in com-
mercial green tea leaves during a long-term storage. Talanta, 129, 32–8.
 Food Aroma Compounds by Capillary Electrophoresis 213

Müller, R. and Rappert, S. (2010). Pyrazines: Occurrence, formation and biodegra-

dation. Applied Microbiology and Biotechnology, 85, 1315–20. doi:10.1007/
s00253-009-2362-4.
Musenga, A., Mandrioli, R., Ferranti, A., D’Orazio, G., Fanali, S., and Raggi, M.A.
(2007). Analysis of aromatic and terpenic constituents of pepper extracts by capil-
lary electrochromatography. Journal of Separation Science, 30, 612–9. doi:10.1002/
jssc.200600456.
Ohashi, M., Omae, H., Hashida, M., Sowa, Y., and Imai, S. (2007). Determination
of vanillin and related flavor compounds in cocoa drink by capillary electro-
phoresis. Journal of Chromatography A, 1138, 262–7. doi:10.1016/j.chroma.
2006.10.031.
Panossian, A., Mamikonyan, G., Torosyan, M., Gabrielyan, E., and Mkhitaryan, S.
(2001). Analysis of aromatic aldehydes in brandy and wine by high-performance
capillary electrophoresis. Analitycal Chemistry, 73, 4379–83. doi:10.1021/
ac0014818.
Papetti, A. and Colombo, R. (in press). High performance Capillary Electrophoresis for
food evaluation. In X. Wang and J. Zhong (Eds.), Evaluation Technologies for Food
Quality. Academic Press Elsevier.
Park, S.T., Kim, J., Choi, K. Lee, H.R., and Chung, D.S. (2012). Headspace-single
drop microextraction with a commercial capillary electrophoresis instrument.
Electrophoresis, 33, 2961–8. doi:10.1002/elps.201200317.
Pawar, R.S. and Grundel, E. (2017). Overview of regulation of dietary supplements in
the USA and issues of adulteration with phenethylamines (PEAs). Drug Testing and
Analysis, 9, 500–17. doi:10.1002/dta.1980.
Pinero, M.-Y., Bauza, R., and Arce, L. (2011). Thirty years of capillary electrophoresis
in food analysis laboratories: Potential applications. Electrophoresis, 32, 1379–93.
doi:10.1002/elps.201000541.
Poinsot, V., Ong-Meang, V., Ric, A., Gavard, P., Perquis, L., and Couderc, F. (2018).
Recent advances in amino acid analysis by capillary electromigration methods.
Electrophoresis, 39, 190–208. doi:10.1002/elps.201700270.
Qiu, J., Wang, J, Xu, Z., Liu, H., and Ren, J. (2017). Quantitation of underivatized
branched-chain amino acids in sport nutritional supplements by capillary electro-
phoresis with direct or indirect UV absorbance detection. PLOs One, 12, e0179892.
doi:10.1371/journal.pone.0179892.
Raab, T., Hauck, T., Knecht, A., Schmitt, U., Holzgrabe, U., and Schwab, W. (2003).
Tautomerism of 4-hydroxy-2,5-dimethyl-3(2H)-furanone: Evidence for its enanti-
oselective biosynthesis. Chirality, 15, 573–8. doi:10.1002/chir.10247.
Ravelo-Pérez, L.M., Asensio-Ramos, M., Hernández-Borges, J., and Rodríguez-
Delgado, M.A. (2009). Recent food safety and food quality applications of CE-MS.
Electrophoresis, 30, 1624–46. doi:10.1002/elps.200800670.
Reijenga, T.P.E.M., Verheggen, J.H.P.A., and Martens, F.M. Everaerts. (1996). Buffer
capacity, ionic strength and heat dissipation in capillary electrophoresis. Journal of
Chromatography A, 744, 147–53. doi:10.1016/0021-9673(96)00273-7.
Rizzi, A. (2001). Fundamental aspects of chiral separations by capillary electrophore-
sis. Electrophoresis, 22, 3079–106. doi:1​0.100​2 /152​2-268​3(200​109)2​2:15<​3079:​
AID-E​LPS30​79>3.​0.CO;​2-F.
Rovio, S., Yli-Kauhaluoma, J., and Sirén, H. (2007). Determination of neutral carbohy-
drates by CZE with direct UV detection. Electrophoresis, 28, 3129–35. doi:10.1002/
elps.200600783.
214 Food Aroma Evolution

Ruiz-Jiménez, J. and Luque de Castro, M.D. (2006). On-line pervaporation-capillary

electrophoresis for the determination of volatile analytes in food slurries. Journal of
Chromatography A, 1128, 251–8. doi:10.1016/j.chroma.2006.06.031.
Slater, G.W., Tessier, F., and Kopecka, K. (2010). The electroosmotic flow (EOF). In M.P.
Hughes and K.F. Hoettges (Eds.), Microengineering in Biotechnology, Methods in
Molecular Biology (pp. 121–34). New York, NY: Humana Press Inc.
Svec, F. (2004). Csaba Horvath’s contribution to the theory and practice of capillary
electrochromatography. Journal of Separation Science, 27, 1255–72. doi:10.1002/
jssc.200401906.
Taga, A., Sato, A., Suzuki, K., Takeda, M., and Kodama, S. (2012). Simple determination
of a strongly aromatic compound, Sotolon, by capillary electrophoresis. Journal of
Oleo Science, 61, 45–8. doi:10.5650/jos.61.45.
Tarongoy, F.M. Jr., Haddad, P.R., and Quirino, J.P. (2018). Recent developments in open
tubular capillary electrochromatography from 2016 to 2017. Electrophoresis, 39,
34–52. doi:10.1002/elps.201700280.
Terabe, S. (2009). Capillary separation: Micellar electrokinetic chromatography. Annual
Review of Analytical Chemistry, 2, 99–120. doi:1​0.114​6/ann​urev.​anche​m.1.0​31207​
.1130​05.
Tominaga, T. and Dubourdieu, D. (2006). A novel method for quantification of 2-methyl-
3-furanthiol and 2-furanmethanethiol in wines made from Vitis vinifera grape
varieties. Journal of Agricultural and Food Chemistry, 54, 29–33. doi:10.1021/
jf050970b.
Trenerry, V.C. (2001). The application of capillary electrophoresis to the analysis of vita-
mins in food and beverages. Electrophoresis, 22, 1468–78. doi:1​0.100​2 /152​2-268​
3(200​105)2​2:8<1​468:A​I D-EL​PS146​8>3.0​.CO;2​- Q.
Tsioupi, D.A., Stefan-Vanstaden, R.I., and Kapnissi-Christodoulou, C.P. (2013). Chiral
selectors in CE: Recent developments and applications. Electrophoresis, 34, 178–
204. doi:10.1002/elps.201200239.
Tůma, P., Málková, K., Samcová, E., and Štulík, K. (2011). Rapid monitoring of mono-
and disaccharides in drinks, foodstuffs and foodstuff additives by capillary electro-
phoresis with contactless conductivity detection. Analytica Chimica Acta, 698, 1–5.
doi:10.1016/j.aca.2011.04.055.
Viglio, S., Fumagalli, M., Ferrari, F., Bardoni, A., Salvini, R., Giuliano, S., and Iadarola,
P. (2012). Recent novel MEKC applications to analyze free amino acids in different
biomatrices: 2009–2010. Electrophoresis, 33, 36–47. doi:10.1002/elps.201100336.
Vitali, L., Della Betta, F., Costa, A.C., Vaz, F.A., Oliveira, M.A., Vistuba, J.P., Fávere,
V.Y., and Micke, G.A. (2014). New multilayer coating using quaternary ammonium
chitosan and κ-carrageenan in capillary electrophoresis: Application in fast analysis
of betaine and methionine. Talanta, 123, 45–53. doi:10.1016/j.talanta.2014.01.047.
Vochyánová, B., Opekar, B.F., Tůma, P., and Štulík, K. (2012). Rapid determinations of
saccharides in high-energy drinks by short-capillary electrophoresis with contact-
less conductivity detection. Analytical and Bioanalytical Chemistry, 404, 1549–54.
doi:10.1007/s00216-012-6242-x.
Wei, X., Handoko, D.D., Pather, L., Methven, L., and Elmore, J.S. (2017). Evaluation of
2-acetyl-1-pyrroline in foods, with an emphasis on rice flavor. Food Chemistry, 232,
531–44. doi:10.1016/j.foodchem.2017.04.005.
Weinberger, R. (2000). Practical Capillary Electrophoresis. Cambridge: Academic Press.
 Food Aroma Compounds by Capillary Electrophoresis 215

Whatley, H. (2001). Basic principles and modes of capillary electrophoresis. In J. Petersen

and A.A. Mohammad (Eds.), Clinical and Forensic Applications of Capillary
Electrophoresis (pp. 21–58). New York, NY: Humana Press.
Zhang, D., Zhang, J., Li, M., Li, W., Aimaiti, G., Tuersun, G., Ye, J., and Chu, Q. (2011).
A novel miniaturized electrophoretic method for determining formaldehyde and
acetaldehyde in food using 2-thiobarbituric acid derivatisation. Food Chemistry,
129, 206–12. doi:10.1016/j.foodchem.2011.04.025.
Zhang, J.-B., Li, M.-J., Li, W.-L., Chu, Q.-C., and Ye, J.-N. (2011). A novel capillary
electrophoretic method for determining aliphatic aldehydes in food samples using
2-thiobarbituric acid derivatization. Electrophoresis, 32, 705–11. doi:10.1002/
elps.201000533
 Chapter 11
Proton-Transfer-Reaction–
Mass Spectrometry
Iuliia Khomenko and Brian Farneti

CONTENTS

11.1 Importance of Direct Injection Mass Spectrometry Analysis of Food VOCs 217
11.2 PTR-MS Technology 219
11.2.1 Drift Tube 221
11.2.2 Switchable Reagent Ion 222
11.2.2.1 Mass Analyzers 223
11.2.2.2 Quadrupole Mass Spectrometer (QMS) 223
11.2.2.3 Time-of-Flight–Mass Spectrometer (ToF-MS) 223
11.2.2.4 Data Analysis 224
11.3 PTR-MS Innovation 226
11.3.1 Toward Increasing of Sensibility 226
11.3.2 Toward High Throughputness 227
11.3.3 Toward Compound Identification 229
11.4 Application of PTR-MS Analysis in Food Science 230
11.4.1 Screening VOC Fingerprinting Analysis 230
11.4.2 Real-Time VOC Evolution Analysis 233
11.5 Challenges and Future Perspectives 233
References 234

11.1 IMPORTANCE OF DIRECT INJECTION MASS

SPECTROMETRY ANALYSIS OF FOOD VOCS

The current mission of the agro-food industry is to guarantee food safety and, at the same
time, improve perceived food quality and fulfill consumer expectations. To address these
issues, a broad and objective quality detection system for food products is needed. One of
the chief quality traits for the agro-food industry is the development of volatile organic com-
pounds (VOCs), associated with the shelf life and taste quality of food products. However,
the so-called “phenotyping bottleneck,” caused by the absence of high-throughput and
non-invasive methodologies, impedes an effective evaluation and prediction of food VOCs
(Furbank and Tester, 2011). The recent advancement of high-throughput and non-invasive
screening technologies stimulated the development of phenomics as a multidisciplinary
study, which links biophysics, biochemistry, and several “omic” analytical techniques, such
as transcriptomics, proteomics, and metabolomics. The latter focuses on the analysis of

217
218 Food Aroma Evolution

metabolites produced by the regulatory processes of the cell as a response of biological

systems to genetic or environmental changes (Fiehn, 2002). VOCs belong to secondary
metabolites. The ability to synthesize specific VOCs has been selected through the course of
evolution in different organisms for different purposes (Pichersky and Gang, 2000). VOCs
present in fruit, vegetables, meat, dairy products, and other foodstuffs are responsible for
their aroma, flavor, and taste. All these factors contribute to the consumer’s preferences
and perception of the food. The rapid development of mass spectrometry (MS) has made
possible the high-resolution characterization of several metabolites from complex matrices
in a single measurement, with a considerable impact on the field of VOC analysis (Herrero
et al., 2012). Cevallos-cevallos et al. (2009) classified metabolomic analyses into targeted
or untargeted. Targeted analyses focus on the identification and quantification of a deter-
mined array of metabolites. On the contrary, untargeted metabolomics put a spotlight on
the detection of as many groups of metabolites as possible without the need to precisely
quantify them. Ibáñez et al. (2013) distinguished two untargeted metabolomic analytical
approaches: (i) “metabolic profiling” refers to analysis of a class of metabolites (chemically
related metabolites or associated with a particular pathway); (ii) “metabolic fingerprinting”
refers to analysis of the total set of metabolites for rapid classification without identifying
individual metabolites. The complexity of the aroma footprint depends on the nonlinear
simultaneous interaction of a mixture of numerous molecules. Their fast and non-invasive
detection can be used for food quality control and for the monitoring of fundamental and
industrial processes (Biasioli et al., 2011a). Cumeras (2017) defined volatilomics as a part
of metabolomics, which focuses on the totality of VOCs produced by living organisms.
According to this, the overall study of aroma compounds in food can be considered as food
volatilomics. Thus, the term “volatilome” is determined as the entire VOC collection of a
sample. Recently this definition has been mentioned in human (Amann et al., 2014), plant
(Bicchi and Maffei, 2012), fruit (Farneti et al., 2017a, 2017b), bacteria (Casaburi et al.,
2015), and fungal (Li et al., 2016) studies. The extreme complexity and large variation of
VOC concentrations in food samples are challenging for any existing analytical technol-
ogy (Ibáñez et al., 2013). The rapid development of mass spectrometry and its application
in metabolomics have had a very significant impact in the field of VOC analysis (Herrero
et al., 2012). Recently, the progress of MS techniques has largely focused on instrumental
improvements to obtain a higher mass resolution, mass accuracy, sensitivity, and enhanced
reproducibility (Ibáñez et al., 2013). According to the techniques and strategies used to
transport VOCs to the instrumentation for further analysis, MS techniques can be divided
into gas chromatographic and direct injection. Direct injection mass spectrometric (DI-MS)
techniques (Biasioli et al., 2011b) have opened up new prospects for food aroma analysis
by decreasing the time needed for sample preparation and analysis. Different DI-MS tech-
niques have witnessed a flurry of developments providing the possibility for rapid, direct,
real-time, and high-throughput volatilome analysis (Huang et al., 2011). These approaches
do not require a chromatographic step prior to MS detection, allowing for the direct
analysis of samples (Ibáñez et al., 2013). DI-MS techniques differ in sample preparation,
sampling, inlet architecture, ionization processes, and detection (Biasioli et al., 2011b).
The most prominent examples are electronic noses, atmospheric-pressure chemical ion-
ization (APCI), direct analysis in real-time–mass spectrometry (DART-MS), ion mobility
spectrometry-mass–spectrometry (IMS-MS), selected ion-flow-tube–mass spectrometry
(SIFT-MS), and proton-transfer-reaction–mass spectrometry (PTR-MS). In the absence of
chromatographic separation, compound identification is only possible up to its chemical
formula and depends on ion chemistry and the resolution of the mass spectrometer (Nozie
et al., 2015). Proton-transfer-reaction–mass spectrometry coupled with a time-of-flight
 Proton-Transfer-Reaction–Mass Spectrometry 219

mass analyzer (PTR-ToF-MS) gives the possibility of detecting and quantifying VOCs in
a direct, simultaneous, and real-time approach at very low levels (pptv) with both high
mass and time resolution (Jordan et al., 2009a). A breakthrough in high-throughput in this
field took place when PTR-ToF-MS was connected for the first time to a multipurpose GC
automatic sampler (Yener et al., 2014). However, one of the major challenges of DI-MS
techniques, and particularly of PTR-MS, is the lack of separation of chemical isomers. For
this reason, a comparison with GC analysis is often mandatory. Nevertheless, there is a
growing interest in developing methods for improving the specificity of DI-MS methods
without losing sensitivity and time resolution.

11.2 PTR-MS TECHNOLOGY

Proton-transfer-reaction–mass spectrometry is a highly sensitive direct injection tech-

nique, developed in the early 1990s for analysis of volatile organic traces in ambient air
(Lindinger et al., 1998). PTR-MS belongs to a chemical ionization technique, which has
been used since the 1960s (Ellis and Mayhew, 2014). This technique became successful
due to its much softer ionization than those available at that time. This type of ionization
allows for the reduction of fragmentation and the observation of the molecular ion of
the compound of interest. The chemical ionization is achieved through various reactions
such as a simple charge transfer (Equation 11.1) or more complicated chemical reactions
(Equations 11.2–11.4).

X+ + M → M+ + X Charge transfer (11.1)

XH + + M → MH + + X Proton transfer (11.2)

X + + MH → M + + HX Hydride transfer (11.3)

X + + M + Z → MX + + Z Adduct formation (11.4)

Nowadays, protonated water (H3O+) is the most used reagent ion in PTR-MS. Proton-
transfer-reactions prevail when protonated water (H3O+) is used as a reagent ion. These
reactions are exothermic and happen when the proton affinity of the compounds exceeds
the proton affinity of the reagent ion. This makes PTR-MS selective to the organic com-
pounds presented in the air at trace concentrations and blind to inorganic gases such as
oxygen, nitrogen, and carbon dioxide. Table 11.1 demonstrates the proton affinity of
compounds presented in food matrices (Hunter et al., 2010). However, propanol and
higher alcohols show an increasing tendency to undergo the hydride transfer reaction on
the proton transfer reaction. Some groups of compounds such as C3 and higher alcohols
trend to fragment, losing their hydroxyl group. This type of reaction is called hydride
transfer.
The typical PTR-MS instrument presented by Lindinger et al. (1998) is composed of
an ion source, a drift tube, and an ion detection system (Figure 11.1). This type of ion
source generates ions with a purity of over 99.5%. An ion source usually consists of a
hollow cathode discharge source where water vapor or other gases are injected.
The electron impact ionization of H 2O produces H 2O+ and the bulk of fragment
ions, such as H+, H 2+, OH+, and O+, are transformed into H3O+ due to the fast reactions
220 Food Aroma Evolution

TABLE 11.1 Proton Affinity of Some VOCs

Substance Molecule Proton Affinity (kJ/mol)
Oxygen O2 421
Nitrogen N2 494
Carbon dioxide CO2 541
Water H 2O 691
Ammonia NH3 854
Methanol CH3OH 754
Ethanol C2H5OH 776
Acetaldehyde C2H4O 769
Propanal C3H6O 786
Acetone C3H6O 812
Ethyl acetate C2H4O2 836
Acetic acid C2H4O2 784
Methanethiol CH4S 773
Source: Adapted from Hunter et al., 2010.

FIGURE 11.1 Schematic drawing of a PTR-MS coupled with a quadrupole mass

spectrometer.

(Equations 11.5–11.16). A lower pressure in the ion source than in the drift tube can pro-
voke the formation of some impurities such as O2+ and NO+ in very small amounts. Their
traces can also be produced in the ion source itself.

e – + H 2O → H 2O + 2e – (11.5)

e – + H 2O → H 2+ + O + 2e – (11.6)
 Proton-Transfer-Reaction–Mass Spectrometry 221

e – + H 2O → H+ + OH + 2e – (11.7)

e – + H 2O → O+ + 2H + 2e – (11.8)

e – + H 2O → OH+ + H + 2e – (11.9)

O+ + H2O → H2O+ + O (11.10)

H+ + H 2O → H 2O+ + H (11.11)

H 2+ + H 2O → H3O+ + H (11.12)

H 2+ + H 2O → H 2O+ + H 2 (11.13)

OH + + H 2O → H3O+ + O (11.14)

OH + + H 2O → H 2O+ + OH (11.15)

H 2O+ + H 2O → H3O+ + OH (11.16)

11.2.1 Drift Tube

The drift tube is a crucial part of the PTR-MS instrument where the chemical ionization
happens. Ions from the ion source are extracted into the drift tube because of an electric
field. There they react with VOCs which are directly injected in the drift tube with the
minimum flow around 40 sccm. The drift tube is filled with air as a buffer gas to dilute
both reagents. Since the drift tube is a chemical reactor, it is possible to calculate the con-
centration of reagents according to the principles of chemical kinetics (Equation 11.17;
Lindinger et al., 1998).

1  VOC.H  1
+

C ( ppbv ) = ⋅ ⋅ ⋅ 109 (11.17)

kt H3O+  N

Where C is the concentration of VOC in the analyte gas (ppbv), k is the reaction rate con-
stant (cm3/s), t is the ion travel time in the drift tube, [VOC.H+] and [H3O+] are signals of
these molecules measured in counts per second (cps). In order to obtain the precise VOC
quantification, without any additional calibration with specific gases, the reactions should
occur under well-defined and controlled conditions, such as the ion travel time t and the gas
density in the drift tube N. In additon to the parameters above, the voltages applied to the
drift tube, the temperature, and the inside pressure play an important role in chemical reac-
tions. Another important drift tube parameter is the reduced electric field E/N, where E is an
electric field across the drift tube, which is made by the voltage applied to the drift tube, and
N is the gas density which is regulated by the drift tube pressure, which is obtained through
continuous pumping. High values of E/N correspond to the higher collision energy, which
increases the ion fragmentation. Lower E/N provokes the cluster formation of a primary ion
such as H3O+(H2O)n. in the case of protonated water (Ellis and Mayhew, 2014).
222 Food Aroma Evolution

11.2.2 Switchable Reagent Ion

Proton-transfer-reactions only allow for the detection of those organic compounds whose
proton affinity is higher than H 2O. For improving compound detection, different reagent
ions should be applied. Jordan et al. (2009a) presented the combination of PTR-MS with a
switchable reagent-ion (SRI) technology. A modified design of the drift tube allows for the
easy and fast switching between H3O+, O2+, and NO+ ions. For some particular applica-
tions, Kr+ can be used as well (Agarwal et al., 2014). Recently some studies have shown the
possibility of measuring with NH4+ as a primary ion (Zhu et al., 2018; Hansel et al., 2018).
The formation of O2+ or NO+ ions in the ion source happens according to the follow-
ing reactions (Equations 11.17–11.23):

e – + O2 → O2+ + 2e – (11.17)

e – + O 2 → O + + O + 2e – (11.18)

e – + N 2 → N 2+ + 2e – (11.19)

e – + N 2 → N + + N + 2e – (11.20)

N 2+ + O2 → O2+ + N 2 , k  kc (11.21)

O+ + N 2 → NO+ + N (11.22)

N + + O2 → NO+ + O (11.23)

In the case of O2+ and NO+ modes, different reaction pathways occur instead of pro-
ton-transfer-reactions. Reactions of charge transfer characterize O2+ as a source of ions
(Equations 11.24–11.25).

O2+ + VOC → VOC + + O2 Non-dissociative (11.24)

( ) (F − fragment; N − natural fragment)

*
O2+ + VOC → VOC + + O2
(11.25)
(VOC ) + * +
→F +N

When NO+ is used, the target volatile may undergo charge transfer, hydride (H –) or
hydroxyl (OH –) ion abstraction or both, according to the conditions of the drift tube. The
reactions of NO+ are given in Equations 11.26–11.27. In these equations, X represents the
abstracted ion (H – or OH –) and B the stabilizing buffer gas.

NO+ + VOC → [VOC − X]+ + XNO (11.26)

NO+ + VOC + B → VOC.NO+ + B (11.27)

NO+ as a primary ion separates isomeric aldehydes and ketones because the former appear
at their parent mass and the latter as H-subtracted masses (Ellis and Mayhew, 2014).
 Proton-Transfer-Reaction–Mass Spectrometry 223

Kr+ possesses higher ionization energies than common air constituents and is respon-
sible for non-dissociative and dissociative charge transfer useful for the ionization of some
inorganic compounds presented in the air, such as CO, CO2 , CH4, NOx, and SO2. Thus,
this reagent ion has found its main application in environmental studies (Sulzer et al., 2012).
11.2.2.1 Mass Analyzers
The third main part of a PTR-MS instrument, after the ion source and the drift tube,
is the ion detection system or mass analyzer as it is shown in Figure 11.1. It is used for
ion separation according to their m/z, which is the mass number of an ion m, divided by
its charge number, z. Each type of mass analyzer is characterized by several important
parameters, such as mass resolution, sensitivity, transmission, and dynamic range. The first
can be explained as the ability to distinguish the signal from ions with a small difference
in m/z values. Not all ions which enter a mass spectrometer will reach the detector. The
transmission is caused by a variety of factors and depends on the m/z value, which is why
it is important to take it into account during VOC quantification. The dynamic range is the
ratio of the highest and the lowest detectable ion signal. Usually, the region with a linear
response to the analyte concentration is considered. The noise level influences a lot on the
recognition of small ion signals. Nowadays, the main mass analyzers coupled to PTR are
the quadrupole, the time-of-flight, and their hybrid quadrupole time-of-flight.
11.2.2.2 Quadrupole Mass Spectrometer (QMS)
First, PTR-MS instruments were equipped with quadrupole mass spectrometers (Ellis and
Mayhew, 2014). A typical QMS consists of four parallel metal rods equally positioned
from the center in the vacuum chamber. The voltage applied along rods accelerates ions
in the system. Each opposing rod is connected electrically, and a voltage with a different
frequency is applied between the different pairs of rods which provoke ion oscillations in
the xy direction. Ions with a specific m/z are in resonance with this oscillation and pass
through the quadrupole until they reach the detector. Ions with other m/z are lost.
The advantages of a QMS system include operation in a low vacuum (10–2 to 10–3 Pa)
and compact size. However, a QMS is characterized by a low mass resolution (m/Δm) such
as up to the nominal mass. For this reason, the separation of isobaric compounds is not
possible with such a mass analyzer. Moreover, QMS has a bell-shape transmission curve,
which means that it is less sensitive to the big molecules rather than to small ones.

11.2.2.3 Time-of-Flight–Mass Spectrometer (ToF-MS)

The time-of-flight–mass spectrometer (ToF-MS) uses the principle of the free flight of
ions in a vacuum system. A bunch of ions are accelerated using a repeller and detected by
a detector (Figure 11.2, Ellis and Mayhew, 2014). In order to prolong the ion path, ToF
analyzers are equipped with a reflector array or simply a reflectron which reverses the
motion of the ions thus doubling the length of ToF analyzer. The reflectron also groups
ions of the same m/z traveling slower or faster due to the reflectron voltages.
The kinetic energy (Ek) of an ion accelerated by a constant voltage depends on its
mass and speed (Equation 11.28).

mv 2
Ek = zeV = (11.28)
2
where z is the charge number, e is the elementary charge, V is the acceleration voltage, m
is the mass of the ion, and v is the velocity of the ion. This is why the time-of-flight of an
ion can be described by the formula in Equation 11.29.
224 Food Aroma Evolution

FIGURE 11.2 Schematic of a time-of-flight–mass spectrometer.

L m 1
tflight = = L (11.29)
v v 2eV
Time-of-flight is proportional to the square root of m/z which means that for ions with a
specific m/z it will take specific time to reach the detector and thus it is possible to recal-
culate the m/z value according to its exact ion time-of-flight.
Among the advantages of using a ToF analyzer are the high mass resolution (up to
15,000), very low detection limit (pptv levels), good transmission, and short time for
whole spectrum acquisition (a second or even less). This type of mass analyzer is usually
big in size, which can considerably increase the size of a whole instrument.
In 2004, for the first time, Blake et al. reported the potential of coupling ToF to a
PTR ion source and published data about their first prototype of such a PTR-ToF-MS
device. Jordan et al. (2009a) introduced the first commercial version of a PTR-ToF-MS
instrument developed by Ionicon (Figure 11.3; Innsbruck, Austria).

11.2.2.4 Data Analysis
PTR-MS instruments produce a lot of information which should be extracted and ana-
lyzed in an appropriate way. In the case of a PTR-QMS system, data extraction is very
easy since the software stores the values of signal and mass peak concentration directly
in an ASCII file.
PTR-ToF-MS data are stored in very complex datasets and require sophisticated analysis
with multiple steps. A typical PTR-ToF-MS mass spectrum, with an upper mass-to-charge
 Proton-Transfer-Reaction–Mass Spectrometry 225

FIGURE 11.3 Schematic drawing of the PTR-ToF-MS instrument. (From Jordan et al.,
2009a.)

(m/z) threshold of 400, consists of around 350,000 data bins. Typical integrated acquisi-
tion speed is 1 spectrum/second. The concentration of a specific mass peak depends on the
intensity of the peak and primary ion and on some drift tube parameters, such as tempera-
ture and voltage. For this reason, the analysis of such datasets needs specific software for a
proper elaboration. Several data processing and analysis software products offer their own
approaches for data elaboration such as PTR-MS Viewer (Ionicon, Innsbruck, Austria),
PTR-ToF Data Analyzer (Müller et al., 2013), University of Innsbruck, Innsbruck, Austria),
PTRwid (Holzinger, 2015; Utrecht University, Utrecht, the Netherlands), TOFOffice
(Edmund Mach Foundation, San Michele all’Adige, Italy), and also several others which
are been developed for the individual needs of each laboratory equipped with PTR-ToF-MS.
Cappellin et al. (2011) highlighted the three main “pillars” of a data analysis meth-
odology: spectra analysis, multivariate analysis and data mining, and analytical informa-
tion (Figure 11.4).
External mass-scale calibration is performed before data acquisition for spectra align-
ment and for facilitating the online monitoring of ongoing experiment. For this reason,
external calibration is done on reference mass peaks of known compounds which are con-
stantly present during all measurement steps, such as primary ions or other peaks belong-
ing to impurities of the instrument (H3O+, O2+, NO+). However, in this case, the calibration
precisely corrects only a part of a spectrum with low mass range. This problem was over-
come by adding a constant reference mass peak at high m/z with a constant intensity
and without any interference with a sample signal. PerMaSCal (Ionicon Analytik GmbH,
Innsbruck, Austria) is a commercially available permeation device. Permeation rate and
respectively peak intensity are strongly temperature dependent. To eliminate any distor-
tion of the VOC sample, PerMaSCal is connected directly to the drift tube where the con-
stant stream of the calibration compound diffuses into the sample gas. 1,3-Diiodobenzene
(C6H4I 2) was selected as a permeation unit because of its low toxicity, simple isotope pat-
tern, ionization with different primary ions, and low probability of overlaps with common
226 Food Aroma Evolution

FIGURE 11.4 Main steps during PTR-ToF-MS data analysis. (From Cappellin et al., 2011.)

VOCs. It was observed as m/z 330.848 and m/z 203.946 (fragment, iodine abstraction) in
the H3O+ mode and as m/z 329.840 in both O2+ and NO+ modes.
The main steps of data pre-processing presented in all software are (i) dead time
correction, (ii) mass-scale calibration, (iii) reference peak determination, (iv) peak extrac-
tion, and (v) calculation of concentration in ppbv.

11.3 PTR-MS INNOVATION

11.3.1 Toward Increasing of Sensibility

The performances of mass spectrometers and detectors are critical for the detection sen-
sitivity of PTR-MS (Blake et al., 2009). However, the small exit aperture at the end of
the drift tube limits the sensibility of the instrument. This aperture is necessary for main-
taining a sufficiently low pressure in the mass spectrometer while sustaining a pressure in
the drift tube, which is several orders of magnitude higher. One of the possible solutions
could be the radio frequency (RF) ion funnel, developed by Shaffer et al. (1997). This
ion funnel uses a series of electrodes with progressively reducing aperture sizes. Different
modifications have been made to the ion funnel design since its initial introduction tak-
ing into consideration the ion motion through trajectory simulations. In 2012, Barber
et al. presented the combination of the RF ion funnel with PTR-MS. The drift tube of
a PTR-MS instrument was modified to allow the operation in both modes: as a conven-
tional drift tube and as an ion funnel.
In 2012, Barber et al. and Sulzer et al. published the implementations of their minor
improvements in a PTR-ToF-MS setup, such as a modified ion source, drift tube, transfer
lens system, and so on.
In 2014, Sulzer et al. introduced a new prototype of a PTR-ToF-MS instrument with
a quadrupole ion guide instead of a transfer lens system. This innovation improves the
efficiency of ion transfer from the drift tube to the mass spectrometer. In combination
 Proton-Transfer-Reaction–Mass Spectrometry 227

with an elevated drift tube pressure of 3.8 mbar this instrument boosts the sensitivity by
over two orders of magnitude compared to the first commercial device released in 2009
(Jordan et al., 2009a).
A more radical redesign of the reactor was implemented in the PTR3 instrument,
which operates at a higher pressure (50−80 mbar), uses a tripole to enhance the ion
kinetic energy, and relies on a large sampling flow to transport ions down the reactor,
thereby allowing for much longer reaction times and more efficient product ion formation
(Krechmer et al., 2018).
In 2018, Krechmer et al. presented a new chemical ionization source called Vocus,
which consists of a discharged reagent-ion source and a focusing ion-molecule reactor
(FIMR) and is coupled to PTR-ToF-MS instrument developed by TOFWERK (Thun,
Switzerland). A FIMR consists of a glass tube with a resistive coating, mounted inside a
radio frequency quadrupole (Figure 11.5). The axial electric field is used to enhance ion
collision energies and limit cluster ion formation. The RF field focuses ions on the central
axis of the reactor and improves the detection efficiency of product ions. Krechmer et al.
(2018) concluded that trajectory calculations and calibrations showed that it increased
the detection efficiency by about an order of magnitude. Nowadays, only H3O+ is used
as a reagent ion, but it is possible to operate the reaction chamber with other ions. The
sensitivity of the instrument does not depend on ambient humidity.

11.3.2 Toward High Throughputness

Since PTR-MS belongs to the class of DI-MS techniques, its main capability is to sample
and analyze gas samples continuously. For this reason, PTR-MS, from the very begin-
ning, was mostly applied in the ambient and environmental studies of air pollutants.
Later it was successfully applied for the headspace VOC analysis of different food prod-
ucts. There are two types of headspace measurements: static and dynamic. During static
headspace sampling, VOCs are taken once with a syringe and introduced into an inlet
using low flows without any carrier gas. One of the drawbacks of this method is a pres-
sure drop inside the vial. During dynamic headspace analysis higher flows pass through
a vial requiring a carrier gas, such as zero air or nitrogen, in order to avoid vacuum
creation (Romano et al., 2015; Figure 11.6). From the very beginning, all the measure-
ments were performed manually, including sample incubation at higher temperatures for
aroma release stimulation. In this case, the analysis was possible only in the presence of

FIGURE 11.5 Design of Vocus consisting of a discharge reagent-ion source and a focus-
ing ion-molecule reactor (FIMR). (From Krechmer et al., 2018.)
228 Food Aroma Evolution

FIGURE 11.6 Schematic representation of the setup for direct VOC measurements.

FIGURE 11.7 Schematic representation of a multipurpose sampler MPS for GC coupled

with PTR-ToF-MS.
personnel who manually measured the samples. This procedure increases the time of the
experiment and the possibility of human factor errors.
Direct injection of VOCs into an instrument allows the analysis of complex aroma
blends in a very short time; thus, manual headspace measurements become a bottle-
neck for experiments. In order to increase the measurement throughputness, automatic
samplers were coupled to PTR-MS instruments. The pioneering work of Makhoul et al.
(2014) was done by connecting a multipurpose sampler (MPS) for GC (Gerstel, Germany)
to PTR-ToF-MS (Ionicon, Innsbruck, Austria) through a MPS purge tool and a head-
space adapter using a conventional syringe-based system (Figure 11.7; Makhoul et al.,
2014). Tray temperature and experiment time sequences are fully controlled by the MPS.
The maximum number of vials can be increased drastically according to the number of
trays connected to the MPS and the programming sequence of the MPS. The purge tool,
together with the headspace adapter, enables the exchange of the gas phase into the head-
space above the sample with a control flushing gas. It can be used both for sample prepa-
ration and measurement. Moreover, it can be used for prevention of oxidation processes
in such products as wine, olive oil, or butter. In this case, a sample should be purged with
nitrogen for the elimination of oxygen from a vial headspace. Such autosamplers give the
possibility of re-measuring the same vials in order to follow the product’s evolution over
time, which is very important in fermentation or aging studies.
Another example of the automatization of PTR-MS measurements is an autosam-
pler for PTR-MS made by Ionicon (Innsbruck, Austria), which is suitable for static and
dynamic headspace analysis up to 270 vials. It is equipped with a heated injection cell
 Proton-Transfer-Reaction–Mass Spectrometry 229

with a direct connection to a PTR-MS inlet (http​://ww​w.ion​icon.​com/p​roduc​t/acc​essor​

ies/a​utosa​mpler​).
The sample size limits the usage of an autosampler. For instance, the 20 mL vial per-
mits the measurement of only small berries, leaves, coffee beans, their powder or puree,
liquid foods like oils, milk, juices, and parts of solid products like chocolate, meat, fish,
and so on. If the sample is too big to enter the vial, which can be used for an autosampler,
or its size cannot be diminished, it should be measured manually (e.g., aroma profiles of
intact apples or strawberries).
The monitoring of online processes which cannot be reduced to a 20 mL vial scale
should be monitored manually. However, if these processes are not very fast and the
experimental setup allows several of them to run simultaneously, it is possible to increase
the throughput of such measurements analyzing one sample after another in a continu-
ous way. It is possible to organize using the automation of the Multivalve Port (Ionicon,
Innsbruck, Austria) or the set of switchable valves.

11.3.3 Toward Compound Identification

PTR-MS, coupled to a ToF analyzer, provides the separation and identification of iso-
baric compounds with precision up to the fourth digit after a comma. However, isomer
compounds still cannot be distinguished in this way. The first trials to distinguish struc-
tural isomeric compounds were made using PTR+SRI-MS technology and switching dif-
ferent reagent ions to distinguish between different classes of compounds, that is, ketones
and aldehydes (Jordan et al., 2009b). This approach works fine with the simple matrices
of several compounds. Food samples consist of a blend of various VOCs, and their sepa-
ration according to different reagent ions becomes very complicated. In this case, these
data represent a food sample fingerprint and can be used in the data analysis for sample
discrimination or characterization (Yener et al., 2015).
The best solution for isomeric compound identification is the application of a fast-chro-
matographic separation. In order not to lose the high analytical throughput of the methodol-
ogy, which is the chief feature of the PTR-MS technique, the gas chromatographic separation
should be as fast as possible. Recently a fastGC add-on (Ionicon, Innsbruck, Austria) was
implemented as an additional inlet system which consisted of four valves and the flow con-
troller (FC N2) (Figure 11.8, Romano et al., 2014). FastGC uses nitrogen as a carrier gas.
Figure 11.8 visualizes all valves in their NO (normally open) state which corresponds to a
fastGC disable mode. FastGC requires higher inlet flows in order to fill the sampling loop.
There are several parameters which have a great influence on fastGC analysis, such as a ther-
mal ramp, sampling time, and carrier gas flow. The polarity of the chromatographic column
should be chosen according to analytical needs and the matrix features.
Several applications of a fastGC add-on coupled to PTR-ToF-MS have been reported
by the scientific community. Materić et al. (2015) showed the potential of this setup for
separating monoterpenes from the standard mixture and biological material using an
80-second run. Romano et al. (2014) applied a fastGC add-on to wine analysis not only
for isomer separation but also for the removal of the ethanol effect without the need
to drastically change the ionization conditions of the experiment. Pallozzi et al. (2016)
studied the implementation of the setup discussed above to detect VOC emissions from
plants; in particular, the assessment of the potential interferences generated on the iso-
prene signal by other biological VOCs, naturally emitted by some species or induced by
abiotic and biotic stresses.
230 Food Aroma Evolution

FIGURE 11.8 Schematic drawing of a fastGC add-on. (From Romano et al., 2014.)

11.4 APPLICATION OF PTR-MS ANALYSIS IN FOOD SCIENCE

The instrumental characteristics of PTR-MS technology make it an ideal tool for rapid,
non-invasive, solvent-free classification of food products according to quality standards
and origin. In the last two decades, PTR-MS has been progressively applied more in food
science, as revealed by the increasing number of peer-reviewed publications (Figure 11.9),
including studies about totally different matrices, from lactic acid fermented products to
fruit and vegetables.

11.4.1 Screening VOC Fingerprinting Analysis

PTR-MS is particularly suited to developing reliable food VOC fingerprints because it

provides handier analytical information (concentration estimation and reduced frag-
mentation) in comparison with the application of MS-e-noses based on electron impact
ionization (Biasioli et al., 2011b). PTR-MS-based e-noses, equipped with multipurpose
autosamplers, provide a rich, informative, and high-throughput fingerprint.
The most efficient way to exploit this spectrometric fingerprint is through unsuper-
vised multivariate methods for data compression and visualization, and through the set-
ting of classification or calibration models by supervised multivariate methods and data
mining. In addition, the association of these VOC fingerprints with sensory analysis by
human testers provides confirmatory information on the use of PTR-MS for food dis-
criminatory analysis such as the classification of food samples based on geographical
location or on production processes.
 Proton-Transfer-Reaction–Mass Spectrometry 231

FIGURE 11.9 Peer-reviewed publications (till the end of 2018) about the use of PTR-MS
technology on food product analysis.

One of the first studies exploiting rapid PTR-MS VOC fingerprinting for food classi-
fication, regarded the discrimination of red orange juices stabilized by different treatments
(Biasioli et al., 2003). Likewise, Gasperi et al. (2009) explored the effects of supercritical CO2
treatments and N2O pasteurization in apple juice VOCs by PTR-MS. Tsevdou et al. (2013)
used PTR-ToF-MS to investigate the effects of thermal or high hydrostatic pressure treatment
on the flavor development of yogurt in the absence or presence of a transglutaminase protein.
232 Food Aroma Evolution

Assessments of food quality using PTR-MS also include the rapid detection of food
spoilage and aging. Aprea et al. (2006) characterized the VOC headspace of several vir-
gin olive oil samples based on their oxidative alteration. Spoiled olive oil samples were
characterized by a significantly increased content of aldehydes such as heptanal, octanal,
and nonanal. Moreover, Aprea et al. (2008) monitored the oxidative alterations of olive
oils online during thermal treatments, following the time-evolution of several aldehydes.
Heenan et al. (2009) used PTR-MS to compare VOC and sensory characteristics of
bread, with the aim of understanding how bread freshness is perceived by the consumer.
Partial last squares regression modeling of VOCs and sensory characteristics confirmed
the possibility of using PTR-MS as a tool to predict bread sensory quality and freshness.
Makhoul et al. (2014) showed, using a PTR-ToF-MS, that the impact of yeast strains on
VOC production during the bread-making process exceeds the impact of the flour.
Positive results of the multivariate modeling of the link between sensory evaluations
by a trained panel and PTR-MS measurement were obtained in discrimination analyses
of different cheese types (Biasioli et al., 2006), butter and butter oil (van Ruth et al.,
2007), tomato (Muilwijk et al., 2015), and chocolate (Deuscher et al., 2019).
Another important application of PTR-MS analysis is the investigation of the geo-
graphical origin of food. In combination with appropriate statistical techniques, PTR-MS
has indeed been shown to be able to provide geographical identification for several food
products, including coffee (Yener et al., 2014, 2015), tea (Yener et al., 2016), chocolate
(Acierno et al., 2016), butter (Maçatelli et al., 2009), olive oil (Araghipour et al., 2008),
truffles ( Vita et al., 2015), unifloral honey (Kuś and van Ruth, 2015), sea roe (Phillips et
al., 2010), saffron (Masi et al., 2016), Dutch cumin cheese (Galle et al., 2011), dry-cured
ham (Sánchez del Pulgar et al., 2011), and wine (Campbell-Sills et al., 2016).
PTR-MS application was recently demonstrated as a powerful phenotyping tool
for aroma assessment in both genetic and quality-related studies of fruit and vegetables
(F&V). PTR-MS was indeed successfully applied to discriminate between aroma vari-
ability in tomato (Farneti et al., 2012, 2013), apple (Farneti et al., 2014, 2015, 2017a,
2017b), blueberry (Farneti et al., 2018), raspberry (Aprea et al., 2009), and pepper (Taiti
et al., 2015). Another important aspect of the F&V production chain is the optimization
of quality upon delivery to the consumer; in this, VOCs should be considered as a central
trait for determining the “from farm to fork” strategies. The end of the “flavor life” often
precedes the end of shelf life as determined by visual and textural features, mainly due to
changes in aroma compound concentration and off-flavors development.
Aroma and flavor are essential for determining the quality of alcohol beverages. It
can help to distinguish raw material variety, geographical origin, fermentation process,
technological regimen, aging, spoilage phenomena, and adulteration. However, when a
molecule is present at too high concentration, such as ethanol in the case alcoholic bever-
ages, the depletion of the parent ion (H3O+) and the formation of undesired ions make the
ionization process more complicated and hinder the application of PTR-MS in enological
studies. Different approaches have been proposed for overcoming these issues. Lindinger,
the father of PTR-MS, proposed the use of ethanol as a carrier gas to completely deplete
hydronium and use protonated ethanol as parent ion. This method was implemented by
Boscaini et al. (2004). However, not only protonated ethanol is formed but also a large
number of charged clusters which make spectra difficult to decipher. Spitaler et al. (2007)
applied a simple dilution of the headspace with the obvious reduction of sensitivity.
Recently Campbell-Sills et al. (2016) demonstrated that dilution with argon directly in
the PTR-MS drift tube destroys ethanol and ethanol/water clusters and cleans the spec-
tra with a reduced loss of sensitivity and allows the discrimination of wines of different
 Proton-Transfer-Reaction–Mass Spectrometry 233

origins. This approach was successfully applied for studying the fermentation of beer
wort (Richter et al., 2018). Romano et al. (2014) tried a fastGC add-on which, without
increasing the minimum analysis time of PTR-MS (about 3 min), allowed for the separa-
tion of ethanol and the use of the standard PTR-MS parameters.

11.4.2 Real-Time VOC Evolution Analysis

The flavor quality of a product is not a stable trait, but it can rapidly and drastically
change over time. The use of PTR-MS as a process analytical technology (PAT) allows
real-time monitoring of VOCs during processing. PTR-MS can be used as an analytical
fingerprinting tool for monitoring food quality during the processing phase.
PTR-ToF-MS has been proposed as a non-invasive technology for the quality control
of milk lactic acid fermentation (Soukoulis et al., 2010). This work, which represents the
first application of PTR-ToF-MS for studying a dynamic biochemical process, has dem-
onstrated the outstanding ability of the instrument to monitor and identify VOCs which
are either formed or depleted during the fermentation process. The successful application
of PTR-MS in the field of lactic acid fermentation helped to continue the investigation
of aroma formation during yogurt and kefir preparation (Benozzi et al., 2015; Yépez et
al., 2019). Similarly, Capozzi et al. (2016) characterized, for the first time, the dynamic
of VOCs associated with aromatic bakery yeasts (Saccharomyces cerevisiae) proposing
several biomarkers helpful for optimizing the aromatic performances of commercial yeast
preparation. In 2017, Khomenko et al. showed the possibility of monitoring the aroma
evolution of wine yeast strains grown aerobically on a solid substrate for 11 days.
PTR-MS analysis was also successfully applied for monitoring coffee VOC dynam-
ics during production steps, mostly during roasting and extraction. The complexity of
the aromas released by coffee beans during roasting was thoroughly investigated using
PTR-MS by comparing different coffee origins and varieties and also different roast-
ing conditions (Mateus et al., 2007; Gloess et al., 2014; Wieland et al., 2012). For the
final coffee quality, extraction of ground coffee with hot water is equally important to
the flavor profile induced by the roasting process. The extraction technique and condi-
tions used for coffee preparation strongly influence the flavor profile in the cup, and it is
often the only parameter that can be influenced by the consumer at home. Sánchez-López
et al. (2014) measured the VOC kinetics produced during coffee extraction by using a
PTR-ToF-MS coupled with a dilution lancet connected to a commercial espresso coffee
machine.
The process of bread baking and toasting was recently studied by Pico et al. (2018).
The authors monitored the aroma development of wheat and gluten-free bread in an
online mode, which gave them the possibility of evaluating the generation of different ten-
tatively identified compounds of lipid oxidation, Maillard reaction, and other processes.
Recently Pedrotti et al. (2018) applied the “aromatic” point of view to the aging pro-
cesses of anhydrous milk fat stored in two different types of packaging.

11.5 CHALLENGES AND FUTURE PERSPECTIVES

PTR-MS technology provides several advantages to VOC analysis in food science and
technology. Its key feature is the possibility of detecting and quantifying VOCs in a
direct, continuous, and real-time way at very low levels (pptv), with both high mass and
234 Food Aroma Evolution

time resolution. The high time resolution allows real-time monitoring of fast food pro-
cesses such as the formation of volatile compounds during thermal processes (i.e., cook-
ing, baking, or roasting), or during food fermentation.
On the other hand, compound identification is still a weak aspect of this technology.
Fragmentation, complex peak structure, and/or the presence of isomeric compounds
may still make this challenge impractical, especially in complex matrices. The ioniza-
tion based on proton transfer provides a soft ionization where most mass peaks appear
on their parent ions; however, some residual fragmentation is not always avoidable
and negligible. Right now, the cutting-edge research on PTR-MS technology focuses
on the improvement of compound identification, instrument sensibility, and analytic
throughputness.
Overall, it has to be noted that the aforementioned characteristics of PTR-MS tech-
nology do not suggest PTR-MS as an alternative to the gas chromatographic method but
as a complementary tool for the study of volatile compounds and a valuable technique
when speed, sensitivity, and online measurements are required.
After more than two decades of its history, PTR-MS found its application not only in
scientific research but also in industrial projects where the high stability, reproducibility,
automation, and throughputness are required. Moreover, with the rise in awareness of
anthropogenic pollution, the absence of any dangerous solvents or reagents needed for
sample preparation and analysis gives PTR-MS an additional value as a green chemistry
technique.

REFERENCES

Acierno, V., Yener, S., Alewijn, M., Biasioli, F., and van Ruth, S., 2016. Factors contrib-
uting to the variation in the volatile composition of chocolate: Botanical and geo-
graphical origins of the cocoa beans, and brand-related formulation and processing.
Food Research International, 84 (1), 86–95.
Agarwal, B., González-Méndez, R., Lanza, M., Sulzer, P., Märk, T. D., Thomas, N.,
and Mayhew, C. A., 2014. Sensitivity and selectivity of switchable reagent ion soft
chemical ionization mass spectrometry for the detection of picric acid. Journal of
Physical Chemistry A, 118 (37), 8229–36.
Amann, A., Costello, B. D. L., Miekisch, W., Schubert, J., Buszewski, B., and Pleil,
J., 2014. The human volatilome: Volatile organic compounds (VOCs) in exhaled
breath, skin emanations, urine, feces and saliva. Journal of Breath Research, 8 (3),
034001. doi:10.1088/1752-7155/8/3/034001.
Aprea, E., Biasioli, F., Carlin, S., Endrizzi, I., and Gasperi, F., 2009. Investigation of
volatile compounds in two raspberry cultivars by two headspace techniques: Solid-
phase microextraction/gas chromatography-mass spectrometry (SPME/GC-MS)
and proton-transfer reaction-mass spectrometry (PTR-MS). Journal of Agricultural
and Food Chemistry, 57, 4011–8.
Aprea, E., Biasioli, F., Sani, G., Cantini, C., Märk, T. D., and Gasperi, F., 2006. Proton
transfer reaction–mass spectrometry (PTR-MS) headspace analysis for rapid detec-
tion of oxidative alteration of olive oil. Journal of Agricultural and Food Chemistry,
54, 7635–40.
Aprea, E., Biasioli, F., Sani, G., Cantini, C., Märk, T. D., and Gasperi, F., 2008. Online
monitoring of olive oil headspace by proton transfer reaction–mass spectrometry.
Rivista Italiana Della Sostanze Grasse, 85, 92–7.
 Proton-Transfer-Reaction–Mass Spectrometry 235

Araghipour, N., Colineau, J., Koot, A., Akkermans, W., Rojas, J. M. M., Beauchamp,
J., Wisthaler, A., Märk, T. D., Downey, G., Guillou, C., Mannina, L., and van
Ruth, S. M., 2008. Geographical origin classification of olive oils by PTR-MS. Food
Chemistry, 108, 374–83.
Barber, S., Blake, R. S., White, I. R., Monks, P. S., Reich, F., Mullock, S., and Ellis, A. M.,
2012. Increased sensitivity in proton transfer reaction mass spectrometry by incor-
poration of a radio frequency ion funnel. Analytical Chemistry, 84 (2012), 5387–91.
Benozzi, E., Romano, A., Capozzi, V., Makhoul, S., Cappellin, L., Khomenko, I., Aprea,
E., Scampicchio, M., Spano, G., Märk, T. D., Gasperi, F., and Biasioli, F., 2015.
Monitoring of lactic fermentation driven by different starter cultures via direct
injection mass spectrometric analysis of flavour-related volatile compounds. Food
Research International, 76 (3), 682–8.
Biasioli, F., Gasperi, F., Aprea, E., Colato, L., Boscaini, E., and Märk, T. D., 2003.
Fingerprinting mass spectrometry by PTR-MS: Heat treatment vs. pressure treat-
ment of red orange juice—A case study. International Journal of Mass Spectrometry,
223–4, 343–53.
Biasioli, F., Gasperi, F., Aprea, E., Endrizzi, I., Framondino, V., Marini, F., Mott, D., and
Märk, T. D., 2006. Correlation of PTR-MS spectral fingerprints with sensory char-
acterisation of flavour and odour profile of “Trentingrana” cheese. Food Quality
and Preference, 17, 63–75.
Biasioli, F., Gasperi, F., Yeretzian, C., and Märk, T. D., 2011a. PTR-MS monitoring of
VOCs and BVOCs in food science and technology. Trends in Analytical Chemistry,
30. doi:10.1016/j.trac.2011.03.009.
Biasioli, F., Yeretzian, C., Märk, T. D., Dewulf, J., and Van Langenhove, H., 2011b.
Direct-injection mass spectrometry adds the time dimension to (B) VOC analysis.
Trends in Analytical Chemistry, 30, 1003–17. doi:10.1016/j.trac.2011.04.005.
Bicchi, C. and Maffei, M., 2012. High-throughput phenotyping in plants. Methods and
Protocols, 918, 289–310. doi:10.1007/978-1-61779-995-2 ISBN 978-1-61779-995-2.
Blake, R. S., Monks, P. S., and Ellis, A. M., 2009. Proton-transfer reaction mass spec-
trometry. Chemical Reviews, 109 (3), 861–96.
Blake, R. S., Whyte, C., Hughes, C. O., Ellis, A. M., and Monks, P. S., 2004. Demonstration
of proton-transfer reaction time-of-flight mass spectrometry for real-time analysis
of trace volatile organic compounds. Analytical Chemistry, 76, 3841–5.
Boscaini, E., Mikoviny, T., Wisthaler, A., von Hartungen, E., and Märk, T. D.,
2004. Characterization of wine with PTR-MS. International Journal of Mass
Spectrometry, 239, 215–9.
Campbell-Sills, H., Capozzi, V., Romano, A., Cappellin, L., Spano, G., Breniaux, M.,
Lucas, P., and Biasioli, F., 2016. Advances in wine analysis by PTR-ToF-MS:
Optimization of the method and discrimination of wines from different geographi-
cal origins and fermented with different malolactic starters. International Journal
of Mass Spectrometry, 397–8, 42–51.
Capozzi, V., Makhoul, S., Aprea, E., Romano, A., Cappellin, L., Sanchez Jimena, A.,
Spano, G., Gasperi, F., Scampicchio, M., and Biasioli, F., 2016. PTR-MS charac-
terization of VOCs associated with commercial aromatic bakery yeasts of wine and
beer origin. Molecules, 21, 483.
Cappellin, L., Biasioli, F., Granitto, P. M., Schuhfried, E., Soukoulis, C., Costa, F., Märk,
T. D., and Gasperi, F., 2011. On data analysis in PTR-ToF-MS: From raw spec-
tra to data mining. Sensors Actuators B: Chemical, 155, 183–90. doi:10.1016/j.
snb.2010.11.044.
236 Food Aroma Evolution

Casaburi, A., Piombino, P., Nychas, G., Villani, F., and Ercolini, D., 2015. Bacterial
populations and the volatilome associated to meat spoilage. Food Microbiology, 45,
83–102. doi:10.1016/j.fm.2014.02.002.
Cevallos-cevallos, J. M., Etxeberria, E., Danyluk, M. D., and Rodrick, G. E., 2009.
Metabolomic analysis in food science: A review. Trends in Food Science &
Technology, 20, 557–66. doi:10.1016/j.tifs.2009.07.002.
Cumeras, R., 2017. Volatilome metabolomics and databases, recent advances and needs.
Current Metabolomics, 5 (2). doi:10.2174/2213235X05666170502103408.
Deuscher, Z., Andriot, I., Sémon, E., Repoux, M., Preys, S., Roger, J.-M., Boulanger,
R., Labouré, H., and Le Quéré, J.-L., 2019. Volatile compounds profiling by using
proton transfer reaction-time of flight-mass spectrometry (PTR-ToF-MS). The case
study of dark chocolates organoleptic differences. Journal of Mass Spectrometry,
54, ii. https://doi.org/10.1002/jms.4214.
Ellis, A. M. and Mayhew, C. A. (eds.), 2014. Proton Transfer Reaction Mass Spectrometry:
Principles and Applications. W. Sussex, UK: Wiley.
Farneti, B., Algarra Alarcón, A., Cristescu, S. M., Costa, G., Harren, F. J. M., Holthuysen,
N. T. E., and Woltering, E. J., 2013. Aroma volatile release kinetics of tomato
genotypes measured by PTR-MS following artificial chewing. Food Research
International, 54, 1579–88.
Farneti, B., Cristescu, S. M., Costa, G., Harren, F. J. M., and Woltering, E. J., 2012. Rapid
tomato volatile profiling by using Proton-Transfer Reaction Mass Spectrometry
(PTR-MS). Journal of Food Science, 77, 551–9.
Farneti, B., Di Guardo, M., Khomenko, I., Cappellin, L., Biasioli, F., Velasco, R., and
Costa, F., 2017. Genome-wide association study unravels the genetic control of
the apple volatilome and its interplay with fruit texture. Journal of Experimental
Botany. ISSN: 0022-0957. doi:10.1093/jxb/erx018.
Farneti, B., Khomenko, I., Cappellin, L., Ting, V. J. L., Costa, G., Biasioli, F., and Costa,
F., 2015. Dynamic volatile organic compound fingerprinting of apple fruit during
processing. LWT—Food Science and Technology, 63, 21–8.
Farneti, B., Khomenko, I., Cappellin, L., Ting, V. J. L., Romano, A., Biasioli, F., Costa,
G., and Costa, F., 2014. Comprehensive VOC profiling of an apple germplasm col-
lection by PTR-ToF-MS. Metabolomics, 11, 838–50.
Farneti, B., Khomenko, I., Grisenti, M., Ajelli, M., Betta, E., Algarra, A., Cappellin,
L., Aprea, E., Gasperi, F., Biasioli, F., and Giongo, L., 2017. Exploring blue-
berry aroma complexity by chromatographic and direct-injection spectro-
metric techniques. Frontiers in Plant Science. ISSN: 1664-462X. doi:10.3389/
fpls.2017.00617.
Fiehn, O., 2002. Metabolomics—The link between genotypes and phenotypes. Plant
Molecular Biology, 48, 155. https://doi.org/10.1023/A:1013713905833.
Furbank, R. T. and Tester, M., 2011. Phenomics—Technologies to relieve the pheno-
typing bottleneck. Trends in Plant Science, 16, 635–44. doi:10.1016/j.tplants.
2011.09.005.
Galle, S. A., Koot, A., Soukoulis, C., Cappellin, L., Biasioli, F., Alewijn, M., and van
Ruth, S. M., 2011. Typicality and geographical origin markers of protected ori-
gin cheese from the Netherlands revealed by PTR-MS. Journal of Agricultural and
Food Chemistry, 59, 2554–63.
Gasperi, F., Aprea, E., Biasioli, F., Carlin, S., Endrizzi, I., Pirretti, G., and Spilimbergo,
S., 2009. Effects of supercritical CO2 and N2O pasteurisation on the quality of fresh
apple juice. Food Chemistry, 115 (1), 129–36.
 Proton-Transfer-Reaction–Mass Spectrometry 237

Gloess, A. N., Vietri, A., Wieland, F., Smrke, S., Schönbächler, B., Sánchez López, J. A.,
Petrozzi, S., Bongers, S., Koziorowski, T., and Yeretzian, C., 2014. Evidence of dif-
ferent flavour formation dynamics by roasting coffee from different origins: On-line
analysis with PTR-ToF-MS. International Journal of Mass Spectrometry, 365–6,
324–37.
Hansel, A., Scholz, W., Mentler, B., Fischer, L., and Berndt, T., 2018. Detection of RO2
radicals and other products from cyclohexene ozonolysis with NH4+ and acetate
chemical ionization mass spectrometry. Atmospheric Environment, 186, 248–55.
Heenan, S. P., Dufour, J. P., Hamid, N., Harvey, W., and Delahunty, C. M., 2009.
Characterisation of fresh bread flavour: Relationships between sensory characteris-
tics and volatile composition. Food Chemistry, 116, 249–57.
Herrero, M., Simó, C., and Cifuentes, A., 2012. Foodomics: MS-based strategies in mod-
ern food science and nutrition. Mass Spectrometry Reviews, 49–69. doi:10.1002/
mas.
Holzinger, R., 2015. PTRwid: A new widget-tool for processing PTR-ToF-MS data.
Atmospheric Measurement Techniques Discussions, 8, 1629–69. doi:10.5194/
amtd-8-1629-2015.
Huang, M., Cheng, S., Cho, Y., and Shiea, J., 2011. Ambient ionization mass spectrom-
etry: A tutorial. Analytica Chimica Acta, 702, 1–15. doi:10.1016/j.aca.2011.06.017.
Hunter, E. P. L., Lias, S. G., Hunter, E. P. L., and Lias, S. G., 2010. Evaluated gas phase
basicities and proton affinities of molecules. Journal of Physical and Chemical
Reference Data, 413. doi:10.1063/1.556018.
Ibáñez, C., García-cañas, V., Valdés, A., and Simó, C., 2013. Novel MS-based approaches
and applications in food metabolomics. Trends in Analytical Chemistry, 52, 100–
11. doi:10.1016/j.trac.2013.06.015.
Jordan, A., Haidacher, S., Hanel, G., Hartungen, E., Herbig, J., Märk, L., Schottkowsky,
R., Seehauser, H., Sulzer, P., and Märk, T. D., 2009b. An online ultra-high sensitiv-
ity Proton-transfer-reaction mass-spectrometer combined with switchable reagent
ion capability (PTR + SRI – MS). International Journal of Mass Spectrometry, 286,
32–8. doi:10.1016/j.ijms.2009.06.006.
Jordan, A., Haidacher, S., Hanel, G., Hartungen, E., Märk, L., Seehauser, H.,
Schottkowsky, R., Sulzer, P., and Märk, T. D., 2009a. A high resolution and
high sensitivity proton-transfer-reaction time-of-flight mass spectrometer (PTR-
ToF-MS). International Journal of Mass Spectrometry, 286, 122–8. doi:10.1016/j.
ijms.2009.07.005.
Khomenko, I., Stefanini, I., Cappellin, L., Cappelletti, V., Franceschi, P., Cavalieri, D.,
Märk, T. D., and Biasioli, F., 2017. Metabolomics, 13, 118. https​://do​i.org​/10.1​0 07/
s​11306​- 017-​1259-​y.
Krechmer, J., Lopez-Hilfiker, F., Koss, A., Hutterli, M., Stoermer, C., Deming, B.,
Kimmel, J., Warneke, C., Holzinger, R., Jayne, J., Worsnop, D., Fuhrer, K., Gonin,
M., and de Gouw, J., 2018. Evaluation of a new reagent-ion source and focusing ion-
molecule reactor for use in proton-transfer-reaction mass spectrometry. Analytical
Chemistry, 90 (20), 12011–8. doi:10.1021/acs.analchem.8b02641.
Kuś, P. M. and van Ruth, S. M., 2015. Discrimination of Polish unifloral honeys using
overall PTR-MS and HPLC fingerprints combined with chemometrics. LWT—Food
Science and Technology, 62, 69–75.
Li, N., Alfiky, A., and Vaughan, M. M., 2016. Stop and smell the fungi: Fungal vola-
tile metabolites are overlooked signals involved in fungal interaction with plants.
Fungal Biology Reviews, 30, 134–44. doi:10.1016/j.fbr.2016.06.004.
238 Food Aroma Evolution

Lindinger, W., Hansel, A., and Jordan, A., 1998. On-line monitoring of volatile organic
compounds at pptv levels by means of proton-transfer-reaction mass spectrom-
etry (PTR-MS) medical applications, food control and environmental research.
International Journal of Mass Spectrometry and Ion Processes, 173, 191–241.
doi:10.1016/S0168-1176(97)00281-4.
Maçatelli, M., Akkermans, W., Koot, A., Buchgraber, M., Paterson, A., and van Ruth,
S. M., 2009. Verification of the geographical origin of European butters using
PTR-MS. Journal of Food Composition and Analysis, 22, 169–75.
Makhoul, S., Romano, A., Cappellin, L., Spano, G., Capozzi, V., Benozzi, E., Märk, T.
D., Aprea, E., Gasperi, F., El-Nakat, H., Guzzo, J., and Biasioli, F., 2014. Proton-
transfer-reaction mass spectrometry for the study of the production of volatile
compounds by bakery yeast starters. Journal of Mass Spectrometry, 49, 850–9.
doi:10.1002/jms.3421.
Masi, E., Taiti, C., Heimler, D., Vignolini, P., Romani, A., and Mancuso, S., 2016. PTR-
ToF-MS and HPLC analysis in the characterization of saffron (Crocus sativus L.)
from Italy and Iran. Food Chemistry, 192, 75–81.
Materić, D., Lanza, M., Sulzer, P., Herbig, J., Bruhn, D., Turner, C., Mason, N., and
Gauci, V., 2015. Monoterpene separation by coupling proton transfer reaction time-
of-flight mass spectrometry with fastGC. Analytical and Bioanalytical Chemistry,
7757–63. doi:10.1007/s00216-015-8942-5.
Mateus, M., Lindinger, C., Gumy, J. C., and Liardon, R., 2007. Release kinetics of vola-
tile organic compounds from roasted and ground coffee: Online measurements by
PTR-MS and mathematical modeling. Journal of Agricultural and Food Chemistry,
55, 10117–28.
Muilwijk, M., Heenan, S., Koot, A., and van Ruth, S. M., 2015. Impact of production
location, production system, and variety on the volatile organic compounds finger-
prints and sensory characteristics of tomatoes. Journal of Chemistry. https://doi.
org/10.1155/2015/981549.
Müller, M., Mikoviny, T., Jud, W., D’Anna, B., and Wisthaler, A., 2013. A new software
tool for the analysis of high resolution PTR-ToF mass spectra. Chemometrics and
Intelligent Laboratory Systems, 127, 158–65. doi:10.1016/j.chemolab.2013.06.011.
Nozie, B., Kalberer, M., Claeys, M., Allan, J., Anna, B. D., Decesari, S., Finessi, E.,
Glasius, M., Grgic, I., Hamilton, J. F., Ho, T., Iinuma, Y., Jaoui, M., Kahnt, A.,
Kampf, C. J., Kourtchev, I., Maenhaut, W., Marsden, N., Saarikoski, S., Schnelle-
kreis, J., Surratt, J. D., Szidat, S., Szmigielski, R., and Wisthaler, A., 2015. The
molecular identification of organic compounds in the atmosphere: State of the art
and challenges. Chemical Reviews, 115 (10), 3919–83. doi:10.1021/cr5003485.
Pallozzi, E., Guidolotti, G., Ciccioli, P., Brilli, F., Feil, S., and Calfapietra, C., 2016. Does
the novel fast-GC coupled with PTR-ToF-MS allow a significant advancement in
detecting VOC emissions from plants? Agricultural and Forest Meteorology, 216,
232–40. doi:10.1016/j.agrformet.2015.10.016.
Pedrotti, M., Khomenko, I., Cappellin, L., Fontana, M., Somenzi, M., Falchero, L.,
Arveda, M., Fogliano, V., and Biasioli, F., 2018. Rapid and noninvasive quality
control of anhydrous milk fat by PTR-MS: The effect of storage time and packag-
ing. Journal of Mass Spectrometry, 53, 753–62. https://doi.org/10.1002/jms.4204.
Phillips, K., Niimi, J., Hamid, N., Silcock, P., Delahunty, C., Barker, M., Sewell, M.,
and Bremer, P., 2010. Sensory and volatile analysis of sea urchin roe from different
geographical regions in New Zealand. LWT—Food Science and Technology, 43,
202–13.
 Proton-Transfer-Reaction–Mass Spectrometry 239

Pichersky, E. and Gang, D. R., 2000. Genetics and biochemistry of secondary metabo-
lites: An evolutionary perspective. Trends in Plant Science, 5, 439–45.
Pico, J., Khomenko, I., Capozzi, V., Navarini, L., Bernal, J., Gómez, M., and Biasioli, F.,
2018. Analysis of volatile organic compounds in crumb and crust of different baked
and toasted gluten-free breads by direct PTR-ToF-MS and fast-GC-PTR-ToF-MS.
Journal of Mass Spectrometry, 53, 893–902.
Richter, T. M., Silcock, P., Algarra, A., Eyres, G. T., Capozzi, V., Bremer, P. J., and
Biasioli, F., 2018. Evaluation of PTR-ToF-MS as a tool to track the behavior of hop-
derived compounds during the fermentation of beer. Food Research International,
111, 582–9.
Romano, A., Fischer, L., Herbig, J., Campbell-Sills, H., Coulon, J., Lucas, P., Cappellin,
L., and Biasioli, F., 2014. Wine analysis by FastGC proton-transfer reaction-time-
of-flight-mass spectrometry. International Journal of Mass Spectrometry, 369, 81–
6. doi:10.1016/j.ijms.2014.06.006.
Romano, A., Gaysinsky, S., Czepa, A., del Pulgar, J. S., Cappellin, L., and Biasioli, F.,
2015. Static and dynamic headspace analysis of instant coffee blends by proton-
transfer-reaction mass spectrometry. Journal of Mass Spectrometry, 50, 1057–62.
doi:10.1002/jms.3619.
Sánchez del Pulgar, J., Soukoulis, C., Biasioli, F., Cappellin, L., García, C., Gasperi, F.,
Granitto, P., Märk, T. D., Piasentier, E., and Schuhfried, E., 2011. Rapid charac-
terization of dry cured ham produced following different PDOs by proton transfer
reaction time of flight mass spectrometry (PTR-ToF-MS). Talanta, 85, 386–93.
Sánchez López, J. A., Zimmermann, R., and Yeretzian, C., 2014. Insight into the time-
resolved extraction of aroma compounds during espresso coffee preparation: Online
monitoring by PTR-ToF-MS. Analytical Chemistry, 86, 11696–704.
Shaffer, S. A., Tang, K., Anderson, G. A., Prior, D. C., Udseth, H. R., and Smith, R. D.,
1997. A novel ion funnel for focusing ions at elevated pressure using electrospray ion-
ization mass spectrometry. Rapid Communications in Mass Spectrometry, 11, 1813–7.
Soukoulis, C., Aprea, E., Biasioli, F., Cappellin, L., Schuhfried, E., Märk, T. D., and
Gasperi, F., 2010. Proton transfer reaction time-of-flight mass spectrometry moni-
toring of the evolution of volatile compounds during lactic acid fermentation of
milk. Rapid Communications in Mass Spectrometry, 24, 2127–34.
Spitaler, R., Araghipour, N., Mikoviny, T., Wisthaler, A., Dalla Via, J., and Märk, T. D.,
2007. PTR-MS in enology: Advances in analytics and data analysis. International
Journal of Mass Spectrometry, 266 (1–3), 1–7.
Sulzer, P., Hartungen, E., Hanel, G., Feil, S., Winkler, K., Mutschlechner, P., Haidacher,
S., Schottkowsky, R., Gunsch, D., Seehauser, H., Striednig, M., Jürschik, S., Breiev,
K., Lanza, M., Herbig, J., Märk, L., Märk, T. D., and Jordan, A., 2014. A pro-
ton transfer reaction-quadrupole interface time-of-flight mass spectrometer (PTR-
QiTOF): High speed due to extreme sensitivity. International Journal of Mass
Spectrometry, 368, 1–5. doi:10.1016/j.ijms.2014.05.004 .
Sulzer, P., Jürschik, S., Agarwal, B., Kassebacher, T., Hartungen, E., Edtbauer, A.,
Petersson, F., Warmer, J., Holl, G., Perry, D., Mayhew, C. A., and Märk, T. D., 2012.
Designer drugs and trace explosives detection with the help of very recent advance-
ments in proton-transfer-reaction mass spectrometry (PTR-MS). Communications
in Computer and Information Science, 318, 366–75.
Taiti, C., Costa, C., Menesatti, P., Comparini, D., Bazihizina, N., Azzarello, E., Masi, E.,
and Mancuso, S., 2015. Class-modeling approach to PTR-TOFMS data: A peppers
case study. Journal of the Science of Food and Agriculture, 95, 1757–63.
240 Food Aroma Evolution

Tsevdou, M., Soukoulis, C., Cappellin, L., Gasperi, F., Taoukis, P. S., and Biasioli, F.,
2013. Monitoring the effect of high pressure and transglutaminase treatment of
milk on the evolution of flavour compounds during lactic acid fermentation using
PTR-ToF-MS. Food Chemistry, 138 (4), 2159–67.
van Ruth, S. M., Koot, A., Akkermans, W., Araghipour, N., Rozijn, M., Baltussen, M.,
Wisthaler, A., Märk, T. D., and Frankhuizen, R., 2007. Butter and butter oil clas-
sification by PTR-MS. European Food Research and Technology, 227, 307. https​://
do​i.org​/10.1​0 07/s​0 0217​- 007-​0724-​7.
Vita, F., Taiti, C., Pompeiano, A., Bazihizina, N., Lucarotti, V., Mancuso, S., and Alpi, A.,
2015. Volatile organic compounds in truffle (Tuber magnatum Pico): Comparison of
samples from different regions of Italy and from different seasons. Science Reports,
5, 12629. doi:10.1038/srep12629.
Wieland, F., Gloess, A. N., Keller, M., Wetzel, A., Schenker, S., and Yeretzian, C., 2012.
Online monitoring of coffee roasting by proton transfer reaction time-of-flight mass
spectrometry (PTR-ToF-MS): Towards a real-time process control for a consistent
roast profile. Analytical and Bioanalytical Chemistry, 402, 2531–43.
Yener, S., Romano, A., Cappellin, L., Granitto, P. M., Aprea, E., Navarini, L., Märk,
T. D., Gasperi, F., and Biasioli, F., 2015. Tracing coffee origin by direct injection
headspace analysis with PTR/SRI-MS. Food Research International, 69, 235–43.
Yener, S., Romano, A., Cappellin, L., Märk, T. D., Sánchez, J., and Gasperi, F., 2014.
PTR-ToF-MS characterisation of roasted coffees (C. arabica ) from different geo-
graphic origins. Journal of Mass Spectrometry, 929–35. doi:10.1002/jms.3455.
Yener, S., Sánchez-Lopez, J. A., Granitto, P. M., Cappellin, L., Märk, T. D., Zimmermann,
R., Bonn, G. K., Yeretzian, C., and Biasioli, F., 2016. Rapid and direct volatile com-
pound profiling of black and green teas (Camellia sinensis) from different countries
with PTR-ToF-MS. Talanta, 152, 45–53.
Yépez, A., Russo, P., Spano, G., Khomenko, I., Biasioli, F., Capozzi, V., and Aznar, R.,
2019. In situ riboflavin fortification of different kefir-like cereal-based beverages
using selected Andean LAB strains. Food Microbiology, 77, 61–8.
Zhu, L., Mikoviny, T., Morken, A. K., Tan, W., and Wisthaler, A., 2018. A compact and
easy-to-use mass spectrometer for online monitoring of amines in the flue gas of a
post-combustion carbon capture plant. International Journal of Greenhouse Gas
Control, 78, 349–53.
 Chapter 12
Stable Isotope Dilution Assay
Hans-Georg Schmarr

CONTENTS

12.1 Definitions and Historical Development 241

12.2 Principle of an Isotope Dilution Assay 242
12.3 Requirements for Labeled Standards and Some Considerations for Their Use 243
12.4 Optimization Strategies for SIDA-Based Applications 248
12.4.1 Non-MS SIDA Applications 248
12.4.2 H/C MDGC with SIDA-Based Quantification 249
12.5 Conclusion and Summary 252
References 254

12.1 DEFINITIONS AND HISTORICAL DEVELOPMENT

Quantitative analysis has various calibration approaches to instrumental methods. One

of these is the internal standard method that utilizes the addition (spiking) of a substance
(the internal standard) in a constant amount to all samples, blanks, and calibration stan-
dards. Calibration then involves plotting the ratio of the analyte signal to the internal
standard signal as a function of the analyte concentration of the standard. This ratio for
the samples is then used to obtain their analyte concentrations from a calibration curve
(Skoog, Holler, and Crouch, 2016). Spiking of the samples is done before further sample
preparation, extraction, and analysis. According to the IUPAC definition, in chromatog-
raphy, this internal standard is a compound added to a sample in a known concentration
to facilitate the qualitative identification and/or quantitative determination of the sample
components (McNaught and Wilkinson, 1997). It is generally acknowledged that such
an internal standard should be as near as possible to the analyte, without existing as such
within the sample matrix. The best available standard is a labeled analyte that does not
exist naturally.
The practice of the quantification of organic compounds utilizing the addition
of a labeled internal standard is called an isotope dilution assay (IDA) and was first
reported in 1940 for the analysis of fatty acids and amino acids in complex mixtures
by Rittenberg and Foster (Rittenberg and Foster, 1940). Furthermore, it was also rec-
ognized as a promising tool for chemical analysis in other areas, for example, following
the decay of radioactive nuclides, with sensitivity potentialities exceeding standard ana-
lytical chemistry procedures (Inghram, 1954). Later, Sweeley et al. described the benefi-
cial use of mass spectrometric (MS) detection for the determination of unresolved gas
chromatographic effluents, here in particular the isotopic derivatives (isotopologues or
also named isotopomers) of glucose and deuterated glucose (Sweeley et al., 1966). The

241
242 Food Aroma Evolution

principle of IDA was also used in the field of pharmacological chemistry (Sadee, Segal,
and Finn, 1973). The first application for quantification of flavorful food components
was described by Cobb in 1969, using radioactively labeled flavor compounds (Cobb,
1969). In 1976, a theoretical basis for isotope dilution was established by Pickup and
McPherson (1976), and it was proposed as a reference method with high accuracy and
precision (Björkhem et al., 1976; Cohen et al., 1980). An important early application
from the field of aroma research was published in 1987 by Schieberle and Grosch (1987),
who quantified acetylpyrazine, 2-methyl-3-ethylpyrazine, and 2-acetyl-l-pyrroline in
wheat bread crust with stable isotopes, thus naming it the “stable isotope dilution assay”
(SIDA). In the same year, Harris et al. published an early application of the trace-level
(ng/L) analysis of potent 2-methoxy-3-alkylpyrazines in wine (Harris et al., 1987). Since
then, in the 1990s, numerous analytical methods with a SIDA-based quantification for
a great number of potent aroma compounds were developed (Guth and Grosch, 1990,
1993; Sen and Grosch, 1991; Sen et al., 1991; Blank, Schieberle, and Grosch, 1993;
Cerny and Grosch, 1993; Grosch, 1993; Allen, Lacey, and Boyd, 1994; Semmelroch
et al., 1995; Kerscher and Grosch, 1998; Wagner and Grosch, 1998; Kotseridis et al.,
1999a,b; Kotseridis, Baumes, and Skouroumounis, 1999; Lin et al., 1999; Pfnuer et al.,
1999), and thereafter. Some of the earlier applications had been summarized together
with other developments in methods for the analysis of flavor compounds and their
precursors (Schieberle, 1995). A perspective paper dealing with important aspects for
the quantification of the sensory-active constituents of foods was published in 2012
(Schieberle and Molyneux, 2012). Noteworthy reviews from the application of SIDA
in the field of flavor analysis were contributed by Mosandl (1992), Schieberle (1995),
Allen and Lacey (1997), Blank et al. (1998), Milo and Blank (1998), and Werkhoff et al.
(2002). A basic description of how to apply a SIDA-based quantification has been sum-
marized by Milo (2005). Calibration issues in SIDA analyses with MS detection, par-
ticularly for a situation when overlapping ions exist in the fragmentation pattern of the
labeled standard and unlabeled analyte, were discussed in various publications (Colby
and McCaman, 1979; Sabot et al., 1988; Sabot and Pinatel, 1993; Sabot, 1994; Fay et
al., 2000; Fay, Metairon, and Baumgartner, 2001; Yang et al., 2006). Also worth men-
tioning are recent reviews targeting application fields beyond flavor analysis (Rychlik
and Asam, 2008, 2009; Rychlik, 2011). Today, SIDA can be considered as the accepted
state-of-the-art method for the most accurate and reliable quantitative determination of
volatile compounds in the flavor research field and elsewhere.

12.2 PRINCIPLE OF AN ISOTOPE DILUTION ASSAY

Most of natural elements show a natural distribution of (stable) isotopes; for example,
carbon consists of 98.9% 12C, 1.1% 13C, and traces of radioactive 14C, or hydrogen that
consists of 99,9885% 1H, 0,0115% 2H (deuterium; D), besides trace amounts of the labile
3H (tritium). With appropriate synthetic approaches, organic compounds (analytes) can

be labeled up to 100% with a minor (stable) isotope in a certain position, for example,
2 H, 13C, or 15 N. The synthesized compound exhibits a higher molecular weight than the

analyte, but almost identical physicochemical properties. Therefore, such “isotopomers”

or “isotopologues” are ideal internal standards for quantitative analysis. Methods based
on the use of labeled internal standards with stable isotopes are called “stable isotope
dilution assays.” The term “dilution” refers to the fact that the analyte is “diluted” with
its isotopomer, the internal standard (Figure 12.1). In a SIDA approach, losses of the
 Stable Isotope Dilution Assay 243

FIGURE 12.1 Origin of the term “dilution” in stable isotope dilution assays: addition of
a standard with a different isotopic distribution—the original isotopic distribution of the
analyte has been “diluted.” (Reprinted with permission from Rychlik and Asam, 2008.)

analyte caused by analytical procedures such as, for example, extraction, distillation, or
degradation are compensated, if the labeled isotopomer is added to the sample prior to the
workup procedure and conditions for reliable equilibration are provided (Figure 12.2).
This way, time-consuming recovery and spiking experiments, necessary with structurally
different internal standards, can be minimized. The quantitation can simply be done by
monitoring the target compound (or a fragment or molecular ion of the analyte) and the
corresponding signal for the isotopomeric internal standard.
The concentration of the analyte and its isotopomer is determined by GC or GC-MS.
In the literature, MS is often used as a detector, mainly due to its ability to discriminate
between the labeled and unlabeled compound, allowing quantification via the peak areas
of selective mass traces. With MS detection, the expression “stable isotope dilution–mass
spectrometry” (SID-MS) may also be found in the literature, whereas “non-MS SIDA”
can be found along with other detectors.

12.3 REQUIREMENTS FOR LABELED STANDARDS

AND SOME CONSIDERATIONS FOR THEIR USE

Since the isotopically labeled analogue of the analyte is used as a standard, some con-
siderations of the labeled standards are necessary. Labeling with deuterium (D) is pref-
erably used because the introduction of this isotope is relatively cheap when compared
with other isotopes. In order to be considered chemically stable, labeling with (stable)
isotopes should be performed in a non-exchangeable position. Under certain circum-
stances, 13C-labeling is strongly recommended, particularly as protons in the α-position
to carbonyl functions are known to enolize and may exchange with protons from the
sample. A possible deuterium/protium (D/H) exchange of such a labeled standard dur-
ing the analytical procedure then has to be ruled out, for example, by testing its stability
244 Food Aroma Evolution

FIGURE 12.2 The ratio of isotopologic analyte and standard remains stable until the
final mass spectroscopic analysis. For a structurally different internal standard, however,
the ratio between standard and analyte can alter during sample preparation. (Reprinted
with permission from Rychlik and Asam, 2008.)

under conditions used for isolating the aroma components, but also throughout the
instrumental analytical process. Examples for critical analytes with exchangeable pro-
tons within the structure that require labeling with 13C were shown, for example, for
4-hydroxy-2,5-dimethyl-3(2H)-furanone (Furaneol®), as illustrated in Figure 12.3 as
c-Ia (Blank et al., 1997) and c-Ib (Blank et al., 1997). On the other hand, its ethyl ana-
logue (homofuraneol) can only be deuterated in a well-defined position, as shown for
2(or 5)-[2,2,2,-2H3]ethyl-4-hydroxy-5(or 2)-methyl-(2H)-furanone (d-II) (Preininger and
Grosch, 1994; Blank et al., 1997). The additional CH 2 within the ethyl group serves as
a barrier that prevents a D/H-exchange. Other examples requiring 13C-labeling are, for
example, 3-hydroxy-4,5-dimethyl-2(5H)furanone (sotolone) c-III (Blank et al., 1996),
or 2,3-butandione (diacetyl) c-IV (Schieberle and Hofmann, 1997). The homologous

FIGURE 12.3 Preferred position and type of labeling of compounds containing an

α,β-dicarbonyl moiety (▪ indicates the labeling position with 13 C, ● with deuterium).
(Reprinted with permission from Milo and Blank, 1998.)
 Stable Isotope Dilution Assay 245

5-ethyl-3-hydroxy-4-methyl-2(5H)-furanone (V) (Blank, Schieberle, and Grosch, 1993),

and 2.3-pentanedione (VI) (Milo and Grosch, 1993) can be deuterated as long as a CH 2
group is located between the enolizing structure and the deuterated moiety.
An interesting observation of a D/H-exchange phenomenon was recently described
for the analysis of 2-aminoacetophenone (2-AAP) in wine, a compound associated with
the so-called atypical aging off-flavor (ATA). The method used 1-(2-​amino​pheny​l)-2,​
2,2-t​rideu​terio​-etha​none (2-AAP-d3) as an internal standard (Figure 12.4), and head-
space solid-phase microextraction (HS-SPME) for sample preparation (Schmarr, Keiser,
and Krautwald, 2016).
In this work, calibration was done with multipoint calibration curves, also including
repetitions over time. The repetition of the calibration curves obtained when samples
were measured in a time series with standards, then kept at a basic pH for longer periods
of time, showed different (increasing) slopes when area ratios were plotted as 2-AAP/2-
AAP-d3. Obviously, 2-AAP-d3 underwent a proton exchange in alkali solution as could
only be verified by analyzing 2-AAP-d3 at pH 9 versus 3.5 (Figure 12.5). In this case,
non-deuterated 2-AAP was built over time (Figure 12.5A). No proton exchange was
observed at a native wine pH of 3.5 (Figure 12.5B). These findings of the D/H-exchange
problem were in accordance with previous work in which 2-AAP was analyzed in milk
powder (Preininger and Ullrich, 2001). In this latter work, the isotopic standard had
actually been prepared via a proton exchange under highly alkaline conditions (pH 14).
Although 2-AAP-d3 seemed to be stable in an aqueous matrix, quantitation in milk pow-
der via SIDA, was not possible due to deuterium exchange. These authors stated that a
deuterated 2-AAP standard must therefore be labeled in non-CH acidic position for use
in milk powder.
Schmarr et al. discussed the differences observed between their work on wine aroma
and the previous study on milk powder, both using 2-AAP-d3 as an internal standard,
with respect to the kinetics for the proton exchange on the one hand and the partition-
ing and extraction between the aqueous and the headspace phases on the other. They
argue that as long as the extraction step is considerably faster than the D/H exchange, a
problem with SIDA-based quantification might not be detected. However, as HS-SPME
usually is a longer process (extraction times are often in the range of about an hour),
this may become critical, and a potential D/H exchange should thus be considered.
Another interesting aspect discussed in this work is the situation with autosamplers
that are often used in routine analyses today. Here, the analysis of multiple samples
might further complicate the story. If samples already spiked with an isotopic standard
remain on the sampler tray waiting for subsequent processing, then excessive proton

FIGURE 12.4 2-AAP and its isotopologue used as an internal standard for SIDA-based
quantification. (Reprinted with permission from Schmarr, Keiser, and Krautwald, 2016.)
246 Food Aroma Evolution

FIGURE 12.5 Stability of area ratios (2-AAP/2-AAP-d3) obtained for buffered model
wines spiked with a fixed amount of 2-AAP-d3. (A) Increasing area ratios over time are
a sign for proton exchange at pH 9. (B) Stable area ratios at a native wine pH (here pH
3.5) allow the use of 2-AAP-d3 as an internal standard for SIDA-based quantification.
(Reprinted with permission from Schmarr, Keiser, and Krautwald, 2016.)

exchange in the aqueous medium has to be expected, as was the case with 2-AAP-d3.
Within the time frame necessary for multipoint calibration runs, but also for a longer
series of samples resting on the sample tray, a back exchange of the deuterium atoms
occurs. Alternatively, in order to prevent (or minimize) the risk of a D/H exchange, the
internal standard would have to be added just before the explicit analysis of each sample.
Today, this might be realized when modern sampler technology is used. New sampler
generations (e.g., from CTC Analytics AG, Zwingen, Switzerland) can be equipped with
a tool exchange station, allowing a timed exchange of an injection tool (e.g., a micro-
liter syringe for spiking versus a SPME fiber tool for extraction). Such equipment then
allows the timed standard addition and allows finally a switching back to the SPME
fiber tool for further processing of the sample. This way, prolonged periods of potential
D/H-back-exchange can be minimized, ruling out the risk of a false SIDA quantification
(Schmarr, Keiser, and Krautwald, 2016).
Detection method should also be well-thought-out when considering the labeling
strategy beforehand. With common MS detection in SIDA applications, in order to
minimize interferences with masses originating from the analyte, the labeling should
 Stable Isotope Dilution Assay 247

increase the molecular weight of the standard by at least two units, preferably three
units, in order to minimize interferences with the natural isotope distribution of the
analyte (Fay et al., 2000; Milo, 2005). Depending on the fragmentation of the molecule
and the ionization method applied (electron impact (EI) or chemical ionization (CI)
with different reactant gases), it is also required that the labeled ion species is accessible,
and in trace-level analyses also of adequate abundance. Some aspects that exploit the
potential of mass spectrometry in this respect have been outlined in detail by Blank et al.
using trans-4,5-epoxy-(E)-2-decenal as an example (Blank et al., 1998; Lin et al., 1999).
Care should also be given to the proper selection of specific masses, as particularly in
trace-level analysis with complex matrices, the low-molecular weight and (sometimes
high) fragmentation of flavor compounds (with common EI) complicate reliable quan-
tification. Often, only low-molecular weight fragment ions of minor selectivity remain
and compete with ubiquitous fragment ions from the matrix. An example of such a
problematic situation was recently shown for α-ionone in wine that was impeded by
co-eluting compounds, although detection was performed with a generally considered
selective (and thus “safe”) detection method using MS/MS with a triple quadrupole
mass spectrometer (Langen, Wegmann-Herr, and Schmarr, 2016). Due to their scarce
presence in nature, labeled fragment ions usually do not exhibit such a problem and
result in far better signal-to-noise ratios for their peaks, but still, the analyte as such
must be detected and quantified without skepticism. With the availability of accurate
mass detection (highResMS), some of the problems discussed above might be circum-
vented, but up to now the high instrument costs prevent their common dissemination,
and applications are usually outside the field of flavor analysis. Still, a recent application
for the quantification of 19 aldehydes in human serum with HS-SPME-GC-highResMS
might serve as an inspiring example. This application used SIDA-based quantification
with 13C-labeled standards (Silva et al., 2018).
In the case of SIDA applications that do not involve mass spectrometric detection
(non-MS SIDA), the required isotope effect necessary for a chromatographic resolu-
tion that allows individual integration of both the standard and analyte depends on
the number of labeled atoms (particularly deuterium atoms as outlined below) incor-
porated, but also on the position within the structure of the molecule. The inverse
isotope effect (the deuterated (heavier) isotopologues are eluted earlier than the non-
deuterated (lighter) ones) in gas–liquid partitioning chromatography (GLC) is caused
by van der Waals (particularly London) dispersion forces. Apolar locations are favor-
able in this respect. With deuterated compounds, the fundamental difference in bond
lengths of C–H and of C–D is that the latter is shorter by about 0.005Å. Therefore,
molar van der Waals volumes are lower, resulting in lower London dispersion interac-
tions and an inverse isotope effect. Compared to 13C-isotopes, the reduction in the
van der Waals volume is more pronounced, and the isotope effect thus stronger with
deuterated compounds. This fundamental understanding of the isotope effect, par-
ticularly that it is higher with deuterated standards than with 13C isotopic compounds,
has to be considered when labeled standards for non-MS SIDA applications have to
be chosen. A detailed discussion on the nature of the isotope effect, examples of its
dependence on the substitution pattern, and the influence of chromatographic condi-
tions, particularly the contribution of the chemical nature of the stationary phase, has
been published recently and might inspire further studies (Schmarr et al., 2012b). The
experience gained in this latter study provided the basis for the optimization of multi-
dimensional GC applications using labeled internal standards, as will be discussed in
the next chapter.
248 Food Aroma Evolution

12.4 OPTIMIZATION STRATEGIES FOR

SIDA-BASED APPLICATIONS

12.4.1 Non-MS SIDA Applications

As already mentioned, besides the almost identical behavior of isotopic labeled standards,
a chromatographic separation of labeled and non-labeled compounds can be achieved and
was demonstrated in the 1960s, at the beginning of the field of GC (Bruner and Cartoni,
1963, 1965; Bruner, Cartoni, and Possanzini, 1969). The nature of the so-called isotope
effect, the elution order of the labeled and non-labeled analyte, was reviewed by Matucha
et al. (1991) and discussed in detail with particular emphasis on novel ionic liquid sta-
tionary phases in a later work (Schmarr et al., 2012b). Based on this knowledge, today,
optimization of the chromatographic separation and proper selection of the degree and
structural variation of the deuteration allows for fine-tuning of the isotopic separation,
and consequently an individual integration of the internal standard and analyte. This
is the prerequisite for non-MS based SIDA applications, as has been presented in recent
flavor analysis work using electron capture detection (ECD) for haloanisoles (Schmarr et
al., 2012a; Slabizki and Schmarr, 2013), pulsed flame photometric detection (PFPD) in a
sulfur mode for sulfur compounds (Koschinski et al., 2010; Schmarr et al., 2010; Ullrich,
Neef, and Schmarr, 2018), or a thermionic detector in a nitrogen-selective mode (NPD)
(Schmarr et al., 2010).
A low-cost, robust, selective, and sensitive non-MS detection, as with the PFPD
in the examples shown in Figure 12.6, favors the development of quantitative non-MS
SIDA-based methods and was recently demonstrated for a routine HS-SPME analysis of
low-molecular weight volatile sulfur compounds in wine (Ullrich, Neef, and Schmarr,
2018). Another application that benefits from the high selectivity and sensitivity of the
ECD is the analysis and SIDA-based quantification of off-flavor haloanisoles in wine
and cork soaks with limits of detection (LODs) in the sub-ng/L range (Schmarr et al.,
2012a; Slabizki and Schmarr, 2013). Based on an automated headspace solid-phase
microextraction (HS-SPME), the latter method only needs marginal sample preparation
and achieved LODs for the most relevant cork off-flavor compounds, such as 2,4,6-tri-
chloroanisole (TCA), 2,3,4,6-tetrachloroanisole (TeCA), and 2,4,6-tribromoanisole
(TBA) well below their sensory threshold values. It is noteworthy that for the complex
matrix situations (here wine), reliable trace-level quantification had only been achieved
after applying heart-cutting multidimensional gas chromatography (H/C MDGC). Such
trace-level non-MS SIDA quantification was possible because the necessary chromato-
graphic resolution of the internal standard and the target analyte peaks were obtained
with highly deuterated [2H 5]-isotopologues and well-chosen chromatographic condi-
tions (Figure 12.7).
Recently, an interesting isotopic separation of the highly volatile compound acet-
aldehyde and its deuterated isotopologue acetaldehyde-2,2,2-d3 was achieved in a
temperature-programmed run on a porous layer open tubular (PLOT) capillary col-
umn coated with particles of divinyl-benzene ethylene glycol/dimethylacrylate (Rt ® -U-
BOND). In a preliminary study, static headspace extraction and gas chromatographic
separation (HS-GC-FID) of acetaldehyde from aqueous solutions was shown as an
application. For detection, the method used a flame ionization detector (FID) that rep-
resents a robust and low-cost alternative to MS detection. Good linearity was obtained
in a calibration range from 0.4 to 40 mg/L, with peak integration benefitting from the
inverse isotope effect encountered on the specific porous polymer (Figure 12.8 and
 Stable Isotope Dilution Assay 249

FIGURE 12.6 Separation of thiols and their [2H10]-isotopologues on a polyethylene gly-

col stationary phase with PFPD detection (S-mode) and enantioseparation of 3MH and
[2H10]-3MH and their isotopologues on AlphaDex 120 (*). Isothermal oven temperatures
were at 120°C; (a) 3MH; polyethylene glycol; (b) 3MHA; polyethylene glycol, 90°C;
(c) 4MMP; polyethylene glycol, 140°C and 100°C; and (d): AlphaDex 120, respectively.
Hydrogen was used as carrier gas at 4 mL min-1; besides AlphaDex 120, which was at
75 kPa constant inlet pressure. Fused silica column dimensions were 30 m × 0.32 mm i.d.
(0.25 mm i.d. for AlphaDex 120). All axis captions as for 3MH (a). (Results presented
earlier in Schmarr et al., 2010.)

12.9). Furthermore, separation of methanol and deuterated methanol (d3) could also be
achieved under the same chromatographic conditions (Figure 12.10) (Schmarr, Wacker,
and Mathes, 2017).
Although non-MS SIDA applications were shown to be promising alternatives to
those with MS detection, their occurrence in the literature is still scarce. Maybe the exam-
ples presented here will inspire future method development in this direction. Common
arguments against the omnipresent detection with MS in the scientific literature are par-
ticularly instrument cost, maintenance costs, and the required operator training level.

12.4.2 H/C MDGC with SIDA-Based Quantification

Fundamental knowledge of the isotope effect might also be utilized for optimization
strategies in H/C MDGC applications, when cut windows should be minimized in order
250 Food Aroma Evolution

FIGURE 12.7 HS-SPME-MDGC-ECD 2D chromatograms of (A) TCA-d5/TCA and

(B) TBA-d5/TBA standards. The indicated shoulder visible at the peak of TCA-d5 indi-
cates co-elution, problematic for quantification on a 35% diphenylpolysiloxane station-
ary phase column. Only minor co-elution (shoulder) of a system background compound
(lowest trace); chromatogram (B) with TBA on TG-1301MS. Integration of TBA is not
hampered for investigated calibration ranges (overlayed traces). (Reprinted with permis-
sion from Slabizki and Schmarr, 2013.)
to avoid co-transfers of potentially interfering matrix compounds from a 1D to the 2D
separation column (Schmarr, Slabizki, and Legrum, 2013) (Figure 12.11).
An example of this optimization strategy favoring narrow-cut windows was presented
with the trace-level analyses of 3-alkyl-2-methoxypyrazines (3-iso-propyl-2-methoxypyr-
azine (IPMP), sec-butyl-2-methoxypyrazine (SBMP), and 3-iso-butyl-2-methoxypyrazine

FIGURE 12.8 Separation of deuterated (d3) and non-deuterated (nd) acetaldehyde after
HS-GC-FID analysis on a 30 m × 0.32 mm i.d. fused silica capillary, coated with 10 µm
of Rt-U-Bond. (Reprinted with permission from Schmarr, Wacker, and Mathes, 2017.)
 Stable Isotope Dilution Assay 251

FIGURE 12.9 Calibration curve shows good linearity for acetaldehyde in the calibrated
range from 0.4 to 40 mg/L. (Reprinted with permission from Schmarr, Wacker, and
Mathes, 2017.)

(IBMP) in a complex matrix (Schmarr, Slabizki, and Legrum, 2013). In addition to SIDA-
based quantification with deuterated internal standards, enantioseparation of the chiral
SBMP occurred on an enantioselective 2D separation column. The authors discussed in
detail the benefit of their minimization strategy in order to reduce the transfer of pos-
sibly co-eluting matrix compounds. A crucial point in their application was to choose a

FIGURE 12.10 Separation of deuterated (d4 or rather d3 after proton exchange in aque-
ous solution) and non-deuterated methanol after HS-GC-FID analysis on a 30 m × 0.32
mm i.d. fused silica capillary, coated with 10 µm of Rt-U-Bond. (Reprinted with permis-
sion from Schmarr, Wacker, and Mathes, 2017.)
252 Food Aroma Evolution

FIGURE 12.11 Strategy to optimize H/C MDGC with SIDA-based quantification. (a)
Isotopic separation due to a pronounced (normal) isotope effect in 1D requires a wide-
cut window. The window size depends on peak widths and the resolution (R s) of the
isotopic standard and the analyte (*). (b) Narrow-cut windows with a 1D separation
that show no or only a marginal isotope effect. Mandatory widths for cut windows to
transfer the analyte (R-H n) or the labeled (here deuterated) internal standard (R-D n)
are indicated with arrows in black, whereas arrows in light gray indicate optional
transfer periods to ensure a complete transfer.

suitable chemical composition for the 1D column stationary phase. A medium polar ionic
liquid phase (SLB-IL60) resulted in an insignificant 1D separation between labeled and
unlabeled compounds. Critical aspects of such narrow-cut windows with a partial trans-
fer versus a complete transfer were discussed, particularly with respect to consequences
for the resulting quantitative data as well as for the calculated enantiomeric composition.
Some of these results will also be presented hereafter.
First, the increase in selectivity with a narrow- (18 s) versus a wide- (42 s) cut win-
dow was obvious and is demonstrated in Figure 12.12. With a transfer period of 42
s, a major co-eluting peak was seen at the 2D retention of SBMP, that was not trans-
ferred with a transfer period of only 18 s (Figure 12.12b), the latter allowing undisturbed
quantification on the corresponding quantifier fragment ion traces (Figure 12.12c). On
the other hand, an excessively narrow-cut window in 1D has to be avoided as often an
intended 1D co-elution is not perfect, and partial transfer of the labeled and unlabeled
compound then results in erroneous data when using SIDA-based quantification (Figure
12.13). Otherwise, in enantio-MDGC with an achiral stationary phase in 1D and an
enantioselective 2D separation, a partial transfer of the 1D peak does not influence the
enantiomeric composition as the achiral 1D separation cannot separate the enantiomers
(Figure 12.13c).

12.5 CONCLUSION AND SUMMARY

Isotopic dilution is probably the most precise and accurate method available today to
quantify flavor compounds, particularly if the internal standard can be homogeneously
distributed within the sample matrix. However, some considerations are necessary
beforehand, concerning the type and chemical nature of the labeling with respect to
the stability of the labeled compound in the sample matrix, but also with respect to the
method used for detection. With common MS detection, an overlap of standard and
 Stable Isotope Dilution Assay 253

FIGURE 12.12 H/C enantio-MDGC with SIDA-based quantification of MPs in galba-

num oil. (Copyright by Springer Nature, reproduced with permission.) (a) 1D Separation
on SLB-IL60 as pre-column; (a*) cut windows. (b) Separation on the enantioselective 2D
column (Lipodex G ®), upper trace with 42 s cut windows (each) or 18 s (lower trace);
absolute scaling intensities given for estimation of real peak size. (c) Quantification on
narrow, 18 s cut windows via quantifier ions: m/z 137 (140), 138 (141), and 124 (127) for
IPMP (d3 -IPMP), SBMP (d3 -SBMP), and IBMP (d3-IBMP), respectively. (Reprinted with
permission from Schmarr, Slabizki, and Legrum, 2013.)

analyte ions should be avoided; otherwise calibration curves have to take the theoreti-
cal and measured isotopic enrichment into account. It is also evident that labeled ions
should be selective and of reasonable abundance, with some applications then demanding
other ionization techniques than standard electron impact ionization. In non-MS SIDA
applications, labeling strategy and chromatographic optimization have to be optimized
to guarantee a good chromatographic separation of standard and analyte, allowing their
individual signal integration. Since H/C MDGC is a common method for quantifying
flavor compounds in complex matrices and/or in trace-level situations, optimization
strategies outlined here allow for the minimization of cut windows, and thus the risk of
transferring potentially co-eluting compounds from 1D to 2D.
254 Food Aroma Evolution

FIGURE 12.13 Consequences of complete or partial transfer in MDGC analysis.

(Copyright by Springer Nature, reproduced with permission.) (a) Co-eluting isotopic MPs
in 1D allow narrow-cut windows, but complete or partial transfer conditions (as indi-
cated by the arrows) must be considered. (b) Influence on peak area ratios of unlabeled/
labeled MP (n=2). (c) Influence on peak area ratios for (S)/(R)-SBMP (n=2). (Reprinted
with permission from Schmarr, Slabizki, and Legrum, 2013.)

REFERENCES

Allen, M. S. and M. J. Lacey. 1997. Stable isotope dilution gas chromatography–mass

spectrometry for determination of methoxypyrazines (“green” aroma) in wine. In
Modern Methods of Plant Analysis, eds. H. F. Linskens and J. F. Jackson, pp. 193–
210. Berlin, Germany: Springer.
Allen, M. S., M. J. Lacey, and S. Boyd. 1994. Determination of methoxypyrazines in red
wines by stable isotope dilution gas chromatography-mass spectrometry. Journal of
Agricultural and Food Chemistry 42 (8):1734–8.
Björkhem, I., R. Blomstrand, O. Lantto, L. Svensson, and G. Ohman. 1976. Toward
absolute methods in clinical chemistry: Application of mass fragmentography to
high-accuracy analyses. Clinical Chemistry 22 (11):1789–801.
Blank, I., C. Milo, J. Lin, and L. B. Fay. 1998. Quantification of aroma-impact com-
ponents by isotope dilution assay—Recent developments. In Flavor Chemistry:
30 Years of Progress, Proceedings of an American Chemical Society Symposium,
Boston, August 23–7, 1998, eds. R. Teranishi, E. L. Wick, and I. Hornstein, pp.
63–74. New York, NY: Kluver Academic/Plenum Publishers, 1999.
Blank, I., J. Lin, R. Fumeaux, D. H. Welti, and L. B. Fay. 1996. Formation of 3-hydroxy-
4,5-dimethyl-2(5H)-furanone (sotolone) from 4-hydroxy-L-isoleucine and 3-ami​
no-4,​5-dim​ethyl​-3,4-​dihyd​ro-2(​5H)-f​urano​ne. Journal of Agricultural and Food
Chemistry 44 (7):1851–6.
 Stable Isotope Dilution Assay 255

Blank, I., L. B. Fay, F. J. Lakner, and M. Schlosser. 1997. Determination of 4-hydroxy-

2,5-dimethyl-3(2H)-furanone and 2(or 5)-ethyl-4-hydroxy-5(or 2)-methyl-3(2H)-
furanone in pentose sugar-based Maillard model systems by isotope dilution assays.
Journal of Agricultural and Food Chemistry 45 (7):2642–8.
Blank, I., P. Schieberle, and W. Grosch. 1993. Quantification of the flavor compounds
3-hydroxy-4,5-dimethyl-2(5H)-furanone and 5-eth​yl-3-​hydro​x y-4-​methy​l-2(5​H)-fu​
ranon​e by a stable isotope dilution assay. In Progress in Flavour and Precursor
Studies, eds. P. Schreier and P. Winterhalter, pp. 103–9. Würzburg, Germany:
Allured Publishing Corporation.
Bruner, F. and G. P. Carton. 1965. Gas chromatographic separation of deuterated meth-
anes. Journal of Chromatography 18 (2):390–1.
Bruner, F. and G. P. Cartoni. 1963. Gas chromatographic separation of benzene and
deuterobenzenes. Journal of Chromatography 10 (3):396.
Bruner, F., G. P. Cartoni, and M. Possanzini. 1969. Separation of isotopic methanes by
gas chromatography. Analytical Chemistry 41 (8):1122–4.
Cerny, C. and W. Grosch. 1993. Quantification of character-impact odor compounds
of roasted beef. Zeitschrift für Lebensmittel-Untersuchung und -Forschung 196
(5):417–22.
Cobb, W. Y. 1969. Quantitation of flavorful food components using isotope dilution.
Journal of Food Science 34 (5):466–9.
Cohen, A., H. S. Hertz, J. Mandel, R. C. Paule, R. Schaffer, L. T. Sniegoski, T. Sun,
M. J. Welch, and E. White. 1980. Total serum cholesterol by isotope dilution/
mass spectrometry: A candidate definitive method. Clinical Chemistry 26 (7):
854–60.
Colby, B. N. and M. W. McCaman. 1979. A comparison of calculation procedures for
isotope dilution determinations using gas chromatography mass spectrometry.
Biomedical Mass Spectrometry 6 (6):225–30.
Fay, L. B., S. Metairon, J. Lin, and I. Blank. 2000. Stable isotope dilution assay mass spec-
trometry in flavour research: Internal standard and calibration issues. In Frontiers
of Flavour Science, Proceedings of the 9th Weurman Flavour Research Symposium,
Freising, Germany, June 22–5, 1999, ed. P. Schieberle, pp. 125–31. Garching,
Germany: Deutsche Forschungsanstalt für Lebensmittelchemie, 2000.
Fay, L. B., S. Metairon, and M. Baumgartner. 2001. Linearization of second-order cali-
bration curves in stable isotope dilution-mass spectrometry. Flavour and Fragrance
Journal 16 (3):164–8.
Grosch, W. 1993. Detection of potent odorants in foods by aroma extract dilution analy-
sis. Trends in Food Science & Technology 4 (3):68–73.
Guth, H. and W. Grosch. 1990. Deterioration of soybean oil: Quantification of primary
flavor compounds using a stable isotope dilution assay. Lebensmittel-Wissenschaft
und -Technologie 23 (6):513–22.
Guth, H. and W. Grosch. 1993. Quantitation of potent odorants of virgin olive oil by
stable-isotope dilution assays. Journal of the American Oil Chemist’ Society 70
(5):513–8.
Harris, R. L. N., M. J. Lacey, W. V. Brown, and M. S. Allen. 1987. Determination of
2-methoxy-3-alkylpyrazines in wine by gas chromatography/mass spectrometry.
Vitis 26:201–7.
Inghram, M. G. 1954. Stable isotope dilution as an analytical tool. Annual Review of
Nuclear Science 4:81–92.
256 Food Aroma Evolution

Kerscher, R. and W. Grosch. 1998. Quantification of 2-methyl-3-furanthiol, 2-furfu-

rylthiol, 3-mercapto-2-pentanone, and 2-mercapto-3-pentanone in heated meat.
Journal of Agricultural and Food Chemistry 46 (5):1954–8.
Koschinski, S., S. Pan, W. Sang, T. Potouridis, U. Fischer, and H.-G. Schmarr. 2010.
Determination of aroma relevant thiols in wines based on gas chromatographic
analysis using large volume injection and pulsed flame photometric detection. In
Frontiers of Flavour Science, Proceedings of the 9th Weurman Flavour Research
Symposium, Freising, Germany, June 22–5, 1999, ed. P. Schieberle, pp. 125–31.
Garching, Germany: Deutsche Forschungsanstalt für Lebensmittelchemie, 2000.
Kotseridis, Y., R. L. Baumes, A. Bertrand, and G. K. Skouroumounis. 1999a. Quantitative
determination of 2-methoxy-3-isobutylpyrazine in red wines and grapes of Bordeaux
using a stable isotope dilution assay. Journal of Chromatography A 841 (2):229–37.
Kotseridis, Y., R. L. Baumes, A. Bertrand, and G. K. Skouroumounis. 1999b. Quantitative
determination of β-ionone in red wines and grapes of Bordeaux using a stable iso-
tope dilution assay. Journal of Chromatography A 848 (1+2):317–25.
Kotseridis, Y., R. L. Baumes, and G. K. Skouroumounis. 1999. Quantitative determina-
tion of free and hydrolytically liberated β-damascenone in red grapes and wines
using a stable isotope dilution assay. Journal of Chromatography A 849 (1):245–54.
Langen, J., P. Wegmann-Herr, and H.-G. Schmarr. 2016. Quantitative determination
of α-ionone, β-ionone, and β-damascenone and enantiodifferentiation of α-ionone
in wine for authenticity control using multidimensional gas chromatography with
tandem mass spectrometric detection. Analytical and Bioanalytical Chemistry 408
(23):6483–96.
Lin, J., L. B. Fay, D. H. Welti, and I. Blank. 1999. Synthesis of trans-4,5-epoxy-(E)-2-de-
cenal and its deuterated analog used for the development of a sensitive and selective
quantification method based on isotope dilution assay with negative chemical ion-
ization. Lipids 34 (10):1117–26.
Matucha, M., W. Jockisch, P. Verner, and G. Anders. 1991. Isotope effect in gas–liq-
uid chromatography of labelled compounds. Journal of Chromatography 588
(1–2):251–8.
McNaught, A. D. and A. Wilkinson. 1997. IUPAC. Compendium of Chemical
Terminology, 2nd ed. (the “Gold Book”). Oxford, UK: Blackwell Scientific
Publications.
Milo, C. 2005. Identification and quantitation of aroma compounds. In Handbook of
Food Analytical Chemistry, Pigments, Colorants, Flavors, Texture, and Bioactive
Food Components, eds. R. E. Wrolstad, T. E. Acree, E. A. Decker, M. H. Penner, D.
S. Reid, S. J. Schwartz, C. F. Shoemaker, D. M. Smith, and P. Sporns, pp. 245–56.
Hoboken, NJ: John Wiley & Sons.
Milo, C. and I. Blank. 1998. Quantification of impact odorants in food by isotope dilu-
tion assay: Strength and limitations. ACS Symposium Series 705:250–9.
Milo, C. and W. Grosch. 1993. Changes in the odorants of boiled trout (Salmo fario)
as affected by the storage of the raw material. Journal of Agricultural and Food
Chemistry 41 (11):2076–81.
Mosandl, A. 1992. Capillary gas chromatography in quality assessment of flavors and
fragrances. Journal of Chromatography 624 (1–2):267–92.
Pfnuer, P., T. Matsui, W. Grosch, H. Guth, T. Hofmann, and P. Schieberle. 1999.
Development of a stable isotope dilution assay for the quantification of 5-methyl-(E)-
2-hepten-4-one: Application to hazelnut oils and hazelnuts. Journal of Agricultural
and Food Chemistry 47 (5):2044–7.
 Stable Isotope Dilution Assay 257

Pickup, J. F. and K. McPherson. 1976. Theoretical considerations in stable isotope dilu-

tion mass spectrometry for organic analysis. Analytical Chemistry 48 (13):1885–90.
Preininger, M. and F. Ullrich. 2001. Trace compound analysis for off-flavor characteriza-
tion of micromilled milk powder. ACS Symposium Series 782:46–61.
Preininger, M. and W. Grosch. 1994. Evaluation of key odorants of the neutral vola-
tiles of Emmentaler cheese by the calculation of odor activity values. Lebensmittel-
Wissenschaft und -Technologie 27 (3):237–44.
Rittenberg, D. and G. L. Foster. 1940. A new procedure for quantitative analysis by
isotope dilution with application to the determination of amino acids and fat acids.
Journal of Biological Chemistry 133:737–44.
Rychlik, M. 2011. Stable isotope dilution assays in vitamin analysis—A review of principles
and applications. In Fortified Foods with Vitamins: Analytical Concepts to Assure
Better and Safer Products, ed. M. Rychlik, pp. 3–18. Weinheim, Germany: Wiley-VCH.
Rychlik, M. and S. Asam. 2008. Stable isotope dilution assays in mycotoxin analysis.
Analytical and Bioanalytical Chemistry 390 (2):617–28.
Rychlik, M. and S. Asam. 2009. Stable isotope dilution assays for quantitation of
organic trace compounds in food analysis. Umweltwissenschaften und Schadstoff-
Forschung 21 (5):470–82.
Sabot, J. F. 1994. Conceptions and misconceptions in stable isotope dilution. Fundamental
mathematical considerations. Analusis 22 (8):381–91.
Sabot, J. F., B. Ribon, L. P. Kouadio-Kouakou, H. Pinatel, and R. Mallein. 1988.
Comparison of two calculation procedures for gas chromatography–mass spectrom-
etry associated with stable isotope dilution. Analyst 113 (12):1843–7.
Sabot, J. F. and H. Pinatel. 1993. Calculation of the confidence range in order to obtain a
linear calibration graph in stable isotope dilution mass spectrometry: Application to
reference methods and pharmacological studies. Analyst 118 (7):831–4.
Sadee, W., J. Segal, and C. Finn. 1973. Diazoxide urine and plasma levels in humans
by stable-isotope dilution-mass fragmentography. Journal of Pharmacokinetics and
Biopharmaceutics 1 (4):295–305.
Schieberle, P. 1995. New developments in methods for analysis of volatile flavor com-
pounds and their precursors. In Characterization of Food—Emerging Methods, ed.
A. G. Gaonkar, pp. 403–31. Amsterdam, the Netherlands: Elsevier.
Schieberle, P. and R. J. Molyneux. 2012. Quantitation of sensory-active and bioactive
constituents of food: A Journal of Agricultural and Food Chemistry perspective.
Journal of Agricultural and Food Chemistry 60 (10):2404–8.
Schieberle, P. and T. Hofmann. 1997. Evaluation of the character impact odorants in
fresh strawberry juice by quantitative measurements and sensory studies on model
mixtures. Journal of Agricultural and Food Chemistry 45 (1):227–32.
Schieberle, P. and W. Grosch. 1987. Quantitative analysis of aroma compounds in wheat
and rye bread crusts using a stable isotope dilution assay. Journal of Agricultural
and Food Chemistry 35 (2):252–7.
Schmarr, H.-G., J. Keiser, and S. Krautwald. 2016. An improved method for the analysis
of 2-aminoacetophenone in wine based on headspace solid-phase microextraction
and heart-cut multidimensional gas chromatography with selective detection by tan-
dem mass spectrometry. Journal of Chromatography A 1477:64–9.
Schmarr, H.-G., S. Koschinski, W. Sang, and P. Slabizki. 2012a. Trace level analysis of
corky off-flavor compounds: Development of a new analytical method based on
solid phase extraction and analysis by multidimensional gas chromatography with
mass spectrometric detection. Journal of Chromatography A 1226:96–102.
258 Food Aroma Evolution

Schmarr, H.-G., C. Sept, U. Fischer, and S. Koschinski. 2010. Stable isotpe dilution
analysis with element-specific rather than mass spectrometric detection. Poster
Presentation, 34th International Symposium on Capillary Chromatography (ISCC)
and the 7th GC×GC Syposium, May 30–June 4, 2010, Riva del Garda, Italy.
Schmarr, H.-G., M. Wacker, and M. Mathes. 2017. Isotopic separation of acetaldehyde
and methanol from their deuterated isotopologues on a porous layer open tubular
column allows quantification by stable isotope dilution without mass spectrometric
detection. Journal of Chromatography A 1481:111–5.
Schmarr, H.-G., P. Slabizki, and C. Legrum. 2013. Optimization in multidimensional gas
chromatography applying quantitative analysis via a stable isotope dilution assay.
Analytical and Bioanalytical Chemistry 405 (20):6589–93.
Schmarr, H.-G., P. Slabizki, S. Müntnich, C. Metzger, and E. Gracia-Moreno. 2012b.
Ionic liquids as novel stationary phases in gas liquid chromatography: Inverse or
normal isotope effect? Journal of Chromatography A 1270 (0):310–7.
Semmelroch, P., G. Laskawy, I. Blank, and W. Grosch. 1995. Determination of potent
odorants in roasted coffee by stable isotope dilution assays. Flavour and Fragrance
Journal 10 (1):1–7.
Sen, A., G. Laskawy, P. Schieberle, and W. Grosch. 1991. Quantitative determination of
β-damascenone in foods using a stable isotope dilution assay. Journal of Agricultural
and Food Chemistry 39 (4):757–9.
Sen, A. and W. Grosch. 1991. Synthesis of six deuterated sulfur-containing odorants
for use as internal standards in quantification assays. Zeitschrift für Lebensmittel-
Untersuchung und-Forschung 192 (6):541–7.
Silva, L. K., G. A. Hile, K. M. Capella, M. F. Espenship, M. M. Smith, V. R. De Jesús,
and B. C. Blount. 2018. Quantification of 19 aldehydes in human serum by head-
space SPME/GC/High-resolution mass spectrometry. Environmental Science &
Technology 52 (18):10571–9.
Skoog, D. A., F. J. Holler, and S. R. Crouch. 2016. Principles of Instrumental Analysis,
7th ed. Boston, MA: Cengage Learning.
Slabizki, P. and H.-G. Schmarr. 2013. Analysis of corky off-flavour compounds at ultra
trace level with multidimensional gas chromatography-electron capture detection.
Journal of Chromatography A 1271 (1):181–4.
Sweeley, C. C., W. H. Elliott, I. Fries, and R. Ryhage. 1966. Mass spectrometric deter-
mination of unresolved components in gas chromatographic effluents. Analytical
Chemistry 38 (11):1549–53.
Ullrich, S., S. K. Neef, and H.-G. Schmarr. 2018. Headspace solid-phase microextrac-
tion and gas chromatographic analysis of low-molecular-weight sulfur volatiles with
pulsed flame photometric detection and quantification by a stable isotope dilution
assay. Journal of Separation Science 41 (4):899–909.
Wagner, R. K. and W. Grosch. 1998. Key odorants of French fries. Journal of the
American Oil Chemists’ Society 75 (10):1385–92.
Werkhoff, P., S. Brennecke, W. Bretschneider, and H.-J. Bertram. 2002. Modern meth-
ods for isolating and quantifying volatile flavor and fragrance compounds. In Food
Science and Technology, ed. R. Marsili, pp. 139–204. Boca Raton, FL: CRC Press.
Yang, Z.-Y., E. Y. Zeng, J.-Z. Wang, and B.-X. Mai. 2006. A numerical scheme to diag-
nose interferences in gas chromatography–mass spectrometry quantitation of coe-
luting isotopically labeled and unlabeled counterparts with partially overlapping ion
profiles. Journal of Chromatography A 1116 (1):265–71.
 Section III
Principles of Processing,
Evolution, and Modification
 Chapter 13
Food Processing, Cooking, and
Aging: A Practical Case Study
Emmanuel Bertrand

CONTENTS

13.1 Introduction 261

13.2 Elaboration of Processed Cheese 262
13.3 Process Making for Processed Cheese 266
13.4 Reactions Occurring during the Cooking of Processed Cheese 268
13.4.1 Lipid Degradation 270
13.4.2 The Maillard Reaction 272
13.4.3 Caramelization Reactions 272
13.5 Identification of Flavor Modifications during Processing 274
13.6 Conclusion 277
References 277

13.1 INTRODUCTION

The conservation of foodstuffs has always been a top priority concern for human beings.
One of the oldest examples of processed food is the discovery of fermented kefir, a dairy
product, in China more than 4000 years ago (Yang et al., 2014). How to ensure the per-
manent supply of perishable goods that are produced in agreement with the seasons? How
to guarantee safe (with respect to the microbiological risk of spoilage) and well-balanced
food (with respect to energy and nutritional needs)? These are two questions that scien-
tists are still trying to answer with the combination of our modern industrial tools and
knowledge but in a context of high demographic pressure and possible resource scarcity
in the future. In the past, the discovery of fire to cook food might have been one of the
explanations for the successful evolution of Homo sapiens (Gibbons, 2007). Traditional
preservation processes such as fermentation (yogurt, cheese, wine, and beer making as
examples), drying (meat, fish, fruits, and vegetables) and sugar addition (jams and marma-
lades) have enabled the preservation of many food products with different consequences:
an acidification of the environment and the installation of a microbiologically safe and
edible microflora (cheese and yogurt), a reduction of water activity (aw), and thus a reduc-
tion of the associated spoilage reactions (Figure 13.1). However, in parallel, it is observed
that the organoleptic properties of the processed products are drastically modified by tex-
ture changes and the apparition of newly formed compounds that may be either desirable
or in some cases rejected by consumers. Modern preservation techniques using hot- and
cold-thermal technologies and alternative technologies (such as high pressure or ionization

261
262 Food Aroma Evolution

FIGURE 13.1 Water activity (aw) and spoilage reaction rates. (Adapted from Labuza
and Dugan, 1971; Filey and Given, 1986, reproduced by Bertrand et al., 2018, with
permission.)

technologies) have helped to overcome some of these aspects by trying to keep foodstuffs
in a condition that is as close as possible to a native and fresh state. More recently, the
development of standardization and industrialization, the development of marketing tech-
niques (and product segmentation), and the analyses of consumer preferences have led to
the development of ultra-processed food. In this case, additives are added for technologi-
cal (to facilitate the production processes) or marketing (colorings and aromas to increase
consumer palatability and trigger product purchase or re-purchase) purposes rather than
nutritional or hedonistic reasons (Monteiro, 2009). In this chapter, we will focus on the
development of the aromatic properties of products during their manufacture and storage
through the example of processed cheese. This example is of interest as processed cheese
comes from the second processing of milk. Its manufacture involves the mixing, heating,
and texturing of dairy (cheese, butter, and milk powders) and non-dairy products (emul-
sifiers, citric acid, and sodium chloride). The result is a homogeneous product, generally
spreadable and with a long shelf life, often over 6 months. Some of the raw materials used
have already been previously processed, either by microbiological (Cheddar cheese) or
thermal (milk powder) processes, which increases the potential for the origin of the newly
formed molecules. During its manufacture and storage, lipid oxidation, caramelization,
and Maillard reactions form odorous compounds, some of which are potentially undesir-
able for the flavor of the product.

13.2 ELABORATION OF PROCESSED CHEESE

Figure 13.2 represents all the ingredients and additives authorized for the production of
processed cheese. Among them, several are optional and provide a specific texture or
aroma to the product. Six ingredients (solid line) are essential: a cheese matrix, milk
powder, milk fat, sodium chloride, water, and emulsifiers (Caric, 2000; Commission du
 Food Processing, Cooking, and Aging: A Case Study 263

FIGURE 13.2 Authorized ingredients for the production of processed cheese (full line:
basic ingredients; dotted line: optional ingredients). (According to Caric, 2000 and
Commission du Codex Alimentarius, 1995.)

Codex Alimentarius, 1995). Because of its success, the cheese matrix is now made specifi-
cally to be processed and is no longer considered as an efficient way to remanufacture
cheeses with visual defects (cracks or holes). Cheddar, Gouda, Emmental, and Mozzarella
are generally used alone or in a mixture. For fresh processed cheeses, the cheese matrix
used is generally a freshly prepared curd without any ripening. The criteria considered for
the selection of the cheese matrix to be processed are the contribution to the flavor, the
contribution to the texture, nutritional, economic, and market considerations. The ripen-
ing time of the cheese matrix is one of the most important parameters. It determines both
the aromatic profile and the content of certain simple sugars. The microbiological reac-
tions that occur during the ripening of the cheese matrix are the origin of several volatile
compounds with interesting flavoring properties, such as diacetyl, or methylated alde-
hydes. Figure 13.3 presents the major biochemical origins of these volatile compounds,
derived from lactose, lipid, and protein catabolism (McSweeney, 2004; Marilley and
Casey, 2004). Some of these volatile compounds can also be obtained using the Maillard
reaction such as, for example, 2,3-butanedione, 2-methylpropanal, and more generally
Strecker aldehydes, 2,5-dimethylpyrazine, and tetramethylpyrazine. Other compounds
can also be obtained by lipid degradation reactions such as butanoic acid or methylated
ketones. Figure 13.4 represents the catabolism of the amino acid leucine that leads to the
formation of 3-methylbutanal. It is also obtained during Strecker degradation when leu-
cine is involved in the reaction. Brickley et al. (2007) obtained confocal microscopic
photographs of three processed cheeses produced under the same manufacturing condi-
tions from Cheddar cheeses ripened for 7 (A), 28 (B), and 168 (C) days, respectively. A
reduction in the average diameter of the fat globules from 120 μm for processed cheese
made from 7-day aged Cheddar to about 2 μm for processed cheese made from 168-day
aged Cheddar was observed. The ripening of the Cheddar cheese matrix resulted in an
increase in small peptides and free amino acids that tend to stabilize the lipid–water inter-
face better than proteins. Bley et al. (1985a,b) measured the intensity of non-enzymatic
browning using the color index (ΔE) resulting from the color difference between samples
manufactured from Cheddar cheeses with different manufacturing and ripening condi-
tions, and a reference sample according to the classical formula ΔE=½ (ΔL 2+Δa2+Δb2).
264 Food Aroma Evolution

FIGURE 13.3 Microbiological reactions occurring during the ripening of the cheese
matrix. (Modified from Marilley and Casey, 2004.)

Where ΔL is the color difference between the product and the reference according to
luminance L (white to black), Δa according to the a-axis (red to green), and Δb according
to the b-axis (yellow to blue). The thermal scales applied during the production of pro-
cessed cheese correspond to those of batch systems: 80°C for a few minutes. This study
makes it possible to relate several factors, such as ripening time, lactose content, galactose
content, salt to relative humidity ratio, and the cooling rate of the processed cheese, with
the intensity of the observed non-enzymatic browning. Galactose causes a browning
more intense than glucose and much moreso than lactose. It is also observed that a short-
ened cooling time limits the phenomenon of non-enzymatic browning. The effect of
Cheddar cheese ripening results in a much lower browning if the starters used are able to
degrade galactose. Therefore, this example highlights the possibility of limiting and con-
trolling the Maillard reaction occurring during the production of processed cheese by
controlling the process parameters and the ingredients used in the formulation. The
amino acid composition of cow milk can be found in the Fox and McSweeney (1998) or
Farrell et al. (2004) studies. The technological processes used make it possible to obtain
a range of milk powders with a broad range of compositions and functional properties.
An extensive description of the technological processes used to obtain these different
milk powders can be found in the Dairy Processing Handbook (Bylund, 1995). Given the
differences in the composition of α-lactalbumin and β-lactoglobulin, a careful selection
of the mixture of milk powders can modulate the ratio between the different amino acids.
In addition to this, the production of powders from whey generally requires significant
mechanical and thermal treatments that lead to possible denaturation of the three-dimen-
sional structure of serum proteins. Amino groups from asparagine, glutamine, lysine,
and arginine will be thus more easily accessible and more responsive to the Maillard reac-
tion. If this reaction is to be minimized, it will be necessary to select “low heat” or even
 Food Processing, Cooking, and Aging: A Case Study 265

FIGURE 13.4 Leucine catabolism. (Modified from Marilley and Casey, 2004.)

“extra low heat” powders. The intensity of the heat treatment undergone to obtain the
powder is equivalent to 70°C for 15 seconds or less (Bylund, 1995). Sithole et al. (2005)
studied non-enzymatic browning occurring during the storage of whey powders obtained
from three different producers. The very different browning rates obtained emphasize the
major importance of the process. However, results obtained by sensory analysis suggest
that the lifetime of these powders is limited by the degradation of their functional proper-
ties rather than by taste, and odors that are not significantly altered during 19 months of
storage at room temperature (Thomas et al., 2004). The fat composition of cow milk is
presented in work by Jensen (2002). Fatty acids are 97% esterified to triglycerides, and to
diglycerides and monoglycerides. The free fatty acid content in milk is less than 0.5%.
Only unsaturated fatty acids are sensitive to oxidation. Linoleic and linolenic acids are
the most sensitive due to their high degree of unsaturation (Jensen, 2002). Phospholipids
also contain polyunsaturated fatty acids. In addition, due to their good emulsifying prop-
erties, they are found at the interface of the emulsion, in a position vulnerable to oxida-
tion. The milk fat used may be butter or anhydrous milk fat. Anhydrous milk fat is
obtained by melting and centrifuging the butter and leads to a product containing more
than 99% fat. To date, butter has more than 230 identified volatile compounds. However,
only a small number of them are considered key odorant molecules for the typical flavor
of butter. These are essentially 2,3-butanedione with a characteristic buttery smell,
butyric acid with a rancid smell, and δ-decalactone with a peachy smell (Mallia et al.,
2008). Emulsifying agents have a major technological role in the production of processed
cheese. They are generally composed of a mixture of citrates, orthophosphates, pyro-
phosphates, and polyphosphates (Caric, 2000). Their role is to supplement the emulsify-
ing capacities of proteins and work schematically as follows: (i) sequestering calcium
bound to caseins, (ii) improving the solubility and dispersion of proteins, (iii) regulating
the pH of the medium, and (iv) improving the hydration of caseins. Polyphosphates also
have the advantage of bacteriostatic properties. Figure 13.5 shows the principle of
266 Food Aroma Evolution

FIGURE 13.5 Chelation of calcium by polyphosphates during cheese making. (Adapted

from Caric, 2000.)

operation of these emulsifying agents. The calcium contained in the calcium–paracasein-

ate complex is removed by the ion exchange properties of phosphates. This increases the
solubility of caseins in water and therefore their emulsifying properties. When the higher
temperatures are reached, bonds between peptides are broken, and the anions of emulsi-
fying agents can bind to proteins. This makes the proteins more hydrophilic, and proteins
are able to bind water molecules. This results in an increase in the viscosity of the mix-
ture, known as the creaming phenomenon (Panouille et al., 2005).

13.3 PROCESS MAKING FOR PROCESSED CHEESE

The production of processed cheese involves many unit operations, as represented in

Figure 13.6.
All these steps can be reduced to five essential unit operations: (i) the mixing of
raw materials, (ii) cooking and temperature control (ensuring the necessary sterilizing
force for the right microbiological preservation of the product), (iii) creaming (to obtain
the desired texture), (iv) cooling, and (v) packaging steps. Thermal operations are par-
ticularly important. Indeed, it is during these stages that major physicochemical modi-
fications regarding oxidation reactions and non-enzymatic browning occur. They are
carried out within a framework of discontinuous or continuous processes. The batch
manufacturing process generally uses a “cutter” type device. Such a system is represented
schematically in Figure 13.7. One of the first patents for such a device was registered in
the United States by Kraft in 1916. It is a large tank of variable volume, sealed tightly by
a cover and equipped with a knife system allowing the mixing of different raw materials.
This type of system ensures all unit operations in the same device. Improvements to the
current cutters (direct or indirect steam injection, partial vacuum) allow temperatures to
quickly increase to around 120 to 130°C. The exact thermal settings to be applied can be
calculated using the parameters of Bigelow or Weibull, once the nature and quantity of
the microorganisms have been determined (van Boekel, 2002; Vanasselt and Zwietering,
2006). For example, Bacillus cereus spores can sometimes be naturally present in an
 Food Processing, Cooking, and Aging: A Case Study 267

FIGURE 13.6 Diagram of the production of processed cheese. (Adapted from Caric, 2000.)

initial processed cheese matrix (Caric, 2000). The duration of these thermal treatments
should be kept as short as possible in order to not generate sensory defects related to
the Maillard reaction occurring at these temperatures and pH conditions. In this case,
however, the product has not reached the texture desired at the end of the cooking step.
A second heat treatment step at moderate temperatures ranging from 80 to 90°C for
about 5 to 30 minutes is then carried out under partial agitation. During this stage, the
combined action of the melting salts sequestering calcium and temperature will allow the
desired texture to be obtained thanks to the creaming phenomenon. The product is then
ready to be packed in different formats and cooled. All of the previously presented unit
operations are adapted for industrial production. A number of patents have been filed to
guarantee each step of the industrial operations. The main processing steps remain very
similar to those of batch production. The raw materials are first prepared and melted.
Storage tanks may be necessary in order to ensure the continuity of production. These
tanks must be maintained at temperatures above 80°C to avoid any gel formation on the
processed cheese. The residence time inside these tanks is variable but can last 1 or 2
hours. In a process proposed by Eyles et al. (1996), the thermal treatment is carried out at
an ultra-high temperature (UHT) using direct steam injection and flash cooling. In this
case, the proportion of vapor incorporated into the cheese matrix is then released dur-
ing the cooling step. The product is then subjected to shear stress to adjust its viscosity.
It is finally packaged into single portions (Weber, 2000; Weber and Didiot, 2009), and
overpackaged into boxes. Depending on the case, it can be cooled quickly at the portion
stage by passing through a cooling tunnel or being cooled more slowly in a cold room.
Figure 13.8 presents the effects of time and temperature on the kinetics of the destruction
of microorganisms and physicochemical changes in the milk. To ensure both a sufficient
sterilizing force (for microbiological safety) and minimal physicochemical damage to
268 Food Aroma Evolution

FIGURE 13.7 Schematic representation of a cutter.

the sensory and nutritional properties of the product, UHT treatment (and direct steam
injection particularly) has proven to be the most efficient of all the thermal processes
compared in Figure 13.8. The thermal performances obtained for three milk sterilization
processes are compared in Figure 13.9. Treatment by direct steam injection ensures the
shortest residence time at a high temperature compared to the other techniques. Storage is
carried out under refrigerated conditions in Europe, North America, and Japan, or under
nonrefrigerated conditions in the case of emerging countries. It is believed that the same
reaction pathways occur; lipid oxidation and the Maillard reaction but at lower reaction
rates than during thermal processing. These reactions are associated with a number of
physicochemical reactions (Schär and Bosset, 2002).

13.4 REACTIONS OCCURRING DURING THE

COOKING OF PROCESSED CHEESE

During the manufacturing and storage stages, processed cheese undergoes chemical, bio-
chemical, and physicochemical modifications. Schär and Bosset (2002), propose eight
 Food Processing, Cooking, and Aging: A Case Study 269

FIGURE 13.8 Destruction of milk microorganisms and temperature-induced changes.

(Adapted from Bylund, 1995 and Fox and McSweeney, 1998.)

categories of physicochemical alterations that may occur during the manufacture and
storage of processed cheese: light-induced reactions, non-enzymatic browning, interac-
tions with the packaging, polyphosphate hydrolysis during storage, changes in the ionic
balance, formation of crystals, water loss leading to variations in water activity, and reac-
tions induced by heat-resistant enzymes. Some of these reactions lead to texture changes
during the storage of processed cheese, while others have a more significant role in the
evolution of flavor during production or storage. From a reactive point of view, the latter
corresponds to the degradation of lipids, Maillard, and caramelization reactions. The

FIGURE 13.9 Typical thermal scales for direct and indirect steam injection and steriliza-
tion in milk bottles used to obtain the same sterilizing force (F), F=40. (Adapted from
Bylund, 1995 and Richardson, 2001.)
270 Food Aroma Evolution

loss of water vapor, in addition to texture changes, can also influence reaction kinetics.
In the case of well-managed processed cheese production, with microbiologically stable
portions, reactions of microbiological origin do not occur. Therefore, we will not provide
much detail on the reactions occurring during ripening.

13.4.1 Lipid Degradation

Lipid degradation generally occurs in two different ways: (i) The hydrolytic pathway
corresponds to the hydrolysis of triglycerides to free fatty acids. The reaction is generally
favored in an acidic or basic environment, with increased heat and humidity. It can be
induced in the manufacturing process, by vigorous agitation, sudden heating, or cooling
as well as by the homogenization processes which can be responsible for damage to the
milk fat globule membrane. Free fatty acids are more sensitive to oxidations than their
corresponding triglycerides. (ii) The oxidative pathway of fat can be divided into three
steps; initiation, propagation, and termination. Figure 13.10 summarizes all the reaction
mechanisms leading to lipid oxidation.
Initiation occurs in three different ways and results in the formation of free radicals.
They can be induced by auto-oxidation, light-induced oxidation, or enzymatic oxidation.
During auto-oxidation, unsaturated lipids LH lose one hydrogen atom to form a lipid
radical L° in the presence of an initiator in Reaction 13.1. The initiation rate increases
with the number of insaturations present in the lipids.

LH + In → L° + InH (13.1)

Light-induced oxidation involves the formation of hydroperoxides in a direct reaction

with an oxygen singleton 1O2 without any radical lipid formation (Reaction 13.2). 1O2
is formed by the activation of oxygen on a sensitizer such as riboflavin (vitamin B2) or

FIGURE 13.10 Mechanisms of lipid oxidation. (Adapted from Eskin and Przybylski, 2001.)
LH: unsaturated lipid; L°: Lipid radical; LOO°: peroxide radical; LOOH: hydroperoxide.
 Food Processing, Cooking, and Aging: A Case Study 271

chlorophyll (Mortensen et al., 2004). In milk, riboflavin is present at levels of 1 to 3 mg

per liter (Causeret and Cirot, 1959). Light and ultraviolet rays can be involved in the
initiation step by activating the riboflavin. A superoxide radical 1O2 is then formed. This
radical is 1450 times more reactive than 3O2 . It will react with an unsaturated lipid to
form hydroperoxides. The hydroperoxides produced in this pathway form cyclic com-
pounds more easily than those induced in the auto-oxidation pathway.

3
sensitizer + 3O2 ® 1sensitizer + 1O2 °
(13.2)
1
O2 ° + LH ® LOOH

Enzymatic oxidation is very important in the case of ripened cheeses. Oxidative enzymes
act most frequently on free unsaturated fatty acids. They catalyze the formation of inter-
mediate hydroperoxides similar to those formed by the non-enzymatic route. However,
the isomers produced are region- and stereospecific. Iron is one of the components of the
lipoxygenases. Propagation takes place in two steps corresponding to the formation of a
hydroperoxide (Reaction 13.3) and its degradation into free radicals (Reaction 13.4). The
lipid radical L° reacts with oxygen to form a free peroxide radical LOO°. This reaction is
favored in the presence of light. The free peroxide radical reacts with another unsaturated
lipid L’H to form a hydroperoxide LOOH and a new lipid radical L’° which will continue
the propagation:

L° + O2 → LOO°
(13.3)
LOO° + L’H → LOOH + L’°

This unstable hydroperoxide LOOH will then degrade into new free radicals:

LOOH → LOO° + H°

LOOH → LO° + OH° (13.4)

2LOOH → LO° + H 2O + LOO°

The free peroxide radicals react with each other to form non-radical products (Reaction
13.5). This termination reaction is favored in an environment where oxygen partial pres-
sure is low. The termination reaction can also take place through the action of antioxi-
dants AH capable of generating free radicals that are stabilized by resonance and which
will not participate in the reaction.

LOO° + L’OO° → LOOL’ + O2

(13.5)
L° + AH → LH + A°
The hydroperoxides formed during the lipid oxidation reaction have no odor nor flavor.
On the other hand, the compounds resulting from their degradation, presented
in Figure 13.10, such as aliphatic aldehydes, are responsible for the flavors resulting
from oxidation. Many different volatile compounds are obtained, such as ketones,
methylated-ketones, aliphatic aldehydes, alcohols, furan compounds, and lactones.
Pentanal and hexanal are two of the most formed products. However, these are generic
compounds that do not specifically originate from a single substrate. Figure 13.11
schematically represents the degradation kinetics of fatty acids and the appearance of
272 Food Aroma Evolution

FIGURE 13.11 Schematic representation of the lipid oxidation kinetics. (Adapted from
Labuza and Dugan, 1971 and Kamal-Eldin et al., 2003.)

lipid oxidation resulting compounds. Many factors increase the lipid oxidation rate.
One such is a higher content in free fatty acids, as they are more easily accessible than
the ones esterified by glycerol. Similarly, long-chain and polyunsaturated fatty acids
are more sensitive to oxidation. Transition metals such as iron, copper, or aluminum
can be accidentally or temporarily introduced by contact with the processing mate-
rial and promote oxidation. On the other hand, phenols and citric acid are known
as sequestering agents of metals and can be used to reduce this undesirable oxida-
tion. Concerning the processing parameters, a higher temperature or a higher oxygen
partial pressure both increases the rate of the oxidation and reduces the induction
period. Finally, it has been established that the oxidation is minimal for water activi-
ties between 0.3 and 0.5.

13.4.2 The Maillard Reaction

The nucleophilic addition of a free amine function with a reducing sugar was discovered
in 1912 by the French chemist and doctor Louis-Camille Maillard. This reaction is part
of the non-enzymatic browning reactions with caramelization due to the formation of
polymers called melanoïdins that cause a characteristic brown color at the advanced
stages of this reaction (Maillard, 1913). This is a very important reaction for the food
industry as it explains a large part of the sensory properties, aroma, and taste of cooked
products. However, it was not before 1953 that Hodge proposed a complete reaction
scheme, which is still in use today. A full description of the reaction can be found in
Bertrand et al. (2018) or in Chapter 14 of this book.

13.4.3 Caramelization Reactions

The caramelization reaction is described by Kroh (1994) as the succession of six reac-
tions leading to the degradation of reducing sugars without the intervention of nitrogen
 Food Processing, Cooking, and Aging: A Case Study 273

FIGURE 13.12 Sugar degradation during caramelization. (Adapted after de Bruijn, 1986.)

compounds. The absence of nitrogen compounds is the main difference between cara-
melization and the Maillard reaction. It is characterized by the formation of a brown
color and aroma compounds typical of caramel. Figure 13.12 shows these reactions.
The first step consists of a reaction of 1,2-enolization of the reducing sugar called Lobry
de Bruyn–Alberda van Ekenstein’s rearrangement, according to the names of the two
researchers who highlighted it in 1885. This reversible step is followed by a second non-
reversible enolization and then the β-elimination of a water molecule. The fourth step is
a dicarbonyl cleavage, which will lead to the formation of carboxylic acids (formic and
acetic acid). The last two steps are retro-aldolization and condensation-aldolization. This
leads to the formation of osuloses, which are highly reactive dicarbonyl compounds, and
to the release of hydrogen ions, causing a decrease in pH over time up to values in the
range of 4 to 5. Osuloses then lead to the formation of compounds that are typical for the
color and flavor of caramel: three oxygenated heterocyclic compounds, 5-hydroxymeth-
ylfurfural, hydroxydimethylfuranone, and hydroxyacetylfurane are formed during the
caramelization of glucose (Coca et al., 2004). The distinction between the caramelization
reaction and the Maillard reaction is extremely delicate when considering the presence
of amino groups in the reaction medium. However, the formation of acid, as shown in
Figure 13.12, seems to be a relevant indicator of the caramelization pathway in such a
situation. The reaction favors high temperatures, generally above 120 or even 150°C,
depending on the sugars involved, and extreme pH conditions, that is, strongly basic (pH
9) or acidic (pH 3) (Kroh, 1994). In a model reaction medium consisting of an equimo-
lar mixture of glucose and glycine buffered at pH 6.8, Martins and van Boekel (2005)
showed that temperatures of treatment above 120°C favor caramelization pathways in
relation to the Maillard reaction path. These thermal scales are higher than those nor-
mally encountered during the manufacture of processed cheeses. In addition, the pH of
processed cheese, between 4 and 7, is not favorable to the caramelization reaction during
the manufacturing process.
274 Food Aroma Evolution

13.5 IDENTIFICATION OF FLAVOR

MODIFICATIONS DURING PROCESSING

Bertrand et al. (2011) studied the evolution of the composition of odorous molecules dur-
ing the cooking of a model processed cheese matrix: 346 volatile compounds were identi-
fied. Many were not specific (branched alkanes, alkenes, terpenes) to particular reactions.
Only 81 structures were significantly influenced by the thermal treatments applied to the
model cheese matrix (p<0.01). These structures were mainly derived from lipid degrada-
tion, involving hydrolysis and oxidation reactions, amino acid degradation, involving the
Maillard reaction, and carbohydrate degradation. Principal component analysis (PCA)
was performed on 27 of the 81 compounds significantly influenced (p<0.001) by heat
treatment and recognized in the literature as markers of fatty acid oxidation, Maillard, or
caramelization reactions. It provided a multivariate view of the evolution of the composi-
tion of the volatile fraction of the matrices during the heat treatments applied (five tem-
perature levels, five treatment times, and three repetitions); this is shown in Figure 13.13.
The study of the main plane of the PCA shows that the temperature rise of the matrix
up to 135°C induces a significant shift in the composition correlated to the variables
present in the quadrant of coordinate [−1, −1] on the circle of correlations. This shift cor-
responds to the formation of a sugar fragmentation product: diacetyl, several Strecker
aldehydes (variables 4 to 7) except methional (variable 8) and several sulfur compounds
(sulfides: variables 9, 10, and 13, thiazole and thiophene: variables 11 and 12). When the
temperature rises to 150°C, there is a break in the evolution of the composition signifi-
cantly correlated to variables 14 and 15 and to all those present in the [−1, 1] quadrant.
This corresponds to the formation of markers for caramelization and Maillard reactions
(furfural, furfuryl alcohol, 5-methylfurfural, maltol, isomaltol, and 5-pyrazines (vari-
ables 14 to 23)). The duration of the heat treatment induces different effects depending on
the temperature. Thus, for final temperatures of 80 and 100°C, whatever the duration, a
limited evolution in the composition can be observed, corresponding to an increase in the
concentrations of diacetyl, Strecker aldehydes, and sulfides. For the final temperatures of
120, 135, and 150°C, the change in composition reflects the formation of the advanced
products of caramelization and Maillard reactions (variables 14 to 23). However, only a
small portion of these compounds are responsible for the characteristic smell of processed
cheese. In order to identify odor-active molecules, as well as those responsible for the
appearance of overcooked defects, Bertrand et al. (2011) used gas chromatography tech-
niques coupled with olfactometry. The products analyzed corresponded to a model raw
cheese matrix and a matrix that had undergone a heat treatment of 1 minute at 150°C.
About 30 odorous zones represented in Figure 13.14 are highlighted. The list of identified
odorant molecules and the predominantly perceived odor is given in Table 13.1.
Under these conditions, the significant differences observed were attributed solely to
the chemical reactions induced by the heat treatment. Of the 16 main odour zones of the
raw cheese matrix (zones: 2, 6, 7, 9, 10, 10, 11, 12, 12, 13, 14, 16, 17, 22, 23, 24, 27, and
29 in Figure 13.13), only 10 had an intensity greater than 1 (on a scale of 5). After cooking,
the aromagram of the cheese matrix was significantly modified. A general increase in the
odor intensity of the different olfactory categories was observed, since 19 odorant mol-
ecules then had an intensity greater than 1; on a qualitative level, cooking greatly ampli-
fied the categories “buttered–cheese–dairy,” “empyreumatic,” and to a lesser extent the
categories “fruity–floral” and “green–vegetable.” These modifications resulted from the
increase in the odor intensity of molecules already present in the raw matrix (zones 2, 9,
11, 17), of which hexanal and octanal (zones 9 and 17 respectively) are the main molecules
 Food Processing, Cooking, and Aging: A Case Study 275

FIGURE 13.13 Principal component analysis plot PC1 versus PC2 of all variables (A) and
samples (B, 80°C; 100°C; 120°C; 135°C; and 150°C). PC1 and PC2 accounted for 60%
and 13.4% of the total variance respectively; “r” stands for the raw matrix (extracted
1 hour at 60°C) and “X” stands for the matrix cooked for 1 hour at 60°C (extracted 1
additional hour at 60°C). Numbers represent the time elapsed at the desired temperature
in minutes. The full line represents the overall time course of the volatile fraction during
the heating period. The dotted line represents its time course during the holding period.
Numerals stand for compounds as follows. 1: 3-hydroxy-2-butanone; 2: 2-cyclopentene-
1,4-dione; 3: 2,3-butanedione; 4: 2-methylpropanal; 5: benzaldehyde; 6: 2-methylbutanal;
7: 3-methylbutanal; 8: methional; 9: dimethylsulfide; 10: dimethyldisulfide; 11: thiazole;
12: thiophene; 13: dimethyl trisulfide; 14: furfural; 15: furfuryl alcohol; 16: pyrazine; 17:
2-methylpyrazine; 18: 2,5-dimethylpyrazine; 19: ethenylpyrazine; 20: 2-ethylpyrazine; 21:
5-methylfurfural; 22: isomaltol; 23: maltol; 24: 2-ethylfuran; 25: hexanal; 26: octanal;
27: nonanal. (Reproduced from Bertrand et al., 2011 with permission.)

responsible for the “fruity–floral” notes perceived by the sniffs. They also resulted from
the appearance of 13 new odorous compounds (zones 1, 3, 4, 5, 5, 8, 15, 18, 19, 20, 21,
25, 26, 28). Of these, four were mainly responsible for the increase in “buttered–cheese–
dairy” odors. These are 2-methylpropanal (zone 1), 2,3-butanedione (zone 2), 2,3-pen-
tanedione (zone 5), and butanoic acid (zone 8). Five molecules were responsible for the
“empyreumatic” note, three of which contributed mainly to the increase in this note after
firing. These were 2-acetylpyrazine (zone 18), furanéol (zone 20), and maltol (zone 21).
The other two molecules, a pyrazine and an unidentified molecule (zones 10 and 13), were
perceived less intensely after firing. The higher intensity of the “green–vegetable” notes
was due to the production after cooking of methional (potato smell, zone 12), dimethyltri-
sulfide (gas, cabbage, zone 15), and an unidentified molecule (zone 28). Other sulfur com-
pounds such as methanethiol, dimethylsulfide, and dimethyldisulfide were not perceived,
276 Food Aroma Evolution

FIGURE 13.14 Model cheese aromagrams before (top; raw sample) and after (bottom;
heated sample) thermal treatment of 1 minute at 150°C. The total olfactory signal by class
is expressed as a mean product of intensity and number of detections of the 8 individual
aromagrams. The breakdown into 10 classes shows the aromagram zones belonging to
the given olfactory classes. Numerals refer to Table 13.1. (Reproduced from Bertrand
et al., 2011, with permission.)

TABLE 13.1 List of Identified Odor-Active Compounds and a Short Description of Their
Perceived Odor by the Panel

Source: Reproduced from Bertrand et al., 2011, with permission.

 Food Processing, Cooking, and Aging: A Case Study 277

probably due to higher detection thresholds than those of the previously mentioned sulfur
compounds. The direct illumination of the cheese matrices indicates that when applying
extreme thermal scales, empyreumatic notes take precedence over buttered notes, fruity
and vegetal. This suggests that during cooking, products of the Maillard reaction affect
the smell of cooked cheese more than the compounds resulting from the oxidation of fatty
acids. Furanéol and maltol can be considered as the main culprits for the “overcooked”
defects found in processed cheeses. Indeed, when firings above 135°C for 5 minutes, the
smell of these compounds begins to be noticeable when the cooked products are lit and
gradually masks the initial smell of the product.

13.6 CONCLUSION

Through this processed-cheese example, we have shown that the formation of flavor
compounds during the manufacturing process is certainly influenced by technological
choices but also and very significantly by the fine-tuned properties of the selected raw
materials (content of certain simple sugars such as galactose, the degree of ripening). This
observation calls for an integrated approach to the entire supply chain from the raw mate-
rials to the consumer not only for safety reasons but also to provide the desired aromatic
profile the consumer is entitled to expect.

REFERENCES

Bertrand, E., El Boustany, P., Faulds, C. B., and Berdagué, J. L., 2018. The Maillard
reaction in food: An introduction. In Reference Module in Food Science, p. 1–10.
Elsevier.
Bertrand, E., Machado-Maturana, E., Chevarin, C., Portanguen, S., Mercier, F.,
Abouelkaram, S., Tournayre, P., Guillard, A. S., Kondjoyan, A., and Berdagué, J.
L., 2011. Heat-induced volatiles and odor-active compounds in a model processed
cheese. International Dairy Journal, 21:806–14.
Bley, M., Johnson, M. E., and Olson, N. F., 1985a. Factors affecting non-enzymatic
browning of process cheese. Journal of Dairy Science, 68:555–61.
Bley, M., Johnson, M. E., and Olson, N. F., 1985b. Predictive test for the tendency of
cheddar cheese to brown after processing. Journal of Dairy Science, 68:2517–20.
Brickley, C. A., Auty, M. A. E., Piraino, P., and McSweeney, P. L. H., 2007. The effect of
natural cheddar cheese ripening on the functional and textural properties of the pro-
cessed cheese manufactured therefrom. Journal of Food Science, 72(9):C483–90.
Bylund, G., 1995. Dairy Processing Handbook. Tetra Pak Processing Systems, Lund, Sweden.
Caric, M., 2000. Processed cheese. In Francis, F. J., editor: Encyclopedia of Food Science
and Technology, second edition, p. 1087–1973. John Wiley & Sons, New York, NY.
Causeret, J. and Cirot, E., 1959. Ebullition domestique et teneur en riboflavine du lait de
vache. Le Lait, 39(383–4):159–65.
Coca, M., Garcia, M. T., Alink, G., Pena, M., and Garcia, J. A., 2004. Study of colored
components formed in sugar beet processing. Food Chemistry, 86:421–33.
Commission du Codex Alimentarius, 1995. Norme générale codex Alimentarius pour les
additifs alimentaires. Codex Alimentarius, Codex Stan, 192:22–3.
de Bruijn, J. M., 1986. Monosaccharides in Alkaline Medium: Isomerization,
Degradation, Oligomerization. PhD Thesis, Delft University, the Netherlands.
278 Food Aroma Evolution

Eskin, N. A. M. and Przybylski, R., 2001. Antioxidant and shelf life of foods. In Eskin, N.
A. M. and Robinson, D. S., editors: Food Shelf Life Stability, Chemical, Biochemical
and Microbiological Changes. CRC Press LLC, Boca Raton, FL.
Eyles, C. T., Anbarci, A., Nassauer, J. S., and Simburger, S., 1996. Process for continuous
production of processed cheese. WO patent no. 96/01567.
Farrell, H. M., Jimenez-Flores, R., Bleck, G. T., Brown, E. M., Butler, J. E., Creamer,
L. K., Arthur, C. L., Hollar, C. M., Ng-Kwai-Hang, K. F., and Swaisgood, H. E.,
2004. Nomenclature of the proteins of cows’ milk—Sixth revision. Journal of Dairy
Science, 87(6):1641–74.
Fox, P. J. and McSweeney, P. L. H., 1998. Dairy Chemistry and Biochemistry. Blackie
Academic and Professional, London, UK.
Gibbons, A., 2007. Food for thought: Did the first cooked meals help fuel the dramatic
evolutionary expansion of the human brain? Science, 316:1558–60.
Hodge, J. E., 1953. Chemistry of browning reactions in model systems. Journal of
Agricultural and Food Chemistry, 1:928–43.
Jensen, R. G., 2002. The composition of bovine milk lipids: January 1995 to December
2000. Journal of Dairy Science, 85(2):295–350.
Kamal-Eldin, A., Mäkinen, M., and Lampi, A. M., 2003. The challenging contribution
of hydroperoxides to the lipid oxidation mechanism. In Kamal-Eldin, A., editor:
Lipid Oxidation Pathways, p. 6–41. AOCS Press, Champaign, IL.
Kroh, L. W., 1994. Caramelisation in food and beverages. Food Chemistry, 51(4):373–9.
Labuza, T. P. and Dugan, L. R., 1971. Kinetics of lipid oxidation in foods. Critical
Reviews in Food Technology, 2(3):355–405.
Maillard, L. C., 1913. Genèse des matières protéiques et des matières humiques. Action
de la glycérine et des sucres sur les acides alpha-aminés. Masson et Cie, Paris,
France.
Mallia, S., Escher F., and Schlichtherle-Cerny, H., 2008. Aroma-active compounds of
butter: A review. European Food Research and Technology, 226(3):315–25.
Marilley, L. and Casey, M. G., 2004. Flavors of cheese products: Metabolic pathways,
analytical tools and identification of producing strains. International Journal of
Food Microbiology, 90(2):139–59.
Martins, S. I. F. S. and van Boekel, M. A. J. S., 2005. Kinetics of the glucose/glycine
Maillard reaction pathways: Influences of pH and reactant initial concentrations.
Food Chemistry, 92(3):437–48.
McSweeney, P. L. H., 2004. Biochemistry of cheese ripening. International Journal of
Dairy Technology, 57(2–3):127–44.
Monteiro, C. A., 2009. Nutrition and health. The issue is not food, nor nutrients, so
much as processing. Public Health and Nutrition, 12:729.
Mortensen, G., Bertelsen, G., Mortensen, B. K., and Stapelfeldt, H., 2004. Light-induced
changes in packaged cheeses—A review. International Dairy Journal, 14(2):85–102.
Panouille, M., Durand, D., Nicolai, T., Larquet, E., and Boisset, N., 2005. Aggregation
and gelation of micellar casein particles. Journal of Colloid and Interface Science,
287(1):85–93.
Richardson, P., editor, 2001. Thermal Technologies in Food Processing. CRC Press, Boca
Raton, FL.
Schär, W. and Bosset, J. O., 2002. Chemical and physicochemical changes in processed
cheese and ready-made fondue during storage: A review. LWT—Food Science and
Technology, 35(1):15–20.
 Food Processing, Cooking, and Aging: A Case Study 279

Sithole, R., Mcdaniel, M. R., and Goddik, L. M., 2005. Rate of Maillard browning in
sweet whey powder. Journal of Dairy Science, 88(5):1636–45.
Thomas, M. E. C., Shär, J., Desobry-Banon, S., and Desobry, S., 2004. Milk powders
aging effect on physical and functional properties. Critical Reviews in Food Science
and Nutrition, 44:297–322.
Vanasselt, E. and Zwietering, M., 2006. A systematic approach to determine global ther-
mal inactivation parameters for various food pathogens. International Journal of
Food Microbiology, 107(1):73–82.
van Boekel, M. A. J. S., 2002. On the use of the Weibull model to describe thermal inac-
tivation of microbial vegetative cells. International Journal of Food Microbiology,
74(1–2):139–59.
Weber, J. C., 2000. Atmospheric package for pasty products. WO 00/17064.
Weber, J. C. and Didiot, L., 2009. Method for packaging a pasty product in an easily
opened, sealed packaging made of plastic material. WO 2009/092966 a1.
Yang, Y., Shevchenko, A., Knaust, A., Abuduresule, I., Li, W., Hu, X., Wang, C., and
Shevchenko, A., 2014. Proteomics evidence for kefir dairy in Early Bronze Age
China. Journal of Archaeological Sciences, 45:178–86.
 Chapter 14
The Maillard Reaction
Joseph Provost

CONTENTS

14.1 Introduction 281

14.2 Maillard Chemistry 282
14.2.1 Initial Stage 282
14.2.2 Intermediate Stage 285
14.2.3 Late Reactions 286
14.3 Pool Theory 287
14.4 Advanced Glycation End Products 288
References 290

14.1 INTRODUCTION

Non-enzymatic catalyzed browning reactions in food and drink are classified into several
distinct reaction types including lipid oxidation, caramelization, ascorbic acid oxidation,
and the Maillard reaction. The last set of reactions, Maillard, are non-enzyme catalyzed
reactions whose product is formed as the result of heat generated during the processing,
preparation, and storage of meat, vegetables, fruit, and liquids. These reactions produce
both desirable and unwanted results for food preparation. Tea, beer, bread, and syrup
are all enhanced by the rich flavor profiles of the Maillard reaction while the organoleptic
properties reduce the qualities and appearance of other flavorants and volatiles in foods
such as milk. Thus, a complete understanding of the Maillard reaction and its implica-
tions in food preparation is very important (Hellwig and Henle, 2014; Bertrand et al.,
2018). The Maillard reaction, a condensation of a reducing sugar with amino groups
of amino acids, peptides, or protein, is a complex reaction with multiple phases and
possible pathways providing a multitude of possible colored and flavored compounds
(Figure 14.1). The initial reaction was first observed by Louis-Camille Maillard as he
investigated peptide synthesis while heating amino acids in the presence of glycerol and
then glucose (Hellwig and Henle, 2014). The key product was a brown precipitate with
the concomitant liberation of carbon dioxide. This work led to further investigations
into understanding the condensation reactions between amino acids and the carbonyls of
reducing sugars (Maillard, 1911, 1916). The first mechanism describing Maillard brown-
ing was proposed by Hodge and Rist (1953) with additional modifications suggested by
others (Eskin and Shahidi, 2013). While there continues to be an active focus on biomedi-
cal applications, where the Maillard reaction is known as glycation producing advanced
glycation end products (AGE) with a number of associated pathologies, much of the work
in the food industry is concerned with the impact of the reaction on Maillard production

281
282 Food Aroma Evolution

FIGURE 14.1 The Maillard reaction.

of both advantageous and toxic products including acrylamide. We will highlight this
complex reaction and describe where AGE products are formed, the addition of caramel-
ization intermediates as substrates of the reaction, the role lipids and other biomolecules
have on the reaction, as well as the factors including pH and water activity that affect the
final products.

14.2 MAILLARD CHEMISTRY

14.2.1 Initial Stage

As initially described by Hodges, the Malliard reaction is a complex set of pathways

consisting of three phases. The initial stage is a condensation between the amino group
and the carbonyl of an oxidized reducing sugar coupled with an Amadori or Heyns rear-
rangement. This stage is followed by an advanced or intermediate phase where sugar
products fragment or dehydrate and amino acids degrade (the Strecker degradation). The
reaction is completed with the final stage of aldol condensation and the formation of
brown melanoid polymers and co-polymers. While the entire set of Maillard reactions
have been described as a “non-classical” chemical reaction format because of the pool of
complex reactions taking place after the initial formation of Heyns and Amadori prod-
ucts (Yaylayan, 1997), they are not linear and vary greatly depending on the conditions
and competing reactants; the initial step remains a simple condensation, dehydration,
 The Maillard Reaction 283

and rearrangement reaction pathway. This initial reaction begins as a nucleophilic substi-
tution by the nitrogen of an amine onto the partially positive carbonyl carbon of an oxi-
dized reducing sugar. This reaction is followed by a reaction which forms an intermediate
amine which will in turn, cyclize to an unstable Schiff base. This will then transform
into an N-substituted glycosylamine. The source of electron-rich nitrogen is commonly
described as the amino acid side group for lysine (ε-amino) and the guanidinium group
of arginine. All free amino acids have the potential to be a Maillard reaction via the free
amino group. Heating ribose with amino acids without nitrogen-containing side-chains
results in the browning reaction. Interestingly, besides lysine, glycine, tryptophan, and
tyrosine all react strongly with reducing sugars, providing a significant Maillard prod-
uct. Protease protection and the nutritional availability of the internal peptide bonds of
proteins are evidence to suggest that the peptide bond imine could react with a reducing
sugar, giving rise to a Maillard product (Horn et al., 1968). The length of the peptide/
protein may impact the rate of the reaction (Kim and Lee, 2009). However, this isn’t as
straightforward as chain length because dipeptide glycine reacted at a greater rate than
free amino acid alone or the tripeptide (Lu et al., 2005). The likely explanation was that
the peptide bond hydrolysis and greater stability of the tripeptide decreased the rate of
the reaction (Kim and Lee, 2009). However, lysine remains the key amino acid thought
to be involved in browning reactions.
Both ketone and aldehyde reducing sugars can participate in the reaction. Aldoses in
an oxidized state will react with forming aldosylamines after the Schiff rearrangement
ultimately giving rise to a 1-amino-1-deoxy-ketose Amadori compound. On the other
hand, ketose sugars will produce a ketosylamine and rearrange into a 2-amino-2-deoxy-
aldose Heyns compound (Figure 14.2). In general, pentoses react with a faster rate than
hexoses, and monosaccharides are significantly more reactive than disaccharides (Eskin
and Shahidi, 2013). The availability of the oxidized anomeric carbon of a reducing sugar
is critical for the nucleophilic amine to attack. Thus, the reaction is enhanced in alkali
conditions when the equilibria of a closed-ring hemi-acetal to an open chain of reducing
simple carbohydrates shifts into the open form of the reducing sugar. A simple study com-
paring the rate of the browning of foods dipped in water (pH 6.90) vs. a dilute solution of
baking soda (pH 9.0) showed that carrot, potato, onion, and chicken all browned two or
even four times faster in basic conditions than neutral (Provost et al., 2016).
In general, lysine with both a side-chain amine and an α-carbon amino group, is
highly reactive with most sugars. However, glycine shows the highest rate of reaction
when heated with fructose, ribose, or lactose (Ashoor and Zent, 1984). When heated in
a 1:1 ratio at pH 9.0, both arginines produced only 30% of the product of either lysine
or glycine with the sugars tested. Surprisingly, several free amino acids reacted ~20% as
effectively as lysine, presumably through the α-carbon amino group (tryptophan, tyro-
sine, proline, leucine, isoleucine, and alanine). Therefore, a protein hydrolysate or foods
with a high concentration of free amino acids (such as shrimp with significant levels of
glycine supporting osmotic pressures) are able to brown easily.
Likewise, the type of sugar has an important impact on the browning reaction.
Monosaccharides react with greater rates than disaccharides, and five-carbon sugars are
more reactive than six-carbon sugars (Pastoria et al., 2018). Generally, sugars react in
the following order, pentoses > hexoses and disaccharides. However, this depends on the
model system used. One report showed that ribose will react with lysine at the greatest
rate with xylulose while moderately reacting with arabinose. Both glucose and fructose
produced an almost undetectable product with protein hydrolysate (Eskin and Shahidi,
2013), and when tested against each amino acid both ribose and lactose showed nearly
284 Food Aroma Evolution

identical rates of browning with most amino acids (lactose slightly > ribose > fructose
> glucose) (Ashoor and Zent, 1984). A conservative estimate of possible reaction combi-
nations of the first step of the Maillard reaction demonstrates the possibility of diverse
aroma, flavor, and color products. Considering each of the amino acids with a free amino
group plus the side groups of lysine and arginine and limiting the reducing sugars to eight
of the more common simple carbohydrates, there are over 576 Amadori or Heyns poten-
tial products at this point alone. The path each intermediate may take through the rest
of the reaction pathway, as well as possibilities for the intermediates of the condensation
product to react with other substrates, highlights the diversity of this reaction. Following
the condensation reaction and loss of water forming an intermediate imine, there are two
potential paths: a reversible cyclization generating a glycosylamine or the formation of a
Schiff base (Figure 14.2). The reaction generating the Schiff base is reversible, especially
so in acidic conditions, where the initial products are easily regenerated. However, the
N-glycosylamine rearrangement into the Amadori or Heyns compound is irreversible, as
the precursor Schiff base can be converted into a cyclic hemiaminal which easily muta-
rotates, whereas the formation of a furanose (Amadori) or ketose (Heyns) hemiacetal
has a similar mutarotation to carbohydrates where the equilibria lies in the cyclic form.
The Schiff base will form an enaminol (enol form) and, depending on the initial sugar
and pH, will have one of three possibilities. At a higher pH, a loss of amine generates a
deoxydicarboyl compound. At a lower pH, a 1–2 enolization results in the generation of
the keto Amadori product, while oxidation and hydrolytic loss of the amine results in a
glucosone cyclic product. Under heat, each of these intermediates can form into flavor/
aroma products via the next set of advanced Maillard reactions.

FIGURE 14.2 Formation of Amadori and Heyns compounds in an early Maillard reaction.
 The Maillard Reaction 285

14.2.2 Intermediate Stage

Depending on the initial condensation and the various pathways already presented,
there are several different fates for the Amadori, Heyns, and other intermediates formed
thus far, in what is classically called the intermediate stage (Figure 14.1). An example
of aroma and flavor compound generation through deoxyosone dehydration includes
the generation of furanones, isomaltol, and maltol in meats (Eskin and Shahidi, 2013).
Although this is perhaps more accurately described as a pool of possible reactants, inter-
mediates, and products, which we will discuss later, Amadori and Heyns products will
degrade into one of several possible pathways depending on pH and temperature. Further
decomposition and dehydration of the three waters of Amadori products in acidic condi-
tions will lead to the formation of a furan ring containing hydroxymethlfurfural and
furfural. Glycine has been reported to increase the formation of this set of products,
presumably as this increases the reaction from the Amadori product over an alternative
degradation of glucose without utilizing the Maillard reaction (Pastoriza et al., 2018).
Also, under acidic conditions, sugar dehydration results in the production of deoxyosone
osloses 1-, 2-, 3, or 4-deoxysones. These can then cyclize into the maltols, and furanones
described earlier, and can in subsequent reactions with ammonia and hydrogen sulfide
produce a number of meaty and other savory flavors. An additional pathway of both
Heyns and Amadori compounds involves dehydration under more neutral or basic con-
ditions resulting in the loss of two water molecules generating the antioxidant-reduced
enediol. The acid-catalyzed fragmentation and loss of amine generates the dicarbonyl
deoxyosones. Further dehydration creates dehydroreductones. Both forms of reductones
serve as a fork in the degradation pathways, where in basic conditions they undergo a
retro-aldolization into acetone compounds, diacetyle, glyoxal, pyruvaldehyde, glycol-
aldehyde, and glyceraldehyde, each with their own potential as a flavorant/odorant, or
they can further react with food compounds on their own or in reactions with amino
acids (Strecker reaction). The presence of the carbonyl group stabilizes the enediol form
as an a-oxo-enediol with a strong acidic tendency leading to a strongly reducing poten-
tial for the compounds. As such, both the reductones and the dehydroreductones play
a key part in browning in subsequent reactions. A third pathway involves the fission or
fragmentation of Amadori/Heyns degradation compounds (Figure 14.1). The reaction is
initiated by an oxidative fission or reversal of the aldol condensation reaction. Through
rearrangement, reduction, and saccharinic rearrangement (migration of alcohol trans-
forming from a dihydroxyl enol to a carboxyl compound), the cleavage reaction forms
small decarbonylated aldehyde, alcohol, and acetic compounds, including acetic acid,
formaldehyde, diacetyl, glyoxal, acetal and acetaldehyde and 2,3 butanedione (Weenen
and Apeldoorn, 1996; Nursten, 2005). An important pathway that involves the loss of
free amino acids and for the most part not peptides or proteins, as observed in the ini-
tial Maillard condensation reaction, is the oxidative degradation of amino acids in the
presence of α-dicarbonyls formed by the Amadori/Heyns compounds (reductones and
fission products). Deoxyosones, diacetyl, pyruvaldehyde, hydroxyacetone, glyoxyal, and
other decomposition products are potential Strecker reactants. The diversity of potential
degradation dicarbonyl compounds and free amino acids combine to make this reaction
a major contributor to the flavor profile of food and beverages. Because this reaction
proceeds at a greater rate with free amino acids, the pathway is more prevalent in foods
high in free amino acids or protein hydrolysates. The reaction involving cysteines and
ribose is responsible for many of the aroma and flavor of cooked meats. Interestingly,
the reactive α-carbonyl of glyoxal can react with the lesser reactive arginine side chain
286 Food Aroma Evolution

FIGURE 14.3 The Strecker reaction.

(in the absence of competing reactant cysteine) preferred over lysine. While the Strecker
degradation reaction imparts a key effect on the flavor profiles of roasted foods includ-
ing meats and cocoa and coffee beans, both the higher- and lower-molecular weight
products of the reaction play a contributory role. It is the further reactions (with a sec-
ond amino acid or condensation of various intermediates) that produce the compounds
involved in the flavor of heated foods. These aroma and flavor products from the Strecker
degradation (Figure 14.3) are often organized by the products: pyrroles, oxazoles and
oxazolines, and thiazole derivatives. Heterocyclic nitrogen-containing compounds such
as pyrazine have been reported to be important in the flavor of many heated and toasted
foods, from meats, broths, and vegetables, most with a low odor threshold. Pyrroles
were first identified in roasted coffee beans and are a key component of the heterocy-
clic compounds formed during non-enzymatic browning. Ribose and b-hydroxy amino
acids are precursors for these compounds, which are further processed into furans and
fururyl-substituted pyrroles. When heated in the presence of serine, threonine, and vari-
ous monosaccharides, the important flavors of heat-treated cereal and popcorn were
identified. Some of the compounds bring about a caramel-like aroma of cooked meats.
The Strecker degradation production of several oxazoles further provides complex green
and vegetable-like notes in cooked meats. Cysteine and methylglyoxyl Strecker degrada-
tion products are examples of thiazole heterocyclic compounds. The nucleophilic attack
of the sulfur of cysteine at the carbon of an imine intermediate formed by the reaction
between ammonia and an aldehyde creates a range of similar cyclic molecules. These
low odor threshold volatiles give sulfurous, onion-like aromas to meats, potato chips,
and roasted peanuts.

14.2.3 Late Reactions

While many intermediates provide both volatile and soluble components of flavor and
aroma, the reactions generate brown-colored compounds as well as flavors. This final
 The Maillard Reaction 287

step is a polymerization of many of the substances into both high and low-molecular
weight melanoids. Much of these are condensation reactions potentially between all of
the intermediates presented thus far. Polymerization of aminated products including pyr-
rols, furaldehyes, and others create a brown poorly defined anionic compound. If the
backbone of the browning pigment is built from a carbohydrate backbone, such as those
found in dark liquids, the products are known as melanosaccharides. The browning reac-
tions found on crusts of bread and baked goods often have protein at the core and are
then called melanoproteins. The constitution of melanoids, in general, is dependent on
the amount of aldehyde and amines involved in the polymerization. Often the condensa-
tion products are polymers of hetercyclic compounds. Proteins, via linkage with arginine
or lysine side groups, act as a scaffold for carbohydrate-involved melanoidins. Thermal
processing of meats shows high levels of carbohydrate-induced browning in this fashion.
Despite a growing role in health as both an antioxidant and other biological effects, very
little has been learned about the structure of these compounds. Most of our understand-
ing comes from using model systems, taking a reductionist approach mixing various
intermediates, and investigating the final products.

14.3 POOL THEORY

The difficulty in determining the intermediates and the final melanoid polymers stems
from the diversity, condition (pH, water availability, temperature), and concentration of
potential reactants in the processed foods. Thus, a particularly interesting and descriptive
theory first described by Yaylayan in 1997, is that a cascade of reactions does not follow
the paths described (Figure 14.4). Rather the products are propagated by chemical pools
generated from various precursors. The pools result from three defined pools of precursor
parents: sugars, amino acids, and Amadori/Heyns products. In addition to the interac-
tion between pool constituents, each parental compound can fragment as they interact
in the “pool.” Depending on the condition, the initial pool will direct the final product

FIGURE 14.4 Pool theory of the Maillard reaction.

288 Food Aroma Evolution

into polymers, dimers, heterocycles ore “other compounds.” As the reactions propagate,
each pool can easily interact and within or between pools, generate new Maillard reac-
tion products. While making it difficult to predict these products, it is a more applicable
description vs. the pathway model.

14.4 ADVANCED GLYCATION END PRODUCTS

For some time, there have been concerns about the loss of nutritional value of foods or
even potential toxicity due to the formation of Maillard products. The loss of lysine as a
nutritional source in processed foods and a decrease in protein function, solubility, and
digestibility after the reaction with Maillard products are examples of the role of these
reactants in food processing. In addition to changes in the nutritional state of foods due to
the Maillard reaction, there are biological health concerns with some of the side reactions
between Maillard intermediates and proteins or lipids. Advanced glycation end products
(AGEs) are the result of side reactions from the Amadori product and α-dicarbonyl com-
pound reaction with proteins or lipids. In general, these products are formed by the reac-
tions of reducing sugars or the degradation of biological macromolecules (carbohydrates,
lipids, proteins) or ascorbic acid. The results are that proteins or lipids that have become
modified (glycated) with carbohydrates in a non-ATP dependent process. Foods espe-
cially high in AGE products include those sterilized, subjected to ultra-high-temperature
processing for pasteurization or roasting. Once absorbed, these compounds have been
implicated as factors in a number of degenerative and aging-related diseases.
Focusing on protein–AGE formation, the end products can result in either a protein–
AGE adduct (mono or polysubstituted) or proteins crosslinked with AGE (protein–AGE–
protein). In addition to late/advanced stage reactants, Amadori products also degrade
into reactive carbonyls, which in turn react with amino groups forming AGE compounds.
Most of the detected AGE products are modifications of primarily lysine or arginine side
groups with a limited set of reactions known involving cysteine side groups (Lund and
Ray, 2017). Production of AGE products via the Hodge pathway involving the degrada-
tion of Amadori or Hayns intermediates can happen in a single step producing a car-
boxylated methyllysine or other AGE products depending on the reducing sugar (Figure
14.5). Formation of reactive dicarbonyls and their condensation with amino groups of
proteins via the Namiki pathway results in another suite of AGE products from glyoxal,
methyl-glyoxal, and 2-deoxy-glucose intermediates.
AGE products are based on a multitude of reactants with highly diverse and het-
erogeneous structures (Figure 14.6). One of the earliest AGE crosslinked proteins and
highly prevalent in tissues and cooking is carboxymethl-lysine (CML) formed by the
degradation of Amadori products or the addition of a reactive glyoxal to a protein’s
lysine residue. Reactions initiated with ribose will result in pentosidine, a crosslinked
AGE formed between an arginine and a lysine. The reaction is a ribose-Amadori product
involving ascorbic acid via the Hodges pathway. The reactive dicarbonyl also can gen-
erate a number of AGE products. Glyoxal-lysine dimer (GOLD) is an imidazolium ion
formed from the cyclic dimerization of two lysine side groups and glyoxal. A similar AGE
product is produced when the crosslinked proteins start instead with methyglyoxyl and
two lysine residues (MOLD). Similarly, dimers of lysine and arginine are formed with
either glyoxal or methyglyoxal. The reaction between arginine and cysteine with glucose
results in 8-hydroxy-5-methyldihydrothiazolo (3,2 alpha) pyridinium-3-carboxylate, also
named Maillard reaction product X (MRX). MRX is found in both prepared food and
 The Maillard Reaction 289

FIGURE 14.5 Maillard production of AGE compounds.

in vivo on long-lived proteins and may be an important participant in the progression of

diabetes. The MRX compound is likely formed from glucose with proteins and has been
identified after cysteine and arginine mixtures were incubated with glucose.
Foods high in fat and protein show the highest levels of AGE products after process-
ing or cooking. Foods cooked in high fat and protein content including those prepared

FIGURE 14.6 Select AGE reactants and crosslinkers+.

290 Food Aroma Evolution

with mayonnaise, olive oils, or almonds all show significant AGE products (Nguyen,
2006). Common mono- and disaccharide glucose displays the slowest rate while fruc-
tose, ribose, and glucose-6-phosphate generate AGE crosslinked proteins at the fastest rate
(Pasupulati et al., 2016). Foods high in AGE products at first were thought to be poorly
absorbed, and the biomedical impact of AGE compounds focused on endogenously pro-
duced glycans. However, measurement of the health impact of a diet high in AGE products
showed increased tissue AGE content, and ingesting foods high in AGE products increased
the risk of heart, kidney, and other diseases including diabetes in mice (Gkogkolou and
Bohm, 2012). Interestingly in diabetic animals, clearance of AGE products was reduced,
indicating a longer transit time concomitant with increased risk of renal-vascular injury
(Nguyun, 2006). Fortunately, a number of approaches have been formulated to create
effective inhibitors of AGE compounds. Trapping the reactive alpha-dicarbonyls using
epicatechin from green tea reduced off-flavors and AGE products in processed milk. A
number of other phenolic flavonoids are under investigation for their trapping and radi-
cal scavenging ability in addition to other approaches limiting high heat processing times
(Lund and Ray, 2017).

REFERENCES

Ashoor, S.H. and J.B. Zent. 1984. Maillard browning of common amino acids and sug-
ars. J. Food Sci. 49(4): 1206–7.
Bertrand, E., P.E. Boustany, C.B. Faulds, and J.L. Berdague. 2018. The Maillard reaction
in food: An introduction. In Reference Module in Food Science, pp. e1–10. Elsevier.
Eskin, M.A. and F. Shahidi. 2013. Biochemistry of Foods, 3rd Ed. Elsevier.
Gkogkolou, P. and M. Bohm. 2012. Advanced glycation end products. Key players in
skin aging? Dermatoendocrinology 4(3): 259–70.
Hellwig, M. and T. Henle. 2014. Baking, ageing, diabetes: A short history of the Maillard
reaction. Angew. Chem. Int. Ed. 53(39): 10316–29.
Hodge, L. and C.E. Rist. 1953. The amadori rearrangement under new conditions and
its significance for non-enzymatic browning reactions. J. Am. Chem. Soc. 75(2):
316–22.
Horn, M.J., H. Lichtenstein, and M. Womack. 1968. Availability of amino acids. A
methionine-fructose compound and its availability to microorganisms and rats. J.
Agric. Food Chem. 16(5): 741–5.
Kim, J.S. and Y.S. Lee. 2009. Study of Maillard reaction products derived from aqueous
model systems with different peptide chain lengths. Food Chem. 116(4): 846–53.
Lund, M.N. and C.A. Ray. 2017. Control of Maillard reactions in foods: Strategies and
chemical mechanisms. J. Agric. Food Chem. 65(23): 4537–52.
Lu, L.C., C. Hao, R. Payne, and C.T. Ho. 2006. Effects of water content on volatile
generation and peptide hydrolysis in Maillard reaction of glycine, diglycine and tri-
glycine. J. Agric. Food Chem. 53: 6443–7.
Maillard, L.C. 1911. Condensation des acides amines en presence de la glycerine:
Cycloglycyglycine et polypeptides. C. R. Hebd. Seances Acad. Sci. 153: 1078–80.
Maillard, L.-C. 1916. Syntheses des matieres humiquesa par action des acides amines sur
les sucres reducteurs. Ann. Chim. 5(0): 258–316.
Nguyen, C.V. 2006. Toxicity of the AGEs generated from the Maillard reaction: On the
relationship of food-AGEs and biological-AGEs. Mol. Nutr. Food Res. 50: 1140–9.
 The Maillard Reaction 291

Nursten, H. 2005. The Maillard Reaction Chemistry, Biochemistry and Implications.

Royal Society of Chemistry, Cambridge.
Pastoriza, S., J. Quesada, and J.A. Rufian-Henares. 2018. Lactose and oligosaccharides:
Maillard reaction. In Reference Module in Food Science, pp. e1–19. Elsevier.
Pasupulati, A.K., P.S. Chitra, and G.B. Reddy. 2016. Advanced glycation end products
mediated cellular and molecular events in the pathology of diabetic nephropathy.
BioMol. Concepts 7(5–6): 293–309.
Provost, J.J., K.L. Colabroy, B.S. Kelly, and M.A. Wallert. 2016. The Science of Cooking:
Understanding the Biology and Chemistry Behind Food and Cooking, 1st Ed. John
Wiley & Sons.
Weenen, H. and W. Apeldoorn. 1996. Carbohydrate cleavage in the Maillard reaction. In
Flavour Science: Recent Developments, A. J. Taylor and D. S. Mottram (eds.), Vol.
197, pp. 211–6. Royal Society of Chemistry, Cambridge.
Yaylayan, V. 1997. Classification of the Maillard reaction: A conceptual approach. Trends
Food Sci. Technol. 8(1): 13–8.
 Chapter 15
Production of Food Aroma
Compounds (Microbial and
Enzymatic Methodologies)
Lorena de Oliveira Felipe, Bruno Nicolau Paulino, Adones
Sales, Gustavo Molina, and Juliano Lemos Bicas

CONTENTS

15.1 Introduction 293

15.2 Microbial-Derived Aroma Compounds 294
15.2.1 Lactic Fermentation 295
15.2.2 Alcoholic Fermentation 297
15.2.3 Microbial Production of Aroma Additives (Bioaromas) 298
15.3 Enzyme-Derived Aroma Compounds 299
15.3.1 Enzymatic Aroma Generation during Processing 300
15.3.2 Enzyme Production of Aroma Additives (Bioaromas) 301
15.4 Final Remarks 302
References 302

15.1 INTRODUCTION

Aromas are part of our daily lives, as well as the history of humanity. They were useful
as a survival tool in prehistory, and nowadays they affect us on a physical, psychological,
and social level (Classen et al., 2002). The presence of aroma in foods, for example, is
an important characteristic related to quality that contributes to memories that influence
our preferences, mood, and even buying decisions (Schab and Cain, 1991; Beyts et al.,
2017; Schoumacker et al., 2017; Wijk et al., 2018). The general perception during food
consumption is formed by a set of physical and chemical sensations perceived by the
senses. Texture, fluidity, or crispness, for example, are perceived by touch and hearing
(Szczesniak, 2002). The palate identifies the taste (salty, sweet, bitter, sour, and umami),
while smell is responsible for the perception of the aroma. The result of these interac-
tions is called flavor (Auvray and Spence, 2008). As part of the flavor perception, aroma
is the result of volatile compounds in foods. This sensation may result from one single
compound, but it usually comes from the interaction between mixtures of volatiles and
is also influenced by the nonvolatile constituents (Taylor and Linforth, 2003). From more
than 7000 volatile compounds already identified in food, only 5% have an impact on
food aroma. These key food odorants (KFOs) have been organized into generalists, inter-
mediaries, and individualists, based on their distribution in food products (Dunkel et

293
294 Food Aroma Evolution

al., 2014). Aroma compounds, usually found in very low concentrations (i.e., in the ppm
range or even lower), may originally be present in the food (i.e., volatiles from raw materi-
als) or produced during food processing or storage. During processing, these compounds
may be formed by enzymatic and non-enzymatic reactions or by fermentation processes
(Longo and Sanromán, 2006). Moreover, aroma compounds may be added to foodstuffs
(i.e., used as additives) to develop the sensory characteristics of these products (Chambers
and Koppel, 2013). In this case, aroma additives might be isolated from nature, synthe-
sized by chemical means, or produced biotechnologically (Gupta et al., 2015).
Food preference, purchase, and consumption are influenced by aromas, which encour-
ages the use of these compounds as additives in the food industry. In 2016, the global
aroma market reached $4.08 billion, and it is expected to grow at a compound annual
growth rate (CAGR) of 6.8% to reach $6.48 billion by 2023 (Statistics: MRC, 2017).
Part of this market includes the so-called “biotech aromas” or “bioaromas,” which are
biotechnologically produced using microorganisms (or other cells) or enzymes, either
native or genetically modified, the process of which is summarized in Figure 15.1. The
interest in such an approach to aroma production is mostly driven by technical, envi-
ronmental, and commercial concerns, for example, milder reaction conditions, no need
for toxic or polluting catalysts, and the higher economic value of the resulting products
(Felipe et al., 2017). This chapter will explore key examples of aroma compounds formed
during processing through the action of microorganisms (fermentation) or enzymes as
well as the production of aroma additives via these two routes.

15.2 MICROBIAL-DERIVED AROMA COMPOUNDS

Fermentation processes yield aroma compounds which are associated with the singular and
well known sensorial perception attributed to fermented products, such as beers, cheeses, and
wines. In addition, the biosynthesis of volatile compounds by microorganisms is the basis
for the biotechnological production of the so-called bioaromas. Recognized as natural com-
pounds, these food additives have gained attention in the food industry due to the increased
interest in food attributes such as freshness, healthfulness, and naturalness (Román et al.,
2017). Therefore, in the following paragraphs, the main aroma compounds produced during
microbial fermentation will be discussed, with a focus on lactic and alcoholic fermentation,
as well as some exemplification related to the bioaromas produced by microorganisms.

FIGURE 15.1 Stages of the biotechnological production of aroma.

 Production of Food Aroma Compounds  295

15.2.1 Lactic Fermentation

Lactic fermentation allows for the production of different compounds that deliver the
singular aromatic identity of some classic products, such as butter, cheese, and yogurt
(Tamime, 2002). Lactic acid bacteria (LAB), especially Lactobacillus sp.; Leuconostoc
sp., and Streptococcus sp., are involved in such a process. LAB are classified as homofer-
mentative or heterofermentative (Gänzle, 2015) depending on the metabolic route used
to produce energy. In homolactic fermentation, LAB such as Streptococcus sp. use the
Embden–Meyerhof–Parnas pathway to convert glucose into lactic acid (C3H6O3) as a sole
product (2 mols per glucose), besides yielding a “fresh” flavor (Martinez et al., 2013).
On the other hand, in heterolatic fermentation, other LAB, such as Leuconostoc, use
the Pentose–Phosphate pathway to convert glucose to produce, ethanol (C2H5OH) or
acetic acid (C2H3O2), as well as lactic acid (1 mol per glucose) and carbon dioxide (CO2).
Additionally, during heterolatic fermentation, the citrate is concomitantly metabolized
with carbohydrates forming aroma compounds: besides the acetaldehyde, C4 carbonyl
molecules, such as diacetyl (C4H6O2), acetoin (C4H8O2), and butanediol (C4H10O2), are
also obtained (Figure 15.2) (Gänzle, 2015). Also produced from the citrate pathway, acet-
aldehyde may result from threonine by the action of threonine aldolase (Zouraria et al.,
1992; Chen et al., 2017).

FIGURE 15.2 Formation of volatile compounds during lactic acid fermentation. (From
Lindsay, 2007.)
296 Food Aroma Evolution

Acetaldehyde (ethanol) is recognized as a character impact compound, associated

with the fresh flavor in yogurt. In such products, milk fermentation is carried out by a
mixed culture of Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bul-
garicus (Zouraria et al., 1992). At the end of the process, more than 90 different volatile
compounds may be identified in this dairy product, such as acetone, diacetyl, ethanol,
and acetaldehyde, the latter volatile being reported as the major compound. In yogurt,
the odor threshold of acetaldehyde is in the range of 0.0079–0.039 mg kg−1. However,
the ideal concentration for consumer acceptability should be between 14 and 20 mg
kg−1 (Chen et al., 2017). Diacetyl (2,3-butanedione) is a key aroma compound in butter
flavor. The aroma and taste threshold values for this compound are 5 ppb (recognition)
and 50 ppm, respectively (Burdock, 2002). A mixed culture of Lactococcus lactis and
Lactococcus diacetilactis is used to ferment milk cream to produce mature or acid butter.
The former strain produces lactic acid; while the latter, which shows a pronounced ability
to metabolize citrate, is responsible for diacetyl production.
Proteolysis and lipolysis are considered key processes in ripened cheese production,
such as blue cheeses (e.g., Bleu d’Auvergne, Cabrales, Danablu, Gorgonzola, Roquefort,
and Stilton), which are characterized by the presence of methyl ketones. Briefly, the fer-
mentation process for blue cheese production can be divided into two parts: primary
fermentation, associated with the activity of LAB, and secondary fermentation (ripening)
carried out by fungi, mainly Penicillium roqueforti (Martin and Cotton, 2016). The main
metabolic pathway for methyl ketone production involves the action of lipases for liber-
ating free fatty acids (FFA), which are further oxidized (β-oxidation), followed by the
decarboxylation of the resulting β-keto acids. Later, some of these methyl ketones may be
reduced to its correspondent alcohols (McSweeney and Sousa, 2000) (Figure 15.3). Some
of the non-converted FFA, such as butanoic (C4H8O2) and caproic acid (C6H12O2), are
also important for the aroma of blue cheese (McSweeney and Sousa, 2000; Ardö, 2011).
Other cheese aromas formed by enzyme action will be discussed in Section 15.3.1.
Many studies have discussed the main parameters that affect the aroma composition
of dry fermented sausages, showing the role of microorganisms such as bacteria, yeasts
and molds, and endogenous enzymes in the different pathways involved in flavor develop-
ment, including carbohydrate metabolism, and the degradation of free amino acids and
fatty acids (Flores et al., 2015; Flores and Olivares, 2015). The LAB present in fermented
meat products are related to carbohydrate fermentation, which is used to produce lactic

FIGURE 15.3 Formation of aroma compounds (short-chain fatty acids and methyl
ketones) in blue cheese.
 Production of Food Aroma Compounds  297

acid and volatile compounds associated with the aroma of these products, including acet-
aldehyde, diacetyl, as well as short-chain alcohols and organic acids (Ravyts et al., 2012).
In this context, lipases and proteinases are considered as the key enzymes associated
with the production by microorganisms from starter cultures of the typical aroma in
fermented meat products (Molly et al., 1997; Cocolin et al., 2006; Franciosa et al., 2018).
Section 15.3.1 also presents other aroma compounds formed by enzyme action in meat.

15.2.2 Alcoholic Fermentation

The main commercially relevant alcoholic fermentations employ yeasts of the

Saccharomyces genus, particularly S. cerevisiae, which produces carbon dioxide (CO2)
and ethanol (C2H6O) as final products of glucose metabolism. In addition to contributing
to the alcoholic aroma note associated to fermented beverages, ethanol is also regarded
as a carrier agent of other aroma active compounds, such as “fusel alcohol” (higher alco-
hols), organic acids, and esters, which are simultaneously produced during the fermenta-
tion process. Together, these odor-active compounds are essential for establishing the
unique aroma of the final product (Hazelwood et al., 2008). Regarding higher alcohol
production, these compounds result from amino acid metabolism via the Ehrlich path-
way. In this case, the amino acids (either from the substrate or produced de novo by the
microorganism) are first transformed into the corresponding α-ketoacids via transamina-
tion, followed by its decarboxylation to the corresponding aldehyde, which undergoes
reduction reactions to yield higher alcohols (Pires et al., 2014). Thus, it is expected that
the quantity and quality of higher alcohols obtained will be correlated to the concentra-
tion of the amino acids originally present or supplemented in the fermentative substrate.
These higher alcohols contribute to different notes in the final product, such as floral,
fruity, herbaceous, pungent, and rose notes (Hazelwood et al., 2008; Pires et al., 2014).
Thereafter, the alcohols initially formed (e.g., ethanol and higher alcohols) can be esteri-
fied to organic acids from yeast metabolism via enzymatic condensation. This results in
the production of esters, which are volatiles associated with fruity notes. For instance,
different esters can be identified (Table 15.1) and most of them are reported as important
odor-active compounds in beer, mainly due to their low threshold values (Hazelwood et
al., 2008; Pires et al., 2014). On the other hand, the diacetyl production is a big concern
to the beer industry, since this product results in buttery notes (off-flavor) in the final
product (Krogerus and Gibson, 2013).

TABLE 15.1 Main Aroma Esters in Beer

Compound Chemical Formula Aroma Description
Ethyl acetate 1 C4H8O2 “Fruity,” “solvent-like” 3
Ethyl hexanoate 1 C8H16O2 “Apple-like” 4
Ethyl octanoate 1 C10H20O2 “Apple” 4
Isoamyl acetate 1 C7H14O2 “Banana” 3
Isobutyl acetate 2 C6H12O2 “Fruity” 3
2-phenylethyl acetate 1 C10H12O2 “Honey,” “roses,” “flowery” 3
Sources: Adapted from 1 Cordente et al., 2012; 2 Pires et al., 2014.
Note: 3 Acetate ester; 4 medium-chain fatty acid (MCFA) ethyl ester.
298 Food Aroma Evolution

TABLE 15.2 Some Examples of Secondary Aroma Compounds

in Wine
Chemical Group Key Examples Aroma Description
Alcohols 1-Hexanol (C6H14O) Grass 1
1-Octen-3-ol (C8H16O) Mushroom 2
Aldehydes Acetaldehyde (C2H4O) “Green” aroma 3
Hexanal (C6H12O)
Octanal (C8H16O) “Citrus” fruit 3
Ketone β-Damascenone (C13H18O) “Baked apple” 1
Sources: Adapted from 1 Escudero et al., 2007; 2 Genovese et al., 2007;
3 Culleré et al., 2011.

In terms of wine, its aroma profile results from the complex combination of primary,
secondary, and tertiary aromas. The first refers to volatiles (mostly terpenes) that are natu-
rally present in different grape varieties and, in general, are associated with floral notes
(Styger et al., 2011). Fermentation and ripening are responsible for secondary and tertiary
aroma production, respectively. In general, the secondary aromas (see some examples in
Table 15.2) are produced in an analogue way, as explained at the beginning of this section.
Besides alcoholic fermentation, malolactic fermentation also takes place in winemak-
ing. This fermentation reduces wine acidity by converting malic acid into lactic acid using
LAB. In such a process, butter notes can be introduced as a result of lactic fermentation
(Betteridge et al., 2015), as already cited in Section 15.2.1. Moreover, other volatiles
may be formed by the action of microorganisms. Englezos et al. (2018a), for instance,
evaluated the volatile profile of red wine produced using mixed fermentations carried
out with Starmerella bacillaris and S. cerevisiae using four red grape varieties and two
different inoculation protocols. The results suggested a higher activity or expression of
β-glycosidase enzymes by the non-Saccharomyces yeast as observed by the release of
monoterpenes (e.g., citronellol, geraniol, linalool, and C13 -norisoprenoid β-damascenone)
from the glycoside precursors present in grapes. The enzymatic formation of these com-
pounds is very important due to their significant contribution to the fruity and floral
aroma of wine (Pons et al., 2017) (some of the examples of enzyme aroma formation in
wine will be presented in Section 15.3.1). The result of different combinations of these
compounds results in a huge variety of wines, which are distinguished by their unique
“bouquet” (Cappello et al., 2017).

15.2.3 Microbial Production of Aroma Additives (Bioaromas)

As mentioned in Section 15.1, aroma compounds may be recovered from nature, pro-
duced by chemical synthesis, or by biotechnological means. The compounds extracted
directly from natural matrices are subjected to seasonality effects, besides usually being
associated with low yields and productivities (very long production times). In contrast,
chemical synthesis is associated with high yields and low costs, but the by-products
eventually formed in the reactions may negatively affect the sensorial perception of the
final product. Moreover, most chemical processes are regarded as non-environmentally
friendly processes. Aroma compounds obtained by biotechnological means, on the
other hand, may be labeled as “natural” and are produced in environmentally friendly
 Production of Food Aroma Compounds  299

TABLE 15.3 Some Aroma Compounds Commercially Produced by

Biotechnological Means
Compound Route Company
Farnesene De novo synthesis Amyris
Nootkatone Biotransformation Evolva, Oxford Biotrans, Isobionics
Patchouli De novo synthesis Amyris, Firmenich
Vanillin Biotransformation Solvay
De novo synthesis IFF, Evolva
α-Santalol De novo synthesis Evolva
Source: Adapted from Sales et al., 2018.

conditions, besides being independent of seasonal and climatic conditions. The biggest
challenge of this process is to overcome the low yields that are usually achieved (Bicas
et al., 2016; Felipe et al., 2017). The aroma compounds produced by microorganisms
can be done in two different ways: by de novo synthesis, where the target molecule(s)
is(are) synthesized from simple substrates following complete metabolic routes; or by a
biotransformation process, where a substrate is converted to a homologous molecule in
one (biotransformation) or a few (bioconversion) catalytic steps. Considering the specific-
ity usually attributed to biological reactions, it is even possible to obtain enantiomerically
pure compounds via such bioprocesses (Bicas et al., 2016, 2009). Another important
motivation for producing “biotech” aromas is related to their market value: although
usually cheaper than their counterparts extracted from nature, the bioaromas are more
valued than their synthetic analogue (Felipe et al., 2017). Consequently, different compa-
nies currently have industrial-scale production processes for the “biotec” aromas, such as
the examples shown in Table 15.3 (Sales et al., 2018).
In this context, it is worth mentioning the growing importance of synthetic biology for
the large-scale microbial production of bioaromas (Meadows et al., 2016). In this sense,
scientists are currently mimicking nature to obtain aroma compounds of interest to the
consumer: by manipulating microorganisms in order to reproduce the metabolic routes
found in plants, the genetically modified strain starts producing the target molecules origi-
nally present in the chosen fruit or vegetable. Moreover, genetic modifications may also be
used to overcome some of the challenges associated with large-scale production processes.
Strategies such as the reduction of substrate toxicity and the increase in yields have been
considered. Thus, some genetically modified microorganisms (e.g., Schizosaccharomyces
pombe, Saccharomyces cerevisiae, and Escherichia coli) are already being employed on an
industrial scale for the commercial production of important flavors and fragrances materi-
als (e.g., vanillin, patchouli, and ambroxide, respectively) (Felipe et al., 2017).

15.3 ENZYME-DERIVED AROMA COMPOUNDS

Besides being employed in the food and beverage industries for a wide range of applica-
tions (Singh and Kumar, 2019), enzymes such as lipases, proteases, peroxidases, lipoxy-
genases, amine oxidase, among others, are related with aroma (and off-flavor) production
in food products. Enzymatic aroma production can take place in situ during the process-
ing and storage of foods, that is, through the action of enzymes endogenously present,
added, or produced in the food, or in vitro, where the synthesized products are further
300 Food Aroma Evolution

used as food additives (Imafidon and Spanier, 1994; Furuya et al., 2017; Sales et al.,
2018). These two topics will be discussed in the next section.

15.3.1 Enzymatic Aroma Generation during Processing

Enzyme-modified cheeses (EMC) are food products in the form of pastes or powders con-
taining concentrated cheese flavors resulting from the action of several enzymes, such as
peptidases and lipase, on mild cheese or fresh cheese curd (Kilcawley et al., 1998; Azarnia
et al., 2010; Miri and Najafi, 2011). The action of peptidases is associated with the pro-
duction of free amino acids, which are important for taste and are the precursors of
aroma compounds (Nuñez, 2016). Moreover, the liberation of free fatty acids from triac-
ylglycerols through the action of lipolytic enzymes is also essential not only to their direct
impact on aroma (short-chain fatty acids) but also for being precursors of different vola-
tile organic compounds (Collins et al., 2003). After oxidation reactions, for instance, free
fatty acids can be converted into β-ketoacids, which are transformed into methyl ketone
when decarboxylated, in a similar sequence of reactions already cited in Section 15.2.1
for blue cheese (Figure 15.3). The catabolism of free fatty acids can also produce second-
ary alcohols, lactones, and acids with different contributions to the aroma of cheeses
(McSweeney and Sousa, 2000; Collins et al., 2003). Moreover, γ- and δ-hydroxylated
free fatty acids produced by the action of lipoxygenase and hydratase enzymes are pre-
cursors of lactones (Dirinck and DeWinne, 1999; Singh et al., 2003). Other important
precursors of aroma compounds in EMC, such as α-keto acids, may also be formed from
amino acids by transamination reactions mediated by aminotransferase (Bertuzzi et al.,
2018). These compounds can be converted into aroma compounds, including branched-
chain and aromatic aldehydes, and methanethiol (Ganesan and Weimer, 2017). Some
aldehydes, such as 2-methylpropanal, 2-methylbutanal, and 3-methylbutanal, are formed
by the transamination of valine, isoleucine, and leucine, respectively, while the forma-
tion of acetoin, diacetyl, or 2,3-butanediol is related to the oxaloacetate formed after
the transamination of aspartic acid (Singh et al., 2003; Ardö, 2006; Le Bars and Yvon,
2008). In meat products, the production of aroma compounds is related to several reac-
tions during processing, including the enzymatic degradation (proteolysis, glycolysis, and
lipolysis) of some constituents, which also has a positive impact on the production of
nonvolatile compounds associated with meat flavor (Toldrá and Flores, 2000; Khan et
al., 2015). Moreover, endogenous enzymes act during the postmortem aging of meat to
tenderize it as well as to generate flavor precursors (free amino acids), which further react
with other degradation products and yield volatile compounds related with meat aroma
(Khan et al., 2015).
The application of glycosidases for wine aroma enhancement has also been con-
sidered. Recent examples include the application of extracellular glycosidase from
yeast for wine aroma enhancement (Hu et al., 2016; Ma et al., 2017; Sun et al., 2018),
including the use of extracellular β-glucosidase from Issatchenkia terricola to signifi-
cantly increase the amount of monoterpenes and norisoprenoids during the fermenta-
tion (González-Pombo et al., 2011). Moreover, the action of β-lyases produced by yeast
during winemaking may also transform nonvolatile precursors, such as cysteine and
glutathione conjugates, on volatile thiols, such as 4-mercapto-4-methyl-2-pentanone
(4MMP), 3-mercapto-1-hexanol (3MH), and acetate 3-mercaptohexyl acetate (3MHA),
which contribute positively to fruit notes in wine aroma (Murat et al., 2001; Englezos
et al., 2018b).
 Production of Food Aroma Compounds  301

15.3.2 Enzyme Production of Aroma Additives (Bioaromas)

The application of isolated enzymes in aroma synthesis is a well-studied process and

several compounds can be produced using this approach. The use of lipases to generate
different flavor esters (e.g., esters of glycerol, aliphatic alcohols, and terpene alcohols),
usually associated with fruity notes, is the classic example. Besides catalyzing hydrolysis
reactions, which are key reactions for the formation of cheese aroma (see Sections 15.2.1
and 15.3.1), lipases may also catalyze esterification and interesterification reactions under
certain circumstances (e.g., low water activity), leading to ester production (Akacha and
Gargouri, 2015). The enzymatic synthesis of butyl stearate and ethyl stearate from stearic
acid and the respective alcohols (n-butanol and ethanol, for instance, has been reported
using Novozym 435 (lipase) as a catalyst (Pereira et al., 2018). The production of ethyl
caprylate from caprylic acid and ethanol using free or immobilized lipase and cyclo-octane
as the solvent is another recent example of such a process (Patel et al., 2015). During
recent years, new approaches have been considered for developing ester flavor production
using lipases. These include the adoption of immobilization techniques (physical adsorp-
tion, entrapment or microencapsulation, and covalent binding) to stabilize the enzyme;
the use of unconventional solvents (supercritical CO2) in the reaction media; and the
application of microwave irradiation techniques (Yadav and Thorat, 2012; Lisboa et al.,
2018), such as the examples given in the following paragraphs. The production of n-butyl
propionate through the esterification of propionic acid with n-butanol, for instance, was
reported using different immobilized enzymes (Lypozyme RM IM, Lypozyme TLIM,
and Novozym 435) under microwave irradiation to reduce the reaction time (Bhavsar
and Yadav, 2018a). Similarly, this enzyme also showed the best performance in the syn-
thesis of ethyl valerate from valeric acid and ethanol as precursors, exhibiting a syner-
gistic effect when microwave irradiation was applied (Bhavsar and Yadav, 2018b). The
microwave-assisted synthesis of isoamyl acetate was also described using two commercial
lipases and different acyl donors, such as acetic acid, acetic anhydride, and ethyl acetate,
as well as the microwave energy levels (Zare et al., 2019). Moreover, the use of super-
critical CO2 as a solvent is another approach being considered for lipase-catalyzed ester
aroma production, such as isoamyl acetate, eugenyl acetate, isobutyl acetate, isoamyl
propionate, isopropyl propionate, isoamyl laureate, geranyl acetate, and others (Santos et
al., 2016, 2017; Dias et al., 2018a,b).
Another use of lipases is for kinetic resolution of racemic mixtures of different
aroma compounds (Todea et al., 2018). Considering that different enantiomers exhibit
a singular odor, the application of enzymes for the kinetic resolution of racemic mix-
tures stands out as an interesting strategy for obtaining enantiomerically pure aroma
compounds for the food, pharmaceutical, and cosmetic industries (Jadhav et al., 2016),
such as the examples presented next. 2-Phenylethanol presents a rose-like odor, while
1-phenylethanol is reported as the predominant aroma compound found in tea flowers
(Hua and Xu, 2011; Zhou et al., 2018). The enzymatic resolution of racemic 1-phenyl-
ethyl acetate to (S)-1-phenylethanol (enantiomeric excess of 97% and conversion of
28.5%) has been achieved using a novel marine microbial GDSL lipase, which exhibited
opposite stereo-selectivity than other common lipases in both transesterification reac-
tions and hydrolysis reactions (Deng et al., 2016). The application of partially puri-
fied cutinase from Fusarium ICT SAC1 was also described in the kinetic resolution of
(R,S)-1-phenylethanol using microwave irradiation to increase conversion efficiency and
enantioselectivity (Kamble et al., 2017). Although the use of different enzymes is well
established for aroma production, further studies are needed, especially focusing on
302 Food Aroma Evolution

the production of new flavor compounds of industrial interest and on discovering new
enzymes that are more adapted to the processing conditions.

15.4 FINAL REMARKS

Aroma compounds have historically been produced in food by empirical processing, such
as in fermented dairy and meat products. Only in recent decades could scientists achieve
greater understanding of the mechanisms behind the origins of these compounds. Some
of the main examples of aromas formed in these traditional fermentations were pre-
sented in this chapter. This chapter also presented key examples of aroma compounds
produced by microorganisms or enzymes which are already a commercial reality, some
of them based on synthetic biology. Therefore, it is expected that genetically engineered
microorganisms or enzymes will still be increasingly employed to develop a better aroma
profile in traditional fermented products as well as to produce new aroma compounds by
biotransformations or de novo synthesis.

REFERENCES

Akacha, N.B. and Gargouri, M. 2015. Microbial and enzymatic technologies used for
the production of natural aroma compounds: Synthesis, recovery modeling, and
bioprocesses. Food Bioprod. Process. 94:675–706.
Ardö, Y. 2006. Flavour formation by amino acid catabolism. Biotechnol. Adv. 24:238–42.
Ardö, Y. 2011. Blue mold cheese. In: Encyclopedia of Dairy Sciences, ed. J.W. Fuquay, p.
767–72. London: Academic Press.
Auvray, M. and Spence, C. 2008. The multisensory perception of flavor. Conscious.
Cogn. 17:1016–31.
Azarnia, S., Lee, B.H., Yaylayan, V., and Kilcawley, K.N. 2010. Proteolysis development
in enzyme-modified cheddar cheese using natural and recombinant enzymes of lac-
tobacillus rhamnosus S93. Food Chem. 120:174–8.
Bertuzzi, A.S., McSweeney, P.L.H., Rea, M.C., and Kilcawley, K.N. 2018. Detection of
volatile compounds of cheese and their contribution to the flavor profile of surface-
ripened cheese. CRFSFS. 17:371–90.
Betteridge, A., Grbin, P., and Jiranek, V. 2015. Improving Oenococcus oeni to overcome
challenges of wine malolactic fermentation. Trends Biotechnol. 33:547–53.
Beyts, C., Chaya, C., Dehrmann, F., James, S., Smart, K., and Hort, J. 2017. A compari-
son of self-reported emotional and implicit responses to aroma in beer. Food Qual.
Prefer. 59:68–80.
Bhavsar, K.V. and Yadav, G.D. 2018a. Microwave assisted solvent-free synthesis of n-butyl
propionate by immobilized lipase as catalyst. Biocatal. Agric. Biotechnol. 14:264–9.
Bhavsar, K.V. and Yadav, G.D. 2018b. Process intensification by microwave irradiation
in immobilized-lipase catalysis in solvent-free synthesis of ethyl valerate. Mol. Catal.
461:34–9.
Bicas, J.L., Dionisio, A.P., and Pastore, G.M. 2009. Bio-oxidation of terpenes: An
approach for the flavor industry. Chem. Rev. 109:4518–31.
Bicas, J.L., Molina, G., Barros, F.F.C., and Pastore, G.M. 2016. Chapter 12. Production of
aroma compounds by white biotechnology. In: White Biotechnology for Sustainable
Chemistry, eds. M.A.Z. Coelho and B.D. Ribeiro, pp. 310–32. Cambridge: Royal
Society of Chemistry.
 Production of Food Aroma Compounds  303

Burdock, G.A. 2002. Fenaroli’s Handbook of Flavor Ingredients. 4th Ed., pp. 424–5.
Boca Raton, FL: CRC Press.
Cappello, M.S., Zapparoli, G., Logrieco, A., and Bartowsky, E.J. 2017. Linking wine
lactic acid bacteria diversity with wine aroma and flavor. Int. J. Food Microbiol.
243:16–27.
Chambers, E.I.V. and Koppel, K. 2013. Associations of volatile compounds with sensory
aroma and flavor: The complex nature of flavor. Molecules. 18:4887–905.
Chen, C., Zhao, S., Hao, G., Yu, H., Tian, H., and Zhao, G. 2017. Role of lactic acid
bacteria on the yogurt flavour: A review. Int. J. Food Prop. 20:S316–30.
Classen, C., Howes, D., and Synnott, A. 2002. Aroma: The Cultural History of Smell.
London: Routledge.
Cocolin, L., Urso, R., Rantsiou, K., Cantoni, C., and Comi, G. 2006. Dynamics and
characterization of yeasts during natural fermentation of Italian sausages. FEMS
Yeast Res. 6:692–701.
Collins, Y.F., McSweeney, P.L.H., and Wilkinson, M.G. 2003. Lipolysis and free
fatty acid catabolism in cheese: A review of current knowledge. Int. Dairy J. 13:
841–66.
Cordente, A.G., Curtin, C.D., Varela, C., and Pretorius, I.S. 2012. Flavour-active wine
yeasts. Appl. Microbiol. Biotechnol. 96:601–18.
Culleré, L., Ferreira, V., and Cacho, J. 2011. Analysis, occurrence and potential sensory
significance of aliphatic aldehydes in white wines. Food Chem. 127:1397–403.
Deng, D., Zhang, Y., Sun, A., Liang, J., and Hu, Y. 2016. Functional characterization
of a novel marine microbial GDSL lipase and its utilization in the resolution of
(±)-1-phenylethanol. Appl. Biochem. Biotechnol. 179:75–93.
Dias, A.L.B., Cunha, G.N., Santos, P., Meireles, M.A., and Martínez, J. 2018a. Fusel oil:
Water adsorption and enzymatic synthesis of acetate esters in supercritical CO2 . J.
Supercrit. Fluids. 142:22–31.
Dias, A.L.B., Santos, P., and Martínez, J. 2018b. Supercritical CO2 technology applied
to the production of flavor ester compounds through lipase-catalyzed reaction: A
review. J. CO2 Utiliz. 23:159–78.
Dirinck, P. and DeWinne, A. 1999. Flavour characterization and classification of cheeses
by gas chromatographic-mass spectrometric profiling. J. Chromatog. A. 847:
203–8.
Dunkel, A., Steinhaus, A., Kotthoff, M., Nowak, B., Krautwurst, D., Schieberle, P., and
Hofmann, T. 2014. Natures chemical signatures in human olfaction. Angew. Chem.
Int. 53:7124–43.
Englezos, V., Rantsiou, K., Cravero, F., Torchio, F., Giacosa, S., Ortiz-Julien, A., Gerbi,
V., Rolle, L., and Cocolin, L. 2018a. Volatile profiles and chromatic characteristics
of red wines produced with Starmerela bacillaris and Saccharomyces cerevisiae.
Food Res. Int. 109:298–309.
Englezos, V., Rantsiou, K., Cravero, F., Torchio, F., Pollon, M., Fracassetti, D., Ortiz-
Julien, A., Gerbi, V., Rolle, L., and Cocolin, L. 2018b. Volatile profile of white wines
fermented with sequential inoculation of Starmerella bacillaris and Saccharomyces
cerevisiae. Food Chem. 257:350–60.
Escudero, A., Campo, E., Fariña, L., Cacho, J., and Ferreira, V. 2007. Analytical char-
acterization of the aroma of five premium red wines. Insights into the role of odor
families and the concept of fruitiness of wines. J. Agric. Food Chem. 55:4501–10.
Felipe, L.O., de Oliveira, A.M., and Bicas, J.L. 2017. Bioaromas—Perspectives for sus-
tainable development. Trends Biotechnol. 62:141–53.
304 Food Aroma Evolution

Flores, M., Corral, S., Cano-García, L., Salvador, A., and Belloch, C. 2015. Yeast strains
as potential aroma enhancers in dry fermented sausages. Int. J. Food Microbiol.
212:16–24.
Flores, M. and Olivares, A. 2015. Flavor. In: Handbook of Fermented Meat and Poultry.
2nd Ed., ed. F. Toldrá, pp. 217–25. John Wiley & Sons.
Franciosa, I., Alessandria, V., Dolci, P., Rantsiou, K., and Cocolin, L. 2018. Sausage fer-
mentation and starter cultures in the era of molecular biology methods. Int. J. Food
Microbiol. 279:26–32.
Furuya, T., Kuroiwa, M., and Kino, K. 2017. Biotechnological production of vanillin
using immobilized enzymes. J. Biotechnol. 243:25–8.
Ganesan, B. and Weimer, B.C. 2017. Amino acid catabolism and its relationship to cheese
flavor outcomes. In: Cheese: Chemistry, Physics, and Microbiology. 4th Ed., eds.
P.L.H. McSweeney, P.F. Fox, P.D. Cotter, and D.W. Everett, pp. 483–516. Academic
Press.
Gänzle, M. 2015. Lactic metabolism revisited: Metabolism of lactic acid bacteria in food
fermentations and food spoilage. Curr. Opin. Food Sci. 2:106–17.
Genovese, A., Gambuti, A., Piombino, B., and Moio, L. 2007. Sensory properties and
aroma compounds of sweet Fiano wine. Food Chem. 96:601–18.
González-Pombo, P., Fariña, L., Carrau, F., Batista-Viera, F., and Brena, B.M. 2011. A
novel extracellular β-glucosidase from Issatchenkia terricola: Isolation, immobiliza-
tion and application for aroma enhancement of white Muscat wine. Proc. Biochem.
46:385–9.
Gupta, C., Prakash, D., and Gupta, S. 2015. A biotechnological approach to microbial
based perfumes and flavours. J. Microbiol. Exp. 2:1–8.
Hazelwood, L.A., Daran, J-M., van Maris, A.J.A., Pronk, J.T., and Dickinson, J.R.
2008. The Ehrlich pathway for fusel alcohol production: A century of research on
Saccharomyces cerevisiae metabolism. Appl. Environ. Microbiol. 74:2259–66.
Hu, K., Zhu, X.L., Mu, H., Ma, Y., Ullah, N., and Tao, Y.S. 2016. A novel extracellu-
lar glycosidase activity from Rhodototula mucilaginosa: Its application potential in
wine aroma enhancement. Lett. Appl. Microbiol. 62:169–76.
Hua, D. and Xu, P. 2011. Recent advances in biotechnological production of 2-phenyl-
ethanol. Biotechnol. Adv. 29:654–60.
Imafidon, G.I. and Spanier, A.M. 1994. Unraveling the secret of meat flavor. Trends
Food Sci. Technol. 5:315–21.
Jadhav, D.D., Patil, H.S., Chaya, P.S., and Thulasiram, H.V. 2016. Fungal mediated
kinetic resolution of racemic actates to (R)-alcohols using Fusarium proliferatum.
Tetrahedron Lett. 57:4563–7.
Kamble, M.P., Chaudhari, S.A., Singhal, R.S., and Yadav, G.D. 2017. Synergism of micro-
wave irradiation and enzyme catalysis in kinetic resolution of (R,S)-1-phenylethanol
by cutinase from novel isolate Fusarium ICT SAC1. Biochem. Eng. J. 117:
121–8.
Khan, M.I., Jo, C., and Tariq, M.R. 2015. Meat flavor precursors and factors influencing
flavor precursors—A systematic review. Meat Sci. 110:278–84.
Kilcawley, K.N., Wilkinson, M.G., and Fox, P.F. 1998. Review: Enzyme-modified cheese.
Int. Dairy J. 8:1–10.
Krogerus, K. and Gibson, B.R. 2013. Diacetyl and its control during brewery fermenta-
tion. J. Inst. Brew. 119:86–97.
Le Bars, D. and Yvon, M. 2008. Formation of diacetyl and acetoin by Lactococcus lactis
via aspartate catabolism. J. Appl. Microbiol. 104:171–7.
 Production of Food Aroma Compounds  305

Lindsay, R.C. 2007. Flavors. In: Fennema’s Food Chemistry. 4th Ed., eds. S. Damodaran,
K.L. Parkin, and O.R. Fennema. CRC Press.
Lisboa, M.C., Rodrigues, C.A., Barbosa, A.S., Mattedi, S., Freitas, L.S., Mendes, A.A.,
Dariva, C., Franceschi, E., Lima, A.S., and Soares, C.M.F. 2018. New perspectives
on the modification of silica aerogel particles with ionic liquid used in lipase immo-
bilization with platform in ethyl esters production. Process. Biochem. 75:157–65.
Longo, M.A. and Sanromán, M.A. 2006. Production of food aroma compounds. Food
Technol. Biotechnol. 44:335–53.
Ma, D., Yan, X., Wang, Q., Zhang, Y., and Tao, Y. 2017. Performance of selected P.
fermentans and its excellular enzyme in co-inoculation with S. cerevisiae for wine
aroma enhancement. LWT—Food Sci. Technol. 86:361–70.
Martin, J.F. and Coton, M. 2016. Chapter 12. Blue cheese: Microbiota and fungal
metabolites. In: Fermented Foods in Health and Disease Prevention, eds. J. Frias,
C. Martinez-Villaluenga, and E. Peñas, pp. 275–303. London: Academic Press.
Martinez, F.A.C., Balciunas, E.M., Salgado, J.M., González, J.M.D., Converti, A., and
Oliveira, R.P.S. 2013. Lactic acid properties, applications and production: A review.
Trends Food Sci. Technol. 30:70–83.
McSweeney, P.L.H. and Sousa, M.J. 2000. Biochemical pathways for the production of
flavor compounds in cheeses during ripening: A review. Le Lait. 80:293–324.
Meadows, A.M., Hawkins, K.M., Tsegaye, Y., et al. 2016. Rewriting yeast central car-
bon metabolism for industrial isoprenoid production. Nature. 537:694–7.
Miri, A.M. and Najafi, H.B.M. 2011. The effect of adding enzyme-modified cheese on
sensory and texture properties of low- and high-fat cream cheeses. Int. J. Dairy
Technol. 64:92–8.
Molly, K., Demeyer, D., Johansson, G., Raemaekers, M., Ghistelinck, M., and Geenen,
I. 1997. The importance of meat enzymes in ripening and flavor generation in dry
fermented sausages. First results of a European project. Food Chem. 59:539–45.
Murat, M.L., Masneuf, I., Darriet, P., Lavigne, V., Tominaga, T., and Dubourdieu, D.
2001. Effect of Saccharomyces cerevisiae strain on the liberation of volatile thiols in
Sauvignon blanc. AJEV 52:136–9.
Nuñez, M. 2016. Enzymes exogenous to milk in dairy technology: Proteinases. In:
Reference Module in Food Science, ed. G. Smithers, p. 1–8. Elsevier.
Patel, V., Gajera, H., Gupta, A., Manocha, L., and Madamwar, D. 2015. Synthesis of
ethyl caprylate in organic media using Candida rugosa lipase immobilized on exfoli-
ated graphene oxide: Process parameters and reusability studies. Biochem. Eng. J.
95:62–70.
Pereira, G.N., Holz, J.P., Giovannini, P.P., Oliveira, J.V., Oliveira, D., and Lerin, L.A.
2018. Enzymatic esterification for the synthesis of butyl stearate and ethyl stearate.
Biocatal. Agric. Biotechnol. 16:373–7.
Pires, E.J., Teixeira, J.A., Brányik,T., and Vicente, A.A. 2014. Yeast: The soul of beer’s
aroma—A review of flavour-active esters and higher alcohols produced by the brew-
ing yeast. Appl. Microbiol. Biotechnol. 98:1937–49.
Pons, A., Allamy, L., Lavigne, V., Dubourdieu, D., and Darriet, P. 2017. Study of the con-
tribution of massoia lactone to the aroma of Merlot and Cabernet Sauvignon musts
and wines. Food Chem. 232:229–36.
Ravyts, F., Vuyst, L.D., and Leroy, F. 2012. Bacterial diversity and functionalities in food
fermentations. Eng. Life Sci. 12:356–72.
Román, S., Sánchez-Siles, L.M., and Siegrist, M. 2017. The importance of food naturalness
for consumers: Results of a systematic review. Trends Food Sci. Technol. 67:44–57.
306 Food Aroma Evolution

Sales, A., Paulino, B.N., Pastore, G.M., and Bicas, J.L. 2018. Biogeneration of aroma
compounds. Curr. Opin. Food Sci. 19:77–84.
Santos, P., Meireles, M.A., and Martínez, J. 2017. Production of ioamyl acetate by
enzymatic reactions in batch and packed bed reactors with supercritical CO2 . J.
Supercrit. Fluids. 127:71–80.
Santos, P., Zabot, G.L., Meireles, M.A.A., Mazutti, M.A., and Martínez, J. 2016.
Synthesis of eugenyl acetate by enzymatic reactions in supercritical carbon dioxide.
Biochem. Eng. J. 114:1–9.
Schab F.R. and Cain W.S. 1991. Memory for odors. In: The Human Sense of Smell, eds.
D.G. Laing, R.L. Doty, and W. Breipohl, pp. 217–40. Berlin: Springer.
Schoumacker, R., Martin, C., Thomas-Danguin, T., Guichard, E., Le Quéré, J.L., and
Labouré, H. 2017. Fat perception in cottage cheese: The contribution of aroma and
tasting temperature. Food Qual. Prefer. 56:241–6.
Singh, P. and Kumar, S. 2019. Microbial enzyme in food biotechnology. In: Enzymes in
Food Biotechnology—Production, Applications, and Future Prospects, pp. 19–28.
Academic Press.
Singh, T.K., Drake, M.A., and Cadwallader, K.R. 2003. Flavor of cheddar cheese: A
chemical and sensory perspective. CRFSFS. 2:166–89.
Statistics MRC. 2017. Aroma Chemicals—Global Market Outlook (2017–2023).
Available in: www.s​trate​gymrc​.com/​repor​t /aro​ma-ch​emica​ls-ma​rket.​ Accessed in
February 6, 2019.
Styger, G., Prior, B., and Bauer, F.F. 2011. Wine flavor and aroma. J. Ind. Microbiol.
Biotechnol. 38:1145–59.
Sun, W.X., Hu, K., Zhang, J.X., Zhu, X.L., and Tao, Y.S. 2018. Aroma modulation of
Cabernet Gernischt dry red wine by optimal enzyme treatment strategy in wine-
making. Food Chem. 245:1248–56.
Szczesniak, A.S. 2002. Texture is a sensory property. Food Qual. Prefer. 13:215–25.
Tamime, A.Y. 2002. Fermented milks: A historical food with modern applications—A
review. Eur. J. Clin. Nutr. 56:S2–15.
Taylo, A.J. and Linforth, R.S.T. 2003. Direct mass spectrometry of complex volatile and
non-volatile flavour mixtures. Int. J. Mass Spectrom. 223:179–91.
Todea, A., Borza, P., Cimporescu, A., Paul, C., and Peter, F. 2018. Continuous kinetic
resolution of aliphatic and aromatic secondary alcohols by sol-gel entrapped lipases
in packed bed bioreactors. Catal. Today. 306:223–32.
Toldrá, F. and Flores, M. 2000. The use of muscle enzymes as predictors of pork meat
quality. Food Chem. 69:387–95.
Wijk, R.A., Smeets, P.A.M., Polet, I.A., Holthuysen, N.T.E., Zoon, J., and Vingerhoeds,
M.H. 2018. Aroma effects on food choice task behavior and brain responses to
bakery food product cues. Food Qual. Prefer. 68:304–14.
Yadav, G.D. and Thorat, P.A. 2012. Microwave assisted lipase catalyzed synthesis of
isoamyl myristate in solvent-free system. J. Molec. Catal. B Enzym. 83:16–22.
Zare, M., Golmakani, M.T., and Niakousari, M. 2019. Lipase synthesis of isoamyl
acetate using different acyl donors: Comparison of novel esterification techniques.
LWT—Food Sci. Technol. 101:214–9.
Zhou, Y., Peng, Q., Zeng, L., Tang, J., Li, J., Dong, F., and Yang, Z. 2018. Study of
the biochemical formation pathway of aroma compound 1-phenylethanol in tea
(Camellia sinensis (L.) O. Kuntze) flowers and other plants. Food Chem. 258:352–8.
Zouraria, A., Accolasa, J.P., and Desmazeauda, M.J. 1992. Metabolism and biochemical
characteristics of yogurt bacteria: A review. Le Lait. 72:1–34.
 Chapter 16
Novel and Emerging Technologies
(Benefits and Limitations)
Mohammad Hassan Kamani, Hanieh Amani, Amir
Yeganehshakib, and Amin Mousavi Khaneghah

CONTENTS

16.1 Introduction 307

16.2 Novel Thermal Processing Techniques (Definition, Principles, and
Applications) 308
16.2.1 Ohmic Heating 309
16.2.1.1 Advantages and Disadvantages of Ohmic Heating 309
16.2.2 Microwave Heating 310
16.2.2.1 Advantages and Disadvantages of Microwave Heating 313
16.2.3 Infrared (IR) Heating 314
16.2.3.1 Advantages and Disadvantages of IR Heating 315
16.2.4 Radio Frequency (RF) Heating 315
16.2.4.1 Advantages and Disadvantages of RF Heating 316
16.3 Novel Non-Thermal Technologies (Definition, Principles, and Applications) 317
16.3.1 Pulsed Electric Field (PEF) 318
16.3.1.1 Advantages and Disadvantages of PEF 320
16.3.2 High-Pressure (HP) Technology 320
16.3.2.1 Advantages and Disadvantages of HP 324
16.3.3 Ultrasound (US) Technology 324
16.3.3.1 Advantages and Disadvantages of US 327
References 327

16.1 INTRODUCTION

The processing of food in order to preserve perishable foods from spoilage goes back to
prehistoric times. The need to improve food quality gradually increased over time, and
more technologies were developed and introduced to the food industry (Lund, 2003). The
primary goal of all these technologies is to make desirable changes to the food matrix.
These changes involve various reactions such as microbial/enzyme inactivation, protein
coagulation, starch swelling, textural softening, and the formation of aroma compo-
nents. However, each technology has its own limitations and may also cause some unde-
sirable changes concerning nutritional and sensory properties (Fellows, 2009; Sun, 2014).
Sensorial characteristics are well known for their importance to consumer acceptabil-
ity (Sun, 2014). Among sensory parameters, aroma plays a distinct role since its profile

307
308 Food Aroma Evolution

determines the flavor of the food that we consume (Parker et al., 2014). The aroma may
directly or indirectly be affected by processing methods. This effect may be positive or
negative, depending on the type of product or the technique used (Caminiti et al., 2011).
This issue has recently encouraged food researchers to devote efforts to implementing
new techniques to minimize the adverse effect of processing (Czechowska et al., 2005;
Sun, 2014). In this respect, various novel methods have been introduced in recent years
which are generally classified into thermal and non-thermal. The latter has gained more
attention due to its higher performance and lower adverse effects on sensory and nutri-
tional qualities (Pereira and Vicente, 2010; Caminiti et al., 2011; Surowsky et al., 2014).
This chapter aims to provide an overview of the principles, applications, and summarized
impacts of each method on the aroma and flavor of food products. In addition, in order
to give better insight into the merits of each method, the benefits and limitations are
provided.

16.2 NOVEL THERMAL PROCESSING TECHNIQUES

(DEFINITION, PRINCIPLES, AND APPLICATIONS)

Thermal processing is the most commonly used technological method, which provides
the required microbiological safety for food products. This technique relies on the gen-
eration of heat outside the food and its transfer inside by convection and conduction
mechanisms. The conventional examples of this method are baking, roasting, steaming,
blanching, boiling, pasteurization, sterilization, extrusion cooking, and different kinds
of drying techniques (e.g., sun, osmotic, oven, drum, or spray drying) (Richardson, 2001;
Rawson et al., 2011). These methods have some limitations that hinder their application
to a wide range of foods. For instance, they might not be able to adequately transfer
heat to the center of semi-solid or solid foods due to the low thermal diffusivities of the
materials. In addition, lengthy exposures to high-temperature heat may lead to a quality
degradation of the food matrix (Jiao et al., 2018).
The methods mentioned above might also be associated with heat-induced reactions
involving changes of flavor, aromatic compounds, and final sensorial quality of the food
(Richardson, 2001). From a flavor point of view, some of these major reactions are (a)
thermal degradation of lipids; (b) dephosphorylation of proteins; (c) hydrolysis of peptide
bonds; (d) Maillard reactions; and (e) interaction of lipid oxidation and Maillard reac-
tion products (Cadwallader and Singh, 2009; He et al., 2013). Some of these reactions/
impacts are desirable in the processing of foods like bread, cereal, coffee, nuts, malt, and
cooked meat due to generation of the specific desired flavor. However, the same impact,
in particular under severe conditions, is considered to be undesirable concerning organo-
leptic attributes for foods like fruit juices, milk, and dairy products (Richardson, 2001;
Perez and Rouseff, 2008; Cadwallader and Singh, 2009). Table 16.1 shows examples of
the negative impacts of thermal pasteurization on the flavor/aroma quality of some food
products.
To tackle this issue, interest has been growing over the past decade to design thermal
techniques that minimize adverse effects and ensure the retention of sensory and nutri-
tional quality. The prime examples of these technologies include, but are not restricted
to, ohmic, microwave, infrared, and radiofrequency heating. They may be used alone or
combined with increased efficiency. The basic idea residing behind all of these process-
ing methods is the mode of heat transfer in a food matrix, which is discussed in detail in
each respective section of this chapter (Rawson et al., 2011; Jiao et al., 2018). These new
 Novel and Emerging Technologies  309

TABLE 16.1 The Negative Effect of Conventional Thermal Methods on the Flavor/Aroma
of Processed Foods
Processing Effect on Flavor/
Product Conditions Aroma Reference
Reduced-calorie carrot Pasteurization Unacceptable cooked (Sinchaipanit et al.,
juice (65°C for 30 min) flavor 2013)
Mixed fruit juices Mild pasteurization Reduced odor score to (Bukvicki et al.,
(orange/apple blend) (70°C for 0.5–1.5 marginal 2014)
min)
Fruit smoothies (apple, Pasteurization Development of a (Hurtado et al., 2015)
orange, strawberry, (85°C for 7 min) cooked-fruit flavor
and banana)
Longan juice Pasteurization The loss in flavor (Zhang et al., 2010)
(100°C for 1 min) compound
Milk Thermal processing, Generation of various (Cadwallader and
in particular, at high cooked, sulfurous Singh, 2009; Eskin
temperature (UHT) and cabbage-like and Shahidi, 2012)
off-flavor
Passion fruit juice Pasteurization Damaged aroma, (Sandi et al., 2003)
(75°C for 1 min) flavor and color

technologies allow the production of high-quality food products with improved heating
and energy efficiency (Pereira and Vicente, 2010; Kaur and Singh, 2016).

16.2.1 Ohmic Heating

Ohmic heating takes its name from Ohm’s law, which describes the relationship between
current, voltage, and resistance (Icier and Ilicali, 2005). It is also known as electrical
resistance heating, Joule heating, electro-heating, direct electrical resistance heating, and
electro-conductive heating (Rawson et al., 2011; Cullen et al., 2012). This technology is
basically a thermal–electrical method, which is based on the passage of an electrical cur-
rent through a food product that has an electrical resistance (Figure 16.1). In other words,
the electrical energy is converted to heat energy within the food (Icier and Ilicali, 2005;
Cullen et al., 2012). It is an instant heating method and has been suggested to be more uni-
form than other electro-heating techniques (Sarang et al., 2008; Rawson et al., 2011). This
technology can mainly be used for liquids and multiphase liquid–solid mixtures, particu-
larly for media that are difficult to process using conventional heat exchangers (Cullen et
al., 2012). In addition, its usage for solid foods like whole turkey meat, beef, and ham has
also been documented (Pongviratchai and Park, 2007; Zell et al., 2010, 2009, 2012). Thus
far, a large number of potential food applications have been introduced for ohmic heating,
including evaporation, extraction, sterilization, pasteurization, fermentation, blanching,
and dehydration (Knirsch et al., 2010). However, only a few studies evaluated the effect
of this technology on food aroma, flavor, and odor, which are represented in Table 16.2.

16.2.1.1 Advantages and Disadvantages of Ohmic Heating

Ohmic heating has significant advantages over conventional thermal methods. These
advantages include (a) shorter treatment time with minimal heat damage and nutrient
310 Food Aroma Evolution

FIGURE 16.1 Basic scheme of ohmic heating system. (From Cappato et al., 2017.)

loss; (b) high temperature with high‌ uniformity; (c) possibility of use for foods with low
thermal conductivity; (d) suitability for continuous processing without heat transfer sur-
faces; (e) better energy efficiency with low maintenance costs; (f) easier precise control
of temperature compared to conventional heating methods; (g) no residual heat transfer
after shut off of the current; (h) higher quality and fresher tasting; (i) reduced fouling; and
(j) its being an environmentally friendly system (Pereira and Vicente, 2010; Sakr and Liu,
2014; Wen, 2014). On the contrary to the mentioned advantages, some disadvantages
have also been reported, which are (a) higher cost of installation and operating system as
compared to conventional methods; (b) insufficient efficacy for the foods containing fat
globules, which act as a non-conductive substance; (c) requested adjustment according
to the conductivity of the liquid product; (d) narrow frequency band; and (e) complex
coupling between temperature and electrical field distribution (Sakr and Liu, 2014; Kaur
and Singh, 2016).

16.2.2 Microwave Heating

Microwave heating is another thermal processing method that has gained considerable
attention in the food industry in recent years (Vadivambal and Jayas, 2010). Microwaves
are electromagnetic waves within a frequency band of 300 MHz to 300 GHz (Regier et
al., 2017). When microwaves impinge on a dielectric food, part of the energy is reflected,
part is transmitted, and part is absorbed by the material, where it is dissipated as heat.
Heating is due to an increase in molecular kinetic energy and associated molecular fric-
tion (Meda et al., 2017). In other words, this technology is based on the transforma-
tion of alternating electromagnetic field energy into thermal energy by affecting the
polar molecules of the food material (Venkatesh and Raghavan, 2004; Vadivambal and
Jayas, 2010). It has the potential to deliver heat instantly throughout the product due to
volumetric heat generation (Vadivambal and Jayas, 2010). Volumetric heat generation
means that materials can absorb microwave energy internally and convert it into heat.
Thus far, many food applications have been suggested for this useful method including
cooking, baking, pre-heating, dehydration/drying, thawing, sterilization, enzyme inac-
tivation, pasteurization, sterilization, blanching, tempering, and extraction of bioactive
compounds (Venkatesh and Raghavan, 2004; Puligundla et al., 2013; Orsat et al., 2017).
 Novel and Emerging Technologies  311

TABLE 16.2 The Effect of Novel Thermal Technologies (Ohmic, Microwave, Infrared, and
Radio Frequency Heating) on the Flavor/Aroma of Processed Foods
Effect on Aroma,
Technology Processing Conditions Product Flavor, and Odor References
Ohmic 70°C/10 min/ Juice blend No negative (Dima et al.,
heating 17.5–20 V (pumpkin, influence on 2015)
carrot, orange, flavor
celery,
grapefruit)
Ohmic 80°C/50 Hz/10 and 33 Concentrated No significant (Tumpanuvatr
heating V cm−1 juice (orange effect on the and Jittanit,
and pineapple) flavor quality of 2012)
juices
Ohmic 120°C/>20 min/50 Orange juice Improved aroma (Leizerson and
heating Hz/8 kV volatile Shimoni,
concentrations 2005)
Ohmic HTST (6 min) to a Cooked ham Lesser flavor and (Zell et al.,
heating target temperature of aroma scores as 2012)
95°C with a total compared to the
residence time of 8 conventional
min; LTLT (5 min method
ohmic heating to 70°C,
8 min holding)/0–250
V, 50 Hz
Microwave Microwave–hot air Garlic cloves Higher retention (Sharma and
heating drying of the volatile Prasad,
(40–70°C/40 W) components 2001)
responsible for
flavor strength
Microwave Microwave roasting Cumin seed Better retention of (Behera et al.,
heating (730 W/10 min) characteristic 2004)
flavor compounds
like total
aldehydes in
microwave-
roasted samples
Microwave Pasteurization of milk Milk No adverse effect (Valero et al.,
heating (80–92°C/15 s/532 W) on flavor 2000)
Microwave Sterilization (915 Skim milk Improved flavor (Clare et al.,
heating MHz/60 kW) quality 2005)
Microwave Blanching (128°C/11 Peanut The occurrence of (Schirack et
heating min/5 kW, 915 MHz) off-flavors (stale/ al., 2006)
floral and ashy
off‐flavors)
observed
(Continued )
312 Food Aroma Evolution

TABLE 16.2 (Continued) The Effect of Novel Thermal Technologies (Ohmic, Microwave,
Infrared, and Radio Frequency Heating) on the Flavor/Aroma of Processed Foods
Effect on Aroma,
Technology Processing Conditions Product Flavor, and Odor References
Microwave Drying in a three-step Black tea Higher sweet (Qu et al.,
heating process: (a) heating for aroma; Promoted 2019)
6 min (Leaf content of total
temperature 91°C), catechins and
followed by 1 h theaflavins;
cooling; (b) heating for helpful to
4 min leaf temperature maintenance
106°C), followed by polyphenols and
30 min cooling; and (c) volatile
heated for 2 min (leaf compounds
temperature
115°C)/700 W
IR Roasting (160 to Sesame seed Good flavor at (Kumar et al.,
220°C/6 kW/1.1 to 1.3 160 to 200°C/ 2009)
μm wavelength of Detection of
radiation) burned flavor
above 200°C
IR Processing of leaves by Green tea leaves Enhanced aroma (Park et al.,
Far‐Infrared (FIR) of leaves in all 2009)
250°C/15 to 25 min samples
and additional 300°C
for 5 min/6.1 W/cm 2/
4.14 µm
IR Roasting (130– Almond No negative (Yang et al.,
150°C/5000 W/m2) influence on 2010)
flavor
IR Thawing by Far-IR Frozen chestnut Prevent the (Hee, 2006)
(170–230°C for 30–60 formation of
min) particular flavors
and aromas
RF Heating (112°C/13.5 White mustard Removal of spicy (Ildikó et al.,
MHz) flavor (pungent) 2006)
by inactivation of
the myrosinase
enzyme
RF Pre-heating for pest Orange Significant change (Birla et al.,
control (48–52°C/0–20 in volatile flavor 2005)
min/12 kW) profiles
RF Sterilization (up to Macaroni and No significant (Wang et al.,
135°C/27 MHz 6/kW) cheese adverse effect on 2003)
flavor quality
RF Post-drying (8–9 Partially baked Slightly uncooked (Koray
min/27.12 MHz/2 kW) cookies flavor observed Palazoğlu et
al., 2012)
(Continued )
 Novel and Emerging Technologies  313

TABLE 16.2 (Continued) The Effect of Novel Thermal Technologies (Ohmic, Microwave,
Infrared, and Radio Frequency Heating) on the Flavor/Aroma of Processed Foods
Effect on Aroma,
Technology Processing Conditions Product Flavor, and Odor References
RF Sterilization (6 kW, Vacuum- No significant (Liu et al.,
27.12 MHz/20 min) packaged effect on the 2015)
Caixin flavor quality
(Brassica
campestris L.)
RF Cooking (73°C for 40 Leg and No significant (Zhang et al.,
min) shoulder hams change in the 2006)
flavor quality but
distinguished
overall
acceptability
observed
RF Pasteurization (75– Vacuum-packed No significant (Orsat et al.,
85°C/10 min/27.12 ham change in the 2004)
MHz /600 W) odor

In conventional methods, energy is transferred due to thermal gradients, but microwave

heating is the conversion transfer of electromagnetic energy to thermal energy through
direct interaction of the incident radiation with the molecules of the target material.
The difference in the way energy is delivered can result in many potential advantages
(Venkatesh and Raghavan, 2004). For instance, in the microwave, heat is generated
throughout the material and leads to a faster heating rate as compared to conventional
cooking. This may result in the prevention of severe damage to the quality attributes, such
as color, flavor, and nutrients of the product (Vadivambal and Jayas, 2010). In addition,
microwaves penetrate a food product and do not act at the surface level. This conversion
of energy into heat throughout the product is more efficient and results in a better final
quality in processed food (Contreras et al., 2017). The output of various studies regarding
the quality improvement of flavor/aroma is presented in Table 16.2. Due to these major
advantages, microwave technology has become very popular for both the industrial and
domestic sectors.

16.2.2.1 Advantages and Disadvantages of Microwave Heating

Apart from the rapidness of the microwave method, there are other quantitative and
qualitative advantages for this heating method, which are (a) shorter start-up time; (b)
greater energy efficiency; (c) precise equipment control; (d) space saving; (e) ease of opera-
tion; (f) lower maintenance; (g) lower noise levels; (h) improved nutritional quality of
processed food; and (i) better retention of taste and color qualities (Richardson, 2001;
Vadivambal and Jayas, 2010; Contreras et al., 2017; Meda et al., 2017; Yolacaner et al.,
2017). Moreover, microwave heating is very advantageous when it is used for the extrac-
tion of bioactive compounds. Reduced nutrient losses and extraction time, improved
extraction yield, rapid and volumetric heating of the absorbing medium, low solvent
consumption, and higher selectivity of target molecules are the examples of benefits of
this method when it is used for extraction purposes (Orsat et al., 2017).
Although microwave technology has considerable advantages for the processing
of food, serious drawbacks have been observed. A major disadvantage is non-uniform
314 Food Aroma Evolution

temperature distribution, which leads to the occurrence of cold and hot spots in the
heated food. This may result in poor quality in the end product (Vadivambal and Jayas,
2010; Dorantes-Alvarez et al., 2017). This issue may also lead to the incomplete destruc-
tion of microbes, which may cause a microbial safety issue in the cold spots (Vadivambal
and Jayas, 2010). In addition, non-uniform microwave heating may cause tough and
leathery crumbs of bakery products due to the occurrence of different patterns of starch
transformation (Yolacaner et al., 2017).
Another weakness of microwave technology is the short baking time when used for
bakery products. Bakery products generally need a long time to complete their physi-
cochemical reactions, while microwave heating has a short overall period, and there-
fore the final quality of microwave-baked products is not desirable. Rapidly generated
gas and steam, insufficient gelatinization, and changes in gluten structure have been
reported for microwave-baked bread as well (Yolacaner et al., 2017). Another issue with
microwave heating is the large number of factors that affect the microwave heat trans-
fer behavior such as the thickness, the geometry, and the dielectric properties of the
food (Vadivambal and Jayas, 2010). The high penetration power of microwave energy
affects heat transfer behavior and may cause overheating of products leading to scorch-
ing, depending on the nature and geometry of the material, dielectric properties, and
oven design (Ekezie et al., 2017). Flavors generated as a result of browning reactions
do not exist in microwave-baked products. During microwave baking, individual fla-
vor components are subjected to losses through distillation, flavor binding by starches
and proteins, and chemical degradation. Many of the nutty, brown, and caramel-type
aromas observed in conventional cakes were lacking in microwave-baked cakes. Also,
during the microwave baking of cakes, undesirable flavors such as flour or egg-like fla-
vors develop. In order to overcome these issues in bakery products, different solutions,
including the use of flavoring agents and the combination of microwave heating with
other methods like hot air, infrared heating, or steam have been proposed (Yolacaner
et al., 2017).

16.2.3 Infrared (IR) Heating

Infrared radiation is a type of energy in the form of an electromagnetic spectrum, which

arises from the movement of electrons in the atoms and molecules inside the exposed
food matrix (Cullen et al., 2012; Rastogi, 2012). IR radiation can fall into three regions
depending on its wavelength. These regions are (a) the near-infrared (NIR; 0.78–1.4
µm); (b) middle-infrared (MIR; 1.4–3 µm); and (c) far-infrared (FIR; 3–1,000 µm)
(Krishnamurthy et al., 2008; Riadh et al., 2015). IR heating has been found to be more
effective when compared to a conventional heating method, and therefore numerous pro-
cessing applications have been suggested for this alternative method. The primary appli-
cations of IR include blanching (e.g., carrots and leaves), thawing (e.g., frozen tuna or
frozen potato purée), drying (e.g., pasta, vegetables, fish, and fruit), heating (e.g., flour),
frying (e.g., meat), roasting (e.g., cereals, coffee, cocoa, chestnuts, hazelnuts, and sesame
seeds), and baking (e.g., pizza, biscuits, and bread) (Richardson, 2001; Hebbar et al.,
2004; Cullen et al., 2012; Rastogi, 2012). IR can also be used for pasteurization and ster-
ilization by inactivating pathogens in foods as well as packaging materials (Richardson,
2001; Krishnamurthy et al., 2008). It can inactivate microorganisms by damaging intra-
cellular components such as DNA, RNA, ribosomes, cell envelopes, and/or proteins in
the cell (Cullen et al., 2012).
 Novel and Emerging Technologies  315

16.2.3.1 Advantages and Disadvantages of IR Heating

IR radiation is generally considered an ideal source of energy for the heating processes
of foods due to its basic characteristics such as high heat transfer capacity, direct heat
penetration into the food product, fast regulation response, and better possibilities for
process control (Richardson, 2001; Cullen et al., 2012; Riadh et al., 2015). Food heated
using an IR treatment can be rapidly cooled, since the infrared radiation mainly heats a
thin layer of the surface, and consequently provides a less negative change in the quality
of the final product (Cullen et al., 2012). In recent years, IR technology has gained signifi-
cant superiority over traditional thermal methods owing to its remarkable benefits. Some
of these major advantages are listed in Table 16.3 (Krishnamurthy et al., 2008; Cullen et
al., 2012; Riadh et al., 2015). Moreover, IR energy in the form of FIR is easily absorbed
by the main components of foods, that is, organic materials and water. This greatly
helps in efficiently pasteurizing food materials since water readily absorbs radiation in
the IR region and this results in rapid temperature increase (Cullen et al., 2012). Using IR
technology for dehydration purposes is also very beneficial. IR heating allows for a high
rate of water evaporation without quality losses such as surface hardening, deformation/
shrinkage, and better quality characteristics concerning the loss of ascorbic acid, color,
and aroma of dried foods (Pan et al., 2008; Riadh et al., 2015). Although IR provides
many benefits for food processing, it has a few disadvantages. The major drawback of IR
technology is its low penetration power. Therefore, it cannot deeply penetrate and con-
tribute to the heating of high-volume samples. Due to this limitation, IR heating is mostly
used where a surface-heating application is required (Cullen et al., 2012). The other limi-
tations of the IR method are provided in Table 16.3 (Krishnamurthy et al., 2008).

16.2.4 Radio Frequency (RF) Heating

Among all electro-technologies, radio frequency is considered a unique technique for

food processing (Piyasena et al., 2003; Jiao et al., 2018). RF heating (also called capaci-
tative dielectric heating) involves the transfer of electromagnetic energy directly into the
product, which results in frictional interaction between molecules (Piyasena et al., 2003).

TABLE 16.3 The Major Advantages and Disadvantages of Infrared Heating

Technology
Advantages Disadvantages
Reduced heating time Fracturing the food materials due to
prolonged exposure
Uniform heating Low penetration power
Equipment compactness
The absence of solute migration in food
material
The lesser quality loss during processing
Preservation of vitamins
Higher energy efficiency and saving space
Intermittent energy source
Cleaner operational environment
Lower operational costs and versatility
316 Food Aroma Evolution

In other words, in this technique, heat is generated within the food materials by molecu-
lar friction using a high frequency alternating electrical energy (Awuah et al., 2014). The
principles of RF heating are very similar to microwave heating. However, the main dif-
ference between these two is the wavelength. The wavelength of RF is designated to be
22 to 360 times as great as microwave frequencies, which allows RF energy to penetrate
dielectric materials more deeply than microwaves (Wang et al., 2003). For this reason,
RF heating is known as a high-frequency dielectric heating system (Piyasena et al., 2003).
The RF band of the electromagnetic spectrum covers a broad range of high frequencies,
typically either in the kHz range (3 kHz < f ≤ 1 MHz) or MHz range (1 MHz < f ≤ 300
MHz) (Piyasena et al., 2003; Liu et al., 2015).
Owing to the great potential of RF technology, it has been successfully applied in var-
ious types of processing in the food sector. These applications include drying (e.g., coffee
beans, cocoa beans, corn, grains, and nuts), thawing (e.g., frozen meat, fishery product,
egg, fruit, and vegetables), post-baking and heating (e.g., cookies, snack foods, bread, and
biscuits), blanching/enzyme inactivation (e.g., a variety of vegetables and fruits), roast-
ing (e.g., salted peanuts and cocoa beans), sterilization (e.g., packaged flours, almond,
spices, and peanut butter crackers), pasteurization (e.g., meat and meat ­products), and
­controlling mold growth (Piyasena et al., 2003; Jiao et al., 2018). Moreover, RF heating
can be applied to efficiently disinfect dry food such as legumes, nuts, stored wheat, milled
rice, and dried fruit. It can selectively heat and kill any pests without damaging the agri-
cultural products (Jiao et al., 2018).

16.2.4.1 Advantages and Disadvantages of RF Heating

As previously discussed, in the RF method, the heat is generated due to the friction of the
rotational movement of the molecules and the space charge displacement in response to
an alternating electric field. This offers considerable advantages over other thermal tech-
niques for food processing (Liu et al., 2015). The general advantages of the RF method
are (a) shorter processing times (which reduces heating time and improves the food qual-
ity significantly); (b) large penetration depth; (c) better bacteriological and sensory qual-
ity during the inactivation of the enzyme and microorganism; (d) reduced water usage
and the capital cost of the equipment; (e) lower energy consumption and saving energy; (f)
reduced cost of waste management; and (g) its being environmentally friendly (Piyasena
et al., 2003; Jiao et al., 2018). Concerning the effect of RF on the flavor of processed
foods, RF showed negligible effects on the alteration of aroma/flavor in most cases of
RF-treated foods (Table 16.2).
RF for the tempering/thawing of frozen foods (e.g., vegetable, frozen fish, or meat)
has superiority when compared to traditional methods with respect to the final quality
of heated food. The traditional methods are very slow and lengthy procedures, which
may lead to significant drip loss, bacteria growth, and finally degradation of quality.
Commercial RF thawing is much faster and can prevent the aforementioned quality
losses. RF reduces food waste and produces quality food with high sensory acceptability
(Jiao et al., 2018). RF energy also has better potential for drying processes compared
to conventional hot-air machines. The conventional methods cannot efficiently remove
the moisture from solid or semi-solid foods during the falling rate period. In addition, it
may cause cracks or hard shells in the dried products. The RF method could successfully
overcome these quality issues using its unique selective and volumetric heating proper-
ties. During the RF drying of moist materials, the water molecules/or wet areas tend to
absorb more energy than the solids because of the differences in their dielectric proper-
ties, and therefore the quality of food can be protected from overheating by evaporating
 Novel and Emerging Technologies  317

only the water (Jiao et al., 2018). Another advantage of RF technology is its application in
low-moisture foods. Low-moisture foods have low thermal diffusivity, and conventional
methods for the bulk heating of these materials are slow and inefficient. In addition, low
water activity makes some pathogens like Salmonella heat tolerant against thermal inac-
tivation. This is considered a technological challenge for controlling pathogens in these
types of foods, which can be solved by RF. RF has unique penetration depth and volu-
metric heating properties, which can be an alternative use for the pathogen inactivation
of low-moisture foods (Jiao et al., 2018).
RF not only has advantages over traditional methods, but it has some specific superi-
ority when it is compared with other novel thermal technologies such as microwave and
ohmic heating. For instance, in contrast to ohmic heating, there is no need for electrodes
to be in contact with the food. Moreover, RF can be applied to both solid and liquid
foods. RF has greater wavelength than microwave frequency, and therefore it would be
more effective in the penetration of treated foods. Furthermore, RF is particularly suit-
able for large industrial applications, owing to its simpler heating system construction as
well as a more straightforward application for continuous processes (Awuah et al., 2014).
Although RF heating has proven its effectiveness in the food industry and research
laboratories, it has some potential shortcomings, which hinder this technology’s use in
the industrial sector. In packaged foods, thermal runaway heating (or hot spot) and arc-
ing (avoiding dielectric breakdown) may occur, which cause packaging failure and prod-
uct destruction (Awuah et al., 2014). Arcing/flashing usually happens with either a too
high-voltage field or the presence of excessive humidity between electrodes (Jiao et al.,
2018). Thermal runaway, the other potential problem, may lead to non-uniformity in the
RF heating system, which may cause insect survival as well as product damage after pro-
cessing (Awuah et al., 2014; Hou et al., 2016). Moreover, some packaging materials may
fail in the RF field, or damage may occur. In this respect, packaging damage may also
occur (Awuah et al., 2014). Another issue which was documented for RF heating is the
thawing of meat products. RF thawing is limited to regular shaped fish and/or meat, and
irregular shaped food cannot be thawed by RF (Jiao et al., 2018). Concerning the cost
of the technology, RF is more expensive than other techniques such as radiation, steam,
or ohmic technology for an equivalent power output (Awuah et al., 2014). Also, more
training courses for the operator may be required to properly handle and troubleshoot
the RF system. These experiences may also be regarded as an additional cost when RF
technology is implemented (Jiao et al., 2018).

16.3 NOVEL NON-THERMAL TECHNOLOGIES

(DEFINITION, PRINCIPLES, AND APPLICATIONS)

Thermal technologies are considered to be an effective method for food processing; how-
ever, due to incidents of overheating, they might be associated with a loss of nutrients
and undesirable sensory quality (Stoica et al., 2013; Ortega-Rivas and Salmerón-Ochoa,
2014). On the other hand, there is a growing demand for high-quality nutritious foods
with “fresh-like” characteristics as well as improved retention of functionalities (Rawson
et al., 2011; Ortega-Rivas and Salmerón-Ochoa, 2014). Therefore, food technologists
have been pursuing improved processing methods that have minimal negative effects on
the nutritional, organoleptic attributes (texture, taste, and aroma) and are free of any
health risks (Rawson et al., 2011; Ortega-Rivas and Salmerón-Ochoa, 2014). To develop
such methods, a reduction in processing temperature is required, since thermal treatment
318 Food Aroma Evolution

is usually responsible for the loss of desired sensory properties and causes various dam-
ages to vitamins, bioactive compounds, and other vital nutrients (Cullen et al., 2012;
Stoica et al., 2013). Therefore, during the last decade, the tendency toward the develop-
ment of novel mild- or non-thermal technology has grown in order to respond to this
demand and replace the heating method with emerging techniques such as pressure, light
pulses, and so on (Pereira and Vicente, 2010; Rawson et al., 2011; Cullen et al., 2012).
The term “non-thermal processing” refers to technologies that are effective at ambi-
ent or sublethal temperatures (Pereira and Vicente, 2010; Cullen et al., 2012). A non-
thermal technique must have at least two criteria to be considered an alternative valid
processing method for thermal processing. The first criterion is equal efficiency to thermal
methods with respect to microorganism/enzyme inactivation. In other words, we need to
know whether the final food product subjected to this novel method is chemically and
microbiologically safe. For instance, thermal technologies can logarithmically decrease
microorganisms during heating. It is expected that the alternative non-thermal method
inactivate the cell population of microorganisms in the same manner. The second valid-
ity aspect is the feasibility of preserving better overall quality attributes than thermal
methods in treated foods (Ortega-Rivas and Salmerón-Ochoa, 2014). Apart from these
criteria, the alternative approach must be economical and have no adverse environmental
impacts (Cullen et al., 2012).
Similar to thermal technologies, non-thermal techniques may also affect sensory
attributes, in particular, in olfactory parameters (flavor, aroma/odor). These param-
eters may be affected by biochemical degradation/composition or their interactions with
other chemical components during processing, and produce a certain kind of flavor/
aroma. For this reason, flavor scientists have recently been focusing on evaluating the
effects of non-thermal techniques on olfactory changes (Ortega-Rivas and Salmerón-
Ochoa, 2014). The prime examples of these current technologies are pulsed electric field,
high-pressure technology, and ultrasound technology. This section is an overview of
these three technologies, their principles, and their applications, as well as their impact
on food aroma/flavor.

16.3.1 Pulsed Electric Field (PEF)

The pulsed electric field is a preservative technology that has drawn considerable atten-
tion as a new non-thermal processing method in various food categories (Cserhalmi et
al., 2006). PEF is an electroporation process that can prolong the shelf life of foods with
minimal detrimental influence on food quality and freshness (Cserhalmi et al., 2006;
Lynch, 2016; Syed et al., 2017). It consists of the delivery of the electrical field in the
form of high-voltage pulses to the sample over a short time period (a few million micro-
seconds) (Soliva-Fortuny et al., 2009). The process depends on the number of delivered
pulses to the food, which is located between two electrodes (Syed et al., 2017). Several
hypotheses have been proposed to explain the rupture mechanism of the cell membrane
in PEF-treated foods. Among them, two hypotheses are generally accepted, which are (a)
electrical breakdown and (b) osmotic imbalances (Buckow et al., 2013). When the food
is affected by the field, its biological cell membranes develop pores. Depending on the
treatment conditions and intensity of the pulses, the pores may be formed in a permanent
or a temporary manner. Later, these formed pores increase membrane permeability and
consequently result in the breakdown of the cell wall, the intrusion of surrounding media,
and finally the loss of cell content (Knorr et al., 2011; Sun, 2014; Lynch, 2016). PEF can
 Novel and Emerging Technologies  319

FIGURE 16.2 Formation of pores in the membrane of a cell after PEF treatment. (Adapted
from Sun, 2014, with slight changes.)

be performed at various temperatures like ambient, sub-, or above-ambient temperatures

(Syed et al., 2017). Figure 16.2 demonstrates the initial stage of pore formation in the cell
membrane after PEF treatment.
PEF is primarily used for two purposes: (a) inactivation of the microbial cell by caus-
ing irreversible loss of cell membrane functionality and (b) decreasing the activity of
various enzymes to improve the quality of the targeted food (Cserhalmi et al., 2006).
PEF is mainly used for the processing of liquid and semi-liquid foods, since the electrical
current can flow more efficiently into liquid media (Stoica et al., 2013; Syed et al., 2017).
However, a number of studies have also been carried out to show the applicability of
the PEF technique in solid food materials (Lynch, 2016). Among all food PEF applica-
tions, microbial control/inactivation has gained a special preference (Syed et al., 2017).
Several studies indicated that PEF enables the inactivation of a wide range of pathogenic
microorganisms (e.g., C. jejuni, E. coli, L. monocytogenes, S. typhimurium, S. enteridi-
tis, Bacillus cereus, Y. enterocolitica, S. aureus, and S. dublin) in various liquid foods
such as juices, liquid eggs, soup, milk, and dairy products (Soliva-Fortuny et al., 2009;
Lynch, 2016; Syed et al., 2017). This inactivation/pasteurization is usually associated
with minimal changes in nutritional and sensory quality (De Silva et al., 2018). PEF
has also showed its capability in the substantial inactivation of many indigenous food
enzymes such as pectin methyl ester, alkaline, phosphatase, polyphenol oxidase, plasmin,
and lipase in processed foods (Buckow et al., 2013; Lynch, 2016). However, this inactiva-
tion mechanism is not yet well understood (Buckow et al., 2013). In addition, it has been
proven that PEF is a useful technique to enhance the efficiency in extracting intracellular
chemical compounds (De Silva et al., 2018). This can be very useful for the extraction of
bioactive components from plant cells (Syed et al., 2017).
With regard to the application of PEF in solid foods, a few reports have demonstrated
that PEF can not only be used for the inactivation of foodborne pathogens on the food
surface but can also increase the yield of juice from raw vegetables/fruits like apple and
sugar beets (Stoica et al., 2013; Lynch, 2016; De Silva et al., 2018). Recently, there has
been a growing interest in the effective application of PEF on food constituents such as
partial modification/alteration of proteins, starch granules, and fatty acid composition
(Knorr et al., 2011). Apart from the aforementioned applications, PEF can be applied as
320 Food Aroma Evolution

a pre- or main treatment for a deep frying process, wastewater, dehydration/drying, bio-
production of certain compounds, freezing/thawing, and foods with low electrical con-
ductivity like less viscous juices (Soliva-Fortuny et al., 2009; Ignat et al., 2015; Wiktor et
al., 2015; Syed et al., 2017; De Silva et al., 2018).

16.3.1.1 Advantages and Disadvantages of PEF

PEF is seen as a key treatment that is more efficient than conventional thermal meth-
ods, owing to its several benefits (Stoica et al., 2013). Although PEF minimizes heating
effects during processing, it has a comparable efficiency with thermal methods in terms of
pathogenic safety (Lynch, 2016; De Silva et al., 2018). Furthermore, the low temperature
used in PEF substantially preserves the nutritional value and provides a better retention
of sensory attributes (fresh taste and flavor, texture, aroma, and color) (Soliva-Fortuny
et al., 2009; Lynch, 2016; Syed et al., 2017). Table 16.4 shows that PEF can improve the
aroma profile and flavor quality of most targeted foods during processing. In addition,
PEF is an eco-friendly technology, has short processing times, and economically improves
the energy usage (Soliva-Fortuny et al., 2009; Knorr et al., 2011; Stoica et al., 2013; Syed
et al., 2017; De Silva et al., 2018). Hence, considering these great advantages and their
successful applications, PEF can be regarded as a popular non-thermal technique for the
processing and preservation of various foods.
On the contrary to the benefits mentioned above, there are some limitations which
hinder PEF from being a perfect technology. PEF is not able to kill bacterial spores during
treatment, which may cause spoilage issues for some foods like milk (Stoica et al., 2013;
Syed et al., 2017). In addition, designing a treatment chamber for PEF-assisted pasteuriza-
tion is a hitch, since each product has its own specific viscosity, electrical conductivity, and
properties, and therefore appropriate optimization is required when it is implemented on an
industrial scale (Knorr et al., 2011). Adaptation of the electrical pulse shape and duration is
also another important matter for PEF, since higher electric field intensities may cause some
electrochemical reactions, which may lead to corrosion of the electrode materials and par-
tial electrolysis of the liquid sample (Knorr et al., 2011). With regard to the liquid sample,
it has been found that the food containing air bubbles cannot be well treated by PEF. Also,
the particle size of the sample should be less than the gap of the treatment region to ensure
an appropriate process (Stoica et al., 2013; Syed et al., 2017). The high initial cost of PEF
systems is another big issue, which has contributed to the resistance of the food industry to
purchasing this technology (Stoica et al., 2013). Another challenge of PEF is marketability
and consumer acceptability of PEF-processed food products. PEF-processed foods have
limited acceptance when compared to thermal products; since the nature of this technol-
ogy has not yet been fully explained to consumers, and clear communication is of prime
importance to them. Furthermore, PEF-treated foods are labeled as “minimally processed
foods,” which induces an adverse image in a consumer’s mind regarding food that is not
well-processed (Syed et al., 2017). The technical and economic drawbacks of PEF have
therefore led to its reduced application when compared to thermal methods.

16.3.2 High-Pressure (HP) Technology

High pressure, sometimes also known as high hydrostatic pressure, is an emerging method,
where food is subjected to elevated pressures, with or without the addition of heat (Cullen
et al., 2012; Ortega-Rivas and Salmerón-Ochoa, 2014). During HP processing, the food
material undergoes physical compression at different pressure levels (ranging from several
 Novel and Emerging Technologies  321

TABLE 16.4 The Effect of Pulse Electric Field (PEF) and High Pressure (HP) on Flavor/
Aroma of Processed Foods
Effect on Aroma, Flavor,
Technology Processing Conditions Product and Odor References
PEF Pasteurization Orange Retained five representative (Yeom
(60.1°C for 9 s/35 kV/ juice flavor compounds et al.,
cm/35 psi) (R-pinene, myrcene, 2000)
octanal, limonene, and
decanal)
PEF Evaluation of retention Strawberry Improved flavor stability of (Aguiló-
of volatile compounds juice the juice Aguayo
contributing to aroma et al.,
profile (35°C–100 2009)
Hz/35 kV/cm for
1700 μs)
PEF Sterilization Litchi juice Better retention of aroma (Li et al.,
(100 Hz/35 profile as compared to 2009)
kV.cm−1/200/μs/18°C) thermal sterilization
PEF Pasteurization Apple Improved retention of (Azhu
(23 kV.cm−1/150 Cider apple cider aroma Valappil
μs/48°C) volatiles and higher et al.,
aroma sensory score 2009)
PEF Pasteurization Orange Improved content of flavor (Min et al.,
(45°C/40 kV/cm/97 ms) juice components (limonene, 2003)
myrcene, decanal, octanal,
and α-pinene)/higher
flavor preference than a
heated sample
PEF Pasteurization(28 kV/ Squeezed No significant change on (Cserhalmi
cm with 50 pulses/less citrus the volatile aroma et al.,
than 34°C) juices components 2006)
PEF Bacterial inactivation Beer Significant loss in flavor/ (Evrendilek
(23 psi/41 kV/ aroma acceptability et al.,
cm/175 μs) 2004)
PEF Pasteurization (35 kV/ Apple juice Improved the retention (Aguilar-
cm/1200 pulses per ofvolatile compounds Rosas
second (pps)/4 μs) responsible for flavor in et al.,
PEF-treated juice 2007)
PEF Pasteurization (90 s/32 Longan Higher content of flavor (Zhang
kV/cm/below juice compounds in PEF-treated et al.,
40°C/10 Hz) juice as compared to a 2010)
thermally treated sample
PEF 40 kV/cm for 57 μs Tomato Preferred flavor score and (Min and
at 45°C juice more retention of the Zhang,
flavor compounds as 2003)
compared to thermally
processed juice
(Continued )
322 Food Aroma Evolution

TABLE 16.4 (Continued) The Effect of Pulse Electric Field (PEF) and High Pressure (HP)
on Flavor/Aroma of Processed Foods
Effect on Aroma, Flavor,
Technology Processing Conditions Product and Odor References
HP Evaluation of enzymatic Strawberry Maintained aromatic (Navarro
activity (400 purée compounds distribution et al.,
MPa/20°C/20 min) 2002)
HP Evaluation of flavor Strawberry The highest stability (Zabetakis
compounds (200 to observed at 400 MPa for et al.,
800 MPa for 15 min acidic flavor compounds, 2000)
at 18–22°C) at 800 MPa for alcohol
compounds, at 200 and
800 MPa for ketone
compounds
HP Sterilization (25°C, 600 Guava Maintained the original (Yen and
MPa, 15 min) juice volatile flavor components Lin, 1999)
HP Quality assessment of Strawberry Improved volatile flavor (Kimura
jam (400–500 MPa, jam components et al.,
10–30 min) 1994)
HP Quality evaluation of Yogurt Highest flavor score (De Ancos
yogurt (100–400 observed at 200 and 300 et al.,
MPa/15 min/at 20°C) MPa 2000)
HP Pasteurization (600 Australian The odor and flavor of (Baxter et
MPa for 60 navel treated juices were al., 2005)
s/18–20°C) orange acceptable and comparable
juices with heated samples
HP 400 and 600 MPa for Beef and No severe changes in (Schindler
15 min at 5°C Chicken aroma properties et al.,
2010)
HP 200–400 Mpa—10 min Alligator No effect on the flavor (Canto
at 20°C meat et al.,
2012)
HP Atmospheric pressures Cherry The loss of intensity in fresh (Viljanen et
and 800 MPa/20– tomato tomato odor at 800 al., 2011)
60°C/10 min purée MPa(20 and 60°C)/
Increased of cooked
tomato odor and tea aroma
HP 400 and 500 MPa for Melon No significant change in (Ma et al.,
20 min/22°C juice flavor 2010)

tens of MPa to several hundreds of MPa, depending on the type of HP unit used) in HP
pasteurization units (Knorr et al., 2011; Cullen et al., 2012). Process temperatures during
HP may range from below zero (to minimize any effects of adiabatic heat) to above 100°C
with various exposure times varying from a millisecond pulse to over 20 minutes. (Pereira
and Vicente, 2010). The main goals of this method are (a) to inactive pathogenic microor-
ganisms at low temperatures; and/or (b) to alter the food attributes in order to produce a
desired-quality food product (Ortega-Rivas and Salmerón-Ochoa, 2014).
During the HP process, food is subjected to isostatic pressure (Cullen et al.,
2012). The sample is first loaded into a high-pressure chamber which is filled with a
 Novel and Emerging Technologies  323

FIGURE 16.3 Scheme of a high-pressure processing unit. (From Bolumar et al., 2015.)

pressure-transmitting media (fluid). The fluid in the chamber is pressurized with a pump,
and this pressure is uniformly and instantaneously transmitted through into the food
(Pereira and Vicente, 2010; Ortega-Rivas and Salmerón-Ochoa, 2014). Owing to the
isostatic principle of pressure transmission in the HP method, all molecules are subjected
to the same amount of pressure at the same time (Knorr et al., 2011). Figure 16.3 depicts
the operation scheme of a HP of food sample during processing.
HP has been shown to be a promising method in several food processes, particularly
in food preservation, with plenty of valuable potential applications. Currently, the largest
commercial application of HP is shelf-life extension via the inactivation of various micro-
organism groups (e.g., viruses, bacteria, yeasts, and molds) (Knorr et al., 2011; Cullen
et al., 2012; Ortega-Rivas and Salmerón-Ochoa, 2014). Microorganism inactivation by
HP occurs by interrupting the cellular functions responsible for the reproduction, survival,
and damage of the membrane proteins (Torres and Velazquez, 2005). The efficacy of this
method has been reported for common microorganisms like S. typhimurium, L. mesen-
teroide, Listeria, and Vibrio vulnificus (Torres and Velazquez, 2005; Knorr et al., 2011).
Apart from microbial inactivation, there are several other interesting applications for HP
that are related to food structure engineering (Knorr et al., 2011). These applications are
(a) improvement of the methods for extraction and isolation of bioactive components (by
increasing yield, reducing time, and improving mass transfer rate); (b) inactivation of vari-
ous enzymes in order to modify the texture of food; (c) improvement of the final yield by
reducing weight loss in various meat and dairy products; (d) improvement of freezing and
thawing in several types of frozen fish products (by acceleration of the freezing process,
decreasing drip loss, and thawing time); (e) stabilization of suspensions and emulsions
by homogenization of the dispersion system; and (f) reduction of oil uptake during fry-
ing and decreasing frying time (Pereira and Vicente, 2010; Knorr et al., 2011; Rawson et
al., 2011; Sun, 2014). HP is also able to rupture cell walls, open the tissue structure, and
increase the permeability of cell membranes. This capability is useful for increasing the
subsequent drying and osmotic dehydration rate of certain food products (Sun, 2014). HP
can affect the native structure of large molecules such as protein and starch (Pereira and
Vicente, 2010; Knorr et al., 2011). This influence can be utilized for food texture engineer-
ing, modifications of food constituents, and physicochemical properties (Sun, 2014). The
prime examples of these modifications are (a) improvement of the coagulating properties
of milk; (b) improvement of the gelation ability in egg, soy, and meat proteins; (c) dena-
turation and alteration of the functionality of proteins such as improving gelatinization
324 Food Aroma Evolution

and digestibility for desired functionality; and (d) gelatinization and swelling of different
starch granules (Pereira and Vicente, 2010; Knorr et al., 2011; Sun, 2014).

16.3.2.1 Advantages and Disadvantages of HP

HP is considered to be a superior method compared to other thermal techniques. The
reason behind this claim is the elimination of heat and its adverse degradation effects
during microorganism/enzyme inactivation process (Pereira and Vicente, 2010; Knorr
et al., 2011; Ortega-Rivas and Salmerón-Ochoa, 2014). In addition, during the HP pro-
cess, the primary structure of low-molecular weight molecules such as peptides, lipids,
vitamins, flavor and aroma components, and saccharides is rarely affected. Therefore,
the minimum change occurs concerning aroma, taste, texture, and nutritional quality (in
particular vitamins) in HP-treated food (Pereira and Vicente, 2010; Knorr et al., 2011;
Cullen et al., 2012; Ortega-Rivas and Salmerón-Ochoa, 2014). Table 16.4 summarizes
the output of research into the desirable effects of HP on the flavor quality and aroma
compounds of food products. HP is independent of the size and geometry of the sample,
and the shape of food can be retained, even at extreme pressures due to uniform pressure
transmission in all directions throughout the food products (Pereira and Vicente, 2010;
Ortega-Rivas and Salmerón-Ochoa, 2014). Another key advantage of the HP method is
its applicability to packaged foods, making efforts to prevent re-contamination or the use
of aseptic filling processes obsolete (Pereira and Vicente, 2010). HP is an environmentally
friendly technology and saves remarkable energy costs when compared to thermal meth-
ods (Pereira and Vicente, 2010; Ortega-Rivas and Salmerón-Ochoa, 2014). Thanks to
the benefits mentioned above, HP presents an exciting opportunity for food processors to
produce value-added food products with the highest freshness quality and nutrient value.
Although several advantages are attributed to HP, a few drawbacks are listed for it as
well. The principal disadvantage is the capital cost of HP technology (i.e., the initial outlay
on equipment), which is higher than thermal equipment (Sun, 2014). It is estimated that the
price of commercial-scale HP equipment ranges from 0.5 to 2.5 million USD, which is very
expensive (Sun, 2014). Many studies have shown that HP has limited efficacy against bac-
terial spores of many species such as B. stearothermophilus, B. subtilis, B. licheniformis,
or C. sporogenes at low temperatures (Torres and Velazquez, 2005; Pereira and Vicente,
2010; Cullen et al., 2012). This is a critical drawback and more research must be done to
tackle these HP obstacles and eliminate this technological limitation.

16.3.3 Ultrasound (US) Technology

During the last decade, ultrasound has received a lot of attention as a processing option
in the food industry (Rawson et al., 2011). Power ultrasound in food treatment is usually
applied in the lower-frequency range (20–100 kHz) with a sound intensity of between 10
and 1,000 W/cm 2 (Figure 16.4) (Cullen et al., 2012). However, it is stated that the most
popular range of frequencies is 20–25 kHz (Paniwnyk, 2016). When a sample is subjected
to US, the wave travels through the medium, resulting in a series of continuous com-
pressions and rarefactions in the material, which finally forms cavitation bubbles. The
formed bubbles subsequently collapse, release energy, and induce various mechanical,
chemical, and biochemical effects in the food sample (Rastogi, 2011; Cullen et al., 2012;
Paniwnyk, 2016). These chemical and physical effects provide a platform for the versatile
applications of US for almost every aspect of food production, ranging from preserva-
tion to modification (Knorr et al., 2011; Ortega-Riva and Salmerón-Ochoa, 2014). US
 Novel and Emerging Technologies  325

FIGURE 16.4 Frequency ranges of sound indicating infrasound (1–16 Hz), human hear-
ing (16 Hz–18 kHz), power ultrasound (20–100 kHz), extended for special applications
(20 kHz–1 MHz), and diagnostic ultrasound (5–10 MHz). (From Rastogi, 2011.)

is mainly used in three areas of food processing, which are (a) inactivation of spoilage
microorganisms and enzymes for shelf-life extension; (b) alteration of food constituents
or ingredients to modify their functionality; and (c) the stimulation of fermentation and
enzyme reactions at low treatment intensities (Knorr et al., 2011).
Various research studies have indicated that US has the ability to inactivate various
pathogenic microorganisms (e.g., Geobacillus stearothermophilus, Saccharomyces cere-
visiae, Escherichia coli, Salmonella enteritidis, Listeria monocytogenes, Pseudomonas
fluorescens, Staphylococcus aureus, Debaryomyces hansenii, and Lactobacillus del-
brueckii) in liquid food products. US can be applied either under pressure or in combina-
tion with a thermal and pressure treatment to efficiently pasteurize/sterilize the treated
sample (Rastogi, 2011; Rawson et al., 2011; Cullen et al., 2012; Paniwnyk, 2016). The
acoustic cavitation produced during US treatment causes shear disruption and forms free
radicals and hydrogen peroxide, leading to physical and/or chemical effects on the micro-
organism structure (Cullen et al., 2012). These effects may cause substantial damages in
the wall and membrane cells of microorganisms and lead to their fracture and leakage
(Knorr et al., 2011; Cullen et al., 2012; Paniwnyk, 2016). Simultaneously, US be used for
the inactivation of various enzymes (e.g., lipoxygenase, pectin methylesterase, alkaline
phosphatase, polygalacturonase, and polyphenol oxidase) by denaturation of proteins
(either by the chemical effects of free radicals or shear forces resulting from the collapse of
cavitating bubbles) (Cullen et al., 2012). US technology has proven to have many benefi-
cial applications in meat processing. It can improve binding capacity and strength, color,
and yield in processed meat. It can also help to tenderize meat tissues by affecting meat
proteins (Knorr et al., 2011; Rastogi, 2011). Sonication can also significantly accelerate
326 Food Aroma Evolution

TABLE 16.5 The Effect of Ultrasound (US) Technology on the Flavor/Aroma of Processed
Foods
Effect on Aroma, Flavor,
Technology Processing Conditions Product and Odor References
US 20 kHz, 263 W, 89.25 Passion Decreased sensory score (Gómez-
μm/8 min/10°C fruit juice for aroma and flavor López et al.,
sonicated juice 2018)
US 330 W/m2/9 min/19 Soursop No significant effect on (Dias et al.,
kHz juice aroma 2015)
US 8 min/20 Calcium- Significant reduction in (Gómez-
kHz/10°C/500 W/ added aroma score López et al.,
Wave amplitude of orange 2010)
89.25 μm juice
US 1, 10, 20, and 30 Orange No significant effect on (Guerrouj
min/24 kHz/105 μm/ juice aroma and flavor at et al., 2016)
below 46°C 1- and 10-min treatment/
rejected aroma and
flavor scores at 20- and
30-min treatment
US 20 kHz/3,6,9 min/20, Apple juice Formation of new (Šimunek
40, and 60°C/ and compounds in the aroma et al., 2013)
amplitude: 60, 90, nectar profile which were not
and 120 mm found in the untreated
samples/and
disappearance some
compounds which existed
in the untreated samples/
The highest sensory value
was observed for apple
juice at 60 mm and 40°C
for 6 min and apple
nectar at 120 mm and
20°C for 3 min
US 400 W/24 kHz/2.5, 5, Milk The generation of rubbery (Riener
10, 15, and 20 aroma in the sonicated et al., 2009)
min/45°C samples
US 30, 60, and 90% Apple juice Higher flavor (Abid et al.,
amplitude acceptability in a 2015)
levels/20 kHz, 20°C sonicated sample at 60
for 3 min and 90% than control/no
significant change at
30% amplitude
US 400 W/24 kHz/50–200 Milk Generation of burnt (Marchesini
s/amplitude 70–100% off-flavor et al., 2012)
US 24 kHz/200 W/0–16 Milk Decline in odor score with (Chouliara
min/15–25°C an increase of treatment et al., 2010)
time
(Continued )
 Novel and Emerging Technologies  327

TABLE 16.5 (Continued) The Effect of Ultrasound (US) Technology on the Flavor/Aroma
of Processed Foods
Effect on Aroma, Flavor,
Technology Processing Conditions Product and Odor References
US 40 kHz/250 Green tea Increase of the extraction (Xia et al.,
W/60°C/40 min yield of aroma 2006)
components and
glycosidic aroma
precursors/US-extracted
tea had fresher flavor as
compared to
conventional extraction

the freezing rate and minimize fouling from surface freezers in high-value foods (Knorr
et al., 2011; Rastogi, 2011). Regarding the fermentation process, US improves fermenta-
tion rates with an increase of mass transfer through the cell wall and membrane and its
influence on boundary layers (Knorr et al., 2011).
Apart from the aforementioned applications, US is also used to improve the opera-
tion of other processes including improving heat transfer during drying, decreasing cook-
ing time; reducing particle sizes, improving efficiency of filtration (improves water flow
through the filter cake); the degassing and destruction of foam, enhancing extraction
yield and efficiency during extraction of valuable compounds from vegetables and seeds;
retaining bioactive ingredients; mixing of materials; and better penetration of liquids into
food materials during marination or pickling (Rastogi, 2011; Cullen et al., 2012; Sun,
2014; Paniwnyk, 2016).

16.3.3.1 Advantages and Disadvantages of US

Similar to other non-thermal technologies, using US for food processing is advanta-
geous, and might have superiority over other conventional processing methods. It can
be useful for the shelf-life extension of heat-sensitive foods, with minimum adverse
quality effects (Rawson et al., 2011; Cullen et al., 2012). It is a safe and non-toxic
process with a short processing time, higher throughput, low energy consumption,
and environmentally friendly (Kentish and Ashokkumar, 2011; Rawson et al., 2011;
Paniwnyk, 2016). Therefore, owing to these benefits, US is strongly considered as a
promising food processing method for the industrial sector. There is some evidence
regarding the destructive effect of US on the quality of food (Rawson et al., 2011;
Cullen et al., 2012). This may happen due to the application of high intensity US for
the inactivation of microorganisms/enzymes that are resistant. This may cause adverse
effects on sensory attributes and limit US application in case of food preservation
(Rastogi, 2011). Table 16.5 shows some of these adverse quality effects on the aroma
profile and flavor compounds of US-treated foods.

REFERENCES

Abid, M., Jabbar, S., Wu, T., Hashim, M. M., Hu, B., Saeeduddin, M., and Zeng, X.
(2015). Qualitative assessment of sonicated apple juice during storage. Journal of
Food Processing and Preservation, 39(6), 1299–308.
328 Food Aroma Evolution

Aguilar-Rosas, S. F., Ballinas-Casarrubias, M. L., Nevarez-Moorillon, G. V., Martín-

Belloso, O., and Ortega-Rivas, E. (2007). Thermal and pulsed electric fields pasteuri-
zation of apple juice: Effects on physicochemical properties and flavour compounds.
Journal of Food Engineering, 83(1), 41–6.
Aguiló-Aguayo, I., Oms-Oliu, G., Soliva-Fortuny, R., and Martín-Belloso, O. (2009).
Flavour retention and related enzyme activities during storage of strawberry juices
processed by high-intensity pulsed electric fields or heat. Food Chemistry, 116(1),
59–65.
Awuah, G. B., Tang, J., and Ramaswamy, H. S. (2014). Radio-Frequency Heating in
Food Processing: Principles and Applications. CRC Press.
Azhu Valappil, Z., Fan, X., Zhang, H. Q., and Rouseff, R. L. (2009). Impact of thermal
and nonthermal processing technologies on unfermented apple cider aroma vola-
tiles. Journal of Agricultural and Food Chemistry, 57(3), 924–9.
Baxter, I. A., Easton, K., Schneebeli, K., and Whitfield, F. B. (2005). High-pressure pro-
cessing of Australian navel orange juices: Sensory analysis and volatile flavor profil-
ing. Innovative Food Science & Emerging Technologies, 6(4), 372–87.
Behera, S., Nagarajan, S., and Rao, L. J. M. (2004). Microwave heating and conventional
roasting of cumin seeds (Cuminum cyminum L.) and effect on chemical composi-
tion of volatiles. Food Chemistry, 87(1), 25–9.
Birla, S. L., Wang, S., Tang, J., Fellman, J. K., Mattinson, D. S., and Lurie, S. (2005).
Quality of oranges as influenced by potential radio frequency heat treatments against
Mediterranean fruit flies. Postharvest Biology and Technology, 38(1), 66–79.
Bolumar, T. , Georget, E. , and Mathys, A. (2015). High pressure processing (HPP) of foods
and its combination with electron beam processing. In Electron Beam Pasteurization
and Complementary Food Processing Technologies. Woodhead Publishing.
Buckow, R., Ng, S., and Toepfl, S. (2013). Pulsed electric field processing of orange juice:
A review on microbial, enzymatic, nutritional, and sensory quality and stability.
Comprehensive Reviews in Food Science and Food Safety, 12(5), 455–67.
Bukvicki, D., Gottardi, D., Tyagi, A. K., Veljic, M., Marin, P. D., Vujisic, L., … and
Vannini, L. (2014). Scapania nemorea liverwort extracts: Investigation on volatile
compounds, in vitro antimicrobial activity and control of Saccharomyces cerevisiae
in fruit juice. LWT-Food Science and Technology, 55(2), 452–8.
Cadwallader, K. R. and Singh, T. K. (2009). Flavours and off-flavours in milk and dairy
products. In Advanced Dairy Chemistry (pp. 631–90). Springer.
Caminiti, I. M., Noci, F., Muñoz, A., Whyte, P., Morgan, D. J., Cronin, D. A., and Lyng,
J. G. (2011). Impact of selected combinations of non-thermal processing technolo-
gies on the quality of an apple and cranberry juice blend. Food Chemistry, 124(4),
1387–92.
Canto, A. C. V. C. S., Lima, B. R. C. C., Cruz, A. G., La´zaro, C. A., Freitas, D. G. C.,
Faria, J. A. F., … and Silva, T. P. J. (2012). Effect of high hydrostatic pressure on the
color and texture parameters of refrigerated Caiman (Caiman crocodilus yacare)
tail meat. Meat Science, 91, 255–60.
Cappato, L. P., Ferreira, M. V. S., Guimaraes, J. T., Portela, J. B., Costa, A. L. R.,
Freitas, M. Q., … and Cruz, A. G. (2017). Ohmic heating in dairy processing:
Relevant aspects for safety and quality. Trends in Food Science & Technology, 62,
104–12.
Chouliara, E., Georgogianni, K. G., Kanellopoulou, N., and Kontominas, M. G. (2010).
Effect of ultrasonication on microbiological, chemical and sensory properties of
raw, thermized and pasteurized milk. International Dairy Journal, 20(5), 307–13.
 Novel and Emerging Technologies  329

Clare, D. A., Bang, W. S., Cartwright, G., Drake, M. A., Coronel, P., and Simunovic,
J. (2005). Comparison of sensory, microbiological, and biochemical parameters of
microwave versus indirect UHT fluid skim milk during storage. Journal of Dairy
Science, 88(12), 4172–82.
Contreras, C., Benlloch-Tinoco, M., Rodrigo, D., and Martínez-Navarrete, N. (2017).
Impact of microwave processing on nutritional, sensory, and other quality attributes.
In The Microwave Processing of Foods (Second Edition, pp. 65–99). Woodhead
Publishing.
Cserhalmi, Z., Sass-Kiss, A., Tóth-Markus, M., and Lechner, N. (2006). Study of
pulsed electric field treated citrus juices. Innovative Food Science & Emerging
Technologies, 7(1–2), 49–54.
Cullen, P.J., Tiwari, K. B., and Valdramidis, V. P. (2012). Novel Thermal and Non-
Thermal Technologies for Fluid Foods (First Edition). Academic Press.
Czechowska-Biskup, R., Rokita, B., Lotfy, S., Ulanski, P., and Rosiak, J. M. (2005).
Degradation of chitosan and starch by 360-kHz ultrasound. Carbohydrate
Polymers, 60(2), 175–84.
De Ancos, B., Cano, M. P., and Gómez, R. (2000). Characteristics of stirred low-fat
yoghurt as affected by high pressure. International Dairy Journal, 10(1–2), 105–11.
De Silva, G. O., Abeysundara, A. T., Minoli, M., and Aponso, W. (2018). Impacts of
pulsed electric field (PEF) technology in different approaches of the food industry: A
review. Journal of Pharmacognosy and Phytochemistry, 7(2), 737–40.
Dias, D. D. R. C., Barros, Z. M. P., de Carvalho, C. B. O., Honorato, F. A., Guerra, N.
B., and Azoubel, P. M. (2015). Effect of sonication on soursop juice quality. LWT-
Food Science and Technology, 62(1), 883–9.
Dima, F., Istrati, D., Garnai, M., Serea, V., and Vizireanu, C. (2015). Study on obtaining
vegetables juices with high antioxidant potential, preserved by ohmic pasteuriza-
tion. Journal of Agroalimentary Processes and Technologies, 21, 67–74.
Dorantes-Alvarez, L., Ortiz-Moreno, A., Guzmán-Gerónimo, R., and Parada-Dorantes,
L. (2017). Microwave-assisted blanching. In The Microwave Processing of Foods
(Second Edition, pp. 179–99). Woodhead Publishing.
Ekezie, F. G. C., Sun, D. W., Han, Z., and Cheng, J. H. (2017). Microwave-assisted
food processing technologies for enhancing product quality and process efficiency:
A review of recent developments. Trends in Food Science & Technology, 67, 58–69.
Eskin, N. M. and Shahidi, F. (2012). Off-Flavors in Milk, Biochemistry of Foods.
Academic Press.
Evrendilek, G. A., Li, S., Dantzer, W. R., & Zhang, Q. H. (2004). Pulsed electric field pro-
cessing of beer: Microbial, sensory, and quality analyses. Journal of Food Science,
69(8), M228–32.
Fellows, P. (2009). Food Processing Technologies: Principles and Practices (Third
Edition). CRC Press.
Gómez-López, V. M., Buitrago, M. E., and Martínez-Yépez, A. (2018). Effect of ultra-
sonication on sensory and chemical stability of passion fruit juice during refriger-
ated storage. Emirates Journal of Food and Agriculture, 30, 85–9.
Gómez-López, V. M., Orsolani, L., Martínez-Yépez, A., and Tapia, M. S. (2010).
Microbiological and sensory quality of sonicated calcium-added orange juice. LWT-
Food Science and Technology, 43(5), 808–13.
Guerrouj, K., Sánchez-Rubio, M., Taboada-Rodríguez, A., Cava-Roda, R. M., and Marín-
Iniesta, F. (2016). Sonication at mild temperatures enhances bioactive compounds and
microbiological quality of orange juice. Food and Bioproducts Processing, 99, 20–8.
330 Food Aroma Evolution

He, J., Vazquez-Landaverde, P. , Qian, M. C. , and Eskin, N. M. (2013). Off-flavors

in milk. In Biochemistry of Foods (Third Edition, pp. 479–500). Academic Press.
Hebbar, H. U., Vishwanathan, K. H., and Ramesh, M. N. (2004). Development of com-
bined infrared and hot air dryer for vegetables. Journal of Food Engineering, 65,
557–63.
Hee, S.C. (2006). Preparation methods of retort roast chestnuts using far infrared ray
thawing. WO Patent 2006/065018 A1.
Hou, L., Johnson, J. A., and Wang, S. (2016). Radio frequency heating for posthar-
vest control of pests in agricultural products: A review. Postharvest Biology and
Technology, 113, 106–18.
Hurtado, A., Picouet, P., Jofré, A., Guàrdia, M. D., Ros, J. M., and Bañón, S. (2015).
Application of high-pressure processing for obtaining “fresh-like” fruit smoothies.
Food and Bioprocess Technology, 8(12), 2470–82.
Icier, F. and Ilicali, C. (2005). Temperature dependent electrical conductivities of fruit
purees during ohmic heating. Food Research International, 38(10), 1135–42.
Ignat, A., Manzocco, L., Brunton, N. P., Nicoli, M. C., and Lyng, J. G. (2015). The effect
of pulsed electric field pre-treatments prior to deep-fat frying on quality aspects of
potato fries. Innovative Food Science & Emerging Technologies, 29, 65–9.
Ildikó, S. G., Klára, K. A., Marianna, T. M., Agnes, B., Zsuzsanna, M. B., and Bálint,
C. (2006). The effect of radio frequency heat treatment on nutritional and colloid-
chemical properties of different white mustard (Sinapis alba L.) varieties. Innovative
Food Science & Emerging Technologies, 7(1–2), 74–9.
Jiao, Y., Tang, J., Wang, Y., and Koral, T. L. (2018). Radio-frequency applications for
food processing and safety. Annual Review of Food Science and Technology, 9,
105–27.
Kaur, N. & Singh, A. K. (2016). Ohmic heating: Concept and applications—A review.
Critical Reviews in Food Science and Nutrition, 56(14), 2338–51.
Kentish, S. and Ashokkumar, M. (2011). The physical and chemical effects of ultrasound.
In Ultrasound Technologies for Food and Bioprocessing (pp. 1–12). Springer, New
York, NY.
Kimura, K., Ida, M., Yosida, Y., Ohki, K., Fukumoto, T., and Sakui, N. (1994).
Comparison of keeping quality between pressure-processed jam and heat-processed
jam: Changes in flavor components, hue, and nutrients during storage. Bioscience,
Biotechnology, and Biochemistry, 58(8), 1386–91.
Knirsch, M. C., Dos Santos, C. A., de Oliveira Soares, A. A. M., and Penna, T. C. V.
(2010). Ohmic heating—A review. Trends in Food Science & Technology, 21(9),
436–41.
Knorr, D., Froehling, A., Jaeger, H., Reineke, K., Schlueter, O., and Schoessler, K. (2011).
Emerging technologies in food processing. Annual Review of Food Science and
Technology, 2, 203–35.
Koray Palazoğlu, T., Coşkun, Y., Kocadağlı, T., and Gökmen, V. (2012). Effect of radio
frequency postdrying of partially baked cookies on acrylamide content, texture, and
color of the final product. Journal of Food Science, 77(5), E113–7.
Krishnamurthy, K., Khurana, H. K., Soojin, J., Irudayaraj, J., and Demirci, A. (2008).
Infrared heating in food processing: An overview. Comprehensive Reviews in Food
Science and Food Safety, 7(1), 2–13.
Kumar, C. M., Appu Rao, A. G., and Singh, S. A. (2009). Effect of infrared heating on
the formation of sesamol and quality of defatted flours from Sesamum indicum L.
Journal of Food Science, 74(4), H105–11.
 Novel and Emerging Technologies  331

Leizerson, S. and Shimoni, E. (2005). Stability and sensory shelf-life of orange juice pas-
teurized by continuous ohmic heating. Journal of Agricultural and Food Chemistry,
53(10), 4012–8.
Li, C., Hao, J., Zhong, H., Dang, M., and Xie, B. (2009). Aroma components at various
stages of litchi juice processing. Journal of the Science of Food and Agriculture,
89(14), 2405–14.
Liu, Q., Zhang, M., Xu, B., Fang, Z., and Zheng, D. (2015). Effect of radio frequency
heating on the sterilization and product quality of vacuum packaged Caixin. Food
and Bioproducts Processing, 95, 47–54.
Lund, D. (2003). Predicting the impact of food processing on food constituents. Journal
of Food Engineering, 56(2–3), 113–7.
Lynch, S. (2016). Pulsed Electric Fields (PEF): Technology, Role in Food Science and
Emerging Applications. Nova Science Publishers, Inc.
Ma, Y., Hu, X., Chen, J., Zhao, G., Liao, X., Chen, F., … and Wang, Z. (2010). Effect of
UHP on enzyme, microorganism and flavor in cantaloupe (Cucumis melo L.) juice.
Journal of Food Process Engineering, 33(3), 540–53.
Marchesini, G., Balzan, S., Montemurro, F., Fasolato, L., Andrighetto, I., Segato, S., and
Novelli, E. (2012). Effect of ultrasound alone or ultrasound coupled with CO2 on
the chemical composition, cheese-making properties and sensory traits of raw milk.
Innovative Food Science & Emerging Technologies, 16, 391–7.
Meda, V., Orsat, V., and Raghavan, V. (2017). Microwave heating and the dielectric
properties of foods. In The Microwave Processing of Foods (Second Edition, pp.
23–43). Woodhead Publishing.
Min, S., Jin, Z. T., Min, S. K., Yeom, H., and Zhang, Q. H. (2003). Commercial‐scale
pulsed electric field processing of orange juice. Journal of Food Science, 68(4),
1265–71.
Min, S. and Zhang, Q. H. (2003). Effects of commercial‐scale pulsed electric field
processing on flavor and color of tomato juice. Journal of Food Science, 68(5),
1600–6.
Navarro, M., Verret, C., Pardon, P., and El Moueffak, A. (2002). Changes in volatile
aromatic compounds of strawberry purée treated by high-pressure during storage.
International Journal of High-Pressure Research, 22(3–4), 693–6.
Orsat, V., Bai, L., Raghavan, G. S. V., and Smith, J. P. (2004). Radio‐frequency heat-
ing of ham to enhance shelf‐life in vacuum packaging. Journal of Food Process
Engineering, 27(4), 267–83.
Orsat, V., Raghavan, G. S. V., and Krishnaswamy, K. (2017). Microwave technology for
food processing: An overview of current and future applications. In The Microwave
Processing of Foods (Second Edition, pp. 100–16). Woodhead Publishing.
Ortega-Rivas, E. and Salmerón-Ochoa, I. (2014). Nonthermal food processing alterna-
tives and their effects on taste and flavor compounds of beverages. Critical Reviews
in Food Science and Nutrition, 54(2), 190–207.
Pan, Z., Shih, C., McHugh, T. H., and Hirschberg, E. (2008). Study of banana dehy-
dration using sequential infrared radiation heating and freeze-drying. LWT-Food
Science and Technology, 41, 1944–51.
Paniwnyk, L. (2016). Applications of ultrasound in processing of liquid foods: A review.
Ultrasonics Sonochemistry. doi: http:​//dx.​doi.o​rg/10​.1016​/j.ul​tsonc​h.201​6.12.​025.
Park, J. H., Lee, J. M., Cho, Y. J., Kim, C. T., Kim, C. J., Nam, K. C., and Lee, S. C.
(2009). Effect of far‐infrared heater on the physicochemical characteristics of green
tea during processing. Journal of Food Biochemistry, 33(2), 149–62.
332 Food Aroma Evolution

Parker, J. K., Elmore, S., and Methven, L. (Eds.). (2014). Flavour Development, Analysis
and Perception in Food and Beverages. Elsevier.
Pereira, R. N. and Vicente, A. A. (2010). Environmental impact of novel thermal and
non-thermal technologies in food processing. Food Research International, 43(7),
1936–43.
Perez-Cacho, P. R. and Rouseff, R. (2008). Processing and storage effects on orange
juice aroma: A review. Journal of Agricultural and Food Chemistry, 56(21),
9785–96.
Piyasena, P., Dussault, C., Koutchma, T., Ramaswamy, H. S., and Awuah, G. B. (2003).
Radio frequency heating of foods: Principles, applications and related properties—A
review. Critical Reviews in Food Science and Nutrition, 43(6), 587–606.
Pongviratchai, P. and Park, J. W. (2007). Electrical conductivity and physical proper-
ties of surimi–potato starch under ohmic heating. Journal of Food Science, 72(9),
E503–7.
Puligundla, P., Abdullah, S. A., Choi, W., Jun, S., Oh, S. E., and Ko, S. (2013). Potentials
of microwave heating technology for select food processing applications: A brief
overview and update. Journal of Food Processing & Technology, 4(11), 278.
Qu, F., Zhu, X., Ai, Z., Ai, Y., Qiu, F., and Ni, D. (2019). Effect of different drying
methods on the sensory quality and chemical components of black tea. LWT—Food
Science and Technology, 99, 112–8.
Rastogi, N. K. (2011). Opportunities and challenges in application of ultrasound in food
processing. Critical Reviews in Food Science and Nutrition, 51, 705–22.
Rastogi, N. K. (2012). Recent trends and developments in infrared heating in food pro-
cessing. Critical Reviews in Food Science and Nutrition, 52(9), 737–60.
Rawson, A., Patras, A., Tiwari, B. K., Noci, F., Koutchma, T., and Brunton, N. (2011).
Effect of thermal and nonthermal processing technologies on the bioactive con-
tent of exotic fruits and their products: Review of recent advances. Food Research
International, 44(7), 1875–7.
Regier, M. , Knoerzer, K. , and Schubert, H. (2017). Introducing microwave-assisted
processing of food: Fundamentals of the technology. In The Microwave Processing
of Foods (Second Edition, pp. 1–22). Woodhead Publishing.
Riadh, M. H., Ahmad, S. A. B., Marhaban, M. H., and Soh, A. C. (2015). Infrared heat-
ing in food drying: An overview. Drying Technology, 33(3), 322–35.
Richardson, P. (2001). Thermal Technologies in Food Processing (First Edition). CRC
Press, Boca Raton, FL.
Riener, J., Noci, F., Cronin, D. A., Morgan, D. J., and Lyng, J. G. (2009). Characterization
of volatile compounds generated in milk by high intensity ultrasound. International
Dairy Journal, 19(4), 269–72.
Sakr, M. and Liu, S. (2014). A comprehensive review on applications of ohmic heating
(OH). Renewable and Sustainable Energy Reviews, 39, 262–9.
Sandi, D., Chaves, J. B. P., Parreiras, J. F. M., De Souza, A. C. G., and Da Silva, M. T.
C. (2003). Avaliação da qualidade sensorial de suco de maracujá amarelo (Passiflora
edulisvar. flavicarpa) submetido à pasteurização e armazenamento. Boll. CPPA, 21,
141–58.
Sarang, S., Sastry, S. K., and Knipe, L. (2008). Electrical conductivity of fruits and meats
during ohmic heating. Journal of Food Engineering, 87(3), 351–6.
Schindler, S., Krings, U., Berger, R. G., and Orlien, V. (2010). Aroma development in
high pressure treated beef and chicken meat compared to raw and heat treated. Meat
Science, 86, 317–23.
 Novel and Emerging Technologies  333

Schirack, A. V., Drake, M. A., Sanders, T. H., and Sandeep, K. P. (2006). Characterization
of aroma active compounds in microwave blanched peanuts. Journal of Food
Science, 71(9), C513–20.
Sharma, G. P. & Prasad, S. (2001). Drying of garlic (Allium sativum) cloves by micro-
wave–hot air combination. Journal of Food Engineering, 50(2), 99–105.
Šimunek, M., Režek Jambrak, A., Petrović, M., Juretić, H., Major, N., Herceg, Z., …
and Vukušić, T. (2013). Aroma profile and sensory properties of ultrasound-treated
apple juice and nectar. Food Technology and Biotechnology, 51(1), 101–11.
Sinchaipanit, P., Kerr, W. L., and Chamchan, R. (2013). Effect of sweeteners and hydro-
colloids on quality attributes of reduced-calorie carrot juice. Journal of the Science
of Food and Agriculture, 93, 3304–11.
Soliva-Fortuny, R., Balasa, A., Knorr, D., and Martín-Belloso, O. (2009). Effects of
pulsed electric fields on bioactive compounds in foods: A review. Trends in Food
Science & Technology, 20(11–2), 544–56.
Stoica, M., Mihalcea, L., Borda, D., and Alexe, P. (2013). Non-thermal novel food
processing technologies. An overview. Journal of Agroalimentary Processes and
Technologies, 19(2), 212–7.
Sun, D. W. (2014). Emerging Technologies for Food Processing. Elsevier.
Surowsky, B., Schluter, O., and Knorr, D. (2014). Interactions of non-thermal atmospheric
pressure plasma with solid and liquid food systems: A review. Food Engineering
Reviews, 7, 82–108.
Syed, Q. A., Ishaq, A., Rahman, U. U., Aslam, S., and Shukat, R. (2017). Pulsed electric
field technology in food preservation: A review. Journal of Nutritional Health and
Food Engineering, 6(6), 1–5.
Torres, J. A. and Velazquez, G. (2005). Commercial opportunities and research chal-
lenges in the high-pressure processing of foods. Journal of Food Engineering, 67(1–
2), 95–12.
Tumpanuvatr, T. and Jittanit, W. (2012). The temperature prediction of some botanical
beverages concentrated juices and purees of orange and pineapple during ohmic
heating. Journal of Food Engineering, 113(2), 226–33.
Vadivambal, R. and Jayas, D. S. (2010). Non-uniform temperature distribution during
microwave heating of food materials—A review. Food and Bioprocess Technology,
3(2), 161–71.
Valero, E., Villamiel, M., Sanz, J., and Martınez-Castro, I. (2000). Chemical and senso-
rial changes in milk pasteurised by microwave and conventional systems during cold
storage. Food Chemistry, 70(1), 77–81.
Venkatesh, M. S. and Raghavan, G. S. V. (2004). An overview of microwave process-
ing and dielectric properties of agri-food materials. Biosystems Engineering, 88(1),
1–18.
Viljanen, K., Lille, M., Heiniö, R., and Buchert, J. (2011). Effect of highpressure process-
ing on volatile composition and odour of cherry tomato purée. Food Chemistry,
129, 1759–65.
Wang, Y., Wig, T. D., Tang, J., and Hallberg, L. M. (2003). Sterilization of foodstuffs
using radio frequency heating. Journal of Food Science, 68(2), 539–44.
Wen, G. (2014). Modeling of Ohmic Heating of Solid Food with Non-uniform Electric
Properties. Master’s Thesis, Tokyo University of Marine Science and Technology.
Wiktor, A., Schulz, M., Voigt, E., Witrowa-Rajchert, D., and Knorr, D. (2015). The
effect of pulsed electric field treatment on immersion freezing, thawing and selected
properties of apple tissue. Journal of Food Engineering, 146, 8–16.
334 Food Aroma Evolution

Xia, T., Shi, S., and Wan, X. (2006). Impact of ultrasonic-assisted extraction on the
chemical and sensory quality of tea infusion. Journal of Food Engineering, 74,
557–60.
Yang, J., Bingol, G., Pan, Z., Brandl, M. T., McHugh, T. H., and Wang, H. (2010).
Infrared heating for dry-roasting and pasteurization of almonds. Journal of Food
Engineering, 101(3), 273–80.
Yen, G. C. and Lin, H. T. (1999). Changes in volatile flavor components of guava juice
with high-pressure treatment and heat processing and during storage. Journal of
Agricultural and Food Chemistry, 47(5), 2082–7.
Yeom, H. W., Streaker, C. B., Zhang, Q. H., and Min, D. B. (2000). Effects of pulsed
electric fields on the quality of orange juice and comparison with heat pasteuriza-
tion. Journal of Agricultural and Food Chemistry, 48(10), 4597–605.
Yolacaner, E. T., Sumnu, G., and Sahin, S. (2017). Microwave-assisted baking. In The
Microwave Processing of Foods (Second Edition, pp. 117–41). Woodhead Publishing.
Zabetakis, I., Koulentianos, A., Orruno, E., and Boyes, I. (2000). The effect of high
hydrostatic pressure on strawberry flavour compounds. Food Chemistry, 71(1),
51–5.
Zell, M., Lyng, J. G., Cronin, D. A., and Morgan, D. J. (2009). Ohmic heating of meats:
Electrical conductivities of whole meats and processed meat ingredients. Meat
Science, 83(3), 563–70.
Zell, M., Lyng, J. G., Cronin, D. A., and Morgan, D. J. (2010). Ohmic cooking of whole
turkey meat—Effect of rapid ohmic heating on selected product parameters. Food
Chemistry, 120(3), 724–9.
Zell, M., Lyng, J. G., Morgan, D. J., and Cronin, D. A. (2012). Quality evaluation of an
ohmically cooked ham product. Food and Bioprocess Technology, 5(1), 265–72.
Zhang, L., Lyng, J. G., and Brunton, N. P. (2006). Quality of radio frequency heated
pork leg and shoulder ham. Journal of Food Engineering, 75(2), 275–87.
Zhang, Y. I., Gao, B. E. I., Zhang, M., Shi, J., and Xu, Y. (2010). Pulsed electric field pro-
cessing effects on physicochemical properties, flavor compounds, and microorgan-
isms of longan juice. Journal of Food Processing and Preservation, 34(6), 1121–38.
 Section IV
Aroma Compounds
in Food Matrices
 Chapter 17
Distillates
Paul S. Hughes

CONTENTS

17.1 Introduction 337

17.2 Major Aroma Volatiles in Distilled Spirits 338
17.3 Raw Materials and Processing 340
17.3.1 Aromas Derived from Carbohydrates 342
17.3.2 Aromas Derived from Cereals 342
17.3.3 Aromas Derived from Fruits 343
17.4 Fermentation 344
17.5 Distillation 346
17.6 Downstream Modification 348
17.7 Undesirable Aromas (Taints) 350
17.8 Future Insights 351
References 351

17.1 INTRODUCTION

Today’s distilled spirit drinks market is a celebration of diversity. A combination of a

broad range of raw materials, distillation technologies, the use of flavors (either dur-
ing distillation or added downstream, Figure 17.1), and other downstream modifications
broaden the sensory scope of the distilled spirits sector further, not least in terms of
flavor. The historical diversity of distilled spirits still exists to this day, so despite the
existence of several global spirit companies, there is still a substantial number of products
that are produced and consumed locally from locally available raw materials.
The evolution of distilled spirits appears to have followed two parallel chronologi-
cal paths: the application of distillation to local fermentations, which rely in turn on the
conversion of locally harvested sugar sources and their conversion to alcohol, and the
discovery of a range of flavor modifications to enhance and differentiate the spirits pro-
duced. The timing of the discovery of distillation is contentious, with some archaeologi-
cal evidence in China pointing to around 2000 BCE, but, in any case, its (re)discovery
by the Arabic scientist Jabir ibn Hayyan in around the 9th century was not really taken
up until his texts were translated into Latin in the 11th century. Fermentation of plant
extracts pre-dated distillation, and so there was a ready supply of fruit- and cereal-based
alcohol sources available, and later the introduction of distillation into Central America
led to the development of Mexican spirits such as tequila and mescal, and rum produc-
tion from sugar cane plantations in the Caribbean and South America.

337
338 Food Aroma Evolution

FIGURE 17.1 Qualitative mapping of distilled spirits based on a carbohydrate source.

The discovery of alcohol as an effective solvent for the dissolution of remedial

plant materials such as juniper led directly to botanical spirits such as gin, aquavit, and
absinthe and, with the observation that the serendipitous storage of spirits in casks gener-
ally enhanced their quality, it is clear that there are many options for introducing flavors
into spirits, whether they are volatile or nonvolatile. In this chapter, we will explore the
evolution of volatile contributions (aromas) to the flavor of major distilled spirits as a
function of the various processes that in turn create fermented extracts, produce distil-
lates, and finally give rise to final products after finishing, through maturation, the addi-
tion of flavors and colors, and filtration.

17.2 MAJOR AROMA VOLATILES IN DISTILLED SPIRITS

The number of aroma compounds identified in distilled spirits has grown substantially,
fueled primarily by the widespread adoption of capillary gas chromatography-mass spec-
trometry (GC-MS) based methods for the analysis of volatiles, the evolution of mass
spectrometry from the realm of the specialist to that of routine analysis, and the develop-
ment of comprehensive mass spectral libraries. In the context of instrumental develop-
ments, relatively early works by Nykanen and Suomalainen (1983) and by Piggott (1983)
arguably provided a springboard for the ever more detailed probing of distilled spirits,
both in terms of aroma and taste, as exemplified by Piggott (2012). Universities and
research laboratories alike have continued to evolve analytical and sensory technologies
and have published a plethora of papers on the volatile composition of a wide range of
distilled spirits.
There is a tenuous overlap between the mechanism of olfactory perception and the
movement of volatiles during GC analysis in that the detection of an aroma requires its
volatilization, either in the nasal cavity to reach the olfactory epithelium or to traverse
the GC column to reach the detector. There is an additional constraint for olfactory
perception, which is that there is a requirement for steric compatibility between at least
one of the 350 or so human olfactory receptors and the aroma molecule; otherwise, the
receptor will not be triggered. Also, if an aroma molecule has a molecular weight much
 Distillates 339

above 300–350 g/mol, it is unlikely to either satisfy any steric criteria required for satis-
factory receptor binding and subsequent perception or to be of insufficient volatility to
reach the receptors.
In the broadest terms, the categories into which aroma volatiles can be classified
are either spirit-based or functional group-based. For the purposes of this review,
the classification will be primarily by functional group (Table 17.1). The flavor activ-
ity of aroma volatiles varies substantially. Using thresholds cited in Burdock and
Fenaroli (2010), which were determined in 48% ABV solution, ester flavor detection,
for instance, can range from 6.9 µg/L (ethyl 3-methylbutanoate) to 32.6 mg/L (ethyl
acetate). The diester, diethyl succinate (derived from the excretion of succinic acid
formed in turn in the tricarboxylic acid cycle during fermentation), has a threshold
of more than 350 mg/L, almost five orders of magnitude higher than that cited above
(Table 17.2). While a given functional group can help us predict the sensory perfor-
mance of an aroma volatile, in practice, even closely related chemical entities can have
distinct aroma qualities. Thus while ethanol and propanol may both be deemed to be
“alcoholic,” they are still readily distinguishable.

TABLE 17.1 Classification of Aroma Volatiles in Distilled Spirits

Volatile Class Examples
Alcohols Ethanol
– Monohydric 2,3,-Butanediol
– Dihydric Glycerol
– Trihydric Phenylethanol
– Aromatic
Carbonyls Acetaldehyde
– Aldehydes 2-Heptanone
– Ketones Diacetyl (2,3-butanedione)
– Dicarbonyls Acetal (1,1-diethoxyethane)
– Acetals
Carboxylic acids Acetic acid
– Monobasic Oxalic acid
– Dibasic Pyruvic acid
– Oxoacids Lactic acid
– Hydroxyacids
Esters
– Monoesters Ethyl butyrate
– Diesters Diethyl succinate
– Lactones
Ethers Diethyl ether
Amines and amides Methylamine
Heterocyclic nitrogen compounds Pyrazine
Heterocyclic oxygen compounds Furan
Terpenes Linalool
Sulfur compounds Dimethyl trisulfide
Halogenated compounds Trichloroanisole
Non-terpene hydrocarbons 2-Methylbutene
Source: After Nykanen and Suomalainen, 1983.
340 Food Aroma Evolution

TABLE 17.2 Typical Threshold Ranges for

Classes of Aroma Volatiles Determined in
48% ABV Solution
Aroma Volatile Class Threshold Range (µg/L)
Esters 6.9–353,000
Carboxylic acids 960–13,700
Alcohols 6.12–179,000
Terpenes 0.12–680
Aldehydes 410–1,200
Ketones 483–1,330
Sulfur compounds 0.36–70
Source: After Burdock and Fenaroli, 2010.

As there are only around 350 olfactory receptors expressed in humans, it is unlikely
that one receptor codes for one compound or even a group of compounds. There is grow-
ing evidence for the so-called combinatorial theory of aroma detection. Here a specific
compound may trigger more than one receptor. A related compound, such as a homolog,
is likely to trigger a similar array of receptors, but not necessarily exactly the same (pre-
sumably due to steric differences). If we then factor in the idea that a compound may or
may not be a perfect fit, so that the probability of a receptor firing may vary, depending
on the aroma ligand, then it is possible to rationalize the vast array of aroma nuances that
humans can detect (Malnic et al., 1999).
While a given aroma compound might be derived from a range of sources, there
are some commonalities from a chemical class perspective (Figure 17.2). In the broadest
terms, fermentation often contributes a significant proportion of the flavor activity of a
spirit, even if we discount the role of ethanol on flavor perception. Notable exceptions
are those distilled spirit drinks based on neutral alcohol. In these instances, the alcohol
base is generally distilled to a purity which substantially excludes all components except
ethanol and water. For unflavored vodkas, this can be sufficient for a work-up to a final
product. For other relevant spirits, such as those flavored with botanical-derived flavors,
cream, or eggs, it is these additions that make major flavor, and aroma, contributions to
the final product. These various sources of aromas will be considered in turn below.

17.3 RAW MATERIALS AND PROCESSING

The production of alcoholic distillates begins with a suitable agricultural source of carbo-
hydrates which can be processed to release fermentability (Table 17.3). Plants store sugars
in various forms as an energy repository both for respiration and for future growth. The
efforts required to release fermentable sugars from these sources vary substantially but
can be broadly classified as fermentable as is or requiring pre-processing before fermen-
tation. In principle aroma compounds that are derived directly from the raw materials
themselves are a function of the variety and provenance of the raw materials, together
with the conditions of processing. Those raw materials that can be fermented as-is are
likely to have a volatiles profile that is more dependent on the provenance of the raw
materials. In this context we can consider molasses to be a raw material, although strictly
speaking it is a by-product of crystallized sugar production. Also, the use of sulfites in
 Distillates 341

FIGURE 17.2 Overview of sources of aromas during the production of distilled spirits.
*MRP stands for Maillard reaction products.

TABLE 17.3 Main Sources of Carbohydrate for Fermentation

Fermentability Releasing
Status Raw Material Principal Sugar Substrates Fermentability
Fermentable Grapes Glucose, fructose, sucrose Crush/strain
as is (simple Other fruits Various including glucose, Crush/mill/strain
sugars) fructose, sucrose
Sugar cane/beet Glucose, fructose, sucrose Crush/strain
Molasses Glucose, fructose, sucrose Dilution
Pre-processing Cereals Starch Malting, milling,
needed mashing
Potatoes Starch Enzymes
Sweet potato, Starch Enzymes
cassava
Agave Inulin Crush, heat, acid
Milk Lactose Specialized yeasts,
enzymes
342 Food Aroma Evolution

some fermentations (e.g., apple, grape, pear) to restrict oxidation can risk its reductive
conversion during fermentation to hydrogen sulfide, especially if sulfur amino acids are
restricted in the fermentation.

17.3.1 Aromas Derived from Carbohydrates

A major source of aroma in many foods and beverages is the thermal degradation of sug-
ars in the presence or absence of amino nitrogen (e.g., amino acids). These pathways are
complex, and can, for instance, ultimately lead to the browning of food and beverages.
The pathways to the production of flavors depend on whether amino nitrogen is pres-
ent. In the absence of amino nitrogen, sugars can caramelize forming brown polymers
and the characteristic caramel flavors. Typical aroma impact characters derived from
caramel can include diacetyl, furans (nutty), maltol (toast), pyruvaldehyde (pungent),
hydroxyacetone (sweet), and ethyl acetate (solvent, sweet). Such sources of flavors are
most likely to occur in spirits made by the fermentation of molasses, such as some Indian
whiskies and some Caribbean rums. Heavily kilned malted cereals may also develop
caramel flavors. One additional aspect to bear in mind is that furfural is often used as a
marker for sugar degradation, particularly fructose, which is generally more thermally
labile. Tequila legislation sets a maximum limit of 4 mg/100 mL pure alcohol for all
final products.
The use of commercial caramels to color spirits does not have any discernible impact
on spirit flavor due to the intensity of color in commercial caramels, and therefore they
are only used in small quantities (typically around 0.1% (v/v), depending on the color
intensity required). It is worth bearing in mind that caramels can be made commercially
in the presence or absence of a nitrogen source (typically an ammonium salt) although
those used for high proof spirits generally are made only with sugar (so-called Type I or
E150a caramels). This is primarily because of their higher solubility in such spirits rela-
tive to other classes.
In the presence of free amino nitrogen, sugars can react to generate an array of aroma
compounds (Figure 17.3, Table 17.4). The resulting mixture of intermediates, color, and
flavor compounds are known collectively as Maillard reaction products (MRPs). Color
formations during caramelization and MRP generation are also known as non-enzy-
mic browning reactions. The generation of these compounds can occur both during raw
material production (e.g., during raw material drying) and during heating and boiling.
While furans are usually markers of other, flavor-active, compounds, the formation of
furaneol, cyclotene, Strecker aldehydes and, under more intense heating, pyrazines can
have a significant influence on final spirit flavor.

17.3.2 Aromas Derived from Cereals

The use of grains or cereals for the production of distilled spirits arguably evolved in
Northern Europe, together with the use of potatoes for spirit production. Generally,
these spirits directly contribute MRPs, as described above, if the cereals are dried suf-
ficiently to allow non-enzymic browning to occur. Additional contributions can come
from dimethyl sulfide (sweetcorn, canned tomato), a thermal degradation product of
S-methylmethionine during kilning, and flavor-active aldehydes caused by the Strecker
degradation of certain amino acids (see Table 17.3).
 Distillates 343

FIGURE 17.3 Representative reaction scheme showing steps for (a) the formation of
aldehydes from sugars and amino acids and (b) subsequent formation of pyrazines and
melanoidins. (Reproduced with permission from Baxter and Hughes, 2001, by the Royal
Society of Chemistry.)

17.3.3 Aromas Derived from Fruits

Many of the volatiles derived from fruits are also found in other sources (such as botani-
cals, see below), including C6-aldehydes and alcohols, terpenes, and the highly flavor-active
compounds α-ionone (violets) and β-damascenone (sweet, stewed fruit). Another highly
flavor-active compound, TPB (E-1-(2,3,6-trimethyl)-1,3,-butadiene) has been found in
­
white wines and Cognac eau-de-vies and has a strong aroma of geraniums. The range of
344 Food Aroma Evolution

TABLE 17.4 Typical Flavor-Impact Heterocyclic

Compound Classes Formed during Maillard-Based
Browning Reactions of Cereals
Compound Aroma Attributes
Pyrazines Cooked, roasted, toasted
Alkylpyridines Green, burned, bitter
Acylpyridines Cracker-like
Pyrroles Cereal-like
Furans, furanones, pyranones Sweet, burned, caramel-like
Oxazoles Green, nutty, sweet
Source: Adapted from van Boekel, 2006.

highly flavor-active compounds that can be derived from fruit allows for substantial oppor-
tunities to tailor a flavor profile, although this also implies the need to carefully select fruit
for fermentation and to manage its storage to ensure flavor quality and consistency.

17.4 FERMENTATION

Much has been written concerning the production of flavor compounds during fermenta-
tion. Not least the role of Saccharomyces cerevisiae in flavor and alcohol production, which
was substantially reviewed by Boulton and Quain (2006). The major aroma compound
formed during fermentation is ethanol. Although it is only modestly flavor-active (thresh-
old of 10–15 g/L in water), typically distilled spirits contain in excess of 300 g/L. In most
cases this ethanol is produced by the yeast fermentation of starch hydrolysates (glucose,
maltose, maltotriose) or directly from sucrose or invert sugars (Figure 17.4). The uptake of
simple sugars is followed by glycolysis, where six-carbon sugars are successively modified to
form two three-carbon species: pyruvic acid (2-oxopropanoic acid) and glycerol. Glycerol
is excreted or used to produce mono-, di-, and triglyceride fats. Pyruvate readily decarbox-
ylates, releasing carbon dioxide, and the reactive acetyl fragment is either stabilized as an
adduct with coenzyme A or is converted to acetaldehyde. Enzymic reduction of acetalde-
hyde yields ethanol, while acetyl coenzyme A can be elaborated to longer-chain fatty acids
or, in conjunction with ethanol and higher alcohols, form esters. Both higher alcohols and
esters are generally flavor-active and found in appreciable quantities in most distilled spir-
its, although the most flavor-relevant esters tend to have lower flavor thresholds.
An alternative outlet for pyruvic acid is the formal addition of acetaldehyde to the
α-carbonyl to form α-acetolactate (Figure 17.5). While this is a key intermediate for the
biosynthesis of the amino acid valine, its oxidative decarboxylation yields the highly
flavor-active diacetyl (2,3-butanedione). If not dealt with, diacetyl can cause problems
with the flavor of the final spirit as its distilling properties are very similar to ethanol (for
a comprehensive review of diacetyl, see Inoue, 2008). Its low flavor threshold and distinc-
tive butterscotch aroma and sweet flavor mean that it can have a significant impact on
spirit quality. The consequence of its distilling properties is that, if present after fermenta-
tion, it can prove difficult to separate in the still.
Yeast can sequentially reduce one or both carbonyls to 3-hydroxy-2-butanone (acet-
oin) and 2,3-butanediol, which are both easier to separate from ethanol during distilla-
tion and have much higher flavor thresholds. A potential issue associated with significant
levels of 2,3-butanediol is its ability to form cyclic acetals with aldehydes, particularly
 Distillates 345

FIGURE 17.4 Summary of biochemical pathways leading to aroma compound formation

during a typical Saccharomyces cerevisiae fermentation. TCA stands for tricarboxylic
acid.

acetaldehyde (Figure 17.6). The resulting dioxolanes derived from 2,3-butanediol and
acetaldehyde have strong sherry- and raisin-like aromas, which may or may not be desired
in the final product. If undesirable, a management strategy is to minimize diacetyl pro-
duction during fermentation, in turn minimizing levels of 2,3-butanediol. Alternatively,
restricting the reduction of diacetyl to acetoin helps to circumvent both elevated levels of
diacetyl and 2,3-butanediol.
The formation of higher alcohols and esters, after ethanol production, are generally
considered to be the most important for fermentation-derived flavors. It is clear that even
a simplified biochemical scheme focused on aroma formation (Figure 17.4) shows the
inter-relationships and complexity of aroma production, not least for esters and higher

FIGURE 17.5 Simplified pathway to diacetyl formation in Saccharomyces cerevisiae

fermentations.
346 Food Aroma Evolution

FIGURE 17.6 Dioxolane formation from aldehydes and 2,3-butanediol. (* Indicates the
undefined chiral center.)
alcohols. The composition of the fermentation medium, both in terms of amino acid pro-
file and the carbon available from sugars, influence the ester and higher alcohol yields
during fermentation. It is worth pointing out that an ester is in equilibrium with its com-
ponent carboxylic acid and alcohol:

R-OH + R ′-CO2H  R ′CO2R + H 2O

Thus, the excretion of acids by yeast can in principle lead to the development of higher
levels of ethyl esters, and higher alcohols can result in more complex esters.
There have been recent reports of certain yeast strains producing low levels of ter-
penes via the mevalonic acid pathway. However, the overall contribution and, therefore,
flavor impact of these terpenes is likely to be minor relative to those sourced from botani-
cals used, say, in the production of gin, absinthe, and spirits derived from fruit-based
fermentations, especially in Saccharomyces cerevisiae fermentations.
S. cerevisiae fermentations also generate small but often significant quantities of
sulfur compounds, including methanethiol (rotting vegetable), dimethyl sulfide (DMS,
sweetcorn), dimethyl disulfide (garlic), dimethyl trisulfide (onion, garlic), propanethiol
(rubber), ethyl thioacetate (cooked vegetable), methionol (raw potato), and methional
(cooked potato). Often unwanted, these compounds are, with the exception of perhaps
DMS, to be avoided. Copper in distillation equipment can scavenge thiols, and DMS is
oxidized over a period of a year or so during maturation, but with these exceptions it is
prudent to manage their levels using fermentation control and during distillation.
The presence of wild yeasts and bacteria during fermentation is often undesirable.
Often, they are present in raw materials and process water. Their presence can cause a
number of problems. For instance, lactic acid bacteria can result in higher levels of diace-
tyl and acidic off-flavors, as well as having an adverse effect on ethanol yield. There are
some exceptions though. Lactic acid bacteria can perform a malo-lactic fermentation to
convert fruit-borne malic acid (tart) into lactic acid (soft, sour) which can be important
for fruit-based spirits, such as brandies. There is also clear evidence that the presence of
adventitious lactic acid bacteria in Scotch whisky fermentation also generates lactic acid,
short-chain fatty acids, and a range of flavor-active lactones (Wilson, 2008).

17.5 DISTILLATION

The production of MRPs as mentioned above can have an impact on the flavor of the
final distillate, presumably due to the need to bring still feed to the boil for subsequent
 Distillates 347

80
70
60

[Ethanol]/% ABV
50
40
30
20
10
0
0 100 200 300 400 500 600
Time (minutes)

FIGURE 17.7 Typical profile of pot distillation of grain-based spirit. The timing for
removal of the early fraction (solid vertical arrow) and the later fraction (dotted vertical
arrow) can have a substantial effect on final spirit quality (adapted with permission from
Buxton and Hughes, 2014, by the Royal Society of Chemistry).

distillation. With some notable exceptions, the volatiles in the still originate from the still
feed. However, some the components in the still feed are too high in concentration for
a final spirit and can have an adverse effect on its quality. To reduce this risk, distilling
practices fall into three categories:

1. Extensive reflux and rectification (e.g., in column stills)

2. Scavenging of undesirable flavors (e.g., using copper)
3. Several distillation stages (typically 2–3)

In any case early- and late-distilling components can be substantially removed by divert-
ing these streams to waste or recycle (Figure 17.7). The early boiling components are typi-
cally compounds such as acetaldehyde, acetone, ethyl acetate, and methanol. The latter
is particularly prevalent in spirits made from stone fruits, grapes, agave, and potatoes,
to the point where it can be harmful, so distillers working with these commodities take
care to ensure adequate methanol removal. This early boiling fraction also contains some
relatively nonvolatile components, such as fatty acids, long-chain esters, phenolics, and
heterocycles. These are the residues from previous distillations which are rinsed out of the
condenser by the heads of the current run.
The switch from heads to spirit production in batch stills tends to be at around
75% ABV. Switching too early means the spirit is likely to contain excessive levels of the
undesirable volatiles mentioned above. Switching too late results in unwarranted losses
of alcohol. The rate of spirit collection has a significant impact on spirit composition and
quality. Heating too rapidly runs the risk of insufficient reflux (i.e., less purification of
the spirit, Watson, 1983) and thermal degradation of the poorly volatile glycerol to form
acrolein (2-propenal), a toxic, lachrymatory compound. Wood maturation of spirits con-
taining significant levels of acrolein does cause a substantial reduction in its levels, but it
can contaminate unmatured spirits if formed.
The decision to switch from spirit to tails collection is made either on the basis of
alcohol content or by nosing the spirit coming from the still. The former works well for
spirit produced consistently, but the sensory method does have merit, especially when
the spirit has a complex composition, such as those produced by distilling botanical
348 Food Aroma Evolution

distillates. Indeed, some distillers may remove a section of spirit mid-run if at that par-
ticular time-point it is deemed to have unacceptable sensory properties.
There are also instances of chemical changes beyond non-enzymic browning reac-
tions in a still, especially if copper is used as a material of construction. Copper can
readily bind free thiol groups, reducing their concentration in the final spirit. Sulfur com-
pounds formed during fermentation, such as hydrogen sulfide (rotten egg), propanethiol
(burned rubber), and methanethiol (rotting vegetable) are often undesirable in spirits,
and so the scavenging properties of copper can improve spirit quality. Additionally, it has
been demonstrated that copper catalyzes the formation of esters during distillation, typi-
cally increasing ester levels by a factor of two, while the concentration of higher alcohols
is essentially unaffected (Watson, 1983). One opportunity to increase copper contact is
to place a still under total reflux in the presence of copper. This is only practically achiev-
able in a column still fitted with a dephleglmator (a small condenser at the top of the
column), which effectively condenses vapors from the top of the column before they pass
to collection. If there is copper in the column, either as a material of construction or as a
packing material, then the additional copper contact under a total reflux regime can help
to remove off-flavors and further stimulate ester formation.
The design of a still has long been believed to affect the composition and quality
of the resulting spirit. Generally, a still with more surface area:volume ratio, with other
aspects being similar (e.g., heat input, state of internal copper surfaces, altitude, decisions
concerning cut-points) will result in a more highly rectified spirit due to greater reflux.
In our laboratory, we observed that there were small but significant differences in the
profiles of % ABV vs. time, with added reflux in the still resulting in a slightly higher
% ABV throughout almost all of the distillation run.
Exploiting chemical reactivity in the still to adjust the distilled spirit flavor was
explored by Rochte (2018). Specifically, an acidic resin was placed in the vapor stream
of the distillation, which allowed the excess butyric acid (cheesy, sweaty) present to react
with ethanol to form the pineapple-like ester ethyl butyrate. This potentially has prom-
ise for both remediating borderline or unacceptable spirits and generating novel flavor
profiles.

17.6 DOWNSTREAM MODIFICATION

There are various ways in which a distilled spirit might be modified (Table 17.5), which
broadly speaking involves introducing elements of botanicals, wood, cream, sugars, or
salts. Each of these contributes different attributes to the final product.
Botanicals can be introduced by either direct addition or after subsequent distilla-
tion. For instance, gin is typically prepared by either soaking (macerating) the botanicals
in spirit then distilling, or by suspending the botanicals above the distilling ethanol/water
mixture (entrainment). Either way, compounds of sufficient volatility are (steam) distilled
from the botanicals and are subsequently recovered in the final spirit. The botanical fla-
vors in gin, derived primarily from juniper berries (or, more accurately, cones), typically
together with coriander seeds, citrus peels, and a range of roots, are primarily terpenoids,
particularly C10 (monoterpenoids) and C15 (sesquiterpenoids), together with some longer-
chain esters (Table 17.6).
The quantity of botanicals used in gin production is typically 4–10 g/L, which, based
on crude estimates of oil content, equates to a maximum of 100–200 mg/L of botanical
oils in gin. Indeed, higher levels can result in the gin forming a haze, or “louching” when
 Distillates 349

TABLE 17.5 Downstream Modifications Commonly Used during Distilled Spirit Production
Approach Major Aroma Contributionsa Example Product Categories
Maceration with botanicals Terpenoids, esters (sugars) Gin, white absinthes, some
and redistillation liqueurs
Maceration with botanicals Terpenoids, esters (sugars, color) Absinthes, some liqueurs
Storage in barrels Esters, higher alcohols, fatty whiskies, brandies, aged
acids, oak lactones (tannins, tequilas, aged rums, aged gins
sugars)
Homogenization with Fatty acids, diacetyl, aldehydes, Cream liqueurs
cream ketones (salts)
Dilution with water to sales (Salts) Most spirits
strength
a Terms in parentheses indicate nonvolatile species.

diluted with an aqueous mixer such as tonic water or cooled/diluted with ice. The levels
of oils in absinthes are substantially higher, perhaps 1 g/L or more. The flavor-active oil
components in absinthe are distinct from gin and, because of their relatively high oil con-
tents, they tend to be marketed at higher alcohol strengths, up to around 74% ABV. In
the context of absinthe louching is desirable, where diluting with ice-chilled water readily
forces a remarkably stable emulsion to form, the principal component being anethole.
There are many other botanical-based spirits, the flavors of which tend to be terpe-
noid- and ester-based. These include the Scandinavian aquavits (flavored with dill and

TABLE 17.6 Common Botanically Derived

Terpenes Found in Spirits
Compound Aroma Attributes
γ-Cadinene Wood
p-Cymene Solvent, gasoline, citrus
Limonene Citrus, lemon, orange
γ-Muurolene Herb, wood, spice
β-Myrcene Balsamic, must, spice
α-Pinene Pine, turpentine
Sabinene Pepper, turpentine, wood
Terpinen-4-ol Turpentine, nutmeg, must
Anethole Anise, fennel
Carvone Mint
Linalool Flower, lavender
Thymol Thyme, woody
Thujone Cedar
Geraniol Rose
Geranyl acetate Rose, geranium
α-Phellandrene Woody, black pepper
β-Caryophyllene Wood, pepper
350 Food Aroma Evolution

FIGURE 17.8 cis- and trans-oak lactones, important flavor-active compounds in most
oak-matured spirits
caraway), pastis and Pernod (flavored with anise), and Dutch/Belgian genevers, where
there is a balance between juniper and the malty flavor of the grain spirit.
Wood maturation is well-recognized as being a major influence on the final sensory
qualities of brown spirits. The contributions of (nonvolatile) color and tannins are essen-
tial for final spirit quality, but in terms of aromas there are three main changes in con-
centrations of flavor-active components. First, the presence of dimethyl sulfide (bp 38°C;
sweetcorn, canned tomato), which originates principally from grains, is substantially lost
after around a year in a cask, being oxidized to the flavor-inactive dimethyl sulfoxide.
Second, acetate is hydrolyzed from the wood surfaces, and this can slowly esterify, as
the release of acetate disturbs the various ester equilibria in which it is involved. Third,
the so-called oak lactones are extracted from the wood (Figure 17.8). These often occur
above their flavor threshold and have flavor descriptors such as coconut and incense.
One further impact of the nonvolatile extractives is their impact on the “structure”
of the alcohol–water matrix. It is well-known that ethanol and water do not form a true
solution above around 17% ABV (Nishimura and Matsuyama, 1989; Nose et al., 2004).
Above this concentration, ethanol has a tendency to form clusters within a depleted alco-
hol continuous phase. The relatively hydrophobic ethanol clusters harbor flavor volatiles
and effectively reduce their aroma activity. Thus, judicious addition of water to a high
strength spirit can indeed release flavor.
Other additions include sugars, cream, and salts. While sugars and salts are not in
themselves appreciably aromatic, there are various reports that at sub-threshold levels
they can have distinct effects on the mouthfeel and other sensory characteristics of dis-
tilled spirits, particularly vodka. The aromas derived from creams include vanillin, diace-
tyl, acetoin, and fatty acid esters, none of which are unique to cream but nonetheless can
make a significant contribution to the flavor of cream liqueurs.

17.7 UNDESIRABLE AROMAS (TAINTS)

While the methods of spirit preparation imply the tailoring of its sensory characteristics
to generate the desired product, adventitious flavors can be deleterious to the quality of a
spirit. The area of food taints has been exhaustively reviewed by Saxby (1996) and spirit
off-flavors by Hughes (2018). Briefly, the presence of one of a range of fungi in casks
can result in the production of trichloroanisole, renowned for its corked or musty taint
in wines closed with contaminated corks. A range of other undesirable fungal-derived
aromas, such as octenone, isoborneol, and geosmin can confer earthy aromas to the
maturing spirit. It is therefore always prudent to “nose” casks before use. Certainly, any
casks found to be contaminated after maturation should be discarded or reworked as it is
unlikely that these compounds can be diluted sufficiently during blending.
 Distillates 351

Another common source of undesirable aromas can come from the poor timing of
cut-points when removing heads and tails from spirit distillations. If the spirit collection
is made too soon, flavor-negative compunds such as acetaldehyde (green apples, sharp)
and ethyl acetate (solvent, nail polish remover) can occur in the final spirit. Extending
spirit collection too far into the tails can result in butyric acid and other fatty acid flavors
(sweaty, cheesy), as well as metallic and leathery aromas.

17.8 FUTURE INSIGHTS

With the blurring of the edges between traditional beer, wine, and spirits categories,
development of rapid maturation techniques and challenges to product definitions (e.g.,
­alcohol-free “whisky”) the distilled spirits industry is innovating probably as fast as it ever
has. The substantial growth in the number of new distilleries from the end of the 20th
century has no doubt fueled innovation across the sector. By way of an example, since
around 2015 the number of wood-aged gin brands has grown to at least 50, ­possibly more.
This is not a new innovation, as gins were moved in barrels during the 18th and 19th
centuries, but rather it is being revisited to create distinction in the category. In the near
future, it is likely that there will be “more of the same” products, crowding an already
busy marketplace.
The move toward low- or zero-alcohol spirits, such as SeedLip and ArKay non-
alcoholic whisky is likely to gain momentum. There are some hurdles to overcome. In
the case of whiskies, the definitions in both the United States and Europe are quite
clear: a whisky should be at least 37.5 and 40% ABV respectively. Arguably difficul-
ties in orientating products like ArKay within established high-value sectors such as
whisky could limit their appeal. For novel products, there are also options for more
radical formulation strategies, for instance, in terms of product formulation (com-
pounding) and dealcoholization. Restricting alcohol production during fermentation
is an unusual strategy for a distiller. But for low- or non-alcoholic products there is
some merit to applying beer brewing approaches, such as high-temperature mashing,
which could gain a foothold in this sector, along with contemporary alcohol removal
technologies such as spinning cone columns, as applied in the wine and kombucha
industries.

REFERENCES

Baxter, E.D. and Hughes, P.S., Beer—Quality, Safety and Nutritional Aspects, Royal
Society of Chemistry, Cambridge, 2001.
van Boekel, M.A.J.S., Formation of flavour compounds in the Maillard reaction,
Biotechnology Advances, 2006, 24, 230–3.
Boulton, C. and Quain, D., Brewing Yeast and Fermentation, Wiley-Blackwell, Hoboken,
NJ, 2006.
Burdock, G.A. and Fenaroli, G., Fenaroli’s Handbook of Flavor Ingredients, CRC Press,
Boca Raton, FL, 2010.
Buxton, I. and Hughes, P.S., The Science and Commerce of Whiskies, Royal Society of
Chemistry, Cambridge, 2014.
Hughes, P.S., Stabilization of distilled spirit, in Post-Fermentation and Distillation
Technology, ed. Bordiga, M., CRC Press, Boca Raton, FL, 2018, pp. 235–49.
352 Food Aroma Evolution

Inoue, T., Diacetyl in Fermented Foods and Beverages, American Society of Brewing
Chemists, St Paul, MN, 2008.
Malnic, B., Hirono, J., Sato, T., and Buck, L.B., Combinatorial receptor codes for odors,
Cell, 1999, 96, 713–23.
Nishimura, K. and Matsuyama, R., Maturation and maturation chemistry, in The Science
and Technology of Whiskies, eds. Piggott, J.R., Sharp, R., and Duncan, R.E.B.,
Longman, Harlow, 1989, pp. 235–63.
Nose, A., Hojo, M., Suzuki, M., and Ueda, T., Solute effects on the interaction between
water and ethanol in aged whiskey, Journal of Agricultural and Food Chemistry,
2004, 52, 5359–65.
Nykanen, L. and Suomalainen, H., Aroma of Beer, Wine and Distilled Alcoholic
Beverages, Reidel, Dordrecht, 1983.
Piggott, J.R., Flavor of Distilled Beverages, Ellis Horwood, Chichester, 1983.
Piggott, J.R. (ed.), Alcoholic Beverages—Sensory Evaluation and Consumer Research,
Woodhead, Philadelphia, PA, 2012.
Rochte, J.D., Distilled Alcoholic Beverage Production Using Reactive Distillation
Techniques, PhD Thesis, Michigan State University, 2018.
Saxby, M.J. (ed.), Food Taints and Off-Flavors, 2nd ed., Springer, New York, NY, 1996.
Watson, D.C., A laboratory apparatus for distillation research, in Proceedings of Current
Developments in Malting, Brewing and Distilling, Institute of Brewing, 1983, pp.
249–55.
Wilson, N.R., The Effect of Lactic Acid Bacteria on Congener Composition and Sensory
Characteristics of Scotch Malt Whisky, PhD Thesis, Heriot-Watt University,
Edinburgh, 2008.
 Chapter 18
Evolution of Beer Aroma
Iztok Jože Košir and Miha Ocvirk

CONTENTS

18.1 Introduction 353

18.2 Aroma Compounds from Hop Essential Oils 354
18.2.1 Hydrocarbon Fraction 355
18.2.2 Alcohols 355
18.2.3 Aldehydes 356
18.2.4 Other Aroma Compounds in Hop Essential Oils 356
18.3 Aroma Compounds Produced during the Brewing Process 357
18.3.1 Higher Alcohols (Fusel Alcohols) 358
18.3.2 Esters 358
18.4 Dimethyl Sulfide 359
18.5 Diacetyl 359
18.6 Phenols 361
18.7 Conclusion 361
References 361

18.1 INTRODUCTION

Typical beer flavor and aroma are the result of a fine and subtle balance between numer-
ous flavor-active compounds that originate from the raw materials used in the brewing
process, together with those originating from yeast during fermentation. On the other
hand, we should not neglect the two groups of volatiles, which can also have an effect on
the beer’s final aroma in both a negative and positive way. Groups of volatiles caused by
microorganism contamination normally impact negatively on beer aroma, but for some
beer types (Lambic), such aromas are desirable. Another group of compounds that can
negatively affect consumers is the stability of volatiles in beer, and their reaction degra-
dation products. A combination of taste and odor is of crucial importance to consumer
acceptance of beer (Ocvirk et al., 2018; Van Opstaele et al., 2010; Jeleǹ et al., 1998)
One volatile component we cannot neglect, and should be mentioned first, is ethanol,
since it is quantitatively second after water. It contributes to the beer’s aroma directly
with its own aroma, which is characterized as warmy, but also influences the contribu-
tion of all other volatiles. Normally, it affects the vapor pressure of other substances and
also the related vitalization. It also has effects on solubility and the reactions among vola-
tile compounds. According to the German Beer Purity Law (“Reinheitsgebot”) from the
year 1516, the only ingredients that could be used in the production of beer were water,
barley, and hops. At that time, the knowledge of yeast’s existence had not yet developed.

353
354 Food Aroma Evolution

Barley is the most abundant raw material in the brewing process. The mashing of ground
barley malt is the first step in the brewing process. Mash is heated to different tempera-
tures to allow particular enzymes to optimally degrade the main malt constituents, such
as beta-glucans, proteins, dextrins, and starch. Temperatures of the mash start at around
45°C and go up to 72°C in four main temperature stages. Volatiles which are responsible
for a malty (worty, husky) aroma can be divided into four groups: volatiles derived from
the oxidation of lipid precursors, volatiles formed in the Maillard reaction, aliphatic
sulfur compounds, and phenols. Wort is actually a growing medium of yeast cells, where
they absorb nutrients, which are vital for living and reproduction. According to the beer
flavor wheel, developed by Dr. Morten Meilgaard in the 1970s, which become a standard
around the world, flavors from barley are divided into cereal, caramelized (roasted), and
phenolic flavors (Meilgaard et al., 1979).

18.2 AROMA COMPOUNDS FROM HOP ESSENTIAL OILS

In the brewing process, quantitatively, hop is a minor ingredient, but of paramount impor-
tance to the brewing industry. With their complex chemistry, hops have been a subject of
investigation for decades (Almaguer et al., 2014). During the brewing process, hops are
added into the boiling wort to provide a bitter taste and aroma to the final product (Šterba
et al., 2015). One of the main reasons for boiling the wort is the isomerization reaction of
the hop alpha-acids into their isomerized forms. The duration of kettle hopping depends
on the time required for the isomerization to take place (Malowicki and Shellhammer,
2005). The addition of hops into the boiling wort is essential with the intent of achieving
the desired bitterness; however, it causes a reduction in the yield of essential oils in the
beer due to evaporation. Consequently, these components are not present in beer in the
same ratios as they are in hops. Some components are highly volatile, and some have low
solubility (Inui et al., 2013). Kettle boiling is also important for the reduction of unde-
sired volatile compounds originated from malt, for example, dimethyl sulfide.
Dry hops normally contain from 0.5 to 4.0% essential oils. Essential oils are con-
stituted of the secondary metabolites of the hop plant (Humulus lupulus L.), located
in the lupulin glands in the hop cones. It has been known for centuries that hop cones
are very aromatic, due to the volatile oil fraction. The hop aroma was first described by
Loiseleur-Deslongchamp as garlic-like. At the beginning of the 20th century, Chapman
did the first systematic examination of hop essential oil (Almaguer et al., 2014). It is
now known that depending on the chemical structure these compounds can be divided
into three groups: the most abundant is the hydrocarbon fraction which forms approxi-
mately 75% of the total oil, the oxygen-bearing compounds forming approximately
25% of the total oil, and the sulfur-containing compounds that are present in much
lower quantities. Generally, myrcene, which is the most abundant component in hop
essential oil, is undesirable in beer, while linalool and geraniol—both from an oxy-
gen fraction present in much lower concentrations than myrcene—provide a pleasant
aroma. The odor threshold is more important than the concentration level of a par-
ticular component (Šterba et al., 2015; Fix, 1999; Bamforth, 2006). Many of the com-
ponents of the essential oils, even if they are present in hops in high concentrations
(for example, myrcene, up to 75% in hops), would not pass the brewing process and
do not contribute to the final beer aroma due to the evaporization, degradation, or
chemical reactions, since concentrations are normally far below the odor threshold
level (Coelhan and Aberl, 2012; Fix, 1999).
 Evolution of Beer Aroma 355

With further investigation and with progress in analytical technology as well as the
increase of knowledge, it is now suggested that there may be over 1000 different compounds,
but only 440 of them have been chemically identified (Almaguer et al., 2014; Roberts et al.,
2004). The composition of hop essential oil is mostly genetically dependent; thus, it can also
be used as an effective tool for characterization of hop varieties (Ocvirk et al., 2016).

18.2.1 Hydrocarbon Fraction

The hydrocarbon fraction that is present in hop oil consists of three groups: monoterpenes,
sesquiterpenes, and aliphatic hydrocarbons. The hydrocarbon group has a low solubility
in a water medium, and consequently in beer as well. During the boiling process, most of
these components can be lost due to their low boiling points. These compounds can eas-
ily oxidize and polymerize into other forms (Sharpe and Laws, 1981). Monoterpenes are
divided into three groups: the acyclic, the monocyclic, and the bicyclic. The most abun-
dant acyclic monoterpene in hop oil is β-myrcene, which can be present in hop oil (up to
75% of the total oil content). Myrcene was first obtained from bay (Laurus nobilis) oil in
1895 by Power and Kleber. It only took a few years to realize that one of the components
in hop essential oils is the same as that from the bay plant. Myrcene, with its sharp smell, is
responsible for the smell of fresh hops. Besides myrcene, acyclic monoterpenes are cis and
trans isomers of β-cymene. p-cymene, α-phellandrene, β–phellandrene, and limonene are
representative of monocyclic terpenes. α-pinene, β–pinene, and sabinene are representative
of bicyclic monoterpenes. Sesquiterpenes in hop oil are acyclic, monocyclic, bicyclic, and
tricyclic. Farnesene is actually the only acyclic sesquiterpene which can be found in hop
oil. Monocyclic sesquiterpenes are β-germacrene, germacrene-D, and α-humulene, which
can be found in hop oil up to 30% of the total oil content. β-cariophyllene (up to 20%),
α- and β-selinene, selinadiene and α-, γ-, δ-, and δ2-cadinene are bicyclic sesquiterpenes.
The tricyclic sesquiterpenes are α-ilangene, α-copaene, and β-copaene. In hop oil, some
diterpenes also can be found. Para- and meta- camphorene originate from a Diels–Alder
reaction from two myrcene molecules. Over the last decade, with better laboratory equip-
ment, new sesquiterpenes were identified: armorphene, curcumene, and bergamotene, but
only in trace amounts. (Almaguer et al., 2014; Kovačevič, 2001; Sharpe and Laws, 1981).
Components with oxygen form approximately one-quarter of the total hop oil. Although
most of these components are present in hop essential oil in trace amounts, these components
are of crucial importance for consumer acceptance of beer aroma. This group of components
with oxygen can be subdivided into the following subgroups according to their chemical
structure on alcohols, aldehydes, ketones, acids, esters, epoxides, and some others. Another
classification of oxygen-bearing compounds into volatile and nonvolatile classes can be found
in the literature. The last are especially important and interesting for brewers because they
can “survive” the boiling process, and they can find an easy way in to beer. During aging,
oxidation processes occur, and hop oil fraction becomes richer in oxygenate compounds. It is
known that over 40 compounds can be produced from myrcene auto-oxidation (Sharpe and
Laws, 1981; Almaguer et al., 2014; Hanke et al., 2008; Coelhan and Aberl, 2012).

18.2.2 Alcohols

The list of different alcohols present in hop essential oil is increasing yearly. Alcohols can be
divided into terpene alcohols, sesquiterpene alcohols, and aliphatic and aromatic alcohols.
356 Food Aroma Evolution

Linalool is the most abundant terpene alcohol in hop oil. Nowadays, it is known that lin-
alool contributes significantly to the floral flavor of beer and is a positive indicator of hop
quality, but it is not entirely clear if levels of free linalool in the essential oil of hops are in
a direct relationship with the concentration of linalool in beer (Čerenak et al., 2011). Post-
linalool aroma depends on the time of hop exposure to elevated temperatures, which leads
to racemization and evaporation, and on the presence of yeast enzymes for degradation of
linalool-containing glucosinolate (Ocvirk and Košir, 2014). Free linalool in beer is degraded
rapidly (Kollmannsberger et al., 2006), and its contribution to beer aroma depends on the
time of hopping. With its low boiling point, high concentrations in beer are unlikely when
the traditional hopping of wort is in use. Other important compounds are 2-methylbutanol,
geraniol, nerol, terpineol, mirtenol, and myrcenol. The sesquiterpene alcohols are nerolidol,
humulol, α-eudesmol, juneol, and humulenol-II. Aliphatic and aromatic alcohols in hop
oils are 2-propanol, butanol, pentanol, heksanol, heptanol, octanol, and nonanol. In addi-
tion to extraction into the wort through the hopping process, the other source of these is
the chemical reaction of the components in which alcohols can be formed. Linalool can be
formed from nerol or geraniol, and beta-citronellol from geraniol. Linalool could also be
cyclized to alpha-terpineol (Riu-Aumatell et al., 2014; Takoi et al., 2010). Although linalool
is not abundant in hops, its contribution to beer aroma is high because of its very low odor
threshold value of only 2.2 µg L –1 (Steinhaus and Schieberle, 2000).

18.2.3 Aldehydes

Aldehydes are responsible for the grassy notes in beer aroma. There is a wide range of
different aldehydes, such as hexanal, heptanal, octanal, benzaldehyde, geranial, neral,
and so on. A large number of ketones present in hop essential oils were also identified,
and acetone is certainly the most common. Humuladienone is supposed to be responsible
for the hoppy aroma in beer. The true brewing value of this compound is still a mystery
because of its high boiling point (Steinhaus and Schieberle, 2000).

18.2.4 Other Aroma Compounds in Hop Essential Oils

There are only a few free organic acids present in hop oil: 3-methylbutanoic acid, 2-meth-
ylpropanoic acid, 4-methyl-3-pentenoic acid, and 4-hydroxi-4-methyl-2-pentenoic acid.
Present epoxides are products of the auto-oxidation of β-cariophyllene and
α-humulene. This is cariophylene oxide, humulene epoxide-I, humulene epoxide-II, and
humulene diepoxide. (Steinhaus and Schieberle, 1981; Kovačevič, 2001.)
Esters are compounds which contribute greatly to hop aroma. There are a lot of dif-
ferent types of esters which are found in beer. Straight esters, saturated branched-chain
esters, or even unsaturated esters. There are many methyl esters up to C13 carboxylic
acids. There were also some esters of terpene alcohol discovered.
Sulfur-containing components have, not surprisingly, a negative effect on beer aroma.
Luckily their amounts are in traces, but their odor threshold is very low—0.1 ppb (Seaton
et al., 1981), so they can easily affect the final aroma of hops and beer. Around 30 sul-
fur-containing compounds were identified in hop oil. The most abundant components are
hydrogen sulfide, dimethylsulfide, methional, and so on, and they have the biggest impact
on beer aroma. Many of these compounds are originated from hop oil components and
sulfur that are normally present in the environment (elementary sulfur or sulfur dioxide) or
 Evolution of Beer Aroma 357

TABLE 18.1 Concentration Range, Odor Threshold in Beer, and Chemical Formula of Some
Components in Hop Essential Oils: Concentration Can Vary Greatly, Depending on the Hop
Cultivar
Chemical Concentration Range Odor Threshold
Formula in Hop Oil (rel. %) in Beer (ppb) Structural Formula
Myrcene C10H16 10–75 30–200

Linalool C10H18O 0.2–1,3 2.2

α-Humulene C15H24 15–42 120

β-Caryophylene C15H24 2.5–18

Farnesene C15H24 <1 NA

are used in various forms of plant protection products. Polysulfides (e.g., DMS) are known
as having the most negative influence on beer aroma and are considered as off-flavors. In
Table 18.1, the five most important hop essential oil components are presented.

18.3 AROMA COMPOUNDS PRODUCED

DURING THE BREWING PROCESS

Beers could be divided into two main groups, depending on the type of yeast strain
used. Lager beers are produced with a yeast strain, known as Saccharomyces pastorianus
or Saccharomyces carlsbergensis, where fermentation runs at low temperatures (around
12°C), while fermentation of ale beers runs with Saccharomyces cerevisiae at tempera-
tures around 22°C. The unique flavor profiles of beer depend on the biochemical activi-
ties during fermentation within the yeast cell, where conversion to volatile compounds
occurs (Olaniran et al., 2017).
Most of these components are formed during alcoholic fermentation and consist of the
metabolic by-products of yeast (Jeleǹ et al., 1998). Although these components are minor
in beer, they have great importance to aroma, because of their low odor thresholds – the
lowest concentration that is perceptible by the human sense of smell. Aroma compounds
358 Food Aroma Evolution

in beer contribute to the quality of beer as a final product (Da Silva et al., 2012). Their
influences on beer aroma are not in proportion with their concentrations. Key flavor-active
components that are produced by yeast and specify beer aroma are mostly higher alcohols
and esters (Charry-Parra et al., 2011; Pires et al., 2014; Bamforth, 1999; Fix, 1999).
Higher alcohols and esters can have both positive and negative impacts on overall
aroma and flavor. That’s why it is believed that all volatiles in beer must be within a
certain concentration range or else some compound may prevail and destroy the balance
(Ocvirk et al., 2018). For example, if the concentration of 2-methylpropanol surpasses
20% of the total amount of n‐propanol, 3-methylbutanol, and 2-methylpropanol, the
quality of beer can be marked as not pleasant.

18.3.1 Higher Alcohols (Fusel Alcohols)

Higher alcohols are the most represented compounds in beer (Pires et al., 2014), and
they are produced from amino acids in two ways. First, from wort through the Ehrlich
mechanism, where amino acids that are present in wort from the malt are absorbed by
yeast. Yeast cells incorporate the taken amino group in its structure. The rest of the
amino acid goes through an irreversible reaction, and the products of these reactions
are the by-product of higher alcohols. The second way of forming higher alcohols is
during sugar metabolism (a biosynthetic mechanism) (Fix, 1999). The most abundant
higher alcohols in beer are n-propanol, isobutanol, 2-methylbutanol, 3-methylbutanol,
and 2-phenylethanol (Fix, 1999; Pires et al., 2014; Thompson-Witrick et al., 2015). Some
higher alcohols can be present in beer in high amounts (>300 mg/L) and can lead to
strong smells, so-called off-flavors. One of the most important volatiles in this group is
3-methylbutanol. With increasing concentrations, the beer aroma becomes heavier, and
this impacts negatively on beer drinkability (Ademola et al., 2017).
The content of higher alcohols, as well as esters, depends mainly on the type of cere-
als used in the brewing process. High concentrations of amino acids in wort increases the
production of higher alcohols. In the synthesis of higher alcohols and esters, the tempera-
ture of fermentation is also a key factor. The concentration of 1-propanol is the highest
in barley beers and the lowest in beer-based beverages, like those made with quinoa, for
example. High values of free amino nitrogen compounds lead to low values of 1-propa-
nol. 2-oxo acids, an intermediate of leucine and isoleucine, are precursors in the anabolic
route of formation of 2- and 3-methylbutanol. The higher the concentrations of leucine
and isoleucine that are in the wort, the lower the values of 2- and 3-methylbutanol that
are in the beer. Another important higher alcohol is 2-phenylethanol, which can suppress
the flavor of dimethyl sulfide. (Deželak et al., 2014)

18.3.2 Esters

Esters are responsible for the fruity/solvent aroma in beer. The formation of esters is
mainly dependent on the type of yeast strain used in the fermentation process and the
type of wort (cereals). The concentration of ethyl esters is also time dependent. Some ester
concentrations increase during the storage of beer (Deželak et al., 2014). Concentrations
of esters in beer are related to the concentration of glucose in wort. The higher the sugar
concentration in wort is, the higher the amounts of esters are. Iso-amyl acetate (banana-
like flavor), ethyl acetate (sweet, solvent-like flavors), 2-phenylethyl acetate (flowery and
 Evolution of Beer Aroma 359

honey-like flavors), iso-butyl acetate (fruity aromas), ethyl hexanoate (red apple aromas),
and phenyl ethyl acetate (rose aromas) are found in beer in trace concentrations, but due
to very low odor threshold concentrations they have a big impact on the final aroma.
Acetaldehyde is a highly volatile aldehyde, which is normally present in beer together
with higher alcohols and esters. Due to its low odor threshold and high volatilization,
it can have a decisive impact on beer aroma. Higher concentrations of esters can have a
negative impact on aromas such as the fruity, sweet flavors of beer.
Ester compounds are the result of a reaction between higher alcohols and organic
acids; they are minor elements in beer, but because of their very low individual odor
thresholds they are the most important aroma elements (Meilgaard, 1975; Pires et al.,
2014; Bamforth, 1999; Fix, 1999). More than 90 esters can be found in beer, but the most
significant among them are ethyl acetate, isoamyl acetate, ethyl octanoate, ethyl decano-
ate, and 2-phenylethyl acetate. Since the aroma is defined by the synergy of multiple com-
pounds, small changes in ester concentrations have a big effect on beer aroma, because
esters in beer are present at around their odor threshold concentrations (Thompson-
Witrick et al., 2015). There are two types of esters found in beer: acetate esters (produced
over acetic acid and ethanol) and medium-chain fatty acid ethyl esters (MCFA), where
the MCFA group is attached to the alcohol radical. Esters are formed during alcoholic
fermentation, but the ester content in beer changes through time (during beer aging), and
it is not necessary that the amounts are reduced. There are two ways of changing the
ester profile: refermentation (by yeast) or by condensation of organic acids with ethanol.
The winy aroma in aged beers is a result of the esterification of methyl butyric acid. It is
known that fruity notes are reduced in aged beers, where sweeter notes prevail. In Table
18.2, some of the most abundant higher alcohols and esters are presented. Concentration
can vary greatly, depending on the yeast strain used in the fermentation process and con-
tent and type of brewing malt (MEBAK, 1996).

18.4 DIMETHYL SULFIDE

Dimethyl sulfide or DMS is one of the most “famous” off-flavors in beer. When this
highly volatile substance is present in beer, even if its concentration is very low, it can
overwhelm other aromas. This happens because of its very low odor threshold in beer.
Generally, DMS can be present in all beer types, as a consequence of the mashing pro-
cedure. It originates from a precursor, S-methylmethionine (SMM), which originates
from the germination of barley. With sustained and vigorous wort boiling, the DMS
can be eliminated or at least reduced to low concentrations, since SMM is heat sensi-
tive. Dimethyl sulfoxide (DMSO) is another source of DMS. DMSO can be transformed
into DMS during fermentation. It is known that calm fermentation positively affects the
formation of DMS. Therefore, light lagers have a higher potential for a DMS off-flavor
(Carpenter, 2015). DMS can be easily detected by the smell of sweetcorn.

18.5 DIACETYL

Vicinal diketones (VDKs) are the products of fermentation. They belong to the organic
group of ketones with an adjacent (vicinal) carbon atom. The most common are 2,3-butane-
dione, also known as diacetyl and 2,3-pentanedione, known as acetyl propionyl. Vicinal
diketones are undesired in beer since they are responsible for a butter-like flavor in the
360 Food Aroma Evolution

TABLE 18.2 Usual Concentration Range of Some Higher Alcohols and Esters in
Lager Beers with Original Extract 11–13%: Concentration Can Vary Greatly,
Depending on the Yeast Strain Used in the Fermentation Process and Content
and Type of Brewing Malt
Chemical Concentration Range Structural Formula
Formula in Beer (ppm)
Acetaldehyde C2H4O 2–10

Ethyl acetate C4H8O2 10–40

n-Propanol C3H8O 5–20

Isoamyl acetate C7H14O2 0.5–3

3-Methyl-1-butanol C5H12O 30–70

2-Phenylacetate C10H10O2 0.1–2

3-Phenylethanol C8H10O 8–40

2-Methylbutanol C5H12O 8–30

Isobutanol C4H10O 5–20

Source: MEBAK, 1996.

beverages. Although VDKs are detectable by humans at fairly low levels, VDKs also play
an important role in the formation of beer aroma during beer maturation. In lagers, diace-
tyl is normally present in the range of 20 to 60 ppb. Concentrations are higher in darker
beers. Brewers around the world use a concentration of diacetyl level as a quality control
parameter for monitoring the technological process. The formation of VDKs is yeast strain
dependent. A well-known way to control and reduce diacetyl concentrations is the so-called
diacetyl rest after fermentation. High diacetyl levels in beer may also be a consequence of
an infection by Pediococcus or Lactobacillus bacteria, and excess oxygen causes diacetyl
production via a redox reaction. (Fix, 1999; White, n.d.)
 Evolution of Beer Aroma 361

18.6 PHENOLS

Volatile phenols in beer originate in the extraction from plant materials, primarily malt
but also from hops. The contents and compositions of phenolic acids vary greatly among
the malt and hop varieties used in the brewing process. The most dominant phenolic
acids are ferulic acid and p-coumaric acid. Hydrocinnamic acids are also present in wort.
Due to their very high flavor threshold, they are not important for beer aroma, since they
are present in wort and beer in ranges below their threshold. However, they can be con-
verted into vinyl and ethyl derivates, with low thresholds and therefore can significantly
contribute to the final aroma of the beer. For example, 4-vinylguaicol and 4-vinylphenol
are decarboxylation products of ferulic acid and p-coumaric acid. The most dominant
volatile phenols in beer are 4-vinylguaicol and 4-ethylguaicol. These components are
responsible for a horsey, leathery, smoky, stable, and of course medicinal aroma in beer
and are all recognized as off-flavors (Lentz, 2018)

18.7 CONCLUSION

Volatile and nonvolatile compounds which are responsible for beer aroma are a quality
parameter and also a parameter of consumers’ decisions. They do not originate only from
raw materials, but also originate as a fermentation or maturation product. Although they
are present in beer at very low concentrations, their odor activity is high because of their
low odor thresholds.

REFERENCES

Almaguer, C., Schönberger, C., Gastl, M., Arendt, K.E., and Becker, T. 2014. Humulus
lupulus—A story that begs to be told. A review. J. Inst. Brew. 120: 289–314.
Bamforth, C.W. 2006. Science Principles of Malting and Brewing, American Society of
Brewing Chemists, St. Paul, MN, pp. 16.
Carpenter, D. 2015. https​://be​erand​brewi​ng.co​m /off​-flav​or-of​-the-​week-​dms/.​
Čerenak, A., Pavlovič, M., Oset Luskar, S., and Košir, I.J. 2011. Characterisation of
Slovenian hop (Humulus lupulus l.) varieties by analysis of essential oil. Hop Bull.
18: 27–32.
Charry-Parra, G., De Jesus-Echevarria, M., and Perez, F.J. 2011. Beer volatile analysis:
Optimization of HS/SPME coupled to GC/MS/FID. J. Food Sci. 76: 205–11.
Coelhan, M. and Aberl, A. 2012. Determination of volatile compounds in different hop
varieties by headspace-trap GC/MS in comparison with conventional hop essential
oil analysis. J. Agric. Food Chem. 60: 2785–92.
Da Silva, G.A., Maretto, D.A., Augusto, F., Poppi, R.J., Bolini, H.M.A., and Teófilo, R.F.
2012. Correlation of quantitative sensorial descriptors and chromatographic signals
of beer using multivariate calibration strategies. Food Chem. 134: 1673–81.
Deželak, M., Zarnkow, M., Becker, T., and Košir, I.J. 2014. Processing of bottom-­
fermented gluten-free beer-like beverages based on buckwheat and quinoa malt with
chemical and sensory characterization. J. Inst. Brew. 120: 360–70.
Fix, G.A. 1999. Wort Boiling in Principles of Brewing Science—A Study of Serious
Brewing Issues, 2nd ed., Brewers Publications, Boulder, CO, pp. 53–78 and 79–126.
362 Food Aroma Evolution

Hanke, S., Herrmann, M., Rückerl, J., Schönberger, C., and Back, W. 2008. Hop volatile
compounds (part II): Transfer rates of hop compounds from hop pellets to wort and
beer. Brew. Sci. 61: 140–4.
Inui, T., Tsuchiya, F., Ishimaru, M., Oka, K., and Komura, H. 2013. Different beers with
different hops. Relevant compounds for their aroma characteristics. J. Agric. Food
Chem. 61: 4758–64.
Jeleǹ, H.H., Wlazɫy, K., Wąsowitz, E., and Kamiǹski, E. 1998. Solid-phase microextrac-
tion for the analysis of some alcohols and esters in beer: Comparison with statistic
headspace method. J. Agric. Food Chem. 46: 1469–73.
Kollmannsberger, H., Biendl, M., and Nitz, S. 2006. Occurence of glycosidically bound
flavour compound in hops, hop products and beer. Monatsschrift Brauwiss. 59:
83–9.
Kovačevič, M. 2001. Določanje sestave eteričnega olja hmelja za razlikovanje med
hmeljnimi kultivarji. Magistrsko delo, Univerza v Ljubljani, Fakulteta za kemijo in
kemijsko tehnoloijo.
Lentz, M. 2018. The impact of simple phenolic compounds on beer aroma and flavor.
Fermentation 4: 20.
Malowicki, M.G. and Shellhammer, T.H. 2005. Isomerization and degradation kinet-
ics of hop (Humulus lupulus) acids in a model wort-boiling system. J. Agric. Food
Chem. 53: 4434–9.
MEBAK. 1996. Gärun​gsneb​enpro​dukte ​— Dest​illat​ionsm​ethod​e, Method 1.1.2,
MEBAK Band III, 2 Auflage, MEBAK, Freising, pp. 5–8.
Meilgaard, M.C. 1975. Flavour chemistry of beer. Part II: Flavor and threshold of 239
aroma volatiles. MBAA Technol. Q. 12: 151–68.
Meilgaard, M.C., Dalgliesh, C.E., and Clapperton, J.F. 1979. Beer flavour terminology.
J. Inst. Brew. 85: 38–42.
Ocvirk, M., Kočar Mlinarič, N., and Košir, I.J. 2018. Comparison of sensory and chemi-
cal evaluation of lager beer aroma by GC and GC/MS. J. Sci. Food Agric. 98(10):
3627–35.
Ocvirk, M. and Košir, I.J. 2014. Pomen linalola v pivovarstvu. Hop Bull. 21: 68–73.
Ocvirk, M., Košir, I.J., and Grdadolnik, J. 2016. Determination of the botanical origin
of hops (Humulus lupulus L.) using different analytical techniques in combination
with statistical methods. J. Inst. Brew. 122: 452–61.
Olaniran, A.O., Hiralal, L., Mokoena, M.P., and Pillay, B. 2017. Flavour‐active volatile
compounds in beer: Production, regulation and control. J. Inst. Brew. 123: 13–23.
Pires, E.J., Teixeira, J.A., Brányik, T., and Vicente, A.A. 2014. Yeast: The soul of beer’s
aroma—A review of flavour-active esters and higher alcohols produced by the brew-
ing yeast. Appl. Microbiol. Biotechnol. 98: 1937–49.
Riu-Aumatell, M., Miró, P., Serra-Cayuela, A., Buxaderas, S., and Lopez-Tamames,
F. 2014. Assessment of the aroma profiles of low-alcohol beers using HS-SPME-
GC-MS. Food Res. Int. 57: 96–202.
Roberts, M.T., Dufour, J.P., and Lewis, A.C. 2004. Application of comprehensive multi-
dimensional gas chromatography combined with time-of-flight mass spectrometry
(GC/GC-TOFMS) for high resolution analysis of hop essential oil. J. Sep. Sci. 27:
473–8.
Seaton, J.C., Suggett, A., and Moir, M. 1981. The flavour contribution of sulphur com-
pounds in hops. MBAA Technol. Q. 18: 26–30.
Sharpe, F.R. and Laws, D.R.J. 1981. The essential oil of hops a review. J. Inst. Brew. 87:
96–107.
 Evolution of Beer Aroma 363

Steinhaus, M. and Schieberle, P. 2000. Comparison of the most odor-active compounds

in fresh and dried hopcones (Humulus lupulus L. variety Spalter Select) based on
GC-olfactometry and odor dilution techniques. J. Agric. Food Chem. 48: 1776–83.
Šterba, K., Čejka, P., Čulik, J., Urkova, M., Krofta, K., Pavlovič, M., Mikyška, A., and
Olšovska, J. 2015. Determination of Linalool in different hop varieties using a new
method based on fluidized—Bed extraction with gas chromatographic mass spec-
trometric detection. J. Am. Soc. Brew. Chem. 73: 151–8.
Takoi, K., Koie, K., Itoga, Y., Katayama, Y., Shimase, M., Nakayama, Y., and Watari,
J. 2010. Biotransformation of hop-derived monoterpene alcohols by lager yeast and
their contribution to the flavor of hopped beer. J. Agric. Food Chem. 58: 5050–8.
Thompson-Witrick, K.A., Rousef, R.L., Cadawallader, K.R., Duncan, S.E., Eigel, W.N.,
Tanko, J.M., and O’Keefe, S.F. 2015. Comparison of two extraction techniques,
solid‐phase microextraction versus continuous liquid–liquid extraction/solvent‐
assisted flavor evaporation, for the analysis of flavor compounds in Gueuze Lambic
Beer. Food Sci. 80: 571–6.
Van Opstaele, F., De Rouck, G., De Clippeleer, J., Areas, G., and De Cooman, L. 2010.
Analytical and sensory assessment of hoppy aroma and bitterness of conventionally
hopped and advanced hopped pilsner beers. J. Inst. Brew. 116: 445–58.
White, C. n.d. Controlling diacetyl. https​://by​o.com​/arti​cle/b​rewin​g-sci​ence-​contr​ollin​
g-dia​cetyl ​/.
 Chapter 19
Coffee Flavor
Sergio Pérez-Burillo, Matteo Bordiga, Silvia
Pastoriza, and José A. Rufián-Henares

CONTENTS

19.1 Introduction 365

19.2 Coffee Production 366
19.2.1 The Fruit 366
19.3 Raw Bean Composition and Flavor Precursor Formation 368
19.4 Coffee Processing: Fermentation and Roasting 373
19.4.1 Influence of Fermentation on Aroma and Different Ways of
Carrying It Out 374
19.4.2 Modulation of Coffee Aroma by Single Microorganisms 376
19.4.3 Influence of Roasting 376
19.4.3.1 Thermal Degradation of Precursors 377
19.4.3.2 The Maillard Reaction 377
19.5 The Impact of the Drying and Storage Processes 377
19.5.1 Drying 378
19.5.2 Storage 378
19.6 Conclusions 378
References 379

19.1 INTRODUCTION

Global coffee consumption reaches about 150 million cups per year and, despite
aspects such as price fluctuations, trade restrictions, and overall inflation, the demand
for both high-quality and specialty coffee is constantly growing (International Coffee
Organization, 2018). The success and popularity of coffee products are related to their
distinctive sensory and pleasant flavor. A key factor of coffee beverage quality is repre-
sented by the postharvest practices applied to obtain dried beans suitable for roasting
(Huch and Franz, 2015). These practices include some complex phases such as harvest-
ing, depulping, drying, and storage. Once on-farm processed, coffee beans can be trans-
ferred to industrial plants, where different products (e.g., semi-manufactured or finished)
are obtained for commercialization (Pereira et al., 2017). Scientific research has focused
on the study of coffee (mainly aroma) for almost a century. To date, over 1,000 volatile
compounds form the complex volatile aroma composition of coffee (Lee and Shibamoto,
2002; Sanz et al., 2002). Thermal reactions, which occur during roasting and brew-
ing, are responsible for generating these compounds. Among them, sulfur compounds,
pyrazines, pyridines, pyrrols, furans, aldehydes, higher alcohols, ketones, and esters
are the most representative (Buffo and Cardelli-Freire, 2004; Rodriguez, Duran, and

365
366 Food Aroma Evolution

Reyes, 2010). It must be noted, however, how the chemical composition of green beans
may affect the quality and value of the coffee. Similarly, chemical modifications also
occur through postharvest processing (Bhumiratana, Adhikari, and Chambers, 2011;
Sunarharum, Williams, and Smyth, 2014). However, how the final coffee quality may
be influenced by different factors such as genotype, cultivation method, and postharvest
treatment is still under study (Knysak, 2017; Pereira et al., 2016).

19.2 COFFEE PRODUCTION

More than 100 species of tropical trees and shrubs of the genus Coffea produce the
berries from which coffee is extracted (Ferrão et al., 2015). The coffee tree is cultivated
throughout the geographic region known as the “coffee belt” located between latitudes
30° N and 30° S. Among coffee producers, Brazil is one of the industry leaders, produc-
ing about a third of the world’s total coffee products. Other main producers are Vietnam,
Indonesia, Colombia, India, Peru, Honduras, Ethiopia, Guatemala, and Mexico, fol-
lowed by more than 50 other countries (minor producers) (Pereira et al., 2017). The
global production of coffee beans has reached about 10 million tons in the last 2 years,
with a turnover greater than US$20 billion (International Coffee Organization, 2018).
This makes coffee one of the most traded and consumed agricultural products worldwide,
second only to oil (Lee et al., 2015). Although several species have been identified, Coffea
arabica L. (Arabica coffee) and Coffea canephora P. (Robusta coffee) account for about
60% and 40%, respectively, of all coffee produced worldwide. Distinctive features such
as smooth, mild, and rich flavors characterize Arabica coffee, while Robusta, lacking in
taste, denotes a flatter flavor, with a muddy odor (Flament, 2002). However, considering
the low price of Robusta coffee, this is commonly blended with Arabica beans to reduce
cost, but enhance cream formation (Folmer, Blank, and Hofmann, 2017) and create dis-
tinctive aroma profiles. However, blending Robusta with Arabica beans inevitably leads
to a loss of aromatic quality. For this reason, its addition must be limited. Generally, cof-
fee blends do not exceed a 50:50 mixture ratio of Arabica and Robusta (Dias et al., 2018).

19.2.1 The Fruit

The structure of the fruit is characterized by an orange-red skin on ripening (exocarp),

a yellow-white pulp and mucilage (mesocarp), and a plain parchment (endocarp) and
silver skin (integument) surrounding the seeds (endosperm) (Figure 19.1). The external
resistance of the fruit is assured by the exocarp (a monocellular layer protected by a waxy
substance). The fruits appear green (unripe) and turn to red/violet, yellow, or orange once
ripe, depending on the genotype. The mesocarp appears like a pulp showing distinctive
features: fleshiness, fibrousness, and sweetness. This is a complex structure characterized
by several compounds such as carbohydrates (mainly glucose, fructose, and pectin), pro-
teins, lipids, tannins, polyphenols, and caffeine (Janissen and Huynh, 2018). The endo-
carp (parchment layer) appears as a thin yellowish polysaccharide, which is composed
essentially of α-cellulose, hemicellulose, and lignin. Polysaccharides (mainly cellulose and
hemicelluloses), monosaccharides, proteins, polyphenols, and other minor compounds
are the main components of the silver skin (Farah and dos Santos, 2014). This layer, char-
acterized by a high antioxidant potential due to the large quantities of total dietary fibers
and phenolic compounds, covers two hemispheres of elliptical seeds inside which is where
 Coffee Flavor 367

FIGURE 19.1 Coffee fruit structure. (Created using information from Farah and dos
Santos, 2014.)

the endosperm and embryos are located (Esquivel and Jiménez, 2012). Harvesting repre-
sents the first step in coffee processing. Commonly, coffee fruits develop heterogeneously,
therefore, leading to a concurrent presence of different maturation stages in the same
coffee tree (green, cherry, raisin representing immature, ripe, and overripe, respectively).
Clearly, harvesting should be initiated when the plant reaches a homogeneous stage of
maturation with a low prevalence of immature fruits. In fact, mature fruits present lower
concentrations of phenolic compounds, thus implying a reduction of astringency; con-
versely, coffee cherries show a higher content of volatile compounds (aldehydes, ketones,
and higher alcohols) in comparison to immature fruits (Wintgens, 2004). Since the fruit
collection step may affect the quality of the final product, harvesting has a notable impor-
tance. The process is predominantly performed by handpicking or by stripping the fruits
onto sheets placed beneath the tree. In addition, the use of mechanical harvesters based
on the vibration of tree branches is spreading more and more worldwide. Clearly, the
choice of approach employed can directly affect the fruit quality used for the further
steps of on-farm processing. Handpicking, thanks to its selectivity, allows the selection
of fruits in their ideal stage of maturation (e.g., coffee cherries). However, applying this
selective method is expensive and laborious. For this reason, many producers choose
between stripping or mechanical harvesting, followed by removal of immature fruits
through sorting (Huch and Franz, 2015). Due to the structure and composition of the
fruit, once harvested, coffee processing should begin as quickly as possible, thus prevent-
ing unfavorable fermentation or mold formation. While the skin and pulp (outer layers)
can be easily removed, the mucilage, parchment, and silver skin are firmly attached to the
beans (De Bruyn et al., 2017). There are different methods that can be used to remove
these layers from the beans: dry, wet, and semi-dry processing (Figure 19.2). In the first
process, seeds are exposed to the sun or air dryers in order to reduce the moisture con-
tent to around 10%. Then, once the fruits are cleaned and dehulled, the dried skin and
pulp are removed. The second process involves a comprehensive series of steps, including
the mechanical removal of both skin and pulp, microbial degradation of the mucilage
layer, followed by sun drying. Thanks to this process, the time required is reduced from
1 month to 10 days; likewise, the area necessary for drying the beans in relation to dry
processing is reduced. The third approach (semi-dry) presents steps of both dry and wet
methods, matching the mechanical removal of the pulp with a final sun-drying phase
368 Food Aroma Evolution

FIGURE 19.2 Schematic representation of postharvest processing methods. (Adapted

from Pereira et al., 2017.)

(Pereira et al., 2017). The drying process can be performed by the sun or by using differ-
ent mechanical dryers (e.g., static, column, round, or forced air dryers). The choice of the
technique used is linked to economic factors and/or to the processing typology. In wet-
processed coffee, for example, the use of mechanical dryers for coffees is increasing more
and more due to the reduction in both drying time and risk of microbial contamination.

19.3 RAW BEAN COMPOSITION AND FLAVOR

PRECURSOR FORMATION

Raw coffee beans are characterized by an unpleasant taste. The distinctive beverage fla-
vor is obtained through thermal reactions (roasting and brewing). In fact, raw beans
only show a basic composition of chemical volatiles when compared to roasted beans.
Generally, in roasted coffee, more than 1,000 volatile compounds can be detected; con-
versely, in raw beans only 200 can be found on average. Table 19.1 shows the main
volatile compounds and the related flavor. The aroma of coffee is strictly related to the
chemical composition of the raw beans and characteristic aromas are generated during
 Coffee Flavor 369

TABLE 19.1 Main Aromatic Compounds Found in Roasted Coffee

Compound Flavor
Aldehydes
(E)-2-Nonenal Buttery
2-e-4-Methylbutanal Buttery
2-Methylbutanal —
2-Methylpropanal Malty/fruity/buttery/oily
3-Methylbutanal Malty
3-Methylpropanal Roasted cocoa
Acetaldehyde —
Hexanal Butter rancid
Methional Cooked potato
p-Anisaldehyde Minty
Phenylacetaldehyde Sweet fruity
Propanal Roasted/fruity
Acids
2-Methylbutyric Sweaty
3-Methylbutyric Sweaty
2-Methyl-1-butanoic Sweaty/acidic
3-Methyl-1-butanoic Sweaty/acidic
4-Methylbutanoic acid Sweet/acidic
Esters
Ethyl-2-methylbutyrate Fruity
Ethyl-3-methylbutyrate Fruity
Furanic Compounds
Furfural Almonds
2-((Methylthio)methyl)furan Smoke/roast
2-Furanemethanol —
2-Methylfuran —
5-Methyl-2-furancarboxyaldehyde —
Furfurylformiate —
Furfurylmethyl —
Furfurylformate —
Furfuryldisulfide —
Dihydro-2-methyl-3(2H)-furanone —
2-Eth​yl-4-​hydro​xy-5-​methy​l-3(2​H)-fu​ranon​e Sweet caramel
3-Hydroxy-4,5-dimethyl-2(5H)-furanone (sotolone) Sweet caramel, seasoning-like
4-Hydroxy-2,5-dimethyl-3(2H)-furanone (furaneol) Sweet caramel
5-Eth​yl-3-​hydro​xy-4-​methy​l-2(5​H)-fu​ranon​e Seasoning-like, caramel-like
(abhexon)
5-Eth​yl-4-​hydro​xy-2-​methy​l-3(2​H)-fu​ranon​e Sweet caramel
2-Ethyl-furaneol
2-Eth​yl-4-​hydro​xy-5-​methy​l-4(5​H)-fu​ranon​e
(Continued )
370 Food Aroma Evolution

TABLE 19.1 (Continued) Main Aromatic Compounds Found in Roasted Coffee

Compound Flavor

Sulfur-Containing Compounds
Dimethyl trisulfide Cabbage-like
Methional Boiled potato-like
Bis(2-methyl-3-furyl)disulfide Meat-like
5-Dimethyl-trisulfide Cabbage-like
Thiols
2-Methyl-3-furanthiol Meaty/boiled
3-Mercapto-3-methylbutylacetate Roasty
3-Methyl-2-butene-1-thiol Amine-like
2-Furfurylthiol Roasty/coffee-like
3-Mercapto-3-methylbutylformate Cat/green/cassis
Methanethiol (mercaptan) Cooked potato
4-Methyl-2-buteno-1-thiol Smoke/roasted
2-Methyl-4-furanthiol (furfuryl mercaptan) Meat
2-Furanmethanethiol Smoke/roasted
2-(Methylthiol)propanal Soy sauce
2-(Methylthio-methyl)furan Smoke/roasted
3,5-Dihydro-4(2H)-thiophenone Smoke/roasted
2-Acetyl-2-thiazoline Roasted
3-Mercapto-3-methylbutanol Hazelnut/roasted
Thiophenes
3-Methylthiophene —
Thiazols
2,4-Dimethyl-5-ethylthiazole Earthy/roasty
Ketones
1-Octen-3-one Mushroom-like
2,3-Hexadione —
3,4-Dimethylcyclopentenol-1-one Caramel-like/sweet
4-(4′-Hydroxyphenyl)-2-butanone Sweet fruity (raspberry ketone)
1-(2-Furanyl)-2-butanone —
2,3-Butanedione Buttery/caramel-like
2,3-Pentanedione Buttery/caramel-like
2-Hyd​roxy-​3,4-d​imeth​yl-2-​cyclo​-pent​en-1-​one Caramel-like
2-Hydroxy-3-methyl-2-ciclopenten-1-one Sweet/caramel
Norisoprenoids
(E)-β-damascenone Honey-like/fruity
Phenolic compounds
Guaiacol Phenolic/burned/
4-Ethylguaiacol Spicy
4-Vinylguaiacol Spicy
Vanillin Vanilla-like
(Continued )
 Coffee Flavor 371

TABLE 19.1 (Continued) Main Aromatic Compounds Found in Roasted Coffee

Compound Flavor
4-Methoxyphenol Phenolic
4-Ethenyl-2-methoxyphenol (4-ethenyl-guaiacol) Phenolic
3-Methylindole Coconut
4-Hydroxy-3-methoxybenzaldehyde (Vaniline) Vanilla
Pyrazines
3-Isopropyl-2-methoxypyrazine Earthy/roasty
2-Ethyl-3,5-dimethylpyrazine Earthy/roasty
3-Isobutyl-2-methoxypyrazine Earthy
2,3-Dimethylpyrazine Hazelnut/roasted
2,5-Dimethylpyrazine Hazelnut/roasted
2-Ethylpyrazine Peanuts/roasted
2-Ethyl-6-methylpyrazine Peanuts/roasted
2,3-Diethyl-5-methylpyrazine Hazelnut/roasted
2-Ethyl-3,5-dimethylpyrazine Earth/hazelnut/roasted
2-Etenyl-3,5-dimethylpyrazine Earth
2-Etenyl-3-ethyl-5-methylpyrazine Earth
6,7-Dihydro-5H-ciclopentapyrazine Hazelnut/roasted
6,7-D​ihydr​o-5-m​ethyl​-5H-c​iclop​entap​yrazi​ne Hazelnut/roasted

roasting. The Maillard reaction, together with Strecker degradation, caramelization, and
oxidation represent the main (complex) reactions of this process (Fisk et al., 2012). Figure
19.3 schematizes the main reactions involved in flavor generation. The chemical com-
position of the unprocessed seeds of the Coffea fruit (raw beans) is extremely complex,
including several compounds characterized by distinctive chemical and physical prop-
erties (Bagchi, Moriyama, and Swaroop, 2016). Carbohydrates represent one of main
aroma precursors in raw beans: both insoluble (cellulose and hemicellulose) and soluble
(e.g., arabinose, fructose, galactose, glucose, sucrose, raffinose, and stachyose) (Fadai
et al., 2017). Other important precursors are represented by nitrogen (N)-containing
compounds and chlorogenic acids (Poisson et al., 2017). During roasting, sucrose, glu-
cose, and fructose (low-molecular weight carbohydrates) can contribute to the formation
of acids and other volatile compounds. Moreover, polysaccharides are key compounds
related to the retention of volatiles and, consequently, flavor formation. Some alkaloids
such as caffeine and trigonelline together with proteins, nonvolatile acids (citric, malic,
and quinic), and volatile ones (acetic, butanoic, decanoic, formic, hexanoic, isovaleric,
and propanoic), are also detected in high concentrations in raw beans. During roast-
ing, these compounds break down, thus generating key flavor-active metabolites such
as pyridines and pyrrols (Poisson et al., 2017). About lipids, although triacylglycerols,
sterols, coffeadiol, arabiol, and tocopherols substantially compose the fraction present
in the raw beans, only a small amount is located in the outer layer of the bean (coffee
wax). Thiols, phenolics, and minerals are the other minor compounds detected in raw
coffee beans. The former are key molecules able to affect the sensory perception of cof-
fee, highlighting the “roasty” and “coffee” notes (Dulsat-Serra, Quintanilla-Casas, and
Vichi, 2016). Phenolic compounds, caffeoylquinic acid-CQA, feruloylquinic acid-FQA,
and dicaffeoylquinic acid-diCQA, are the most representative molecules, and are also
characterized by a well-known antioxidant activity and several properties that support
372 Food Aroma Evolution

FIGURE 19.3 Scheme of the main reactions involved in flavor generation.

human health (Ogawa, 2014). Clearly, mineral composition appears strictly related to
soil constitution as well as to other factors, such as altitude, humidity, and temperature.
Generally, potassium shows higher amounts, followed by phosphorus, magnesium, and
calcium (Carvalho Neto et al., 2017). It is well known that heterocyclic compounds,
key molecules in providing the distinctive flavor of coffee, are usually not found in raw
 Coffee Flavor 373

beans. For example, in a previous study, only 2-methoxy-3-(2-methylpropyl)-pyrazine

was detected in raw beans when compared to the compounds (more than 350 heterocyclic
compounds) found in roasted beans (Lee and Shibamoto, 2002). Among these, the main
classes of compound are pyrrols, furans, pyrazines, thiazols, oxazols, thiopheones, and
imidazols. However, although less complex than roasted beans, the volatile fingerprint
of raw beans comprise several compounds such as hydrocarbons, higher alcohols, alde-
hydes, ketones, acids, esters, lactones, sulfur compounds, furans, and phenols. Among
these, aldehydes and alkanes are the most abundant chemical classes detected. In a study
focused on Turkish raw beans, isoamyl alcohol (10.4%), hexanal (10.4%), and hexaco-
sane (8.2%) were shown to be the most predominant (Poyraz et al., 2016). In another
study analyzing Hawaiian raw coffee samples, 3-methyl butanoic acid (32.8%), phenyl
ethyl alcohol (17.3%), hexanol (7.2%), 4-hydroxy-3-methylacetophenone (3.7%), and
3-methyl butanol (3.6%) were the main constituents detected (Lee and Shibamoto, 2002).
High concentrations of both aldehydes (hexanal and benzaldehyde) and alkanes (tetra-
decane and cyclotetrasiloxane, octamethyl) were found in Thai raw beans (Somporn et
al., 2011). It has been proven that many volatiles, mainly those of non-thermal origin,
are lost during the roasting process. Hexanal, isoamyl alcohol, 1-hexanol, 1-pentanol,
2-heptanol, 1-octen-3ol, benzyl alcohol, and benzaldehyde fall into this group (Poyraz
et al., 2016). On the other hand, γ-butyrolactone, linalool, guaiacol, pyridine, furfural,
5-methylfurfural, 1-methylpyrrole, and β-damascenone are reported to be common com-
pounds detected in both raw and roasted samples (Gonzalez-Rios et al., 2007). These are
responsible for distinctive notes found in the final coffee beverage (e.g., “spicy,” “green,”
and “vegetable-like”) (Poyraz et al., 2016). Furthermore, some compounds such as geos-
min, 2,4,6-trichloroanisol-phenol, and 4-heptenal, externally caused by insects or the
immaturity of beans, are defined as off-flavors, negatively affecting coffee quality.

19.4 COFFEE PROCESSING: FERMENTATION AND ROASTING

Coffee flavor is greatly influenced by the roasting and brewing process. However, pre-
harvest and postharvest procedures also have a great deal of influence on flavor (Table
19.2). After coffee beans are harvested, they are submitted to a process that eliminates

TABLE 19.2 Main Compounds in Relation to Postharvest Treatment

Wet Treatment G/R Dry Treatment G/R
2-Hydroxy-3-methyl-cyclopenten-1-one Roasted 2,3-Butanediol Green
2-Phenylethanol Green Acetic acid Green
3-Methyl-2-cyclopenten-1-one Roasted Dimethyl sulfide Green
4-Ethylguaiacol Roasted Dimethyl sulfoxide Green
4-Methoxyphenol Roasted
4-Vinylguaiacol Roasted
5-Methylfurfural Roasted
Ethanol Green
Ethyl acetate Green
Ethyl isovalerate Green
Furfuryl formate Roasted
Methyl acetate Green
374 Food Aroma Evolution

the pulp and mucilage and then dried. All the metabolic reactions involved in such steps
affect the quality of the coffee (Kleinwächter, Bytof, and Selmar, 2015; Lee et al., 2015)
Green coffee processing aims to eliminate the flesh (exo- and mesocarp) and the parch-
ment (endocarp) and to dry the coffee beans thereafter. As explained, this process can be
performed in three different ways: dry processing, wet processing, or a combination of
both. In wet processing, the pulp is mechanically removed from the fruit and the remain-
ing mucilages are degraded by fermentation. Then, the beans are dried and the parch-
ment and testa removed by hulling. On the other hand, dry processing consists of drying
the whole coffee fruit before removing the non-desirable layers by hulling (Kleinwächter,
Bytof, and Selmar, 2015). The latter is mostly used in areas where climate conditions
are fit for drying. The type of processing that is applied is going to influence the coffee
flavor and its quality. Therefore, wet-processed beans are characterized by their full
aroma and pleasant acidity. On the other hand, beans that are submitted to the dry pro-
cess usually possess a so-called full body flavor. When sensory analysis was performed,
it was detected that wet-processed coffees were more aromatic with fruity and acidic
attributes, whereas dry-processed coffees had a higher degree of bitter, burned, and
woody notes (Leloup et al., 2004; Duarte, Pereira, and Farah, 2010). According to these
authors, such differences are mostly attributed to the fermentation step. However, it is
a difficult process to study and the effects of aroma profiles have not been entirely elu-
cidated. Fermentation is usually controlled by detecting when it is ready with the naked
eye. This makes the process difficult to standardize and also dangerous since under or
over fermentation can easily happen. However, there are some research projects that
have focused on establishing standard parameters, using known microorganism cultures
rather than relying on autochthone microbiota, and so on (Lee et al., 2015). Roasting,
on the other hand, is the main contributor to coffee flavor. Such process involves many
chemical reactions that are not yet well understood related to chemical browning such
as the Maillard reaction and caramelization. This process is usually carried out at tem-
peratures above 200°C. These chemical reactions modify/degrade substances present in
green coffee leading to the appearance of new compounds responsible for the character-
istic flavor of brewed coffee. As a consequence of these reactions, hundreds of volatile
compounds appear. There are also nonvolatile compounds that participate in the coffee
flavor, being responsible for bitterness, astringency, and sweetness, though they can also
be precursors for aromatic compounds (Toledo et al., 2016). Among these nonvolatile
compounds, caffeine, trigonelline, cholorogenic acids, lipids, proteins, and polysaccha-
rides are found. The main parameter is the degree of roasting; the higher the tempera-
ture and the time, the darker the coffee bean will be. The degree that the coffee bean has
been roasted is usually measured by its color and/or the loss of weight (Kleinwächter,
Bytof, and Selmar, 2015).

19.4.1 Influence of Fermentation on Aroma and Different Ways of Carrying It Out

The impact of fermentation on coffee aroma is often underestimated, since the main
objective of this process is not to influence it but rather just clean the coffee beans of
mucilage layers. However, during this process mucilages are metabolized and degraded,
thereby yielding new compounds (Lee et al., 2015). The fermentation of coffee beans can
be carried out with chemicals or microorganisms, which is the most common technique
(Toledo et al., 2016). Many different aerobic bacteria have been isolated from the fer-
mentation phase: Klebsiella azaenae, K. oxytoca, Hafnia spp., Enterobacter aerogenes,
 Coffee Flavor 375

and lactic acid bacteria such as Leuconostoc mesenteroides or Lactobacillus brevis, and
others including Erwinia herbicola and E. dissolvens. Some yeasts have also been isolated
such as Hanseniaspora uvarum, Saccharomyces cerevisiae, Torulaspora delbrueckii,
Debarymyces hansenii, Kloeckera apis apicualata, Candida guilliermondii, C. parapsilo-
sis, C. tropicalis, Criptococcus albidus, C. laurentii, Pichia anomala, and P. kluyveri (Lee
et al., 2015). On the other hand, pectinolytic microbiota has also been isolated during fer-
mentation. This would suggest that they play a role during such a process. Nevertheless, it
is still not well known whether bacteria or yeasts share the responsibilities here or whether
one has more weight than the other (Toledo et al., 2016). During fermentation both kinds
of microorganisms seem to grow, but as the pH starts to drop yeasts become predomi-
nant (Lee et al., 2015). The main problem associated with fermentation is the difficulty
in keeping it properly controlled. This is an important issue because even though a well
carried fermentation is going to improve coffee flavor, a lack of control over it is going
to yield non-desirable flavors (Jackels and Jackels, 2005). Usually, the end point of the
fermentation is just based on observation, which makes it very subjective and easy to get
wrong (Jackels et al., 2006). This misjudgment can cause problems with the fermentation
either due to over fermentation or under fermentation. Each of these is going to gener-
ate undesirable substances. When under fermentation happens, some mucilage is still not
degraded which could cause undesirable bacteria to grow. On the other hand, when over
fermentation takes place, black or “stinker” beans are produced which are characterized
by a rather poor appearance and by undesired flavors such as fruity, floral, sour, or alco-
hol (Lee et al., 2015). Moreover, fermentation usually relies on autochthone microbiota,
which is highly variable depending on previous processing, climate conditions, geographic
locations, and so on. Additionally, this microbiota is very complex making it very dif-
ficult to control (Lee et al., 2015). There have been some research projects focused on
solving such fermentation issues, mostly by defining an objective end point and using
known starter cultures. According to a study, pH could be a reliable tool for controlling
fermentation. In this study, a decrease in the pH (from 5.5 to 4) yielded the same results in
different batches (Jackels and Jackels, 2005). According to the authors, pH control would
prevent undesirable fermentation. Moreover, aside from pH, other parameters that could
help with controlling fermentation are temperature and humidity. On the other hand, the
use of certain starter cultures, as well as pectinolytic enzymes, to gain more control over
the fermentation has also been studied. A study in which cellulase and Aspergillus niger
were combined and used as starter yielded a higher amount of reducing sugars, which are
essential substrates for the Maillard reaction and caramelization that take place during
roasting (Lee et al., 2015). As a consequence, more caramel and sweet flavors were found
in the resulting coffee brews. The use of pectinolytic enzymes has proven to be useful in
removing the mucilage layer producing coffee brews with better attributes than when such
enzymes are not used. In addition, the suitability of the indigenous yeast microbiota vs. the
use of a selected yeast starter was studied. As result, it was observed that the latter yielded
a high-quality coffee brew in terms of aroma. Among the compounds generated by such a
starter were acetaldehyde, ethanol, ethyl acetate, isoamyl acetate, and notes such as butter,
fruits, or caramel, which are well accepted (Evangelista et al., 2014). There are also other
means for performing fermentation which help with its modulation. Some of them involve
a digestive step in which an animal ingests the coffee beans. Therefore, these beans are
subjected to an acidic and enzymatic hydrolysis and fermentation as a result of the transi-
tion through the digestive system of the animal. On the one hand, there is the production
of Kopi Luwak, one of the most expensive coffees in the world. In this case, coffee brews
are obtained from coffee beans that have been ingested by a civet cat (Marcone, 2004).
376 Food Aroma Evolution

According to this author, protein hydrolysis achieved within the gastrointestinal tract of
the civet cat would, on the one hand, reduce bitterness and, on the other hand, release
amino acids, which would have an important role during the Maillard reaction. A similar
case is black ivory coffee, which uses coffee beans that have passed through the gastro-
intestinal tract of an elephant, whose own characteristics cause the final coffee to differ
from the coffee obtained from the civet cat. Another fermentation methodology is the so-
called monsooning, which achieves a monsooned coffee. This practice takes place mostly
in India and consists of a fermentation on the surface of the beans that happens during
their transportation during the monsoon period (Ahmad and Magan, 2002). What makes
this methodology special is the specific microbiota that appears due to the monsoon condi-
tions and makes monsooned coffee different from non-monsooned coffee. Accordingly, a
different flavor profile is achieved, though there is little known about the role that bacteria
and yeasts have on such flavors.

19.4.2 Modulation of Coffee Aroma by Single Microorganisms

Modulating coffee aroma by using specific microorganisms has also been attempted.
Accordingly, Rhizopus oligosporus could be used to ferment coffee beans and modulate
coffee flavor (Lee et al., 2016). These authors studied the volatile and nonvolatile frac-
tion of the green coffees obtained. They observed a higher hydrolysis of proteins, which
gave rise to a higher concentration of proline and aspartic acid, involved in the Maillard
reaction. Moreover, they showed a higher degradation of ferulic and caffeic acids, which
in turned produced higher amounts of volatile compounds derived from phenolic com-
pounds. Therefore, the fermentation of coffee beans using R. oligosporus prompted
important changes in the volatile profile of green coffees and the precursors of volatile
compounds that appear during roasting having thus a great influence on coffee aroma.
Second, Lee et al. (2017) attempted to modulate coffee aroma by using Yarrowia lipo-
lytica as a starter culture for coffee bean fermentation. These authors observed a decrease
of sugars, amino acids, and phenolic compounds after fermentation. As result of phenolic
metabolism, a higher concentration of volatile phenolics appeared. Moreover, the metab-
olism of this yeast yielded higher concentrations of 2-phenyletanol and at the same time
the levels of alkanes, acids, and aldehydes were reduced. These metabolic outputs were
attributed to the unique hydrophobic pathways found in Y. lipolytica. Consequently, fer-
mentation using this yeast has proven to be able to modulate coffee flavor.

19.4.3 Influence of Roasting

Roasting is a very complex process in which coffee beans are submitted to high tempera-
tures that can reach up to 300°C. During this step, most of the aromatic compounds
responsible for coffee flavor are going to appear. Roasting involves a large number of
transformations of different natures: physical, physicochemical, and chemical. During
roasting both volatile and nonvolatile compounds are formed. Nonvolatile compounds
are also important since they are responsible for bitterness, sweetness, astringency, and
so on, but they are also precursors of volatile compounds. Among these nonvolatile
­compounds we find caffeine, lipids, proteins, phenolics, and trigonelline (Toledo et al.,
2016). The main reactions involved in roasting are the Maillard reaction and pyrolysis or
thermal degradation.
 Coffee Flavor 377

19.4.3.1 Thermal Degradation of Precursors

During roasting, due to the high temperatures applied, thermal degradation of proteins,
sugars, and phenolic compounds takes place. The degradation of carbohydrates, com-
plex or simple, participate in the chemical browning reaction called caramelization and
therefore give rise to products of such a reaction and caramel flavors (Toledo et al., 2016).
Moreover, phenolic compounds present in green coffee beans are also degraded due to
high temperatures. Therefore, chlorogenic acids, other phenolics such as feruloylquinic
acid, are broken down into their respective hydroxycinnamates. Further, the resulting
hydroxycinnamates can produce a second degradation and other chemical reactions
resulting in volatile phenolics such as guaiacol, p-vinylguaiacol, and others (Lee et al.,
2015). It is not just the compounds mentioned above that can be degraded during roast-
ing but also trigonelline, which is an alkaloid. As a result, pyrrols and pyridines are
produced. Another significant pathway to generate such compounds is the thermal deg-
radation of hydroxy amino acids (Lee et al., 2015).

19.4.3.2 The Maillard Reaction

The Maillard reaction is the main chemical pathway involved in the development of
coffee flavor. A great deal of different compounds appear during its course: pyrazines,
pyrrols, thiols, furanones, pyridines, thiophenes (Lee et al., 2015). The Maillard reaction
takes place when the carbonil group of a reducing sugar reacts with the free amino group
of an amino acid or protein (Rufián-Henares, García-Villanova, and Guerra-Hernández,
2008). Therefore, Maillard reaction development is going to rely on the amounts of such
precursors. As has been stated before, the fermentation step has a great influence on
the concentration of such compounds, and thus fermentation is going to have a great
influence on roasting. Briefly, the Maillard reaction is developed through several steps
(Rufián-Henares and Pastoriza, 2016):

• Reaction of the sugars with the amino acids (Schiff base).

• Amadori rearrangement or Heyns rearrangement.
• Degradation of Amadori and Heyns products producing aromatic compounds
such as reductones, furanic compounds, pyranones, and pyrrols; formation of
acetal or acetaldehyde.
• Interaction of amino acids with decarbonyl, acetal, acetaldehyde. The Strecker
degradation occurs, giving rise to new aldehydes with very high aromatic
properties.
• As part of the final stages, more aromatic compounds are formed, without the
participation of amino acids, and pyrazines, pyrrols, thiazols, and thiopehenes
appear.
• Finally, melanoidins are formed. These compounds are large polymers that are
brown in color, derived from the condensation of different molecules.

19.5 THE IMPACT OF THE DRYING AND STORAGE PROCESSES

Until this point, it is clear that roasting generates most of the aromatic compounds asso-
ciated with coffee. However, it is also clear that the precursors for such aromatic com-
pounds can be influenced by the treatments that take place before roasting, mostly by
fermentation. This process can modify the content in proteins, carbohydrates, trigonel-
line, phenolic compounds, and so on. In fact, some studies have proven that differences in
378 Food Aroma Evolution

the concentrations of such compounds give different coffee qualities after tasting (Franca,
Mendonça, and Oliveira, 2005). In another study, it was found that coffee quality was
positively related with trigonelline and 3,4-dicaffeoylquinic acid, but it was negatively
related with monoesters of phenolics such as 3-caffeoylquinic acid (Farah et al., 2006).
However, among the different postharvest processes, the two phases of drying and stor-
age are also able to influence the quality level of the coffee.

19.5.1 Drying

During this process, coffee beans remain viable, characterized by intense metabolic activ-
ity. Low-molecular weight sugars such as glucose, fructose, and mannose are subject to
reactions of interconversion. Similarly, proteins are hydrolyzed, resulting in the accu-
mulation of a wide variety of free amino acids (Jöet et al., 2010). However, currently,
there is not a study that has investigated changes in specific volatile compounds through
the drying process, and only a small amount of research has focused only on the major
chemical compounds such as sugars and proteins. Clearly, novel studies that are more
specific could assist in the understanding of volatile losses caused by evaporation and/or
oxidation reactions during drying. After the drying process, the freshly processed coffee
beans can be stored for a period of up to 3 years.

19.5.2 Storage

This phase must be maintained under ideal conditions of humidity (about 11%), under
low temperature, and in an inert atmosphere, thus preserving bean quality. During the
storage process, beans can remain viable for up to 6 months (up to 1 year if stored within
the parchment layer). After this period, the beans start a series of reactions (the senes-
cence process) (Toci et al., 2013). A reduction in cup quality is generally linked to the loss
of bean viability. This decline is correlated with chemical reactions occurring during the
senescence reactions, such as chlorogenic acid oxidation, leading to the development of
a bluish-green color in the beans. The carbohydrate fraction present in freshly processed
coffee beans has a direct influence on the germination process during storage, and its
content is subject to both positive and negative changes during this period (Selmar, Bytof,
and Knopp, 2008). Due to oxidation processes, undesired changes within the lipid frac-
tion also occur during storage (accumulation of reactive oxygen species) (Rendón, Salva,
and Bragagnolo, 2014). Compounds such as carbohydrates, lipids, and proteins repre-
sent key precursors in the development of chemical aromas during the roasting process.
Coffees prepared using beans stored for only 6 months show raspy, woody, or stale notes,
while after 1 year of storage flat aromas and old and woody notes are detected (Rendón,
Salva, and Bragagnolo, 2014). This evidence proves a possible direct influence of volatile
constituents formed and/or generated during the storage process.

19.6 CONCLUSIONS

The findings presented in this chapter show that the main odor-active compounds in cof-
fee beverages come from the roasting process and are consequently not detected in raw
green coffee beans. As a result, green coffee bean quality is determined by the major,
 Coffee Flavor 379

nonvolatile constituents present in the raw material, such as sugar, amino acids, and
lipids. These aroma precursors will further undergo modifications in the postharvest pro-
cessing steps due to the seed germination process and lipid oxidation; in fact, pre-roasting
processes also play an important role in coffee flavor, especially coffee fermentation.
Among the different steps in postharvest coffee processing, microbial mucilage removal
has a major influence on the volatile composition of the processed beans. Moreover, fer-
mentation represents a critical step since coffee beans can develop odd flavors due to a
lack of monitoring. For this reason, different fermentation methodologies have been stud-
ied to improve coffee quality. Research has shown that microbial-derived metabolites can
diffuse into seeds, thus remaining after the roasting process (e.g., esters, higher alcohols,
aldehydes, ketone, and terpenoids). Among these compounds, flavor-active esters show a
great potential for influencing the quality of the final coffee beverage.

REFERENCES

Ahmad, R. and Magan, N. (2002). Microfloral contamination and hydrolytic enzyme

differences between monsooned and non-monsooned coffees. Letters in Applied
Microbiology, 34(4), 279–82.
Bagchi, D., Moriyama, H., and Swaroop, A. (2016). Green Coffee Bean Extract in
Human Health. Boca Raton, FL: CRC Press.
Bhumiratana, N., Adhikari, K., and Chambers, E. (2011). Evolution of sensory aroma
attributes from coffee beans to brewed coffee. LWT—Food Science and Technology,
44, 2185–92.
Buffo, R.A. and Cardelli-Freire, C. (2004). Coffee flavour: An overview. Flavour and
Fragrance Journal, 19, 99–104.
Carvalho Neto, D.P., Pereira, G.V.M., Tanobe, V.O.A., Soccol, V.T., da Silva, B.J.G.,
Rodrigues, C., and Soccol, C.R. (2017). Yeast diversity and physicochemical char-
acteristics associated with coffee bean fermentation from the Brazilian Cerrado
Mineiro region. Fermentation, 3, 1–11.
De Bruyn, F., Zhang, S.J., Pothakos, V., Torres, J., Lambot, C., Moroni, A.V., Callanan,
M., Sybesma, W., Weckx, S., and De Vuyst, L. (2017). Exploring the impacts of
postharvest processing on the microbiota and metabolite profiles during green cof-
fee bean production. Applied and Environmental Microbiology, 83, e02398–416.
Dias, R.C.E., Valderrama, P., Março, P.H., dos Santos Scholz, M.B., Edelmann, M., and
Yeretzian, C. (2018). Quantitative assessment of specific defects in roasted ground
coffee via infrared-photoacoustic spectroscopy. Food Chemistry, 255, 132–8.
Duarte, G.S., Pereira, A.A., and Farah, A. (2010). Chlorogenic acids and other relevant
compounds in Brazilian coffees processed by semi-dry and wet post-harvesting
methods. Food Chemistry, 118(3), 851–5.
Dulsat-Serra, N., Quintanilla-Casas, B., and Vichi, S. (2016). Volatile thiols in coffee: A
review on their formation, degradation, assessment and influence on coffee sensory
quality. Food Research International, 89, 982–8.
Esquivel, P. and Jiménez, V.M. (2012). Functional properties of coffee and coffee by-
products. Food Research International, 46, 488–95.
Evangelista, S.R., Silva, C.F., Miguel, M.G.P. da C., Cordeiro, C. de S., Pinheiro, A.C.M.,
Duarte, W.F., and Schwan, R.F. (2014). Improvement of coffee beverage quality by
using selected yeasts strains during the fermentation in dry process. Food Research
International, 61, 183–95.
380 Food Aroma Evolution

Fadai, N.T., Melrose, J., Please, C.P., Schulman, A., and Van Gorden, R.A. (2017). A
heat and mass transfer study of coffee bean roasting. International Journal of Heat
and Mass Transfer, 104, 787–99.
Farah, A. and dos Santos, T.F. (2014). The coffee plant and beans: An introduction. In
V.R. Preedy (Ed.). Coffee in Health and Disease Prevention (pp. 73–81). Academic
Press.
Farah, A., Monteiro, M.C., Calado, V., Franca, A.S., and Trugo, L.C. (2006). Correlation
between cup quality and chemical attributes of Brazilian coffee. Food Chemistry,
98(2), 373–80.
Ferrão, L.F.V., Caixeta, E.T., Pena, G., Zambolim, E.M., Cruz, C.D., Zambolim, L.,
Ferrão, M.A.G., and Sakiyama, N.S. (2015). New EST–SSR markers of Coffea ara-
bica: Transferability and application to studies of molecular characterization and
genetic mapping. Molecular Breeding, 35, 31.
Fisk, I.D., Kettle, A., Hofmeister, S., Virdie, A., and Kenny, J.S. (2012). Discrimination
of roast and ground coffee aroma. Flavour, 1, 14.
Flament, I. (2002). Coffee Flavor Chemistry (Chapter 5). West Sussex: John Wiley &
Sons.
Folmer, B., Blank, I., and Hofmann, T. (2017). Chapter 17—Crema—Formation, sta-
bilization, and sensation. In F. Britta (Ed.). The Craft and Science of Coffee (pp.
399–417). London: Academic Press.
Franca, A.S., Mendonça, J.C.F., and Oliveira, S.D. (2005). Composition of green and
roasted coffees of different cup qualities. LWT—Food Science and Technology,
38(7), 709–15.
Gonzalez-Rios, O., Suarez-Quiroz, M.L., Boulanger, R., Barel, M., Guyot, B., Guiraud,
J.P., and Schorr-Galindo, S. (2007). Impact of “ecological” post-harvest processing
on the volatile fraction of coffee beans. I. Green coffee. Journal of Food Composition
and Analysis, 20, 289–96.
Huch, M. and Franz, C.M.A.P. (2015). Coffee: Fermentation and microbiota. In W.
Holzapfel (Ed.). Advances in Fermented Foods and Beverages (pp. 501–13).
Woodhead Publishing.
International Coffee Organization. (2018). The Current State of the Global Coffee Trade
|CoffeeTradeStats. Accessed date: September 21, 2018.
Jackels, S.C. and Jackels, C.F. (2005). Characterization of the coffee mucilage fermenta-
tion process using chemical indicators: A field study in Nicaragua. Journal of Food
Science, 70(5), C321–5.
Jackels, S.C., Jackels, C.F., Vallejos, C., Kleven, S.H., Rivas, R., and Fraser-Dauphinee,
S. (2006). Control of the Coffee Fermentation Process and Quality of Resulting
Roasted Coffee : Studies in the Field Laboratory and on Small Farms in Nicaragua
During the 2005–06 Harvest. Retrieved September 11, 2018, from /pape​r/Con​trol-​
of-th​e -Cof​fee-F​ermen​t atio​n-Pro​c ess-​a nd-o​f-%3A​-Jack​els-J​ackel​s /0ed​f 9efe​87dce​
b4e97​6ea76​c9b84​a8af2​d043c​0f.
Janissen, B. and Huynh, T. (2018). Chemical composition and value-adding applications
of coffee industry by-products: A review. Resources, Conservation and Recycling,
128, 110–7.
Jöet, T., Laffargue, A., Descroix, F., Doulbeau, S., Bertrand, B., de Kochko, A., and
Dussert, S. (2010). Influence of environmental factors, wet processing and their
interactions on the biochemical composition of green Arabica coffee beans. Food
Chemistry, 118, 693–701.
 Coffee Flavor 381

Kleinwächter, M., Bytof, G., and Selmar, D. (2015). Coffee processing. In Victor R.
Preedy (Ed.). Coffee in Health and Disease Prevention (pp. 73–80). London, UK:
Elsevier.
Knysak, D. (2017). Volatile compounds profiles in unroasted Coffea arabica and Coffea
canephora beans from different countries. Food Science and Technology, 37, 444–8.
Lee, K.-G. and Shibamoto, T. (2002). Analysis of volatile components isolated from
Hawaiian green coffee beans (Coffea arabica L.). Food and Fragrance Journal, 17,
349–51.
Lee, L.W., Cheong, M.W., Curran, P., Yu, B., and Liu, S.Q. (2015). Coffee fermentation
and flavor—An intricate and delicate relationship. Food Chemistry, 185, 182–91.
Lee, L.W., Cheong, M.W., Curran, P., Yu, B., and Liu, S.Q. (2016). Modulation of cof-
fee aroma via the fermentation of green coffee beans with Rhizopus oligosporus: I.
Green coffee. Food Chemistry, 211, 916–24.
Lee, L.W., Tay, G.Y., Cheong, M.W., Curran, P., Yu, B., and Liu, S.Q. (2017). Modulation
of the volatile and non-volatile profiles of coffee fermented with Yarrowia lipolytica:
I. Green coffee. LWT, 77, 225–32.
Leloup, V., Gancel, C., Liardon, R., Rytz, A., and Pithon, A. (2004). Impact of wet and
dry process on green coffee composition and sensory characteristics. Amino Acids,
93–101.
Marcone, M.F. (2004). Composition and properties of Indonesian palm civet coffee (Kopi
Luwak) and Ethiopian civet coffee. Food Research International, 37(9), 901–12.
Ogawa, M. (2014). Coffee and hippuric acid. In V.R. Preedy (Ed.). Coffee in Health and
Disease Prevention (pp. 209–15). Academic Press.
Pereira, G.V.M., Carvalho Neto, D.P., Medeiros, A.B.P., Soccol, V.T., Neto, E.,
Woiciechowski, A.L., and Soccol, C.R. (2016). Potential of lactic acid bacte-
ria to improve the fermentation and quality of coffee during on-farm processing.
International Journal of Food Science and Technology, 51, 1689–95.
Pereira, G.V.M., Soccol, V.T., Brar, S.K., Neto, E., and Soccol, C.R. (2017). Microbial
ecology and starter culture technology in coffee processing. Critical Reviews in
Food Science and Nutrition, 57, 2775–88.
Poisson, L., Auzanneau, N., Mestdagh, F., Blank, I., and Davidek, T. (2017). New insight
into the role of sucrose in the generation of α-diketones upon coffee roasting. Journal
of Agricultural and Food Chemistry, 66, 2422–31.
Poyraz, I.E., O z̈ tu r̈ k, N., Kiyan, H.T., and Demirci, B. (2016). Volatile compounds of
Coffea arabica L. Green and roasted beans. Anadolu University Journal of Science
and Technology, 5, 31–5.
Rendón, M.Y., Salva, T.J.G., and Bragagnolo, N. (2014). Impact of chemical changes on
the sensory characteristics of coffee beans during storage. Food Chemistry, 147,
279–86.
Rodriguez, J., Duran, C., and Reyes, A. (2010). Electronic nose for quality control of
Colombian coffee through the detection of defects in “cup tests.” Sensors, 10, 36–46.
Rufián-Henares, J.A., García-Villanova, B., and Guerra-Hernández, E. (2008).
Occurrence of furosine and hydroxymethylfurfural as markers of thermal dam-
age in dehydrated vegetables. European Food Research and Technology, 228(2),
249–56.
Rufián-Henares, J.A. and Pastoriza, S. (2016). Maillard reaction. In B. Caballero, P.
Finglas, and F. Toldrá (Eds.). The Encyclopaedia of Food and Health (Vol. 3, pp.
593–600). Oxford: Academic Press.
382 Food Aroma Evolution

Sanz, C., Czerny, M., Cid, C., and Schieberle, P. (2002). Comparison of potent odorants
in a filtered coffee brew and in an instant coffee beverage by aroma extract dilution
analysis. European Food Research and Technology, 214, 299–302.
Selmar, D., Bytof, G., and Knopp, S.-E. (2008). The storage of green coffee (Coffea ara-
bica): Decrease of viability and changes of potential aroma precursors. Annals of
Botany, 101, 31–8.
Somporn, C., Kamtuo, A., Theerakulpisut, P., and Siriamornpun, S. (2011). Effects of
roasting degree on radical scavenging activity, phenolics and volatile compounds of
Arabica coffee beans (Coffea arabica L. cv. Catimor). International Journal of Food
Science and Technology, 46, 2287–96.
Sunarharum, W.B., Williams, D.J., and Smyth, H.E. (2014). Complexity of coffee flavor:
A compositional and sensory perspective. Food Research International, 62, 315–25.
Toci, A.T., Neto, V.J.M.F., Torres, A.G., and Farah, A. (2013). Changes in triacylglycer-
ols and free fatty acids composition during storage of roasted coffee. LWT—Food
Science and Technology, 50, 581–90.
Toledo, P.R.A.B., Pezza, L., Pezza, H.R., and Toci, A.T. (2016). Relationship between
the different aspects related to coffee quality and their volatile compounds.
Comprehensive Reviews in Food Science and Food Safety, 15(4), 705–19.
Wintgens, J.N. (2004). Factors influencing the quality of green coffee. In J.N. Wintgens
(Ed.). Coffee: Growing, Processing, Sustainable Production (pp. 789–809).
Weinheim: Wiley—VCH Verlag GmbH & Co.
 Chapter 20
Aroma Evolution in
Chocolate Production
Roberta Ascrizzi, Luisa Pistelli, and Guido Flamini

CONTENTS

20.1 The Food of the Gods 383

20.2 The Volatiles of Chocolate Aroma 384
20.2.1 Pyrazines 385
20.2.2 Esters 386
20.2.3 Alcohols and Phenols 388
20.2.4 Aldehydes and Ketones 391
20.2.5 Furanones and Pyrones 393
20.2.6 Acids 395
20.3 The Importance of the Starting Material 397
20.3.1 The Cocoa Varieties 397
20.3.1.1 The “Fine Grade” Varieties: Criollo, Trinitario, and
Nacional 398
20.3.1.2 The Forastero “Bulk Grade” Cocoa Variety 399
20.3.2 The Geographical Origins of Cocoa 399
20.4 Primary Processing: Postharvest Fermentation and Drying 399
20.4.1 Fermentation of the Beans 400
20.4.2 The Drying Phase 403
20.5 The Secondary Processing Phases 403
20.5.1 The Alkalization Process 404
20.5.2 The Roasting Process 404
20.5.3 The Grinding and Refining Phases 406
20.5.4 The Conching Phase 408
References 409

20.1 THE FOOD OF THE GODS

Few other plant species have had the same worldwide success among consumers through-
out the millennia as cocoa. In 1753, the Swedish founder of the binomial nomenclature
Carl Linnaeus named this tree Theobroma cacao (which in Greek means “food of the
gods”) to memorialize the Mexican legend of Quetzalcoatl, the benevolent God who
gifted the cocoa tree to the Mexican people as a sign of his love, as it came from the gar-
den of heaven (González, 2007). It was a crop of utmost importance in the pre-Columbian
Mesoamerican communities: its fruits, called “pods,” were used as the core ingredient of

383
384 Food Aroma Evolution

a bitter beverage and as a currency (Nair, 2010). A recent study, though, identified the
area of its first domestication and consumption in the upper Amazonas regions of South
America, from which it was then traded in the Central America region, where its use was
only established 1.5 millennia later than in the Amazon basin (Zarrillo et al., 2018).
Its main use is undoubtedly the preparation of chocolate, which represents one of
the most traded commodities in the world. The reason for its success among consumers
is its aroma, a complex bouquet of flavors whose development is due to many factors.
Chocolate aroma development, indeed, depends on several factors (Figure 20.1) (Afoakwa
et al., 2008; Engeseth and Ac Pangan, 2018; Aprotosoaie, Luca, and Miron, 2016):

1. The starting material, which includes the genotype and the environmental grow-
ing conditions of the processed cocoa
2. The primary processing, including all the required postharvest treatments (i.e.,
pulp management, fermentation, and drying)
3. The secondary processing, which consists of the pre-roasting treatments (i.e.,
cleaning, shells breaking, and winnowing), followed by the actual chocolate
preparation (i.e., alkalization, roasting, and grinding)

Each of these factors contributes to the flavor; thus, knowledge of the processes involved
in the development of the aroma compounds in each of them is of utmost importance in
the production of a successful final product.

20.2 THE VOLATILES OF CHOCOLATE AROMA

Several volatile organic compound (VOC) chemical classes contribute to the distinctive
chocolate aroma: most develop from aroma precursors, namely reducing sugars and
free amino acids, whose production is thermally induced once the fermentation begins
and the enzyme pools are triggered. Carbohydrates constitute almost 20% of the raw,
unfermented cocoa beans: 2 to 4% is represented by free sugars, among which sucrose
accounts for up to 90%, while polysaccharides add up to circa 12% (Aprotosoaie, Luca,

FIGURE 20.1 Main factors influencing chocolate aroma development throughout its
processing chain.
 Aroma Evolution in Chocolate Production 385

and Miron, 2016). The invertase activity of the bean enzymes converts sucrose into fruc-
tose and glucose during the fermentation phase (Biehl and Ziegleder, 2003a; Afoakwa
et al., 2008): these reducing sugars are of fundamental importance in the production of
the typical chocolate flavor-active compounds by means of the Maillard reaction, which
takes place during the roasting phase (Ho, Zhao, and Fleet, 2014).
Over 600 VOCs have been reported in the chocolate headspace (Ziegleder, 1990),
of which pyrazines are the most important compounds involved in its distinctive flavor,
with a detected pool of circa 80 different molecules belonging to this class (Aprotosoaie,
Luca, and Miron, 2016; Afoakwa et al., 2008). Esters follow as the second most relevant
chemical group of compounds in the final product’s aromatic quality (Aprotosoaie, Luca,
and Miron, 2016). Other favorable aroma-contributing volatiles are alcohols, aldehydes,
ketones, furanones, and pyrones (Aprotosoaie, Luca, and Miron, 2016). Phenols and
acids, instead, confer unpleasant notes: the former are reported as smoky-like, while the
latter are described as rancid and vinegar-like (Aprotosoaie, Luca, and Miron, 2016;
Jinap, Dimick, and Hollender, 1995).

20.2.1 Pyrazines

Pyrazines are heterocyclic compounds with two nitrogen atoms in a 6-membered ring,
and they represent the key compounds involved in the pleasant chocolate flavor (Beckett,
2015). They are heterocyclic volatiles produced by the Maillard reaction, which is trig-
gered in conditions of low humidity and high temperatures (Yu and Zhang, 2010; Ho,
Zhao, and Fleet, 2014), and their synthesis is linked with that of aldehydes. The Maillard
reaction generates α-dicarbonyl compounds, which are responsible for the browning and
for the toasted flavor of the cooked food. Under high temperature conditions, the Strecker
degradation occurs between α-dicarbonyl compounds and α-amino acids: the amino acid
skeleton is rearranged in a volatile aldehyde, while its carboxylic group is eliminated as
CO2, and the sugar molecule retains its amino group (Figure 20.2).
At this point, the two sugar residues, identical or not, condense, and undergo oxida-
tion, which ultimately produces a substituted pyrazine (Rodriguez-Campos et al., 2012)
(Figure 20.3).
These reactions are strongly influenced by the temperature and the duration of the
thermal reaction (Aprotosoaie, Luca, and Miron, 2016). The pyrazine synthesis rate is
reported as steadily increasing for the first 30 minutes of a 150°C roasting temperature,
while they decrement thereafter, for either a depletion of the precursors or their involve-
ment in other reactions, or for volatilization of the pyrazines (Reineccius, Keeney, and
Weissberger, 1972).
The structure numbering of pyrazines is reported in Figure 20.4, and the main pyr-
azines developed during the chocolate production are reported in Table 20.1.
Alkyl-substituted pyrazines are the most relevantly involved in the chocolate flavor
bouquet, with tetra- (TMP) and tri-methyl (TrMP) pyrazines as the most important, of
which the former represents over 90% of the total detected pyrazines and is considered
to have cocoa flavor-enhancing properties (Rodriguez-Campos et al., 2012; Ziegleder,
1990; Aprotosoaie, Luca, and Miron, 2016). TMP confers, indeed, roasted cocoa- and
coffee-like notes (Bonvehí, 2005; Rodriguez-Campos et al., 2011; Afoakwa, 2010). Earthy
and peanut-like aromatic notes are attributed to TrMP, which is also a very distinctive
chocolate flavor-active compound (Frauendorfer and Schieberle, 2006, 2008; Bonvehí,
2005). Other compounds of this class conferring nutty and peanut-like notes are 2-methyl,
386 Food Aroma Evolution

FIGURE 20.2 The reaction between the α-dicarbonyl compound from the Maillard reac-
tion and an α-amino acid to produce an aminoreductone, a precursor of pyrazines.

2-ethyl, 2,5- and 2,6-dimethyl, 2-ethyl-5-methyl, and 2,3-diethyl pyrazines (Bonvehí,

2005; Afoakwa, 2010). Sweeter, caramel- and candy-like attributes are, instead, character-
istic of 2,3-dimethyl and 2,3,5-tri-methyl-6-ethyl pyrazines (Bonvehí, 2005; Rodriguez-
Campos et al., 2012). Roasted and earthy flavors, perceived as undesirable, are exhibited
by 2-ethyl-3,5-dimethyl and 2,3-dietyl-5-methyl pyrazines (Afoakwa et al., 2013).

20.2.2 Esters

Methyl-, ethyl-esters, and acetates are the most involved VOCs of this chemical class
of compounds in chocolate headspace emission (Rodriguez-Campos et al., 2011; Ramli

FIGURE 20.3 Strecker degradation leading to the formation of heterocyclic compounds

and, ultimately, substituted pyrazines.
 Aroma Evolution in Chocolate Production 387

FIGURE 20.4 Structure numbering of pyrazines.

TABLE 20.1 Main Aroma Contributions Attributed to the Pyrazines Developed during the
Processing Chain of Chocolate
Aroma Contribution Responsible Pyrazine Reference
Favorable Notes
Roasted cocoa- and Tetramethyl pyrazine Bonvehí, 2005; Rodriguez-
coffee-like (R1=R2=R3=R4=CH3) Campos et al., 2011; Afoakwa,
2010
Earthy and Tri-methyl pyrazine (R1=R2=R3=CH3, Frauendorfer and Schieberle,
peanuts-like R4=H) 2006, 2008; Bonvehí, 2005
Nutty and 2-Methyl pyrazine (R1=CH3, Bonvehí, 2005; Afoakwa, 2010
peanuts-like R2=R3=R4=H)
2-Ethyl pyrazine (R1=CH2CH3,
R2=R3=R4=H)
2,5-Dimethyl pyrazine (R1=R3=CH3,
R2=R4=H)
2,6-Dimethyl pyrazine (R1=R4=CH3,
R2=R3=H)
2-Ehyl-5-methyl pyrazine
(R1=CH2CH3, R3=CH3, R2=R4=H)
2,3-Diethyl pyrazine
(R1=R2=CH2CH3, R3=R4=H)
Sweet, caramel-, and 2,3-Dimethyl pyrazine (R1=R2=CH3, Bonvehí, 2005; Rodriguez-
candy-like R3=R4=H) Campos et al., 2012
2,3,5-Tri-methyl-6-ethyl pyrazine
(R1=R2=R4=CH3, R3=CH2CH3)
Undesirable Notes
Roasted and earthy 2-Ethyl-3,5-dimethyl pyrazine Afoakwa et al., 2013
(R1=R4=CH3, R2=CH2CH3, R3=H)
2,3-Dietyl-5-methyl pyrazine
(R1=R2= CH2CH3, R3=CH3, R4=H)
388 Food Aroma Evolution

et al., 2006). In general, fruity and floral notes are attributed to esters, but some com-
pounds of this class can also represent off-flavors. Among acetates, for example, a relevant
presence of 2-phenylethyl and ethylphenyl (Figure 20.5) is positively correlated with the
pleasantness of chocolate (Rodriguez-Campos et al., 2012). In particular, 2-phenylethyl
acetate has a pleasant honey-like and flowery aroma, while ethylphenyl acetate exhibits a
sweeter contribution, with fruity notes (Rodriguez-Campos et al., 2011).
Other esters detected in chocolate confer a fruity aroma (Figure 20.6): ethyl octano-
ate is described as pineapple-like, ethyl decanoate has pear and grape reminiscences, ethyl
hexanoate is associated with an apple-like flavor (Bonvehí, 2005). A balsamic flavor is
attributed to isoamyl benzoate (Rodriguez-Campos et al., 2012) and methyl cinnamate
(Bonvehí, 2005).
Methyl- and ethyl-esters of myristic and stearic acids (Figure 20.7) are, instead, esters
exerting fatty, waxy, and oily aroma contributions: their presence, is, indeed, undesirable
in the final product (Bonvehí, 2005).

20.2.3 Alcohols and Phenols

Alcohols are mainly developed during the fermentation phase, as a result of both the micro-
bial carbohydrate metabolism and the thermal degradation of amino acids (Rodriguez-
Campos et al., 2011; Aprotosoaie, Luca, and Miron, 2016). The main compounds of this
chemical class involved in chocolate aroma are listed in Table 20.2.

FIGURE 20.5 Main acetates involved in pleasant chocolate aroma.

FIGURE 20.6 Other esters involved in the pleasantness of chocolate aroma.

 Aroma Evolution in Chocolate Production 389

FIGURE 20.7 Methyl (R1=CH3) and ethyl (R1=CH 2CH3) esters of myristic (R 2=C13) and
stearic (R 2=C17) acids.

A high alcohol content is reported as a desirable attribute in chocolate, as it confers

candy and flowery flavors, as well as green and floral notes (Rodriguez-Campos et al.,
2012; Aprotosoaie, Luca, and Miron, 2016). In particular, the presence of a relevant
content of 2,3-butanediol (Figure 20.8), which is responsible for the natural odor of
cocoa butter, is favorable, and it is reported as a fairly stable presence throughout the
whole processing chain of high-quality Criollo products (Ramos et al., 2014; Schwan
and Wheals, 2004; Ascrizzi et al., 2017). Other compounds of this class exhibiting sweet
and honey-like notes are 1-propanol, phenylethanol, and benzyl alcohol (Figure 20.8)
(Rodriguez-Campos et al., 2012). A fresher and citrus-like contribution is reported for
2-methyl-1-butanol, trans-3-hexen-1-ol, 1- and 2-hexanol, and 2-heptanol (Figure 20.8)
(Frauendorfer and Schieberle, 2008; Bonvehí, 2005; Ramos et al., 2014; Rodriguez-
Campos et al., 2012).

TABLE 20.2 Main Aroma Contributions Attributed to the Volatile Alcohols

Developed during the Processing Chain of Chocolate
Aroma Contribution Responsible Alcohol Reference
Favorable Notes
Cocoa butter odor 2,3-Butanediol Ramos et al., 2014; Schwan and Wheals,
2004; Ascrizzi et al., 2017
Sweet, honey-like 1-Propanol Rodriguez-Campos et al., 2012
Phenylethanol
Benzyl alcohol
Fresh, citrus-like 2-Methyl-1-butanol Frauendorfer and Schieberle, 2008;
trans-3-Hexen-1-ol Bonvehí, 2005; Ramos et al., 2014;
1-Hexanol Rodriguez-Campos et al., 2012
2-Hexanol
2-Heptanol
Floral Linalool
trans-Linalool oxide
(furanoid)
Undesirable Notes
Medicinal 2-, 3-, and Bonvehí, 2005
4-Methylphenols
2-Butanol
Repulsive 1-Heptanol Rodriguez-Campos et al., 2012; Bonvehí,
1,2-Propanediol 2005; Ramos et al., 2014
Smoky Phenol Rodriguez-Campos et al., 2012; Bonvehí,
2005; Ramos et al., 2014
390 Food Aroma Evolution

FIGURE 20.8 Volatile alcohols involved in the pleasantness of chocolate aroma.

Some derivatives, instead, exhibit an undesirable contribution to the overall chocolate

flavor perception: 2-, 3-, and 4-methylphenols have a medicinal and unpleasant aroma,
as well as 2-butanol (Figure 20.9) (Bonvehí, 2005). Moreover, phenol, 1-heptanol, and
1,2-propanediol (Figure 20.9) are perceived as repulsive flavors (Rodriguez-Campos et
al., 2012; Bonvehí, 2005; Ramos et al., 2014). In general, phenols exert aroma-damag-
ing properties, with smoky notes: their development has been linked to contamination
with burning wood or charcoal smoke during the drying phase or in storage facilities

FIGURE 20.9 Volatile alcohols linked exhibiting an unpleasant aroma to chocolate.

 Aroma Evolution in Chocolate Production 391

FIGURE 20.10 Main terpene alcohols involved in chocolate aroma.

(Aprotosoaie, Luca, and Miron, 2016). Their absence in the final product is mandatory
for high-quality cocoa (Jinap et al., 1999).
Among the terpene alcohols, linalool (Figure 20.10) shows a relevant presence in fine
grade cocoa, while being far less represented in the bulk varieties (Ziegleder, 1990; Biehl
and Ziegleder, 2003a; Ascrizzi et al., 2017). Its floral flavor is also given by its oxidized
derivative, trans-linalool oxide (furanoid) (Figure 20.10), which was also reported in fine
quality cocoa of the Criollo variety (Bonvehí, 2005; Ascrizzi et al., 2017).

20.2.4 Aldehydes and Ketones

Relevant contents of aldehydes and ketones are reported as desirable attributes for a
pleasant chocolate product, with a good cocoa flavor (Aprotosoaie, Luca, and Miron,
2016; Rodriguez-Campos et al., 2012). Aldehydes, in particular the aliphatic ones, enrich
the chocolate aroma bouquet with fruity and flowery notes (Bonvehí, 2005).
Among aldehydes, 2-methylbutanal (Figure 20.11) is unaffected by the duration
and the temperature of the fermentation process (Rodriguez-Campos et al., 2012). Its

FIGURE 20.11 Main aldehydes conferring pleasant aroma notes to chocolate.

392 Food Aroma Evolution

FIGURE 20.12 Volatile aldehydes whose decrement is favorable in chocolate processing.

positive contribution to the product is described as malty, with chocolate-like notes

(Frauendorfer and Schieberle, 2006; Afoakwa, 2010). Other cocoa-like aldehydes such as
3-methylbutanal and 2-methylpropanal (Figure 20.11) (Rodriguez-Campos et al., 2012)
are among the aliphatic compounds; 2-phenyl-2-butenal, 4-methyl-2-phenyl-2-pentenal,
and 5-methyl-2-phenyl-2-hexenal are among the aromatic ones (Figure 20.11) (Bonvehí,
2005; Afoakwa, 2010). The latter, in particular, is responsible for the intense bitter flavor
of some cocoas, and it has been reported in samples from different geographical origins
(Bonvehí, 2005; Ascrizzi et al., 2017).
Acetaldehyde, benzaldehyde, and n-pentanal (Figure 20.12), on the other hand, con-
fer a pungent and bitter characteristic to chocolate; thus, their decrement throughout
the processing chain is favorable (Ramos et al., 2014; Rodriguez-Campos et al., 2011;
Bonvehí, 2005).
Fruity and, more prominently, floral flavor contributions are caused by the presence of
ketones like 2-pentanone, acetophenone, 2-hydroxy, and 4-methyl acetophenones (Figure
20.13) (Rodriguez-Campos et al., 2011; Rodriguez-Campos et al., 2012; Bonvehí, 2005).
2-heptanone (Figure 20.14) exhibits a sweeter flavor, reported as a positive fruity
aroma, but also as a cheese-like and unpleasant note (Rodriguez-Campos et al., 2012;

FIGURE 20.13 Volatile ketones conferring fruity and floral aroma properties to chocolate.
 Aroma Evolution in Chocolate Production 393

FIGURE 20.14 Main volatile ketones responsible for unpleasant fat and pungent notes
in chocolate.

Schwab, Davidovich-Rikanati, and Lewinsohn, 2008). Among the undesirable ketone

contributions, a fat and buttery aroma is caused by 2,3-butanedione, while pungent notes
are due to benzylideneacetone and 2,3-pentanedione (Figure 20.14) (Rodriguez-Campos
et al., 2012; Bonvehí, 2005).
The main compounds of the chemical classes involved in chocolate aroma are listed
in Table 20.3.

20.2.5 Furanones and Pyrones

The monosaccharide degradation that occurs in the drying and roasting process gener-
ates furanones and pyrones (Aprotosoaie, Luca, and Miron, 2016). These compounds
add depth and complexity to the overall chocolate flavor, with aroma notes ranging
from nutty to sweet: the main compounds belonging to the chemical classes that exert
an aroma-active behavior (both positively and negatively) in chocolate are reported in
Table 20.4.
Furaneol and 5-(1-hydroxyethyl)-2-furanone (Figure 20.15) are perceived as fruity
(Bonvehí, 2005; Krings, Berger, and Banavara, 2003), while a nutty perception is asso-
ciated with maltol, 2-acetyl-5-methylfuran, and 5,6-dihydro-6-pentyl-2-pyrone (Figure
20.15) (Krings, Berger, and Banavara, 2003; Bonvehí, 2005).
A completely opposite sensorial reaction is, instead, linked to other pyrones (i.e.,
5,6-dihydro-4-methyl-2-, 3,5-dihydroxy-6-methyl-4-, and 2,3-d​ihydr​o-3,5​-dihy​droxy​
-6-me​thyl-​4 -pyr​ones,​all reported in Figure 20.16), which are perceived as repulsive,
unpleasant, sour, and over-roasted aroma notes (Krings, Berger, and Banavara, 2003;
Bonvehí, 2005).
394 Food Aroma Evolution

TABLE 20.3 Main Aroma Contributions Attributed to the Volatile Aldehydes and Ketones
Developed during the Processing Chain of Chocolate
Aroma Contribution Responsible Aldehyde/Ketone Reference
Favorable Notes
Malty, chocolate-like 2-Methylbutanal Rodriguez-Campos et al., 2012;
Frauendorfer and Schieberle,
2006; Afoakwa, 2010
Typical cocoa-like aroma 3-Methylbutanal Rodriguez-Campos et al., 2012;
2-Methylpropanal Bonvehí, 2005; Afoakwa, 2010
2-Phenyl-2-butenal
4-Methyl-2-phenyl-2-pentenal
Intense bitter flavor of some 5-Methyl-2-phenyl-2-hexenal Bonvehí, 2005; Ascrizzi et al.,
cocoa 2017
Fruity and floral 2-Pentanone Rodriguez-Campos et al., 2011,
Acetophenone 2012; Bonvehí, 2005
2-Hydroxyacetophenone
4-Methylacetophenone
Undesirable Notes
Pungent, bitter Acetaldehyde Ramos et al., 2014; Rodriguez-
Benzaldehyde Campos et al., 2011, 2012;
n-Pentanal Bonvehí, 2005
Benzylideneacetone
2,3-Pentanedione
Pleasant and fruity in small 2-Heptanone Rodriguez-Campos et al., 2012;
amounts; sweet and Schwab, Davidovich-Rikanati,
cheese-like if more present and Lewinsohn, 2008
Fat, buttery 2,3-Butanedione Rodriguez-Campos et al., 2012;
Bonvehí, 2005

TABLE 20.4 Main Aroma Contributions Attributed to the Volatile Furanones and
Pyranones Developed during the Processing Chain of Chocolate
Aroma
Contribution Responsible Furanone/Pyrone Reference
Favorable Notes
Fruity Furaneol Bonvehí, 2005; Krings,
5-(1-Hydroxyethyl)-2-furanone Berger, and Banavara, 2003
Nutty Maltol Krings, Berger, and
2-Acetyl-5-methylfuran Banavara, 2003; Bonvehí,
5,6-Dihydro-6-pentyl-2-pyrone 2005
Undesirable Notes
Repulsive, sour, 5,6-Dihydro-4-methyl-2-pyrone Krings, Berger, and
over-roasted 3,5-Dihydroxy-6-methyl-4-pyrone Banavara, 2003; Bonvehí,
2,3-D​ihydr​o-3,5​-dihy​droxy​-6-me​thyl-​4-pyr​one 2005
 Aroma Evolution in Chocolate Production 395

FIGURE 20.15 Furanones and pyrones associated with favorable aroma notes in
chocolate.

20.2.6 Acids

During fermentation, the short-chain carboxylic acids arise as a result of the bacteria car-
bohydrate metabolism, as well as products of the Strecker aldehyde degradation (Crafack
et al., 2014). Their flavor contributions are not only perceived as repulsive, but they also
exhibit low perception thresholds; thus, their release before the packaging of the final
product is desirable (Aprotosoaie, Luca, and Miron, 2016). Other than the vinegar-like
and pungent notes of acetic and propionic acids, sweaty and rancid flavors are reported
for butyric, isobutyric, 2-methylbutyric, and isovaleric acids (Figure 20.17) (Rodriguez-
Campos et al., 2011, 2012; Krings, Berger, and Banavara, 2003).
For acids with alkyl chains longer than 4 units (i.e., hexanoic, heptanoic, octanoic,
and decanoic acids, all reported in Figure 20.18), the aroma contribution gives off more
fatty and rancid characters, with sour attributes (Rodriguez-Campos et al., 2012).
Only two volatile acids detected in chocolate aroma are perceived as pleasant notes:
2-methyl and 3-phenyl propionic acids (Figure 20.19), conferring floral, rose-like notes to
the product (Krings, Berger, and Banavara, 2003; Bonvehí, 2005).

FIGURE 20.16 Volatile pyrones linked to unpleasant notes in chocolate aroma.

396 Food Aroma Evolution

FIGURE 20.17 Shorter-chain volatile acids conferring unpleasant aroma attributes to

chocolate.

FIGURE 20.18 Volatile alcohols with longer alkyl chains exhibiting rancid aroma notes.

FIGURE 20.19 Volatile acids conferring favorable aroma notes to chocolate.

 Aroma Evolution in Chocolate Production 397

20.3 THE IMPORTANCE OF THE STARTING MATERIAL

According to the APG (Angiosperm Phylogeny Group) III system (2009), Theobroma
cacao L. belongs to the Sterculioideae sub-family of the Malvaceae family (APG, 2009;
Chase and Reveal, 2009). The Cronquist system previously listed it as part of the now
obsolete Sterculiaceae family (Cronquist, 1981).
As a relatively small tree, its 4–8 m height can reach up to more than 10 m when
shaded by other forest trees, such as coconut and banana trees, which ensures protec-
tion from direct sunlight and wind (Afoakwa, 2016). Cocoa’s ideal growth habitats are
located between 20° north and south of the equatorial plane, in year-long rainy and
humid environments with temperatures ranging from 20 to 30°C, mostly in plantations
between 400 and 700 m a.s.l. (Beckett, 2015; McShea et al., 2008). The cocoa plant
begins producing its fruits after 2 to 3 years, with a full yield only after 6 to 7 years: as a
yearlong flower-bearing species, it exhibits two major flowerings which produces better
fruit (Ascrizzi et al., 2017; Beckett, 2015). The fruit consists of 15 to 25 cm long pods
growing directly on the trunk and on the bigger branches, weighing between 300 and
700 g (Ascrizzi et al., 2017). Each pod is constituted by an external husk containing 30
to 50 seeds embedded in a mucilaginous and thick pulp, rich in carbohydrates, and with a
bittersweet taste (Afoakwa, 2016). An adult cocoa tree yields approximately 2 kg of seeds
per year (Ascrizzi et al., 2017). The morphological features of the pods, together with
their geographical area of origin and their flavor characters, are used in order to classify
the many varieties of this species (Biehl and Ziegleder, 2003b).

20.3.1 The Cocoa Varieties

Cocoa is native to the humid forests of the Andean lower-eastern equatorial slopes
in South America (Afoakwa, 2010): the largest cocoa tree biodiversity, indeed, is still
reported in the Amazon basin (Motamayor et al., 2008). The controversial genetic
classification of this species is due to the breeding selection performed by cocoa grow-
ers once its domestication spread out (Warren and Kennedy, 1991): while the wild
individuals are self-incompatible, actually, the cultivated ones are not (Motamayor
et al., 2008).
Traditional classification clustered cocoa varieties into three main groups: Criollo,
Forastero, and Trinitario, of which the latter consists of Criollo×Forastero hybrids
(Cheesman, 1944). A fourth group was later added to this classification: the Nacional,
which is only grown in Ecuador (Despréaux, 1998). The genotyping of 1241 individuals
for more than 100 markers, though, seemed to evidence the need for a more complex clas-
sification system: Motamayor et al. (2008), indeed, proposed 10 major clusters to reflect
the actual available genetic biodiversity more accurately, rather than the traditional geo-
graphical origin–driven classification. In the present chapter, however, the classification
of the four main varieties (Criollo, Forastero, Trinitario, and Nacional) has been taken
into account. With the exception of Forastero, which is considered a “bulk grade” cocoa,
the other three are “fine grade” (Chetschik et al., 2018). While “bulk grade” cocoa is per-
ceived as harsher and stronger, the “fine grade” ones have a smoother and more delicate
aroma (Kattenberg and Kemming, 1998). The cocoa blends used in chocolate production
are generally dominated by bulk varieties, with the addition of only small quantities of
fine grade cocoa, which are sufficient to confer a distinctive aroma to the overall flavor
(Afoakwa et al., 2008).
398 Food Aroma Evolution

The starting material genotype induces variations in both the quality and the inten-
sity of the flavor of chocolate: aroma precursors and the enzyme pools involved in their
chemical modification cause the varietal flavor differences, reported as primarily quali-
tative rather than quantitative (Luna et al., 2002; Counet et al., 2004; Reineccius and
Heath, 2005). The variety-specific proteins and carbohydrates composing the fruit tissues
are the substrates of the degradation performed by the specific enzyme pools once the
fermentation takes place: the products of these degradation reactions are reducing sug-
ars and amino acids, which are the flavor precursors of chocolate (Schwan and Wheals,
2004; Ziegleder, 1990; Afoakwa et al., 2008; Kadow et al., 2013). The fruit pulp can,
indeed, be considered as a “reservoir” of chocolate aroma compounds, which migrate
into the seeds during the fermentation phase (Eskes et al., 2007).
“Fine grade” cocoa is, in general, perceived as a higher quality product according
to consumer preference, even though its volatile headspace emission was found to be
less potent when compared to “bulk grade” (Counet et al., 2004; Jinap, Dimick, and
Hollender, 1995). This can be explained as a “bouquet effect,” in which the compounds
act simultaneously, balancing each other, which ultimately results in a more pleasant
flavor perception (Jinap, Dimick, and Hollender, 1995).

20.3.1.1 The “Fine Grade” Varieties: Criollo, Trinitario, and Nacional

Criollo represents only 3% of global cocoa production, and it is mostly produced in Venezuela
(Aprotosoaie, Luca, and Miron, 2016). It has lower yields, and it is more susceptible to
pests and climatic shifts (Ziegleder, 1990). It is regarded as a higher quality product; thus,
it is more expensive compared to the other varieties (Afoakwa, 2016; Rodriguez-Campos
et al., 2011; Schwan and Wheals, 2004). Criollo cocoa is referred to as the “fine flavor
variety” or “fine grade,” as its characteristic floral and fruity notes are considered high-
quality attributes, which are not exhibited by “bulk” cocoa (Sukha et al., 2008; Ascrizzi
et al., 2017): monoterpene compounds have been proposed as the compounds responsible
for these pleasant notes (Ziegleder, 1990). Indeed, a higher linalool (Figure 20.10) emission
has been reported for fine grade cocoa seeds compared to bulk seeds, followed by myrcene
and ocimene (Ziegleder, 1990). These compounds have been detected in the pulps of ripe
cocoa fruit, thus attributing the pleasant aroma notes to the pulp composition of this variety
(Pino, Ceballos, and Quijano, 2010; Eskes et al., 2007). In Criollo cocoa, the mucilaginous
pulp is particularly sweet, with caramel- and nutty-like aroma notes, which migrate from
the pulp to the seeds during the fermentation phase, in which the permeability of the seeds
is enhanced (Kadow et al., 2013; Eskes et al., 2007). Methylketones (such as 2-heptanone,
shown in Figure 20.14, and 2-nonanone), secondary alcohols (such as 2-heptanol, Figure
20.8), and their respective esters (such as 2-heptanol acetate) are other aroma-active com-
pounds reported as already present in the cocoa fruit pulp of Criollo cocoa (Kadow et al.,
2013). Moreover, this variety exhibited a higher release of volatile acids (mainly acetic acid)
in its unfermented cocoa bean headspace emission (Qin et al., 2016): the sour and vinegar-
like aroma of acetic acid (Eskes et al., 2007) confers to this variety a more relevant final
acidic note (Frauendorfer and Schieberle, 2008; Acierno et al., 2016; Ascrizzi et al., 2017). In
general, Criollo exhibits higher content of aroma precursors, mainly reducing sugars, which
ultimately lead to a product richer in pyrazines (Giacometti, Mazor Jolić, and Josić, 2015).
Trinitario cocoa is a hybrid variety of Criollo and Forastero, which only grows as
a cultivated plant (Wood and Lass, 2001). Trinitario has inherited a higher yield and
less susceptibility to pest diseases from the Forastero variety (Jahurul et al., 2013). This
hybrid, however, exhibits a more complex aromatic bouquet compared to bulk cocoa,
with some wine-like flavors typical of its fine grade cocoa parent, Criollo (Giacometti,
 Aroma Evolution in Chocolate Production 399

Mazor Jolić, and Josić, 2015). Its acidic volatile content before fermentation was higher
than the bulk grade cocoa, as well as its volatile alcohol (such as 2-pentanol) and mono-
terpene content (such as linalool, see Figure 20.10) (Qin et al., 2016). Aldehydes (such
as hexanal and nonanal) and esters (such as benzyl and phenethyl acetates) in Trinitario
unfermented beans are even more abundant than in Criollo (Qin et al., 2016).
The Nacional fine grade variety, also known as “Arriba,” is only cultivated in
Ecuador (Aprotosoaie, Luca, and Miron, 2016). Its bean exhibits peculiar spicy and flo-
ral aromatic notes (Despréaux, 1998; Luna et al., 2002; Counet et al., 2004). Among the
fine grade varieties, it has the lowest pyrazine content: it is, indeed, the variety with the
lowest content of reducing sugars after the fermentation process (Afoakwa et al., 2008;
Giacometti, Mazor Jolić, and Josić, 2015). It is quite rare as a pure variety; in fact, it
accounts for only 5% of global cocoa production (Afoakwa, 2016).

20.3.1.2 The Forastero “Bulk Grade” Cocoa Variety

This resilient and productive variety represents over 90% of global cocoa production
(Aprotosoaie, Luca, and Miron, 2016). Its massive use has led to its large worldwide
distribution: Forastero cocoa exhibits a large genetic variability, with several subvariet-
ies, of which “Amelonado” is the most known (Jahurul et al., 2013). This high genotypic
variability causes a less consistent characterization of the volatile emissions from the
unfermented cocoa beans, although it is in general poorer in the peculiar volatile organic
compounds emitted from the other varieties of cocoa bean (Qin et al., 2016).

20.3.2 The Geographical Origins of Cocoa

Even when dealing with the same cocoa variety, its country of origin plays a dramatic role
in the final aroma of the packaged chocolate.
West Africa is the area of origin of the starting materials considered to be the ideal
standard in terms of flavor balance, with a pronounced chocolate character and nutty
undertones (Afoakwa, 2010). Nigeria, Côte d’Ivoire, and Ghana are the main producers
in West Africa: the latter, in particular, is known for producing less variable cocoa beans,
considered to be a premium raw material. Côte d’Ivoire, instead, is known for its beans
with low bitterness and low astringency (Afoakwa, 2010).
Among the South American cocoa, Brazil is known for its highly acidic beans, with-
out a significant cocoa impact but characterized by sourness and astringency (Urbanski,
1992; Fowler, 1999). This might be explained by the predominance of the lactic bacteria
during the fermentation phase, coupled with the lack of yeast activity, which ultimately
causes the complete lack of pleasant chocolate side notes such as nutty and/or fruity
(Afoakwa, 2010). The Ecuadorian beans are more balanced, although they do not exhibit
the same pronounced chocolate attributes as the Ghanaian ones (Afoakwa, 2010).

20.4 PRIMARY PROCESSING: POSTHARVEST

FERMENTATION AND DRYING

The harvesting of the cocoa fruits is performed every 2 to 4 weeks throughout several
months of the year, as not all the pods simultaneously show the same degree of ripen-
ing: in general, 7 to 10 days are necessary for complete fruit ripening, but harvesting can
be performed after 2 weeks. The harvesting time is a parameter to carefully evaluate: if
400 Food Aroma Evolution

performed too late, it already bears germinated beans or the formation of molds, thus
making the fruits unsuitable for chocolate production (Afoakwa, 2016). The cocoa pods
grown on the lower branches or trunk segments can be gathered by hand, while a knife,
cutlass, or machete can be used to remove those grown on the tallest sections of the plant
(Afoakwa, 2016; Ascrizzi et al., 2017).
Once harvested, the pods can be either cut immediately, or stored for a few days
before opening: cocoa fruit storage has shown a positive contribution to flavor develop-
ment, although it increments the mold formation and pest contamination occurrences if
not performed in a carefully controlled environment (Afoakwa, 2016). The opening of
the pods consists of cracking the outer shell and separating the bean from the mucilagi-
nous pulp in which it is embedded (Beckett, 2015). In this phase, defective cocoa beans
are rejected: common defects are flatness (the cocoa beans are oval), parasitic diseases,
or already triggered germination (Afoakwa, 2016). These processes produce the starting
material on which the primary processing phases will take place (Figure 20.20).
From this point on, the postharvest treatments take place: generally, they are per-
formed in the country of origin of the plant, as they must take place after a short
period following harvesting. The primary postharvest processing phases of the cocoa
beans ensure the development of the aroma precursors, which will be transformed
in the desirable chocolate aroma notes later on in the secondary processing phases
(Afoakwa, 2010).

20.4.1 Fermentation of the Beans

The fermentation phase takes place after the opening of the cocoa seeds. It is a crucial
step, as the fermentation-induced release of aroma precursors is of fundamental impor-
tance in the final product’s aromatic quality: scarcely or badly fermented beans lead to
a poor quality chocolate, as the building blocks of the flavor notes have not been prop-
erly developed (Schwan and Wheals, 2004; Tran et al., 2015). Moreover, unfermented
cocoa beans produce a sour, bitter, and astringent chocolate (Schwan and Wheals, 2004;
Ardhana, 2003; Afoakwa, 2010; Beckett, 2009). The duration of this process varies
based on the cocoa variety, but the only case in which it is not performed at all is for
beans used to manufacture cocoa butter only (Beckett, 2009).

FIGURE 20.20 Flow diagram for the preparation of primary processing starting material.
 Aroma Evolution in Chocolate Production 401

The cocoa bean consists of an outer shell, representing 10–14% of its dry weight,
and the kernel (or cotyledon), which accounts for up to 86–90% of the total bean mass:
the latter is the portion bearing the distinctive chocolate aroma (Osman, Nasarudin, and
Lee, 2004; Afoakwa, 2010). The cotyledon, indeed, contains globulin and albumin as
storage proteins, of which the former is degraded during fermentation, developing the
free amino acids, later used as aroma precursors during the roasting phase (Böle Biehl,
Wewetzer, and Passern, 1982; Voigt, Biehl, and Wazir, 1993; Pettipher, 1990). The sug-
ary tissue remaining on the seeds after the opening of the pods is a mucilaginous pulp: it is
rich in sugars, which represent the substrate for the fermentation process. Carbohydrates
constitute 10–15% of its weight, but it is mainly composed of water (82–87%), thus
representing an ideal medium for microbial growth (Afoakwa, 2010; Amoa-Awua et al.,
2007; Beckett, 2009; Nielsen, 2006; Thompson et al., 2013).
An overview of the main methods and phases of the fermentation of the beans is
shown in Figure 20.21. After harvesting and opening, the pods, with their residual adher-
ing pulp, can be fermented using different techniques: the most popular are box and
heap fermentation, but baskets, trays, or drying platforms can also be used (Afoakwa,
2016). Heap fermentation is performed mostly in the African cocoa-producing coun-
tries: the beans are heaped and covered with plantain leaves; they are mixed and turned
several times to avoid mold growth (Afoakwa, 2016). Smaller heaps tend to produce
better-flavored cocoa seeds (Beckett, 2015). Box fermentation is the most common tech-
nique, consisting of wooden or concrete boxes in which the cocoa beans are inserted:
these boxes have holes in their sides to ensure air circulation, and they are raised above
ground level (Afoakwa, 2016). Plantain or banana leaves are frequently inserted to cover
the seeds, and the boxes are then closed with a weighted board (Afoakwa, 2016). The
beans are mixed and turned in this case as well. Small-scale producers generally practice
basket fermentation, which involves woven baskets, with the usual banana or plantain
leaves covering the beans (Afoakwa, 2016). The tray fermentation, instead, is less labor-
demanding, as the beans do not need to be turned because they are laid on a single level
and the trays have holes on the bottom to ensure airflow (Afoakwa, 2016).
The fermentation process usually lasts 5 to 6 days for most cocoa varieties, but for
Criollo, it is reduced to 1 to 3 days (Afoakwa, 2010). During the first day, the residual
pulp layer liquefies and drains off, inducing a steady increment of the temperature inside
the bean (Afoakwa, 2016). The digestion of the pulp is performed by the resident micro-
organisms, which were naturally inoculated from the environment once the cocoa pods

FIGURE 20.21 Main methods and chemical modifications involved in the fermentation
phase.
402 Food Aroma Evolution

were cracked (Beckett, 2009). The fermentation is made up of three stages, which par-
tially overlap one another:

1. Anaerobic yeast metabolism (24–36 h): Sugars are converted to alcohol.

2. Lactic acid bacteria metabolism (48–96 h): Sugars and some organic acids are
converted into lactic acid.
3. Acetic acid bacteria metabolism: The alcohols are converted into acetic acid, in a
strong exothermic reaction which induces a marked temperature increase, reach-
ing up to 50°C in some fermentations (Beckett, 2009; Afoakwa, 2016).

Alcohol and acetic acid induces the death of the bean, avoiding its germination and caus-
ing cell structure breakage, which leads to the release of the enzymes (Beckett, 2009).
During the first (anaerobic) stage of fermentation, the invertase enzyme is active, and it
converts sucrose into glucose and fructose (Afoakwa, 2010). The proteinase enzymes
are active as well; they attack the proteins, leading to the development of peptides and
free amino acids (Afoakwa, 2016). Free amino acids and reducing sugars are the aroma
precursors whose presence is of fundamental importance in the development of choco-
late flavor, mostly due to the Maillard reaction and Strecker degradation, two reactions
which will take place during the roasting process.
In a well-performed fermentation, time is certainly a key factor: a duration suf-
ficient for the development of pleasant aroma compounds must be balanced with the
avoidance of producing unpleasant volatiles. Volatile organic acids are, indeed, released
as a result of the metabolism of the sugar occurring in this process (Rodriguez-Campos
et al., 2011): their development is enhanced if it lasts more than 6–8 days (Rodriguez-
Campos et al., 2012). The fermented cocoa beans exhibit the most significant content
of organic acids in their volatile emission (Frauendorfer and Schieberle, 2006). This
is also true for Criollo beans, whose fermentation phase is slightly shorter compared
to the Forastero variety (Ascrizzi et al., 2017): being processed in a less acidic envi-
ronment, the Criollo beans are characterized by a mild flavor variety (Beckett, 2009;
Wood and Lass, 2001).
Tetramethyl pyrazine is reported as the only pyrazine whose content is already rel-
evant in the unroasted, fermented beans (Ascrizzi et al., 2017; Reineccius, Keeney, and
Weissberger, 1972). Its presence in unroasted beans has been proposed as a result of
thermally triggered reactions following the microbial synthesis taking place during the
fermentation, particularly as a metabolic product of Bacillus subtilis (Ramli et al., 2006).
During this phase, indeed, the induction of the Maillard reaction is ensured by a rise in
the temperature, which can reach 50°C in the center of a fermenting mass (Reineccius,
Keeney, and Weissberger, 1972). The roasting of beans fermented without yeast has been
reported as a factor leading to a product with a less chocolaty character, due to the lower
amount of pyrazines (Ho, Zhao, and Fleet, 2014).
The fermentation phase also leads to the production of the esters, for which
amino acids are the main source (Biehl and Ziegleder, 2003a). The yeast metabolism
is, indeed, an important source of desirable compounds like 2-phenylethyl acetate
and ethyl-2-methyl butanoate (Aprotosoaie, Luca, and Miron, 2016; Afoakwa et al.,
2008). Moreover, defects in the fermentation process can cause the formation of amyl
acetates, which are considered off-odor VOCs in chocolate (Rodriguez-Campos et al.,
2012).
To ensure the development of the favorable contribution of aldehydes and ketones,
the ideal fermentation duration is 6 to 8 days (Rodriguez-Campos et al., 2012).
 Aroma Evolution in Chocolate Production 403

20.4.2 The Drying Phase

The procedure of drying the fermented beans is performed to reduce the moisture content
of the material in order to prevent mold growth and thus ensuring safe storage prior to
processing (Afoakwa, 2010). The moisture content drops from 55% to around 7.5%:
levels of moisture above 8% causes mold to develop (Fowler, Leheup, and Cordier, 1998;
Awua, 2002), but reaching levels below 6% confers a brittle aspect to the beans, making
them more prone to breakage during their handling in the subsequent processing phases
(Afoakwa, 2010; Beckett, 2009).
The control of the two main parameters involved in the drying phase, namely tem-
perature and humidity, is of the utmost importance in the production of high-quality
chocolate. They can also be adapted to obtain desired characteristics of the final prod-
uct. Different techniques can be used to perform the drying process of the cocoa beans;
optimal performances are obtained with the natural method, but artificial methods are
available for producing countries where the climate does not allow sun drying (Beckett,
2015). The cocoa beans are laid on wooden boxes or raised platforms, and they are
generally covered with bamboo (Afoakwa, 2016). However, many studies reported sun
drying as the best technique to ensure higher ratings in terms of consumer preference for
chocolate aroma development; moreover, the sun-dried beans had fewer smoky off-notes
(Buyukpamukcu et al., 2001; Amoye, 2006; Granvogl, Bugan, and Schieberle, 2006).
The latter can be a result of the contamination of the beans with the smoke from the fire
used to produce the heat permeating the drying chamber (Beckett, 2009).
The drying phase is also important in chocolate aroma evolution, as it is also meant
to improve flavor development (Afoakwa, 2010). Together with roasting, it is meant,
indeed, to reduce the acidity and astringency of cocoa, by means of the volatilization
of acetic acid and the oxidation of phenols, respectively (Afoakwa, 2010; Kadow et al.,
2013; Rodriguez-Campos et al., 2011). In terms of the reduction of unpleasant flavors,
the drying process consents the first decrement of short-chain organic volatile acids,
such as acetic, propionic, butyric, and isobutyric acids (Páramo et al., 2010). The hard-
ening of the bean cases can inhibit elimination through volatilization of these detrimen-
tal compounds: this can happen if drying is too harsh, generally if artificially performed
(Dimick and Hoskin, 1999). Rodriguez-Campos et al. (2012) reported 70°C as the ideal
drying temperature for the release of aldehydes and ketones. For the development of
pyrones and furanones, high residual humidity and moderate temperatures are desirable
(Ziegleder, 2009).

20.5 THE SECONDARY PROCESSING PHASES

The fermented cocoa beans are composed of the cocoa nibs or kernels (inner core) and
the shell (outer layer). The shell is not used in the production of chocolate and it also
retains the majority of the external contaminants (Knezevic, 1983); it does not confer
pleasant flavor to chocolate, and it can also damage the grinding machines, as it is fibrous
and hard and it retains silica particles (Beckett, 2009, 2015). Before storing the nibs, the
cocoa bean is subjected to a cleaning process, which removes dirt particles and contami-
nants, followed by the breaking of their shells by means of mechanical agitation, and
then winnowed to remove the unwanted material and only keep the nibs (Beckett, 2015;
Afoakwa, 2016). The cocoa nibs are the initial material used for the subsequent phases
taking place in the processing facilities: they are generally stored in jute bags, which
404 Food Aroma Evolution

allow the volatilization of the water vapor, before being sent to the country of destination
(Beckett, 2009).

20.5.1 The Alkalization Process

The alkalization process is mainly performed if the starting material is addressed to the
production of the cocoa powder and, in rare cases, this phase is carried out to enrich
chocolate with specific flavors (Beckett, 2009, 2015). This process is also known under
the name “Dutching”: it was invented in the Netherlands in the 19th century as a method
of obtaining a cocoa powder that would not sink to the bottom of a liquid medium, such
as water or milk, when preparing liquid chocolate (Beckett, 2015).
An alkali solution, generally potassium carbonate, is added to the cocoa nib prior
to the roasting procedure: the alkaline solution dose is important, as an over-alkalized
product gives soapy flavors and the addition of small amounts of acetic or tartaric
acid might need to be performed to balance the pH (Beckett, 2015). Sometimes, alka-
lization is performed on very acidic beans, and this treatment could be beneficial in
order to reduce the pungency and sourness of the final product (Beckett, 2015). The
alkalization process is performed in rotating drums, in which the alkali solution is
sprayed on the nibs; then they are gently and steadily dried at temperatures below
100°C (Afoakwa, 2016).
Among the volatile compounds exhibiting an aroma-active presence, furanones and
pyrones are strongly affected by the alkalization process, since it destroys these com-
pounds (Rodriguez-Campos et al., 2012).

20.5.2 The Roasting Process

Roasting, together with the fermentation process, is the most important phase in the
development of chocolate aroma: the possible starting substrates and the main events
occurring in this process are shown in Figure 20.22.
The roasting process can be performed on three different substrates (Afoakwa, 2010):

a. The whole beans: In this case, the winnowing procedure is performed after the
roasting takes place.
b. The cocoa nibs.
c. The cocoa liquor: In this case, the cocoa nibs are ground into cocoa liquor prior
to roasting.

With the exception of the whole beans, the other roasting substrates require a physical
(thermal and/or mechanical) pre-treatment to remove the shells.
During the roasting phase, many processes of different natures take place (Afoakwa,
2016):

1. If performed on whole beans, the shells are loosened and fall off the beans,
exposing the nibs.
2. The moisture drops to 2%, thus closer to the final desired moisture content of the
chocolate.
3. The nibs become more friable, and their color darkens.
 Aroma Evolution in Chocolate Production 405

FIGURE 20.22 Flowchart of the roasting possible substrates and main events.

4. The microbial title drops again: this is particularly relevant for products like
cocoa butter, liquor, and powder, for which the microbiological purity is more
stringent.
5. The proteins undergo denaturation, and the amino acids are degraded; the reduc-
ing sugars are also destroyed in this phase, as they are used as substrates of the
Maillard reaction.
6. A significant volatilization of the residual acidic content takes place, thus making
this phase particularly relevant for the reduction of the astringent, pungent, and
bitter notes conferred to chocolate by the volatile acids.

This process is performed in roasters, which can operate in batch or continuous flow,
and are generally drum-shaped (Beckett, 2015). The key parameters to control in order
to obtain a well-performed roasting procedure are its duration, the operating tempera-
tures, and the rate of moisture loss: each of these factors must be carefully regulated and
checked to ensure the production of a good quality chocolate (Awua, 2002).
In an early stage, before the actual roasting temperatures are reached, the roaster
generates vapor from water and heat, which is key in killing the residual bacteria
(Beckett, 2015). Before the proper roasting, however, the beans or nibs must be gently
dried: as for the drying phase, however, over-drying is not advisable, because it makes
the substrate more brittle and friable, thus more difficult to work with (Beckett, 2009,
2015; Afoakwa, 2010).
Once the moisture content is below 3%, the proper roasting takes place: the tem-
perature rises to the actual operating conditions, which generally range between 110 and
406 Food Aroma Evolution

140°C for 45 minutes to 1 hour; the material is subsequently cooled in an external cooler
(Beckett, 2015).
The most important chemical modifications that take place during this phase are (i)
the volatilization of the unpleasant volatile organic acids and (ii) the Maillard reaction
(Beckett, 2015). After the drying process, indeed, this phase allows for another signifi-
cant drop in the unpleasant volatile acid content, as up to 70% of the acetic acid occurs
during roasting (Rodriguez-Campos et al., 2012).
The Maillard reaction is important for both the non-enzymatic browning of the
baked, toasted, and roasted foods, as well as for the production of the distinctive aroma
flavor-active compounds (Beckett, 2015). It requires the presence of water, heat, reduc-
ing sugars, and free amino acids and a pH above 3 to be triggered (Beckett, 2015). The
alkyl skeleton of the amino acids is volatilized in the form of a volatile aldehyde through
the Strecker degradation: aldehydes are, actually, important contributors to the choco-
late aroma bouquet originating from odorless amino acids (Beckett, 2015). Prolonged
roasting procedures, though, performed at high temperatures, are not advisable in the
production of a rich and flavorful chocolate, as they cause a decrement in the aldehyde
content. Even though their synthesis is triggered by heat, if the temperature is too elevated
they might evaporate faster than they are synthesized (Aprotosoaie, Luca, and Miron,
2016; Owusu, Petersen, and Heimdal, 2013). The reducing sugars are split up into small
carbon chains. The most important amino acid involved in pyrazine synthesis is glycine,
which reacts with a 1,2-dioxo compound (glyoxal) (Beckett, 2015). Other amino acids
whose presence is relevant for the formation of aroma compounds are leucine, threonine,
valine, and glutamine, which all react with glucose when the temperature reaches 100°C
(Beckett, 2015). In particular, aroma notes reported as sweet are conferred by the inter-
action between leucine and glucose, while more chocolaty notes are described for threo-
nine and glutamine with glucose once the temperature reaches 100°C; finally, stronger
and more penetrating chocolate flavors are exerted by valine and glucose once heated
above 180°C (Dimick and Hoskin, 1999).
Tetra- and tri-methyl pyrazine contribution to a pleasant final product is so impor-
tant that their relative ratio is considered as an indicator of a correct roasting process: the
ideal degree induces a TMP/TrMP ratio ranging from 1.5 to 2.5, while values below 1
indicate an over-roasted product (Ziegleder, 1990). The roasting temperature influences
the esters content, as well; excessively high temperatures applied during this phase, indeed,
negatively affect the presence of these compounds, thus leading to a less pleasant product
(Ramli et al., 2006). A temperature of 130°C kept for 20 minutes ensures the ideal condi-
tions for the release of compounds like furanones and pyrones (Ziegleder, 1991).
With the exception of pyrazines, the effect of the roasting procedure on cocoa flavor
is quantitatively relevant, rather than qualitatively, as it mostly affects a small set of key
compounds released in the previous steps (Frauendorfer and Schieberle, 2008).

20.5.3 The Grinding and Refining Phases

After the roasting phase, a winnowing phase might be performed if the nib shells were
not removed in the previous phases; otherwise, the roasted cocoa nibs are directly ground
and refined with the other ingredients (combined grinding), such as granulated sugar,
necessary for the final chocolate recipe (Beckett, 2009; Dimick and Hoskin, 1981).
Alternatively, the mixing can be performed after each ingredient has undergone grinding
by itself (separate grinding): in this case, the manufacturer can set the optimal particle
 Aroma Evolution in Chocolate Production 407

sizing for each ingredient, but both the texture and the aroma of the final product will
exhibit differences compared to the combined grinding. The influence of this phase on
flavor is due to the influence exerted by the texture on the dynamic of the release of the
aroma compounds during the consumption of the final product (Beckett, 2009).
The grinding procedure consists of an irreversible process of the mechanical size
reduction of the particles involved. Performing this process is easier for material which
has previously been roasted and alkalized (Niediek, 1994). A general flowchart evidenc-
ing the phases of these processes is shown in Figure 20.23.
The grinding can be performed by several procedures, which take advantage of dif-
ferent forces to obtain the size reduction of the particles:

1. Crushers, which use direct mechanical compression on the material.

2. Different types of mills:
a. Stirred mills: The grinding media (beads, rods, or balls) are composed of
loose elements, whose relative motion imparts the breakage of the particles,
which relies on an impact force. Moreover, an impeller can be added to
enhance the flow rate of the grinding media introducing an agitation, and
thus a shear action to the system.
b. Impact mills: Hammers or pins impart impact stress on particles, generally
forcing them to pass through grated screens which act as both sieves and
impact sites.
c. Jet fluid: Air or steam are used to impart acceleration to the particles, thus
enhancing the particle reduction (Beckett, 2009).

The grinding phase produces the cocoa liquor, a liquid medium generated by the release
of the cocoa butter, a fat substance, from the disrupted cell structures of the cocoa nibs,
of which cocoa butter represents around 55% (Beckett, 1994). Contrary to roasting, it
is difficult to over grind the cocoa nibs, but, although in this process the use of tempera-
tures higher than the refining phase, the color and flavor can be altered by high grinding
temperatures (Niediek, 1994; Kuster, 1984).
Once ground, the cocoa liquor undergoes the refining process, during which the par-
ticle size is further reduced: in Europe, the diameter of the particles is generally dropped
to as low as 15–22 μm, while in the United States this value ranges between 20 and 30
μm (Beckett, 2009). This procedure ensures the reduction of the sugar and milk particles
(if present), providing a more thorough mixing of the ingredients, thus enhancing the
interspersion of the solids and the fats for a more pleasant sensorial experience (Beckett,
2009). This process is performed by means of a five roller refiner, which imparts compres-
sion and shear force to the refining media.

FIGURE 20.23 Flowchart of the main characteristics of the grinding and refining
processes.
408 Food Aroma Evolution

For improving the flow properties of chocolate, grinding and refining are of the
utmost importance to the sensory pleasantness profile of the final product: chocolate is
a solid which melts at body temperature in the mouth, and the dynamic of this melting,
coupled with the interaction with saliva, is responsible for its aroma and taste percep-
tions. A longer flavor persistence in the mouth was reported for several finer chocolate
samples, although they shared the same amount and size of cocoa solids; the coarser
ones, instead, had a shorter residence time in the mouth, and thus a lower perceived flavor
intensity (Mongia, 1997).
The physical modification of the materials induced through the grinding and the
refining phases are far more studied than their direct impact on the volatile emission of
the flavor-active compounds of the chocolate material. As an energy intense process, in
which shear, impact, and compressive forces are at play, the heat developed during these
operations is to be taken into account for its effect on the most volatile compounds. As
reported by Ascrizzi et al. (2017), the evaluation of the dynamics of the volatiles released
all throughout the processing chain of a premium dark chocolate showed a new and
significant release of volatile acids in the grainding and refining phases. Moreover, cocoa
liquor and refined chocolate showed a higher emission of both alcohols and aldehydes,
thus evidencing the direct importance residing in particle size reduction processes on
the volatiles, in terms of both the elimination of off-flavors (i.e., volatile acids) and their
impact on pleasant aroma notes (Ascrizzi et al., 2017).

20.5.4 The Conching Phase

The conching procedure was invented in 1878 in Switzerland by Randi Lindt (Beckett,
2015). Its main purpose is the further removal of unwanted flavors, as well as the improve-
ment of the physical properties of the final product, namely the reduction of its viscosity
and its smoothing (Owusu, Petersen, and Heimdal, 2013). It is performed in tanks with
stirring arms, and it consists of a combination of mixing the cocoa liquor with cocoa but-
ter and lecithin, strong agitation, and aeration of the chocolate undertaken while heating
it: as a thermally aided process, it is costly (Owusu, Petersen, and Heimdal, 2013). It is
not always performed, especially because nowadays the fine grinders are able to produce
a liquor with the right amount of smoothness and viscosity, with a more rounded mouth-
feel (Beckett, 2015). Moreover, moisture is also reduced by 30% (Beckett, 2009).
The reports on the influence of this phase on aroma development are conflicting.
No new flavor compound synthesis is triggered during conching, but quantitative dif-
ferences may occur in terms of an increment of branched pyrazines and a loss of low-
molecular weight aldehydes due to their volatilization (Counet et al., 2002; Schnermann
and Schieberle, 1997). The quantitative effect of conching on alcohols is not clear, as
the literature is contradictory: while some studies report alcohols as another chemical
class of volatile compounds whose content is incremented when the conching is performed
(Crafack et al., 2014), other works described a pronounced decrement in their content
when the conching process was carried out (Owusu, Petersen, and Heimdal, 2013). The
amino acid content, instead, is not affected by conching; moreover, their content, as well
as that of sugars available after the roasting, combined with the temperatures reached dur-
ing this process, are not able to trigger the Maillard reaction (Hoskin and Dimick, 1984).
It is reported, however, that as much as 80% of the total volatile emission is lost after
the first few hours of this treatment: an over conched product, therefore, will exhibit
a significantly reduced flavor bouquet (Beckett, 2015). The volatilization of these acid
 Aroma Evolution in Chocolate Production 409

compounds might induce an increment in the perception of the other flavor notes (Mohr,
Niediek, and Lange, 1968; Mohr, 1978; Maniere and Dimick, 1979). On the other hand,
this phase might help to eliminate unwanted acidic notes in the case of more acidic start-
ing material, or to remove off-flavors such as phenols (Beckett, 2015).

REFERENCES

Acierno, Valentina, Sine Yener, Martin Alewijn, Franco Biasioli, and Saskia Van Ruth.
2016. “Factors Contributing to the Variation in the Volatile Composition of
Chocolate: Botanical and Geographical Origins of the Cocoa Beans, and Brand-
Related Formulation and Processing.” Food Research International 84. Elsevier
B.V.: 86–95. doi:10.1016/j.foodres.2016.03.022.
Afoakwa, Emmanuel Ohene. 2010. Chocolate Science and Technology. Chichester, UK:
John Wiley & Sons. doi:10.1002/9781444319880.
Afoakwa, Emmanuel Ohene. 2016. Cocoa Production and Processing Technology, 1st
ed. CRC Press.
Afoakwa, Emmanuel Ohene, Alistair Paterson, Mark Fowler, and Angela Ryan.
2008. “Flavor Formation and Character in Cocoa and Chocolate: A Critical
Review.” Critical Reviews in Food Science and Nutrition 48 (9): 840–57.
doi:10.1080/10408390701719272.
Afoakwa, Emmanuel Ohene, Jennifer Quao, Jemmy Takrama, Agnes Simpson Budu,
and Firibu Kwesi Saalia. 2013. “Chemical Composition and Physical Quality
Characteristics of Ghanaian Cocoa Beans as Affected by Pulp Pre-Conditioning and
Fermentation.” Journal of Food Science and Technology 50 (6). Springer: 1097–
105. doi:10.1007/s13197-011-0446-5.
Amoa-Awua, W.K., M. Madsen, J. Takrama, A. Olaiya, L. Ban-Koffi, and M. Jakobsen.
2007. Quality Manual for Production & Primary Processing of Cocoa. https​://cs​
irspa​ce.fo​odres​earch​gh.si​te/ha​ndle/​12345​6789/​1248.​
Amoye, S. 2006. “Cocoa Sourcing, World Economics and Supply.” The Manufacturing
Confectioner 86 (1): 81–85.
APG. 2009. “An Update of the Angiosperm Phylogeny Group Classification for the Orders
and Families of Flowering Plants: APG III.” Botanical Journal of the Linnean Society
161 (2). John Wiley & Sons (10.1111): 105–21. doi:10.1111/j.1095-8339.2009.00996.x.
Aprotosoaie, Ana Clara, Simon Vlad Luca, and Anca Miron. 2016. “Flavor Chemistry
of Cocoa and Cocoa Products-An Overview.” Comprehensive Reviews in Food
Science and Food Safety 15 (1): 73–91. doi:10.1111/1541-4337.12180.
Ardhana, M. 2003. “The Microbial Ecology of Cocoa Bean Fermentations in Indonesia.”
International Journal of Food Microbiology 86 (1–2). Elsevier: 87–99. doi:10.1016/
S0168-1605(03)00081-3.
Ascrizzi, Roberta, Guido Flamini, Cecilia Tessieri, and Luisa Pistelli. 2017. “From the
Raw Seed to Chocolate: Volatile Profile of Blanco de Criollo in Different Phases of
the Processing Chain.” Microchemical Journal 133 (July). Elsevier B.V.: 474–79.
doi:10.1016/j.microc.2017.04.024.
Awua, P.K. 2002. Cocoa Processing and Chocolate Manufacture in Ghana. Saffron
Walden, Essex, UK: David Jamieson and Associates Press.
Beckett, S.T. 1994. “Control of Particle Size Reduction during Chocolate Grinding.”
Manufacturing Confectioner 74 (5): 90–97.
410 Food Aroma Evolution

Beckett, S.T. 2009. Industrial Chocolate Manufacture and Use, 4th ed.
doi:10.1002/9781444301588.
Beckett, S.T. 2015. The Science of Chocolate, 2nd ed. Cambridge, UK: Royal Society of
Chemistry. https​://pu​bs.rs​c.org ​/en/c​onten​t /ebo​ok/97​8-1-7​8801-​235-5​.
Biehl, B. and G. Ziegleder. 2003a. “COCOA | Chemistry of Processing.” In Encyclopedia
of Food Sciences and Nutrition, edited by B. Caballero, 2nd ed., pp. 1436–48.
Elsevier. doi:10.1016/B0-12-227055-X/00261-3.
Biehl, B. and G. Ziegleder. 2003b. “COCOA | Production, Products, and Use.” In
Encyclopedia of Food Sciences and Nutrition, pp. 1448–63. Elsevier. doi:10.1016/
B0-12-227055-X/00262-5.
Biehl, Böle, Christa Wewetzer, and Detlef Passern. 1982. “Vacuolar (Storage) Proteins
of Cocoa Seeds and Their Degradation during Germination and Fermentation.”
Journal of the Science of Food and Agriculture 33 (12). John Wiley & Sons: 1291–
304. doi:10.1002/jsfa.2740331216.
Bonvehí, J. Serra. 2005. “Investigation of Aromatic Compounds in Roasted Cocoa
Powder.” European Food Research and Technology 221 (1–2): 19–29. doi:10.1007/
s00217-005-1147-y.
Buyukpamukcu, Elif, David M. Goodall, Carl-Erik Hansen, Brendan J. Keely, Sunil
Kochhar, and Hans Wille. 2001. “Characterization of Peptides Formed during
Fermentation of Cocoa Bean.” Journal of Agricultural and Food Chemistry 49 (12).
American Chemical Society: 5822–27. doi:10.1021/jf0104127.
Chase, M.W. and J.L. Reveal. 2009. “A Phylogenetic Classification of the Land Plants to
Accompany APG III.” Botanical Journal of the Linnean Society 161 (2). John Wiley
& Sons (10.1111): 122–27. doi:10.1111/j.1095-8339.2009.01002.x.
Cheesman, E. 1944. “Notes on the Nomenclature, Classification and Possible
Relationships of Cococa Populations.” Tropical Agriculture 21: 144–59.
Chetschik, Irene, Markus Kneubühl, Karin Chatelain, Ansgar Schlüter, Konrad Bernath,
and Tilo Hühn. 2018. “Investigations on the Aroma of Cocoa Pulp (Theobroma
cacao L.) and Its Influence on the Odor of Fermented Cocoa Beans.” Journal of
Agricultural and Food Chemistry 66 (10): 2467–72. doi:10.1021/acs.jafc.6b05008.
Counet, Christine, Caroline Ouwerx, Delphine Rosoux, and Sonia Collin. 2004.
“Relationship between Procyanidin and Flavor Contents of Cocoa Liquors from
Different Origins.” Journal of Agricultural and Food Chemistry 52 (20): 6243–49.
doi:10.1021/jf040105b.
Counet, Christine, Delphine Callemien, Caroline Ouwerx, and Sonia Collin. 2002. “Use
of Gas Chromatography−Olfactometry To Identify Key Odorant Compounds in
Dark Chocolate. Comparison of Samples before and after Conching.” Journal of
Agricultural and Food Chemistry 50 (8). American Chemical Society: 2385–91.
doi:10.1021/jf0114177.
Crafack, Michael, Hanna Keul, Carl Emil Eskildsen, Mikael A. Petersen, Sofie Saerens,
Andreas Blennow, Mathias Skovmand-Larsen, et al. 2014. “Impact of Starter
Cultures and Fermentation Techniques on the Volatile Aroma and Sensory
Profile of Chocolate.” Food Research International 63: 306–16. doi:10.1016/j.
foodres.2014.04.032.
Cronquist, Arthur. 1981. An Integrated System of Classification of Flowering Plants.
New York, NY: Columbia University Press.
Despréaux, D. 1998. “Le Cacaoyer et La Cacaoculture.” In Cacao et Chocolatsproduction,
Utilisation, Caractéristiques, edited by J. Pontillon, pp. 44–93. Paris, France:
Technique & Documentation Lavoisier.
 Aroma Evolution in Chocolate Production 411

Dimick, P.S. and J.C. Hoskin. 1999. “Industrial Chocolate Manufacture and Use.” In
Industrial Chocolate Manufacture and Use, edited by S.T. Beckett, 3rd ed. Oxford,
Malden, MA: Blackwell Publishing.
Dimick, P.S. and J.M. Hoskin. 1981. “Chemico-Physical Aspects of Chocolate
Processing—A Review.” Canadian Institute of Food Science and Technology
Journal 14 (4). Elsevier: 269–82. doi:10.1016/S0315-5463(81)72927-4.
Engeseth, Nicki J. and Marlon Fernando Ac Pangan. 2018. “Current Context on
Chocolate Flavor Development — A Review.” Current Opinion in Food Science 21
(June). Elsevier: 84–91. doi:10.1016/j.cofs.2018.07.002.
Eskes, A.B., S.D. Guarda, C.L. Garcia, and R. Garcia. 2007. “Is Genetic Variation for
Sensory Traits of Cocoa Pulp Related to Fine Flavour Cocoa Traits?” INGENIC
Newsletters 11 (June): 22–28. doi:10.1002/art.37993.
Fowler, M.S. 1999. “Cocoa Beans: From Tree to Factory.” In Industrial Chocolate
Manufacture and Use, edited by S.T. Beckett, pp. 8–35. Oxford, UK: Wiley-Blackwell.
Fowler, M.S., P. Leheup, and J.-L. Cordier. 1998. “Cocoa, Coffee and Tea.” In
Microbiology of Fermented Foods, pp. 128–47. Boston, MA: Springer US.
doi:10.1007/978-1-4613-0309-1_5.
Frauendorfer, Felix and Peter Schieberle. 2006. “Identification of the Key Aroma
Compounds in Cocoa Powder Based on Molecular Sensory Correlations.” Journal
of Agricultural and Food Chemistry 54 (15): 5521–29. doi:10.1021/jf060728k.
Frauendorfer, Felix and Peter Schieberle. 2008. “Changes in Key Aroma Compounds of
Criollo Cocoa Beans during Roasting.” Journal of Agricultural and Food Chemistry
56 (21): 10244–51. doi:10.1021/jf802098f.
Giacometti, J., S. Mazor Jolić, and D. Josić. 2015. “Cocoa Processing and Impact on
Composition.” In Processing and Impact on Active Components in Food, edited by V.
Preedy, pp. 605–12. London, UK: Academic Press. doi:10.1016/C2012-0-02526-4.
González, Elaine. 2007. “Chocolate—Mexico’s Living Legacy.” The Manufacturing
Confectioner.
Granvogl, Michael, Susanne Bugan, and Peter Schieberle. 2006. “Formation of Amines
and Aldehydes from Parent Amino Acids during Thermal Processing of Cocoa and
Model Systems: New Insights into Pathways of the Strecker Reaction.” Journal of
Agricultural and Food Chemistry 54 (5). American Chemical Society: 1730–39.
doi:10.1021/jf0525939.
HoVan Thi Thuy, Jian Zhao, and Graham Fleet. 2014. “Yeasts Are Essential for Cocoa
Bean Fermentation.” International Journal of Food Microbiology 174 (March).
Elsevier: 72–87. doi:10.1016/J.IJFOODMICRO.2013.12.014.
Hoskin, J.C. and P.S. Dimick. 1984. “Role of Nonenzymatic Browning during the
Processing of Chocolate—A Review.” Process Biochemistry 11: 92–104.
Jahurul, M.H.A., I.S.M. Zaidul, N.A.N. Norulaini, F. Sahena, S. Jinap, J. Azmir,
K.M. Sharif, and A.K. Mohd Omar. 2013. “Cocoa Butter Fats and Possibilities of
Substitution in Food Products Concerning Cocoa Varieties, Alternative Sources,
Extraction Methods, Composition, and Characteristics.” Journal of Food
Engineering 117 (4). Elsevier: 467–76. doi:10.1016/j.jfoodeng.2012.09.024.
Jinap, S., P.S. Dimick, and R. Hollender. 1995. “Flavour Evaluation of Chocolate
Formulated from Cocoa Beans from Different Countries.” Food Control 6 (2): 105–
10. doi:10.1016/0956-7135(95)98914-M.
Jinap, S, W.I. Wan Rosli, A.R. Russly, and L.M. Nordin. 1999. “Effect of Roasting Time
and Temperature on Volatile Component Profiles during Nib Roasting of Cocoa Beans
(Theobroma Cacao).” Journal of the Science of Food and Agriculture 77 (4): 441–48.
412 Food Aroma Evolution

Kadow, Daniel, Joerg Bohlmann, Wilberth Phillips, and Reinhard Lieberei. 2013.
“Identification of Main Fine or Flavour Components in Two Genotypes of the
Cocoa Tree (Theobroma Cacao L.).” Journal of Applied Botany and Food Quality
86: 90–98. doi:10.5073/JABFQ.2013.086.013.
Kattenberg, H. and A. Kemming. 1998. “The Flavor of Cocoa in Relation to the Origin
and Processing of the Cocoa Beans.” In Food Flavors, Ingredients and Composition,
edited by G. Charalambous, pp. 1–22. Amsterdam, the Netherlands: Elsevier Science.
Knezevic, G. 1983. “Über Den Metallgehalt in Kakaobohnen Und-Schalen.” Zucker-
Und Süßwaren Wirtschaft 36 (10): 319–20.
Krings, Ulrich, Ralf G. Berger, and Dattatreya S. Banavara. 2003. “Thin Layer High
Vacuum Distillation to Isolate the Flavor of High-Fat Food.” European Food
Research and Technology 217 (1): 70–73. doi:10.1007/s00217-003-0700-9.
Kuster, W. 1984. “Liquor Grinding.” Manufacturing Confectioner 64 (8): 47–55.
Luna, Fabienne, Dominique Crouzillat, Loïc Cirou, and Peter Bucheli. 2002. “Chemical
Composition and Flavor of Ecuadorian Cocoa Liquor.” Journal of Agricultural and
Food Chemistry 50 (12). American Chemical Society: 3527–32. doi:10.1021/jf0116597.
Maniere, F.Y. and P.S. Dimick. 1979. “Effects of Conching on the Flavour and Volatile
Components of Dark Semi-Sweet Chocolate.” Lebensmittel-Wissenschaft u.
Technologie 12: 102–07.
McShea, Andrew, Emma Ramiro-Puig, Sandra B. Munro, Gemma Casadesus, Margarida
Castell, and Mark A. Smith. 2008. “Clinical Benefit and Preservation of Flavonols
in Dark Chocolate Manufacturing.” Nutrition Reviews 66 (11). Oxford University
Press: 630–41. doi:10.1111/j.1753-4887.2008.00114.x.
Mohr, W. 1978. “Developments in Cocoa Technology.” Lebensmittelchemie u
Gerichtliche Chemie 32: 27–31.
Mohr, W., E.A. Niediek, and H. Lange. 1968. “Conchieren von Milchfreien
Schokolademassen.” Süsswaren 24: 1375–83.
Mongia, G. 1997. Particle Size Distribution Affects the Rheology and Sensory Attributes
of Milk Chocolate. University Park, PA: Pennsylvania State University.
Motamayor, Juan C., Philippe Lachenaud, Jay Wallace da Silva e Mota, Rey Loor, David
N. Kuhn, J. Steven Brown, and Raymond J. Schnell. 2008. “Geographic and Genetic
Population Differentiation of the Amazonian Chocolate Tree (Theobroma Cacao
L.). Edited by Justin O. Borevitz.” PLoS ONE 3 (10). Public Library of Science:
e3311. doi:10.1371/journal.pone.0003311.
Nair, K.P. Prabhakaran. 2010. “Cocoa (Theobroma Cacao L.).” In The Agronomy and
Economy of Important Tree Crops of the Developing World, pp. 131–80. Elsevier.
doi:10.1016/B978-0-12-384677-8.00005-9.
Niediek, E.A. 1994. “Particle Size Reduction.” In Industrial Chocolate Manufacture
and Use, edited by S.T. Beckett, pp. 83–101. Boston, MA: Springer US.
doi:10.1007/978-1-4615-2111-2_7.
Nielsen, D.S. 2006. The Microbiology of Ghanaian Cocoa Fermentations. Frederiksberg,
Denmark: Royal Veterinary and Agricultural University.
Osman, H., R. Nasarudin, and S.L. Lee. 2004. “Extracts of Cocoa (Theobroma Cacao
L.) Leaves and Their Antioxidation Potential.” Food Chemistry 86 (1). Elsevier:
41–46. doi:10.1016/j.foodchem.2003.08.026.
Owusu, Margaret, Mikael Agerlin Petersen, and Hanne Heimdal. 2013. “Relationship
of Sensory and Instrumental Aroma Measurements of Dark Chocolate as Influenced
by Fermentation Method, Roasting and Conching Conditions.” Journal of Food
Science and Technology 50 (5). doi:10.1007/s13197-011-0420-2.
 Aroma Evolution in Chocolate Production 413

Páramo, D., P. García-Alamilla, M.A. Salgado-Cervantes, V.J. Robles-Olvera, G.C.

Rodríguez-Jimenes, and M.A. García-Alvarado. 2010. “Mass Transfer of Water and
Volatile Fatty Acids in Cocoa Beans during Drying.” Journal of Food Engineering
99 (3). Elsevier: 276–83. doi:10.1016/j.jfoodeng.2010.02.028.
Pettipher, G.L. 1990. “The Extraction and Partial Purification of Cocoa Storage
Proteins.” Café, Cacao, Thé 34 (1): 23–26.
Pino, Jorge A., Liena Ceballos, and Clara E. Quijano. 2010. “Headspace Volatiles of
Theobroma cacao L. Pulp from Colombia.” Journal of Essential Oil Research 22 (2).
Taylor & Francis Group: 113–15. doi:10.1080/10412905.2010.9700276.
Qin Xiao-Wei, Jian-Xiong Lai, Le-He Tan, Chao-Yun Hao, Fu-Peng Li, Shu-Zhen He, and
Ying-Hui Song. 2016. “Characterization of Volatile Compounds in Criollo, Forastero
and Trinitario Cocoa Seeds (Theobroma cacao L.) in China.” International Journal
of Food Properties 2912 (October). Taylor & Francis: 10942912.2016.1236270. doi
:10.1080/10942912.2016.1236270.
Ramli, N., O. Hassan, M. Said, W. Samsudin, and N.A. Idris. 2006. “Influence of
Roasting Conditions on Volatile Flavor of Roasted Malaysian Cocoa Beans.”
Journal of Food Processing and Preservation 30 (3). John Wiley & Sons (10.1111):
280–98. doi:10.1111/j.1745-4549.2006.00065.x.
RamosCíntia Lacerda, Disney Ribeiro Dias, Maria Gabriela da Cruz Pedrozo Miguel,
and Rosane Freitas Schwan. 2014. “Impact of Different Cocoa Hybrids (Theobroma
Cacao L.) and S. Cerevisiae UFLA CA11 Inoculation on Microbial Communities
and Volatile Compounds of Cocoa Fermentation.” Food Research International 64
(October). Elsevier: 908–18. doi:10.1016/j.foodres.2014.08.033.
Reineccius, Gary and Henry B. Heath. 2005. Flavor Chemistry and Technology, 2nd ed.
Taylor & Francis.
Reineccius, Gary A., Philip G. Keeney, and Wendy Weissberger. 1972. “Factors
Affecting the Concentration of Pyrazines in Cocoa Beans.” Journal of Agricultural
and Food Chemistry 20 (2). American Chemical Society: 202–06. doi:10.1021/
jf60180a032.
Rodriguez-Campos, J., H.B. Escalona-Buendía, I. Orozco-Avila, E. Lugo-Cervantes, and
M.E. Jaramillo-Flores. 2011. “Dynamics of Volatile and Non-Volatile Compounds
in Cocoa (Theobroma cacao L.) during Fermentation and Drying Processes Using
Principal Components Analysis.” Food Research International 44 (1): 250–58.
doi:10.1016/j.foodres.2010.10.028.
Rodriguez-Campos, J., H.B.B. Escalona-Buendía, S.M.M. Contreras-Ramos, I. Orozco-
Avila, E. Jaramillo-Flores, and E. Lugo-Cervantes. 2012. “Effect of Fermentation
Time and Drying Temperature on Volatile Compounds in Cocoa.” Food Chemistry
132 (1). Elsevier: 277–88. doi:10.1016/j.foodchem.2011.10.078.
Schnermann, Petra and Peter Schieberle. 1997. “Evaluation of Key Odorants in Milk
Chocolate and Cocoa Mass by Aroma Extract Dilution Analyses.” Journal of
Agricultural and Food Chemistry 45 (3). American Chemical Society: 867–72.
doi:10.1021/jf960670h.
Schwab, Wilfried, Rachel Davidovich-Rikanati, and Efraim Lewinsohn. 2008.
“Biosynthesis of Plant-Derived Flavor Compounds.” Plant Journal 54 (4): 712–32.
doi:10.1111/j.1365-313X.2008.03446.x.
Schwan, Rosane F. and Alan E. Wheals. 2004. “The Microbiology of Cocoa Fermentation
and Its Role in Chocolate Quality.” Critical Reviews in Food Science and Nutrition
44 (4): 205–21. doi:10.1080/10408690490464104.
414 Food Aroma Evolution

Sukha, Darin A., David R. Butler, Pathmanathan Umaharan, and Emma Boult. 2008.
“The Use of an Optimised Organoleptic Assessment Protocol to Describe and
Quantify Different Flavour Attributes of Cocoa Liquors Made from Ghana and
Trinitario Beans.” European Food Research and Technology 226 (3): 405–13.
doi:10.1007/s00217-006-0551-2.
Thompson, Sterling S., Kenneth B. Miller, Alex S. Lopez, and Nicholas Camu. 2013.
“Cocoa and Coffee.” In Food Microbiology, pp. 881–99. American Society of
Microbiology. doi:10.1128/9781555818463.ch35.
TranPhuong Diem, Davy Van de Walle, Nathalie De Clercq, Ann De Winne, Daniel
Kadow, Reinhard Lieberei, Kathy Messens, Dung Nhan Tran, Koen Dewettinck,
and Jim Van Durme. 2015. “Assessing Cocoa Aroma Quality by Multiple Analytical
Approaches.” Food Research International 77. Elsevier: 657–69. doi:10.1016/j.
foodres.2015.09.019.
Urbanski, J.J. 1992. “Chocolate Flavor/Origins and Descriptions on the Effects of Process
and Bean Source.” The Manufacturing Confectioner 11: 69–82.
Voigt, Jürgen, Böle Biehl, and Syed Kamaruddin Syed Wazir. 1993. “The Major Seed
Proteins of Theobroma cacao L.” Food Chemistry 47 (2). Elsevier: 145–51.
doi:10.1016/0308-8146(93)90236-9.
Warren, J.M. and A.J. Kennedy. 1991. “Cocoa Breeding Revisited.” Cocoa Growers’
Bulletin 44: 18–24.
Wood, G.A.R. and R.A. Lass. 2001. Cocoa, 4th ed. Blackwell Science. doi:10.1002/
9780470698983.
Yu, Ai Nong and Ai Dong Zhang. 2010. “The Effect of PH on the Formation of Aroma
Compounds Produced by Heating a Model System Containing L-Ascorbic Acid
with l-Threonine/l-Serine.” Food Chemistry 119 (1). Elsevier: 214–19. doi:10.1016/j.
foodchem.2009.06.026.
Zarrillo, Sonia, Nilesh Gaikwad, Claire Lanaud, Terry Powis, Christopher Viot, Isabelle
Lesur, Olivier Fouet, et al. 2018. “The Use and Domestication of Theobroma cacao
during the Mid-Holocene in the Upper Amazon.” Nature Ecology & Evolution 2
(12): 1879–88. doi:10.1038/s41559-018-0697-x.
Ziegleder, G. 2009. “Flavour Development in Cocoa and Chocolate.” In Industrial
Chocolate Manufacture and Use, edited by Stephen T. Beckett, pp. 169–91. Oxford,
UK: Wiley-Blackwell. doi:10.1002/9781444301588.ch8.
Ziegleder, Gottfried. 1990. “Linalool Contents as Characteristic of Some Flavor Grade
Cocoas.” Zeitschrift Für Lebensmittel-Untersuchung Und Forschung 191 (4–5).
Springer-Verlag: 306–09. doi:10.1007/BF01202432.
Ziegleder, Gottfried. 1991. “Composition of Flavor Extracts of Raw and Roasted
Cocoas.” Zeitschrift Für Lebensmittel-Untersuchung Und-Forschung 192 (6).
Springer-Verlag: 521–25. doi:10.1007/BF01202506.
 Chapter 21
Bakery Products
Joana Pico and Juan Carlos Diego

CONTENTS

21.1 Importance of Aroma in the Baking Market 415

21.2 The Technology of Making Main Baked Goods 416
21.2.1 Bread and Toast Elaboration 418
21.2.2 Cake Elaboration 418
21.2.3 Biscuit and Cracker Elaboration 420
21.3 The Aroma of Baked Goods from a Biological Point of View 421
21.3.1 Fermentation Process 422
21.3.2 Lipid Oxidation Process 423
21.3.3 Maillard Reaction and Caramelization 424
21.4 Evolution of Volatile Compounds during Processing and Storage 426
21.4.1 From the Raw Materials to the Baked Product 427
21.4.2 Toasting of Bread 430
21.4.3 Changes in the Aroma Quality during Storage 431
21.5 General Conclusions 432
References 433

21.1 IMPORTANCE OF AROMA IN THE BAKING MARKET

The baking of bread is considered to be one of the oldest human activities (Cauvain,
1998). Bread and other baked products are simply a mixture of water, flour, and yeast, or
another leavening agent, with more ingredients that enhance its flavor and texture. The
ingredients themselves and, above all, the processing steps (e.g., kneading, fermentation,
baking) and storage conditions, determine the final flavor of the baked good. It is a fact
that aroma is the most important characteristic of a food’s flavor (Picó, 2012); indeed,
consumers’ acceptance and the commercial values of food products depend considerably
on the composition and amount of aroma substances (Cserháti, 2010). On the other
hand, nowadays, leading chefs consider bread and bakery goods not solely as a comple-
ment to the dishes they serve but also a fundamental aspect of their innovative offerings
(Martinez-Monzo, Garcia-Segovia, and Albors-Garrigos, 2014). Thus, it is clear that
baked goods and their corresponding aromas play a significant role in the current food
market. Overall, the world is experiencing an increase in demand for bakery products
with a compound annual growth rate (CAGR) of 4.5% from 2013 to 2017. Moreover, it
is expected to have a CAGR of 5.7% between 2018 and 2022, with global sales reaching
US$464.4 billion (Agrifood Canada website). Breads are the largest segment, followed by
cakes and pastries, as is indicated in Table 21.1. In 2017, the expenditure per capita in the

415
416 Food Aroma Evolution

TABLE 21.1 Historic Expenditure per Capita on Baked

Goods in the United States in US$ from 2013 to 2017
Category 2013 2014 2015 2016 2017
Bread 72.5 71.9 71.6 71.9 71.8
Cakes 47.2 48.5 49.7 51.7 52.6
Dessert mixes 5.5 5.2 4.7 4.5 4.4
Frozen baked goods 7.3 7.3 7.2 7.1 6.9
Pastries 40.0 40.9 42.0 43.2 44.2
Total baked goods 172.5 173.8 175.2 178.4 179.9
Source: Agrifood Canada website.

United States for bread was US$71.8, for cakes it was US$52.6, and for pastries it was
US$44.2 (Agrifood Canada website).
The modern baking industry is mostly based on wholesale bakeries, which mainly
sell bread, cookies, crackers, cakes, and related frozen bakery products. These wholesale
bakeries are equipped with extensive production facilities and reach consumers through
retail locations (Martinez-Monzo, Garcia-Segovia, and Albors-Garrigos, 2014). In order
to get attractive prices for consumers, wholesale industry tends to reduce costs, which
usually involves lessening the production time and sometimes the quality of the raw
materials or nutritional properties. Consequently, lower fermentation times (Zehentbauer
and Grosch, 1998b), lower fermentation temperatures associated to higher yeast concen-
tration (Birch, Petersen, and Hansen, 2013), or cheaper ways of baking, like the electrical
oven instead of wood oven (Bianchi et al., 2008), are employed, compromising the flavor
of the baked product. On the other hand, the market is also challenged by health and diet
concerns, like sugar, fat, or salt-reduced content, gluten-free products, or high-fiber level.
Consumers are searching for better-for-you treats that still taste good; they are avoiding
processed or artificial ingredients, but they are not willing to sacrifice the flavor (Agrifood
Canada website). Therefore, the analysis of volatile compounds in baked goods becomes
essential to understand how the processing steps, new ingredients, and storage affects the
aroma, so as to change the production accordingly and improve the final flavor.

21.2 THE TECHNOLOGY OF MAKING MAIN BAKED GOODS

Understanding the processing steps of baked goods is of utmost importance in order to

explain and foresee the consequent changes in the aroma quality. This makes it possible to
improve the processing or storage stages, maintaining a pleasant aroma in the final product.
Although there are a lot of schemes to classify bakery products, the most common
way for cereal chemists has been based on the source of leavening. The four main catego-
ries are (Matz, 1960): (i) yeast leavened bakery goods, which includes products in which
yeast serves principally as the aerating agent (i.e., white bread, rolls bread, panettone,
sweet bread, whole wheat bread or sour bread like rye bread) and products in which the
aeration action is secondary and yeasts help to dough conditioning and flavor developing
(i.e., Danish pastry); (ii) chemically leavened bakery goods, mainly using sodium bicar-
bonate with or without leavening acids or baking powder (i.e., white layer cake, muffins
or chocolate cake); (iii) air leavened bakery goods, which can incorporate the air using
highly emulsified shortenings, whipping eggs and sugar, creaming shortening and sugar,
 Bakery Products 417

aerated flour or mixing under pressure (i.e., angel food cake, sponge cake, chiffon cake,
pound cake, or wafer biscuits); (iv) unleavened bakery goods, without chemical or bio-
logical leavening agents added and the use of very weak flours with low protein content
(i.e., pie crust, fried pie, crust puff pastry, cookies, or brownies).
In general, the making of baked goods consists of mixing and kneading the ingredi-
ents, proofing the dough or batter using yeast or physical/chemical agents, and baking the
final product. All these steps lead to changes in the composition and structure from the
dough or batter to the final product that, undoubtedly, affect the generation of the aroma.
Mixing is the initial step, in which the type of mixer and the intensity of time of mixing
will determine the state of dispersion of the ingredients and, in turn, the efficiency of pro-
cessing and the quality of the final product. When water is added to the dry flour, around
30–35% is bounded to the flour, generating a “one-phase system,” and the rest is going
to be free water. This liquid phase dissolves the soluble components of flour, provides the
medium for the reaction to take place, and dissolves the carbon dioxide for diffusing the
gas cells (Blanshard, Frazier, and Galliard, 1985). The occlusion of air is crucial in the
mixing step for cake-making, where whipping is necessary to introduce the air necessary
for the batter expansion. However, it is also crucial in bread since the bubbles are the
nuclei of the gas cells that locate the carbon dioxide generated during fermentation. This
air becomes really important for the generation of volatile compounds, since it can be used
for the yeast to grow or for the lipoxygenase during lipid oxidation. As has been indicated,
there are different ways of proofing the dough or batter, which are also decisive for the
generation of volatile compounds or its precursors. Yeast produces volatile compounds
directly through the fermentation of sugars, the glycolysis of pyruvic acid, or the Ehrlich
pathway (Pico, Bernal, and Gómez, 2015), but it also affects the texture of the dough and
baked product since it provides the carbon dioxide necessary for the dough expansion
and the creation of the crumb structure (Blanshard, Frazier, and Galliard, 1985; Pico,
2018). For chemically leavened goods, the source of carbon dioxide is sodium bicarbonate
or potassium bicarbonate in sodium-free diets. Sodium bicarbonate is tasteless and when
dissolved in water, gives a mixture of ions and carbon dioxide at a pH higher than 8. To
control the rate of evolution and yield of the gas, acids are added to the dough, but they
are also tasteless (e.g., phosphates). Baking powder is usually employed, which presents a
mixture of sodium bicarbonate and acids. In these products, soft flours with less exten-
sible proteins are employed, which makes the use of shortenings in the batters necessary
to entrap the air during mixing (Matz, 1960). Finally, the baking step is similar in all the
baked goods, differing mainly in the temperature and time of baking as well as in the rela-
tive humidity (RH %) of the oven. During baking, as the temperature goes up, important
physical, chemical, and biological changes happen (Pico, 2018): (i) the activity of yeast (if
added) and of the endogenous or added enzymes is finished, which leads to the supres-
sion of the generation of volatile compounds arising from fermentation; (ii) the ethanol
is evaporated (when yeast was added); (iii) the water is evaporated, giving crispiness and
color to the crust but also reducing some volatile compounds that are soluble in the water
vapor; (iv) the Maillard reaction between reducing sugars and amino acids generates not
only color but also a great amount of pleasant aroma compounds, starting at 110°C; and
(v) starch is gelatinized, starting at 60°C. Starch gelatinization is the most perceptible
change in a dough or batter during baking and explains its transformation from the vis-
cous dough or batter into the solid baked product. The degree of gelatinization strongly
depends on the water available, progressing more completely when more water is avail-
able. Then, the degree of gelatinization is different between baked products: in biscuits,
the starch granule can still be observed; in wafers, it is totally destroyed; and in bread,
418 Food Aroma Evolution

some of them are intact while others are disrupted and can be enzymatically degraded.
Fats, salt, and emulsifiers delay the gelatinization, while the effect of sugar is not clear
(Blanshard, Frazier, and Galliard, 1985). The state of the starch is really important for
determining the interaction between the volatile compounds and amylose, which impacts
the release of the volatile compounds from the baked matrix (Arvisenet et al., 2002).

21.2.1 Bread and Toast Elaboration

Bread is the most sold baked good, even though satisfactory white bread can be made
at home with just flour, water, yeast (or sourdough), and salt. Italian white bread reci-
pes are usually based on Italian white bread recipes are usually based on 60% water,
1.75% compressed yeast, and 1.75% salt, on a percentage of flour basis (Matz, 1960).
Traditional French bread has been characterized by a very crispy crust, an irregular and
aerated crumb, and an authentic bread taste. The French government, in 1993, decreed
that traditional French bread can only be made with four ingredients: traditional wheat
flour, water, salt, and yeast and/or a leavening agent. It is usually based on 60% water,
1.8% salt, 1.5% yeast, and 30% of a previously fermented dough prepared by mixing
wheat flour, yeast (1.5%), salt (1.8%), and water (66%), all the ingredients as % of flour
basis. Also, these traditional French baguettes cannot be frozen at any stage; neither do
they contain additives nor preservatives (Jouquand et al., 2018). The raw materials, the
addition of emulsifiers and additives (i.e., ascorbic acid, bromated, or azodicarbonamide),
the use of enzymes (e.g., amylases, proteases, lipoxygenases or lipases), the formulation
as well as the bread-making steps, influence the final quality of bread (Belitz, Grosch,
and Schieberle, 2009; Cauvain, 2003; Cho and Peterson, 2010; Martínez-Anaya, 1996).
Indeed, there is a relationship between the processing stages and the changes in the com-
position and structure from the dough to the final bread, as can be seen in Figure 21.1.
The main steps in the elaboration of bread are (Pico, 2018): (i) kneading, where
the gluten structure starts to be created, and the air creates bubbles in the dough that
provide the oxygen necessary for the yeast to grow and for the lipoxygenase to inititate
the oxidation reaction; (ii) fermentation, usually using Saccharomyces cerevisiae. The
starch is progressively converted into sugar by amylase action, which is employed by yeast
to produce, mainly, carbon dioxide and ethanol. Fermentation is usually carried out in
a controlled atmosphere between 30°C and 45°C and 85% of relative humidity (RH);
(iii) baking, at standard temperatures between 190°C and 250°C, the core temperature
being around 92–96°C. There are different physicochemical changes involved, as it was
previously explained, with the formation of the crust and generation of color and a huge
number of aroma compounds by Maillard reaction. If a slice of bread is placed in front
of a high-temperature predominantly radiation-based source for a certain period, toasted
bread is obtained. Toasting is considered a wonderful option compared with fresh bread,
due to the easier digestion, a delay in staling, and increased flavor as a consequence of
Maillard and caramelization reactions (Kirit, Erdogdu, and Ozdemir, 2013).

21.2.2 Cake Elaboration

Cakes are prepared mainly through the baking of a batter, which is an oil–water type
emulsion with air bubbles entrapped in the fat phase and the rest of the ingredients dis-
persed or dissolved in the water phase. When the batter is baked, the volume of the cake
 Bakery Products 419

FIGURE 21.1 Relationship between the processing stages and the changes in the com-
position and structure from the dough to the final bread. (From Joana Pico, Doctoral
Thesis, 2018, http:​//uva​doc.u​va.es​/ hand​le/10​324/2​9499.)

increases around 3.5 times during baking as a consequence of the air and/or carbon
dioxide employed (10% of the expansion) and the evaporation of water within the bubble
(90% of the expansion). Fine bubbles of a homogeneous size are required for a suitable
textured cake (Bennion, Stewart, and Bamford, 1966). The coalescence of the bubbles
leads to losses of gas, which also implies losses of volatile compounds by swiping.
The main methods for preparing a cake are (Bennion, Stewart, and Bamford, 1966):
(i) the sugar batter method, where fat and castor sugar are creamed together for about 10
minutes, followed by stirring with egg for 5–7 minutes and lastly the addition of flour,
milk, and water; (ii) the flour batter method, where the fat is creamed with the same
amount of flour as fat and the egg is whipped with the same amount of sugar as egg,
and all the ingredients carefully whisked for a creamy mass; (iii) the sugar/flour batter
method, similar to the flour batter method but with the egg and sugar mixed as a solu-
tion; (iv) the dry crumbly batter method; (v) the blending crumbly at stage I method,
which consists of three steps. First, sieved sugar, dried egg, milk powder, salt, fat, and
its weight in flour are slowly mixed. Then, water is added in equal weight to the fat and
flour and mixed for 5–7 minutes, and in the last step the rest of the ingredients are added;
(vi) the blending paste at stage I method, used for high-ratio cakes. In the first step, the
flour, fat, baking powder, and special ingredients are mixed into a smooth paste. Then,
the sugar, salt, milk, and water (40% of the flour weight) are mixed for 1–2 minutes, and
in the last step the eggs are mixed at a slow speed; (vii) the continuous mixing method,
with all the ingredients together in the machine, with an efficient dispersion; (vii) the
all-in high-speed method, with all the ingredients whisked in a high speed dough mixer.
Examples of chemically leavened cakes are white layer cakes, with 25.6% flour,
25.6% sugar, 11.2% shortening, 16.0% egg whites, 10.2% milk powder, 1.6% baking
powder, 0.6% salt, and 9.2% water (Matz, 1960). Sponge cakes are the most common
type of air leavened cake, which are prepared by mixing 10 eggs, an amount of sugar
420 Food Aroma Evolution

equivalent to the weight of 10 eggs, and an amount of flour equivalent to the weight of
six eggs. The mixture is beaten for 30 minutes and usually baked for another 30 minutes
(Matz, 1960). A special type of cake is the brioche, since it is usually fermented for 30
minutes at 26°C, and commonly prepared with flour (19.4%), yeast (6.1%), warm milk
(24.2%), butter (19.4%), eggs (19.4%), sugar (4.8%), and salt (6.7%).

21.2.3 Biscuit and Cracker Elaboration

Biscuits are cereal products baked to a moisture content of less than 5%, enriched mainly
with fat and sugar and other secondary ingredients such as chocolate. The method of
dough piece forming depends on the enrichment of the formulation with fat and sugar,
while the secondary processing adds greatly to the enjoyment of biscuits (Manley, 2011).
The terminology “biscuit” is usually employed in the UK and Ireland, while “cookie” is
used in the United States and Canada. There is great controversy about the use of biscuit
and cookie as synonyms since in the UK the term cookie usually refers to a chocolate chip
cookie. According to other traditional recipes from Eastern European countries cookies
are jam-filled, and in other parts of Europe they include a percentage of almond flour.
Moreover, when biscuits are more or less unsweetened, salty, and crispy, they are called
crackers, which are considered bread substitutes usually eaten with butter, cheese, or cold
meat as a snack.
The main biscuits consumed nowadays are (Manley, 2011; Matz, 1960): (i) crack-
ers, which are an unsweetened mixture of flour, fat, and salt, sometimes fermented with
yeast, and sheeted and laminated prior to gauging, cutting, and baking. Depending on
the degree of sweetness, if they are fermented or not, and the percentage of fat in the
dough, it is possible to talk about matzos, puff crackers, cream crackers, soda crackers,
or savory crackers. As an example, the typical all-in dough cream cracker recipe consists
of 50% strong flour, 50% weak flour and 1.0% malt flour, diastatic, 1.0% sugar, 1.5%
malt extract non-diastatic, 16.0% fat, 0.2% sodium carbonate, 1.7% yeast, 1.2% salt,
34.0% water (on a percentage of 100% flour basis), and a fermentation time of between
4 and 16 hours; (ii) puff biscuits, which are made from non-fermented and undeveloped
dough with a non-homogeneous distribution of fat. The lamination of the dough cre-
ates layers that are separated during baking, leading to a flaky structure; (iii) hard sweet
biscuits, formulations low in fat and sugar for allowing the hydrated flour proteins to be
worked into extensible doughs that with reducing chemicals does not need lamination.
The large amount of moisture lost during baking leads to great losses of volatile com-
pounds; thus, these biscuits are sprayed with flavored oil after they come out of the oven.
The most common recipe for Marie biscuits consists of 19% sugar, 13% fat, 2.0% malt
extract, 1.7% skimmed milk powder, 1.0% salt, 0.4% sodium bicarbonate, 1.5% ammo-
nium bicarbonate, 0.26% lecithin, 0.03% sodium metabisulfite, and 24% water (on a
percentage of 100% flour basis); (iv) short dough biscuits, which are distinguished by the
lack of extensibility. They present relatively high levels of sugar and fat (usually butter).
Two-stage mixing methods are normally employed, without fermentation, and they are
sheeted, gauged, and cut. A special way of piecing this dough is the use of extrusion and
co-extrusion; (v) soft dough biscuits, considered a fancy or luxury type for the expensive
ingredients. Butter at 17°C, fine sugar, eggs, milk, water, ground almonds, coconut flour,
and cocoa are widely employed. The mixing is short and gentle, and the pieces are formed
by extrusion; (vi) wafer biscuits, characterized for being produced from a very fluid bat-
ter which is baked between heavy hot plates to produce thin sheets. They are commonly
 Bakery Products 421

sandwiched with cream or caramel before cutting, and the recipe is rich in sugar, fat, and
egg allowing forming and rolling immediately after baking and before cooling. The typi-
cal recipe is 3.5% sugar, 2.7% fat or oil, 3.1% skimmed milk powder, 0.33% dried egg
powder, 0.18% salt, 0.29% soda, 0.83% ammonium bicarbonate, 0.05% fluid lecithin,
and 145% water (on a percentage of flour basis); (vii) Danish pastries, which have the
distinguished feature of the interleaving of dough sheets with layers of fat and the leav-
ening with yeast for 20–30 minutes at a temperature below room temperature to give a
porous structure to the crumb. For separating the dough layers until the product goes to
the oven, Danish pastries are rolled while cold. The common formula is 50% hard wheat
flour, 50% soft wheat flour, 1.4% yeast, 10% sugar, 25% cold milk, and 12% whole egg
(on a percentage of 100% flour basis).

21.3 THE AROMA OF BAKED GOODS FROM

A BIOLOGICAL POINT OF VIEW

The aroma of the crumb or the most internal part of the baking product is mainly gener-
ated by fermentation, including the glycolysis of pyruvic acid and the Ehrlich pathway,
lipid oxidation, as well as the reactions in the yeast cells catalyzed by acetyltransferases
for esters. The aroma of the most external part of the baked product, usually called the
crust, is mostly generated through Maillard reactions, including the Strecker degrada-
tion, caramelization, or thermal degradation of sugars and amino acids (Pico, Bernal,
and Gómez, 2015). In baked products with very low moisture content, such as toasted
bread or Marie biscuits, the main reactions are also Maillard reactions, Strecker degrada-
tion, caramelization, or thermal degradation of sugars (Rychlik and Grosch, 1996). The
ingredients of the baked product contain the precursors and enzymes that participate
in these reactions: flours are rich in starch, which is hydrolyzed mainly by α-amylases
and α-amyloglucosidases into glucose and other small reducing sugars (Martínez-Anaya,
1996). Yeast consumes a great part of the glucose during fermentation, but as a reducing
sugar, it can also participate in Maillard reactions, caramelization, and thermal degra-
dation reactions, just as the other reducing sugars. Sucrose, commonly added to cakes,
biscuits, and gluten-free bread, cannot participate in Maillard reactions, but it can be
hydrolyzed by the invertase of the yeast cells into glucose and fructose (Matz, 1960),
being both able to participate in Maillard reactions, caramelization, and thermal degra-
dation as well as glucose in fermentation; milk and to a lesser extent eggs also contain
sugars that take part in all these reactions. On the other hand, flours, but above all eggs
and milk added to cakes and biscuits, also enclose amino acids, necessary for the Maillard
reaction, Strecker degradation, Ehrlich pathway, and the thermal degradation of amino
acids (Pico, Bernal, and Gómez, 2015). Likewise, proteases can hydrolyze proteins into
amino acids that will participate in these reactions. Finally, the butter or oil added to the
recipe enable the generation of volatile compounds from lipid oxidation. Lipoxygenases
play a key role in the oxidation of lipids containing cis, cis-1,4-pentadiene groups, which
usually come from the action of lipases that hydrolyze triacylglycerides into unsaturated
fatty acids (Guinet and Godon, 1966).
Undoubtedly, the conditions of the processing steps, such as mixing speed, fermen-
tation time and temperature, as well as baking time and temperature, will influence the
extent of all the described reactions. However, although hundreds of compounds are gen-
erated in baked goods production, only a small proportion contributes to the final aroma,
those whose concentration is higher than their odor threshold (OT) (Cho and Peterson,
422 Food Aroma Evolution

2010). The impact of these volatile compounds can be positive, such as 3-methyl-1-buta-
nol, 2-methyl-1-butanol, phenylethyl alcohol, 2,3-butanedione, acetoin, hexyl acetate, or
2-acetyl-1-pyrroline, while other volatile compounds have a negative impact when their
concentration increases, like 1-octen-3-ol, 2,4-(E,E)-decadienal, benzaldehyde, hexanal,
methional, or butyric acid (Pico, Bernal, and Gómez, 2015).

21.3.1 Fermentation Process

Important volatile compounds are generated in bread, brioche, crackers, and Danish pas-
tries through fermentation reactions. The most common volatile compounds found in
fermented baked products are shown in Table 21.2.
During the fermentation process, 95% of the sugars are fermented by Saccharomyces
cerevisiae into ethanol and carbon dioxide. Only the remaining 5% participate in sec-
ondary reactions, which are the glycolisis of pyruvic acid (Czerny and Schieberle, 2002)
and the Ehrlich pathway (Birch et al., 2013). A huge number of volatile compounds are
produced from both reactions (Figure 21.2), mainly short-chain alcohols, esters, and
carbonyl compounds during the glycolysis of pyruvic acid (Drapon and Richard-Molar,
1979) and higher-molecular weight alcohols, aldehydes, and esters from the Ehrlich

TABLE 21.2 Representative Volatile Compounds Found in Baked Products

from Fermentation Processes
Odor Thresholdb,c Organoleptic
Volatile Compound a Origin (µg Kg–1) Characteristicsb,c,d,e
3-Methyl-1-butanol Ehrlich pathway 250 Balsamic, alcohol
2,3-Butanedione Fermentation 6.5 Buttery
3-Methylbutanal Ehrlich pathway 0.2 Malty
Acetic acid Fermentation 32,300 Vinegar-like
Ethanol Fermentation 100,000 Alcohol
2-Methyl-1-propanol Ehrlich pathway 3200 Wine, malty
Benzaldehyde Fermentation 350 Bitter almond
Ethyl hexanoate Glycolisis 1 Fruity, juicy
1-Propanol Glycolisis 6600 Fruity, plastic
Furfural Fermentation 3000 Woody, almond
Phenylacetaldehyde Ehrlich pathway 4 Honey-like
2-Phenylethanol Ehrlich pathway 1100 Rose-like
3-Methylbutanoic acid Fermentation 120 Rancid, sweaty
2-Methyl-1-butanol Ehrlich pathway 40,000 Sweet
Acetoin Fermentation 800 Buttery
Butyric acid Fermentation 240 Rancid, sweaty
Ethyl octanoate Fermentation 92 Fruity, floral
2-Pentylfuran Fermentation 6 Floral, fruity
a Pico, Bernal, and Gómez (2015).
b https​://pu​bchem​.ncbi​.nlm.​nih.g​ov/co​mpoun​d/.
c Birch, Petersen, and Hansen (2013).
d http://www.pherobase.com.
e http://www.thegoodscentscompany.com.
 Bakery Products 423

FIGURE 21.2 Volatile compounds generated during fermentation by the glycolysis of

pyruvic acid (A) and by the Ehrlich pathway of phenylalanine (B). (From Joana Pico,
Doctoral Thesis, 2018, http:​//uva​doc.u​va.es​/ hand​le/10​324/2​9499.)

pathway (Birch et al., 2013). The Ehrlich pathway implies the transamination, decarbox-
ylation and reduction/oxidation of amino acids inside the yeast cell, leading to the gen-
eration of 3-methylbutanal, 3-methyl-1-butanol, and 3-methylbutanoate (from leucine);
2-methylpropanal, 2-methylpropanol, and 2-methylpropanoate (from valine); 2-methyl-
butanal, 2-methyl-1-butanol, and 2-methylbutanoate (from isoleucine); 2-phenylethanal,
2-phenylethanol, and 2-phenylethanoate (from phenylalanine); and methional, methi-
onol, and 3-(methylthio)-propanoate from methionine (Hazelwood et al., 2008).
Lactic acid bacteria (LAB), endogenous in flour but also added to sourdough for
its acidification, are also able to ferment sugars and generate volatile compounds.
Lactobacillus and Kluyveromyces are the most commonly employed for the elaboration
of sourdough bread (Ur-Rehman, Paterson, and Piggott, 2006). The volatile composition
in the crumb of bread with and without sourdough is similar, but the amount of volatile
compounds, especially alcohols like 3-methyl-1-butanol, phenylethyl alcohol, or benzyl
alcohol, is much higher (Hansen and Schieberle, 2005). Indeed, the coexistence of yeasts
and LAB implies a higher number of volatile compounds than when only LAB have been
added (Hansen and Hansen, 1994).

21.3.2 Lipid Oxidation Process

This reaction plays an important role in baked products with a high content of fat, both
butter and oil, such as cakes, soft dough biscuits, short dough biscuits, Danish pas-
tries, or gluten-free bread. Aldehydes, ketones, alcohols, and esters are generated through
the action of lipoxygenases, commonly giving an off-flavor if they are present in high
concentrations (Martínez-Anaya, 1996). As it is reflected in Figure 21.3, lipoxygenases
424 Food Aroma Evolution

FIGURE 21.3 Volatile compounds generated through the lipid oxidation of linoleic acid.
(From Joana Pico, Doctoral Thesis, 2018, http:​//uva​doc.u​va.es​/ hand​le/10​324/2​9499.)​

use oxygen to produce free radicals from cis, cis-1,4-unsaturated fatty acids, leading to
hydroperoxides that are unstable and degrade during baking into a great number of vola-
tile compounds, mainly hexanal, hexenal, and pentanol (Pico, Bernal, and Gómez, 2015).
Other volatile compounds, like 2,4-(E,E)-decadienal, nonanal, 2-(E)-nonenal,
octanal, heptanal, 1-hexanol, 1-octen-3-one, or 2-pentylfuran are produced by subse-
quent reactions (Birch, Petersen, and Hansen, 2014). The most common volatile com-
pounds resulting from lipid oxidation are shown in Table 21.3.
The concentration of volatile compounds from lipid oxidation could be considered
a balance between the amount of lipids present in the dough from the flour, butter, and
other ingredients, and the lipoxygenase activity. However, the level of antioxidants, like
flavonoids and vitamin E, is also really important for avoiding the action of lipoxygen-
ases (Pico, Hansen, and Petersen, 2017).

21.3.3 Maillard Reaction and Caramelization

The Maillard reaction can be considered one of the most important reactions for aroma
generation, since it takes place during baking for all the described baked products. The
Maillard reaction entails a complex non-enzymatic process where an amino acid and a
reducing sugar, such as glucose and fructose, react at temperatures between 110°C and
150°C, leading to the formation of brown pigments (melanoidines) and many volatile
compounds (Figure 21.4). Furans, pyrazines, pyrroles, pyrrolines, oxazoles, thiophenes,
thiopyranes, thiazolines, and sulfuric compounds (Table 21.4) are the main compounds
generated in the crust or the most external surface of baked products during Maillard
reactions (Pico, 2018). The Strecker degradation is a special stage of the Maillard reac-
tion where aldehydes, like acetaldehyde, formaldehyde, or glyoxal, are generated as a
 Bakery Products 425

TABLE 21.3 Representative Volatile Compounds Found in Baked

Products from Lipid Oxidation
Odor Thresholdb,c
Volatile Compound a (µg Kg–1) Organoleptic Characteristicsb,c,d,e
Hexanal 4.5 Green grass
2-(E)-Nonenal 0.08 Green, tallow
1-Hexanol 2500 Sweet alcohol
Benzaldehyde 350 Bitter almond
1-Pentanol 4000 Fusel-like
2,4-(E,E)-Decadienal 0.1 Fatty, deep-fried
Nonanal 1 Waxy, green, fatty
Hexanoic acid 3000 Fatty
Heptanal 3 Fatty, pungent
1-Octen-3-one 0.01 Mushroom
2-Pentylfuran 6 Floral, fruity
2-Ethyl-1-hexanol 138 Sweet, floral
Benzyl alcohol 10,000 Fruity, balsamic
2-Octenal 3 Fatty, nutty
1-Octen-3-ol 1 Mushroom
Octanal 0.7 Citrus
2,4-(E,E)- 0.06 Fatty, waxy
nonadienal
a Pico, Bernal, and Gómez (2015).
b https​://pu​bchem​.ncbi​.nlm.​nih.g​ov/co​mpoun​d/.
c Birch, Petersen, and Hansen (2013).
d http://www.pherobase.com.
e http://www.thegoodscentscompany.com.

consequence of the reaction between an amino acid and a dehydroreductone (Guinet and
Godon, 1996). Some aldehydes can be generated both from the Strecker reaction or the
Ehrlich pathway (i.e., 2-methylpropanal, 3-methylbutanal, 2-methylbutanal, phenylacet-
aldehyde, and methional) (Rooney, Salem, and Johnson, 1967; Martínez-Anaya, 1996).
Indeed, the baking step is responsible for the final aroma of the baked product, inas-
much as volatile compounds generated in the crumb during fermentation can be volatil-
ized or transferred to the crust (Onishi et al., 2011), some hydroperoxides generated
during lipid oxidation are broken into volatile compounds (Pico, Bernal, and Gómez,
2015) and the key Maillard reaction takes place, together with caramelization and the
thermal degradation of amino acids and sugars.
The rate of the Maillard reaction depends on the type of sugar, xylose being the most
reactive, while the aroma profile relies on the type of amino acid (Kiely, Nowlin, and
Moriarty, 1960). 2-acetyl-1-pyrroline has been considered the key aroma compound of
the Maillard reaction, due to its high odor activity value (OAV) and its pleasant roasty
notes (Zehentbauer and Grosch, 1998a). There are also compounds that can be gener-
ated both by the Maillard reaction and other sources, like 2-methylbutanal, 3-methyl-
butanal, and methional (also generated by the Ehrlich pathway), 3-methylbutyric acid,
2,3-butanedione (also from fermentation), 4-hydroxy-2,5-dimethyl-3(2H)-furanone,
and 2-methyl-propanal. Furan is generated by the Maillard reaction but also by lipid
426 Food Aroma Evolution

FIGURE 21.4 Volatile compounds produced by means of Maillard reactions. (From

Doctoral Thesis Joana Pico, 2018, http:​//uva​doc.u​va.es​/ hand​le/10​324/2​9499.)

oxidation and thermal degradation of ascorbic acid, while furfural, 5-hydroxymethylfur-

fural, 5-methyl-2-furaldehyde, and furfuryl alcohol can be generated by Maillard reac-
tions and caramelization (Rannou et al., 2016; Ait Ameur et al., 2008). Caramelization
is a thermal reaction that takes place at temperatures higher than 150°C, when the sugars
are heating above their melting point (Hadiyanto et al., 2007).

21.4 EVOLUTION OF VOLATILE COMPOUNDS

DURING PROCESSING AND STORAGE

The generation of volatile compounds in baked products mainly depends on the ingre-
dients and processing steps. Flour is the main ingredient in baked products, but it is
well known that its contribution to the final flavor is very limited (Taylor, Guichard,
and Cayot, 2007). Pico (2018) compared the volatile compounds analyzed in gluten-free
bread elaborated with quinoa flour, teff flour, and corn starch with the volatile com-
pounds of the corresponding flour or starch and found that the volatile profiles were
completely different (Pico et al., 2017; Pico, Hansen, and Petersen, 2017). Most of the
volatile compounds that were relevant in quinoa and corn starch crumb were important
in teff flour and vice versa (Pico, 2018), suggesting that the main purpose of the flours is
providing precursors for the reactions that generate the volatile compounds throughout
 Bakery Products 427

TABLE 21.4 Representative Volatile Compounds Found in Baked Products from

Maillard Reactions
Odor Thresholdb,c Organoleptic
Volatile Compound a Origin (µg kg–1) Characteristicsb,c,d,e
2,3-Butanedione Maillard 6.5 Buttery
3-Methylbutanal Strecker degradation 0.2 Malty
Acetic acid Maillard 32,300 Vinegar-like
Benzaldehyde Maillard 350 Bitter almond
Furfural Maillard 3000 Woody, almond
Phenyacetaldehyde Strecker degradation 4 Honey-like
Acetoin Maillard 800 Buttery
2-Acetyl-1-pyrroline Maillard 0.053 Roasted, popcorn
1-Methylpyrrol Maillard 37 Toasted
Furfuryl alcohol Maillard 8 Coffee
2-Pentylfuran Maillard 6 Floral, fruity
Benzyl alcohol Maillard 10,000 Fruity, balsamic
Furan Maillard 10,060 Ethereal
5-Hydroxymethylfurfural Maillard nf Herbal
2,5-Dimethylpyrazine Maillard 800 Chocolate, earthy
2-Ethylpyrazine Maillard 6000 Musty, nutty, peanut
2-Ethyl-3-methylpyrazine Maillard 0.4 Potato, burned nutty
a Pico, Bernal, and Gómez (2015).
b https​://pu​bchem​.ncbi​.nlm.​nih.g​ov/co​mpoun​d/.
c Birch, Petersen, and Hansen (2013).
d http://www.pherobase.com.
e http://www.thegoodscentscompany.com.

nf: Not found.

the processing steps. Indeed, there can be volatile compounds that are only present when
certain flours are employed. For example, the addition of orange-fleshed potato flour
to the traditional sourdough of panettone, made of wheat flour, led to the generation
of volatile compounds like 2-octenal-2-butyl, dimethyl-decane, or 2-chloro-octane that
were only found in panettones made with orange-fleshed potato flour (Aparecida Pereira
et al., 2019). As explained before, there are several stages in which these compounds
can be produced: through enzymatic activity and non-enzymatic lipid oxidation during
kneading, through dough fermentation by yeast (including Ehrlich pathway and glycoly-
sis of pyruvic acid) and lactic bacteria, and through lipid oxidation reactions and thermal
reactions taking place during baking mainly through the Maillard and caramelization
reactions. There is a great number of studies explaining how the ingredients and condi-
tions affect the final aroma of each step, but the evolution of volatile compounds over
time is not so well known.

21.4.1 From the Raw Materials to the Baked Product

In bread, most efforts have been made for understanding the evolution of volatile
compounds during the dough leavening. To this end, Makhoul et al. (2015) employed
428 Food Aroma Evolution

TABLE 21.5 Evolution of Volatile Compounds from Dough (0 Minutes, 45 Minutes, and 90
Minutes of Fermentation) to Crumb (40 Minutes Baking) in Wheat Bread
Behavior from Dough to
Crumb Volatile Compounds
Evaporation from dough to 2,3-Butanedione, 1-propanol, 2-methyl-1-propanol, 3-methyl-
crumb 1-butanol, 2-methyl-1-butanol, Ethyl octanoate,
Continuous increase from Ethyl lactate, Acetoin, Acetic acid, 2,3-butanediol, nonanal,
dough to crumb 2-(e)-nonenal, Phenylethyl alcohol
Increase from 45 to 90 minutes 1-Hexanol, Isobutyric acid, Butyric acid, 3-methylbutyric acid,
fermentation and during 2-methylbutyric acid, 2,4-decadienal,4-vinylguaiacol
baking
Increase from 0 to 45 minutes Methional, 1-Octen-3-ol, Butyrolactone
fermentation and during
baking
Increase from 0 to 45 minutes Phenylacetaldehyde
fermentation and then steady
Steady during fermentation, Hexanal, Hexanoic acid, Benzaldehyde, Benzyl alcohol,
increase during baking Furfuryl alcohol, Furfural
Source: Pico, Martinez, Bernal, and Gomez (2017).

PTR-MS (proton transfer reaction–mass spectrometry) to study the interaction between

the flour and yeast in bread dough. They found that some selected mass peaks continu-
ously increased over the fermentation time, while other mass peaks experienced a great
increase in the first stage of fermentation and then a progressive decrease. However, due
to the limitations of the technique, the mass peaks were only tentatively identified.
Pico et al. (2017a) studied the complete evolution from the beginning of fermentation
to the end of baking in gluten-free bread using GC-MS (gas chromatography–MS), as is
summarized in Table 21.5.
As expected, the dough was characterized by volatile compounds from the fermen-
tation and small amounts of volatile compounds from non-enzymatic lipid oxidation.
1-propanol, 2-methyl-1-propanol, 3-methyl-1-butanol, and 2-methyl-1-butanol, and
ethyl lactate increased along the entire fermentation process as well as acetoin, acetic
acid, 2,3-butanediol, phenylethyl alcohol (also generated by Maillard reactions), and
nonanal and 2-(E)-nonenal (also generated by lipid oxidation). 2,4-(E,E)-decadienal,
1-hexanol, 3/2-methylbutyric acid, butyric acid, 2,3-butanedione, and acetoin were
more intensively generated at the end of fermentation, between 45 and 90 minutes.
Methional and phenylacetaldehyde increased up to 45 minutes of fermentation and
then kept steady due to the depletion of the amino acids that originate them, methi-
onine and phenylalanine, respectively. The redox couples hexanal–hexanoic acid,
furfuryl alcohol–furfural as well as benzyl alcohol–benzaldehyde did not suffer sig-
nificant changes during fermentation. When the dough was baked, it was found that
2,3-butanedione and ethyl octanoate, as well as the alcohols 1-propanol, 2-methyl-
1-propanol, 3-methyl-1-butanol, and 2-methyl-1-butanol were partially evaporated due
to their high volatility, while 2-pentylfuran was exclusively produced through baking.
The rest of the studied volatile compounds increased during baking, as they could also
be generated by the Maillard reaction (e.g., acetoin, acetic acid, 3-methylbutyric acid),
caramelization (e.g., furfural), and enzymatic lipid oxidation (e.g., nonanal, 2,4-(E,E)-
decadienal, hexanal). The final aroma of bread is, therefore, a consequence of all these
 Bakery Products 429

processes of generation of volatile compounds during the whole bread-making and pos-
sible evaporation during baking.
The examination of the aroma evolution in cakes and cookies has been focused on
the baking step, mainly on the study of furanic compounds and volatiles from lipid oxi-
dation. The few studies found in the literature analyzing the complete volatile profile
of cakes and cookies during baking have employed online SPME-GC/MS (solid-phase
microextraction-GC/MS). Rega et al. (2009) observed that during the baking of a sponge
cake, there was a slight increase in the concentration in the vapor of volatile compounds
from lipid oxidation, such as hexanal, nonanal, or 2,4-(E,E)-decadienal; indeed, aliphatic
aldehydes and 1-octanol were already present in relatively high amounts at the first 5 min-
utes of baking. Regarding Maillard compounds, including increasing Strecker aldehydes,
pyrazines, furanic compounds, and enolization markers, there was a general linear trend
of them being completely absent in vapors at the beginning of baking and then gradu-
ally increasing. Only 5-hydroxymethylfurfural (HMF) was formed at the beginning and
released mainly at the end of the baking process, following an exponential curve.
Using the same technique, Ait Ameur et al. (2008) studied the main volatile com-
pounds dynamically extracted from the vapors of cookies baking. They noticed that
furanic compounds, concretely HMF, furfural, and 2-methylfurfural, exponentially
increased after 4 minutes of baking, with a content 20 and 600 times higher than ketones
and aldehydes, respectively. This increase of furanic compounds in the vapor phase
implied a decrease of HMF and furfural and a small increase of 2-methylfurfural in the
baked matrix at the end of baking. Ketones, mainly 1-hydroxy-2-propanone and 2,3-d​
ihydr​o-3,5​-dihy​droxy​- 6-me​thyl-​4(H)-​pyran​- 4-on​e, followed the same tendency than
furanic compounds, slowly accumulating during the first 4 minutes and then a great
increase at the end of baking. The aliphatic aldehydes from lipid oxidation, nonanal, and
2,4-(E,E)-decadienal, presented an early formation during the first minutes and then
decayed in concentration at 6–8 minutes of baking to the end.
Volatile compounds from lipid oxidation have also been frequently analyzed in cakes
and biscuits as a result of their rancid-unpleasant notes, enhanced by the addition of oil
and/or shortenings to the recipe; it is well known that the polyunsaturated fatty acids
from the lipid fraction are oxidized by lipoxygenases during kneading, requiring oxygen
and low activation energy (Maire et al., 2013), and subsequent degradation of the hydro-
peroxides to these volatile compounds during baking (Caponio et al., 2008). Maire et al.
(2013) found that the amount of hexanal in the dough was higher than those globally
found in the employed ingredients, demonstrating that lipid oxidation occurs really early
in the dough preparation and in a minor extent during baking.
Due to health concerns, the analysis of HMF and furan, as well as acrylamide, has
been commonly extended to the aroma evolution of cakes and biscuits during baking.
The main reason is the complex recipe of both baked products, which includes ingredi-
ents rich in reducing sugars (sweeteners) and amino acids (eggs and milk). Acrylamide is
produced by the Maillard reaction from the reaction of asparagine and reducing sugars,
mainly fructose (Nguyen, Fels-klerx, and Boekel, 2017), and it is potentially carcinogenic
and neurotoxic. HMF is a degradation product of an Amadori rearrangement of the
Maillard reaction between asparagine and reducing sugars, and it is also potentially car-
cinogenic. In biscuits, it can also be generated by caramelization due to the high content
of sucrose (Ait Ameur et al., 2008). Furan, for its part, can be generated by carameliza-
tion, Maillard reaction, and, less favored, by lipid oxidation. Its generation is enhanced
in the presence of glucose (Srivastava et al., 2017) and egg yolk (Cepeda-Vazquez et al.,
2017), ingredients typically employed in cakes. Nguyen, Fels-Klerx, and Boekel (2017)
430 Food Aroma Evolution

studied the changes in acrylamide ad HMF concentration in different biscuits during

baking. They found that the isomerization of glucose to fructose leads to an increase in
the concentration of acrylamide and the caramelization of glucose and fructose to an
increase of HMF, but the formation of both compounds is only visible after 10 minutes of
baking. Petisca et al. (2013) found that the increase in the baking time led to an increase
in the concentration of HMF in traditional ovens and microwaves, but the concentra-
tion kept steady when using steam ovens. Although the baking of biscuits can vary from
2 to 15 minutes, the most common baking time is 6 minutes (Cauvain and BakeTran,
2016); thus, the concentration of HMF or acrylamide should not be a risky issue. As
a matter of fact, acrylamide was not detected by Jouquand et al. (2018) in traditional
French baguettes whatever the baking time (from 12 to 22 minutes), although the limit of
detection was relatively high, 18 μg/kg. The French baguette-making process, with long
enough dough fermentation, seems to favor the consumption of free asparagine by the
yeast, inducing to the lower amount of acrylamide in the crust.
Finally, losses of exogenous flavors added to sweet baked products during manufac-
turing have also been examined. Flavoring is a practice widely used for cakes and pas-
tries, and it implies the addition of natural flavors like vanilla, essential oils like lemon
oil, or artificial flavors like diacetyl or vanillin (Hazelton, Desrochers, and Walker, 2003;
Taylor, Guichard, and Cayot, 2007). Although flavoring is used to generate a desired
aromatic note, some added volatile compounds could be lost or modified during the elab-
oration process. It could be a consequence of evaporation or retention of the volatile com-
pounds by the matrix, reducing the availability of the odor during consumption (Taylor,
Guichard, and Cayot, 2007). Pozo-Bayon, Guichard, and Cayot (2006) quantified the
aroma compounds added to the dough and retained in sponge cakes after processing
and found that their concentration ranged between 29% (losses during elaboration) and
170% (generation during elaboration). They also concluded that the absence of fatty
material dramatically decreases the retention of the added aroma compounds.

21.4.2 Toasting of Bread

The toasting of bread occupies a considerable segment of the food industry since it allows
easier digestion and increases flavor and edibility compared with fresh bread, especially
after staling starts (Kirit, Erdogdu, and Ozdemir, 2013). The chemical reactions that
take place during toasting are mainly Maillard reactions (above 50°C) and carameliza-
tion (above 120°C), and the important volatile compounds found in toasted bread are
2-acetyl-1-pyrroline, 4-hydroxy-2,5-dimethyl-3(2H)-furanone, 3-methylbutyric acid,
2,3-butanedione, and 2-(E)-nonenal (Rychlik and Grosch, 1996; Kirit, Erdogdu, and
Ozdemir, 2013). Rychlik and Grosch (1996) reported that 2-acetyl-1-pyrroline quickly
increased at the beginning of toasting, while 4-hydroxy-2,5-dimethyl-3(2H)-furanone
was mainly produced at medium browning. The accumulation of HMF in toasted bread
has also been considered and it was reported that the accumulation and distribution
of this compound depended on the temperature evolution and heat transfer mechanism
in the slice: the crumb section was free of HMF until 118 seconds of toasting, and in
the crust it started an exponential increase, while the accumulation in the crumb sec-
tion becomes higher than in the crust section after 215 seconds (Kirit, Erdogdu, and
Ozdemir, 2013). Pico et al. (2018) reported an increase in all volatile compounds found in
toasted gluten-free breadcrumbs and crusts regarding the corresponding baked samples.
This was attributed to the generation of volatile compounds from the Maillard reaction,
 Bakery Products 431

caramelization, and lipid oxidation or to the transference from the crumb to the crust of
volatile compounds already generated from lipid oxidation and fermentation, leading to
easier extraction during analysis (Pico et al., 2018).
They also concluded that the degree of enhancement of volatile compounds was dif-
ferent among the diverse toasted gluten-free bread, including quinoa, teff and rice flours,
and corn and wheat starches; in the case of Maillard reactions, it was explained by the
different kinetics and depletion of precursors of each bread.

21.4.3 Changes in the Aroma Quality during Storage

Baking products have short shelf-lives, leading to considerable economic losses annually.
The main cause is the “staling,” a complex physicochemical process that leads to degra-
dation of flavor and overall bread quality, followed by mold spoilage (Plessas et al., 2011).
The examination of aroma evolution of baked products during storage at room tempera-
ture has been focused mainly on the flavor of the bread, and it has been referred to as a
loss of pleasant volatile compounds along with the generation of off-flavors through lipid
oxidation (Latou et al., 2010). The improvement of bread shelf life in terms of aroma has
been achieved using special packaging (Latou et al., 2010) or by the addition of sour-
dough with different starter cultures (Plessas et al., 2011, 2008).
Along general lines, there is a global decrease of volatile compounds during short
storage periods, which is higher in the crust than in the crumb. Chiavaro et al. (2008)
reported a decrease in the volatile compounds of bread stored for 8 days, with a reduction
between 2–5 times in the crusts of artisanal breads, 3–10 times in the crust of industrial
breads, and 1.5–3 times in the crumb of both artisanal and industrial breads. This general
decrease over the first week of storage was in concordance with the study of Pico et al.
(2017b), and it was attributed to evaporation from bread surface, oxidative reactions, as
well as diffusion from the crust to the center of the crumb and the subsequent entrapment
by proteins and starch (Schieberle and Grosch, 1992). Only acetoin has been reported to
increase during the first week of storage (Pico et al., 2017b; Jensen et al., 2011), prob-
ably due to the decarboxylation of 2-acetolactate (Birch, Petersen, and Hansen, 2013).
Ethanol, due to its high volatility, has been commonly reported as decreasing over the
time (Pico et al., 2017b; Jensen et al., 2011; Plessas et al., 2011, 2008), although Latou
et al. (2010) observed an increase. However, when the aroma evolution during storage
is examined for longer periods of time, there are changes that can lead to increases or
decreases of certain volatile compounds (Pico et al., 2017b; Jensen et al., 2011). Jensen
et al. (2011) concluded that after 1 week of storage bread was characterized by volatile
compounds from fermentation and Maillard reactions, while after 2–3 weeks of storage
the volatile compounds present in higher concentration were those of lipid oxidation. The
general tendencies observed by Jensen et al. (2011) were similarly detected by Pico et al.
(2017b) during the study of frozen storage, but with a delay usually due to the mitigation
of reactions at freezing temperatures. During the first week, there was an overall decrease
of the volatile compounds, with preservation of 34% regarding room temperature. As
depicted in Figure 21.5, the total content of alcohols, acids, and ketones followed the
same general tendency during freezing, with the decrease in the total area during the
first week, but an increase during the second week and a final decrease leading up to the
fourth week.
In both studies, the alcohols showed exactly the same behavior over 2 weeks, with
a decrease in the first week and an increase in the second week up to the initial value of
432 Food Aroma Evolution

FIGURE 21.5 Evolution of the main groups of volatile compounds in the wheat crumb
stored for 1 week at room temperature (black lines) and in the wheat crumb frozen for 1,
2, and 4 weeks (gray lines). The results are the sum of the peak areas of the ketones (con-
tinuous line, 106), aldehydes (discontinuous line, 106), alcohols (scratch doubly spotted
line, 107), and acids (spotted line, 107). (From Pico, Martínez, Bernal, and Gómez 2017.)

the fresh sample. Regarding the acids, Jensen et al. (2011) reported an increase during
the first week and a decrease during the second week, while Pico et al. (2017b) results
showed an increase during the second week and then a decrease again leading up to the
fourth week. It was explained through the oxidation of the aldehydes, a reaction that can
be delayed during freezing. Ketones exhibited the same general tendency of increasing at
the beginning and then decreasing. Only the behavior of aldehydes was different in both
studies: Jensen et al. (2011) reported the continuous increase of aldehydes as a conse-
quence of lipid oxidation, a reaction that seems to be inhibited during freezing due to the
continuous decreasing of the concentration of aldehydes in Pico et al.’s (2017b) research.
In the case of cakes and biscuits, the loss of quality during storage has been attrib-
uted to the increase of off-flavors from lipid oxidation (Gebreselassie and Hall, 2016),
since oil and shortenings are frequently added. Yang et al. (2013) observed a continuous
increase in the rancid markers 2,4-decadienal and 2,4-heptadienal in biscuits stored for
8 weeks, along with a continuous decrease of HMF. Cognat et al. (2014) also observed a
decrease in the absolute content of hexanal and 1-pentene in oat biscuits stored over 28
weeks. Finally, the same increase in the aldehydes content was reported by Sakač et al.
(2016) in gluten-free cookies stored for 16 weeks at room temperature, increase that was
delayed in packed cookies regarding unpacked ones.

21.5 GENERAL CONCLUSIONS

The evolution of volatile compounds during baked goods production has been demon-
strated as dependent on the ingredients and processing steps. The aroma precursors pres-
ent in flour, as well as the yeast, determine the volatile compounds from fermentation
 Bakery Products 433

and lipid oxidation that are generated in the crumb of the bread, and which can be
partially lost during baking or can increase if they can be generated not only by fer-
mentation but also by Maillard reactions in the crust. In the case of cakes and cookies,
Maillard reactions, caramelization, and lipid oxidation have been described as the main
processes for the generation of volatile compounds due to the source of reducing sug-
ars and amino acids found in the added sweeteners and eggs, as well as the fatty acids
detected in shortenings. Due to the high temperatures applied during baking, the evolu-
tion of 5- HMF and furan during baking have been reported to be of utmost importance
as a consequence of their carcinogenic characteristics. Finally, the storage of bread has
been indicated as one of the main concerns in the aroma quality of bread due to its short
shelf life, with general losses of pleasant volatile compounds. In the case of cakes and
cookies, the use of shortenings leads to the generation of off-flavors from lipid oxidation
during storage.

REFERENCES

Agrifood Canada. Website: http:​//www​5.agr​.gc.c​a /eng ​/indu​stry-​marke​ts-an​d-tra​de/in​

terna​t iona​l-agr​i-foo​d-mar​ket-i​ntell​igenc​e /sec​tor-t​rend-​a naly​sis-b​a ked-​goods​-in-t​
he-un​ited-​state​s-and​- cana​da/?i​d=153​73812​72039​. Last accessed January 20, 2019.
Ait Ameur, Lamia, Barbara Rega, Pierre Giampaoli, Gilles Trystram, and Inès Birlouez-
Aragon. 2008. “The Fate of Furfurals and Other Volatile Markers during the Baking
Process of a Model Cookie.” Food Chemistry 111 (3): 758–63.
Aparecida Pereira, Ana Paula, María Teresa Pedrosa Silva Clerici, Marcio Schmiele, Luis
Carlos Gioia Júnior, Marina Akemi Nojima, Caroline Joy Steel, Yoon Kil Chang,
Glaucia Maria Pastore, and Elizabeth Harumi Nabeshima. 2019. “Orange-Fleshed
Sweet Potato Flour as a Precursor of Aroma and Color of Sourdough Panettones.”
LWT—Food Science and Technology 101: 145–51.
Arvisenet, Gaelle, Patricia Le Bail, Andree Voilley, and Nathalie Cayot. 2002. “Influence
of Physicochemical Interactions between Amylose and Aroma Compounds on the
Retention of Aroma in Food-like Matrices.” Journal of Agricultural and Food
Chemistry 50 (24): 7088–93.
Belitz, Hans Dieter, Werner Grosch, and Peter Schieberle. 2009. Food Chemistry. 4th ed.
Springer-Verlag, Berlin.
Bennion, Edmund B., James Stewart, and G. S. T. Bamford. 1966. Cake Making. Leonard
Hill Books, London.
Bianchi, Federica, Mariella Careri, Emma Chiavaro, Marilena Musci, and Elena
Vittadini. 2008. “Gas Chromatographic-Mass Spectrometric Characterisation of
the Italian Protected Designation of Origin ‘Altamura’ Bread Volatile Profile.” Food
Chemistry 110: 787–93.
Birch, Anja N., Mikael A. Petersen, and Åse S. Hansen. 2013. “The Aroma Profile
of Wheat Bread Crumb Influenced by Yeast Concentration and Fermentation
Temperature.” LWT—Food Science and Technology 50: 480–8.
Birch, Anja N., Mikael A. Petersen, and Åse S. Hansen. 2014. “Aroma of Wheat Bread
Crumb.” Cereal Chemistry 91 (2): 105–14.
Birch, Anja N., Mikael A. Petersen, Nils Arneborg, and Åse S. Hansen. 2013. “Influence of
Commercial Baker’s Yeasts on Bread Aroma Profiles.” Food Research International
52: 160–6.
434 Food Aroma Evolution

Blanshard, J. M. V., P. J. Frazier, and T. Galliard. 1985. Chemistry and Physics of Baking.
The Royal Society of Chemistry, London.
Caponio, Francesco, Carmine Summo, Antonella Pasqualone, and Maria T. Bilancia.
2008. “Effect of Kneading and Baking on the Degradation of the Lipid Fraction of
Biscuits.” Journal of Cereal Science 48 (2): 407–12.
Cauvain, Stanley P. 1998. Technology of Breadmaking. Springer, Berlin.
Cauvain, Stanley P. 2003. Bread Making. Improving Quality. CRC Press, FL.
Cauvain, Stanley P. and Witney BakeTran. 2016. Cookies, Biscuits and Crackers:
Formulation, Processing and Characteristics. Encyclopedia of Food Grains. 2nd
ed. Elsevier, Amsterdam.
Cepeda-Vazquez, Mayela, Barbara Rega, Nicolas Descharles, and Valerie Camel. 2017.
“How Ingredients Influence Furan and Aroma Generation in Sponge Cake.” Food
Chemistry 245: 1025–33.
Chiavaro, Emma, Elena Vittadini, Marilena Musci, Federica Bianchi, and Elena Curti.
2008. “Shelf-Life Stability of Artisanally and Industrially Produced Durum Wheat
Sourdough Bread (‘Altamura Bread’).” LWT—Food Science and Technology 41 (1):
58–70.
Cho, In Hee and Devin G. Peterson. 2010. “Chemistry of Bread Aroma: A Review.” Food
Science and Biotechnology 19 (3): 575–82.
Cognat, Claudine, Tom Shepherd, Susan R. Verrall, and Derek Stewart. 2014.
“Relationship between Volatile Profile and Sensory Development of an Oat-Based
Biscuit.” Food Chemistry 160: 72–81.
Cserháti, Tibor. 2010. Chromatography of Aroma Compounds and Fragances. Springer,
Berlin.
Czerny, Michael and Peter Schieberle. 2002. “Important Aroma Compounds in Freshly
Ground Wholemeal and White Wheat Flour. Identification and Quantitative
Changes during Sourdough Fermentation.” Journal of Agricultural and Food
Chemistry 50: 6835–40.
Drapon, R. and D. Richard-Molard. 1979. Influence de divers procédés technologiques
sur la formation de l'arôme du pain. Répercussions sur sa qualité. Actes du colloque
CNERNA. Ed. du Centre National de la Recherche Scientifique, Paris.
Gebreselassie, Etsehiwot and Clifford Hall. 2016. Oxidative Stability and Shelf Life of
Crackers, Cookies, and Biscuits. Elsevier Inc.
Guinet, R. and B. Godon. 1996. La Panification Française. Montagud, Barcelona.
Hadiyanto, A. G. Asselman, R. M. Van Straten, D. C. Boom, D. C. Esveld, and A. J. B.
Van Boxtel. 2007. “Quality Prediction of Bakery Products in the Initial Phase of
Process Design.” Innovative Food Science & Emerging Technologies 8: 285–98.
Hansen, Åse ̊ and Peter Schieberle. 2005. “Generation of Aroma Compounds during
Sourdough Fermentation: Applied and Fundamental Aspects.” Trends in Food
Science and Technology 16: 85–94.
Hansen, Birgit and Åse ̊ Hansen. 1994. “Volatile Compounds in Wheat Sourdoughs
Produced by Lactic Acid Bacteria and Sourdough Yeasts.” Zeitschrift Für
Lebensmittel-Untersuchung Und -Forschung 198: 202–9.
Hazelton, J. L., J. L. Desrochers, and C. E. Walker. 2003. Chemistry of Biscuit Making,
pp. 533–9. Elsevier, Amsterdam.
Hazelwood, L. A., J. M. Daran, A. J. van Maris, J. T. Pronk, and J. R. Dickinson.
2008. “The Ehrlich Pathway for Fusel Alcohol Production: A Century of Research
on Saccharomyces cerevisiae Metabolism (Vol 74, Pg 2259, 2008).” Applied and
Environmental Microbiology 74 (8): 3920.
 Bakery Products 435

Jensen, Sidsel, Henrik Oestdal, Leif H. Skibsted, Erik Larsen, and Anette K. Thybo. 2011.
“Chemical Changes in Wheat Pan Bread during Storage and How It Affects the Sensory
Perception of Aroma, Flavour, and Taste.” Journal of Cereal Science 53 (2): 259–68.
Jouquand Céline, Céline Niquet-Léridon, Philippe Jacolot, Noémie Petit, Marier David,
and Pascale Gadonna-Widehem. 2018. “Effects of Maillard Reaction Products on
Sensory and Nutritional Qualities of the Traditional French Baguette.” Journal of
Food Science 83 (10): 2424–31.
Kiely, R. J., A. C. Nowlin, and J. H. Moriarty. 1960. “Bread Aromatics from Browning
Systems.” Cereal Science Today 5: 273–4.
Kirit, Ayse Basak, Ferruh Erdogdu, and Yuksel Ozdemir. 2013. “Accumulation of
5-Hydroxymethyl-2-Furfural during Toasting of White Bread Slices.” Journal of
Food Process Engineering 36: 241–6.
Latou, E., S. F. Mexis, A. V. Badeka, and M. G. Kontominas. 2010. “Shelf Life Extension
of Sliced Wheat Bread Using Either an Ethanol Emitter or an Ethanol Emitter
Combined with an Oxygen Absorber as Alternatives to Chemical Preservatives.”
Journal of Cereal Science 52 (3): 457–65.
Maire, Murielle, Barbara Rega, Marie-Elisabeth Cuvelier, Paola Soto, and Pierre
Giampaoli. 2013. “Lipid Oxidation in Baked Products : Impact of Formula and
Process on the Generation of Volatile Compounds.” Food Chemistry 141: 3510–8.
Makhoul, Salim, Andrea Romano, Vittorio Capozzi, Giuseppe Spano, Eugenio Aprea,
Luca Cappellin, Elisabetta Benozzi, Matteo Scampicchio, Tilmann D. Märk,
Flavia Gasperi, Hanna El-Nakat, Jean Guzzo, and Franco Biasioli. 2015. “Volatile
Compound Production During the Bread-Making Process: Effect of Flour, Yeast
and Their Interaction.” Food and Bioprocess Technology 8 (9): 1925–37.
Manley, D. 2011. Manley’s Technology of Biscuits, Crackers and Cookies. Woodhead
Publishing, Cambridge.
Martínez-Anaya, M. Antonia. 1996. “Enzymes and Bread Flavor.” Journal of Agricultural
and Food Chemistry 44 (9): 2469–80.
Martinez-Monzo, J., P. Garcia-Segovia, and J. Albors-Garrigos. 2014. “Trends and
Innovations in Bread, Bakery and Pastry.” Journal of Culinary Science & Technology
11: 37–41.
Matz, Samuel A. 1960. Bakery. Technology and Engineering. The Avi Publising
Company, CT.
Nguyen, H. T., H. J. Ine Van Der Fels-Klerx, and M. A. J. S. Van Boekel. 2017. “Acrylamide
and 5-Hydroxymethylfurfural Formation during Biscuit Baking. Part II : Effect of
the Ratio of Reducing Sugars and Asparagine.” Food Chemistry 230: 14–23.
Onishi, Masanobu, Michiko Inoue, Tetsuya Araki, Hisakatsu Iwabuchi, and Yasuyuki
Sagara. 2011. “Odorant Transfer Characteristics of White Bread during Baking.”
Bioscience, Biotechnology, and Biochemistry 75 (2): 261–7.
Petisca, Catarina, Ana Rita, Trinidad Pérez-palacios, Olívia Pinho, and Isabel Ferreira.
2013. “Study of Hydroxymethylfurfural and Furfural Formation in Cakes dur-
ing Baking in Different Ovens, Using a Validated Multiple-Stage Extraction-Based
Analytical Method.” Food Chemistry 141: 3349–56.
Pico, J. 2018. Analysis of Volatile Compounds in Bread and Related Products.
Improvement of Gluten-free Breads Aroma. Doctoral Thesis. http://uvadoc.uva.es/
handle/10324/29499. Last accessed January 20, 2019.
Pico, Joana, Åse S. Hansen, and Mikael A. Petersen. 2017. “Comparison of the Volatile
Profiles of the Crumb of Gluten-Free Breads by DHE-GC/MS.” Journal of Cereal
Science 76: 280–8.
436 Food Aroma Evolution

Pico, Joana, Jesus Tapia, José Bernal, and Manuel Gómez. 2017. “Comparison of Different
Extraction Methodologies for the Analysis of Volatile Compounds in Gluten-Free
Flours and Corn Starch by GC/QTOF.” Food Chemistry 54 (6): 1433–41.
Pico, Joana, José Bernal, Iuliia Khomenko, Manuel Gómez, Vittorio Capozzi, and
Luciano Navarini. 2018. “Analysis of Volatile Organic Compounds in Crumb and
Crust of Different Baked and Toasted Gluten-Free Breads by Direct PTR–ToF–MS
and Fast—GC–PTR–ToF–MS.” Journal of Mass Spectrometry 53 (9): 893–902.
Pico, Joana, José Bernal, and Manuel Gómez. 2015. “Wheat Bread Aroma Compounds
in Crumb and Crust : A Review.” Food Research International 75: 200–15.
Pico, Joana, Mario M. Martínez, José Bernal, and Manuel Gómez. 2017a. “Evolution
of Volatile Compounds in Gluten-Free Bread: From Dough to Crumb.” Food
Chemistry 227: 179–86.
Pico, Joana, Mario M. Martínez, José Bernal, and Manuel Gómez. 2017b. “Impact
of Frozen Storage Time on the Volatile Profile of Wheat Bread Crumb.” Food
Chemistry 232: 185–90.
Picó, Yolanda. 2012. Chemical Analysis of Food: Techniques and Applications. Elsevier,
Amsterdam.
Plessas, Stavros, Argyro Bekatorou, J. Gallanagh, Poonam Nigam, Athanasios A.
Koutinas, and C. Psarianos. 2008. “Evolution of Aroma Volatiles during Storage
of Sourdough Breads Made by Mixed Cultures of Kluyveromyces marxianus
and Lactobacillus delbrueckii ssp. Bulgaricus or Lactobacillus helveticus.” Food
Chemistry 107 (2): 883–9.
Plessas, Stavros, Athanasios Alexopoulos, Argyro Bekatorou, Ioanna Mantzourani,
Athanasios A. Koutinas, and Eugenia Bezirtzoglou. 2011. “Examination of
Freshness Degradation of Sourdough Bread Made with Kefir through Monitoring
the Aroma Volatile Composition during Storage.” Food Chemistry 124 (2):
627–33.
Pozo-Bayon, Maria Angeles, Elisabeth Guichard, and Nathalie Cayot. 2006. “Feasibility
and Application of Solvent Assisted Flavour Evaporation and Standard Addition
Method to Quantify the Aroma Compounds in Flavoured Baked Matrices.” Food
Chemistry 99: 416–23.
Rannou, Cécile, Delphine Laroque, Emilie Renault, Carole Prost, and Thierry Sérot.
2016. “Mitigation Strategies of Acrylamide, Furans, Heterocyclic Amines and
Browning during the Maillard Reaction in Foods.” Food Research International
90: 154–76.
Rega, Barbara, Aurélie Guerard, Julien Delarue, Muriel Maire, and Pierre Giampaoli.
2009. “On-Line Dynamic HS-SPME for Monitoring Endogenous Aroma Compounds
Released during the Baking of a Model Cake.” Food Chemistry 112: 9–17.
Rooney, L. W., A. Salem, and J. A. Johnson. 1967. “Studies of the Carbonyl Compounds
Produced by Sugar-Amino Acid Reactions. I. Model Systems.” Cereal Chemistry
44: 576–83.
Rychlik, Michael and Werner Grosch. 1996. “Identification and Quantification of
Potent Odorants Formed by Toasting of Wheat Bread.” LWT—Food Science and
Technology 29: 515–25.
Sakač, Marijana, Mladenka Pestori, Anamarija Mandic, Aleksandra Misan, Natasa
Nedelkkovic, Dubravka Jambrec, Pavle Jovanov, Vera Lazic, Lato Pezo, and Ivana
Sedej. 2016. “Shelf-Life Prediction of Gluten-Free Rice-Buckwheat Cookies.”
Journal of Cereal Science 69: 336–43.
 Bakery Products 437

Schieberle, Peter and Werner Grosch. 1992. “Changes in the Concentrations of Potent
Crust Odourants during Storage of White Bread.” Flavour and Fragance Journal 7
(4): 213–8.
Srivastava, R., J. Bousquières, S. Roux, C. Bonazzi, and B. Rega. 2017. “Kinetic Study of
Furan and Furfural Generation during Baking of Cake Models.” Food Chemistry
267: 329–36.
Taylor, Publisher, E. Guichard, and N. Cayot. 2007. “Flavor Control in Baked Cereal
Products.” Food Reviews International 22 (4): 37–41.
Ur-Rehman, Salim, Alistair Paterson, and John R. Piggott. 2006. “Flavour in Sourdough
Breads: A Review.” Trends in Food Science and Technology 17: 557–66.
Yang, Ni, Joanne Hort, Robert Linforth, Keith Brown, Stuart Walsh, and Ian D. Fisk.
2013. “Impact of Flavour Solvent (Propylene Glycol or Triacetin) on Vanillin,
Parameters and Sensory Perception of Shortcake Biscuits over Accelerated Shelf Life
Testing.” Food Chemistry 141: 1354–60.
Zehentbauer, G. and Werner Grosch. 1998a. “Crust Aroma of Baguettes I. Key Odorants
of Baguettes Prepared in Two Different Ways.” Journal of Cereal Science 28: 81–92.
https://doi.org/10.1006/jcrs.1998.0184.
Zehentbauer, G. and Werner Grosch. 1998b. “Crust Aroma of Baguettes II. Dependence
of the Concentrations of Key Odorants on Yeast Level and Dough Processing.”
Journal of Cereal Science 28: 93–6.
 Chapter 22
Recent Advances in the Study
of Grape and Wine Volatile
Composition: Varietal,
Fermentative, and Aging
Aroma Compounds
Pilar Rubio-Bretón, Maria Rosario Salinas, Ignacio Nevares,
Eva Pilar Pérez-Álvarez, Maria del Álamo-Sanza, Sandra Marín-San
Román, Gonzalo Luis Alonso, and Teresa Garde-Cerdán

CONTENTS

22.1 Introduction 439

22.2 Varietal and Pre-Fermentative Aromas 440
22.3 Fermentative Aromas 445
22.4 Aging Aromas 451
22.5 Conclusion 455
Acknowledgments 455
References 455

22.1 INTRODUCTION

Wine aroma is very complex due to the large number of volatile compounds that it con-
tains. More than 800 volatile compounds have been found in wine, and some of them
provide a relevant response in the olfactory system with a wide range of concentrations
that varies from a few ng/L to hundreds of mg/L (de Castilhos et al., 2019). These com-
pounds are widely known thanks to the large number of analytical techniques available
(Flanzy, 2000). Wine aroma is important for determining wine quality and therefore
wine value (Aleixandre-Tudo et al., 2015; Gambetta et al., 2014; Mihnea et al., 2015).
Various aromatic compounds can be distinguished in wine, and they depend on several
factors such as grape variety, season, terroir, grape maturity, viticultural practices, fer-
mentation conditions, and conservation, with grape maturity and alcoholic fermentation
being the most critical stages. In addition, the release and perception of the aroma largely
depend on physical and environmental aspects, such as the wine temperature or the shape
of the glass (Ayestarán et al., 2019; Gambetta et al., 2014; Styger, Prior, and Bauer, 2011).

439
440 Food Aroma Evolution

The constituents of wine aroma have been classified according to their origin:

• Varietal aromas: These come from the grape.

• Pre-fermentative aromas: These are formed during harvest and last until the
beginning of the alcoholic fermentation.
• Fermentative aromas: These are formed by yeasts in the alcoholic fermentation
or by lactic bacteria in the malolactic fermentation.
• Aging aromas: These come from the wine conservation stage.

This chapter is divided into three sections; the first corresponds to the primary aromas
(varietal and pre-fermentative), the second to the fermentative or secondary aromas, and
the third to the aging or tertiary aromas, the origin of which and the main factors affect-
ing their content in grapes and wines are described.

22.2 VARIETAL AND PRE-FERMENTATIVE AROMAS

The varietal aromas of wine are composed by the odorous compounds that come from
the grape’s own metabolism. Therefore, they depend on the grape variety and are influ-
enced by the vineyard’s edaphic and climatic conditions and by the viticultural prac-
tices carried out. The grape aroma metabolites constitute a very complex group of
substances that can be found in free form, that is, as volatile molecules and therefore
odorants, or in bound form, that is, as nonvolatile precursors and therefore odorless
molecules. There are some wines whose varietal aromatic notes correspond to their
grapes, among them are the Muscat varieties, in which the first odorant compounds of
the terpenes family were identified.
However, most wine grapes have no smell and, therefore, odorless precursors of
aroma are the most abundant and important compounds in the aromatic group of
grapes. These precursors begin their transformation in their corresponding odorant
molecules from the moment the grapes are collected, throughout the different stages
of the winemaking process, especially during alcoholic fermentation and during wine
aging. Aroma precursors are organic molecules of glycosidic, amino acidic, and lipidic
natures, which provide the volatile compounds by means of the enzymes of the grape
itself or of the microorganisms that participate in the different stages of the winemak-
ing, fundamentally the yeasts. Also, the wine’s chemical conditions, especially its pH
and temperature, participate in the hydrolysis of odorants from their precursors.
The odorant compounds of wines that come from grapes can be grouped accord-
ing to their chemical family in methoxypyrazines, terpenes, C13 norisoprenoids, benze-
noids (phenylpropanoids), thiols, and C 6 compounds. Among them, methoxypyrazines
in grapes are found only as free volatiles and thiols only as precursors. Other com-
pounds are found in the grape, as free volatiles or as precursors, with the bound form
being predominant.
Methoxypyrazines exist in grape berries as volatile free compounds and have a veg-
etable aroma that is only acceptable in some white wines. Their presence is detrimen-
tal in some red wines, especially those from the Cabernet family (Ward et al., 2015).
There are several types of methoxypyrazines, but 3-isobutyl-2-methoxypyrazine (IBMP),
in particular, is the most abundant, and exceeds its low olfactory perception threshold
(OPT) (Table 22.1) in wines from cool climates, especially if they come from early har-
vests, since these compounds diminish considerably with maturation until they disappear
 Grape and Wine Volatile Composition 441

TABLE 22.1 Some Varietal and Pre-Fermentative Aroma Compounds in Wines

Compounds Aromatic Descriptor ODT (µg/L)
Methoxypyrazines 3-Isobuthy-2-methoxypyrazine Green pepper, pea pod 0.001a
(IBMP) Green pepper, pea 0.002a
3-Isopropyl-2-methoxypyrazine pod, earthy
(IPMP)
Monoterpenes Geraniol Geranium, rose 30b
Linalool Coriander seed, rose 15b
Nerol Floral, orange flowers 15c
Citronellol Rose, lemongrass 100c
cis-Rose oxide Floral, green 8a
Sesquiterpenes Farnesol Floral 1000d
Nerolidol Floral 15b
Rotundone Black or white pepper 0.016e
C13 β-Damascenone Apple sauce, rose 0.05b
norisoprenoids β-Ionone Violet 0.09f
(E)-1​-(2,3​,6-Tr​imeth​ylphe​nyl) Geranium leaf 0.024a
butan-​1,3-d​iene (TBP)
Benzenoids Benzyl alcohol Sweet, fruity, chemical 200000b
2-Phenylethanol Floral, rose 10000b
Methyl vanillate Honey, vanilla 3000c
Ethyl vanillate Sweet, honey, vanilla 990c
Syringol Smoky 2000f
Thiols 3-Mercaptohexan-1-ol (3mh) Passion fruit, 0.06a
3-Mercaptohexyl acetate (3mha) grapefruit 0.004a
4-Mercapto-4-methylpentan-2-one Boxwood, passion 0.0008a
(4MMP) fruit
Boxwood, broom
C6 compounds (E)-2-Hexenal Bitter almond 4f
(Z)-3-Hexenol Herbaceous 400f
1-Hexanol Herbaceous 8000f
Olfactory detection threshold (in wine or model solution): ODT; aDarriet et al. (2012); bGuth (1997);
cEtievant (1991); dFranco et al. (2004); eWood et al. (2008); fFerreira et al. (2000).

(Sidhu et al., 2015). Another factor that can affect IBPM concentrations is the vine water
status, as it has been shown that irrigation of the vines increases the content of IBPM
(Mendez-Costabel et al., 2014).
The terpenes found in most grape cultivars are alcohols and oxides of 10 carbon
atoms, such as geraniol, linalool, citronellol, nerol, and rose oxide, among others. They
have floral aromas and a low OPT (Table 22.1). Terpenes significantly exceed their OPT
in the grapes and wines of the Muscat family and have an important role in the wine
aroma of other white varieties, such as Albariño, Gewürztraminer, and Chardonnay, but
not in wines from red varieties of grape. Among terpenes, however, there are synergistic
effects that could make them effectively participate in the aroma, even without exceed-
ing their OPT. They are synthesized during grape ripening and are located mainly in the
grape skin. Although there is no unanimity in this regard, free volatiles accumulate before
industrial maturity, while precursors continue to increase until overripe (Hjelmeland and
Ebeler, 2015). Terpenes are sensitive to viticultural practices, soil type, exposure to light,
UV-B radiation, water deficit, basal leaf removal, crop thinning, and pests (Robinson et
442 Food Aroma Evolution

al., 2014). In wine, they undergo chemical reactions and rearrangements to form other
terpenes, which explains the aroma aging suffered by certain wines (Black et al., 2015).
Although less studied, terpenes of 15 carbon atoms, or sesquiterpenes, are also pres-
ent in grapes and wines. Among them, farnesol and nerolidol, linear sesquiterpenes that
have floral aromas, are the most abundant, but their OPTs are higher than the C10 com-
pounds, which make them less important for the wine aroma (Table 22.1). However,
a recent study suggests that the content of farnesol and nerolidol in acidic conditions
decreases during aging due to cycling, which may contribute to the balsamic and spicy
aroma of Corvina grapes and wines (Slaghenaufi and Ugliano, 2018). A sesquiterpene
which has been further investigated recently is rotundone, which is a powerful odorant
with a spiced aroma (black or white pepper). This compound was initially associated with
the Australian cool-climate Shiraz grapes, but is also found in other red varieties, such as
Cabernet Sauvignon, Graciano, or Pinot Noir, among others (Mattivi et al., 2011).
Norisoprenoids come from the oxidative degradation of the carotenoids, which
give rise to compounds of 13 carbon atoms. Carotenoids share their biosynthetic onset
with sesquiterpenes, having the compound farnesyl pyrophosphate in common, as a
starting point, so usually the set of monoterpenes, sesquiterpenes, and C13 norisopren-
oids are called terpenoids. The most important C13 norisoprenoids are those that have
the megastigmane structure and have a wide range of odors, from floral and fruity to
scents reminiscent of tobacco and kerosene, but the most abundant in grapes and in
most wines are those with pleasant aromas, especially the oxygenated megastigmanes
β-damascenone and β-ionone, both with very low OPTs (Table 22.1). Recently, a positive
correlation between C13 norisoprenoids and fruity aromas, especially β-damascenone,
as an enhancer of this aroma, has been found (Sáenz-Navajas et al., 2015), and so they
have been the focus of many current studies. C13 norisoprenoids are derived from pho-
toactive carotenoids (Chen et al., 2017), and therefore an increase in the penetration
of light through the canopy, achieved through various agronomic practices, influences
their formation (Hernández-Orte et al., 2015), although this could also be attributed to
the impact of temperature on the cluster that could be confused as being the result of
sunlight-related effects.
Although dependent on grape varieties, edaphic and climatic conditions, and viticul-
tural practices, the accumulation of norisoprenoids has been reported to begin earlier than
the optimal technological ripening. This has been used to mitigate the effect of climate
change on the quality of wines, since early-harvested grapes produce wines with lower
alcohol content and optimum aromatic quality (Asproudi et al., 2018). Other important
oxygenated C13 norisoprenoids are 3-oxo-α-ionol (tobacco aroma), β-damascone (tobacco
and fruity), and 3-hydroxy-β-damascone (tea and tobacco), but their levels in wines are
usually very low, and their OPTs have not been defined. However, it has recently been
shown that a synergistic effect between 3-oxo-α-ionol, β-damascenone, and TBP (the
major representative of non-oxygenated C13 norisoprenoids) is responsible for the tobacco
aroma of certain wines (Slaghenaufi and Ugliano, 2018).
Terpenoids (monoterpenes, sesquiterpenes, and C13 norisoprenoids) are found in
grapes mainly as odorless precursors in glycosylated or bound form. The free aroma, or
aglycone, joins a sugar moiety through the process of glycosylation, which is one of the
predominant reactions in plants catalyzed by a group of enzymes called glycosyltransfer-
ases. Glycosylation is a mechanism used by plants to facilitate the solubilization, trans-
port, and accumulation of lipophilic compounds, as well as a detoxification process. In
grapes, the sugars are mainly glucoside or disaccharide or trisaccharide glycosides, where
the other sugars are arabinose, rhamnose, apiose, and xylose conjugated to the glucose.
 Grape and Wine Volatile Composition 443

Not all glycosides are present in different cultivars, and they differ in their proportions;
therefore, they have been proposed for use in varietal differentiation (Ghaste et al., 2015).
These glycoconjugates are of great importance in winemaking since they are extracted
from the grapes at the beginning of the process, and, in acidic conditions and reactions
catalyzed by glycolytic enzymes, can release volatile aglycones and participate in the wine
aroma. The highest release of volatile aglycones takes place during alcoholic fermenta-
tion, followed by malolactic fermentation, although sugar and alcohol limit the glycosi-
dase activity of yeasts and bacteria. However, this fact is favored by the acidic pH of the
wine, which is why hydrolysis takes place alongside aging. The glycoconjugate hydrolysis
yields volatile aglycones, but while in the case of glycosylated monoterpenes and sesqui-
terpenes this hydrolysis directly releases free volatiles, in the case of the glycosylated C13
norisoprenoids, it produces non-odorous or less-odorous compounds, which require new
chemical rearrangements to give odorous compounds. This fact is well demonstrated for
β-damascenone (Hjelmeland and Ebeler, 2015).
The accumulation of glycosylated terpenoids begins at ripening, but some studies
have shown a correlation between these compounds and the enological classical param-
eters of the grapes at such a stage. This is possibly due to the difficulty of its determina-
tion, since the glycoconjugate aromas are not analyzed as intact molecules, but indirectly
through the volatile aglycones. Once the glycoconjugates are isolated, they are hydrolyzed
to release the volatile aglycones, which are subsequently determined by gas chromatogra-
phy (GC). However, if the hydrolysis is enzymatic, the specificity of the enzymes implies
the release of only some aglycones, and if the hydrolysis is acidic, important structural
changes occur in the aglycones that do not reflect the original chemical nature of the gly-
cosides (Hjelmeland and Ebeler, 2015; Liu et al., 2017). In recent years, a simple method
has been developed to determinate the glycosylated aroma precursors of white and red
grapes, musts, and wines, using a previous method as a reference that was proposed sev-
eral years ago by Williams et al. (1995). It is based on analysis using high-performance
liquid chromatography-refractive index detection (HPLC-IR) of the glucose released by
acidic hydrolysis of such precursors, called glycosyl–glucose (G-G), a moiety common in
all grape aroma glycoconjugate compounds (Salinas et al., 2012) (Figure 22.1). Therefore,
this G–G analysis allows for the estimation of all of these compounds, and the method
has been adapted to be able to apply it in wineries using UV–VIS spectrometry (Serrano
de la Hoz et al., 2014), which provides the varietal aroma potential index (IPAv). The
evolution of the aroma glycoconjugate compounds by the IPAv parameter during three
consecutive vintages was estimated by Crespo et al. (2018). They observed a high paral-
lelism between the evolution of the IPAv and °Brix, as well as its relation with the climatic
fluctuations during the period of the grape ripening.
In order for the glycosylation to take place, it is necessary that the aglycones have an
alcohol, phenol, or acid function, so in grapes there are a large number of glycosylated

FIGURE 22.1 Hydrolytic breakdown of the glycosidic bonds of the glycosylated precur-
sor of linalool and release of sugars and the volatile aglycone. Glucose and the glycosyl–
glucose moiety (G–G) are shown in red.
444 Food Aroma Evolution

precursors in addition to the terpenoids. Thus, many benzenoids, especially phenylpro-

panoids, which are present in grapes in very low concentrations as free volatiles, can be
found in important levels in the form of glycosylated precursors. Glycosylated forms of
volatile phenols such as eugenol, guaiacol, vanillin, and their derivatives, can contrib-
ute to the spicy aroma of wines, without these having been subjected to a process of
aging in contact with oak wood (Table 22.1). It is also noted that the balsamic aroma
associated with the “terroir” of many wines can be attributed to these phenylglycosyl-
ated compounds. The mechanism of glycosylation as a detoxifying process of plants has
been demonstrated recently by Hayasaka et al. (2013) in relation to the effect that the
smoke from forests fires next to vineyards produced in their wines. These wines showed
a clear smoky odor, as a consequence of the hydrolysis of the glycosylated precursors
generated by the plant with volatile phenols present in the smoke, especially syringol and
guaiacol. The same effect was observed when vines were treated with oak lactones and
with oak extracts (Pardo-García et al., 2015), where a previous step to glycosylation was
observed, consisting in the opening of the lactone ring to give the corresponding hydroxy-
acid. Coumaric and ferulic cinnamic acids, located mainly in the grape skins, are also
precursors of volatile phenols, such as vinyl and ethylphenol and vinyl and ethylguaiacol,
for which enzymes with decarboxylation activity are required. This activity is inhibited
when there are catechins; therefore, the presence of these volatile phenols is more impor-
tant in wines of short maceration, like white wines, than in those of long maceration, as
happens with red wines (Darriet, Thibon, and Dubourdieu, 2012).
Some wines have pleasant aromas of sulfur, which exclusively exist in grapes in the
form of non-odorous precursors of cysteine and glutathione. They are located in the
pulp or in the skin, depending on the type of precursor. These compounds, called poly-
functional thiols, are absent in musts and are formed mainly during the alcoholic fer-
mentation, as long as the yeasts possess a specific lyase activity for the S-cysteine and
glutathione conjugates. The most important thiols are 3-mercaptohexan-1-ol (3MH),
3-mercaptohexyl acetate (3MHA), and 4-mercapto-4-methyl-pentan-2-one (4MMP),
powerful odorants to which the typical aromas of some Sauvignon Blanc and Verdejo
wines are attributed, with tropical notes reminiscent of grapefruit, boxwood, and pas-
sion fruit. They have also been found in many other varieties, reds and whites, although
they do not seem to define the aroma of their wines. It has been observed that they are
dependent on the vine agronomic practices (Helwi et al., 2015) and that, although they
are released at the beginning of the alcoholic fermentation, they are chemically unstable
and very easily oxidized, and even in reducing environments, they can be transformed
into undesirable aromas. Thus, for their preservation, a compromise between the oxy-
gen content of the wine and the reduction conditions is necessary (Herbst-Johnstone,
Nicolau, and Kilmartin, 2011).
All the grapes have, as free volatiles, other types of compounds, the so-called C6
compounds, which are referred to as “green leaf volatiles,” since they are characterized
by a “herbaceous” and “green” aroma. C6 compounds mainly consist of alcohols and
aldehydes, such as 1-hexanol, (Z)-3-hexenol, (E)-3-hexenol, and (E)-2-hexenal. The alco-
hols have a high OPT, so they are among the less odorant volatiles in grapes (Table 22.1).
The evolution of these compounds in grapes is different, as aldehydes decrease toward the
end of ripening, and alcohols increase, which is a positive aspect, because alcohols have a
higher OPT than aldehydes, resulting in a less pronounced herbaceous character (Kalua
and Boss, 2010). In addition, their content can be increased from the moment in which
the berry is separated from the cluster and in all the other pre-fermentative stages, which
is why they are called pre-fermentative volatiles. Since they come from the grape variety,
 Grape and Wine Volatile Composition 445

many authors consider them as varietal aromas; however, they are common in all grapes,
and therefore in this chapter they are considered pre-fermentative aromas. The main agent
that triggers their formation is oxygen, which activates enzymatic systems that generate
aldehydes and alcohols from polyunsaturated fatty acids; these reactions are favored by
temperature. In the grapes, C6 compounds can also be found as non-odorous glycosylated
precursors, but their concentration is much lower than those of the free forms, since to
be generated is necessary oxygen from the air as the initiator of their synthesis from fatty
acids. The increase of C6 compounds in musts and wines can be avoided by reducing the
contact of the harvest with the oxygen and keeping it at low temperatures. This is the
reason why mechanical harvesting increases the herbaceous character. In the same way,
night-harvesting and use of inert gases are imposed to transport the harvest to the cellars,
and avoid uncontrolled oxidation in other pre-fermentative stages (Boselli et al., 2010).
During alcoholic fermentation, yeasts reduce these aldehydes to alcohols, which are much
less odorous and normally do not exceed their OPT in wines, but if they do, it should be
considered a wine defect.
The amino acids of the grapes play a decisive role in the process of the alcoholic
fermentation, since they are nitrogen sources for yeasts. Since the amino acid profile of
the grapes is a varietal characteristic, the fermentative aromatic profile generated by the
amino acids of the grapes is also a varietal characteristic. The amino acids of the grapes
are therefore precursors of fermentative aromas; such will be studied in the next section.

22.3 FERMENTATIVE AROMAS

Fermentative aromas are all those wine volatile compounds that are formed during the
vinification. The first step in this process is the alcoholic fermentation, which is car-
ried out by yeasts (Saccharomyces and non-Saccharomyces); and the second step is the
malolactic fermentation, which is carried out by lactic bacteria. For the wine aroma, the
alcoholic fermentation is a very important process, since it is responsible for the wine
notes common to every wine. In addition, the volatile compounds formed during this
stage quantitatively represent most of the constituents of the wine aroma. Conversely,
the malolactic fermentation subtly modifies the wine aroma (Flanzy, 2000). The volatile
compounds formed in the fermentative stage belong to the following groups: alcohols,
acids, esters, carbonyl compounds, sulfur compounds, lactones, and volatile phenols
(Flanzy, 2000). Due to their contribution to the wine aroma, the higher alcohols and
their acetates, and the fatty acids and their ethyl esters, are the most studied compounds
(Molina et al., 2007; Wang et al., 2018).
Higher alcohols are quantitatively dominated by 2- and 3-methyl-1-butanol (known
as isoamyl alcohols), n-propanol, isobutanol, n-butanol, n-pentanol, 2-phenylethanol,
3-methylthio-1-propanol, tyrosol, and tryptophol. Their average total content in the
wine is of the order of 300–400 mg/L, above these values they can contribute in a
negative way to the wine aroma. The formation of these compounds is linked to the
sugar and amino acid metabolism, and is therefore strongly influenced by grape nitrogen
composition. In this way, the formation of these compounds depends on the yeast nitro-
gen demand (Flanzy, 2000). Isobutanol and isoamyl alcohol suppress fruity and woody
notes, but not leather, animal, and ink notes, and can impart a negative quality to the
wine. The presence of 2-phenylethanol, which provides honey and rose notes, can con-
tribute to the wine aroma (Wang et al., 2018). The most significant acetates are isobutyl
acetate, which contributes to the fruity aroma, isoamyl acetate, which contributes to the
446 Food Aroma Evolution

banana flavor, and 2-phenylethyl acetate, which contributes to the floral aroma. These
esters are derived from their corresponding higher alcohols, isobutanol (derived from the
amino acid valine), isoamyl alcohol (derived from leucine), and 2-phenylethanol (derived
from phenylalanine) (Garde-Cerdán and Ancín-Azpilicueta, 2008; Gutiérrez-Gamboa
et al., 2017).
The fatty acids and their ethyl esters are, together with the higher alcohols and their
acetates, the main markers of the fermentative aroma. The fatty acids and their ethyl
esters are produced in high quantities in wines, presenting a pleasant aroma; they are
actually the principal esters responsible for the smells judged as pleasant, mainly fruity.
Conversely, although the fatty acids have been related as not desirable odors, they are
considered necessary for a good balance of the wine aroma (Flanzy, 2000). Hexanoic,
octanoic, and decanoic acids are common volatile fatty acids in wine, with octanoic acid
being the most abundant. These fatty acids contribute to fatty, rancid, and cheese flavors
(Wang et al., 2018). The ethyl esters contribute to the pleasant aroma, and are formed
during the alcoholic and malolactic fermentations, with those coming from the action of
the yeasts in alcoholic fermentation being the most important for the wine aroma (Belda
et al., 2017). The formation of esters, higher alcohols, and fatty acids from sugars or
amino acids (Ehrlich’s pathway) is shown in Figure 22.2.
There are different factors that can affect the volatile compound content in wines.
These factors include viticultural practices, among them, basal defoliation and foliar
application; pre-fermentative techniques, such as the addition of enological tannins and
different types of maceration; fermentative techniques, such as the use of different strains
of yeasts and lactic bacteria, as well as fermentation conditions; and, moreover, the grape
nitrogen composition, which is another important factor. Table 22.2 shows a summary of
how the factors and practices studied affect the content of fermentative aromas.

FIGURE 22.2 Simplified metabolic map of ester formation. Genes that codify enzymes
involved in ester formation are indicated. The substrate availability depends mainly
on carbon, nitrogen, and fatty acid metabolism, while the ester synthase activity is
mainly determined by the activity of the corresponding genes. Thus, factors that affect
yeast metabolism or ester synthase gene regulation will influence ester synthesis. (From
Belda et al., 2017.)
 Grape and Wine Volatile Composition 447

TABLE 22.2 Summary of the Factors and Practices That Can Affect the Content of the
Wine Fermentative Volatile Compounds
Factors Practices Volatile Compounds Improved Reference
Viticultural Basal defoliation 1-Hexanol, 2-phenylethanol, (Wang et al., 2018)
techniques 2-phenylethyl acetate, decanoic acid,
ethyl octanoate
Foliar application Amino acids (Sánchez-Gómez
et al., 2016)
Sunlight exposure Alcohols, ethyl esters, lactones, (Vilanova et al.,
terpenes, C13 norisoprenoids 2017)
Regulated deficit Benzaldehyde, benzyl alcohol, phenyl- (Ju et al., 2018)
irrigation (RDI) ethyl alcohol, 2-methyl-1-propanol, (García-Esparza
3-methy-l-butanol, amino acids et al., 2018)
Volatile compounds
Pre- Addition of Higher alcohols, isoamyl acetate, (Chen et al., 2016)
fermentative tannins 2-phenylethanol (Chen et al., 2018)
techniques Higher alcohols
Maceration Alcohols, carbonyl compounds (Ayestarán et al., 2019)
Acetaldehyde, ethyl acetate, C6 (Lukić et al., 2017)
compounds, esters (Mihnea et al., 2015)
Alcohols, ethyl vanillate (Aleixandre-Tudo
Alcohols et al., 2015)
Fermentative Type of yeast Lactones, minor ethyl esters (Ramírez et al., 2016)
techniques Ethyl acetate, esters, higher alcohols, (Varela et al., 2017)
sulfur compounds (Puertas et al., 2018)
Ethyl propanoate, ethyl (Blanco et al., 2013)
2-methylpropanoate, ethyl (Ma et al., 2017)
hydrocinnamate, 2-phenyl acetate,
2-methylpropyl ethanoate
Higher alcohols, 2-phenylethanol,
esters (ethyl 2-hydroxypropanoate)
and fatty acids
Acetates, ethyl esters, fatty acids
Type of bacteria Compounds related to fruity aroma (Styger et al., 2011)
and butter note
Nitrogen Volatile compounds in general (Fairbairn et al.,
compounds Esters 2017)
(Rollero et al., 2015)
Fermentation 15°C: Compounds related to a fresh (Molina et al., 2007)
conditions (Tª, and fruity aroma (Rollero et al., 2018)
pH, agitation) 28°C: Compounds related to floral
aromas
80 rpm: Optimal for the production
of fermentative aromas
Others (grape 2-Phenylethyl acetate, C6 alcohols (de Castilhos et al.,
pre-drying, Lipids: Esters, higher alcohols, volatile 2019)
submerged cap, acids (Varela et al., 2012)
supplementation Oxygen: Higher alcohols
with O2 and Lipids + Oxygen: Higher alcohols
lipids)
448 Food Aroma Evolution

Viticultural practices are those carried out in the vineyard that can affect plant
growth and grape composition. Among these practices, basal defoliation, one of the
most common current viticulture management practices, is used in order to modify the
effect of the climate in the vineyard and is applied with the aim of improving the wine
quality. The effects of basal defoliation in certain fermentative volatile compounds
(2-phenylethanol, 2-phenylethyl acetate, decanoic acid, and ethyl octanoate) mainly
depend on grape maturity, and, to a lesser extent, on grape variety and climatic condi-
tions (Wang et al., 2018). Grape fatty acid and amino acid levels are modified by envi-
ronmental factors, such as exposure to sunlight (Wang et al., 2018). In the same way,
the sunlight exposure facilitated by the training system chosen in the vineyard could
affect wine volatile composition, as observed by Vilanova et al. (2017) in Albariño
wines. This fact may be related to the effective canopy surface area and the density
that the training system allows. Thus, wines from the Geneva Double-Curtain training
system were found to have the highest total concentration of alcohols, ethyl esters, and
lactones, while wines from the Scott-Henry training system had the highest concentra-
tion of terpenes and C13 norisoprenoids.
On the other hand, Ju et al. (2018) studied the effect of different regulated deficit
irrigation (RDI) strategies on Cabernet Sauvignon grapes and wine volatile compounds.
They observed that RDI treatments increased the levels of aromatic compounds such
as benzaldehyde, benzyl alcohol, and phenylethyl alcohol with respect to the highest
watered treatment (100% grapevine estimated evapotranspiration, ETc). Also detected
were increments of phenylethyl alcohol, 2-methyl-1-propanol, and 3-methyl-1-butanol
content in wines elaborated from RDI treatment. They related these increases to the
effects caused by the irrigation strategy used on the concentration of the amino acids,
especially those precursors of volatile compounds. Also, García-Esparza et al. (2018)
observed that low watered treatments (rainfed or irrigation at 25 or 50% of ETc) would
result in a more adequate strategy due to the increase of the volatile compound content,
unlike the most irrigated treatment which seemed to produce an increase of herbaceous
related aromas (hexanal and (E)-2-hexenal), which are not desirable for wines. Another
viticultural technique widely used is foliar application, which consists of enriching
plants with different nutrients through the leaves. It has been shown that the application
of different aqueous wood extracts increases the amino acids content, and therefore the
formation of volatile compounds during alcoholic fermentation (Sánchez-Gómez et al.,
2016). However, the wine aromatic profile was scarcely affected by the application of
several nitrogen sources as fertilizers (Rubio-Bretón et al., 2018). The pre-fermentative
techniques are those that are made in the cellar before starting the winemaking. One of
the most carried out pre-fermentation techniques is the addition of enological tannins.
In this sense, it has been observed that treatments with tannins significantly improve
wine aromatic complexity, inhibiting the formation of higher alcohols in alcoholic fer-
mentation and increasing isoamyl acetate (banana) and 2-phenylethanol (floral aromas)
content (Chen et al., 2016). However, this technique affects the synthesis of methanol
and diacetyl, which increases excessively. On the other hand, it has been seen that the
addition of these compounds is negative for floral and fruity aromas, but positive for the
formation of higher alcohols in fermentations involving Saccharomyces cerevisiae and
less effective in those involving non-Saccharomyces yeasts (Chen et al., 2018). Another
pre-fermentative technique that affects the content of fermentative aromas is the dif-
ference in maceration type, as several works have studied. The wines produced by car-
bonic maceration have a higher content of alcohols and carbonyl compounds, but lower
 Grape and Wine Volatile Composition 449

content of C6 alcohols and volatile acids than conventionally produced wines (Ayestarán
et al., 2019). On the other hand, it has been reported that standard macerated wines
contain a higher amount of higher alcohols, that wines from cold maceration prior to
the fermentation showed the highest levels of acetaldehyde and ethyl acetate, and that
maceration at high temperature was the best for increasing the amount of esters and
fatty acids in wine (Lukić et al., 2017). In another study, fermentation with the skins,
and contact with the skins before fermentation, led to lower levels of terpenes, esters,
acids, and thiols, and higher levels of alcohols, compared to fermented wines without
any type of contact with the skins (Aleixandre-Tudo et al., 2015). Fermentative tech-
niques are those carried out during fermentation. The most significant fermentative
factors are the different types of yeasts (Figure 22.3) and bacteria and the fermentation
conditions (temperature, pH, level of agitation).
Yeasts are responsible for the transformations that occur during the alcoholic fermen-
tation. These microorganisms degrade the two fermentable sugars in the grapes, glucose
and fructose, to produce ethanol and carbon dioxide. During fermentation, numerous
yeast species can coexist, with the S. cerevisiae species being the most common. In recent
years, the effect of these yeasts on the aromatic wine composition has been studied.
A study reveals that wines fermented with Torulaspora delbrueckii had dry fruity and
pastry aromas, and low intensities of fresh fruit aromas, due to the reduction in the
content of the main esters and increase in the content of lactones and some ethyl esters
(Ramírez et al., 2016). Another study, in which two types of yeast were used, showed
that wines produced by Metschnikowia pulcherrima contained 1.0% v/v less ethanol
than wines fermented with S. cerevisiae and higher concentrations of ethyl acetate, total
esters, total higher alcohols, and total sulfur compounds, while wines fermented with
S. uvarum showed a reduction of 1.7% v/v in ethanol and a higher total concentra-
tion of higher alcohols (Varela et al., 2017). On the other hand, the impact of different

FIGURE 22.3 Aromatic compounds formed by yeasts during the alcoholic fermentation.
(From Styger et al., 2011.)
450 Food Aroma Evolution

fermentative strategies on two white wines was evaluated: inoculation with S. cerevisiae
yeast, sequential inoculation (T. delbrueckii/S. cerevisiae), and spontaneous fermenta-
tion. The wines inoculated with S. cerevisiae and T. delbrueckii improved the nuances of
the ripe fruit and showed a higher content of ethyl propanoate, ethyl 2-methylpropano-
ate, and ethyl dihydrocinnamate. Thus, the spontaneous fermentation of wines presented
improved nuances of “stone fruit” (peach, nectarine, apricot) related to the higher con-
tents of 2-phenylethyl acetate and 2-methylpropyl ethanoate (Puertas et al., 2018). It
has also been shown that wines produced by a strain of S. cerevisiae, as well as wines
produced by spontaneous fermentation, produced high concentrations of esters, mainly
ethyl 2-hydroxypropanoate, and fatty acids (Blanco et al., 2013). In another work, vola-
tile content was compared in fermentations carried out with S. cerevisiae and mixed
fermentations (Pichia fermentans/S. cerevisiae). The results revealed that mixed fermen-
tations increased the content of acetates, ethyl esters, fatty acids, and many other volatile
fermentative compounds (Ma et al., 2017).
Another factor of great importance in the production of fermentative volatile com-
pounds is the fermentation conditions. One of the conditions that this most affects is the
temperature of the fermentation, but there are also other factors, such as pH or agitation
speed. Regarding the temperature of fermentation, its effect has been evaluated and it
has been found that at lower temperatures, higher concentrations of compounds related
to fresh and fruity aromas were found, while at higher temperatures, higher concentra-
tions of floral aromas (Molina et al., 2007) and esters were found (Rollero et al., 2015).
The agitation speed during fermentation is another factor that influences the production
of volatile aromas; it was found that 80 rpm was more effective than 125 and 40 rpm
for the production of fermentative aromas (Rollero et al., 2018). The factors explored
up to this point are the most important, but there are other factors that can modify the
content of fermentative volatile aromas in wine; some of these are the changes in the
winemaking process, where wines that have been previously dried presented a note of
marmalade, possibly due to the presence of 2-phenylethyl acetate (de Castilhos et al.,
2019), and supplementation with oxygen and lipids, where the addition of lipids increased
the concentration of esters, higher alcohols, and volatile acids, the addition of oxygen
increased the concentration of higher alcohols, and the addition of oxygen together with
lipids, showed an additive effect on the concentration of higher alcohols, but oxygen
suppressed the potentiating effect of lipids in the formation of volatile esters and acids
(Varela et al., 2012). After alcoholic fermentation, some wines may undergo malolactic
fermentation. The agents of these malolactic transformations are lactic bacteria, mainly
of the Oenococcus oeni species and to a lesser extent the Pediococcus damnosus and
Lactobacillus species. It has been shown that bacteria can influence the aroma, pro-
ducing volatile metabolites, and modifying the aroma compounds derived from grapes
and yeast metabolism (Styger et al., 2011). In general, it has been found that malolactic
fermentation can improve the fruity aroma and the note of butter, and is able to reduce
the green aroma of the wine (Styger et al., 2011). Finally, the level of nitrogen com-
pounds in the must can affect the volatile composition of the wine, since amino acids (the
main source of nitrogen for yeasts), are precursors of volatile fermentative compounds
(Sánchez-Gómez et al., 2016). The availability and utilization of nitrogen by S. cerevisiae
significantly influence the fermentation kinetics (Arias-Gil, Garde-Cerdán, and Ancín-
Azpilicueta, 2007) and the production of volatile compounds important for wine aroma
(Garde-Cerdán and Ancín-Azpilicueta, 2008). It has been shown that the concentration
of volatile compounds in wine was directly related to the amount of amino acids present
 Grape and Wine Volatile Composition 451

in the must, and that nitrogen has a double role, promoting yeast growth to carry out the
alcoholic fermentation, and producing volatile compounds (Fairbairn et al., 2017).

22.4 AGING AROMAS

The aging period for a wine involves the development of a series of processes that deter-
mine the properties of the end product. The wine can be subjected to some of these pro-
cesses during the maturing period, such as contact with wood, storage in containers of
different characteristics (stainless steel, concrete, granite, or wood), storage in bottles, or
all of these. The aging stage is the final and longest period involved in producing wine,
in which the traits acquired in the prior processes are consolidated and new properties,
especially if the wine is kept in contact with wood, are added. During the storage/matura-
tion of wine, tertiary aromas may be generated by the interaction of wine with the com-
pounds yielded by the wood, such as in the case of the aging of wine in barrels, or with
innovative aging technologies, as the conservation of wine in contact with wood frag-
ments, aging on lees, or the application of micro-oxygenation (MOX). The main groups
of volatile substances that wood provides to wine are (Table 22.3): furanic compounds
(furfural, 5-methylfurfural, 5-hydroxymethylfurfural, and their corresponding alco-
hols), phenolic aldehydes (vanillin and syringaldehyde), lactones (cis and trans-β-methyl-
γ-octalactone), volatile phenols (eugenol, guaiacol, 4-methylguaiacol, 4-propilguaiacol,
isoeugenol, 4-vinylguaiacol, syringol, and 4-alilsyringol), and acetic acid, which is not
positive for the aromatic quality of the wine and leads to an increase in volatile acidity.
Most of these compounds are specific substances from the wood that are not found in
the base wine (guaiacol, 4-methylguaiacol, and oak lactones). But some compounds, such
TABLE 22.3 Main Volatile Odorant Compounds Added to Wine by Oak, Aromatic
Descriptors, and Perception Thresholds in Red Wine (µg/L)
Compound Aromatic Descriptor ODT (µg/L)
Furanic compounds Furfural Almond 20,000a,c,e
5-Methylfurfural Toasted almond 450,00a,c,e
5-Hydroxymethylfurfural Almond 100,000d
Phenolic aldehydes Vanillin Vanilla 320a
Syringaldehyde Spices, smoke 50,000a
Lactones cis-Oak lactone Wood, coconut, vanilla 74c,e /54b
trans-Oak lactone Wood, coconut, spices 320c,e / 370b
Volatile phenols Eugenol Spices (clove) 500a,c,e
Guaiacol Smoke, toasted 75a,c,e
4-Methylguaiacol Smoke, burned 65a,c,e
4-Propilguaiacol Leather, animal 10d
Isoeugenol Spices 6d
4-Vinylguaiacol Carnation, pepper 380a,c,e
Syringol Smoke 2000c
Ethylphenols 4-Ethylphenol Horse stable, medicinal 620c
4-Ethylguaiacol Leather, phenolic 150a,c,e
Olfactory detection threshold (in wine or model solution): ODT; aBoidron et al. (1988);
bBrown et al. (2006); cChatonnet et al. (1992); dSáenz-Navajas et al. (2010); eSpillman et al.

(2004a).
452 Food Aroma Evolution

as vanillin, syringaldehyde, and eugenol, can be present in the wine in traces, and their
concentration can significantly increase due to the wood contribution (Flanzy, 2000).
Furfural and 5-methylfurfural have, on their own, a minor impact. However, furfu-
ral has been reported to have an important modifying effect on the aroma perception of
oak lactones, decreasing wood aroma as the levels of furfural are increased (Chira and
Teissedre, 2015). Oak lactones contribute to the olfactory characteristics of wine with
coconut, woody, and oak notes, whereas vanillin is related to vanilla and coffee smells,
and volatile phenols to spicy and smoked flavors (Prida and Chatonnet, 2010). Therefore,
during the process of conservation and aging of the wines, the fruity aromas characteris-
tic of young wines, coming from esters, alcohols, and terpenes, decrease, with more com-
plex aromas that possess other sensory characteristics appearing. The extraction process
of volatile compounds that takes place in oak barrels depends on many factors, among
which stand out the oak species, the geographical origin of the oak, the silvicultural
treatment of the tree, the wood processing in cooperage (seasoning method and oak
toasting intensity during barrel manufacturing), the contact time between the wood and
the wine, and the number of times that the barrels have been used previously (González-
Centeno, Chira, and Teissedre, 2016; Navarro et al., 2018). Besides the characteristics of
the barrel, the grape variety and wine composition have a real impact on the extractive
capacity of wine. Several authors (Garde-Cerdán, Torrea-Goñi, and Ancín-Azpilicueta,
2004; González-Centeno et al., 2016) suggest that the extraction of woody aroma com-
pounds from barrels increased with the increasing degree of alcohol in the wine. Moreover,
according to other authors (Garde-Cerdán et al., 2004; Rodríguez-Rodríguez and
Gómez-Plaza, 2012), high alcohol content and acidity could favor the alcoholysis of lig-
nin, and a higher amount of guaiacol and 4-methylguaiacol could then be released. These
same authors (Rodríguez-Rodríguez and Gómez-Plaza, 2012) also observed a significant
negative correlation between lactone concentration and alcohol content, titratable acid-
ity, and volatile acidity, as well as a significant positive correlation between the vanillin
and the alcohol content and titratable acidity. Oak is the most commonly used barrel
material in the wine aging process. Oak species and their geographical origins consider-
ably affect the volatile composition of wines. The species used most widely in cooperage
are Quercus alba from America and Q. petraea and Q. robur from Europe, mainly from
France. Martínez-Gil et al. (2018) observed that the wines macerated in contact with Q.
alba oak chips showed greater content of furfural, guaiacol, vanillin, syringol, and cis-
oak lactone than the one macerated with Q. petraea oak chips. Also, other authors (Chira
and Teissedre, 2015; Spillman, Sefton, and Gawel, 2004b) suggested than Q. robur oak
contains less eugenol than Q. petraea. Moreover, it is well known that wines aged in
American oak barrels have a higher cis/trans-oak lactone ratio than those aged in French
oak barrels (Chira and Teissedre, 2015; Garde-Cerdán and Ancín-Azpilicueta, 2006a, a;
Gómez-Plaza et al., 2004). The aromatic composition of other oak geographical origins
and wood species have been studied, showing interesting oenological potential as an
alternative species for coopering: Colombian oak (Q. humboldtii) (Martínez-Gil et al.,
2018); Spanish oak (Q. pyrenaica) (Gallego et al., 2012); Slavonia oak (Q. petraea and
Q. robur) (Chira and Teissedre, 2015); acacia (Robinia pseudoacacia) (Fernández de
Simón et al., 2014; Kyraleou et al., 2015); cherry (Prunus avium) (Fernández de Simón et
al., 2014); chestnut (Castanea sativa) (Fernández de Simón et al., 2014); and ash (Fraxinus
excelsior) (Fernández de Simón et al., 2014). Wood seasoning in cooperage affects the
final chemical composition of the wood and therefore the volatile composition of wines
aged in the barrels. The effect of various oak seasoning methods, including natural sea-
soning in open air, artificial seasoning in a kiln, and a mixed method that combines
 Grape and Wine Volatile Composition 453

open-air and kiln drying was studied by Martínez et al. (2008). These authors observed
that the naturally seasoned wood had higher concentrations of volatile phenols, phenolic
aldehydes, furanic compounds, and cis- and trans-oak lactones, than mixed or artificial
methods. The toasting degree of barrels can be classified as light, medium, or high.
Several authors (Dumitriu et al., 2017; Herrero et al., 2016) observed that wines aged in
barrels with a high toasted level showed the highest concentration of volatile aroma com-
pounds highlighting those aromas extracted from the wood. Guaiacol and 4-methyl-
guaiacol have smoky aromas and are indicators of the wood toasting level, since they are
formed by the thermal degradation of lignin during the toasting process. As it has been
observed in many works (Chira and Teissedre, 2013b; Gómez García-Carpintero et al.,
2012; Guchu et al., 2006; Herrero et al., 2016; Vichi et al., 2007), higher toasting levels
correspond to higher concentrations of these compounds, due to exposure of the wood to
heat and fire. On the other hand, several authors (Bosso et al., 2008; Chira and Teissedre,
2013b; Herrero et al., 2016) have observed that the maximum concentrations of syringal-
dehyde, vanillin, furfural, and guaiacol were found in wines that were in contact with
medium toasting level woods. Moreover, the toasting process significantly affects lactone
levels, obtaining more oak lactone content in light toasted than in medium toasted barrels
(Bosso et al., 2008; Chira and Teissedre, 2015), probably due to the thermodegradation
of these heat-sensitive compounds or their loss by volatilization when the oak wood is
subjected to high temperatures or even charring (Chira and Teissedre, 2013b). Moreover,
the watering process during toasting enhances furanic compounds, vanillin, and oak
lactone extraction (Chira and Teissedre, 2015). Because oak extractives from a barrel are
finite, the volatile composition of wines aged in used barrels has been studied by several
authors (Garde-Cerdán, Rodrı ́guez-Mozaz, and Ancín-Azpilicueta, 2002; Gómez-Plaza
et al., 2004). The quantity of these compounds and their rate of extraction diminish with
the utilization of the barrel during successive years. The concentrations of volatile aro-
matic compounds found in wines matured in second-fill barrels are lower than in new
barrels, even if the maturing time is longer. Another factor that affects the concentration
of aromatic compounds in wines is the contact time with the wood. Many works (Chira
and Teissedre, 2013b; González-Centeno et al., 2016; Herrero et al., 2016) show how the
concentration of the main volatile compounds extracted from wood to wine (vanillin,
furfural, guaiacol, eugenol, and lactones) increases over time. In several studies (Chira
and Teissedre, 2013a; Rodríguez-Rodríguez and Gómez-Plaza, 2012), the extraction rate
of furanic compounds in wines in contact with wood reached a maximum after 3 or 6
months of maceration, and after 12 months these compounds were exhausted. This is due
to the fact that the biochemical reduction of furfural and 5-methylfurfural to the corre-
sponding alcohols can be higher than their extraction from wood. Moreover, the maxi-
mum concentrations for the vanillin and guaiacol were at 9 or 12 months, and in the case
of eugenol, isoeugenol, and both lactones, an increase during the whole oak maturation
period was observed. According to Herrero et al. (2016), only methyl vanillate and vinyl-
phenols decreased over time. Methyl vanillate decreased over time because it can be easily
degraded during aging (Jarauta, Cacho, and Ferreira, 2005), while vinylphenols may
decrease during the aging period with the presence of Bretanomyces/Dekkera yeasts,
which promote their transformation to ethylphenols (4-ethylguaiacol and 4-ethylphenol).
These compounds contribute negatively to wine aroma due to the odor associated with
them described as phenolic, horse sweat, or stable odors (Table 22.3). The matter of when
the oak fragments are added also influences the concentration of wine tertiary aromas.
Thus, Kyraleou et al. (2016) observed that the addition of wood chips after fermentation
enhanced the levels of guaiacol, 4-methylguaiacol, and vanillin, compared with the wines
454 Food Aroma Evolution

fermented with chips. On the other hand, the presence of lees during aging can modify
the wine aromatic properties. Thus, the conservation of white wine on yeast lees in oak
barrels significantly reduces the concentration of volatile compounds that can be extracted
from wood. For example, during the aging on lees, the biochemical reduction of vanillin,
with a high contribution to the wine aroma due to its low detection threshold (Table
22.3), to vanillyl alcohol, a very little aromatic compound (Flanzy, 2000). Besides, the
addition of lees to wine aged in oak wood decreases the concentration of phenols, furanic
compounds, phenolic aldehydes, and lactones, due to their binding to the lees, which are
the compounds with the most affinity for the lees eugenol, 4-propylguaiacol, 4-methyl-
guaiacol, furfural, and 5-methylfurfural. This retention is incremented with increased
concentration of lees. On the other hand, guaiacol and γ–nonalactone were the only
compounds that were not bound by the lees (Jiménez-Moreno and Ancín-Azpilicueta,
2007). For all this, aging wine in oak barrels in the presence of lees attenuates the impact
of wood on the wine aroma (Liberatore et al., 2010; Loira et al., 2013). Moreover, during
the aging on the lees of red wines in oak barrels furfurylthiol can be detected, with coffee
aroma, which comes from the reaction between furfural from the wood and hydrogen
sulfide, originating from the lees (Blanchard, Tominaga, and Dubourdieu, 2001).
During the maturation of wine, it is subjected to natural oxygenation, as is the case
with aging in barrels or other containers of permeable materials, such as concrete, gran-
ite, and polyethylene; or to induced oxygenation, such as when it is stored and micro-oxy-
genated in stainless steel tanks (del Álamo-Sanza, Laurie, and Nevares, 2015; Nevares
and del Álamo-Sanza, 2018). The role of oxygen in modifying the wine aroma during
aging is an aspect widely studied during the bottle storage process (Caillé et al., 2010;
Ferreira et al., 2014; Lopes et al., 2009; Ugliano, 2013; Ugliano et al., 2012; Vasserot,
Jacopin, and Jeandet, 2001). However, few papers evaluating its effect on the aroma of
wines aged in oak barrels or in contact with oak alternatives have been found (del Álamo-
Sanza et al., 2010; Fernández de Simón et al., 2010; Gallego et al., 2012; Oberholster et
al., 2015) and none assessing the effect of oxygen on the aroma of wines stored in granite
or concrete. The interaction between the compounds responsible for the wine aroma
and the oxygen available during aging is determined by the type of wood used in the
process, the environmental conditions of storage, and the quantity of oxygen consumed
by the wine. Thus, when red wine is aged in oak barrels for 6 months using the alterna-
tives (oak chips and oak staves) and MOX, those treated with Q. alba, Q. petraea, or
Q. pyrenaica oak staves, together with MOX, consume more oxygen (25.7, 19.8, and
12.5 mg/L, respectively) than those aged with Q. alba+MOX, Q. petraea+MOX, or Q.
pyrenaica+MOX (16.8, 9, and 6.7 mg/L, respectively) oak chips according to a paper
by del Álamo-Sanza et al. (2010). The paper stated that the volatile compound content
from the wood was closely related to oxygen consumption in the wine. So, higher oxygen
consumption by wines treated with staves+MOX was related to higher 5-methylfurfural,
furfural, 2-furanmethanol, cis- and trans-oak lactones, maltol, guaiacol, 4-ethylphenol,
4-ethylguaiacol, vanillin, and syringaldehyde content in all the wines studied, regardless
of the type of oak used: Q. alba, Q. petraea, or Q. pyrenaica (del Álamo-Sanza et al.,
2019; Fernández de Simón et al., 2010). Different studies confirm that oxygen reaches the
wine in a barrel via the joins between the staves and via the wood (Nevares et al., 2016;
Nevares and del Álamo-Sanza, 2014). This oxygen transfer (OTR) follows a dynamic
trend by decreasing with aging, basically due to the wine impregnating the wood (del
Álamo-Sanza, Cárcel, and Nevares, 2017; del Álamo-Sanza and Nevares, 2014, 2017;
Nevares and del Álamo-Sanza, 2014). The type of wood chosen, and the cooperage treat-
ments, determine the wine–wood interaction and therefore the final traits of the wine.
 Grape and Wine Volatile Composition 455

The UVaMOX group is working on the relationships that exist between woods of vary-
ing permeability to oxygen and aromatic composition, as both aspects are decisive to the
characteristics of wine matured in wood. Woodgrain is normally selected according to
its direct relationship with the aromatic profile which the wood will potentially transfer
to the wine, depending on cooperage treatments. Collins et al. (2015) showed that dur-
ing toasting, there are specific differences in the temperature profiles of each oak barrel,
and these differences probably result in differences in the composition of each barrel.
Therefore, the effect of temperature gradients is crucial to controlling the aromatic poten-
tial of wood. Nevertheless, the wood, its anatomy, and its properties without a doubt
determine the aromatic potential, which will be transferred to the wine. Recent studies
by the UVaMOX group have shown that the OTR to the wine through French oak bar-
rels determines the volatile compound quantity ceded to the wine during maturation.
This could be explained by considering the mechanisms that regulate the oxygen transfer
through the wood: this is a process whereby oxygen is diffused through the structure of
the wood into the wine. Therefore, the anatomy of the wood (the proportion of early- and
latewoods, the medullary radius and its inclination) and the density are the main factors
defining its OTR. As the structure of high oxygenation woods differs significantly from
that of low oxygenation ones, the volatile compounds profile that will potentially be
transferred to the wines will also differ greatly.

22.5 CONCLUSION

Aromatic composition is a key aspect of wine quality. Varietal and pre-fermentative

aromas are mainly influenced by grape variety and their state of maturity, viticultural
practices, and edaphic and climatic conditions. For its part, fermentative aroma content
depends on the handling of technologies during pre-fermentation and alcoholic and malo-
lactic fermentation. The choice and management of every one of these aspects are funda-
mental to the final balance of the wine aroma. Knowledge concerning the type of wood
and oxygen during wine maturation is also essential to controlling the process, since they
determine the final aromatic traits obtained. It is crucial to highlight that the choice of
wood according to its physicochemical properties and capacity for oxygenating wine is
also a decisive factor for the aromatic profile acquired by wine during the aging process.

ACKNOWLEDGMENTS

P. R.-B., E. P. P.-Á., and T. G.-C. thank MINECO for funding their doctoral contracts.
S. M.-S. thanks Gobierno de La Rioja for her pre-doctoral contract. M. A.-S. and I.
N. thank Junta de Castilla y León (VA315P18), MINECO (AGL2017-87373-C3-2-R),
FEDER, and the Interreg España-Portugal Programme (Iberphenol) for the funding.

REFERENCES

Aleixandre-Tudo, J. L., Weightman, C., Panzeri, V., Nieuwoudt, H. H., and du Toit, W.
J. (2015). Effect of skin contact before and during alcoholic fermentation on the
chemical and sensory profile of South African Chenin blanc white wines. South
African Journal of Enology and Viticulture, 36(3), 366–77.
456 Food Aroma Evolution

Arias-Gil, M., Garde-Cerdán, T., and Ancín-Azpilicueta, C. (2007). Influence of addi-

tion of ammonium and different amino acid concentrations on nitrogen metabolism
in spontaneous must fermentation. Food Chemistry, 103(4), 1312–8.
Asproudi, A., Ferrandino, A., Bonello, F., Vaudano, E., Pollon, M., and Petrozziello, M.
(2018). Key norisoprenoid compounds in wines from early-harvested grapes in view
of climate change. Food Chemistry, 268, 143–52.
Ayestarán, B., Martínez-Lapuente, L., Guadalupe, Z., Canals, C., Adell, E., and Vilanova,
M. (2019). Effect of the winemaking process on the volatile composition and aro-
matic profile of Tempranillo Blanco wines. Food Chemistry, 276, 187–94.
Belda, I., Ruiz, J., Esteban-Fernández, A., Navascués, E., Marquina, D., Santos, A.,
and Moreno-Arribas, M. V. (2017). Microbial contribution to wine aroma and its
intended use for wine quality improvement. Molecules, 22(2), 189.
Black, C. A., Parker, M., Siebert, T. E., Capone, D. L., and Francis, I. L. (2015). Terpenoids
and their role in wine flavour: Recent advances. Australian Journal of Grape and
Wine Research, 21, 582–600.
Blanchard, L., Tominaga, T., and Dubourdieu, D. (2001). Formation of furfurylthiol
exhibiting a strong coffee aroma during oak barrel fermentation from furfural
released by toasted staves. Journal of Agricultural and Food Chemistry, 49(10),
4833–5.
Blanco, P., Mirás-Avalos, J. M., Pereira, E., and Orriols, I. (2013). Fermentative aroma
compounds and sensory profiles of godello and albariño wines as influenced by
Saccharomyces cerevisiae yeast strains. Journal of the Science of Food and
Agriculture, 93(11), 2849–57.
Boidron, J. N., Chatonnet, P., and Pons, M. (1988). Influence du bois sur certaines sub-
stances odorantes des vins. Connaissance Vigne Vin, 22(4), 275–94.
Boselli, E., di Lecce, G., Alberti, F., and Frega, N. G. (2010). Nitrogen gas affects the
quality and the phenolic profile of must obtained from vacuum-pressed white grapes.
LWT—Food Science and Technology, 43(10), 1494–500.
Bosso, A., Petrozziello, M., Santini, D., Motta, S., Guaita, M., and Marulli, C. (2008).
Effect of grain type and toasting conditions of barrels on the concentration of
the volatile substances released by the wood and on the sensory characteristics of
Montepulciano d’Abruzzo. Journal of Food Science, 73(7), 462–70.
Brown, R. C., Sefton, M. A., Taylor, D. K., and Elsey, G. M. (2006). An odour detection
threshold determination of all four possible stereoisomers of oak lactone in a white
and a red wine. Australian Journal of Grape and Wine Research, 12(2), 115–8.
Caillé, S., Samson, A., Wirth, J., Diéval, J. B., Vidal, S., and Cheynier, V. (2010). Sensory
characteristics changes of red Grenache wines submitted to different oxygen expo-
sures pre and post bottling. Analytica Chimica Acta, 660(1–2), 35–42.
Chatonnet, P., Dubourdieu, D., and Boidron, J. N. (1992). Incidence des conditions de
fermentation et d’élevage des vins blancs secs en barriques sur leur composition en
substances cédées par le bois de chêne. Sciences Des Aliments, 12(4), 665–85.
Chen, K., Escott, C., Loira, I., del Fresno, J. M., Morata, A., Tesfaye, W., Calderon, F.,
Benito, S., and Suárez-Lepe, J. A. (2016). The effects of pre-fermentative addition
of oenological tannins on wine components and sensorial qualities of red wine.
Molecules, 21(11), 1445.
Chen, K., Escott, C., Loira, I., del Fresno, J. M., Morata, A., Tesfaye, W., Calderon, F.,
Suárez-Lepe, J. A., Han, S., and Benito, S. (2018). Use of non-Saccharomyces yeasts
and oenological tannin in red winemaking: Influence on colour, aroma and senso-
rial properties of young wines. Food Microbiology, 69, 51–63.
 Grape and Wine Volatile Composition 457

Chen, W. K., Yu, K. J., Liu, B., Lan, Y. B., Sun, R. Z., Li, Q., He, F., Pan, Q., Duan,
C. Q., and Wang, J. (2017). Comparison of transcriptional expression patterns of
carotenoid metabolism in “Cabernet Sauvignon” grapes from two regions with dis-
tinct climate. Journal of Plant Physiology, 213, 75–86.
Chira, K. and Teissedre, P. L. (2013a). Extraction of oak volatiles and ellagitannins
compounds and sensory profile of wine aged with French winewoods subjected to
different toasting methods: Behaviour during storage. Food Chemistry, 140(1–2),
168–77.
Chira, K. and Teissedre, P. L. (2013b). Relation between volatile composition, ellagitannin
content and sensory perception of oak wood chips representing different toasting
processes. European Food Research and Technology, 236(4), 462–70.
Chira, K. and Teissedre, P. L. (2015). Chemical and sensory evaluation of wine matured
in oak barrel: Effect of oak species involved and toasting process. European Food
Research and Technology, 240(3), 533–47.
Collins, T. S., Miles, J. L., Boulton, R. B., and Ebeler, S. E. (2015). Targeted volatile
composition of oak wood samples taken during toasting at a commercial cooperage.
Tetrahedron, 71(20), 2971–82.
Crespo, J., Rigou, P., Romero, V., García, M., Arroyo, T., and Cabellos, J. M. (2018).
Effect of seasonal climate fluctuations on the evolution of glycoconjugates during
the ripening period of grapevine cv. Muscat à petits grains blancs berries. Journal of
the Science of Food and Agriculture, 98(5), 1803–12.
Darriet, P., Thibon, C., and Dubourdieu, D. (2012). Aroma and aroma precursors in
grape berry. In: H. Gerós, M. Cahves, and S. Delrot (Eds.), The Biochemistry of the
Grape Berry (pp. 111–36). Bentham Books.
de Castilhos, M. B. M., Del Bianchi, V. L., Gómez-Alonso, S., García-Romero, E., and
Hermosín-Gutiérrez, I. (2019). Sensory descriptive and comprehensive GC–MS
as suitable tools to characterize the effects of alternative winemaking procedures
on wine aroma. Part I: BRS Carmem and BRS Violeta. Food Chemistry, 272,
462–70.
del Álamo-Sanza, M., Cárcel, L. M., and Nevares, I. (2017). Characterization of the oxy-
gen transmission rate of oak wood species used in cooperage. Journal of Agricultural
and Food Chemistry, 65(3), 648–55.
del Álamo-Sanza, M., Laurie, V. F., and Nevares, I. (2015). Wine evolution and spatial
distribution of oxygen during storage in high-density polyethylene tanks. Journal of
the Science of Food and Agriculture, 95(6), 1313–20.
del Álamo-Sanza, M. and Nevares, I. (2014). Recent advances in the evaluation of the
oxygen transfer rate in oak barrels. Journal of Agricultural and Food Chemistry,
62(35), 8892–9.
del Álamo-Sanza, M. and Nevares, I. (2017). Oak wine barrel as an active vessel: A criti-
cal review of past and current knowledge. Critical Reviews in Food Science and
Nutrition, 0, 1–16.
del Álamo-Sanza, M., Nevares, I., Gallego, L., Fernández de Simón, B., and Cadahía,
E. (2010). Micro-oxygenation strategy depends on origin and size of oak chips
or staves during accelerated red wine aging. Analytica Chimica Acta, 660(1–2),
92–101.
del Álamo-Sanza, M., Nevares, I., Martínez-Gil, A., Rubio-Bretón, P., and Garde-
Cerdán, T. (2019). Impact of long bottle aging (10 years) on volatile composition
of red wines micro-oxygenated with oak alternatives. LWT—Food Science and
Technology, 101, 395–403.
458 Food Aroma Evolution

Dumitriu, G. D., López de Lerma, N., Zamfir, C. I., Cotea, V. V., and Peinado, R. A.
(2017). Volatile and phenolic composition of red wines subjected to aging in oak
cask of different toast degree during two periods of time. LWT—Food Science and
Technology, 86, 643–51.
Etievant, P. X. (1991). Wine. In H. Maarse (Ed.), Volatile Compounds in Foods and
Beverages (pp. 483–546). New York, NY: Marcell Dekker Inc.
Fairbairn, S., McKinnon, A., Musarurwa, H. T., Ferreira, A. C., and Bauer, F. F. (2017).
The impact of single amino acids on growth and volatile aroma production by
Saccharomyces cerevisiae strains. Frontiers in Microbiology, 8, 2554.
Fernández de Simón, B., Cadahía, E., Muiño, I., del Álamo-Sanza, M., and Nevares, I.
(2010). Volatile composition of toasted oak chips and staves and of red wine aged
with them. American Journal of Enology and Viticulture, 61(2), 157–65.
Fernández de Simón, B., Martínez, J., Sanz, M., Cadahía, E., Esteruelas, E., and Muñoz,
M. (2014). Volatile compounds and sensorial characterisation of red wine aged in
cherry, chestnut, false acacia, ash and oak wood barrels. Food Chemistry, 147,
346–56.
Ferreira, V., Bueno, M., Franco-Luesma, E., Culleré, L., and Fernández-Zurbano, P.
(2014). Key changes in wine aroma active compounds during bottle storage of
Spanish red wines under different oxygen levels. Journal of Agricultural and Food
Chemistry, 62(41), 10015–27.
Ferreira, V., López, R., and Cacho, J. (2000). Quantitative determination of the odorants
of young red wines from different grape varieties. Journal of the Science of Food
and Agriculture, 80(11), 1659–67.
Flanzy, C. (2000). Enología: Fundamentos científicos y tecnológicos. Madrid:
Mundi-Prensa.
Franco, M., Peinado, R. A., Medina, M., and Moreno, J. (2004). Off-vine grape drying
effect on volatile compounds and aromatic series in must from Pedro Ximénez grape
variety. Journal of Agricultural and Food Chemistry, 52(12), 3905–10.
Gallego, L., del Álamo-Sanza, M., Nevares, I., Fernández, J., Fernández de Simón,
B., and Cadahía, E. (2012). Phenolic compounds and sensorial characterization
of wines aged with alternative to barrel products made of Spanish oak wood
(Quercus pyrenaica Willd.). Food Science and Technology International, 18(2),
151–65.
Gambetta, J. M., Bastian, S. E. P., Cozzolino, D., and Jeffery, D. W. (2014). Factors influ-
encing the aroma composition of Chardonnay wines. Journal of Agricultural and
Food Chemistry, 62(28), 6512–34.
García-Esparza, M. J., Abrisqueta, I., Escriche, I., Intrigliolo, D. S., Álvarez, I., and
Lizama, V. (2018). Volatile compounds and phenolic composition of skins and seeds
of “Cabernet Sauvignon” grapes under different deficit irrigation regimes. Vitis—
Journal of Grapevine Research, 91(3), 83–91.
Garde-Cerdán, T. and Ancín-Azpilicueta, C. (2006a). Effect of oak barrel type on the
volatile composition of wine: Storage time optimization. LWT—Food Science and
Technology, 39(3), 199–205.
Garde-Cerdán, T. and Ancín-Azpilicueta, C. (2006b). Review of quality factors on wine
ageing in oak barrels. Trends in Food Science & Technology, 17(8), 438–47.
Garde-Cerdán, T. and Ancín-Azpilicueta, C. (2008). Effect of the addition of differ-
ent quantities of amino acids to nitrogen-deficient must on the formation of esters,
alcohols, and acids during wine alcoholic fermentation. LWT—Food Science and
Technology, 41(3), 501–10.
 Grape and Wine Volatile Composition 459

Garde-Cerdán, T., Rodrı ́guez-Mozaz, S., and Ancín-Azpilicueta, C. (2002). Volatile

composition of aged wine in used barrels of French oak and of American oak. Food
Research International, 35(7), 603–10.
Garde-Cerdán, T., Torrea-Goñi, D., and Ancín-Azpilicueta, C. (2004). Accumulation
of volatile compounds during aging of two red wines with different composition.
Journal of Food Engineering, 65(3), 349–56.
Ghaste, M., Narduzzi, L., Carlin, S., Vrhovsek, U., Shulaev, V., and Mattivi, F. (2015).
Chemical composition of volatile aroma metabolites and their glycosylated precur-
sors that can uniquely differentiate individual grape cultivars. Food Chemistry, 188,
309–19.
Gómez García-Carpintero, E., Gómez Gallego, M. A., Sánchez-Palomo, E., and González-
Viñas, M. A. (2012). Impact of alternative technique to ageing using oak chips in
alcoholic or in malolactic fermentation on volatile and sensory composition of red
wines. Food Chemistry, 134(2), 851–63.
Gómez-Plaza, E., Pérez-Prieto, L. J., Fernández-Fernández, J. I., and López-Roca, J.
M. (2004). The effect of successive uses of oak barrels on the extraction of oak-
related volatile compounds from wine. International Journal of Food Science and
Technology, 39(10), 1069–78.
González-Centeno, M. R., Chira, K., and Teissedre, P. L. (2016). Ellagitannin content,
volatile composition and sensory profile of wines from different countries matured in
oak barrels subjected to different toasting methods. Food Chemistry, 210, 500–11.
Guchu, E., Díaz-Maroto, M. C., Pérez-Coello, M. S., González-Viñas, M. A., and
Cabezudo, M. D. (2006). Volatile composition and sensory characteristics of
Chardonnay wines treated with American and Hungarian oak chips. Food
Chemistry, 99(2), 350–9.
Guth, H. (1997). Quantitation and sensory studies of character impact odorants of dif-
ferent white wine varieties. Journal of Agricultural and Food Chemistry, 45(8),
3027–32.
Gutiérrez-Gamboa, G., Garde-Cerdán, T., Portu, J., Moreno-Simunovic, Y., and
Martínez-Gil, A. M. (2017). Foliar nitrogen application in Cabernet Sauvignon vines:
Effects on wine flavonoid and amino acid content. Food Research International, 96,
46–53.
Hayasaka, Y., Parker, M., Baldock, G. A., Pardon, K. H., Black, C. A., Jeffery, D. W.,
and Herderich, M. J. (2013). Assessing the impact of smoke exposure in grapes:
Development and validation of a HPLC-MS/MS method for the quantitative analy-
sis of smoke-derived phenolic glycosides in grapes and wine. Journal of Agricultural
and Food Chemistry, 61(1), 25–33.
Helwi, P., Thibon, C., Habran, A., Hilbert, G., Guillaumie, S., Delrot, S., Darriet, P.,
and Van Leeuwen, C. (2015). Effect of vine nitrogen status, grapevine variety and
rootstock on the levels of berry S-glutathionylated and S-cysteinylated precursors
of 3-sulfanylhexan-1-ol. Journal International des Sciences de La Vigne et du Vin,
49(4), 253–65.
Herbst-Johnstone, M., Nicolau, L., and Kilmartin, P. A. (2011). Stability of varietal
thiols in commercial sauvignon blanc wines. American Journal of Enology and
Viticulture, 62(4), 495–502.
Hernández-Orte, P., Concejero, B., Astrain, J., Lacau, B., Cacho, J., and Ferreira, V.
(2015). Influence of viticulture practices on grape aroma precursors and their
relation with wine aroma. Journal of the Science of Food and Agriculture, 95(4),
688–701.
460 Food Aroma Evolution

Herrero, P., Sáenz-Navajas, M. P., Avizcuri, J. M., Culleré, L., Balda, P., Antón, E. C.,
Ferreira, V., and Escudero, A. (2016). Study of Chardonnay and Sauvignon blanc
wines from D.O.Ca Rioja (Spain) aged in different French oak wood barrels:
Chemical and aroma quality aspects. Food Research International, 89, 227–36.
Hjelmeland, A. K. and Ebeler, S. E. (2015). Glycosidically bound volatile aroma com-
pounds in grapes and wine: A review. American Journal of Enology and Viticulture,
66(1), 1–11.
Jarauta, I., Cacho, J., and Ferreira, V. (2005). Concurrent phenomena contributing to
the formation of the aroma of wine during aging in oak wood: An analytical study.
Journal of Agricultural and Food Chemistry, 53(10), 4166–77.
Jiménez-Moreno, N. and Ancín-Azpilicueta, C. (2007). Binding of oak volatile com-
pounds by wine lees during simulation of wine ageing. LWT—Food Science and
Technology, 40(4), 619–24.
Ju, Y. L., Xu, G. Q., Yue, X. F., Zhao, X. F., Tu, T. Y., Zhang, J. X., and Fang, Y.
L. (2018). Effects of regulated deficit irrigation on amino acid profiles and their
derived volatile compounds in Cabernet Sauvignon (Vitis vinifera L.) grapes and
wines. Molecules, 23(8), 1983.
Kalua, C. M. and Boss, P. K. (2010). Comparison of major volatile compounds from
Riesling and Cabernet Sauvignon grapes (Vitis vinifera L.) from fruitset to harvest.
Australian Journal of Grape and Wine Research, 16(2), 337–48.
Kyraleou, M., Kallithraka, S., Chira, K., Tzanakouli, E., Ligas, I., and Kotseridis, Y.
(2015). Differentiation of wines treated with wood chips based on their phenolic
content, volatile composition, and sensory parameters. Journal of Food Science,
80(12), C2701–10.
Kyraleou, M., Teissedre, P. L., Tzanakouli, E., Kotseridis, Y., Proxenia, N., Chira, K.,
Ligas, I., and Kallithraka, S. (2016). Addition of wood chips in red wine during and
after alcoholic fermentation: Differences in color parameters, phenolic content and
volatile composition. Journal International des Sciences de La Vigne et du Vin,
50(4), 209–22.
Liberatore, M. T., Pati, S., Nobile, M. A. D., and Notte, E. L. (2010). Aroma quality
improvement of Chardonnay white wine by fermentation and ageing in barrique on
lees. Food Research International, 43(4), 996–1002.
Liu, J., Zhu, X. L., Ullah, N., and Tao, Y. S. (2017). Aroma glycosides in grapes and
wine. Journal of Food Science, 82(2), 248–59.
Loira, I., Vejarano, R., Morata, A., Ricardo-da-Silva, J. M., Laureano, O., González, M.
C., and Suárez-Lepe, J. A. (2013). Effect of Saccharomyces strains on the quality of
red wines aged on lees. Food Chemistry, 139(1–4), 1044–51.
Lopes, P., Silva, M. A., Pons, A., Tominaga, T., Lavigne, V., Saucier, C., Darriet, P.,
Teissedre, P. L., and Dubourdieu, D. (2009). Impact of oxygen dissolved at bot-
tling and transmitted through closures on the composition and sensory properties
of a Sauvignon blanc wine during bottle storage. Journal of Agricultural and Food
Chemistry, 57(21), 10261–70.
Lukić, I., Budić-Leto, I., Bubola, M., Damijanić, K., and Staver, M. (2017). Pre-
fermentative cold maceration, saignée, and various thermal treatments as options
for modulating volatile aroma and phenol profiles of red wine. Food Chemistry,
224, 251–61.
Ma, D., Yan, X., Wang, Q., Zhang, Y., and Tao, Y. (2017). Performance of selected P.
fermentans and its excellular enzyme in co-inoculation with S. cerevisiae for wine
aroma enhancement. LWT—Food Science and Technology, 86, 361–70.
 Grape and Wine Volatile Composition 461

Martínez, J., Cadahía, E., Fernández de Simón, B., Ojeda, S., and Rubio, P. (2008). Effect
of the seasoning method on the chemical composition of oak heartwood to cooper-
age. Journal of Agricultural and Food Chemistry, 56, 3089–96.
Martínez-Gil, A. M., del Álamo-Sanza, M., Gutiérrez-Gamboa, G., Moreno-Simunovic,
Y., and Nevares, I. (2018). Volatile composition and sensory characteristics of
Carménère wines macerating with Colombian (Quercus humboldtii) oak chips
compared to wines macerated with American (Q. alba) and European (Q. petraea)
oak chips. Food Chemistry, 266, 90–100.
Mattivi, F., Caputi, L., Carlin, S., Lanza, T., Minozzi, M., Nanni, D., Valenti, L., and
Vrhovsek, U. (2011). Effective analysis of rotundone at below-threshold levels in
red and white wines using solid-phase microextraction gas chromatography/tandem
mass spectrometry. Rapid Communications in Mass Spectrometry, 25(4), 483–8.
Mendez-Costabel, M. P., Wilkinson, K. L., Bastian, S. E. P., Jordans, C., Mccarthy, M., Ford,
C. M., and Dokoozlian, N. K. (2014). Effect of increased irrigation and additional nitro-
gen fertilisation on the concentration of green aroma compounds in Vitis vinifera L.
Merlot fruit and wine. Australian Journal of Grape and Wine Research, 20(1), 80–90.
Mihnea, M., González-SanJosé, M. L., Ortega-Heras, M., and Pérez-Magariño, S. (2015).
A comparative study of the volatile content of Mencía wines obtained using differ-
ent pre-fermentative maceration techniques. LWT—Food Science and Technology,
64(1), 32–41.
Molina, A. M., Swiegers, J. H., Varela, C., Pretorius, I. S., and Agosin, E. (2007). Influence
of wine fermentation temperature on the synthesis of yeast-derived volatile aroma
compounds. Applied Microbiology and Biotechnology, 77(3), 675–87.
Navarro, M., Kontoudakis, N., Gómez-Alonso, S., García-Romero, E., Canals, J. M.,
Hermosín-Gutiérrez, I., and Zamora, F. (2018). Influence of the volatile substances
released by oak barrels into a Cabernet Sauvignon red wine and a discolored
Macabeo white wine on sensory appreciation by a trained panel. European Food
Research and Technology, 244(2), 245–58.
Nevares, I. and del Álamo-Sanza, M. (2014). Oxygène et barriques: Actualisation des
connaissances. Quantité et voies de pénétration de l’oxygène dans la barrique.
Revue des Oenologues et des Techniques Vitinicoles et Oenologicques: Magazine
Trimestriel d’information Professionnelle, 41(153), 41–4.
Nevares, I. and del Álamo-Sanza, M. (2018). New materials for the aging of wines and
beverages: Evaluation and comparison. In Food Packaging and Preservation (Vol. 9,
pp. 375–407). Academic Press.
Nevares, I., Mayr, T., Baro, J. A., Ehgartner, J., Crespo, R., and del Álamo-Sanza, M.
(2016). Ratiometric oxygen imaging to predict oxygen diffusivity in oak wood dur-
ing red wine barrel aging. Food and Bioprocess Technology, 9(6), 1049–59.
Oberholster, A., Elmendorf, B. L., Lerno, L. A., King, E. S., Heymann, H., Brenneman,
C. E., and Boulton, R. B. (2015). Barrel maturation, oak alternatives and micro-oxy-
genation: Influence on red wine aging and quality. Food Chemistry, 173, 1250–8.
Pardo-García, A. I., Wilkinson, K. L., Culbert, J. A., Lloyd, N. D. R., Alonso, G. L., and
Salinas, M. R. (2015). Accumulation of glycoconjugates of 3-methyl-4-hydroxyoc-
tanoic acid in fruits, leaves, and shoots of Vitis vinifera cv. Monastrell following
foliar applications of oak extract or oak lactone. Journal of Agricultural and Food
Chemistry, 63(18), 4533–8.
Prida, A. and Chatonnet, P. (2010). Impact of oak-derived compounds on the olfactory
perception of barrel-aged wines. American Journal of Enology and Viticulture,
61(3), 408–13.
462 Food Aroma Evolution

Puertas, B., Jiménez-Hierro, M. J., Cantos-Villar, E., Marrufo-Curtido, A., Carbú, M.,
Cuevas, F. J., Moreno-Rojas, J. M., González-Rodríguez, V. E., Cantoral, J. M.,
and Ruiz-Moreno, M. J. (2018). The influence of yeast on chemical composition and
sensory properties of dry white wines. Food Chemistry, 253, 227–35.
Ramírez, M., Velázquez, R., Maqueda, M., Zamora, E., López-Piñeiro, A., and
Hernández, L. M. (2016). Influence of the dominance of must fermentation by
Torulaspora delbrueckii on the malolactic fermentation and organoleptic quality of
red table wine. International Journal of Food Microbiology, 238, 311–9.
Robinson, A. L., Boss, P. K., Solomon, P. S., Trengove, R. D., Heymann, H., and Ebeler,
S. E. (2014). Origins of grape and wine aroma. Part 2. Chemical and sensory analy-
sis. American Journal of Enology and Viticulture, 65(1), 25–42.
Rodríguez-Rodríguez, P. and Gómez-Plaza, E. (2012). Dependence of oak-related vola-
tile compounds on the physicochemical characteristics of barrel-aged wines. Food
Technology and Biotechnology, 50(1), 59–65.
Rollero, S., Bloem, A., Camarasa, C., Sánchez, I., Ortiz-Julien, A., Sablayrolles, J. M.,
Dequin, S., and Mouret, J. R. (2015). Combined effects of nutrients and tempera-
ture on the production of fermentative aromas by Saccharomyces cerevisiae during
wine fermentation. Applied Microbiology and Biotechnology, 99(5), 2291–304.
Rollero, S., Roberts, S., Bauer, F. F., and Divol, B. (2018). Agitation impacts fermen-
tation performance as well as carbon and nitrogen metabolism in Saccharomyces
cerevisiae under winemaking conditions. Australian Journal of Grape and Wine
Research, 24(3), 360–7.
Rubio-Bretón, P., Gutiérrez-Gamboa, G., Pérez-Álvarez, E. P., Santamaría, P., and
Garde-Cerdán, T. (2018). Foliar application of several nitrogen sources as fertilis-
ers to Tempranillo grapevines: Effect on wine volatile composition. South African
Journal of Enology and Viticulture, 39(2), 235–45.
Sáenz-Navajas, M. P., Avizcuri, J. M., Ballester, J., Fernández-Zurbano, P., Ferreira, V.,
Peyron, D., and Valentin, D. (2015). Sensory-active compounds influencing wine
experts’ and consumers’ perception of red wine intrinsic quality. LWT—Food
Science and Technology, 60(1), 400–11.
Sáenz-Navajas, M. P., Campo, E., Fernández-Zurbano, P., Valentin, D., and Ferreira, V.
(2010). An assessment of the effects of wine volatiles on the perception of taste and
astringency in wine. Food Chemistry, 121(4), 1139–49.
Salinas, M. R., Serrano de la Hoz, K., Zalacain, A., Lara, J. F., and Garde-Cerdán, T.
(2012). Analysis of red grape glycosidic aroma precursors by glycosyl glucose quan-
tification. Talanta, 89, 396–400.
Sánchez-Gómez, R., Garde-Cerdán, T., Zalacain, A., García, R., Cabrita, M. J., and
Salinas, M. R. (2016). Vine-shoot waste aqueous extract applied as foliar fertil-
izer to grapevines: Effect on amino acids and fermentative volatile content. Food
Chemistry, 197, 132–40.
Serrano de la Hoz, K., Carmona, M., Zalacain, A., Alonso, G., and Salinas, M. (2014).
The varietal aroma potential index (IPAv): A tool to evaluate the quality of grape
and wines, white and red. In 37th OIV Congress.
Sidhu, D., Lund, J., Kotseridis, Y., and Saucier, C. (2015). Methoxypyrazine analysis and
influence of viticultural and enological procedures on their levels in grapes, musts,
and wines. Critical Reviews in Food Science and Nutrition, 55(4), 485–502.
Slaghenaufi, D. and Ugliano, M. (2018). Norisoprenoids, sesquiterpenes and terpenoids con-
tent of Valpolicella wines during aging: Investigating aroma potential in relationship to
evolution of tobacco and balsamic aroma in aged wine. Frontiers in Chemistry, 6, 1–13.
 Grape and Wine Volatile Composition 463

Spillman, P. J., Sefton, M. A., and Gawel, R. (2004a). The contribution of volatile com-
pounds derived during oak barrel maturation to the aroma of a Chardonnay and
Cabernet Sauvignon wine. Australian Journal of Grape and Wine Research, 10(3),
227–35.
Spillman, P. J., Sefton, M. A., and Gawel, R. (2004b). The effect of oak wood source,
location of seasoning and coopering on the composition of volatile compounds in
oak-matured wines. Australian Journal of Grape and Wine Research, 3, 216–26.
Styger, G., Prior, B., and Bauer, F. F. (2011). Wine flavor and aroma. Journal of Industrial
Microbiology and Biotechnology, 38(9), 1145–59.
Ugliano, M. (2013). Oxygen contribution to wine aroma evolution during bottle aging.
Journal of Agricultural and Food Chemistry, 61(26), 6125–36.
Ugliano, M., Diéval, J. B., Siebert, T. E., Kwiatkowski, M., Aagaard, O., Vidal, S., and
Waters, E. J. (2012). Oxygen consumption and development of volatile sulfur com-
pounds during bottle aging of two Shiraz wines. Influence of pre- and postbottling con-
trolled oxygen exposure. Journal of Agricultural and Food Chemistry, 60(35), 8561–70.
Varela, C., Barker, A., Tran, T., Borneman, A., and Curtin, C. (2017). Sensory profile
and volatile aroma composition of reduced alcohol Merlot wines fermented with
Metschnikowia pulcherrima and Saccharomyces uvarum. International Journal of
Food Microbiology, 252, 1–9.
Varela, C., Torrea, D., Schmidt, S. A., Ancín-Azpilicueta, C., and Henschke, P. A. (2012).
Effect of oxygen and lipid supplementation on the volatile composition of chemi-
cally defined medium and Chardonnay wine fermented with Saccharomyces cerevi-
siae. Food Chemistry, 135(4), 2863–71.
Vasserot, Y., Jacopin, C., and Jeandet, P. (2001). Effect of bottle capacity and bottle-cap
permeability to oxygen on dimethylsulfide formation in champagne wines during
aging on the lees. American Journal of Enology and Viticulture, 52(1), 54–5.
Vichi, S., Santini, C., Natali, N., Riponi, C., López-Tamames, E., and Buxaderas, S.
(2007). Volatile and semi-volatile components of oak wood chips analysed by
Accelerated Solvent Extraction (ASE) coupled to Gas Chromatography-Mass
Spectrometry (GC–MS). Food Chemistry, 102(4), 1260–9.
Vilanova, M., Genisheva, Z., Tubio, M., Álvarez, K., Lissarrague, J. R., and Oliveira,
J. M. (2017). Effect of vertical shoot-positioned, Scott-Henry, Geneva double-cur-
tain, arch-cane, and parral training systems on the volatile composition of Albariño
wines. Molecules, 22(9), 1500.
Wang, Y., He, L., Pan, Q., Duan, C., and Wang, J. (2018). Effects of basal defoliation on
wine aromas: A meta-analysis. Molecules, 23(4), 779.
Ward, S. C., Petrie, P. R., Johnson, T. E., Boss, P. K., and Bastian, S. E. P. (2015). Unripe
berries and petioles in Vitis vinifera cv. Cabernet Sauvignon fermentations affect
sensory and chemical profiles. American Journal of Enology and Viticulture, 66(4),
435–43.
Williams, P. J., Cynkar, W., Francis, I. L., Gray, J. D., Iland, P. G., and Coombe, B. G.
(1995). Quantification of glycosides in grapes, juices, and wines through a deter-
mination of glycosyl glucose. Journal of Agricultural and Food Chemistry, 43(1),
121–8.
Wood, C., Siebert, T. E., Parker, M., Capone, D. L., Elsey, G. M., Pollnitz, A. P., Eggers,
M., Meier, M., Vössing, T., Widder, S., Krammer, G., Sefton, M. A., and Herderich,
M. J. (2008). From wine to pepper: Rotundone, an obscure sesquiterpene, is a
potent spicy aroma compound. Journal of Agricultural and Food Chemistry, 56(10),
3738–44.
 Chapter 23
Milk/Dairy
Kieran Kilcawley

CONTENTS

23.1 Introduction 465

23.2 Sensory Analysis 466
23.2.1 Affective Studies 466
23.2.2 Difference Testing 466
23.2.3 Descriptive Methods 467
23.3 Flavor Chemistry Methodologies 467
23.3.1 Solvent-Assisted Flavor Extraction 468
23.3.2 Headspace Solid-Phase Microextraction 468
23.3.3 Thermal Desorption 468
23.4 Milk 469
23.4.1 Bovine Milk 469
23.4.2 Ovine and Caprine Milk 470
23.4.3 Cheese and Yogurt 470
23.4.3.1 Glycolysis 471
23.4.3.2 Lipolysis 471
23.4.3.3 Proteolysis 472
23.5 Main Volatile Compounds in Milk, Cheese, and Yogurt 472
23.5.1 Acids 472
23.5.2 Aldehydes 473
23.5.3 Ketones 475
23.5.4 Alcohols 476
23.5.5 Esters 477
23.5.6 Sulfur Compounds 477
23.5.7 Lactones 479
23.5.8 Miscellaneous Volatile Compounds 479
References 480

23.1 INTRODUCTION

The aroma of milk and dairy products is related to the presence of volatile compounds
influencing sensory perception. Volatile compounds in milk can be affected by a wide
range of factors from the animal (breed, health status, age), diet, on-farm practices, post-
farm storage, and processing. The aroma of dairy products produced from milk is also
dependent upon milk quality (microbial load, storage conditions), processing conditions,
subsequent fermentation, and storage. In general terms, it is estimated that flavor consists

465
466 Food Aroma Evolution

of approximately 80% aroma and 20% taste (Spence, 2015) and this is mainly based on
the fact that many more aromatic compounds exist than nonvolatile tastants, rather than
a definitive science. However, it is estimated that only 3–5% of aroma compounds pres-
ent in food actually contribute to sensory perception, as the abundance of most are below
their odor thresholds (OTs) (Dunkel et al., 2014). Even though a considerable amount of
research has been undertaken on the flavor of milk and dairy products, it is surprising
how little information exists on the key character aromatic compounds that influence the
sensory perception of milk and many specific dairy products. This is mainly due to the
presence of significant amounts of potential odor-active volatiles that differ in molecular
weight, polarity, vapor pressure and stability; the difficulty in extracting, concentrat-
ing, and analyzing them; and the inherent variability of milk and dairy products. The
analytical route of choice for volatile compounds has been gas chromatography–mass
spectrometry (GC-MS). In order to best determine aroma active compounds influencing
sensory perception, a chemometric approach is useful for finding associations between
volatile and sensory data.

23.2 SENSORY ANALYSIS

Different techniques exist to evaluate the sensory properties of food, and these can be
roughly separated into three categories (a) affective, (b) difference, and (c) descriptive
techniques (O’Sullivan, 2017). The choice is very important, and a range of different
techniques have been used to evaluate milk and dairy products (Drake, 2007; Schiano
et al., 2017; Kilcawley et al., 2018).

23.2.1 Affective Studies

Affective studies use hedonics (liking) to capture responses of either consumers or asses-
sors. Consumers/assessors score on their preference or liking of attributes such as, “appear-
ance,” “flavor,” or “texture,” and ultimately their “overall acceptability” of a product
(O’Sullivan, 2017). Ideally, a minimum of 50 consumers/assessors is required to provide
meaningful conclusions in part because consumers are highly variable (Drake, 2007). It
has also been noted that in some sensory studies of milk, differences identified by descrip-
tive sensory analysis were not perceived by consumers using affective studies (Khanal et al.,
2005; Croissant et al., 2007). This is not surprising in that highly trained panelists were
indirectly compared to untrained consumers. The use of affective sensory data with aroma
chemical data is of limited use; however, the incorporation of hedonic data with descriptive
sensory data can provide useful information in relation to “likeness” and “acceptability.”

23.2.2 Difference Testing

Difference methods are the most basic form of sensory testing. Products are compared
directly, and assessors are asked to determine if they are the same or different (O’Sullivan,
2017). The number of panelists required varies depending upon the test chosen, but gen-
erally 25 to 50 is acceptable (Drake, 2007). Dubroeucq et al. (2002) used triangle tests to
discern differences in raw bovine milk produced by different diets and found that panel-
ists were able to discern milk from some different diets. Bovolenta et al. (2009) also used
 Milk/Dairy 467

a triangle test to evaluate raw bovine milk from cows on different diets but did not find
any significant difference. The use of difference sensory testing with flavor chemical data
alone is of limited value.

23.2.3 Descriptive Methods

Descriptive methods involve the training of panelists to quantitatively determine the sen-
sory attributes in a sample or more usually a selection of samples (O’Sullivan, 2017).
Initially, panelists use focus groups to determine a list of specific attributes/lexicons that
best represent the sensory properties of the sample. These are fine-tuned and evaluated
over subsequent sessions. The assessors are trained to measure attributes associated with
“appearance,” “aroma,” “flavor,” “texture,” “taste,” and “aftertaste.” Descriptive analy-
sis allows screened individuals to be trained to give a more standardized quantitative
sensory response. A range of different methods for descriptive profiling exist (O’Sullivan,
2017). Quite a lot of research has been undertaken on milk and cheese using descriptive
analysis as it is easier to look for correlations between specific attributes/lexicons and fla-
vor chemical data. For example, Villeneuve et al. (2013), used this approach to determine
that diet influenced the sensory perception of pasteurized bovine milk, as did Bonanno
et al. (2013) to assess the impact of farming practices and cheesemaking processes on
the flavor of Caciocavallo cheese. As mentioned earlier, it can be useful to include affec-
tive, sensory assessment with descriptive and flavor chemical data as you can gain some
insight into the consumer acceptability of your product.

23.3 FLAVOR CHEMISTRY METHODOLOGIES

The most widely used approach to identify aromatic volatiles in dairy products is by
GC-MS. GC is used because of the volatile nature of the aromatic compounds of interest
and MS for identification purposes. Most GC-MS systems use quadrupole MS, and these
can be run in full scan mode for untargeted analyses to find all the volatiles present or in
selected ion monitoring for target analysis (knowns), specific compounds.
A key aspect in relation to volatile profiling by GC-MS is the polarity of the GC
column and choice of volatile extraction technique. Polar, nonpolar, and mid polar GC
column phases exist, and the choice will mainly depend upon the volatile compounds of
interest and column stability. Key factors that need to be taken into account in relation
to the choice of the extraction technique are the effectiveness of concentration, practi-
cality, automation, and flexibility. Different automated extraction platforms are avail-
able with multiple extractions and concentration/enrichment options in one system. This
capability/flexibility is important as to date no single extraction technique can provide a
complete volatile profile as they all confer a degree of bias. Most techniques for volatile
analysis by GC-MS are headspace techniques, possibly because of the ease of automation,
but also due to the fact that there is no need to undertake complex sample preparation
or use solvents in the extraction process. Some headspace techniques can be dynamic
where an inert gas is flushed through the sample to aid recovery of the volatiles of inter-
est. Direct immersive extraction and solvent extraction techniques are also widely used.
Figure 23.1 highlights the typical procedure for extracting volatiles from cheese and its
subsequent analysis by GCMS. Some of the more useful and widely used extraction/
concentration techniques to identify aromatic volatiles in cheese were recently reviewed
468 Food Aroma Evolution

FIGURE 23.1 Typical procedure for the extraction/concentration of volatiles from cheese
and subsequent analysis by GCMS.

(Bertuzzi et al., 2018). Three of the most common current methods used in aromatic
dairy research are solvent-assisted flavor evaporation (SAFE), headspace solid-phase
microextraction (HS-SPME), and thermal desorption (TD).

23.3.1 Solvent-Assisted Flavor Extraction

SAFE is an enclosed distillation extraction technique under vacuum where the extract is
cryogenically cooled in liquid nitrogen (Kilcawley, 2017). The extract can be concentrated
by purging with an inert gas such as nitrogen and then injected into the GC. An advantage
is that is relatively cost-effective, but a little impractical and is thought to be less effective for
highly volatile compounds (Bertuzzi et al., 2018).

23.3.2 Headspace Solid-Phase Microextraction

HS-SPME is easily the most widely used technique for aromatic research, which is in part
due to the length of time the technique has been available (Pawliszyn, 1997), but also mainly
because of its ease of use (automation) and the wide choice of available sorption phases. The
principal of the technique involves exposing a fiber on the end of a needle which is coated
in sorption phase(s) to the heated headspace of a sample in an enclosed vial. HS-SPME is
a passive technique and the sample size to headspace volume in the vial, equilibration time
and temperature, extraction time and temperature, and rate of agitation can all impact on
the efficiency of the process. It is best to optimize these conditions and the fiber for each
sample type (Mondello et al., 2005). The fiber is subsequently desorbed at high temperature
in the GC inlet and the fiber is reconditioned before the next sample. A major drawback of
HS-SPME is the volume of sorption phase on the fiber, which, depending upon the nature
and concentration of the volatiles present in your sample, can result in volatile discrimina-
tion based on affinity to the fiber and volatility. However, a new version, SPME Arrow, is
thought to overcome this issue by significantly increasing the volume of the sorption phase.

23.3.3 Thermal Desorption

TD is a dynamic technique where an inert gas is purged through the sample at a specific
temperature. The volatiles in the gas phase are passed through a sorption phase inside a
 Milk/Dairy 469

packed tube. The tube is subsequently desorbed, and all the volatiles are passed directly
into the GC via a transfer line. A range of different options are available to concentrate
or enrich volatiles from single or multiple samples to specific sorbent traps, where the
extract can be preserved or recollected depending upon choice. The main advantages are
the wide range and volume of sorption phases available; the high degree of control in rela-
tion to temperatures, gas pressures, and split flows used to concentrate volatiles; and the
high degree of automation. Sorbent tubes can also be reconditioned for multiple usages.
Potential disadvantages are that moisture removal can be problematic in enclosed systems
and can potentially damage columns, flow controllers, and MS detectors. Yuceer et al.
(2009) used TD to determine aroma-active volatiles in Ezine cheese by olfactometry.

23.4 MILK

The flavor of bovine, caprine, and ovine milk is influenced by a range of animal (breed,
genetics, health status, and stage of lactation), dietary (type of forage, forage conserva-
tion methods/practices, grassland management, and type of concentrate), and process-
ing (heat treatment, homogenization, storage time, agitation, etc.) factors. According to
Badings (1991) there are three basic elements responsible for the sensory properties of
milk: (a) pleasant mouth-feel due to presence of macromolecules such as colloidal pro-
teins and fat globules, (b) sweet and salty taste due to lactose and milk salts, respectively,
and (c) a weak and delicate aroma profile due to the correct balance of numerous volatile
constituents found in low abundance. These elements, along with the color and appear-
ance, are the main determinants of the sensory quality of milk (Cadwallader, 2010). Heat
treatment of milk also influences flavor in proportion to the extent of heat treatment,
with high heat treatments influencing both flavor and color (Singh et al., 2007).
Badings and Neeter (1980) suggested that the aroma of milk is determined by many
volatile compounds sometimes present in very low concentrations, some transferred from
the feed, others the result of minor conversions of milk constituents by chemical (oxida-
tive or thermal), microbial, and enzymatic reactions. Volatile compounds in forage and
feed enter milk through two routes; the main route is absorbed in the digestive tract,
that is, the rumen and/or intestine, before diffusing into the blood and then reaching the
mammary gland. The second route is the pulmonary route, where volatiles diffuse into
the air and are inhaled, absorbed into the lungs, enter the bloodstream, and subsequently
diffuse into the mammary gland (Viallon et al., 2000).

23.4.1 Bovine Milk

Bovine milk composition and flavor variations have been attributed to feed, geographi-
cal location, seasonal variation, and animal breed (Bendall, 2001; Bugaud et al., 2001;
Croissant et al., 2007; Coppa et al., 2011; Larsen et al., 2013; Villeneuve et al., 2013;
Faulkner et al., 2018). Many volatile aromatic compounds are present but typically at
concentrations below their OT (Marsili, 2003). This makes it difficult to determine those
volatiles that influence sensory perception. In addition, the extraction/concentration
technique, polarity of the GC column and sensitivity of the detector must be taken into
consideration as these will both positively and negatively discriminate against certain
individual volatiles and volatile chemical classes. Thus, the potential exists to over or
underestimate the relative importance of certain volatile compounds.
470 Food Aroma Evolution

The following aldehydes (acetaldehyde, heptanal, (z)-4-heptenal, nonanal, and

octanal), esters (ethyl acetate, ethyl butanoate, ethyl hexanoate, ethyl isovalerate, and
ethyl octanoate), ketones (acetoin, acetone, 2-butanone, and diacetyl), acids (butanoic
acid and octanoic acid), alcohols (ethanol and 1-octen-3-ol), sulfur (dimethyl sulfone),
and miscellaneous (indole) volatile compounds were postulated as potentially important
volatiles in bovine milk (Moio et al., 1993; Bendall and Olney, 2001; Toso et al., 2002;
Ai et al., 2015). A key aspect in relation to the significance of each volatile is its OT and
level of abundance. Other studies have shown a clear influence of diet on the volatile
profile and sensory properties of bovine milk. Faulkner et al. (2018) identified p-cresol as
an important volatile derived from pasture feed milk that potentially influences sensory
perception. These authors also found that acetic acid, hexanal, 2-butanone, 1-pentanol,
dimethyl sulfone, and toluene were impacted by diet in raw bovine milk. Coppa et al.
(2011) found that benzeneacetaldehdye, dimethyl sulfone, 4-hydroxy-4-methyl 2-penta-
none (diacetyl alcohol), octanoic acid, 1-octadecene, skatole, toluene, and undecanoic
acid in bovine milk were also influenced by diet. Croissant et al. (2007) found differences
in the volatile profiles of milk from total mixed ration and pasture diets, with the greatest
differences associated with acetic acid, 2-butanone, dimethyl sulfide, 2-furanmethanol,
2-heptanone, hexanoic acid, maltol, octanoic acid, and toluene. Villeneuve et al. (2013)
also found that acetone, 2-butanone, dimethyl sulfone, γ-lactones, pentanal, 1-pentanol,
α-pinene, and toluene differed in milk where cows were fed hay or pasture or silage, but
this did not impact sensory perception. A detailed list of potentially important volatiles
derived from diet in bovine milk was provided by Kilcawley et al. (2018), including infor-
mation on source and odor descriptors.

23.4.2 Ovine and Caprine Milk

Caprine and ovine milk differ considerably to bovine milk, mainly in relation to pH,
color, and sensory characteristics (Jandal, 1996). From a sensory perspective, the main
difference in caprine and ovine milk compared to bovine milk is the fatty acid profile.
The short-chain water-soluble fatty acids, heptanoic, octanoic, decanoic, and dodecanoic
are significantly more abundant in ovine and caprine milk (Park et al., 2007) and in the
free form are highly odor active. Other very characteristic volatiles particularly associ-
ated with ovine milk are the 4-alkyl-branched-chain fatty acids, which are products of
lipolysis and are thus higher in cheese than in milk (Kaffarnik et al., 2014). The two most
important are 4-methyl octanoic acid and 4-ethyl octanoic acid, which are very odor
active and have a goaty aroma.

23.4.3 Cheese and Yogurt

The diversity within cheese in relation to sensory characteristics is significant, with an

estimated 500 or more varieties commercially available. It is well established that three
main biochemical pathways; glycolysis, lipolysis, and proteolysis influence cheese flavor
development and in general, the extent of each is characteristic of the individual cheese
variety. Mulder (1952) postulated, the “component balance theory,” that cheese flavor
is the result of the correct balance and concentration of a wide variety of volatile flavor
compounds. This statement remains valid, except that it relates only to the aroma, as
nonvolatile compounds also impact on taste and thus have an important role (Kilcawley
 Milk/Dairy 471

and O’Sullivan, 2018). The aroma of cheese is impacted by potentially hundreds of vola-
tile compounds that have been identified in different cheese varieties; however, those
influencing sensory perception must be present at a concentration above their OT. Thus,
in most cases, the vast majority of volatile compounds present do not impact sensory
perception. The source of most volatile compounds in cheese are the result of the activ-
ity of microbial populations added during the cheesemaking procedure, either as starter
cultures, adjunct cultures, or as secondary cultures for surface or mold-ripened cheeses,
advantageous non-starter lactic acid bacteria or resulting from lipolysis from indigenous
milk lipases (lipoprotein lipase) or added animal lipases/esterases. Some volatile com-
pounds in the milk from diet can also influence the volatile profile of cheese but are less
significant for longer ripened cheeses where microbial activity tends to dictate the volatile
profile. An exception may be for cheeses produced from milk where animals have been
fed wild pasture containing dicotyledon plants, and thus significant concentrations of
terpenes may transfer into the cheese and potentially impact on the sensory properties of
the cheese (Kilcawley et al., 2018; Basdagianni et al., 2019). The same applies to yogurt
except that the starter cultures added are the main driving force of flavor development in
natural yogurts due to the optimized processing conditions for carbohydrate fermenta-
tion. Yogurts derived from ovine and caprine milk have a characteristic aroma mainly
related to the unique ratio of free fatty acids present in these milk. The pH of the final
product also has a major impact on odor perception as it can influence the odor activity
of some cheese volatiles (Kilcawley and O’Sullivan, 2018).

23.4.3.1 Glycolysis
In glycolysis, starter cultures readily metabolize lactose as it has a higher energy value
than other substrates present and thus aids their growth and survival during cheese
production. Glycolysis is important in cheese flavor development and is the main flavor
generation pathway in yogurt. In most cheese varieties lactose is utilized quite quickly
(McSweeney, 2004). Citrate levels are relatively low in milk, but it can also be a sub-
strate for key volatiles for certain cheese varieties and especially yogurt. The main
aromatic compounds generated by glycolysis from lactose or citrate are; acetaldehyde,
acetic acid, acetoin, 2,3-butanediol, 2-butanone, diacetyl, ethanol, 2-butanol, and pro-
pionic acid.

23.4.3.2 Lipolysis
Lipolysis is the hydrolysis of tri-, di-, and mono-acylglycerides producing free fatty acids
and is carried out by two hydrolytic enzymes: esterases and lipases. The sources of
lipolytic enzymes in cheese are relatively widespread, and they can originate from: (a)
the indigenous milk lipase (lipoprotein lipase), (b) pregastric esterases/rennet pastes, (c)
starter lactic acid bacteria, (d) adjunct starter bacteria, (e) non-starter advantageous
lactic acid bacteria, (f) yeasts and molds, or (g) exogenous lipases (Deeth and Fitzgerald,
1995; Fox and Wallace, 1997; McSweeney and Sousa, 2000). The extent of lipolysis var-
ies considerably between cheese types and within cheese types due to seasonal effects on
milk, animal breed, differences in manufacture, and ripening times (Woo et al., 1984;
Woo and Lindsay, 1984; Kilcawley and O’Sullivan, 2018). The OT of free fatty acids is
influenced substantially by pH and cheese composition. Free fatty acid anions at high
pH are less flavor active and less volatile than at lower pH. The main volatile compounds
arising from metabolic activity, chemical activity, or enzymatic activity on lipids are
esters, aldehydes, methyl ketones, secondary alcohols, and lactones by bacteria, yeasts,
and molds.
472 Food Aroma Evolution

23.4.3.3 Proteolysis
The main products of proteolysis are free amino acids, and these are readily metabolized
in cheese (over-ripening once carbohydrate sources have been depleted) and to a lesser
extent in yogurt. In cheese, the metabolism of amino acids is undertaken by starter bacte-
ria, secondary cultures, yeasts, molds, and advantageous non-starter lactic acid bacteria
(Kilcawley and O’Sullivan, 2018). In milk, some protein metabolic compounds can be
derived from diet, rumen metabolism of amino acids, or by the growth of advantageous
bacteria (Kilcawley et al., 2018). The main amino acid substrates for flavor generation
by microbial activity are aromatic amino acids, branched-chain amino acids, aspartic
acid, and sulfur amino acids. Amino acid transamination results in the formation of
α-keto acids, which are key intermediates in the biochemical process (Ganesan et al.,
2007). α-Ketoglutaric acid is the most widely studied α-keto acid acceptor for amino
acid transamination and has been identified as a rate-limiting factor in flavor develop-
ment in different cheeses (Banks et al., 2001; Yvon et al., 1998; Yvon and Rijnen, 2001).
α-Ketoglutaric acid is produced by the glutamate dehydrogenase pathway by oxidative
deamination of glutamate (Helinck et al., 2004; Tanous et al., 2002), and this is likely
to be the main mechanism of formation. A wide range of volatile compounds are pro-
duced from amino acids that are important cheese flavor compounds (Smit et al., 2005;
Kilcawley and O’Sullivan, 2018).

23.5 MAIN VOLATILE COMPOUNDS IN MILK, CHEESE, AND YOGURT

23.5.1 Acids

The main acids that influence the sensory character of milk and/or cheese and yogurt
are shown in Table 23.1. A major characteristic of animal milk fat that is different to
vegetable fat is the presence of water-soluble short-chain fatty acids from butanoic acid
to dodecanoic acid. All of these acids are important aromatic volatiles in milk and dairy
products, mainly due to their abundance. Most fatty acids in milk are bound to glycerol
as tri-acylglycerides, di-glycerides, or mono-glycerides, but a small portion are in free
form. These acids are only aroma active in free form (Kilcawley and O’Sullivan, 2018).
Butanoic acid in milk is an indicator of lipolytic rancidity and has a very strong sensory
characteristic and is an important odor compound in cheese (Ha and Lindsay, 1991).
Butanoic, heptanoic, and octanoic acids are all very important volatile compounds in
many cheese varieties partly because of their abundance, as they are relatively easily
hydrolyzed due to their position on the glyceride backbone (mainly Sn-3) (Kilcawley and
O’Sullivan, 2018). The OT of heptanoic, octanoic acids are slightly lower than decanoic
and dodecanoic acids but are much higher than the main 4-alkyl-branched-chain fatty
acids which are also thought to be important characteristic volatiles for caprine and ovine
milk and cheese (Salles et al., 2002; Delgado et al., 2013; Kaffarnik et al., 2014). Both
4-methyl octanoic acid and 4-ethyl octanoic acid have characteristic goaty aromas and
low to very low OT. Hexanoic acid has a “cheesy, goaty, sharp” aroma with a low OT
and is important in ovine and caprine cheeses.
Other acids such as formic acid, acetic acid, and propionic acid are chemically
similar to these other fatty acids but arise primarily from a carbohydrate metabolism
rather than from fat hydrolysis. Acetic acid is also a product of amino acid metabolism
and has a characteristic “vinegar,” “sour,” “pungent” aroma and has a relatively high
 Milk/Dairy 473

TABLE 23.1 Odor Thresholds of Important Acids in Dairy Flavor

CAS Odor Threshold
Volatile Acid Number in Water (Ppm) Odor Description
Ethanoic acid (Acetic acid) 64-19-7 22 a Vinegar, sour, pungentc
Propionic acid 79-09-4 2.19a Sour pungentc
Butanoic acid 107-92-6 1.274a Rancid, cheesy, sharpc
Pentanoic acid (Valeric acid) 109-52-4 1.207a Cheesy, sour, meaty, sweatyc
Hexanoic acid 142-62-1 0.0356a Cheesy, goaty, sharpc
Heptanoic acid 111-14-6 3b Cheesy, goaty, rancidc
Octanoic acid 124-07-2 1.405a Cheesy, sweatyc
Nonanoic acid 112-05-0 2.4–8.8c Fatty, greenc
Decanoic acid 334-48-5 10a Soapy, waxyc
Dodecanoic acid 143-07-7 2.2–16c Soapy, metallicc
2-Methyl propionic acid 79-31-2 5.3d Cheesy, rancid, caramelc
(Isobutyric acid)
2-Methyl butanoic acid 116-53-0 0.07c Cheesy, rancid, sour, sweatyc
3-Methyl butanoic acid 503-74-2 0.07d Sharp, sweaty, sweet, fruityf
(Isovaleric acid)
4-Ethyl octanoic acid 16493-80-4 0.0018–2.4d Characteristic goatyf
4-Methyl octanoic acid 54947-74-9 0.02d Goaty, muttonye
Benzoic acid 65-85-0 n/a Green,fattyg
Phenylacetic acid 103-82-2 0.14e Waxy, rosya
a Karagül-Yüceer et al., 2003; bAbilleira et al., 2010; cQian and Burbank, 2007; dMolimard and
Spinnler, 1996; eCzerny et al., 2008; fGanesan et al., 2007; gMoio et al., 2000.
n/a: Not available.

OT. Propionic acid directly contributes to the flavor of Swiss cheeses and is produced
by propionibacteria that metabolize l-lactate. It has a “sour,” “pungent” aroma and a
medium OT. It is also a product of amino acid metabolism. Branched-chain acids such
as ­isobutyric acid, 2-methyl butanoic acid, and isovaleric acid derived from the metabo-
lism of branch chain amino acids also have a low OT and are widespread in cheeses and
yogurts (Ha and Lindsay, 1991; Curioni and Bosset, 2002; Smit et al., 2005; Routray and
Mishra, 2011). Phenylacetic acid is derived from aromatic amino acid metabolism and
also has a low OT and is widely found in cheese and yogurt (Curioni and Bosset, 2002;
Routray and Mishra, 2011; Kilcawley and O’Sullivan, 2018).

23.5.2 Aldehydes

The main sources of aldehydes in milk and dairy products varies from carbohydrate
metabolism, amino acid metabolism, and lipid oxidation (Kilcawley et al., 2018). The
main aldehydes thought to influence the sensory characteristics of milk, cheese, and
yogurts are listed in Table 23.2.
Acetaldehyde is mainly produced from glycolysis and is an important volatile com-
pound in yogurt and in some cheeses. Both benzaldehyde and phenylacetaldehyde are
widespread in many kinds of cheese and in yogurt (Curioni and Bosset, 2002; Smit et al.,
2005; Routray and Mishra, 2011; Bertuzzi et al., 2018; Kilcawley and O’Sullivan, 2018)
474 Food Aroma Evolution

TABLE 23.2 Odor Thresholds of Important Aldehydes in Dairy Flavor

CAS Odor Threshold
Volatile Aldehyde Number in Water (ppm) Odor Description
Propanal 123-38-6 0.037a Pungent, acrid, solventa
Butanal 123-72-8 0.018a Pungent, malty, greena
Pentanal 110-62-3 0.012–0.07a Malty, apple, greena
Hexanal 66-25-1 0.045b Grassy, green, tallowa
Heptanal 111-71-7 0.003b Fatty, green, woody, fruitya
Octanal 124-13-0 0.0007b Fatty, citrusa
Nonanal 124-19-6 0.001c Citrus, green, fatty, florala
Decanal 112-31-2 0.002a Waxy, floral, citrusa
Dodecanal 112-54-9 0.0005–0.002a Citrus, powerfula
Acetaldehyde 75-07-0 0.025a Pungent, penetrating, fruitya
Benzaldehyde 100-52-7 0.003d Bitter, almond oil, sweet cherryh
Phenylacetaldehyde 122-78-1 0.001e Floral, hyacinth, greena
(Benzeneacetaldehyde)
2-Methyl propanal 78-84-2 0.0007b Malty, cocoa, green, pungenta
2-Methyl butanal 96-17-3 0.0009b Cocoa, coffee, almond, maltya
3-Methyl butanal 590-86-3 0.012e Malty, cocoaa
Vanillin 121-33-5 0.64f Vanillad
Furfural 98-01-1 3a Sweet, almond, penetratinga
(Z)-4-Heptenal 6728-31-0 0.00006g Putty-like, biscuit-like, potato-
like, green, tallow and creamy
a Qian and Burbank, 2007; bVazquez-Landaverde et al., 2005; cAbilleira et al., 2010; dKara-
hadian et al., 1985; eMoio et al., 2000; fKaragül-Yüceer et al., 2003; gBendall and Olney,
2001; hSmit et al., 2005.

and have quite a low OT and very characteristic aromas, “bitter,” “almond oil,” “sweet
cherry” and “floral,” “hyacinth,” “green,” respectively. These aldehydes are products of
aromatic amino acid metabolism (Kilcawley and O’Sullivan, 2018).
Many aldehydes in milk and dairy products are derived from fatty acid autoxidation
(Kilcawley and O’Sullivan, 2018). In general, they have a low to very low OT but are
readily reduced and thus their role in cheese flavor may not be as significant as other lipid
volatile compounds. Many aldehydes confer green, grass-like aromas or malty nuances
with most saturated and unsaturated aldehydes derived from the oxidation of saturated
and unsaturated free fatty acids, respectively. This is not a prevalent mechanism in many
kinds of cheese due to low redox potential. However, aldehydes from lipid oxidation are
major aroma compounds in dairy powders due to their association with oxidation off-
flavors (Croissant et al., 2011). Hexanal is associated with “fatty,” “green,” “woody,”
“fruity” aromas but has a relatively low OT. Pentanal is has a “malty,” “apple,” “green”
aroma and a low OT. Bendal and Olney (2001) identified (z)-4-heptenal another lipid
oxidation product as a key aroma compound of bovine milk. The OT of (z)-4-heptenal is
very low; however, it has not been widely identified in many milk samples, although this
may be due to the choice of extraction procedures used rather than its absence.
Aldehydes derived from the metabolism of branched-chain amino acids (2-methyl
propanal, 2-methyl butanal, and 3-methyl butanal) have a low OT and in general “cocoa,”
“malty” aroma attributes. They are found in most cheese varieties and in yogurt (Curioni
 Milk/Dairy 475

and Bosset, 2002; McSweeney, 2004; Smit et al., 2005; Delgado et al., 2011; Routray and
Mishra, 2011; Kilcawley and O’Sullivan, 2018).

23.5.3 Ketones

The main ketones thought to influence the aromatic properties of milk and dairy prod-
ucts are listed in Table 23.3. Ketones are either derived from carbohydrate metabolism,
oxidation of fatty acids, or metabolism of amino acids (Kilcawley and O’Sullivan, 2018).
Acetoin, 2-butanone, and diacetyl are derived from glycolysis. The OT of 2-butanone
is relatively high and therefore unlikely to impact on sensory perception at low levels of
abundance and has an “acetone-like” aroma. Both acetoin and diacetyl are much more
odor active. Diacetyl, which has a “butter” sensory character and both acetoin and diace-
tyl are important compounds in the flavor of yogurt (Routray and Mishra, 2011). Diacetyl
is also a significant aroma compound in many cheese types and is especially important in
cottage cheese and quarg. Acetophenone a product of the metabolism of aromatic acids
is an important aroma compound in surface mold-ripened cheese (Bertuzzi et al., 2018)
with a low OT and an “orange blossom,” “sweet,” “pungent” aroma.
Methyl ketones are major contributors to the odor of many kinds of cheese but espe-
cially blue-veined cheese. Methyl ketones are formed by β-oxidation of mainly short and
medium free fatty acids; however, other mechanisms have been suggested for their for-
mation in cheese due to their abundance in some cheese types in relation to the free fatty
acid content. Alewijn (2006) postulated that some methyl ketones in cheeses like Gouda

TABLE 23.3 Odor Thresholds of Important Ketones in Dairy Flavor

Odor Threshold
Volatile Ketones CAS Number in Water (ppm) Odor Description
2-Propanone (Acetone) 67-64-1 40.9–500a Acetone-like, pungentf
2-Butanone 78-93-3 50a Acetone-like, ethericf
2-Pentanone 107-87-9 2.3a Floral, fruity, wine,
acetone-likef
2-Hexanone 591-78-6 0.93a Floral, fruityf
2-Heptanone 110-43-0 0.009b Blue cheese, fruity, sweetf
2-Octanone 111-13-7 1c Fruity, musty, unripe, apple,
greenf
2-Nonanone 821-55-6 0.02b Fruity, musty, rose, tea-likef
2-Decanone 693-54-9 0.19a Fruity, mustyf
2-Undecanone 112-12-9 0.007b Floral, herbaceous, fruityf
Acetophenone 98-86-2 0.65–0.17a Orange blossom, sweet,
pungentf
1-Octen-3-one 4312-99-6 0.009a Mushrooma
Acetoin 513-86-0 0.75*d Butterya
(3-Hydroxy-2-Butanone)
Diacetyl (2,3-Butanedione) 431-03-8 0.005e Butterya
a Molimard and Spinnler, 1996; bMoio et al., 2000; cKarahadian et al., 1985; dLarsen and Poll,
1990; eVazquez-Landaverde et al., 2005; fQian and Burbank, 2007.
* Reference odor threshold in raspberries.
476 Food Aroma Evolution

may arise directly from esterified β-keto acids and Forss (1979) postulated that heating
of milk may also induce ketone formation. The aroma of methyl ketones is varied, with
many having a fruity dimension and a low to very low OT. The ethyl ketone 1-octen-3-
one is thought to be an important component in Parmesan, Grana Padano, and Pecorino
cheeses and in yogurt and has a mushroom-like earthy aroma (Kubickova and Grosch,
1998; Qian and Burbank, 2007; Routray and Mishra, 2011). 1-Octen-3-one is a product
of lipid oxidation with a very low OT (Table 23.3). Many other ketones exist which are
produced from lipid oxidation in cheese, but their presence varies considerably between
cheese types. Overall, it is thought that methyl and ethyl ketones have the biggest impact
on the flavor of blue-type cheeses and surface mold-ripened cheeses (Bertuzzi et al., 2018).

23.5.4 Alcohols

Alcohols can also be produced from glycolysis, lipid oxidation, and amino acid metabo-
lism. The main alcohols thought to influence the sensory character of milk, cheese, and
yogurt are listed in Table 23.4.
Ethanol is a product of carbohydrate metabolism (Kilcawley and O’Sullivan, 2018)
and is more important as a key component of ethyl esters rather than as a stand-alone

TABLE 23.4 Odor Thresholds of Important Alcohols in Dairy Flavor

Odor Threshold
Volatile Alcohols CAS Number in Water (ppm) Odor Description
Ethanol 64-17-5 100–200a Alcoholica
1-Propanol 71-23-8 9–45a Alcoholic, fruity, sweeta
1-Butanol 71-36-3 0.5–7.5a Fruity, green, medicinala
1-Pentanol 71-41-0 0.73b Green, fusel oil, woodya
1-Hexanol 111-27-3 0.05c Green, florala
1-Heptanol 111-70-6 0.52–2.4a Fatty, sweet, greena
1-Octanol 111-87-5 0.13d Fatty, waxyd
2-Butanol 78-92-2 0.525d Fruitya
2-Pentanol 6032-29-7 8.5a Fruity, green, fusel oila
2-Heptanol 543-49-7 0.005b Fruity, brassy, herbaceousa
2-Octanol 123-96-6 0.18e Greene
2-Nonanol 628-99-9 0.075b Fatty, mild, green, melonb
2,3-Butanediol 513-85-9 50f* Fruityi
1-Octen-3-ol 3391-86-4 0.001b Mushrooma
3-Methyl butanol 123-51-3 0.25g Alcoholic, green, floral,
maltya
2-Methyl butanol 137-32-6 0.3b Winej
2-Methyl propanol 78-83-1 3.2–12.5h Penetrating, alcohol,
wine-likej
2-Phenylethanol 60-12-8 0.12–1a Honey, spicy, florala
a Qian and Burbank, 2007; bMoio et al., 2000; cAbilleira et al., 2010; dJuric et al.,
2003; eMolimard and Spinnler, 1996); fDahl et al., 2000; gVazquez-Landaverde et
al., 2005; hCzerny et al., 2008; iCurioni and Bosset, 2002; jSingh et al., 2003.
* Reference odor threshold in Serra da Estrela cheese.
 Milk/Dairy 477

aroma active volatile, due to its very high OT. 2-3-Butanediol, another product of gly-
colysis, also has a high OT but a “fruity” aroma. The secondary alcohol, 2-butanol is
produced from 2-butanone also derived from glycolysis, it is also present in many cheese
types and has low OT with an “acetone-like” aroma.
The main primary alcohols resulting from lipid oxidation are 1-propanol, 1-penta-
nol, 1-hexanol, and 1-heptanol. Their odor varies, having “fruity,” “sweet,” “green,”
“floral,” “fatty,” and “waxy” characteristics with a low OT, and they are found in milk,
cheese, and yogurt (Kilcawley et al., 2018; Kilcawley and O’Sullivan, 2018; Curioni and
Bosset, 2002; Routray and Mishra, 2011). Secondary alcohols are formed by the reduc-
tion of their corresponding methyl ketones (Engels and Visser, 1997). The most common
secondary alcohols in dairy products are 2-pentanol, 2-heptanol, 2-octanol, and 2-nona-
nol, and again they tend to have a “fruity,” “green” aroma character and a range of OTs.
2-Heptanol has been identified as a key odourant of Gorgonzola, and Grana Padano
cheese (Curioni and Bosset, 2002) and Penicillium species are directly responsible for
the production of 2-pentanol, 2-heptanol, and 2-nonanol in blue-vein cheeses (Collins et
al., 2003). 1-Octen-3-ol has a mushroom-like aroma and a very low OT and is derived
from 1-octen-3-one and may have a role in the flavor of Camembert, Parmesan, Grana
Padano, and Pecorino cheeses, but may also provide a metallic aroma at high concentra-
tions (Moio and Addeo, 1998).
Alcohols derived from the metabolism of branched-chain amino acids (3-methyl buta-
nol, 2-methyl butanol, and 2-methyl propanol) have a “alcoholic wine-like” aroma and
low to medium OT and are also widespread in cheeses (Curioni and Bosset, 2002; Smit et
al., 2005; Delgado et al., 2011; Kilcawley and O’Sullivan, 2018). 2-Phenylethanol derived
from the metabolism of aromatic amino acids has “penetrating,” “alcohol,” “wine-like”
aroma and a low OT, and is also present in many kinds of cheese (Curioni and Bosset,
2002; Singh et al., 2003; Bertuzzi et al., 2018; Kilcawley and O’Sullivan, 2018).

23.5.5 Esters

Esters are formed by two potential pathways, esterification (formation of esters from
alcohols and carboxylic acids) and alcoholysis (production of esters from alcohols and
acylglycerols or from alcohols and fatty acyl-CoAs derived from the metabolism of fatty
acids, amino acids and/or carbohydrates) (Kilcawley and O’Sullivan, 2018). The main
esters thought to influence the sensory properties of milk and dairy products are listed
in Table 23.5. A rate-limiting step in ester formation in cheese is alcohol availability, and
ethyl esters are the most common because of the presence of ethanol from carbohydrate
metabolism. Methyl, propyl, butyl, and isobutyl esters have all been reported in cheese
but are less common as generation is dependent upon the availability of the relevant
alcohols from different metabolic pathways. All esters are generally described as having a
“fruity” aroma and have a low OT. Esters are characteristic compounds of many cheese
types (Barbieri et al., 1994; Sablé and Cottenceau, 1999; Curioni and Bosset, 2002; Singh
et al., 2003; Delgado et al., 2011; Bertuzzi et al., 2018; Kilcawley and O’Sullivan, 2018).

23.5.6 Sulfur Compounds

The main sulfur compounds thought to influence the sensory properties of milk and
dairy products are shown in Table 23.6. Sulfur compounds in general are quite odor
478 Food Aroma Evolution

TABLE 23.5 Odor Thresholds of Important Esters in Dairy Flavor

Odor Threshold
Volatile Esters CAS Number in Water (ppm) Odor Description
Ethyl acetate 141-78-6 0.005 a Solvent, fruityf
Ethyl propanoate 105-37-3 0.0099b Pineapple, fruityf
Ethyl butanoate 105-54-4 0.001c Pineapple, banana,
melonf
Ethyl hexanoate 123-66-0 0.001c Pineapple, banana,
fruityf
Ethyl octanoate 106-32-1 0.07d Apricot, fruity, fatty,
floralf
Ethyl decanoate 110-38-3 0.122d Fruity, grape (cognac)f
Ethyl benzoate 93-89-0 0.06c Florale
Ethyl isovalerate (Ethyl-3-methyl 108-64-5 0.0002d Apple, fruityg
butanoate)
Ethyl isobutanoate (Ethyl 97-62-1 0.003e Fruity, bananae
isobutyrate)
Butyl acetate 123-86-4 0.066d Pineappleb
Butyl butanoate 109-21-7 0.10d Pineapple, banana,
sweeth
Butyl hexanoate 626-82-4 0.70d Fruity, wine-likei
Propyl butanoate 105-66-8 0.124d Pineapple, bananab
Isoamyl acetate (3-methyl butyl 123-92-2 0.002b Pear, banana,
acetate) powerfulf
Amyl acetate 628-63-7 0.001c Banana, pearg
Methyl hexanoate 106-70-7 0.05F Pineapple, fruityf
2-Phenylethyl acetate 103-45-7 19F Floral, rose, honeyf
a Vazquez-Landaverde et al., 2005; bMolimard and Spinnler, 1996; cMoio et al., 2000; dAbilleira et
al., 2010; eCurioni and Bosset, 2002; fQian and Burbank, 2007; gStyger et al., 2011; hBarron et al.,
2005; iMoio and Addeo, 1998.

TABLE 23.6 Odor Thresholds of Important Sulfur in Dairy Flavor

Odor
Threshold in
Volatile Sulfur Compounds CAS Number Water (ppm) Odor Description
Dimethyl sulfide 75-18-3 0.002a Boiled cabbageb
Dimethyl disulfide 624-92-0 0.00016b Cabbage-like, strong onionb
Dimethyl trisulfide 3658-80-8 0.00001b Very ripe cheese, garlicb
Methanethiol 74-93-1 0.002c Rotten cabbage, fecalb
Methional 3268-49-3 0.0002b Boiled/baked potatob
Dimethyl sulfone 67-71-0 n/a*d Sulfurous, hot milk, burnedd
3-Methyl thiopropanol (Methionol) 505-10-2 0.00043e Baked/boiled potatof
a Vazquez-Landaverde et al., 2005; bQian and Burbank, 2007; cMolimard and Spinnler, 1996;
dKilcawley et al., 2018; eCzerny et al., 2008; fSingh et al., 2003.

* Odor threshold is low.

n/a: Not available.
 Milk/Dairy 479

active, with a very low OT and are the result of the metabolism of the amino acids methi-
onine or cysteine (Kilcawley and O’Sullivan, 2018). These sulfur compounds are present
in many kinds of cheese and in yogurt (Curioni and Bosset, 2002; Smit et al., 2005; Qian
and Burbank, 2007; Routray and Mishra, 2011; Kilcawley and O’Sullivan, 2018). The
sensory characters are in general negative, but they all contribute positively to sensory
perception when present at low abundance.

23.5.7 Lactones

Some of the main lactones thought to influence the sensory properties of milk and dairy
products are given in Table 23.7. Lactones are widely found in milk (Urbach, 1997) and
yogurt (Routray and Mishra, 2011) and to a lesser extent in cheese apart from Gouda,
Parmesan, and Grana Padano (Alewijn et al., 2005; Qian and Burbank, 2007). Lactones
can be derived by heat in the presence of water and hydroxyacids, with γ- and δ-lactones
derived from 4- and 5-hydroxy fatty acids, respectively being more stable (Alewijn, 2006).
Alewijn et al. (2007) suggested that lactones may also be formed by a one-step non-
enzymatic reaction in which a hydroxyl fatty acid esterified in a triacylglycerol undergoes
an intramolecular transesterification to release a lactone directly. It also reported that
lactones may be derived directly from keto acids after they are reduced to hydroxyacids
(Wong et al., 1975). The formation of lactones in bovine milk appears to be influenced by
diet, season, stage of lactation, and breed (Fox et al., 2000). Both γ- and δ-dodecalactone
have been identified as potentially important compounds in yogurt aroma (Routray and
Misha, 2011) and have a low OT.

23.5.8 Miscellaneous Volatile Compounds

Some of the main volatile compounds not classified as acids, aldehydes, alcohols, esters,
ketones, lactones, or sulfur compounds that are thought to influence the sensory prop-
erties of milk and dairy products are listed in Table 23.8. The phenolic compounds p-­
cresol and phenol have a different OT, with phenol much higher than p-cresol. Both have

TABLE 23.7 Odor Thresholds of Important Lactones in Dairy Flavor

Odor Threshold
Volatile Lactones CAS Number in Water (ppm) Odor Description
γ-Hexalactone 695-06-7 1.6–13a Coconut, fruity, sweeta
γ-Heptalactone 105-21-5 0.52a Coconut, fruity, nuttya
δ-Octalactone 698-76-0 0.57b Coconut, animala
γ-Octalactone 104-50-7 0.095–0.4b Coconut, fruitya
γ-Nonalactone 104-61-0 0.065a Coconut, peacha
δ-Decalactone 705-86-2 0.03c Coconut, apricota
γ-Decalactone 706-14-9 0.09b Coconut, apricot, fattya
δ-Dodecalactone 713-95-1 0.0046c Fresh fruit, peacha
γ-Dodecalactone 2305-05-7 0.007a Peach, butter, sweet, florala
a Qian and Burbank, 2007; bMolimard and Spinnler, 1996; cKaragül-Yüceer et al.,
2003.
480 Food Aroma Evolution

TABLE 23.8 Odor Thresholds of Important Miscellaneous Compounds

in Dairy Flavor
Volatile
Miscellaneous Odor Threshold
Compounds CAS Number in Water (ppm) Odor Description
Indole 120-72-9 0.021a Mothball-likea
Toluene 108-88-3 n/ab* Nutty, bitter almondf
α-Pinene 80-56-8 n/ab** Pine, greenb
Skatole 83-34-1 0.05c Unclean, mothballg
Phenol 108-95-2 10d Floral, medicalh
p-Cresol 106-44-5 0.001–0.055e Phenolic, medicinali
a Karagül-Yüceer et al., 2003; bCurioni and Bosset, 2002; cUrbach et al.,
1972; dKarahadian et al., 1985; eCzerny et al., 2008; fArora et al., 1995;
gSingh et al., 2003. hMolimard and Spinnler, 1996; iKilcawley and O’Sullivan,

2018.
*    Odor threshold is high.
** Odor threshold is medium.
n/a: Not available.

“phenolic medicinal” aromas and are also relatively common in cheese (Ha and Lindsay,
1991; Urbach, 1997; Curioni and Bosset, 2002; Singh et al., 2003; Bertuzzi et al., 2018).
p-Cresol can result from a number of sources and is linked to diet (Faulkner et al., 2018).
Both indole and skatole have low OT and “mothball” aromas and are generally undesir-
able components in cheese and yogurt except in surface mold-ripened cheeses (Bertuzzi
et al., 2018). Toluene is potentially from the metabolism of a range of substrates, includ-
ing β-carotene (Kilcawley et al., 2018), and has a “nutty,” “bitter almond” aroma but
a high OT and therefore unlikely to influence flavor unless very abundant. Skatole is a
product of aromatic amino acid metabolism (Smit et al., 2005) and is characterized by an
“unclean, mothball” aroma with a low OT. α-Pinene is a monoterpene derived from diet
(Kilcawley et al., 2018) and has a “pine,” “green” aroma and a medium OT and relatively
common in milk and cheese (Urbach, 1997; Curioni and Bosset, 2002; Villeneuve et al.,
2013; Basdagianni et al., 2019).

REFERENCES

Abilleira, E., Schlichtherle-Cerny, H., Virto, M., de Renobales, M., and Barron, L.J.R.
2010. Volatile composition and aroma-active compounds of farmhouse Idiazabal
cheese made in winter and spring. Int. Dairy J. 20:537–44.
Ai, N.S., Liu, H.L., Wang, J., Zhang, X.M., Zhang, H.J., Chen, H.T., Huang, M.Q.,
Liu, Y.G., Zeng, F.P., and Sun, B.G. 2015. Triple-channel comparative analysis of
volatile flavour composition in raw whole milk and skim milk via electronic nose,
GC-MS and GC-O. Anal. Meth. 7:4278–84.
Alewijn, M. 2006. The Formation of Fat-derived Flavour Compounds during Ripening
of GOUDA-type Cheese. PhD Thesis, Wagenigen University, the Netherlands.
Alewijn, M., Sliwinkski, E.L., and Wouters, J.T.M. 2005. Production of fat-derived
(flavour) compounds during the ripening of Gouda cheese. Int. Dairy J.
15:733–40.
 Milk/Dairy 481

Alewijn, M., Smit, B.A., Sliwinkski, E.L., and Wouters, J.T.M. 2007. Formation mecha-
nism of lactones in Gouda cheese. Int. Dairy J. 17:59–66.
Arora, G., Cormier, F., and Lee, B. 1995. Analysis of odor-active volatiles in ched-
dar cheese headspace by multidimensional gc/ms/sniffing. J. Agri. Food Chem.
43:748–52.
Badings, H.T. 1991. Milk. In Volatile Compounds in Food and Beverages, ed. H. Maarse,
pp. 91–6. New York, NY: Marcel Dekker.
Badings, H.T. and Neeter, R. 1980. Recent advances in the study of aroma compounds of
milk and dairy products. Neth. Milk Dairy J. 34:9–30.
Banks, J.M., Yvon, M., Gripon, J.C., Fuente, M.A.D.L., Brechany, E.Y., Williams, A.G.,
and Muir, D.D. 2001. Enhancement of amino acid catabolism in Cheddar cheese
using α-ketoglutarate: Amino acid degradation in relation to volatile compounds
and other aroma character. Int. Dairy J. 11:235–43.
Barbieri, G.L., Bolzoni, M., Careri, A., Mangia, G., Parolari, S., Spagnoli, S., and Virgili,
R. 1994. Study of the volatile fraction of parmesan cheese. J. Agric. Food Chem.
42:1170–6.
Barron, L.J.R., Redondo, Y., Aramburu, M., Pérez-Elortondo, F.J., Albisu, M., Nájera,
A.I., and de Renobales, M. 2005. Variations in volatile compounds and flavour in
idiazabal cheese manufactured from ewe’s milk in farmhouse and factory. J. Sci.
Food Agric. 85:1660–71.
Basdagianni, Z., Papaloukas, L., Kyriakou, G., Karaiskou, C., Parissi, Z., Sinapis, E.,
and Kasapidou, E. 2019. A comparative study of the fatty acid and terpene profiles
of ovine and caprine milk from Greek mountain sheep breeds and a local goat breed
raised under a semi-extensive production system. Food Chem. 278:625–9.
Bendall, J.G. 2001. Aroma compounds of fresh milk from New Zealand cows fed differ-
ent diets. J. Agric. Food Chem. 4910:4825–32.
Bendall, J.G. and Olney, S.D. 2001. Hept-cis-4-enal: Analysis and flavour contribution to
fresh milk. Int. Dairy J. 11:855–64.
Bertuzzi, A.S., Walsh, A.M., Sheehan, J.J., Cotter, P.D., Crispie, F., McSweeney, P.L.H.,
Kilcawley, K.N., and Rea, M.C. 2018. Omics-based insights into flavour devel-
opment and microbial succession within surface-ripened cheese. mSystems 3(1):
1–15.
Bonanno, A., Tornambè, G., Bellina, V., De Pasquale, C., Mazza, F., Maniaci, G., and
Di Grigoli, A. 2013. Effect of farming system and cheesemaking technology on the
physicochemical characteristics, fatty acid profile, and sensory properties of cacio-
cavallo palermitano cheese. J. Dairy Sci. 961:710–24.
Bovolenta, S., Corazzin, M., Saccá, E., Gasperi, F., Biasioli, F., and Ventura, W. 2009.
Performance and cheese quality of brown cows grazing on mountain pasture fed
two different levels of supplementation. Livestock Sci. 124:58–65.
Bugaud, C., Buchin, S., Hauwuy, A., and Coulon, J.-B. 2001. Relationships between
flavour and chemical composition of abondance cheese derived from different types
of pastures. Le Lait 816:757–73.
Cadwallader, K. 2010. Instrumental measurement of milk flavour and colour. In
Improving the Safety and Quality of Milk, Volume 2: Improving Quality in
Milk Products, ed. M.W. Griffiths, pp. 181–206. Oxford, Cambridge, New Delhi:
Woodhead Publishing Limited.
Collins, Y.E., McSweeney, P.L.H., and Wilkinson, M.G. 2003. Lipolysis and free
fatty acid catabolism in cheese: A review of current knowledge. Int. Dairy J. 13:
841–66.
482 Food Aroma Evolution

Coppa, M., Martin, B., Pradel, P., Leotta, B., Priolo, A., and Vasta, V. 2011. Effect of a
hay-based diet or different upland grazing systems on milk volatile compounds. J.
Agric. Food Chem. 599:4947–54.
Croissant, A.E., Washburn, S., Dean, L., and Drake, M.A. 2007. Chemical properties
and consumer perception of fluid milk from conventional and pasture-based pro-
duction systems. J. Dairy Sci. 90:4942–53.
Croissant, A.E., Watson, D.M., and Drake, M.A. 2011. Application of sensory and
instrumental volatile analysis to dairy products. Annu. Rev. Food Sci. Technol.
2:395–421.
Curioni, P.M.G. and Bosset, J.O. 2002. Key odorants in various cheese types as deter-
mined by gas chromatography-olfactometry. Int. Dairy J. 12:959–84.
Czerny, M., Christbauer, M., Christbauer, M., Fischer, A., Granvogl, M., Hammer, M.,
Hartl, C., Hernandez, N.M., and Schieberle, P. 2008. Re-investigation on odour
thresholds of key food aroma compounds and development of an aroma language
based on odour qualities of defined aqueous odorant solutions. Eur. Food Res.
Technol. 228:265–73.
Dahl, S., Tavaria, F.K., and Malcata, F.X. 2000. Relationships between flavour and
microbiological profiles in serra da estrela cheese throughout ripening. Int. Dairy J.
10:255–62.
Deeth, H.C. and FitzGerald, C.H. 1995. Lipolytic enzymes and hydrolytic rancidity in
milk and milk products. In Advanced Dairy Chemistry, Volume 2: Lipids, ed. P.F.
Fox, pp. 247–308. London: Chapman & Hall.
Delgado, F.J., González-Crespo, J., Cava, R., and Ramírez, R. 2011. Formation of the
aroma of a raw goat milk cheese during maturation analysed by SPME-GC-MS.
Food Chem. 129:1156–63.
Delgado, F.J., González-Crespo, J., Ramírez, R., and Cava, R. 2013. The aromatic pro-
file of cheese during ripening: A focus on goats cheese. In Handbook of Cheese
in Health, Production, Nutrition and Medical Sciences, eds. V.R. Preedy, R.R.
Watson, and V.B. Patel, pp. 467–80. Wagenigen: Academic Publishers.
Drake, M.A. 2007. Invited review: Sensory analysis of dairy foods. J. Dairy Sci.
90:4925–37.
Dubroeucq, H., Martin, B., Ferlay, A., Pradel, P., Verdier-Metz, I., Chilliard, Y., Agabriel,
J., and Coulon, J. 2002. Cow’s feeding may modify sensory properties of milk.
Rencontres Recherches Ruminants 9:351–4.
Dunkel, A., Steinhaus, M., Kotthoff, M., Nowak, B., Krautwurst, D., Schieberle, P., and
Hofmann, T. 2014. Nature’s chemical signatures in human olfaction: A foodborne
perspective for future biotechnology. Angew. Chem. Int. Ed. 53:7124–43.
Engels, W.J.M. and Visser, S. 1997. A comparative study of volatile compounds in the
water-soluble fraction of various types of ripened cheese. Int. Dairy J. 48:127–40.
Faulkner, H., O’Callaghan, T.F., McAuliffe, S., Hennessy, D., Stanton, C., O’Sullivan,
M.G., Kerry, J.P., and Kilcawley, K.N. 2018. Effect of different forage types on the
volatile and sensory properties of bovine milk. J. Dairy. Sci. 101:1034–47.
Forss, D. 1979. Review of the progress of dairy science: Mechanisms of formation of
aroma compounds in milk and milk products. J. Dairy Res. 46:691–706.
Fox, P.F., Guinee, T.P., Cogan, T.M., and McSweeney, P.L.H. 2000. Biochemistry of
cheese ripening. In Fundamentals of Cheese Science, eds. P.F. Fox, T.P. Guinee,
T.M. Cogan, and P.L.H. McSweeney, pp. 236–78. USA: Aspen Publishers.
Fox, P.F. and Wallace, J.M. 1997. Formation of flavor compounds in cheese. Adv. Appl.
Microbiol. 45:17–85.
 Milk/Dairy 483

Ganesan, B., Weimer, B.C., Qian, M.C., and Burbank, H.M. 2007. Compounds associ-
ated with cheese flavour. In Improving the Flavour of Cheese, ed. B.C. Weimer, pp.
26–51. Cambridge, England: Woodhead Publishing.
Ha, J.K. and Lindsay, R.C. 1991. Contributions of cow, sheep, and goat milks to charac-
terising branched-chain fatty acid and phenolic flavors in varietal cheeses. J. Dairy
Sci. 74:3267–74.
Helinck, S., Le Bars, D., Moreau, D., and Yvon, M. 2004. Ability of thermophilic lac-
tic acid bacteria to produce aroma compounds from amino acids. Appl. Environ.
Microbiol. 70:3855–61.
Jandal, J.M. 1996. Comparative aspects of goat and sheep milk. Small Ruminant Res.
22:177–85.
Juric, M., Bertelsen, G., Mortensen, G., and Pertersen, M.A. 2003. Light-induced colour
and aroma changes in sliced, modified atmosphere packaged semi-hard cheeses. Int.
Dairy J. 13:239–49.
Kaffarnik, S., Kayademir, Y., Heid, C., and Vetter, W. 2014. Concentrations of volatile
4-alkyl-branched chain fatty acids in sheep and goat milk and dairy products. J.
Food Sci. 79:C2209–14.
Karagül-Yüceer, Y., Vlahovich, K.N., Drake, M.A., and Cadwallader, K.R. 2003.
Characteristic aroma compounds of rennet casein. J. Agri. Food Chem. 51:6797–801.
Karahadian, C., Josephson, D.B., and Linday, R.C. 1985. Contribution of penicillium sp.
to the flavors of brie and camembert cheese. J. Dairy Sci. 68:1865–77.
Khanal, R.C., Dhiman T.R., Ure A.L., Brennand C.P., Boman R.L., and McMahon D.J.
2005. Consumer acceptability of conjugated linoleic acid-enriched milk and ched-
dar cheese from cows grazing on pasture. J. Dairy Sci. 88:1837–47.
Kilcawley, K. 2017. Cheese flavour. In Fundamentals of Cheese Science, 2nd edition, eds.
P.F. Fox, T.P. Guinee, T.M. Cogan, and P.L.H. McSweeney, pp. 443–74. New York,
NY: Springer.
Kilcawley, K.N., Faulkner, H., Clarke, H.J., O’Sullivan, M.G., and Kerry, J.P. 2018.
Factors influencing the flavour of bovine milk and cheese from grass based versus
non-grass based milk production systems. Foods 7:1–43.
Kilcawley, K.N. and O’Sullivan, M.G. 2018. Cheese favour development and sen-
sory characteristics. In Global Cheesemaking Technology: Cheese Quality and
Characteristics, eds. P. Papademas and T. Bintsis, pp. 45–70. T. Wiley.
Kubickova, L. and Grosch, W. 1998. Evaluation of flavour compounds of Camembert
cheese. Int. Dairy J. 8:11–6.
Larsen, M. and Poll, L. 1990. Odour thresholds of some important compounds in rasp-
berries. Z. Lebensm. Unters. Forsch. 191:129–31.
Larsen, M.K., Kidmose, U., Kristensen, T., Beaumont, P., and Mortensen, G. 2013.
Chemical composition and sensory quality of bovine milk as affected by type
of forage and proportion of concentrate in the feed ration. J. Sci. Food Agric.
931:93–9.
Marsili, R. 2003. Flavors and off-flavors in dairy foods. In Encyclopedia of Dairy Science,
eds. H. Roginski, J.W. Fuquay, and P.F. Fox, pp. 1069–81. London: Academic Press.
McSweeney, P.L.H. 2004. Biochemistry of cheese ripening: Introduction and overview.
In Cheese, Chemistry, Physics and Microbiology, 3rd edition, Volume 1: General
Aspects, eds. P. Fox, P. McSweeney, T. Cogan, and T. Guinee, pp. 347–60. San
Diego, CA: Academic Press.
McSweeney, P.L.H. and Sousa, M.J. 2000. Biochemical pathways for the production of
flavour compounds in cheeses during ripening: A review. Le Lait 80:293–324.
484 Food Aroma Evolution

Moio, L. and Addeo, F. 1998. Grana pandano cheese aroma. J. Dairy Res. 65:317–33.
Moio, L., Dekimpe, J., Etievant, P., and Addeo, F. 1993. Neutral volatile compounds in
the raw milks from different species. J. Dairy Res. 60:199–213.
Moio, L., Piombino, P., and Addeo, F. 2000. Odour-impact compounds of gorgonzola
cheese. J. Dairy Res. 67:273–83.
Molimard, P. and Spinnler, H.E. 1996. Review: Compounds involved in the flavour of
surface mould-ripened cheeses: Origins and properties. J. Dairy Sci. 79:169–84.
Mondello, L., Costa, R., Tranchida, P.Q., Chiofalo, B., Zumbo, A., Dugo, P., and Dugo,
G. 2005. Determination of flavor components in Sicilian goat cheese by automated
HS-SPME-GC. Flavour Frag. J. 20:659–65.
Mulder, H. 1952. Taste and flavour-forming substances in cheese. Neth. Milk Dairy J.
6:157–67.
O’Sullivan, M.G. 2017. A Handbook for Sensory and Consumer-driven New Product
Development: Innovative Technologies for the Food and Beverage Industry.
Amsterdam, Boston, Cambridge, Heidelberg, London, New York, Oxford, Paris,
San Diego, San Francisco, Singapore, Sydney, Tokyo: Woodhead Publishing Limited.
Park, Y.W., Juárez, M., Ramos, M., and Haenlein, G.F.W. 2007. Physico-chemical char-
acteristics of goat and sheep milk. Small Ruminant Res. 68:88–113.
Pawliszyn, J. 1997. Solid Phase Microextraction: Theory and Practice. New York, NY:
Wiley-VCH.
Qian, M.C. and Burbank, H.M. 2007. Hard Italian cheeses: Parmigiano-reggiano and
grana padano. In Improving the Flavour of Cheese, ed. B.C. Weimer, pp. 421–43.
Cambridge, England: Woodhead Publishing.
Routray, W. and Mishra, H.N. 2011. Scientific and technical aspects of yogurt aroma
and taste: A review. Comp. Rev. Food Sci. Food Safety 10:208–20.
Sablé, S. and Cottenceau, G. 1999. Current knowledge of soft cheeses flavour and related
compounds. J. Agric. Food Chem. 47:4825–36.
Salles, C., Somerer, N., Septier, C., Issanchou, S., Chabanet, C., Garem, A., and Le
Quéré, J.-L. 2002. Goat cheese flavour: Sensory evaluation of branched-chain fatty
acids and small peptides. J. Food Sci. 67:835–41.
Schiano, A.N., Harwood, W.S., and Drake, M.A. 2017. A 100-year review: Sensory
analysis of milk. J. Dairy Sci. 100:9966–86.
Singh, T.K., Cadwallader, K.R., and Drake, M.A. 2007. Biochemical processes in the
production of flavor in milk and milk products. In Handbook of Food Products
Manufacturing, Section XIV: Milk and Milk Products, ed. Y.H. Hui, pp. 715–48.
NJ: John Wiley & Sons.
Singh, T.K., Drake, M.A., and Cadwallader, K.R. 2003. Flavor of cheddar cheese: A
chemical and sensory perspective. Comp. Rev. Food Sci. Food Safety 2:139–62.
Smit, G., Smit, B.A., and Engels, W.J.M. 2005. Flavour formation by lactic acid bacte-
ria and biochemical flavour profiling of cheese products. FEMS Microbiol. Rev.
29:591–610.
Spence, C. 2015. Just how much of what we taste derives from the sense of smell? Flavour
4:30.
Styger, G., Prior, B., and Bauer, F.F. 2011. Wine flavour and aroma. J. Indust. Micrbiol.
Biotechnol. 38:1145–59.
Tanous, C., Kieronczyk, A., Helinck, S., Chambellon, E., and Yvon, M. 2002. Glutamate
dehydrogenase activity: A major criterion of the selection of flavour-producing lactic
acid bacteria strains. Anton. Leeuw. 82:271–8.
 Milk/Dairy 485

Toso, B., Procida, G., and Stefanon, B. 2002. Determination of volatile compounds in
cows’ milk using headspace GC-MS. J. Dairy Res. 694:569–77.
Urbach, G. 1997. The flavour of milk and dairy products. II. Cheese: Contribution of
volatile compounds. Int. J. Dairy Technol. 50:79–89.
Urbach, G., Stark, D., and Forss, A. 1972. Volatile compounds in butter oil. II. Flavour
and flavour thresholds of lactones, fatty acids, phenols, indole and skatole in deodor-
ized synthetic butter. J. Dairy Sci. 39:35–47.
Vazquez-Landaverde, P.A., Velazquez, G., Torres, J.A., and Qian, M.C. 2005. Quantitative
determination of thermally derived off-flavor compounds in milk using solid-phase
microextraction and gas chromatography. J. Dairy Sci. 88:3764–72.
Viallon, C., Martin, B., Verdier-Metz, I., Pradel, P., Garel, J.-P., Coulon, J.-B., and
Berdagué, J.-L. 2000. Transfer of monoterpenes and sesquiterpenes from forages
into milk fat. Le Lait 806:635–41.
Villeneuve, M.-P., Lebeuf, Y., Gervais, R., Tremblay, G., Vuillemard, J., Fortin, J., and
Chouinard, P. 2013. Milk volatile organic compounds and fatty acid profile in cows
fed timothy as hay, pasture, or silage. J. Dairy Sci. 96:7181–94.
Wong, N.P., Ellis, R., and LaCroix, D.E. 1975. Quantitative determination of lactones in
cheddar cheese. J. Dairy Sci. 58:1437–41.
Woo, A.H., Kollodges, S., and Lindsay, R.C. 1984. Modified techniques for improved
gas chromatographic quantification of free fatty acids in dairy foods. J. Dairy Sci.
67:1517–20.
Woo, A.H. and Lindsay, R.C. 1984. Concentrations of major free fatty acids and flavour
development in Italian cheese varieties. J. Dairy Sci. 67:960–8.
Yuceer, Y.K., Tuncel, B., Guneser, O., Engin, B., Isleten, M., Yasar, K., and Mendes, M.
2009. Characterization of aroma-active compounds, sensory properties, and prote-
olysis in ezine cheese. J. Dairy Sci. 92:4146–57.
Yvon, M., Berthelot, S., and Gripon, J.C. 1998. Adding α-ketoglutarate to semi-hard
cheese curd highly enhances the conversion of amino acids to aroma compounds.
Int. Dairy J. 8:889–98.
Yvon, M. and Rijnen, L. 2001. Cheese flavour formation by amino acid catabolism. Int.
Dairy J. 11:185–201.
 Chapter 24
Meat
Mónica Bueno, Thais Devincenzi, and Virginia Celia Resconi

CONTENTS

24.1 Introduction 487

24.2 Analysis of Aroma Compounds in Meat 489
24.2.1 Extraction Techniques 489
24.2.2 Sensory-Directed Location of Important Aromatic Compounds 490
24.2.3 Odorant Identification 491
24.2.4 Quantification and Comparative Studies 491
24.2.5 Hierarchy of Aroma Compounds 494
24.2.6 Overall Aroma Perception 495
24.3 The Most Relevant Aroma Compounds in Meat 495
24.4 Factors Influencing Meat Aroma 497
24.4.1 Animal Species 497
24.4.2 Production Factors 499
24.4.2.1 Breed 499
24.4.2.2 Sex 500
24.4.2.3 Age at Slaughter 500
24.4.2.4 Animal Feeding 501
24.4.3 Processing Factors 502
24.4.3.1 Bacterial Spoilage 502
24.4.3.2 Aging 503
24.4.3.3 Packaging Systems at Display 503
24.4.3.4 Freezing and Thawing 504
24.4.3.5 Cooking 504
24.4.3.6 Storage of Cooked Meat 505
References 505

24.1 INTRODUCTION

Food aroma plays a major role in the satisfaction gained from eating, and meat is no
exception. By contrast, some aromas are responsible for the aversion of consumers to
certain products. In the case of meat, a clear example of this is the boar taint in pork,
an unpleasant odor and flavor that was traditionally handled by surgical castration, but
this practice is now questioned by animal welfare organizations, so new on-farm strate-
gies are under development (Font-i-Furnols, 2012; Heyrman et al., 2018). Generally,
the focus of the beef sector to improve sensory quality has been more directed toward
tenderness, but consumers are starting to rate flavor as more important (Henchion,

487
488 Food Aroma Evolution

McCarthy, and Resconi, 2017), which may be more challenging to address. We all agree
that consumers generally prefer tender meat, but finding the flavor attributes they pre-
fer, their optimum intensity, and a balanced proportion between the others is com-
plex (O’Quinn et al., 2016), and there is no single flavor characteristic desirable by all
consumers. Therefore, the strategy should be in producing differentiated products with
unique flavors in order to satisfy different niches of consumers or single individuals who
wish to explore diverse experiences, such as when traveling and trying regional products
or when going to a restaurant or preparing a special meal at home. In this sense, studies
conducted to identify aromas (and tastes) responsible for meat flavors in a given product,
although laborious and expensive, help to find effective strategies for controlling unde-
sirable aromas and promoting desired ones. Although great analytical advances have
been achieved, more studies are still necessary to reveal the aromas of singular products
linked to production and processing factors. One study, for example, has shown that
the “spicy” aromas found in cooked lamb produced in Iceland were associated with
terpenoids, such as β-phellandrene and α-pinene, which arise from animal grazing on
an aromatic pasture which grows naturally in the region (Thorkelsson et al., 2009).
Moreover, the identification of aroma compounds linked to certain products, but also
applicable to volatiles with no odor impact, could be of particular interest in the search
for authentication methods (Devincenzi et al., 2014). For all these reasons, research into
meat aroma is still needed, especially directed toward the promotion of pleasant aromas,
to offer consumers high-quality meat with distinct and customized flavors (Aaslyng and
Meinert, 2017) (Figure 24.1).
This review first discusses the ways in which meat aroma analyses have been con-
ducted. Then, the ranking of key aroma compounds, revealed by gas chromatography
analysis, from different species is presented. And finally, variation in meat flavor by
means of production and processing factors is addressed.

FIGURE 24.1 Overview of meat aroma analysis.

 Meat 489

24.2 ANALYSIS OF AROMA COMPOUNDS IN MEAT

24.2.1 Extraction Techniques

Extraction is a major and critical issue in the analyses of solid samples, particularly in the
study of raw or cooked meat and, in general, it can be said that there are better and worse
alternatives, but, unfortunately, there is no perfect extraction methodology. Key compounds
can be lost or undergo transformations associated with enzyme activity, lipid oxidation, or
microbial growth during the extraction process. Therefore, for aroma studies, one of the
main objectives must be to obtain extracts representative of the headspace composition of
the samples. The most commonly applied methods of volatile isolation include dynamic
headspace analysis (DHS), solvent-assisted flavor evaporation (SAFE), and headspace solid-
phase microextraction (HS-SPME). In the case of meat products, other traditional sam-
pling methods are steam distillation solvent extraction (SDE) (Ramarathnam, Rubin, and
Diosady, 1991a, 1992b) and static headspace (SHS) (Ang and Liu, 1996; Wu, Hamouz, and
Schnepf, 1998). The aroma extracts obtained by SDE may lead to artifact formation due
to the elevated temperatures applied during distillation; whereas a disadvantage of SHS is
that a large volume of headspace has to be injected to obtain clear olfactometric signals.
Nevertheless, they can be used as complementary techniques of a total extraction strategy.
In DHS, also referred to as the “purge and trap” technique, aroma volatiles are purged
using a gentle stream of nitrogen and usually trapped in Tenax sorption traps (Thomas
et al., 2014b; Wu et al., 2015). Finally, the volatiles are thermally desorbed directly in
the chromatographic column. In this strategy, a new aliquot of sample is required in each
chromatographic analysis. Sometimes, liquid extracts are required, so other strategies were
developed to trap the volatiles in solid-phase extraction (SPE) cartridges of LiChrolut® EN
resins, which were then eluted with solvents (Resconi et al., 2010). This sorbent can be up
to 200 times more efficient than Tenax TA (Lopez et al., 2007), making it possible to purge
volatiles using a large stream of nitrogen over a very long period of time. As for drawbacks,
this strategy presents difficulties in the transfer of some polar compounds to the trap. On
the other hand, this type of strategy allows for some very interesting applications, such as
the capture of the aroma that emerges while eating by sampling the exhaled air from the
panelists consuming the meat (Bueno et al., 2011). Distillation in the SAFE apparatus is
among the most versatile methods used today in food flavor isolation. It has been applied
for the isolation of volatiles from multiple meat products such as sausages (Corral et al.,
2016), ham (Wang et al., 2018a), beef jerky (Jie et al., 2016), donkey (Tian et al., 2018),
duck (Strasser and Schieberle, 2014), and chicken (Duan et al., 2015). However, recoveries
for very highly volatile compounds can be better with other extraction techniques (Xu, Fan,
and Qian, 2007). SPME has been the technique of choice in many recent works, such as Sun
et al. (2018), Wang et al. (2018b), and Watkins et al. (2012), among others. This technique
is clean, very easy to use, and provides a good concentration of different volatiles in the
headspace. Nevertheless, the technique does not provide an extract, and on the other hand it
is quite difficult to optimize and validate and therefore to assess the reliability of the results.
Another important aspect prior to developing a SPME method is fiber selection. Different
meat research has demonstrated differences in the aromatic profile obtained depending
on the fiber used (Corral et al., 2016; Machiels and Istasse, 2003; Mansur et al., 2018).
Notwithstanding, for a general extraction of volatiles with different polarities, a divin​ylben​
zene/​carbo​xen/p​olydi​methy​lsilo​xane (DVB/CAR/PDMS) fiber coating can be a good choice
(Acevedo et al., 2012), meanwhile for the extraction of a specific group of compounds
490 Food Aroma Evolution

further studies must be carried out. For instance, a polydimethylsiloxane/divinylbenzene

(PDMS/DVB) fiber gives optimum results for the extraction of oxidation-related carbonyl
compounds in meat (Bueno et al., 2019). Moreover, different extraction techniques have
been compared for the extraction of meat volatile compounds. The HS-SPME method has
become popular because it provides wider and faster information compared to other meth-
ods (Liu, Xu, and Zhou, 2007; Rivas-Cañedo et al., 2011). The main conclusion of the
many diverse studies is that different techniques were complementary and necessary to
reproduce the overall meat aroma, such as SAFE and HS-SPME for goat aroma (Corral et
al., 2016) or HS-SPME and SDE for duck aroma (Liu, Xu, and Zhou, 2007). Another con-
clusion to be taken into account is that DHS and SPME methods provided better extraction
for volatiles with low molecular masses, while SDE extracted more high boiling and more
heavy volatiles (Liu, Xu, and Zhou, 2007; Madruga et al., 2009). In addition, in recent
years, new analytical procedures have been developed to shorten the time analysis, includ-
ing the use of direct mass spectrometry (MS). However, these procedures do not provide a
single liquid extract that can be used along the whole gas chromatography–olfactometry
(GC-O) process. Several direct MS techniques which avoid the chromatographic separation
have been applied to meat products, such as proton transfer reaction–MS (PTR-MS) (Holm
et al., 2013), selected ion flow tube–MS (SIFT-MS) (Carrapiso et al., 2015; Olivares et al.,
2012), and atmospheric pressure chemical ionization–MS (APCI-MS) (Carrapiso, 2007).

24.2.2 Sensory-Directed Location of Important Aromatic Compounds

Since all aroma molecules are more or less volatile, the technique that is best suited to
screen the odor-active molecules from the rest of the molecules is GC-O. This technique
makes use of the human nose as a detector for the compounds eluting out of the chromato-
graphic column. There are several different approaches for GC-O, differing in the way
in which the olfactometric signals are recorded and processed. These methods are com-
monly classified in dilution such as CHARM (combined hedonic aroma response method)
or AEDA (aroma extract dilution analysis), in time-intensity such as OSME, in detection
frequency such as NIF (nasal impact frequency), and in posterior intensity or frequency
and intensity combinations such as MF (modified frequency) calculation. This section will
briefly explain the most commonly used approach: AEDA and MF. In AEDA (Schieberle
and Grosch, 1987; Ullrich and Grosch, 1987), a concentrated aroma extract is analyzed to
identify odor peaks. The extract is then serially diluted 1:1 and re-analyzed until no odor is
detected within the dilution. The impact of an odor-active compound is given by its flavor
dilution factor (FD) value. The latter is calculated by dividing the largest volume analyzed
by the lowest volume in which the respective odor-active compound is still detectable. By
working properly (using a panel of judges and 1/10 dilutions), results are obtained with rea-
sonable effort. These results are mostly used for comparative purposes. MF combines both
odor detection frequency and odor intensity scoring in the same way, but the intensities are
scored using a prefixed scale. Moreover, data treatment takes into account the frequency of
reporting a note by the panel as adjusted by the intensities assigned to it (Dravnieks, 1985).

MF (%) = F (%) × I (%)

A minimum of six trained judges that smell the extract once is required in this method;
this is one of its main advantages, and it is also time-saving. On the other hand, in
 Meat 491

this method, as in all methods involving the use of a single dilution of the extract, it is
important to adequately choose the concentration factor for such an extract in order
to ensure that only a reduced number of odorants reach odor intensities approaching
the maximum on the scale. Nowadays, in general, sensory panels are commonly used,
which is crucial due to the documented existence of many odorants for which a part of
the population shows specific anosmia and the large differences in individual thresholds
(Ferreira and Cacho, 2009). Although these described methods still have limitations,
such as the non-consideration of odorant losses during the isolation procedure and of
synergistic or suppressive effects of distinct compounds in a flavor mixture, the great
number of published works relating to the GC-O analysis of meat (Table 24.1) reveals
that this technique can be used to effectively identify the key aroma compounds. GC-O
has been successfully used to determine the aroma profile of several meat-related matri-
ces, such as boiled, roasted, and fried beef, ham, sausages, turkey breast meat, and so
on (Zellner et al., 2008).

24.2.3 Odorant Identification

To identify the compounds responsible for an aroma, analysis by GC is mandatory.

Capillary gas chromatography provides the separation to isolate each one of the hundreds
of compounds that form the extract.
The basic steps in the identification of the key aroma compounds are the following:

• Standardization of retention times. Retention times of the important odor zones

have to be normalized by those of n-alkanes.
• Getting retention data from the important odor zones in a second chromato-
graphic column with a phase of different polarity.
• Running the extract on a GC-MS system.
• The latter two steps can be combined if the extract is running in a compre-
hensive two-dimensional gas chromatography coupled to mass spectrometry
(GC × GC-MS).
• Confirmation of candidates. This will be conducted by the injection of pure
standards.

24.2.4 Quantification and Comparative Studies

Often, the extract obtained for GC-O analysis to elucidate the impact odors cannot
be used for a robust quantification process. The quantification of aroma compounds in
meat presents several difficulties, since diverse groups constitute the aromatic substances,
some of these compounds are very reactive. These are found in different concentration
levels, and some are still unknown. For these reasons, it is absolutely necessary to use
an array of analytical methods and detectors for their determination. Some volatiles
can be determined by direct GC-MS using similar, already discussed pre-concentration
techniques (Section 24.2.1). However, there are analytes of interest that cannot be eas-
ily determined using a single non-selective-pre-concentration step. This happens when
the analytes are difficult to extract because they are very polar or very volatile. For
instance, different alternatives such as SPME or DHS have been proposed for the extrac-
tion of carbonyl compounds of low molecular weight such as hexanal, 1-octen-3-one,
TABLE 24.1 The Most Relevant Aroma Compounds Detected by GC-O in Meat
492

Aroma Number Overall Sheep/Lamb Beef Chicken/Hen Pork

Roast/
Rankinga of Papersb Rankingc Raw1 Pressure1 Grill2 Broth3 Boil4 Stew5 Grill6 Fried7 Broth8 Pressure9 Boil10 Grill11 Raw12 Broth13 Roast14

Methional 11 19 1 xx xx xx xxx xx xx xxx xx xx xx xx x xxx

Hexanal 12 19 2 xx x xx xxx xx x x xxx xx xx xx xxx
x
(E,E)-2,4-decadienal 7 16 3 xx xx xx xx xx xx xx xx xx xxx xxx xxx
(E)-2-nonenal 15 17 4 xx xx xx xx xxx x xx xx xx xxx xx
2-Methyl-3-furanthiol 4 12 5 xx xxx xxx xxx xxx xx x xxx xxx
Dimethyl trisulfide 13 12 6 xxx x x xx xx x xx
1-Octen-3-one 14 12 7 xx xx xx xx x xxx xx x xx
2-Furfurylthiol 6 10 8 xx xxx xx xxx xxx xxx x xxx
Furaneol 9 10 9 x xxx xx xxx xx xx xx xxx xxx xx
2,3-Butanedione 10 10 10 xx x xx xx xx xx xx xx xxx
Octanal 23 13 11 xxx xx x xx x xx xx xx
3-Methylbutanal 17 10 12 x xx xx xx x xx xxx
Heptanal 17 10 12 xx x xx xx x x xx
Food Aroma Evolution

(E,E)-2,4-nonadienal 27 12 13 xx x x x xx xx xx xx xx xx
Nonanal 28 14 13 x xx xx xx xx x xxx x
(E)-2-heptenal 18 9 14 xxx xx xx x x
Butanoic acid 19 9 15 xx x xx xx xxx xx x xxx xx
(E)-2-decenal 5 6 16 xxx xx xx x xx
2-Ethyl-3,(5/6)-dimethylpyrazine 21 9 17 xx xx xx xxx xx
1-Octen-3-ol 25 10 18
Phenylacetaldehyde 16 8 19 xxx xx xxx x xx
2-Methylbutanal 26 10 20 xx xx
2-Octanone 8 6 21 xx xx xx
(Continued )
TABLE 24.1 (Continued) The Most Relevant Aroma Compounds Detected by GC-O in Meat
Aroma Number Overall Sheep/Lamb Beef Chicken/Hen Pork

Roast/
Rankinga of Papersb Rankingc Raw1 Pressure1 Grill2 Broth3 Boil4 Stew5 Grill6 Fried7 Broth8 Pressure9 Boil10 Grill11 Raw12 Broth13 Roast14
Pentanal 22 8 22 xx xx x x xx xxx
2,5(and 2,6)-Dimethyl pyrazine 1 2 23 xxx
2,3-Diethyl-5-methylpyrazine 19 7 24 x xxx xx xxx
(E)-2-octenal 20 7 25 xx xx x xxx
2-Acetyl-2-thiazoline 24 8 26 xx xx xx xx xx xx xx xx
Ethanal 3 3 27 xxx xxx xx
(E,E)-2,4-octadienal 2 2 28 xxx xxx
(Z)-2-nonenal 16 6 28 xxx xx x xx x
a The aroma ranking was obtained by (1) recording a ranking of the aroma compounds within each study/type of meat (by the given values of FD fac-
tors, MF, detection frequency, odor intensity, etc.) from 1 (least important) to the maximum level given in the article (only odor zones referring to
individually identified compounds were considered); (2) rescaling the previous ranking from 1 (least important) to 3 (more important) within the
study/type of meat); (3) calculating the mean of the scores of each compound from all the papers (only considering values higher than 0). b Number
Meat

of papers from the 23 listed above, where each compound was detected as relevant by GC-O (papers were counted even if a compound was identified
together with others within an odor zone). c The Aroma Ranking was multiplied by an arbitrary factor according to the number of the paper where
the compound was cited in order to give a stronger weight to the odorants more cited; a factor of 1 was given for the minimum paper cited (1) and 3
to the maximum number of papers (19).
1 (Rota and Schieberle, 2005), 2 (Bravo-Lamas et al., 2018; Bueno et al., 2011; Resconi et al., 2010), 3 (Gasser and Grosch, 1990), 4 (Gasser and Grosch,
1988; Kerscher et al., 1997), 5(Guth and Grosch, 1994), 6 (Resconi et al., 2012a,b), 7 (Kerler and Grosch, 1996; Machiels et al., 2004, 2003; Specht
and Baltes, 1994), 8 (Fan et al., 2018; Feng et al., 2018; Gasser and Grosch, 1990; Zhang et al., 2018), 9 (Farkaš et al., 1997), 10 (Kerler and Grosch,
1997), 11(Aliani and Farmer, 2005b), 12 (Zareian et al., 2018), 13 (Zhao et al., 2017), 14 (Xie et al., 2008).
493
494 Food Aroma Evolution

or (E,E)-2,4-decadienal, most of them making use of the formation of derivatives with

pentafluorobenzylhydroxylamine (Bueno et al., 2013; Resconi et al., 2012a) or with pen-
tafluorophenyl hydrazine (Poole et al., 2016; Rochat and Chaintreau, 2005). In both
cases, the presence of fluorine atoms in the derivative favors the sensibility when the MS
source is operating in negative chemical ionization (NCI) (Zapata et al., 2010). Another
well-known example is the analysis of fatty acid methyl esters (FAMEs), which can be
determined by a gas chromatography-flame ionization detector (GC-FID). Readers are
encouraged to carefully read specific reviews for more information about methanolic
reagents used for FAME analysis (Murrieta, Hess, and Rule, 2003; Seppanen-Laakso,
Laakso, and Hiltunen, 2002). Other analytes require the use of highly sensitive and
selective detectors when they are present at very low levels. Volatile sulfur and nitro-
gen compounds in meat products, which are in the ppb to ppt range (Thomas et al.,
2014b), require the use of flame photometric (FPD), pulsed flame photometric (PFPD),
sulfur chemiluminescence (SCD), or nitrogen–phosphorus (NPD) detectors (Corral et
al., 2016; Fan, Sommers, and Sokorai, 2004). Regarding the presentation of final results,
different approaches are carried out. For liquid extracts that come from SAFE or DHS,
the addition of an internal standard (IS) could control the quantification process. Some
considerations must be taken into account to find an appropriate IS. For instance, the
IS must have similar volatility and similar distribution coefficients as the analytes, and
no coeluting with any other compound (Machiels et al., 2003) with the same mass-to-
charge ratio. Stable isotope dilution assays (SIDAs), adding different isotopically labeled
standards (Kosowska et al., 2018; Soellner and Schieberle, 2009), are the most suitable
solution, but when a large number of molecules are required, their high cost makes one
consider other options. In SPME analysis of solid meat samples, comparisons among
samples are usually found using just raw areas (Pérez et al., 2008; Saraiva et al., 2015)
or area percentages (Pia Gianelli et al., 2012). In the first case, there is no control over
changes related to fiber adsorption–desorption or to detector sensitivity. In the second
case, the problem arises when all the compounds vary in the same way and, therefore, no
differences between samples are observed (Bueno et al., 2013). Problems might be solved
by standard addition. However, in solid matrices, the mass-transfer mechanism can be
different for the standards added and the native analytes (Ouyang and Pawliszyn, 2008).
To overcome the difficulties, new procedures have been developed in recent years for the
direct analysis of solid meat samples. For example, Ioannidis et al. (2018) described a
headspace thermal desorption gas chromatography (HS-TD-GC-MS) in which sorbent
tubes used for sampling were initially loaded with a gaseous internal standard. Bueno et
al. (2019) developed an external calibration for SPME that uses ISs in the control solu-
tion, which provides extra control over the external response factor and also controls the
system (fiber, column, MS) in extended experiments.

24.2.5 Hierarchy of Aroma Compounds

Thousands of volatile compounds were identified as volatile constituents, but the use of
olfactometry methodologies and mass spectrometry contributed to the increase in knowl-
edge of the main odors present in meat. More precisely, the hierarchy of odor-active
compounds is based on the odor activity value (OAV) concept. OAV is defined as the
ratio of the volatile concentration to its odor detection threshold. OAVs > 1 indicates that
the compound contributes to the aroma profile of a food because it is present above its
threshold, so, the higher the OAV, the higher its contribution.
 Meat 495

24.2.6 Overall Aroma Perception

Since interaction between compounds released from the matrix might occur, the best
way to confirm that all the compounds identified and correctly quantified actually
contribute to the aroma in a food system is by an aroma reconstitution analysis. The
aroma of the recombination should match with the original extract according to a
sensory panel. Finally, compounds that truly have an impact on the aroma profile can
be determined by the systematic removal of each compound from the recombinant
in suppression studies, analyzing the tolerance of the overall aroma to small changes
in the concentration of each volatile. With this methodology, 12-methyltridecanal
which has been previously identified between the 10 most relevant odorants in boiled
and stewed beef by GC-O analyses (Guth and Grosch, 1994; Kerscher and Grosch,
1997), was found to not critically contribute to the overall aroma in a model system
(Grosch, 2001). In another study, meat was spiked with different concentrations of
the target compounds not present in the beef sample used and their contribution was
evaluated with a trained and a consumer panel (Prescott, Young, and O’Neill, 2001).
With this approach, branched-chain fatty acids (BCFA) were found as contributors of
barnyard/milky/sour/sheep meat flavors, whereas skatole gave grassy and sweet flavors.
Furthermore, BCFAs decreased the acceptability rates of Japanese consumers but did
not affect the scores of consumers more familiar with sheep meat from New Zealand
(Prescott, Young, and O’Neill, 2001).
Multivariate statistical tools such as principal component analysis (PCA) or partial
least squares regression (PLSR) are widely used to develop models and to study rela-
tionships between analytical data of specific aroma-active compounds and sensory data;
examples are found in studies on lamb, beef, chicken, and a meat product (Bueno et al.,
2013; Holm et al., 2010; Resconi et al., 2010, 2012b; Zhan et al., 2013; Zhang et al.,
2018). Nevertheless, learning about the role of the different components in the formation
of the aromatic concept and especially the interaction of families of compounds with
meat flavor (Bueno et al., 2014) opens a new field of study. Recently, the risk of dislike has
been calculated by combining consumer data in relation to meat concentration of skatole
and androstenone with the distribution of both compounds in the pig population with a
bivariate log-normal model (Christensen, Nielsen, and Aaslyng, 2019). With this model,
sorting limits of concentrations for the two compounds, deemed responsible for boar
taint, can be established. Therefore, classification of carcasses according to risk of dislike
can be settled. Great variability in consumer response was observed in this study, as it has
been in others (Panella-Riera et al., 2016), in part due to different odor sensitivities and
decisions based on those perceptions among consumers.

24.3 THE MOST RELEVANT AROMA COMPOUNDS IN MEAT

Table 24.1 shows the 31 aroma compounds most important in meat from ovine, bovine,
poultry, and porcine species, as revealed by the GC-O analysis published in 23 papers.
The compounds are ordered according to a ranking relating to their mean aroma impact
and the number of papers in which each compound was acknowledged. The papers ana-
lyzed used samples which were either raw or cooked in several ways (boiled meat, broth,
stewed juice, grilled meat, oven meat, fried meat), and aromas were collected during or after
cooking. The extraction procedures involved several strategies, including solvent distilla-
tion extraction (SDE), dynamic headspace—solid-phase extraction, comprising a modeled
496 Food Aroma Evolution

mouth-space and desorbed thermally or with solvents; static headspace (SHS)—SPE or

SPME, extraction-high vacuum distillation (EVD), solvent-assisted flavor evaporation, and
even an in vivo mouth-space. Further differences within each approach can be found, such
as the amount of sample, headspace volume, the shape of the extractions apparatus, sen-
sibility of the equipment, and sensory panel, expressed a unit of the odor relevance (FD
factors, MF, headspace volume, detection frequency, intensity, etc.). Therefore, global infor-
mation based on such methodologies makes us believe that the ranking presented is robust,
although direct comparisons should be carefully undertaken. It makes us believe that the
ranking presented is robust, although direct comparisons should be carefully undertaken.
The most relevant odor is methional, which is a Strecker aldehyde with sulfur, and the
amino acid precursor of which is methionine. This compound smells like cooked potato,
cooked vegetable, and meat (Bueno et al., 2011; Machiels, Istasse, and van Ruth, 2004;
Rota and Schieberle, 2005), has a low odor threshold, and was recognized as one of the
sulfur compounds that is most important to the aroma of beef meat when it has been oven
cooked (Rochat, Laumer, and Chaintreau, 2007), with other significant compounds (e.g.,
methanethiol, dimethyldisulfide) being derived from its breakdown (Resconi, Escudero,
and Campo, 2013). It was also found to be a key odor in an omission study of boiled beef
(Grosch, 2001). As shown in Table 24.1, methional is ubiquitous in several animal species
and cooking procedures, including raw sheep meat, but highlighted in beef broth. This
compound was detected in raw lamb as well, but only at a high temperature of extraction
(Vasta, Ratel, and Engel, 2007), and wasn’t reported in a comprehensive review of volatiles
assessed in raw meat (Casaburi et al., 2015). In cooked goat meat, Madruga et al. (2009),
found higher concentrations of this compound in SDE compared to DHS and SPME extrac-
tion procedures. Broths from ox and cow meat had higher FD factors of methional than
broth from chicken (Gasser and Grosch, 1990), and a higher concentration of this com-
pound was reported in beef and chicken, compared to pork (Kerscher and Grosch, 2000).
Methional also increased in cooked, refrigerated meat (Kerler and Grosch, 1996, 1997).
Hexanal is second in the ranking (Table 24.1), sharing the number of papers where it was
regarded as a relevant odorant in meat with methional (19 out of 23 papers consulted), but
having a lower mean odor impact. Since hexanal is a major product of lipid oxidation, and
this reaction takes place in raw, cooked, and stored meat from all the species, it is not sur-
prising it is highly cited. The odor characteristic of hexanal is described as grassy, green, and
fatty (Bravo-Lamas et al., 2018; Kerler and Grosch, 1996; Zareian et al., 2018). Whether
its contribution to the meat aroma is positive or negative depends on several factors, such
as the concentration, the food matrix (raw or cooked meat), and the type and concentration
of the rest of the odorants. An odor threshold of 4.5 ppb was described in water (Kerler
and Grosch, 1996), while it was 5870 ppb in a meat system (Brewer and Vega, 1995). In
raw meat, the amount of hexanal was reported to range from 0.14 to 94.43 ppb (Casaburi
et al., 2015), whereas in cooked meat it reached 269 ppb in beef and 1596 ppb in chicken,
increasing after storage to 2329 ppb and 11,332 ppb, respectively (Kerler and Grosch, 1996,
1997). Thus, its odor contribution, particularly in stored cooked meat, seems outstanding.
In raw meat stored under a high-oxygen atmosphere, pentanoic and hexanoic acids were
recently proposed to be better odor shelf-life markers in beef compared to hexanal and
other odor compounds usually present in raw meat (Resconi et al., 2018). After hexanal,
two other aldehydes, derived from lipid oxidation, were identified as the most relevant
aroma compounds: (E,E)-2,4-decadienal, described as having a rancid, meaty, fried and
fatty odor, and (E)-2-nonenal, described as having a fatty and tallowy odor (Bueno et al.,
2011; Gasser and Grosch, 1988). Since these two compounds (along with hexanal) originate
mainly from n-6 linoleic acid (Whitfield, 1992), meat with a higher concentration of such
 Meat 497

fatty acids should lead to higher concentrations of these compounds, as seen in the study
by Lorenz et al. (2002). However, the results are sometimes contradictory because meat
composition and reactions are complex and reactions are interlinked: n-3 polyunsaturated
fatty acids, for example, could initiate free radical oxidation of n-6 and n-9 acids (Elmore
et al., 2002); antioxidants such as vitamin E that are present in the meat independently of
the fatty acid composition can decrease overall lipid oxidation reactions (Resconi et al.,
2010); or the interactions of lipid oxidation between proteolysis, protein oxidation, and
the Maillard reaction; loss of volatiles; interactions between the meat matrix; and so on,
might take place (Estévez et al., 2003; Resconi et al., 2013). The lipid-derived aldehydes
(E,E)-2,4-decadienal, octanal, and nonanal, but not (E)- and (Z)-2-nonenal, were found
to be essential to the aroma of boiled meat in an omission experiment (Grosch, 2001). All
of these compounds were at higher concentrations in beef compared to pork and chicken
(Kerscher and Grosch, 2000). Another compound present in cooked meat, moreso in beef
than in other species, that evokes meaty notes is 2-methyl-furanthiol (Kerscher and Grosch,
2000), which is the key aroma in simulated beef flavorings (Moon, Cliff, and Li-Chan,
2006). This sulfur compound is thought to come mainly from the degradation of thiamine
or other pathways such as the Maillard reaction between cysteine and ribose, the Strecker
reaction from sulfur amino acids, and/or interactions among them (Cerny and Briffod,
2007; Güntert et al., 1990). Finally, in the top 10 in Table 24.1, the other two sulfur com-
pounds were present: dimethyl trisulfide and 2-furfutylthiol, the lipid oxidation–derived
ketone 1-octen-3-one, 2,3-butanedione (originated by bacterial and Maillard reaction), and
furaneol. Interestingly, this last compound was present in the mouth-space of consumers
but was not released into the headspace from meat while grilling (Bueno et al., 2011).

24.4 FACTORS INFLUENCING MEAT AROMA

Knowing the key aroma compounds and understanding the aspects that are involved in
the variation of their precursors (amino acids, peptides, reducing sugars, and lipids) gives
an important clue to identifying how factors influence meat aroma (Figure 24.2).
The postmortem modifications that occur in muscle are a source of variation for
aroma precursor content in meat products. After slaughter, the glycogen stored in mus-
cle is the main source of energy for maintaining homeostasis. Once the bloodstream
is interrupted, ATP synthesis occurs in an anaerobic pathway, and lactic acid starts to
accumulate, which consequently leads to a reduction of the pH values in muscular tis-
sue. Additionally, the postmortem proteolysis performed by calcium-dependent enzymes
results in the disruption of cell content and the release of several amino acids that are
precursors of aroma compounds. Moreover, there is a loss of antioxidant enzyme func-
tion, which favors oxidation processes. In meat, lipids are found in the cell membranes
(phospholipids) and as reserves in the myocytes and adipocytes (mainly triacylglycerol).
Both forms are important sources of aroma precursors and aroma reservoirs. In light of
the above, it seems reasonable to suggest that factors that affect meat composition and
structure are prone to affect aroma development and release from the meat when we eat.

24.4.1 Animal Species

The reactions between reducing sugars and amino acids, thiamine degradation, and lipid
oxidation are the main pathways in the formation of aroma compounds in cooked meat.
498 Food Aroma Evolution

FIGURE 24.2 Factors affecting the aroma development in meat. (Adapted from Erasmus,
Muller, and Hoffman, 2017.)

Therefore, the concentration of some precursors, which are variable between animal spe-
cies, could explain the species-dependent aroma in meat. Lamb meat, for example, has
a reducing sugar content that is lower than in pork and beef (Macy, Naumann, and
Bailey, 1964), which may lead to lower Maillard reaction products. Thiamine (vitamin
B1) concentration also contrasts between species. Thiamine is a water-soluble vitamin
that is very susceptible to thermal degradation and is responsible for the formation of
several sulfur aroma compounds such as 2-methyl-3-furantiol. The addition of thiamine
to cooked ham was directly linked with an increase in this key aroma (Thomas et al.,
2014a). However, although thiamine concentration in raw meat is about 0.08–0.11
mg/100 g in beef, 0.17–0.18 mg/100 g in lamb, 0.23 mg/100 g in chicken, and 0.81–0.88
mg/100 g in pork (Aliani and Farmer, 2005a; Badiani et al., 1998; Dawson, Unklesbay,
and Hedrick, 1988; Gerber, Scheeder, and Wenk, 2009), 2-methyl-3-furanthiol was pres-
ent in higher concentrations in beef compared to pork and chicken (Kerscher and Grosch,
2000). The presence of other components and the pH level influence the degradation of
this important precursor (Resconi et al., 2013). Other water-soluble compounds such
as inosine and inosine 5’-monophosphate (IMP) can also contribute to particular spe-
cies aroma. Madruga et al. (2010) found that inosine concentrations were four and five
times higher in goat meat than the values reported in beef (Koutsidis et al., 2008a) and
in chicken (Aliani et al., 2005a), respectively. Inosine is formed by the cleavage of IMP,
and it is suggested that the rates of these reactions are different between species, explain-
ing the differences described by Madruga et al. (2010). Both compounds are considered
precursors of meat aroma because they react with Strecker products such as hydrogen
sulfide and ammonia to form thiophenes and thiazoles. Lipid content and composition
 Meat 499

also differs between species and they are a source of species-specific aroma compounds.
For lamb meat, for example, branched-chain fatty acids, such as 4-ethyloctanoic acid,
that are stored on the triglycerides are related to sheep meat odor (Rota and Schieberle,
2005). The degree of unsaturation of the fatty acids in intramuscular fat is related to
the volatiles concentrations derived from lipid oxidation reactions (Elmore et al., 1999;
Resconi et al., 2013). Poultry and raw pork meat have higher levels of polyunsaturated
fatty acids than lamb or beef, for example (Calkins and Hodgen, 2007). However, the
typical aroma related to each species may be due to more quantitative rather than qualita-
tive differences in the volatile compounds between other species (Kerscher and Grosch,
2000; Resconi et al., 2013).

24.4.2 Production Factors

In general, livestock animals for meat production are selected and raised with the aim of
maximizing economic efficiency. The choices of the type of animal, gender, age, and car-
cass cover at slaughter or feeding systems are traditionally made according to the farmer
or industry expectations in accordance with the possibilities to access resources like land,
feed, genetics, and other technologies. However, differences in production factors could
be related to variations in meat qualities, such as composition and sensory characteristics,
including meat aroma.

24.4.2.1 Breed
Differences in intramuscular fat (IMF) could be one important source of variation in
the aroma compounds between breeds (though not the only one). In cattle, early matu-
rity breeds, such as Angus, Wagyu, and Holstein, deposit more lipids than late maturity
breeds, like Blonde d´Aquitane, Charolais, or Belgian Blue (Bonnet et al., 2010). In beef,
overall liking usually increases with the increase in IMF (Hunt et al., 2016). Recently,
highly marbled beef from Wagyu and Angus cattle (IMF>10%), has been highly valued
by a large number of consumers (Corbin et al., 2015; Legako et al., 2016). Besides the
mechanical role of IMF, the concentration of water-soluble compounds such as 2-methyl
propanal, 2/3-methylbutanal, and 2,3-butanedione increased when IMF is higher than
10%, while the concentration of fat-soluble and lipophilic volatile compounds (hexanal,
octanal, nonanal) decreased, as seen in mouth models (Frank et al., 2016) and in in vivo
mouth release (Frank et al., 2017), and this was associated with a more intense flavor
perception. Furthermore, while grilling the beef with the higher IMF, more carbonyl
compounds derived from lipid oxidation and Strecker/Maillard reactions were produced
since samples required more time to reach the same internal temperature than meat with
lower IMF, but lipid-derived compounds were also more independent of the cooking time
(Frank et al., 2017). In a study with beef cattle comparing Aberdeen Angus, Belgian Blue,
and Limousin steers, at the same age at slaughter and receiving the same diet, Machiels,
Istasse, and van Ruth (2004), with a model mouth apparatus with artificial saliva and
GC-O analysis, found that 21 odor-active compounds were significantly affected by the
breed. Ten of them were Maillard reaction products, five derived from lipids, and the
rest were not identified. Belgian Blue, the breed with the highest levels of crude pro-
tein and less intramuscular fat, developed a wider range of odor-active molecules such
as 2-octanone and 2,3-diethyl-5-methylpyrazine. The authors suggest that these differ-
ences could be due to the complexity of the muscle matrix and cross-reactions between
Maillard derived products and lipid degradation products. The effect of breeds in volatile
500 Food Aroma Evolution

compounds was also verified in lambs, comparing Soays and Suffolk breeds (Elmore et
al., 2000), and Scottish Blackface (SB) versus Texel×Scottish Blackface (Gkarane et al.,
2018). Fan et al. (2018) reported breed effects in chicken when comparing the aroma
compounds in broths from Beijing Youji, a native Chinese chicken breed, with the broth
obtained by fast-growing chickens. In pigs, Estévez et al. (2003) found a different volatile
profile in the cooked meat from Iberian pigs compared to lean pigs, which also differed
in the flavor deterioration of the stored meat. Different accumulations of skatole due to
genetics has also been suggested (Borrisser-Pairó et al., 2016).

24.4.2.2 Sex
The effect of sex in meat aroma compounds is very well studied, particularly in pigs.
The boar taint is an off-flavor that compromises swine meat and product acceptance
(Font-i-Furnols, 2012; Wauters et al., 2017). Together, skatole (3-methyl-indole) and a
sexual steroid, androstenone (5α-androst-16-en-3-one) are mainly responsible for boar
taint (Deslandes, Gariépy, and Houde, 2001) in all male pigs. Castration of piglets was
typically recommended for preventing the taint; however, this practice has been discour-
aged, as commented on in the introduction of this chapter. Nowadays, boar taint still
presents a risk of consumer rejection (Aluwé, Tuyttens, and Millet, 2015; Borrisser-Pairó
et al., 2016; Christensen, Nielsen, and Aaslyng, 2019). For ruminants, the effect of sex on
the aroma compounds is also reported. Gender differences could be related to the dynam-
ics and to the composition of adipose tissue deposition, which differs between males
and females of the same age (Bauchart and Thomas, 2010). Heifer IMF presents higher
contents of triglycerides and lower phospholipids than bulls, and this difference may
instigate changes in the composition of aroma compounds produced from lipid oxida-
tion, like aldehydes, alcohols, and ketones (Gorraiz et al., 2002). Moreover, the indirect
effects of testosterone can be involved in the sex-specific aromas in meat. Testosterone
could be associated with higher physical activity in uncastrated males, and therefore to
a lower glycogen stock in the muscle at slaughter and higher ultimate pH, which favors
the formation of thiazoles and pyrazines (Madruga and Mottram, 1995). In ovines, meat
from castrated animals reached higher flavor acceptability scores than rams (Gravador
et al., 2018) and differences in the proportion of several odorous volatile compounds
were found, including aldehydes, pyrazines, and 4-methyl-octanoic acid (Gkarane et al.,
2018). The last compound increased 13-fold in adipose tissue from rams compared to
wethers at 30 weeks of age in a previous study (Sutherland and Ames, 1996). Another
compound, 4,6-dimethyl-1,3-oxathiane, resembling a stale/wet animal odor, has also
been linked to non-castrated animals (Sutherland and Ames, 1995).

24.4.2.3 Age at Slaughter
The intensity of meat flavor usually increases with the age of the animals. After birth and
during growth until the maturity of the animal, the muscle characteristics alter its com-
position, metabolic profile, and oxidative capacity (Dingboom and Weijs, 2004). This
alteration in muscle composition modifies amino acid, peptide, reducing sugar, and lipid
contents, which are the main aroma precursors in meat aroma. Myoglobin, for example,
is found in higher concentration in the muscles of older animals. The heme-iron present
in this molecule is thought to be a pro-oxidant that promotes lipid oxidation and there-
fore the formation of volatile compounds such as aldehydes and ketones (Calkins and
Hodgen, 2007). Furthermore, precursor or flavorsome lipid-soluble compounds could
be accumulated in their fat throughout their life. The effect of age on meat aroma is
pronounced in ovines, which is related to the increase in branched-chain fatty acids, such
 Meat 501

as 4-ethyloctanoic acid (Rota and Schieberle, 2005), and could greatly explain the low
value of cull ewes. Recently, consumers rated meat from lambs slaughtered at 385 days
with stronger off-flavors than lambs slaughtered at 196 days (Gravador et al., 2018). It is
important to note that age at slaughter can sometimes be confounded with other factors
like growth rate or feeding systems. In cull beef cows reared on pastures, meat off-flavors
were reduced by finishing animals in grain (Stelzleni and Johnson, 2008). In pork, boar
taint is higher as the age/slaughter weight of the animal increases (Borrisser-Pairó et al.,
2016; Zamaratskaia et al., 2004).

24.4.2.4 Animal Feeding
The effect of animal feed on meat quality, sensory profile, and consumer liking is well
studied. During the last decade, efforts have also been made to elucidate the effects of
production systems on the aroma compounds of meat products. Animal feeding systems
can change the metabolic muscle status, and therefore glycogen stocks, and consequently
changes in final pH or in the amount of precursors available for Maillard reactions,
which could affect meat aroma (Braggins, 1996; Raes et al., 2003; Vestergaard, Oksbjerg,
and Henckel, 2000). Some aroma precursors are at the same time tastants (Imafidon
and Spanier, 1994). In poultry, a lower protein or energy level in the diet plus exercise
decreases free glutamine, which degrades the taste of the meat (Fujimura and Kadowaki,
2006). Steers finished with concentrates showed higher and lower concentrations in the
meat of sugars and amino acids, respectively, compared with animals fed with grass
silage (Koutsidis et al., 2008b). However, differences in volatile compounds of grilled
meat were more related to the fatty acid composition than to aroma precursor differences
(Elmore et al., 2004). According to Farmer, Kennedy, and Hagan (2009), the natural
concentrations of sugars, such as glucose and glucose-6-phosphate, need to be triplicated
to observe a significant change in the aroma of the cooked meat. However, in the study
by Koutsidis et al. (2008b), all diets were formulated as isoenergetic, meaning differences
in precursors could be more obvious in more extreme comparisons, so we cannot under-
estimate the relevance of these taste/aroma precursors. Animal diets can also affect the
amount of intramuscular fat, some consequences of which for meat flavor have already
been discussed, but changes in the fatty acid profile, particularly in monogastrics, were
also observed in ruminants. A more unsaturated fatty acid profile leads to a higher sus-
ceptibility to lipid oxidation, and the type of fatty acid present in the diet produces a
certain type of derived volatile. Feedstuffs can also be a source of antioxidants or even
some aroma compounds or precursors (Bou et al., 2001; Brodowska et al., 2016a; Fruet
et al., 2018; Schreurs et al., 2008; Wood et al., 2004). Grass vs. concentrate diets in rumi-
nants have been proven to change the proportion of some fatty acids, especially polyun-
saturated fatty acids (PUFA) like linolenic acid (C18:3 n-3), generally increasing when
animals were fed with pasture diets (Realini et al., 2004). On the other hand, typical
concentrate-rich diets present higher linoleic acid (C18:2 n-6), which is related to higher
concentrations of 1-octen-3-one, hexanal, (E)-2-nonenal, (Z)-2-nonenal, and (E,E)-2,4-
decadienal (Lorenz et al., 2002; Mezgebo et al., 2017). But concentrates could be for-
mulated with linseed, oils, and marine products like fish oils and algae, resulting in high
C18:3 n-3, C20:5 n ­ -3, and C22:6 n-3 fatty acid contents in meat, which easily undergo
oxidative reactions, and therefore, more (Z)-4-heptenal could be seen in the volatile frac-
tion compared to traditional concentrates (Elmore et al., 2000). Although the fatty acid
profile could be improved in terms of human nutrition with such ingredients, rancid or
fishy notes could be found in the meat, decreasing acceptability of the product, such as
has been reported for chicken meat (Jayasena et al., 2013). How a specific meat fatty acid
502 Food Aroma Evolution

profile could influence (or not) pork flavor attributes has been studied as well (Alonso et
al., 2012; Tikk et al., 2007). Diet is also implicated in the production of 3-methylindole,
or skatole, which is associated with a fecal odor and is especially important in ovine and
porcine meat aromas (Deslandes, Gariépy, and Houde, 2001; Devincenzi et al., 2014;
Young et al., 2003). Skatole is formed by the microbial deamination and decarboxylation
of tryptophan in the rumen of ruminants and in the caecum and colon of pigs (Deslandes,
Gariépy, and Houde, 2001; Schreurs et al., 2008). Legume-rich diets, like white clover
and alfalfa pastures, are associated with a greater concentration of skatole in ruminant
adipose tissue, particularly when diets present high contents of readily digestible proteins
(Devincenzi et al., 2019; Rousset-Akrim, Young, and Berdagué, 1997). On the other
hand, legumes rich in tannins, like Lotus corniculatus, can significantly decrease skatole
synthesis in rumen (Schreurs et al., 2007) and its concentration in meat (Rivaroli et al.,
2019). The inclusion of prebiotics from chicory roots or derived prebiotics (inulin and
oligofructose) has been proven to be an effective method of reducing this undesirable
aroma in pigs (Aluwé et al., 2017; Heyrman et al., 2018). Finally, diet can alter the aro-
matic profile of meat by a direct transfer of aroma compounds from the diet to the meat,
like terpenes, for example. Terpenes are a group of secondary plant metabolites and are
found at greater concentration and richness in dicotyledonous than in monocotyledonous
plants (Mariaca et al., 1997). Terpenes are not transformed by an animal’s metabolism;
therefore, the terpene profile of the meat is modulated by the terpene profile of the diet
(Prache, 2009). Nonetheless, differences in terpene concentration are found depending
on the tissue because these compounds are lipophilic and are accumulated in the adipo-
cytes (Serrano et al., 2007).

24.4.3 Processing Factors

Storage time and conditions are important factors that affect meat aroma. These factors
are important in raw meat because consumers should not be exposed to an unpleasant
smell when opening the package at home, but they also affect the aroma development
and acceptability of cooked meat. Therefore, knowing the mechanism involved in the
formation of aroma compounds during storage is indispensable. Spoilage caused by the
enzymatic activity of microorganisms during storage, the proteolysis of the muscle dur-
ing the aging process, and lipid oxidation are the main pathways that lead to a different
aroma profile during the storage of raw meat.

24.4.3.1 Bacterial Spoilage
Meat can be contaminated by animal microbiota, by the abattoir facilities, or by the pro-
cessing environment, and it is also prone to natural spoilage due to its physicochemical
composition. Differences in microbial population lead to different metabolic pathways
and result in different aroma compounds (Casaburi et al., 2015). The microbial com-
munity present in raw meat is dynamic and multifactorial (Lyte et al., 2016). It can vary
according to physicochemical conditions after slaughter (final pH, water content, cold
chain maintenance), from the origin of the meat (slaughter house plant), and from the
packaging atmosphere (Fraqueza, Ferreira, and Barreto, 2008; Pothakos et al., 2015;
Reis et al., 2016). The main aroma compounds formed by bacterial activity in raw meat
originate from glucose fermentation (ethanol, acetic acid, 3-hydroxy-2-butanone, and
2,3 butanediol) and from amino acid catabolism (3-methylbutanal 3-methyl-butanol,
2-methyl-butanol, and dimethyl-disulfide (Reis et al., 2016).
 Meat 503

24.4.3.2 Aging
The biochemical transformations that occur during postmortem proteolysis generally
have a positive impact on meat quality, especially in meats considered too firm, such as
beef and pork (Herrera-Mendez et al., 2006). During aging, tenderization of the meat
takes place, and the water-soluble aroma precursors like amino acids, sugars, nucleo-
tides, and free fatty acids increase (Hood and Allen, 1971; Koutsidis et al., 2008a;
Meinert et al., 2009). The effects of aging on the sensory attributes and aroma com-
pounds of raw (Casaburi et al., 2015; Reis et al., 2016; Resconi et al., 2018) and cooked
meat (Aaslyng and Meinert, 2017; Gorraiz et al., 2002; Monsón, Sañudo, and Sierra,
2005; Watanabe et al., 2015) have been described. Vacuum packaging, also known as
wet-aging, minimizes lipid oxidation and spoilage by aerobic bacteria. Nevertheless,
the occurrence of confinement odor is a risk for vacuum-packed raw meat (Johnson,
1991). This odor in wet-aged lamb meat is mainly associated with 3-methyl-butanal,
3-hydroxy-2-butanone, and sulfur dioxide, which are products from fermentative bac-
teria activities in raw meat (Reis et al., 2016). In wet-aged cooked meat, lipid oxidation
was promoted for up to 7 days, and this was regarded as beneficial in terms of flavor
(Gorraiz et al., 2002). In meat stored for longer (up to 30 days) and cooked at high
temperature, an increase in nitrogen-containing heterocyclic compounds and some
aldehydes suggests that other reactions such as Maillard and Strecker reactions are
promoted by aging the meat (Watanabe et al., 2015). Dry aging is becoming popular in
some butcheries and restaurants as it offers a distinguished product with a unique fla-
vor, even though it is a very costly technology (Dashdorj et al., 2016). Kim et al. (2016)
found higher glutamate levels (related to the umami taste) in dry-aged beef when com-
paring to wet-aged beef and suggested that this compound could play an important role
in consumer preference for dry-aged beef. However, even if dry-aging is a traditional
process of storing meat (Berger et al., 2018), studies identifying the volatile compounds
in dry-aged beef are still scarce. More than 30 days of dry aging are necessary to obtain
the “dry-aged” flavor and although more common in beef, it has also been explored in
pork (Aaslyng and Meinert, 2017).

24.4.3.3 Packaging Systems at Display

Packaging technologies were developed to keep meat acceptable for longer periods in a
convenient way for consumers. In red meat, the reductive environment that occurs under
vacuum packaging leads to changes in meat color, which are less appreciated by consum-
ers when purchasing. In this sense, modified atmosphere packaging (MAP) with high-
oxygen concentrations is interesting because it keeps the red coloration in beef during
display exposition, but oxygen-permeable overwrap is also commonly used for retail. In
high-oxygen MAP, however, aromatic compounds originating from lipid oxidation and
bacteria can enlarge the formation of the volatiles responsible for undesirable aromas,
like pentanoic, hexanoic, and heptanoic acids, 1-hexanol, (E)-2-undecenal, ethyl octano-
ate, 2-heptanone, and 2-pentylfuran (Resconi et al., 2018). In a recent review, the origins
of aroma compounds in raw meat according to the type of packaging and bacteria popu-
lation were discussed (Casaburi et al., 2015). Furthermore, rancid odors developed in
raw meat can persist and give an unacceptable rancid flavor to the cooked beef (Resconi
et al., 2012b). In fact, meat displayed without oxygen has an extended shelf life and bet-
ter quality at consumption (Aaslyng, Tørngren, and Madsen, 2010), and for that reason
some premium quality beef is nowadays sold in vacuum skin packs. Aroma transfer from
the packaging to the meat also needs to be considered.
504 Food Aroma Evolution

24.4.3.4 Freezing and Thawing

Freezing is one of the most applied methods for preserving meat in both domestic and
industrial environments. Few studies have reported the direct effects of freezing and thaw-
ing (methods, temperature, freezing rate, duration) on meat aroma profiles (Bueno et al.,
2011, 2013). However, clues about the effect of freezing and thawing can be hypothesized
because both processes alter meat composition and the structure of the cells. Freezing and
thawing change the water fraction of meat; therefore, the concentration of the solutes,
like proteins, carbohydrates, lipids, vitamins, and minerals, increases, and so potential
changes on the concentration of meat aroma precursors can be expected. Bueno et al.
(2013) found an absence of furaneol, a strong odorant compound, in cooked thawed
meat, which could be associated with a loss of its precursors when water is drained dur-
ing the thawing process. Furthermore, freezing and thawing promote oxidation reactions
(Brodowska et al., 2016b; Hansen et al., 2004; Muela et al., 2015; Pinheiro et al., 2019),
which will probably lead to a volatile profile with higher concentrations of aldehydes,
ketones, alcohols, and so on. Moreover, fermentation processes can occur during freez-
ing. This reaction was suggested by Bueno et al. (2011), who found higher concentrations
of butanoic and 3-methylbutanoic acids in frozen lamb meat while fresh lamb meat had
higher levels of pyrazines. However, beef can be stored under vacuum and frozen for up
to 90 days without any appreciable loss in the aroma when cooked (Vieira et al., 2009).
24.4.3.5 Cooking
Meat is generally consumed after a heating process that increases palatability but that
is also an effective way to inactivate pathogenic microorganisms, and therefore improve
its hygienic quality. Although some key odorants that exist or develop in raw meat can
impact on the aroma of cooked meat (Table 24.1), important chemical reactions are ther-
mally dependent; hence, variations in temperature, cooking method (grilled, roasted,
fried, broiled, etc.), and duration of cooking will significantly affect the total quantity
and type of aroma compounds present in the cooked meat (Domínguez et al., 2014).
Furthermore, the release of volatiles when we eat warm meat is higher compared to cold
meat. Cooking under higher temperatures leads to greater concentrations of Maillard-
derived compounds (Byrne et al., 2002; Meinert et al., 2007). This could be due to a
higher release of substrates for these reactions (Lorenzen et al., 2005) and to the fact that
high temperatures induce the formation of antioxidants, controlling the lipid oxidation
reaction (Byrne et al., 2002). This could explain why, out of the 31 compounds most
relevant to the aroma of meat mentioned earlier, three pyrazines predominate in roast-
ing, grilling and frying cooking methods (Table 24.1), although 2-ethyl-3,(5/6)-dimethyl
pyrazine was also found in chicken broth and boiled hen (Fan et al., 2018; Farkaš et
al., 1997). In well-done grilled pork meat, pyrazines are the main volatile compounds
obtained compared to grilling to a lower degree of doneness, or roasting and boiling
cooking methods (Mottram, 1985). More recently, in foal steaks with the same internal
temperature (70°C), pyrazines were only found after fried or grilled cooking procedures,
but not when roasted or microwaved, while the highest lipid oxidation product levels
were reached after roasting (Domínguez et al., 2014). Furthermore, even the kind of
stewpot can affect the aroma in meat. Because the rate of heat dispersion differs between
materials, the reactions that lead to aroma compound formation occur at different rates.
This fact can be confirmed by a recent study with chicken broth (Zhang et al., 2018). The
authors found different aroma profiles in chicken broth when it was cooked in different
types of stewpots (traditional clay stewpot, ceramic electrical stewpot, or in an electrical
stewpot with temperature modulations).
 Meat 505

24.4.3.6 Storage of Cooked Meat

The reheating of stored, cooked meat, even under refrigeration (4°C), can generate sev-
eral aroma impact compounds from lipid oxidation reactions. After reheating, hexanal
and trans-4,5-epoxy-(E)-2-decenal were increased (Kerler and Grosch, 1996), and were
related to the appearance of warmed-over flavors (WOF) in beef. In chicken patties,
Ferreira et al. (2016) also verified an increase of aroma compounds during pre-cooking,
chilling, and reheating, originating from lipid oxidation at each stage. In pork, the key
warmed-over flavors identified were pentanal, 2-pentylfuran, octanal, nonanal, 1-octen-
3-ol, and hexanal (O’Sullivan et al., 2003). Moreover, after reheating stored cooked meat,
there is a decrease in sulfur-containing compounds and furanones, which could reduce
the meaty flavor in reheated meat (Ferreira et al., 2016; Kerler and Grosch, 1996). The
temperature and type of cooking can affect the formation compounds involved in WOF
and even the meat size (Lepper-Blilie et al., 2014). Higher cooking temperatures prevent
the formation of these compounds, probably because of the formation of antioxidant
molecules like melanoidins and pre-melanoidins (Byrne et al., 2002). Feeding animals
with antioxidants was shown to reduce the problem (Yang et al., 2002) plus the use of
other processing conditions (vacuum, low temperature, active packaging, etc.).

REFERENCES

Aaslyng, M. D. and Meinert, L. (2017). Meat flavour in pork and beef—From animal to
meal. Meat Science, 132, 112–7.
Aaslyng, M. D., Tørngren, M. A., and Madsen, N. T. (2010). Scandinavian consumer
preference for beef steaks packed with or without oxygen. Meat Science, 85(3),
519–24.
Acevedo, C. A., Creixell, W., Pavez-Barra, C., Sanchez, E., Albornoz, F., and Young, M.
E. (2012). Modeling volatile organic compounds released by bovine fresh meat using
an integration of solid phase microextraction and databases. Food and Bioprocess
Technology, 5(6), 2557–67.
Aliani, M. and Farmer, L. J. (2005a). Precursors of chicken flavor. I. Determination
of some flavor precursors in chicken muscle. Journal of Agricultural and Food
Chemistry, 53(15), 6067–72.
Aliani, M. and Farmer, L. J. (2005b). Precursors of chicken flavor. II. Identification of
key flavor precursors using sensory methods. Journal of Agricultural and Food
Chemistry, 53(16), 6455–62.
Alonso, V., Najes, L. M., Provincial, L., Guillén, E., Gil, M., Roncalés, P., and Beltrán,
J. A. (2012). Influence of dietary fat on pork eating quality. Meat Science, 92(4),
366–73.
Aluwé, M., Heyrman, E., Theis, S., Sieland, C., Thurman, K., and Millet, S. (2017).
Chicory fructans in pig diet reduce skatole in back fat of entire male pigs. Research
in Veterinary Science, 115, 340–4.
Aluwé, M., Tuyttens, F. A. M., and Millet, S. (2015). Field experience with surgical
castration with anaesthesia, analgesia, immunocastration and production of entire
male pigs: Performance, carcass traits and boar taint prevalence. Animal, 9(3),
500–8.
Ang, C. Y. W. and Liu, F. (1996). Static headspace capillary GC method for determining
changes of volatiles from heated chicken breast meat. Journal of Muscle Foods, 7(1),
131–38.
506 Food Aroma Evolution

Badiani, A., Nanni, N., Gatta, P. P., Bitossi, F., Tolomelli, B., and Manfredini, M. (1998).
Nutrient content and retention in selected roasted cuts from 3-month-old ram lambs.
Food Chemistry, 61(1–2), 89–100.
Bauchart, D. and Thomas, A. (2010). Facteurs d’élevage et valeur santé des acides gras des
viandes musculaires. In: D. Bauchart and B. Picard, Muscle et Viande de Ruminant
(pp. 131–42). Versailles: Quae.
Berger, J., Kim, Y. H. B., Legako, J. F., Martini, S., Lee, J., Ebner, P., and Zuelly, S. M.
S. (2018). Dry-aging improves meat quality attributes of grass-fed beef loins. Meat
Science, 145, 285–91.
Bonnet, M., Cassar-Malek, I., Chilliard, Y., and Picard, B. (2010). Ontogenesis of muscle
and adipose tissues and their interactions in ruminants and other species. Animal,
4(7), 1093–109.
Borrisser-Pairó, F., Panella-Riera, N., Zammerini, D., Olivares, A., Garrido, M. D.,
Martínez, B., Gil, M., García-Regueiro, J. A., and Oliver, M. A. (2016). Prevalence
of boar taint in commercial pigs from Spanish farms. Meat Science, 111, 177–82.
Bou, R., Guardiola, F., Grau, A., Grimpa, S., Manich, A. M., Barroeta, A., and Codony,
R. (2001). Influence of dietary fat source, alpha-tocopherol, and ascorbic acid sup-
plementation on sensory quality of dark chicken meat. Poultry Science Association,
80, 800–7.
Braggins, T. J. (1996). Effect of stress-related changes in sheep meat ultimate pH on cooked
odor and flavor. Journal of Agricultural and Food Chemistry, 44(8), 2352–60.
Bravo-Lamas, L., Barron, L. J. R., Farmer, L., and Aldai, N. (2018). Fatty acid composi-
tion of intramuscular fat and odour-active compounds of lamb commercialized in
northern Spain. Meat Science, 139, 231–8.
Brewer, M. S. and Vega, J. D. (1995). Detectable odor thresholds of selected lipid oxida-
tion compounds in a meat model system. Journal of Food Science, 60(3), 592–5.
Brodowska, M., Guzek, D., Kołota, A., Głąbska, D., Górska-Horczyczak, E., Wojtasik,
I. M., and Wierzbicka, A. (2016a). Effect of diet on oxidation and profile of volatile
compounds of pork after freezing storage. Journal of Food and Nutrition Research,
55(1), 40–7.
Brodowska, M., Guzek, D., Kołota, A., Głąbska, D., Górska-Horczyczak, E., Wojtasik-
Kalinowska, I., and Wierzbicka, A. (2016b). Effect of diet on oxidation and profile
of volatile compounds of pork after freezing storage. Journal of Food and Nutrition
Research, 55(1), 40–7.
Bueno, M., Campo, M. M., Cacho, J., Ferreira, V., and Escudero, A. (2014). A model
explaining and predicting lamb flavour from the aroma-active chemical compounds
released upon grilling light lamb loins. Meat Science, 98(4), 622–8.
Bueno, M., Resconi, V. C., Campo, M. M., Cacho, J., Ferreira, V., and Escudero, A.
(2011). Gas chromatographic-olfactometric characterisation of headspace and
mouth space key aroma compounds in fresh and frozen lamb meat. Food Chemistry,
129(4), 1909–18.
Bueno, M., Resconi, V. C., Campo, M. M., Cacho, J., Ferreira, V., and Escudero, A.
(2013). Effect of freezing method and frozen storage duration on odor-active
compounds and sensory perception of lamb. Food Research International, 54(1),
772–80.
Bueno, M., Resconi, V. C., Campo, M. M., Ferreira, V., and Escudero, A. (2019).
Development of a robust HS-SPME-GC-MS method for the analysis of solid food
samples. Analysis of volatile compounds in fresh raw beef of differing lipid oxida-
tion degrees. Food Chemistry, 281, 49–56.
 Meat 507

Byrne, D. V., Bredie, W. L. P., Mottram, D. S., and Martens, M. (2002). Sensory and
chemical investigations on the effect of oven cooking on warmed-over flavour devel-
opment in chicken meat. Meat Science, 61(2), 127–39.
Calkins, C. R. and Hodgen, J. M. (2007). A fresh look at meat flavor. Meat Science,
77(1), 63–80.
Carrapiso, A. I. (2007). Effect of fat content on flavour release from sausages. Food
Chemistry, 103(2), 396–403.
Carrapiso, A. I., Noseda, B., Garcia, C., Reina, R., Sanchez del Pulgar, J., and Devlieghere,
F. (2015). SIFT-MS analysis of Iberian hams from pigs reared under different condi-
tions. Meat Science, 104, 8–13.
Casaburi, A., Piombino, P., Nychas, G.-J., Villani, F., and Ercolini, D. (2015). Bacterial
populations and the volatilome associated to meat spoilage. Food Microbiology,
45(Part A), 83–102.
Cerny, C. and Briffod, M. (2007). Effect of pH on the Maillard reaction of [13C5]
xylose, cysteine, and thiamin. Journal of Agricultural and Food Chemistry, 55(4),
1552–6.
Christensen, R. H., Nielsen, D. B., and Aaslyng, M. D. (2019). Estimating the risk of
dislike: An industry tool for setting sorting limits for boar taint compounds. Food
Quality and Preference, 71, 209–16.
Corbin, C. H., O’Quinn, T. G., Garmyn, A. J., Legako, J. F., Hunt, M. R., Dinh, T. T.
N., Rathmann, R. J., Brooks, J. C., and Miller, M. F. (2015). Sensory evaluation of
tender beef strip loin steaks of varying marbling levels and quality treatments. Meat
Science, 100, 24–31.
Corral, S., Leitner, E., Siegmund, B., and Flores, M. (2016). Determination of sulfur and
nitrogen compounds during the processing of dry fermented sausages and their rela-
tion to amino acid generation. Food Chemistry, 190, 657–64.
Dashdorj, D., Tripathi, V. K., Cho, S., Kim, Y., and Hwang, I. (2016). Dry aging of beef:
Review. Journal of Animal Science and Technology, 58, 20 .
Dawson, K. R., Unklesbay, N. F., and Hedrick, H. B. (1988). HPLC determination of
riboflavin, niacin, and thiamin in beef, pork, and lamb after alternate heat-process-
ing methods. Journal of Agricultural and Food Chemistry, 36(6), 1176–9.
Deslandes, B., Gariépy, C., and Houde, A. (2001). Review of microbiological and bio-
chemical effects of skatole on animal production. Livestock Production Science,
71(2), 193–200.
Devincenzi, T., Delfosse, O., Andueza, D., Nabinger, C., and Prache, S. (2014). Dose-
dependent response of nitrogen stable isotope ratio to proportion of legumes in diet
to authenticate lamb meat produced from legume-rich diets. Food Chemistry, 152,
456–61.
Devincenzi, T., Prunier, A., Meteau, K., and Prache, S. (2019). How does barley supple-
mentation in lambs grazing alfalfa affect meat sensory quality and authentication?
Animal, 13(2), 427–34.
Dingboom, E. G. and Weijs, W. A. (2004). The effect of growth and exercise on muscle
characteristics in relation to meat quality. In: M. F. W. t. Pas, M. E. Everts, and H. P.
Haagsman, Muscle Development of Livestock Animals: Physiology, Genetics, and
Meat Quality. CABI Publishing.
Domínguez, R., Gómez, M., Fonseca, S., and Lorenzo, J. M. (2014). Influence of thermal
treatment on formation of volatile compounds, cooking loss and lipid oxidation in
foal meat. LWT—Food Science and Technology, 58(2), 439–45.
Dravnieks, A., ed. (1985). Atlas of Odor Character Profiles. Philadelphia, PA: ASTM.
508 Food Aroma Evolution

Duan, Y., Zheng, F., Chen, H., Huang, M., Xie, J., Chen, F., and Sun, B. (2015). Analysis
of volatiles in Dezhou Braised Chicken by comprehensive two-dimensional gas chro-
matography/high resolution-time of flight mass spectrometry. LWT—Food Science
and Technology, 60(2, Part 2), 1235–42.
Elmore, J. S., Campo, M. M., Enser, M., and Mottram, D. S. (2002). Effect of lipid
composition on meat-like model systems containing cysteine, ribose, and polyun-
saturated fatty acids. Journal of Agricultural and Food Chemistry, 50(5), 1126–32.
Elmore, J. S., Mottram, D. S., Enser, M., and Wood, J. D. (1999). Effect of the polyun-
saturated fatty acid composition of beef muscle on the profile of aroma volatiles.
Journal of Agricultural and Food Chemistry, 47(4), 1619–25.
Elmore, J. S., Mottram, D. S., Enser, M., and Wood, J. D. (2000). The effects of diet and
breed on the volatile compounds of cooked lamb. Meat Science, 55(2), 149–59.
Elmore, J. S., Warren, H. E., Mottram, D. S., Scollan, N. D., Enser, M., Richardson, R.
I., and Wood, J. D. (2004). A comparison of the aroma volatiles and fatty acid com-
positions of grilled beef muscle from Aberdeen Angus and Holstein-Friesian steers
fed diets based on silage or concentrates. Meat Science, 68, 27–33.
Erasmus, S. W., Muller, M., and Hoffman, L. C. (2017). Authentic sheep meat in the
European Union: Factors influencing and validating its unique meat quality. Journal
of the Science of Food and Agriculture, 97(7), 1979–96.
Estévez, M., Morcuende, D., Ventanas, S., and Cava, R. (2003). Analysis of volatiles
in meat from Iberian pigs and lean pigs after refrigeration and cooking by using
SPME-GC-MS. Journal of Agricultural and Food Chemistry, 51(11), 3429–35.
Fan, M., Xiao, Q., Xie, J., Cheng, J., Sun, B., Du, W., Wang, Y., and Wang, T. (2018).
Aroma compounds in chicken broths of Beijing Youji and commercial broilers.
Journal of Agricultural and Food Chemistry, 66(39), 10242–51.
Fan, X. T., Sommers, C. H., and Sokorai, K. J. B. (2004). Ionizing radiation and anti-
oxidants affect volatile sulfur compounds, lipid oxidation, and color of ready-to-eat
Turkey Bologna. Journal of Agricultural and Food Chemistry, 52(11), 3509–15.
Farkaš, P., Sádecká, J., Kováč, M., Siegmund, B., Leitner, E., and Pfannhauser, W. (1997).
Key odourants of pressure-cooked hen meat. Food Chemistry, 60(4), 617–21.
Farmer, L. J., Kennedy, J., and Hagan, T. (2009). Contribution of aqueous precursors
to the odour and flavour of cooked meats. In: ICoMST. Copenhague, Dinamarca.
Feng, Y., Cai, Y., Fu, X., Zheng, L., Xiao, Z., and Zhao, M. (2018). Comparison of
aroma-active compounds in broiler broth and native chicken broth by aroma extract
dilution analysis (AEDA), odor activity value (OAV) and omission experiment. Food
Chemistry, 265, 274–80.
Ferreira, V. and Cacho, J. (2009). Identification of impact odorants in wine. In: M. V.
Moreno Arribas and M. C. Polo, Wine Chemistry and Biochemistry (pp. 393–415).
New York, NY: Springer Science + Business Media.
Ferreira, V. C. S., Morcuende, D., Madruga, M. S., Hernández-López, S. H., Silva, F. A.
P., Ventanas, S., and Estévez, M. (2016). Effect of pre-cooking methods on the chem-
ical and sensory deterioration of ready-to-eat chicken patties during chilled storage
and microwave reheating. Journal of Food Science and Technology, 53(6), 2760–9.
Font-i-Furnols, M. (2012). Consumer studies on sensory acceptability of boar taint: A
review. Meat Science, 92(4), 319–29.
Frank, D., Ball, A., Hughes, J., Krishnamurthy, R., Piyasiri, U., Stark, J., Watkins, P., and
Warner, R. (2016). Sensory and flavor chemistry characteristics of Australian beef:
Influence of intramuscular fat, feed, and breed. Journal of Agricultural and Food
Chemistry, 64(21), 4299–311.
 Meat 509

Frank, D., Kaczmarska, K., Paterson, J., Piyasiri, U., and Warner, R. (2017). Effect of
marbling on volatile generation, oral breakdown and in mouth flavor release of
grilled beef. Meat Science, 133, 61–8.
Fraqueza, M. J., Ferreira, M. C., and Barreto, A. S. (2008). Spoilage of light (PSE-like)
and dark turkey meat under aerobic or modified atmosphere package: Microbial
indicators and their relationship with total volatile basic nitrogen AU—Fraqueza,
M.J. British Poultry Science, 49(1), 12–20.
Fruet, A. P. B., Trombetta, F., Stefanello, F. S., Speroni, C. S., Donadel, J. Z., De Souza, A.
N. M., Rosado Júnior, A., Tonetto, C. J., Wagner, R., De Mello, A., and Nörnberg,
J. L. (2018). Effects of feeding legume-grass pasture and different concentrate levels
on fatty acid profile, volatile compounds, and off-flavor of the M. longissimus tho-
racis. Meat Science, 140, 112–8.
Fujimura, S. and Kadowaki, M. (2006). Improvement of meat taste by dietary compo-
nents. Bulletin of the Faculty of Agriculture, Niigata University, 58(2), 151–3.
Gasser, U. and Grosch, W. (1988). Identification of volatile flavour compounds with high
aroma values from cooked beef. Zeitschrift für Lebensmittel-Untersuchung und
Forschung, 186, 489–94.
Gasser, U. and Grosch, W. (1990). Primary odorants of chicken broth. A comparative
study with meat broths from cow and ox. Zeitschrift für Lebensmittel-Untersuchung
und Forschung, 190, 3–8.
Gerber, N., Scheeder, M. R. L., and Wenk, C. (2009). The influence of cooking and
fat trimming on the actual nutrient intake from meat. Meat Science, 81(1),
148–54.
Gkarane, V., Brunton, N. P., Harrison, S. M., Gravador, R. S., Allen, P., Claffey, N.
A., Diskin, M. G., Fahey, A. G., Farmer, L. J., Moloney, A. P., and Monahan, F. J.
(2018). Volatile profile of grilled lamb as affected by castration and age at slaughter
in two breeds. Journal of Food Science, 83(10), 2466–77.
Gorraiz, C., Beriain, M. J., Chasco, J., and Insausti, K. (2002). Effect of aging time on
volatile compounds, odor, and flavor of cooked beef from Pirenaica and Friesian
bulls and Heifers. Journal of Food Science, 67(3), 916–22.
Gravador, R. S., Pace, E., Mooney, B. R., Jaeger, S. R., Gkarane, V., Fahey, A. G., Brunton,
N. P., Claffey, N. A., Allen, P., Diskin, M. G., Moloney, A. P., Farmer, L. J., and
Monahan, F. J. (2018). A consumer study of the effect of castration and slaugh-
ter age of lambs on the sensory quality of meat. Small Ruminant Research, 169,
148–53.
Grosch, W. (2001). Evaluation of the key odorants of foods by dilution experiments,
aroma models and omission. Chemical Senses, 26(5), 533–45.
Güntert, M., Brüning, J., Emberger, R., Köpsel, M., Kuhn, W., Thielmann, T., and
Werkhoff, P. (1990). Identification and formation of some selected sulfur-containing
flavor compounds in various meat model systems. Journal of Agricultural and Food
Chemistry, 38(11), 2027–41.
Guth, H. and Grosch, W. (1994). Identification of the character impact odorants of stewed
beef juice by instrumental analyses and sensory studies. Journal of Agricultural and
Food Chemistry, 42(12), 2862–6.
Hansen, E., Juncher, D., Henckel, P., Karlsson, A., Bertelsen, G., and Skibsted, L. H.
(2004). Oxidative stability of chilled pork chops following long term freeze storage.
Meat Science, 68(3), 479–84.
Henchion, M. M., McCarthy, M., and Resconi, V. C. (2017). Beef quality attributes: A
systematic review of consumer perspectives. Meat Science, 128, 1–7.
510 Food Aroma Evolution

Herrera-Mendez, C. H., Becila, S., Boudjellal, A., and Ouali, A. (2006). Meat ageing:
Reconsideration of the current concept. Trends in Food Science & Technology,
17(8), 394–405.
Heyrman, E., Millet, S., Tuyttens, F. A. M., Ampe, B., Janssens, S., Buys, N., Wauters,
J., Vanhaecke, L., and Aluwé, M. (2018). On farm intervention studies on reduction
of boar taint prevalence: Feeding strategies, presence of gilts and time in lairage.
Research in Veterinary Science, 118, 508–16.
Holm, E. S., Adamsen, A. P. S., Feilberg, A., Schäfer, A., Lokke, M. M., and Petersen,
M. A. (2013). Quality changes during storage of cooked and sliced meat products
measured with PTR-MS and HS-GC-MS. Meat Science, 95(2), 302–10.
Holm, E. S., Schäfer, A., Skov, T., Koch, A. G., and Petersen, M. A. (2010). Identification
of chemical markers for the sensory shelf-life of saveloy. Meat Science, 90(2),
314–22.
Hood, R. L. and Allen, E. (1971). Influence of sex and postmortem aging on intramuscu-
lar and subcutaneous bovine lipids. Journal of Food Science, 36(5), 786–90.
Hunt, M. R., Legako, J. F., Dinh, T. T. N., Garmyn, A. J., O’Quinn, T. G., Corbin, C.
H., Rathmann, R. J., Brooks, J. C., and Miller, M. F. (2016). Assessment of volatile
compounds, neutral and polar lipid fatty acids of four beef muscles from USDA
Choice and Select graded carcasses and their relationships with consumer palatabil-
ity scores and intramuscular fat content. Meat Science, 116, 91–101.
Imafidon, G. I. and Spanier, A. M. (1994). Unraveling the secret of meat flavor. Trends in
Food Science & Technology, 5(10), 315–21.
Ioannidis, A. G., Walgraeve, C., Vanderroost, M., Van Langenhove, H., Devlieghere, F.,
and De Meulenaer, B. (2018). Non-destructive measurement of volatile organic com-
pounds in modified atmosphere packaged poultry using SPME-SIFT-MS in tandem
with headspace TD-GC-MS. Food Analytical Methods, 11(3), 848–61.
Jayasena, D. D., Ahn, D. U., Nam, K. C., and Jo, C. (2013). Flavour chemistry of chicken
meat: A review. Asian-Australasian Journal of Animal Sciences, 26(5), 732–42.
Jie, S., Dan-dan, P., Hai-tao, C., Bao-guo, S., and Yu-yu, Z. (2016). Analysis of volatile
components in spiced beef jerky by SAFE-GC-MS. Science and Technology of Food
Industry, 17, 281–7.
Johnson, B. Y. (1991). Weep and Other Characteristics of Stored and Transported
Chilled Meat. Australia: CSIRO Food & Nutritional Sciences.
Kerler, J. and Grosch, W. (1996). Odorants contributing to Warmed-Over Flavor (WOF)
of refrigerated cooked beef. Journal of Food Science, 61(6), 1271–5.
Kerler, J. and Grosch, W. (1997). Character impact odorants of boiled chicken: Changes
during refrigerated storage and reheating. Zeitschrift für Lebensmitteluntersuchung
und -Forschung A, 205(3), 232–8.
Kerscher, R. and Grosch, W. (1997). Comparative evaluation of potent odorants of
boiled beef by aroma extract dilution and concentration analysis. Zeitschrift für
Lebensmitteluntersuchung und -Forschung A, 204(1), 3–6. Freising, Germany: Dt.
Forschungsanstalt für Lebensmittelchemie.
Kerscher, R. and Grosch, W. (2000). Comparison of the aromas of cooked beef, pork and
chicken. In: P. Schieberle and K. Engel, Frontiers of Flavour Science (pp. 17–20).
Kim, Y. H. B., Kemp, R., and Samuelsson, L. M. (2016). Effects of dry-aging on meat
quality attributes and metabolite profiles of beef loins. Meat Science, 111, 168–76.
Kosowska, M., Majcher, M. A., Jelen, H. H., and Fortuna, T. (2018). Key aroma com-
pounds in smoked cooked loin. Journal of Agricultural and Food Chemistry, 66(14),
3683–90.
 Meat 511

Koutsidis, G., Elmore, J. S., Oruna-Concha, M. J., Campo, M. M., Wood, J. D., and
Mottram, D. S. (2008a). Water-soluble precursors of beef flavour. Part II: Effect of
post-mortem conditioning. Meat Science, 79(2), 270–7.
Koutsidis, G., Elmore, J. S., Oruna-Concha, M. J., Campo, M. M., Wood, J. D., and
Mottram, D. S. (2008b). Water-soluble precursors of beef flavour: I. Effect of diet
and breed. Meat Science, 79(1), 124–30.
Legako, J. F., Dinh, T. T. N., Miller, M. F., Adhikari, K., and Brooks, J. C. (2016).
Consumer palatability scores, sensory descriptive attributes, and volatile compounds
of grilled beef steaks from three USDA Quality Grades. Meat Science, 112, 77–85.
Lepper-Blilie, A. N., Berg, E. P., Buchanan, D. S., Keller, W. L., Maddock-Carlin, K. R.,
and Berg, P. T. (2014). Effectiveness of oxygen barrier oven bags in low temperature
cooking on reduction of warmed-over flavor in beef roasts. Meat Science, 96(3),
1361–4.
Liu, Y., Xu, X. L., and Zhou, G. H. (2007). Comparative study of volatile compounds
in traditional Chinese Nanjing marinated duck by different extraction techniques.
International Journal of Food Science and Technology, 42(5), 543–50.
Lopez, P., Batlle, R., Nerin, C., Cacho, J., and Ferreira, V. (2007). Use of new gen-
eration poly(styrene-divinylbenzene) resins for gas-phase trapping-thermal desorp-
tion—Application to the retention of seven volatile organic compounds. Journal of
Chromatography A, 1139(1), 36–44.
Lorenz, S., Buettner, A., Ender, K., Nürnberg, G., Papstein, H.-J., Schieberle, P., and
Nürnberg, K. (2002). Influence of keeping system on the fatty acid composition
in the longissimus muscle of bulls and odorants formed after pressure-cooking.
European Food Research and Technology, 214(2), 112–8.
Lorenzen, C. L., Davuluri, V. K., Adhikari, K., and Grün, I. U. (2005). Effect of end-
point temperature and degree of doneness on sensory and instrumental flavor profile
of beefsteaks. Journal of Food Science, 70(2), S113–8.
Lyte, J. M., Legako, J. F., Martin, J. N., Thompson, L., Surowiec, K., and Brooks, J.
C. (2016). Volatile compound characterization of modified atmosphere packaged
ground beef held under temperature abuse. Food Control, 59, 1–6.
Machiels, D. and Istasse, L. (2003). Evaluation of two commercial solid-phase micro-
extraction fibres for the analysis of target aroma compounds in cooked beef meat.
Talanta, 61(4), 529–37.
Machiels, D., Istasse, L., and van Ruth, S. M. (2004). Gas chromatography-olfactometry
analysis of beef meat originating from differently fed Belgian Blue, Limousin and
Aberdeen Angus bulls. Food Chemistry, 86(3), 377–83.
Machiels, D., van Ruth, S. M., Posthumus, M. A., and Istasse, L. (2003). Gas chroma-
tography-olfactometry analysis of the volatile compounds of two commercial Irish
beef meats. Talanta, 60(4), 755–64.
Macy, R. L., Naumann, H. D., and Bailey, M. E. (1964). Water-soluble flavor and odor
precursors of meat. II. Effects of heating on amino nitrogen constituents and car-
bohydrates in lyophilized diffusates from aqueous extracts of beef, pork, and lamb.
Journal of Food Science, 29(2), 142–8.
Madruga, M. S., Elmore, J. S., Dodson, A. T., and Mottram, D. S. (2009). Volatile flavour
profile of goat meat extracted by three widely used techniques. Food Chemistry,
115(3), 1081–7.
Madruga, M. S., Elmore, J. S., Oruna-Concha, M. J., Balagiannis, D., and Mottram, D.
S. (2010). Determination of some water-soluble aroma precursors in goat meat and
their enrolment on flavour profile of goat meat. Food Chemistry, 123(2), 513–20.
512 Food Aroma Evolution

Madruga, M. S. and Mottram, D. S. (1995). The effect of pH on the formation of Maillard-

derived aroma volatiles using a cooked meat system. Journal of the Science of Food
and Agriculture, 68(3), 305–10.
Mansur, A. R., Lee, H. J., Choi, H. K., Lim, T. G., Yoo, M., Jang, H. W., and Nam, T. G.
(2018). Comparison of two commercial solid-phase microextraction fibers for the
headspace analysis of volatile compounds in different pork and beef cuts. Journal of
Food Processing and Preservation, 42(10), 1–14.
Mariaca, R. l. G., Berger, T. F. H., Gauch, R., Imhof, M. I., Jeangros, B., and Bosset,
J. O. (1997). Occurrence of volatile mono- and Sesquiterpenoids in highland and
lowland plant species as possible precursors for flavor compounds in milk and dairy
products. Journal of Agricultural and Food Chemistry, 45(11), 4423–34.
Meinert, L., Andersen, L. T., Bredie, W. L. P., Bjergegaard, C., and Aaslyng, M. D.
(2007). Chemical and sensory characterisation of pan-fried pork flavour: Interactions
between raw meat quality, ageing and frying temperature. Meat Science, 75(2),
229–42.
Meinert, L., Tikk, K., Tikk, M., Brockhoff, P. B., Bredie, W. L. P., Bjergegaard, C., and
Aaslyng, M. D. (2009). Flavour development in pork. Influence of flavour precur-
sor concentrations in longissimus dorsi from pigs with different raw meat qualities.
Meat Science, 81(1), 255–62.
Mezgebo, G. B., Monahan, F. J., McGee, M., O’Riordan, E. G., Richardson, I. R.,
Brunton, N. P., and Moloney, A. P. (2017). Fatty acid, volatile and sensory char-
acteristics of beef as affected by grass silage or pasture in the bovine diet. Food
Chemistry, 235, 86–97.
Monsón, F., Sañudo, C., and Sierra, I. (2005). Influence of breed and ageing time on the
sensory meat quality and consumer acceptability in intensively reared beef. Meat
Science, 71(3), 471–9.
Moon, S.-Y., Cliff, M. A., and Li-Chan, E. C. Y. (2006). Odour-active components of
simulated beef flavour analysed by solid phase microextraction and gas chromatog-
raphy-mass spectrometry and -olfactometry. Food Research International, 39(3),
294–308.
Mottram, D. S. (1985). The effect of cooking conditions on the formation of volatile
heterocyclic compounds in pork. Journal of the Science of Food and Agriculture,
36(5), 377–82.
Muela, E., Monge, P., Sañudo, C., Campo, M. M., and Beltrán, J. A. (2015). Meat quality
of lamb frozen stored up to 21months: Instrumental analyses on thawed meat dur-
ing display. Meat Science, 102, 35–40.
Murrieta, C. M., Hess, B. W., and Rule, D. C. (2003). Comparison of acidic and alkaline
catalysts for preparation of fatty acid methyl esters from ovine muscle with emphasis
on conjugated linoleic acid. Meat Science, 65(1), 523–9.
Olivares, A., Dryahina, K., Spanel, P., and Flores, M. (2012). Rapid detection of lipid
oxidation in beef muscle packed under modified atmosphere by measuring volatile
organic compounds using SIFT-MS. Food Chemistry, 135(3), 1801–8.
O’Quinn, T. G., Woerner, D. R., Engle, T. E., Chapman, P. L., Legako, J. F., Brooks,
J. C., Belk, K. E., and Tatum, J. D. (2016). Identifying consumer preferences for
specific beef flavor characteristics in relation to cattle production and postmortem
processing parameters. Meat Science, 112, 90–102.
O’Sullivan, M. G., Byrne, D. V., Jensen, M. T., Andersen, H. J., and Vestergaard, J.
(2003). A comparison of warmed-over flavour in pork by sensory analysis, GC/MS
and the electronic nose. Meat Science, 65(3), 1125–38.
 Meat 513

Ouyang, G. F. and Pawliszyn, J. (2008). A critical review in calibration methods for solid-
phase microextraction. Analytica Chimica Acta, 627(2), 184–97.
Panella-Riera, N., Blanch, M., Kallas, Z., Chevillon, P., Garavaldi, A., Gil, M., Gil, J.
M., Font-i-Furnols, M., and Oliver, M. A. (2016). Consumers’ segmentation based
on the acceptability of meat from entire male pigs with different boar taint levels in
four European countries: France, Italy, Spain and United Kingdom. Meat Science,
114, 137–45.
Pérez, R. A., Rojo, M. D., González, G., and De Lorenzo, C. (2008). Solid-phase microex-
traction for the determination of volatile compounds in the spoilage of raw ground
beef. Journal of AOAC International, 91(6), 1409–15.
Pia Gianelli, M., Salazar, V., Mojica, L., and Friz, M. (2012). Volatile compounds pres-
ent in traditional meat products (charqui and longaniza sausage) in Chile. Brazilian
Archives of Biology and Technology, 55(4), 603–12.
Pinheiro, R. S. B., Francisco, C. L., Lino, D. M., and Borba, H. (2019). Meat quality of Santa
IInês lamb chilled-then-frozen storage up to 12 months. Meat Science, 148, 72–8.
Poole, J. J., Grandy, J. J., Gómez-Rios, G. A., Gionfriddo, E., and Pawliszyn, J. (2016).
Solid Phase microextraction on-fiber derivatization using a stable, portable, and
reusable pentafluorophenyl hydrazine standard gas generating vial. Analytical
Chemistry, 88(13), 6859–66.
Pothakos, V., Devlieghere, F., Villani, F., Björkroth, J., and Ercolini, D. (2015). Lactic
acid bacteria and their controversial role in fresh meat spoilage. Meat Science, 109,
66–74.
Prache, S. (2009). Diet authentication in sheep from the composition of animal tissues
and products. Revista Brasileira de Zootecnia, 38, 362–70.
Prescott, J., Young, O., and O’Neill, L. (2001). The impact of variations in flavour com-
pounds on meat acceptability: A comparison of Japanese and New Zealand con-
sumers. Food Quality and Preference, 12(4), 257–64.
Raes, K., Balcaen, A., Dirinck, P., De Winne, A., Claeys, E., Demeyer, D., and De Smet,
S. (2003). Meat quality, fatty acid composition and flavour analysis in Belgian retail
beef. Meat Science, 65(4), 1237–46.
Ramarathnam, N., Rubin, L. J., and Diosady, L. L. (1991a). Studies on meat flavor. 2.
A quantitative investigation of the volatile carbonyls and hydrocarbons in uncured
and cured beef and chicken. Journal of Agricultural and Food Chemistry, 39(10),
1839–47.
Ramarathnam, N., Rubin, L. J., and Diosady, L. L. (1991b). Studies on meat flavor.
1. Qualitative and quantitative differences in uncured and cured pork. Journal of
Agricultural and Food Chemistry, 39(2), 344–50.
Realini, C. E., Duckett, S. K., Brito, G. W., Dalla Rizza, M., and De Mattos, D. (2004).
Effect of pasture vs. concentrate feeding with or without antioxidants on car-
cass characteristics, fatty acid composition, and quality of Uruguayan beef. Meat
Science, 66(3), 567–77.
Reis, M. M., Reis, M. G., Mills, J., Ross, C., and Brightwell, G. (2016). Characterization
of volatile metabolites associated with confinement odour during the shelf-life of
vacuum packed lamb meat under different storage conditions. Meat Science, 113,
80–91.
Resconi, V., Campo, M. d. M., Montossi, F., Ferreira, V., Sañudo, C., and Escudero, A.
(2012a). Gas chromatographic-olfactometric aroma profile and quantitative analy-
sis of volatile carbonyls of grilled beef from different finishing feed systems. Journal
of Food Science, 77(6), S240–6.
514 Food Aroma Evolution

Resconi, V., Escudero, A., and Campo, M. (2013). The development of aromas in rumi-
nant meat. Molecules, 18(6), 6748.
Resconi, V. C., Bueno, M., Escudero, A., Magalhaes, D., Ferreira, V., and Campo, M. M.
(2018). Ageing and retail display time in raw beef odour according to the degree of
lipid oxidation. Food Chemistry, 242, 288–300.
Resconi, V. C., Campo, M. M., Montossi, F., Ferreira, V., Sañudo, C., and Escudero, A.
(2010). Relationship between odour-active compounds and flavour perception in
meat from lambs fed different diets. Meat Science, 85(4), 700–6.
Resconi, V. C., Escudero, A., Beltrán, J. A., Olleta, J. L., Sañudo, C., and Campo, M.
M. (2012b). Color, lipid oxidation, sensory quality and aroma compounds of beef
steaks displayed under different levels of oxygen in a modified atmosphere package.
Journal of Food Science, 77(1), S10–8.
Rivaroli, D., Prunier, A., Meteau, K., do Prado, I. N., and Prache, S. (2019). Tannin-rich
sainfoin pellet supplementation reduces fat volatile indoles content and delays diges-
tive parasitism in lambs grazing alfalfa. Animal, 13(9), 1–8.
Rivas-Cañedo, A., Juez-Ojeda, C., Nuñez, M., and Fernández-García, E. (2011). Volatile
compounds in ground beef subjected to high pressure processing: A comparison
of dynamic headspace and solid-phase microextraction. Food Chemistry, 124(3),
1201–7.
Rochat, S. and Chaintreau, A. (2005). Carbonyl odorants contributing to the in-oven
roast beef top note. Journal of Agricultural and Food Chemistry, 53(24), 9578–85.
Rochat, S., Laumer, J.-Y. d. S., and Chaintreau, A. (2007). Analysis of sulfur compounds
from the in-oven roast beef aroma by comprehensive two-dimensional gas chroma-
tography. Journal of Chromatography A, 1147(1), 85–94.
Rota, V. and Schieberle, P. (2005). Changes in key odorants of sheep meat induced
by cooking. In: Food Lipids, vol. 920 (pp. 73–83). Washington, DC: American
Chemical Society.
Rousset-Akrim, S., Young, O. A., and Berdagué, J. L. (1997). Diet and growth effects
in panel assessment of sheepmeat odour and flavour. Meat Science, 45(2), 169–81.
Saraiva, C., Oliveira, I., Silva, J. A., Martins, C., Ventanas, J., and Garcia, C. (2015).
Implementation of multivariate techniques for the selection of volatile compounds as
indicators of sensory quality of raw beef. Journal of Food Science and Technology-
Mysore, 52(6), 3887–98.
Schieberle, P. and Grosch, W. (1987). Evaluation of the flavor of wheat and rye bread
by aroma extract dilution analysis. Zeitschrift für Lebensmittel-Untersuchung und
Forschung, 185(2), 111–3.
Schreurs, N. M., Lane, G. A., Tavendale, M. H., Barry, T. N., and McNabb, W. C.
(2008). Pastoral flavour in meat products from ruminants fed fresh forages and its
amelioration by forage condensed tannins. Animal Feed Science and Technology,
146(3–4), 193–221.
Schreurs, N. M., McNabb, W. C., Tavendale, M. H., Lane, G. A., Barry, T. N., Cummings,
T., Fraser, K., López-Villalobos, N., and Ramírez-Restrepo, C. A. (2007). Skatole
and indole concentration and the odour of fat from lambs that had grazed peren-
nial ryegrass/white clover pasture or Lotus corniculatus. Animal Feed Science and
Technology, 138(3–4), 254–71.
Seppanen-Laakso, T., Laakso, I., and Hiltunen, R. (2002). Analysis of fatty acids by gas
chromatography, and its relevance to research on health and nutrition. Analytica
Chimica Acta, 465(1–2), 39–62.
 Meat 515

Serrano, E., Cornu, A., Kondjoyan, N., Figueredo, G., Agabriel, J., and Micol, D. (2007).
Terpene accumulation in muscle and fatty tissues of calves supplemented with essen-
tial oils. Journal of Animal and Feed Science, 16(2), 168–79.
Soellner, K. and Schieberle, P. (2009). Decoding the key aroma compounds of a Hungarian-
type salami by molecular sensory science approaches. Journal of Agricultural and
Food Chemistry, 57(10), 4319–27.
Specht, K. and Baltes, W. (1994). Identification of volatile flavor compounds with high
aroma values from shallow-fried beef. Journal of Agricultural and Food Chemistry,
42(10), 2246–53.
Stelzleni, A. M. and Johnson, D. D. (2008). Effect of days on concentrate feed on sen-
sory off-flavor score, off-flavor descriptor and fatty acid profiles for selected muscles
from cull beef cows. Meat Science, 79(2), 382–93.
Strasser, S. and Schieberle, P. (2014). Characterization of the key aroma compounds in
roasted duck liver by means of aroma extract dilution analysis: Comparison with
beef and pork livers. European Food Research and Technology, 238(2), 307–13.
Sun, Y., Fu, M., Li, Z., and Peng, X. (2018). Evaluation of freshness in determination
of volatile organic compounds released from pork by HS-SPME-GC-MS. Food
Analytical Methods, 11(5), 1321–9.
Sutherland, M. M. and Ames, J. M. (1995). The effect of castration on the headspace
aroma components of cooked lamb. Journal of the Science of Food and Agriculture,
69(4), 403–13.
Sutherland, M. M. and Ames, J. M. (1996). Free fatty acid composition of the adipose
tissue of intact and castrated lambs slaughtered at 12 and 30 weeks of age. Journal
of Agricultural and Food Chemistry, 44(10), 3113–6.
Thomas, C., Mercier, F., Tournayre, P., Martin, J.-L., and Berdagué, J.-L. (2014a).
Identification and origin of odorous sulfur compounds in cooked ham. Food
Chemistry, 155, 207–13.
Thomas, C., Mercier, F., Tournayre, P., Martin, J. L., and Berdagué, J. L. (2014b).
Identification and origin of odorous sulfur compounds in cooked ham. Food
Chemistry, 155, 207–13.
Thorkelsson, G., Jonsdottir, R., Hilmarsson, O. T., Olafsdottir, A., and Martinsdottir,
E. (2009). The influence of grazing time on Angelica archangelica on volatile com-
pounds and sensory quality of meat from pasture lambs. In: ICoMST. Copenhague,
Denmark.
Tian, X., Dan, W., Mingjuan, M., Haitao, C., Baoguo, S., Ning, Z., and Yuyu, Z. (2018).
Identification of flavor-active compounds in spiced donkey meat by odor activity
value (OVA) calculation and gas chromatography-olfactometry-mass spectrometry.
Food Science, China, 39(8), 123–8.
Tikk, K., Tikk, M., Aaslyng, M. D., Karlsson, A. H., Lindahl, G., and Andersen, H.
J. (2007). Significance of fat supplemented diets on pork quality—Connections
between specific fatty acids and sensory attributes of pork. Meat Science, 77(2),
275–86.
Ullrich, F. and Grosch, W. (1987). Identification of the most intense volatile flavour com-
pounds formed during autoxidation of linoleic acid. Zeitschrift für Lebensmittel-
Untersuchung und -Forschung, 184(4), 277–82.
Vasta, V., Ratel, J., and Engel, E. (2007). Mass spectrometry analysis of volatile com-
pounds in raw meat for the authentication of the feeding background of farm ani-
mals. Journal of Agricultural and Food Chemistry, 55(12), 4630–9.
516 Food Aroma Evolution

Vestergaard, M., Oksbjerg, N., and Henckel, P. (2000). Influence of feeding intensity,
grazing and finishing feeding on muscle fibre characteristics and meat colour of
semitendinosus, longissimus dorsi and supraspinatus muscles of young bulls. Meat
Science, 54(2), 177–85.
Vieira, C., Diaz, M. T., Martínez, B., and García-Cachán, M. D. (2009). Effect of frozen stor-
age conditions (temperature and length of storage) on microbiological and sensory qual-
ity of rustic crossbred beef at different states of ageing. Meat Science, 83(3), 398–404.
Wang, W., Feng, X., Zhang, D., Li, B. i., Sun, B., Tian, H., and Liu, Y. (2018a). Analysis
of volatile compounds in Chinese dry-cured hams by comprehensive two-dimen-
sional gas chromatography with high-resolution time-of-flight mass spectrometry.
Meat Science, 140, 14–25.
Wang, X., Zhu, L., Han, Y., Xu, L., Jin, J., Cai, Y., and Wang, H. (2018b). Analysis of
volatile compounds between raw and cooked beef by HS-SPME–GC–MS. Journal
of Food Processing and Preservation, 42(2), e13503.
Watanabe, A., Kamada, G., Imanari, M., Shiba, N., Yonai, M., and Muramoto, T. (2015).
Effect of aging on volatile compounds in cooked beef. Meat Science, 107, 12–9.
Watkins, P. J., Rose, G., Warner, R. D., Dunshea, F. R., and Pethick, D. W. (2012). A
comparison of solid-phase microextraction (SPME) with simultaneous distillation-
extraction (SDE) for the analysis of volatile compounds in heated beef and sheep
fats. Meat Science, 91(2), 99–107.
Wauters, J., Verplanken, K., Vercruysse, V., Ampe, B., Aluwé, M., and Vanhaecke,
L. (2017). Sensory evaluation of boar meat products by trained experts. Food
Chemistry, 237, 516–24.
Whitfield, F. B. (1992). Volatiles from interactions of Maillard reactions and lipids.
Critical Reviews in Food Science and Nutrition, 31(1/2), 1–58.
Wood, J. D., Nute, G. R., Richardson, R. I., Whittington, F. M., Southwood, O., Plastow,
G., Mansbridge, R., da Costa, N., and Chang, K. C. (2004). Effects of breed, diet
and muscle on fat deposition and eating quality in pigs. Meat Science, 67(4), 651–67.
Wu, H., Zhuang, H., Zhang, Y., Tang, J., Yu, X., Long, M., wang, J., and Zhang, J.
(2015). Influence of partial replacement of NaCl with KCl on profiles of volatile
compounds in dry-cured bacon during processing. Food Chemistry, 172, 391–99.
Wu, Y., Hamouz, F., and Schnepf, M. (1998). Factors affecting static headspace-gas chro-
matographic analysis of lipid oxidation in precooked meat. Journal of Agricultural
and Food Chemistry, 46(9), 3677–82.
Xie, J., Sun, B., Zheng, F., and Wang, S. (2008). Volatile flavor constituents in roasted
pork of Mini-pig. Food Chemistry, 109(3), 506–14.
Xu, Y., Fan, W., and Qian, M. C. (2007). Characterization of aroma compounds in apple
cider using solvent-assisted flavor evaporation and headspace solid-phase microex-
traction. Journal of Agricultural and Food Chemistry, 55(8), 3051–7.
Yang, A., Brewster, M. J., Beilken, S. L., Lanari, M. C., Taylor, D. G., and Tume, R. K.
(2002). Warmed-over flavor and lipid stability of beef: Effects of prior nutrition.
Journal of Food Science, 67(9), 3309–13.
Young, O. A., Lane, G. A., Priolo, A., and Fraser, K. (2003). Pastoral and species flavour
in lambs raised on pasture, lucerne or maize. Journal of the Science of Food and
Agriculture, 83(2), 93–104.
Zamaratskaia, G., Babol, J., Andersson, H., and Lundström, K. (2004). Plasma skatole
and androstenone levels in entire male pigs and relationship between boar taint com-
pounds, sex steroids and thyroxine at various ages. Livestock Production Science,
87(2), 91–8.
 Meat 517

Zapata, J., Mateo-Vivaracho, L., Cacho, J., and Ferreira, V. (2010). Comparison of
extraction techniques and mass spectrometric ionization modes in the analysis of
wine volatile carbonyls. Analytica Chimica Acta, 660(1–2), 197–205.
Zareian, M., Böhner, N., Loos, H. M., Silcock, P., Bremer, P., and Beauchamp, J. (2018).
Evaluation of volatile organic compound release in modified atmosphere-packaged
minced raw pork in relation to shelf-life. Food Packaging and Shelf Life, 18, 51–61.
Zellner, B. A., Dugo, P., Dugo, G., and Mondello, L. (2008). Gas chromatography-
olfactometry in food flavour analysis. Journal of Chromatography A, 1186(1–2),
123–43.
Zhan, P., Tian, H., Zhang, X., and Wang, L. (2013). Contribution to aroma character-
istics of mutton process flavor from the enzymatic hydrolysate of sheep bone pro-
tein assessed by descriptive sensory analysis and gas chromatography olfactometry.
Journal of Chromatography B-Analytical Technologies in the Biomedical and Life
Sciences, 921, 1–8.
Zhang, M., Chen, X., Hayat, K., Duhoranimana, E., Zhang, X., Xia, S., Yu, J., and
Xing, F. (2018). Characterization of odor-active compounds of chicken broth and
improved flavor by thermal modulation in electrical stewpots. Food Research
International, 109, 72–81.
Zhao, J., Wang, M., Xie, J., Zhao, M., Hou, L., Liang, J., Wang, S., and Cheng, J. (2017).
Volatile flavor constituents in the pork broth of black-pig. Food Chemistry, 226,
51–60.
 Chapter 25
Fish
Asghar Amanpour, Gamze Guclu, and Serkan Selli

CONTENTS

Abbreviations 519
25.1 Introduction 520
25.2 Fish Aroma 521
25.2.1 Fresh Fish Aroma 522
25.2.2 Effect of Postmortem Alterations on Fish Aroma 522
25.2.3 Effect of Fish Species on Fish Aroma 523
25.2.4 Effect of Environmental Conditions and Seasonal Impacts on
Fish Aroma 525
25.2.5 Effect of pH Levels on Fish Aroma 526
25.2.6 Effect of Fat/Oil Amount on Fish Aroma 526
25.2.7 Effect of Storage Conditions on Fish Aroma 527
25.2.7.1 Effect of Microbial Degradation on Fish Aroma 528
25.2.7.2 Effect of Lipid Oxidation on Fish Aroma 533
25.2.8 Effect of Processing on Fish Aroma 533
25.2.8.1 Effect of the Ripening Process on Fish Aroma 534
25.2.8.2 Effect of Smoking Process on Fish Aroma 535
25.2.8.3 Effect of the Cooking Process on Fish Aroma 536
25.3 Conclusion 537
References 537

ABBREVIATIONS

CHD: coronary heart disease

CVD: cardiovascular disease
GC-MS: gas chromatography-mass spectroscopy
UFA: unsaturated fatty acids
PUFA: polyunsaturated fatty acids
ATP: adenosinetriphosphate
Hx: hypoxhanthin
SSO: specific spoilage organisms
NPN: non-protein nitrogen-containing compounds
TMAO: trimethylamineoxide
TMA: trimethyamine
DMA: dimethylamine
DMS: dimethyl sulfide
EPA: eicosa-5,8,ll,14,17-pentaenoic acid

519
520 Food Aroma Evolution

H2S: hydrogen sulfide

TVC: total viable counts
TVB-N: total volatile bases-nitrogen
Cu: copper
Fe: iron
OAVs: odor activity values

25.1 INTRODUCTION

Fish or fishery products are consumed all around the world due not only to their nutri-
tional advantages to health but also to their distinguishable and delicate flavors. The
health and nutritional importance of fishery products originated in a study on the Eskimos
which showed that despite their fat-rich consumption based on seafood they had lower
rates of coronary heart disease (CHD) and less risk of cancer. Further epidemiological
investigations have also asserted that people who eat fishery products have less risk of
cardiovascular disease (CVD) than those who did not (Shahidi and Cadwallader, 1997).
Moreover, more studies have exhibited that fresh and high-quality fish consumption also
provides positive benefits for medical conditions such as hypertension, obesity, CVD,
some cancers, and constipation (Mohan et al., 2016). Apart from the health benefits,
fresh fish includes a proper balance of lipid and lipid-derived compounds, free amino
acids, protein, nucleotides, carbohydrates, and organic acids contributing to the distinct
and elegant flavor of seafood, which makes it different from other meat flavors (Shahidi
and Cadwallader, 1997). Unfortunately, fresh fish is highly perishable due to enzymatic
and chemical reactions, microbial activities, improper handling and storage conditions,
and far shorter shelf life than terrestrial meats (Ramezani et al., 2015). In order to con-
serve the quality of fresh fishery products during their preparation, storage, and distribu-
tion, some strategies comprising super or deep chilling, cooling, freezing, salting, drying,
chemical treatments, cooking, smoking, canning, packaging, gamma irradiation, high-
pressure treatment, and a pulsed electric field have been reported (Sampels, 2015; Mohan
et al., 2016).
Fish quality can be interpreted as combining the properties of freshness, integrity,
and wholesomeness (Martin, 1988). Healthiness is related to the quality of a food in
terms of being suitable to eat, uncontaminated, and packed and stored in a hygienic con-
dition. Integrity is linked with the quality of a food product being appropriate to the sup-
pliers’ claims. Freshness is related to the quality of appearance, taste, and texture in food
products. In fact, fish quality is a complicated concept entailing an entire scope of issues
in view of the consumers, including quality, nutrition, safety, accessibility, freshness,
eating quality, physical properties, size, integrity, convenience, and the type of product
(Bisogni et al., 1987; Botta, 1995; Olafsdottir et al., 1997a; Bremner, 2000). Nutritional
and sensory features along with freshness are three vital properties indicating the quality
of fish flesh as appreciated by consumers (Grigorakis, 2007).
Both nutritional and organoleptic values in fish are fundamentally connected with its
chemical composition, which is subsequently dependent on numerous quality influencing
issues, namely, inherent fish features like species, age, and sex; environmental parameters
such as temperature and salinity; dietary ingredient history including the composition
of diet and ratio of feeding; and harvesting and post-harvesting processes (Huss, 1995;
Grigorakis, 2007). As a consequence, fish quality is dependent on the complex interaction
of multiple different factors (Caggiano, 2000; Grigorakis, 2007).
 Fish 521

The nutritional composition of a variety of fish species has previously been thor-
oughly reviewed in detail (Robertson, 2009; Rehbein and Oehlenschlager, 2009). Apart
from the nutritional contents of fish and fishery products, the flavor is of great impor-
tance. Flavor is usually defined as the sensation emerging from the interaction of signals
formed as a result of recognizing smell and taste from a food product (Ho and Hartman,
1994; Laing and Jinks, 1996). It is mostly made up of the volatiles distinguished by the
nose, not only orthonasally but also retronasally, nonvolatile compounds recognized on
the tongue, and compounds that are perceived in the mouth as texture or mouthfeel.
Aroma is considered more remarkable than taste in defining flavor (Hognadottir, 2000).
Carbohydrates, proteins, and lipids are the key resources of food flavor. From them,
lipids can substantially contribute to the formation of food flavor. Lipids result in rancid-
ity, oxidized, and stale flavors in plenty of fat-rich foodstuffs and are considered to be
responsible for a warmed-over meat flavor. Furthermore, they can be accountable for the
delightful flavors in plenty of deep-fried foodstuffs, dairy products, vegetables, and fruit
(Ho and Hartman, 1994).
This chapter chiefly emphasizes the effect of postmortem alterations, fish species,
environmental conditions, lipids, storage conditions, microbial degradation, and process-
ing conditions on fish aroma.

25.2 FISH AROMA

Fishery products possess complicated flavor structures consisting of correspondingly sig-

nificant aroma, taste-, and aroma-active compounds. The taste-active compounds, which
mostly come from nonvolatile compounds, for example, nucleotides, free amino acids,
mineral salts, and sugars, have been paid substantial attention and their significance to
seafood flavor is clearly documented by many different studies (Pigott, 1990; Whitfield,
1990; Lindsay, 1994; Kawai and Sakaguchi, 1996; Shahidi and Cadwallader, 1997).
Aroma is one of the vital factors determining fish and fishery product quality. Flavor
investigation into fresh seafood has been given considerable attention owing to the signif-
icance of aroma to consumer satisfactoriness and preferences in fresh seafood (Lindsay,
1994; Pigott, 1990). The overall aroma of fish species is the result of the combined effect
of many different volatiles with quantities varying between nanograms and milligrams.
Lipoxygenase-derived lipid-based volatiles may predominantly result in the fresh seafood
aroma profile. Nevertheless, apart from enzymatic pathways, amines, including princi-
pally trimethylamine in addition to environmentally induced flavors, can contribute to
the fresh seafood aroma (Kawai and Sakaguchi, 1996; Whitfield, 1990). Several exten-
sive review documents on fresh fish volatiles can be found elsewhere (Lindsay, 1994;
Shahidi and Cadwallader, 1997). Moreover, Olafsdottir et al. (1997a) provided a com-
prehensive review of the principal classifications of fish odor such as fresh fish odor,
environmentally derived odor, oxidized odor, microbial spoilage odor, and treating odor.
The fresh seafood odor is acceptable for the first few days after catching. Then, microbial
metabolites and oxidation products can alter the overall fish aroma (Olafsdottir et al.,
1997b; Hognadottir, 2000).
Aroma is one of the foremost key considerations for assessing seafood freshness and
quality. The development of distinctive odors in seafood through chilled storage pro-
duced by microbial growth has been documented in other studies (Shewan et al., 1953;
Castell and Greenough, 1957). The organoleptic descriptors for the development of aroma
alterations through fish storage were depicted as the following: fresh → flat → sweet →
522 Food Aroma Evolution

stale → putrid (Elliott, 1947). Aroma alterations have been related to the generation of
volatile compounds formed by leading spoilage organisms (Miller et al., 1973; Herbert
and Shewan, 1976). The postmortem, a kind of complex process in the fish, results in
loss of freshness and spoilage. As well as the interactions of chemical, biochemical, and
microbiological combination in fish, it can be changeable regarding different fish species,
diverse extrinsic parameters such as handling and various storage conditions may affect
the spoilage array and produce unpleasant odors (Huss, 1995).

25.2.1 Fresh Fish Aroma

Fresh fish aroma generally has sweet, mild, green, plant-, and fishy-like odor notes. Its
characteristic aroma may be quite different from processed and stored fish. The vola-
tile compounds related to fresh fish odor are mainly C6, C8, and C9 alcohols, ketones,
and aldehydes originated from the unsaturated fatty acids (UFA) through lipoxygenase
pathways. The C6 compounds like hexanal, (E)-2-hexenal, and (Z)-3-hexenal give green,
grassy, and herbal plant odor notes. These compounds are associated with freshwater
fish species and are generally not detected in saltwater fish species. The C8 compounds
such as 1-octen-3-one, 1-octen-3-ol, 1-(Z)-5-octadien-3-one, and 1-(Z)-5-octadien-3-ol
can be found in many seafood and fish species, providing high-density plant and metal-
lic odor notes. The C9 compounds, for example, 3,6-nonadienal, and 2,6-nonadienal
can give cucumber, green, and fresh odor notes to some kinds of seafood, especially
freshwater fish species. The fresh fish aroma compounds discussed above are identical to
those detected in several vegetables, as can be deduced from their distinctive odor notes.
These compounds can be formed throughout lipoxygenases in fish as well as in plants,
while the pathways may be rather different. The C8 ketones and alcohols detected in
many fish species can also emerge in some mushroom species. Even though they solely
possess mushroom or geranium odor notes, and they provide powerful plant odor notes
in freshly cached seafood. The C9 aroma compounds present in melons and cucumber
fruits can give melon and cucumber odor notes to the fish species in which they are avail-
able. 2-Hexenal and hexanal provide green and herbal plant odor notes. Although both
compounds are present in all freshwater fishes, hexanal has been detected in saltwater
fishes within 5 and 6 days after catching (Hall and Andersson, 1983; Josephson, 1985;
Lindsay, 1990).

25.2.2 Effect of Postmortem Alterations on Fish Aroma

The alterations of fish postmortem primarily consist of autolytic activity comprising

nucleotides deterioration, active taste inosine generation, hypoxanthine (Hx) aggrega-
tion, endogenous enzyme activity, and pH reduction, followed by the propagation spe-
cific spoilage organisms (SSO) and the progress of volatiles leading to spoilage alterations
as well as impacting the final product quality (Botta, 1995; Huss, 1995). Therefore, more
studies are required to clarify the chemical elucidation of spoilage processes in meat prod-
ucts as well as the connection of organoleptic properties with microbiological alterations
(Dainty, 1996). Postmortem alterations resulting in the progress of odors (PUFA: poly-
unsaturated fatty acids, ATP: adenosinetriphosphate, Hx: hypoxanthine, SSO: specific
spoilage organisms, NPN: non-protein nitrogen-containing compounds, and TMAO:
trimethylamineoxide) in chilled fish are summarized in-depth below (Josephson et al.,
 Fish 523

1983; Lindsay et al., 1987; Hsieh et al., 1988; Olafsdottir et al., 1998a; Jørgensen et al.,
2001):

1. Lipoxygenase pathway: lipids → PUFA → volatile compounds

2. Endogenous enzymes:
a. Glycolysis; pH↓ and lactic acid↑
b. Rigor mortis; ATP → inosine → Hx → urea
c. Autolysis; cathepsin and calpain → texture alterations
3. Oxidation: protein → peptides → amino acids → free fatty acids → volatile
compounds
4. SSO: soluble substances in the muscle → sarcoplasmic proteins → nucleotides →
NPN → TMAO

It is well-known that PUFA via the enzymatic pathway can be converted into volatile com-
pounds, which is the initial step of the generation of fresh fish odors resulting in fresh
and plant aroma notes (Josephson et al., 1983; Hsieh et al., 1988). During the storage
period, aroma profile alterations can occur by the extra conversion and degradation of fish
fillet compositions, including membrane-bound phospholipids and sarcoplasmic proteins
through intrinsic enzymatic activity, microbial growth, and autoxidation. The pool con-
stituents are comprised of mainly soluble compounds including non-protein nitrogenous
components (NPN), small peptides such as amino acids, anserine, and carnosine, and
asnucleotides such as guanidin. These compounds are generally formed due to microbial
growth and metabolism. TMAO and creatine can be tainted and result in off-flavors in
the fish flesh (Ikeda, 1980). Some compounds discussed earlier can affect fish taste such as
individual amino acids, including glutamic acid, alanine, valine, and valine, in addition to
the degradation compounds of the nucleotides such as inosine. In addition, it was reported
that anserine-like tastes contribute to a “mouth satisfaction” feeling (Hodge, 1967).

25.2.3 Effect of Fish Species on Fish Aroma

Fish species may be divided into three categories concerning their habitat, such as fresh
saltwater or marine fish, freshwater fish, and euryhaline fish (Rehbein and Oehlenschlager,
2009). Fish species each have a distinct odor, and its alteration is dependent on many dif-
ferent agents. Content and odor thresholds make the compounds influential and substan-
tial agents, as some compounds may illustrate synergistic impacts and affect the quality
of the global fish odor. Some of the volatile compounds are pleasant in low amounts.
However, increasing their content through the enzymatic pathway, particularly in long-
chain ketones and alcohols derivatives, may result in the formation of an off-odor to
the overall fresh fish aroma. Additionally, the accumulation of greater amounts of such
compounds due to the auto-oxidation pathway may lead to oxidized and fishy odor notes
(Maarse, 2017).
Fresh saltwater, freshwater, and euryhaline species odor alterations occur due to
diverse enzymatic reactions (Kawai and Sakaguchi, 1996). Lipoxygenase enzyme activity
has been detected on the gills and skin of both fresh saltwater and freshwater fish species
such as sardines, river trout, and rainbow trout, resulting in the generation of volatile
compounds, contributing to a green attribute with a desirable fish aroma (Zhang et al.,
1992; German et al., 1985, 1986). Unsaturated carbonyl compounds and alcohols con-
taining six, eight, or nine carbon atoms available in the headspace of fish in low amounts
524 Food Aroma Evolution

(ppb) may form character impact odors in fish, including plant, cucumber, melon, and
mushroom notes (Josephson et al., 1984; Lindsay et al., 1987; Milo and Grosch, 1993).
The enzymatic activity which causes the formation of distinct green and plant odor notes
is more prevalent in freshwater and euryhaline fish species than in saltwater fish species.
Fresh saltwater fish are roughly without odor because they contain a low concentra-
tion of volatile compounds. Fresh saltwater fish commonly emit an indistinct odor, imme-
diately after being caught in the ocean. This property relates to all live saltwater seafood
comprising Alaska pollack, sardine, bonito, Pacific mackerel, herring, redfish, saithe, and
cod (Kawai and Sakaguchi, 1996; Rehbein and Oehlenschlager, 2009). Nitrogen-holding
compounds including trimethylamine (TMA), dimethylamine (DMA), and ammonia;
sulfur-comprising compounds like dimethyl sulfide (DMS) and methanethiol; and some
acids such as propionic, formic and acetic acids can be found in the faint odor of a vari-
ety of fresh saltwater fish. Of the compounds discussed, even low amounts of TMA can
result in very distinct odor notes in fresh saltwater fish products. Additionally, combining
the TMA and further-mentioned compounds can give rise to an additional saltwater fish
odor than just TMA alone. The dilutions of the TMA and DMS contribute to the dried
or boiled fish-like and seashore product-like odor notes. Yet the blend of the TMA and
DMS compounds hold a distinct odor note comparatively related to a seafood flavor.
Some volatile compounds found in several saltwater fish are outlined in a previous review
(Kawai and Sakaguchi, 1996). Recently, the volatile compounds of wild gilthead sea
bream (Sparus aurata) were evaluated by sensory and instrumental analyses. Aldehydes
and alcohols were found as the most predominant volatile compounds in sea bream, as
they accounted for the main proportion. Hexanal, heptanal, nonanal, (Z)-4-heptenal,
octanal, (E)-2-nonenal, decanal, benzenacetaldehyde, (E,E)-2,4-decadienal, 1-octen-
3-ol and (E)-1,5-octadien-3-ol were potent volatiles with regard to odor activity values
(OAVs) (Selli and Cayhan, 2009).
In comparison with fresh saltwater fish, plenty of freshwater fish inhabiting lakes,
rivers, and ponds comprising salmon, catfish, carp, tench, pikeperch, perch, pike, even
cultured trout possess an earthy odor note. The volatile compounds derived from piperi-
dine and its combination of ethanal, likely 1,1-bispiperidino-ethane, can be accountable
for the overall odor of fresh salmon and carp. In fact, a complex of piperidine and etha-
nal is accountable for an earthy odor note, and that complex, in addition to TMA, is
accountable for the indistinct odor of fresh saltwater fish. Piperidine and pyrrolidine have
already been reported in a study on fresh carp flesh and 12 diverse products of canned
fresh saltwater fish and freshwater fish. Although pyrrolidine was detected in greater
amounts in all the studied fish, piperidine was found in trace levels in pond smelt and
two canned red salmon, yet not in carp flesh. Hence, piperidine is not a compound com-
mon to ordinary fresh saltwater and freshwater fishes. Additionally, piperidine concerns/
relates to the spawning or the succeeding mortality of grass carp and salmon. Piperidine
and pyrrolidine can be formed through a cyclization pathway from two diaminoalkanes,
namely cadaverine (1,5-diaminopentane) and putrescine (1,4-diaminobutane), respec-
tively. 1,5-diaminopentane was investigated in snake-head, loach, eel, carp, and rainbow
trout samples, and only detected in loach, with nearly 2 mg per 100 g, whereas 1,4-diami-
nobutane was detected in snake-head, carp, rainbow trout at approximately 1 mg per
100 g. The distinct earthy odor character of freshwater fish originates from geosmin and
2-methylisoborneol. Inhabitant microorganisms such as blue-green algae and actinomy-
cetes (mold-like bacteria) generate both geosmin and 2-methyl-isoborneol compounds.
Plenty of freshwater fish dwelling in lakes and ponds might contain these compounds
in very trace amounts. However, as these compounds have low odor threshold values,
 Fish 525

freshwater fish are therefore established for their characteristic earthy odor notes (Kawai
and Sakaguchi, 1996). Similarly, the odor-active and off-odor compounds of rainbow
trout (Oncorhynchus mykiss) were determined using GC-MS coupled with olfactometry.
Based on this study, 2-methylisoborneol (musty) and geosmin (earthy) (Figure 25.1) were
found as the most important off-odor contributors to the rainbow trout aroma (Selli et
al., 2009).
The four common compounds detected in saltwater species including 1-octen-3-ol,
hexanal, 2,5-octadien-1-ol, and 1,5-octadien-3-ol contribute to the moderate, faint odor
notes in saltwater species. Moreover, the unsaturated C9 carbonyl compounds comprising
2,6-nonadienal with strong plant, green, melon, and cucumber odor notes were found in
freshwater and euryhaline fish. Generally, fresh fish contains very low amounts of aroma
compounds in its headspace. In addition to low concentration, some of them possess very
low odor thresholds. However, they still impact the fish aroma and drastic changes in
their concentration influences the overall aroma of fish (Josephson et al., 1984).

25.2.4 Effect of Environmental Conditions and Seasonal Impacts on Fish Aroma

Ecological parameters and seasonal impacts, including spawning, may affect the qual-
ity of fish odor. In mature salmon, their volatile template alters when they migrate from
the sea for spawning. The C9 lipoxygenase-originated compounds were detected in
greater amounts in spawning freshwater and euryhaline fish (Josephson et al., 1984).
Similarly, seasonal impacts have been studied on capelin, a kind of saltwater species,
which give a very distinct cucumber odor note when spawning. In capelin, 2,6-nona-
dienal was found as the most dominate aroma-active compound with a cucumber odor
note (Olafsdottir et al., 1997b). Additionally, investigations on hydroperoxide accu-
mulation in fish flesh demonstrated that the contribution of such accumulations to
the formation of fresh fish odor is related to seasonal changes. Specific hydroperoxide
isomer accumulation happened together with the course of development of distinctive
odor in sweet smelt. Therefore, the C9 volatile compounds such as 3,6-nonadien-1-ol,
(E,Z)-2,6-nonadienal, and (E)-2-nonenal in sweet smelt flesh may be derived from the
hydroperoxide isomers (Kaewsrithong et al., 2001).

FIGURE 25.1 Chemical structures of geosmin and 2-methylisoborneol (MIB).

526 Food Aroma Evolution

25.2.5 Effect of pH Levels on Fish Aroma

The pH level of food strongly influences flavor alterations. The importance of pH impact
on fish odor has recently been reviewed by different researchers (Kawai and Sakaguchi,
1996; Schultz, 1967). In fish products, the hydrogen ions formed in postmortem mus-
cles arise from ATP hydrolysis rather than from lactate production; however, there is
an intense correlation between the decline in pH and concentration of lactate in the fish
fillet (Hultin, 1985). The lactate concentration generated is, after all, mostly dependent
on the ATP concentration formed through the glycolytic cycle. Moreover, the lactic acid
concentration generated is proportional to the stored carbohydrate (glycogen) concentra-
tion in the tissue of the living fish (Hall, 2012). Stress prior to capture accelerates the pH
reduction in the postmortem muscles (Chiba et al., 1991). Nevertheless, the ultimate pH
reached is concerned with the kind of muscle and species of fish, yet, contrary to mam-
malian muscles, it is not linked with the level of exercise or stress before death (Love
and Muslemuddin, 1972). The production of lactate owing to struggle during capture is
simply and gently eliminated from the fish muscle. The ultimate muscle pH of lean white
fish such as cod is generally between 6.2 and 6.6, while that of red meat species such as
mackerels and tuna is lower and ranges between 5.5 and 5.9 (Lougovois and Kyrana,
2005). The muscles of fish normally have rather lower amounts of glycogen in compari-
son with mammals, originally owing to alterations in the feeding template over the sea-
son. Consequently, the ultimate pH in fish is generally greater than in animals with warm
blood. The decrease of the pH postmortem has the main impact on different parameters
of muscle quality, such as microbial growth resistance, the capacity of water binding, and
texture (Lougovois and Kyrana, 2005).
Three main mechanisms for the generation of fresh fish aroma compounds such as
oxidative reactions with omega-3 PUFAs, biosynthetic reactions of eicosa-5,8,ll,14,17-
pentaenoic acid (EPA) with lipoxygenases and hydroperoxide lyases, and spontaneous
retro-aldol condensation are outlined. Each generation mechanism is affected by the pH
level. Moreover, pH influences the formation of some low-molecular weight compounds,
namely TMA from the fish muscle, DMS from saltwater fish products, and hydrogen
sulfide (H 2S) from heated meats and proteins. The TMA and acid combination can alter
the quality of the odor. Both TMA and the acids and TMA and DMS have been used to
manufacture fish flavor reactions (Kawai and Sakaguchi, 1996). The pH in foodstuffs
has a role in the flavor’s progress through the Maillard reaction. Increasing pH results in
the increasing of favored color and polymeric compounds as well as nitrogen-containing
compounds such as pyrazines (Mottram and Madruga, 1994).

25.2.6 Effect of Fat/Oil Amount on Fish Aroma

Both fresh fish and saltwater fish species may be categorized into four groups concern-
ing their total fat amount in edible fillet, ranging in certain species between 1 and
30%, based on their maturity period. The four fish groups classified by their total fat
content are lean fish species (less than 1%), low fat content fish species (between 1 and
5%), medium fatty fish species (between 5 and 10%), and fatty fish species (more than
10%) (Oehlenschlager and Rehbein, 2009). The protein content generally stays con-
stant in every fish, while fat content changes significantly in terms of quantity and fatty
acid profile. The fat content is not only principally associated with biological stage of
maturity, but also with nutritional conditions, harvesting season and ground, and age.
 Fish 527

Additionally, the fat distribution in the bodies of fish is not homogenous. It is deposited
in the liver as an energy resource store in lean fish species, and in the muscle fillet as a
subcutaneous layer in the crust under the skin or in the intestines of fatty fish species.
In a great deal of fatty fish species, there is a linear correlation between the water and
fat extent of muscle fillet. Lean fish species contain a greater content of polar lipids
such as phosphatidylethanolamine and phosphatidylcholine than fatty fish species, in
which the fat is composed mostly of neutral lipids or triacylglycerols. Fish lipids alter
from those of terrestrial animals largely in their elevated extent of long-chain, exceed-
ingly unsaturated fatty acids of the n–3 series, such as eicosapentaenoic acid (20:5)
and docosahexaenoic acid (22:6), frequently related to as polyunsaturated fatty acids.
The PUFAs content in fatty fish species such as dogfish, herring, mackerel, salmon, and
tuna are 3, 2.3, 4.6, 2.3, and 2.1%, respectively. The extremely unsaturated attributes
of PUFAs make them susceptible and lead them to oxidative breakdown and lipid oxi-
dation. Hence, fatty fish species tend to display both rancid odors and tastes after a
restricted storing period (Oehlenschlager and Rehbein, 2009).
Lipids of seafood species are of great importance to not only pleasant but also the
unpleasant flavor quality of these fishery products. Although seafood lipids can influ-
ence the freshly farmed fish flavors through undergoing lipoxygenase pathway oxidation,
a long storage period of fishery products containing lipids gives rise to flavor deterio-
ration and off-flavors. Accordingly, autoxidation inhibition may be a leading criterion,
while the fishery products are consumed as a food source or food ingredient (Shahidi and
Cadwallader, 1997). For preventing the oxidative deterioration and aroma reversion of
fishery products, the use of antioxidants, appropriate packaging methods, and applying
low temperatures during the storage period are employed. Additionally, in the case of
seafood oil products, the microencapsulation method, which can both prolong the shelf-
life of products and be simply added into foodstuffs, is used (Shahidi and Han, 1993).

25.2.7 Effect of Storage Conditions on Fish Aroma

Throughout the storage period, microbial and autolytic reactions can deteriorate fresh
fish odors and change the metallic, plant, and fresh flavors into a flat flavor. Fatty acids,
phenols, and sulfur compounds can emerge quickly through the growth of microbes,
which provide putrid and spoiled off-odor notes. The negative fishy odors such as dried
and stale fish and old, reminiscent odors appear with the breakdown of TMAO into
TMA through microbial reactions which are not present in freshly harvested fish. In
other respects, DMA is formed through the enzymatic pathways in the fish muscle, caus-
ing an ammoniacal aroma. Apart from microbially produced volatiles, another reaction
responsible for undesirable odors is the autoxidation of fish lipids. Aldehydes, critical
compounds due to their comparatively lower odor thresholds, are generally produced
as a result of lipid oxidation. These compounds are generally responsible for the green,
fatty, plant-like, and fruity odor notes in fresh and stored seafood (Alasalvar et al., 2005).
Beside aldehydes, ketone groups make a substantial contribution to the overall aroma of
stored fish and sea products. Ketones with lower odor threshold values make a higher
contribution to aroma generally with buttery, fatty, and fishy notes (Table 25.1). These
compounds are mainly originated from the oxidation of fatty acids, microbial oxidation,
and amino acid degradation pathways during storage (Chung and Cadwallader, 1994;
Pan and Kou, 1994). A summary of the volatile compounds detected in fish throughout
storage with an increment in concentration development is displayed in Table 25.1.
528 Food Aroma Evolution

TABLE 25.1 Changes in Seafood Volatile Compounds during Storage Periods

Fish Storage Detected Volatiles Odor Attributes Reference
Conditions
European For 15 days Thiophene Cooked vegetables, sulfur Leduc et al.
seabass at 4˚C in ice Hexanal Fruity, green apple (2012)
Octanal Fruity, lemon, sweet
1-Nonen-3-ol Mushroom
Raw For 9 days at 2,3-Butanedione Buttery, caramel Prost et al.
sardine 4˚C in ice Heptanal Green, floral (2004)
(E)-2-Hexenal Green
(Z)-4-Heptenal Cooked fish, potato
(Z)-3-Hexen-1-ol Woody, marine
1-Octen-3-ol Mushroom
Methional Boiled potato
2-Nonanol Plastic, fruity
Boiled For 20 weeks 2,3-Pentanedione Buttery Milo and
cod at –13˚C Methional Boiled potato Grosch (1996)
Dimethyl trisulfide Cabbage-like
1-Octen-3-one Mushroom
(E,E)-2,4-Nonadienal Fatty
2-Acetyl-1-pyrroline Roasted

25.2.7.1 Effect of Microbial Degradation on Fish Aroma

Microbial spoilage odors of fish are caused by the degradation of fish compounds, espe-
cially amino acids, through the microbial pathway. In the headspace of fish during spoil-
age, the general spoilage compounds including sulfides, methyl mercaptan, hydrogen
sulfide, ethanol, TMA, and ammonia give putrid, rotten, stale, and fishy odor notes
(Herbert et al., 1975). Volatile compounds produced by spoilage microorganisms were
generally summarized in Table 25.2.
TMAO is commonly not present in freshwater fish and TMA does not exist in freshly
caught saltwater fish (Figure 25.2). Fish with higher TMA amounts are not very desir-
able for consumers. TMAO can have an osmoregulatory role in marine fish, while it is
generally not detected in freshwater fish. DMA with an ammoniacal odor note can be
generated together with formaldehyde from the enzymatic reaction in fish fillet. These
enzymatic reactions may result in fish muscle toughening. The crosslinks of formalde-
hyde with proteins lead to texture changes. Both methyl mercaptan and dimethyl sul-
fide contribute to off-odors and are normally created through microbial activity in fish
(Josephson et al., 1983, 1984; Lindsay et al., 1987; Lindsay, 1990; Kawai and Sakaguchi,
1996). Ethyl butyrate, methyl (methyl thio) methyl sulfide, carbon disulfide, methyl
disulfide, o-2-methylpropyl hydroxylamine, o-3-methylbutyl hydroxylamine, 2-methyl-
2-propanamine, and trimethylamine have been reported as the main aroma compounds
detected in the headspace of some tropical prawns after less than 8 days storage in ice.
These aroma compounds are mostly generated by bacterial degradation of bioactive com-
pounds, particularly amino acids (Chinivasagam et al., 1998). Moreover, several other
aroma compounds formed through bacterial spoilage have also been recommended to
estimate the generation of spoilage odors taking place in fish products under various
 Fish 529

TABLE 25.2 Detected Volatile Compounds in Seafood Associated with Spoilage

Microorganisms
Volatile Compounds Related Spoilage Seafood Storage Conditions References
Microorganism
Methanethiol, Pseudomonas fragi Prawn For less than 8 Chinivasagam
1-propanethiol, extract days in ice or ice et al. (1998)
dimethyl sulfide, slurry
dimethyl trisulfide,
ethyl acetate,
methyl acetate
1,2-Butanediol, Pseudoalteromonas Brown Refrigerated at 4°C Broekaert
dimethyl disulfide, spp. shrimp et al. (2013)
methanethiol,
acetone,
3-methylbutanal, tma
2-Butanone, Psychrobacter spp. Brown Refrigerated at 4°C Broekaert
2‑methyl‑2-propanol, shrimp et al. (2013)
acetaldehyde, acetone,
isopropyl alcohol,
tma
3-Methyl-1-butanal, Brochothrix Cooked MAP storage for Jaffrès et al.
2-methyl-1-propanol, thermosphacta shrimp 27 days at 8°C (2011)
2-methyl-1-butanal,
2,3-butanedione,
methanethiol,
ethyl acetate
3-Methyl-2-butanone, Carnobacterium Cooked MAP storage for Jaffrès et al.
2-methyl-1-butanol, maltaromaticum shrimp 27 days at 8°C (2011)
acetaldehyde,
3-methyl-1-butanal,
methanethiol,
ethyl acetate
1-Propanol, 2-Butanol, Serratia liquefaciens Cooked MAP storage for Jaffrès et al.
2-propanone, shrimp 27 days at 8°C (2011)
2,3-butanedione,
methanethiol,
ethyl acetate
3-Methyl-2-butanol, Photobacterium Raw MAP storage for 8 Macé et al.
2-methyl-1-propanol, phosphoreum salmon days at 8°C (2013)
isobutyraldehyde,
benzaldehyde,
acetic acid,
3-hydroxybutanone,
ethyl acetate
(Continued )
530 Food Aroma Evolution

TABLE 25.2 (Continued) Detected Volatile Compounds in Seafood Associated with

Spoilage Microorganisms
Volatile Compounds Related Spoilage Seafood Storage Conditions References
Microorganism
TMA, Pseudomonas Black For 15 days at Miller et al.
methyl mercaptan, putrefaciens rockfish 1–2°C (1973)
dimethyl disulfide,
dimethyl trisulfide,
3-methyl-1-butanol
3-Methyl-1-butanol, Brochothrix Cooked MAP storage for Laursen et al.
1-octen-3-ol, thermosphacta peeled 10 days at 5°C (2006)
1-penten-3-ol, shrimp
3-methyl-1-butanal,
2,3-butanedione,
dimethylbenzene
2-Ethyl-1-hexanol, Carnobacterium Cooked MAP storage for Laursen et al.
3-methyl-1-butanol, spp. peeled 10 days at 5°C (2006)
1-pentanol, shrimp
1-octen‑3-ol, 1-penten-
3-ol, 3-methyl-1-
butanal,
2,3-butanedione,
4-methyl-2-pentanone,
2-heptanone,
2,4,6-trimethylpyridine
Ethanol, Pseudomonas spp. Sea Air and MAP Parlapani et
2-ethyl‑1‑hexanol, bream storage at 0 and al. (2017)
3-methyl-1-butanol, fillets 15°C
hexanal,
octanal,
6-methyl-5-hepten-
2‑one, 5-ethyl-2-
methylbutyrate,
ethyl octanoate
1-Pentanol, Shewanella spp. Sea Air and MAP Parlapani et
1-heptanol, bream storage at 0 and al. (2017)
hexanal, fillets 15°C
octanal,
nonanal,
2-pentanone,
(E)-2-octenal

storage conditions (Lindsay et al., 1987; Olafsdottir et al., 2000, 2002). Therefore, the
number of volatile compounds generated by microbial activity can be increased by spoil-
ing fish during storage (Lindsay et al., 1987).
TMA and a mixture of several compounds including sulfur compounds, amines and
alcohols have already been recommended as indices of freshness and spoilage by dem-
onstrating various alterations taking place in storage (Lindsay et al., 1987; Jørgensen
et al., 2001; Alasalvar et al., 2005). Additionally, the contribution of specific spoilage
 Fish 531

FIGURE 25.2 The microbial transformation of TMA to TMAO and DMA.

organisms such as Photobacterium phosphoreum, Pseudomonas ssp., and Shewanella

putrefaciens progress the microbial metabolites, resulting in the spoilage of fish, and are
of great importance. For this reason, the measurement of SSOs is used to estimate the
spoilage degree and freshness more than total viable count (TVC) data in fishery products
(Dalgaard, 2000; Gram and Dalgaard, 2002).
Accumulation of short-chain esters, carbonyls, and alcohols in fish fillets in elevated
amounts over a chilled storage period can occur due to the microbial activities. Short-
chain alcohols such as 3-methyl-1-butanol, butanol, and ethanol have already been sug-
gested as indicators of quality for refrigerated fish spoilage. Further, propanol is reported
as an indicator in fish fillet packed by means of modified atmosphere packaging tech-
niques (Lindsay et al., 1987). The primary formation of ethanol in the spoilage of fishery
products has been involved with the use of carbohydrate resources, while the production
of branched-chain aldehydes and alcohols, for example, 3-methyl-butanal, 3-methyl-
1-butanol, and 2-methyl-1-propanol, most likely stem from the breakdown of leucine
and valine, respectively. Furthermore, 2-methyl-1-propanol and 3-methyl-1-butanol, that
supply fruity and alcoholic odor notes, have been detected as increasing throughout the
storage of cod (Olafsdottir et al., 1998b).
Ketones may affect the global odors owing to their usual odors and their low odor
thresholds. Acetoin (3-hydroxy-2-butanone) generation is representative of the spoilage
of chilled cod muscles packed in styrofoam packages and associated with P. phospho-
reum growth. First, acetoin amounts increased more than TMA, and so can be used as a
primary indicator of spoilage to identify loss of freshness. The acetoin level is far greater
than the further lipid-originated ketones found such as 3-pentanone, and 2-butanone, as
well as the carotenoid 6-methyl-5-heptene-2-one, originated in cod during storage. No
clear enhancement took place during continued storage or until the end of the shelf life
(Olafsdottir et al., 1998b).
TMA, acetic acid, dimethyl trisulfide, dimethyl disulfide, methanethiol, piperidine,
1-penten-3-ol, and 3-methyl-1-butanol have already been identified as indicators of spoil-
age in wild and cultured sea bream stored in ice for 23 days (Alasalvar et al., 2005).
Former investigations on the generation of aroma breakdown compounds exhibited
that pseudomonads, particularly P. fragi, were accountable for quality alterations and
the progress of fruity and sweet off-odors in chilled rockfish, haddock, and cod from the
North Atlantic region (Miller et al., 1973).
Esters, ketones, sulfides, and amines are the major volatile compound groups attrib-
uted to the presence and growth of S. putrefaciens and P. fragi in prawns (Chinivasagam
et al., 1998). The significance of Pseudomonas species in spoilage over the storage period
532 Food Aroma Evolution

has been shown in some fish species, such as American plaice (Lauzon, 2000) and some
Mediterranean Sea fish (Koutsoumanis and Nychas, 2000).
That amines increase during the spoilage of fish is well established, and the evalua-
tion of the very volatile amine compounds like total volatile bases (TVB-N) and TMA
have been utilized in the fish industry as quality indicators for seafood. Increments of
fishy and ammonia-like off-odors have been linked with fish spoilage bacteria such as
P. phosphoreum and Shewanella putrefaciens, which may degrade TMAO to TMA
(Jørgensen and Huss, 1989).
The volatile sulfur compounds, for example, dimethyl disulfide, methyl sulfide,
methyl mercaptan, and hydrogen sulfide have been introduced as the principal source of
putrid spoilage odors in seafood (Herbert et al., 1971). Dimethyl trisulfide was already
related to fish spoilage as well as being accountable for a characteristic undesirable onion
odor note in prawns (Whitfield and Tindale, 1984). Additionally, dimethyl trisulfide in
the headspace of boiled cod was found as the most powerful odourant leading to off-odors
in cod generated once the fresh fish was unsuitably stored. Through increased spoilage of
seafood, the amount of the microbially produced sulfur compounds can advance. Besides
providing the entire putrid spoilage odor notes of other aroma compounds like acids and
amines, the volatile sulfur compounds, possessing very low odor thresholds and distinct
odor notes, have a tendency to dominate the putrid spoilage odor (Milo and Grosch,
1996).
Certain acids, for example, propionic, formic, and acetic acids synthesized from the
breakdown of amino acids and lipids are, too, included in the flavor of fish. The above-
mentioned acids containing a short aliphatic chain may be related with an odor of fresh-
ness in numerous species such as sea bream (Pristimoides sieboldi), Spanish mackerel
(Scombrus japonicus), and bonito (Sarda sarda) (Kawai and Sakaguchi, 1996). Moreover,
volatile acids formation like acetic and formic acids in tuna are related to spoilage. Through
chilled storage, acetic acid was found in advancing amounts in cod fillets (Quaranta et al.,
1984). The presence of acids in seafood shift the TMA odor quality, and, with enhancing
acid content, the TMA volatility reduces, and the quality of the odor alters (Kawai and
Sakaguchi, 1996). Diverse benzene-derived compounds have been disclosed as a fraction
of the total volatiles in fermented anchovy, thermally processed crab, and chilled fish
(Alasalvar et al., 2005). Once investigating the volatile compounds in cod muscles, chlo-
roform, and styrene were most prevailing of the aromatic compounds found and were
detected to enhance over a stored period. Styrene was, too, determined in cultured and
wild sea bream over a chilled storage period, using polystyrene packages (Alasalvar et al.,
2005). The styrene odor is perceived as a kerosene note and has been related to off-odors
in surimi originated products linked with the growth of yeast (Koide et al., 1992).
Despite acids, esters, sulfur compounds, ketones, aldehydes, and alcohols being of
chief concern as indicators of fish spoilage; further groups of volatile compounds can
also play a role in evaluating the global headspace odor of seafood. The amount of the
straight-chain alkanes, for instance, undecane, decane, and nonane, seem to be identi-
cal in chilled cod muscles throughout storage time (Olafsdottir et al., 1998a). Although
several branched-chain alkanes were found, they are not considered of concern as indica-
tors of fish quality as they are not odor-active compounds. Numerous terpene-derived
compounds have been determined in seafood, such as limonene found throughout storage
in sea bream fish (Alasalvar et al., 2005). The limonene compound might stem from fish
that were fed from plant or algae sources. Based on a lower odor threshold, giving a fresh
lemon aroma note, limonene can play an important role in the global aroma note in fish
muscles (Olafsdottir et al., 1998a).
 Fish 533

25.2.7.2 Effect of Lipid Oxidation on Fish Aroma

Lipid-derived volatile compounds are important aroma compounds characteristic of fish
species. Lipid oxidation initiation in fish is largely related to polyunsaturated fatty acids
in phospholipids of muscle cell membranes, known to be more sensitive to oxidation than
triacylglycerols in fat accumulations (Decker and Xu, 1998). Lipid autoxidation in fish
during storage results in unpleasant odor notes. Indeed, oxidative procedures taking place
during fish storage may cause saturated and unsaturated aldehyde accumulation including
2,4,7-decadienal, 2,4-heptadienal, (Z)-4-heptenal, and hexanal that provide rancid odor
notes (Aro et al., 2003). By breakdown of the (E,Z)-2,6-nonadienal through autoxidizing
lipid systems, (Z)-4-heptenal is generated with linseed oil, painty, and putty odor attributes
at elevated amounts and a cardboard odor note at decreased amounts, as well as a boiled
potato odor note in general. Moreover, 2,4,7-decatrienals give fishy cod liver oil and burned
odor attributes in fish (Josephson et al., 1984; Lindsay, 1990; Kawai and Sakaguchi, 1996).
Investigations on the progress of volatile compounds in chilled cod muscles packed in
styrofoam boxes during the stored period at 0°C revealed that lipid-originated saturated
aldehydes that are oxidation products like decanal, heptanal, and hexanal were identified
in the muscles with increasing concentration throughout the storage period. Oxidation-
derived compounds bring about the distinctive fishy and sweet odor notes of chilled cod
muscles in addition to further carbonyls such 6-methyl-5-hepten-2-one, 3-pentanone,
2-butanone, 3-methyl butanal, and 3-hydroxy-2-butanone. Aldehydes mostly possess low
odor thresholds, and, consequently, their impression was bigger than ketones and alco-
hols, even though their total amounts were lower in fish species. The 6-methyl-5-hepten-
2-one compound which stems from carotenoids was depicted as flowery and spicy via
GC-O, and gave rise to further aldehydes and ketones in the specific sweet odor note in
the cod muscle (Olafsdottir et al., 2005). The effect of further odor-active compounds
exists in lower concentrations like the unsaturated aldehydes such as 2,4,7-decatrienal
and 2,4-heptadienal, and ought not to be disregarded. The compounds discussed above
have been related to dried fish, and rancid odor notes, but the sampling methods employed
were not adequate to permit quantification of these compounds (Nollet and Toldra, 2009).
Diverse antioxidants may affect the constancy of the fish fillet, and they have already
been investigated concerning the phospholipids oxidative constancy. The principal mem-
brane, bound in the lipids of seafood, are phospholipids, high unsaturated, and delicate to
oxidation, which can develop more during storage and further pre-processing procedures
(Hultin, 1994). In order to recognize the oxidation mechanisms in the fish muscle, the effect
of antioxidants such as glutathione peroxidase, ascorbic acid, and α-tocopherol, as well as
aqueous pro-oxidants comprising blood constituents such as inorganic metals like copper
(Cu) and iron (Fe), have been extensively investigated (Hultin and Kelleher, 2000). Therefore,
the amount of pro- and antioxidants in the muscle besides the primary, secondary, and ter-
tiary degradation compounds of lipid oxidation can be applied as possible quality indicators
in fishery products (Undeland, 1997). Proteolysis in fish has a vital part in postmortem alter-
ations bringing about unpleasant texture variations. The activity of endogenous enzymes
such as neutral calcium-dependent proteases (calpains) and lysosomal proteases (cathepsins)
gives rise to the breakdown of fish fillets (Delbarre-Ladrat et al., 2004).

25.2.8 Effect of Processing on Fish Aroma

The progress of odor in processed fishery products is a consequence of composite lipolytic

and proteolytic reactions caused by diverse processing factors such as temperature and
534 Food Aroma Evolution

enzymes. Lipid oxidation, thermal degradation, and the Maillard reaction, comprising the
Strecker degradation, is vital in the creation of complex processing flavors. Thermally pro-
duced key odorants by means of the Maillard reaction such as pyrazines are distinctive in
enzymatically hydrolyzed fishery products such as the byproducts of crayfish processing
(Baek and Cadwallader, 1996). Volatile compounds such as furans have been detected in
spray-dried shrimp powder, and shrimp hydrolysate, as well as sulfur-containing and alkyl-
pyrazines compounds, has been detected in cooked crustaceans (Pan and Kuo, 1994). Key
odorants characterized in enzymatically formed fishery product odorants are produced by
way of the Maillard reaction and Strecker breakdown of amino acids. Such compounds are
methional, the Strecker aldehyde created from methionine with a distinctive potato odor
note, and 2-acetyl-1-pyrroline with a popcorn odor note may be thermally formed. Besides,
lipid-originated compounds, such as 2,4-decadienal, 2,4-heptanal, hexanal, 2,6-nonadi-
enal, 1-octen-3-ol, and (Z)-4-heptenal, can lead to the overall aroma of fish and fishery
products (Jonsdottir et al., 2007). Aldehydes derived from lipids have a principal part in
odor generation and have been disclosed as bringing about the specific sweet and fish odor
notes of processed fishery products such as those in smoked salmon (Varlet et al., 2006,
2007; Jonsdottir et al., 2008). Decrement in the content of the fresh fish flavor in pickled fish
is associated with lower concentrations of fish carbonyls and alcohols. The residual fresh
fish carbonyls and alcohols bring about the slight but individual pickled fish flavor (Nollet
and Toldra, 2009). Processed fish odor has already been investigated in diverse fishery prod-
ucts like those that have been hydrolyzed, smoked, cooked, pickled, and ripened (Milo and
Grosch, 1996; Joffraud et al., 2001; Jonsdottir et al., 2007; Cayhan and Selli, 2011).

25.2.8.1 Effect of the Ripening Process on Fish Aroma

Several lipolytic and proteolytic reactions affect the progress of wanted flavor and texture
in ripened products (Toldra, 1998; Toldra et al., 2000). Aroma construction is a result of
a complicated mixture of chemical or enzymatic reactions comprising the Strecker deg-
radation, Maillard reactions, and lipid oxidation. Several volatiles have been identified in
ripened products, the utmost of them are produced from the enzymatic or chemical oxi-
dation of unsaturated fatty acids and supplementary interactions with free amino acids,
peptides, and proteins. Throughout processing, a thorough lipolysis has been confirmed.
Free fatty acids are formed on account of phospholipid hydrolysis, demonstrating part
of phospholipases, whereas the triglycerides remain virtually intact. The accumulation
of free fatty acids may occur in the course of the process as a consequence of oxida-
tive processes, and subsequently volatile compounds like ketones, alcohols, aldehydes,
and hydrocarbons (Toldra, 1998). Protease activity throughout processing initiates the
development of free amino acids and small peptides that can enhance the taste (Toldra
and Flores, 1998). Free amino acids too may perform as substrates for Maillard reactions
and Strecker breakdowns with sugars (Toldra, 1998). Identical procedures have been
testified to in ripened fishery products where methional originated from methionine, and
2,6-nonadienal from the oxidation of fatty acids were the foremost odor compounds
in sugar salted ripened roe products (Jonsdottir et al., 2004). In another study, simi-
larly, 3,5-octadien-2-one and 2,4-heptadienal compounds were related to the advance-
ment of characteristic aroma attained after anchovy ripening and consequently it was
recommended that lipid autoxidation throughout ripening was predominantly account-
able for aroma expansion. Yet with ripened products, manufacturers have perceived that
a certain level of proteolysis is indispensable for the aroma profile to improve. Also,
(Z)-1,5-octadien-3-one and methional were recognized as effective odor compounds
in ripened anchovy (Triqui and Guth, 1997), in addition to aldehyde compounds like
 Fish 535

3-methylbutanal, 2-methylpropanal, and acetaldehyde were fundamental, and the vola-

tile compounds of ripened anchovy were generally found to be derived from amino acids.
The malty odor note in salt-ripened herring as a wanted attribute has been distinguished
(Gudmundsdottir and Stefansson, 1997). Although the derivation of such flavors in her-
ring has not been identified, in numerous foodstuffs the existence of branched-chain
aldehydes, which have a low odor threshold, has been related to the degradation of amino
acids (Moore et al., 1976). For instance, isobutyraldehyde giving a typical malty odor
is formed from valine through microbial activity. Microbial activity has not been given
much attention in the ripening process of herring. However, the ripening flavor may be
caused by the degradation of free amino acids.
The traditional salted cod products from North Atlantic fisheries are extremely
esteemed as ripened seafood products in numerous territories; in addition to those origi-
nating from the Mediterranean region. Throughout salted cod ripening, the anticipated
texture and flavour improve because of the breakdown of fat and protein. In a survey,
the maturing of salted cod (Gadus morhua) manufactured by diverse salting procedures
exhibited potato, cucumber, and rancid odor notes, and boiled potato odor attributes
were observed in the highest odor concentrations. The boiled potato odor note is asso-
ciated with heptanal besides potato, and rancid odor notes are connected with (Z)-4-
heptenal (Nollet and Toldra, 2009; Oliveira et al., 2012). Also, methional originated
from methionine along with heptanal and (Z)-4-heptenal, which can be accountable for
the boiled potato odor note. In salted cod, further potent aroma compounds are formed
from the oxidation of lipids, for instance, 2-butanone, hexanal, and 1-penten-3-ol. A con-
vincing level of lipid oxidation is not only compulsory but also anticipated for the product
to satisfactorily ripen. Still, the process should be measured to attain the desired stage of
ripening as to consumer inclinations (Lauritzsen and Olsen, 2004; Oliveira et al., 2012).

25.2.8.2 Effect of Smoking Process on Fish Aroma

Although the characteristic smoke flavor comes from a number of compounds deriving
from smoke, it is largely linked with the volatile phenolic compounds (Sikorski et al.,
1998). Volatile phenolic compounds are vital for conservation and aroma attributes in
smoked products. Most of them are generated by lignin pyrolysis. The level of phenolic
compounds of such products is associated with their woody nature (Nollet and Toldra,
2009). 4-Methylguaiacol and guaiacol have been recognized as the key phenols in herring
(Clupea harengus) smoked muscles irrespective of the smoke procedure applied (Serot et
al., 2004). In order to evaluate smoked fish product quality by applying the electronic
nose technique, numerous quality samples were opted for to investigate results from dis-
tinctive volatile compounds of both the smoking process and microbial pathway using gas
chromatography. The leading aroma profile and groups in cold-smoked salmon displayed
an influence of the particular groups to the entire headspace. Guaiacol was regarded as
the key compound bringing about the typical smokehouse odor note. In addition, fur-
fural was found in elevated amounts in certain products. Therefore, furfural is a fragile
odorant and does not cause a distinctive smoked odor.
The principal compounds bringing about the smoked salmon odor come from the
breakdown of compounds through lipid oxidation and Maillard reactions (Toldra,
1998). Although the characteristic smoked salmon odor stems from a great deal of
chemical compounds detected in the smoke, it is largely linked with the phenolic com-
pounds. Syringol (2,6-dimethoxyphenol) and guaiacol (2-methoxyphenol), derived
from phenolic compounds, have been regarded as the main typical smoke-attributed
compounds in smoked salmon (Salmo salar) (Varlet et al., 2007; Jonsdottir et al., 2008)
536 Food Aroma Evolution

and in smoked herring (Clupea harengus) (Serot et al., 2004). The headspace com-
pounds of smoked swordfish and cod, exhibiting phenol pyrolysis classes, are the main
volatile compounds (Guillen et al., 2006). Together with phenols, furan compounds are
responsible for the smoked odor note in smoked salmon, while carbonyl compounds
like (E,Z)-2,6-nonadienal and heptanal were distinctive in unsmoked fish with its char-
acteristic fishy odor note (Varlet et al., 2006, 2007). Heptanal and (Z)-4-heptenal com-
pounds derived from an oxidative pathway with potato and rancid odor notes together
with 1-octen-3-ol with a mushroom odor note lead to the strongest odors in smoked
salmon that give fishy, earthy, fatty, and rancid odor notes (Jonsdottir et al., 2008).
Decanal, nonanal, hexanal, and 1-penten-3-ol are the main oxidatively originated
volatiles. It is obvious that they give rise to the specific fish odor in smoked salmon.
Alcohols, aldehydes, and ketones, microbial formed chemical compounds, such as 1-pro-
panol, 1-penten-3-ol, 3-methyl-1-butanol, 2-methyl-1-butanol, and 3-methyl butanal,
are plentiful in the cold-smoked salmon headspace over a storage period attributed to
spoilage off-flavors (Joffraud et al., 2001). Some of the compounds discussed above
were suggested as the main spoilage factors for smoked salmon regarding their elevated
concentrations with fruity spoilage off-odors, such as 3-methyl-1-butanol, 3-hydroxy-
2-butanone, 3-methyl-butanal, 2-pentanone, 2-butanone, and ethanol (Jonsdottir et
al., 2008). Furthermore, it was confirmed that the chosen principal volatiles can be
used over traditional microbial and chemical variables to elualterations in organolep-
tic properties such as off-odor, off-flavor, and sour/sweet, rancid, and smoked notes
(Nollet and Toldra, 2009).

25.2.8.3 Effect of the Cooking Process on Fish Aroma

During the cooking process, the flavor of fishery products alters intensely. The thermally
induced alterations lead to cooked meaty odor notes, which are often specific to fish
species. Strecker and Maillard degradation reactions have a principal role in evolving
the meaty odorants of cooked fishery products. Further reactions like the retro-aldol
condensation of dienals and lipid oxidation result in several significant odor compounds.
A mixture of the reactions discussed above is crucial for the evolution of the character
impact odor compounds in cooked fish, shellfish, and crustaceans, in addition to fur-
ther seafood (Shahidi and Cadwallader, 1997). Diverse reactions taking place during the
heat processing of muscle-containing foods have been associated with the development
of meaty flavors. In addition, the contribution of lipids, amino acids, and ribose to the
development of heat-produced flavors has also been shown (Lindsay, 1990). 2-Methyl-
3-furanthiol exhibiting a powerful meaty odor attribute has been observed in tuna and
is accountable for the typical meaty flavor in canned tuna (Withycombe and Mussinan,
1988). Unsaturated carbonyl compounds were the most prevailing compounds in canned
and boiled fish, particularly in trout, sulfur compounds were also present and caused off-
odors. Methional with a distinctive boiled potato odor note in the headspace of boiled
trout was observed as the dominant odor of the aldehyde fraction (Milo and Grosch,
1993). With regard to odor assessment in boiled cod, 3-methyl-butanal together with
(E,E)-2,4-decadienal, (E,Z)-2,6-nonadieanal, (Z)-1,5-octadien-3-one, methional, and
acetaldehyde were elucidated as character impact odorants (Milo and Grosch, 1996). In
a study, key odorants of cooked gray mullet (Mugil cephalus) were analyzed by GC-MS-
olfactometry by Cayhan and Selli (2011). Aldehydes were qualitatively and quanti-
tatively found as the most predominant volatile compounds in gray mullet. Based on
AEDA results, the most potent aroma-active compounds detected in the cooked sam-
ples were (Z)-4-heptenal and nonanal, which were depicted as the main cooked fish and
 Fish 537

green-fruity odor notes, respectively (Cayhan and Selli, 2011). In another study, raw red
mullet (Mullus barbatus) and its cooked samples, acquired from steam and oven cook-
ing, were subjected to aroma and key odorant analysis applying GC-MS-olfactometry. By
applying aroma extract dilution analysis, 8 and 13 aroma-active compounds were found
in raw and cooked fish samples, respectively. The most prominent differences between
raw and cooked fish samples were as follows: 3-hydroxybutan-2-one, 2,3-octadienone,
(E,E)-2,4-heptadienal, linalool, γ-butyrolactone, 1-methylpyrrolidin-2-one, 2H-furan-
5-one, and pyrrolidin-2-one were only found in cooked samples whereas hexanal and
2-phenoxyethanol in only raw fish samples (Salum et al., 2017).

25.3 CONCLUSION

Fish or fishery products are preferred all around the world due not only to their nutri-
tional advantages to health but also their distinguishable and delicate flavors. The aroma
profile is one of the important parameters for determining fish quality properties. Fish
aroma quality is affected by many factors, including enzymatic and chemical reactions,
microbial activities, improper handling, and storage conditions. In order to prevent the
decline in aroma and quality of fresh fish and fishery products, some traditional tech-
niques have been applied including super or deep chilling, cooling, freezing, salting and
drying, chemical treatments, cooking, smoking, canning, and packaging. In addition,
some novel applications used for this purpose have included gamma irradiation, ultra-
sound, high-pressure treatment, and a pulsed electric field.

REFERENCES

Alasalvar, C., Taylor, K.D.A., and Shahidi, F. (2005). Comparison of volatiles of cultured
and wild sea bream (Sparus aurata) during storage in ice by dynamic headspace
analysis/gas chromatography–mass spectrometry. J Agric Food Chem 53, 2616–22.
Aro, T., Tahvonen, R., Koskinen, L., and Kallio, H. (2003). Volatile compounds of Baltic
herring analysed by dynamic headspace sampling–gas chromatography–mass spec-
trometry. Eur Food Res Technol 216, 483–8.
Baek, H.H. and Cadwallader, K.R. (1996). Volatile compounds in flavor concentrates
produced from crayfish-processing byproducts with and without protease treat-
ment. J Agric Food Chem 44, 3262–7.
Bisogni, C.A., Ryan, G.J., and Regenstein, J.M. (1987). What is fish quality? Can we
incorporate consumer perceptions? In Seafood Quality Determination, Proceedings
of the International Symposium on Seafood Quality Determination, edited by D.E.
Kramer and J. Liston, Coordinated by the University of Alaska Sea Grant Collage
Program, Anchorage, AK.
Botta, J.R. (1995). Evaluation of Seafood Freshness Quality. New York, NY: VCH
Publishers Inc.
Bremner, H.A. (2000). Toward practical definitions of quality for food science. Crit Rev
Food Sci Nutr 40, 83–90.
Broekaert, K., Noseda, B., Heyndrickx, M., et al. (2013). Volatile compounds associated
with Psychrobacter spp. and Pseudoalteromonas spp., the dominant microbiota of
brown shrimp (Crangon crangon) during aerobic storage. Int J Food Microbiol 166,
487–93.
538 Food Aroma Evolution

Caggiano, M. (2000). Quality in harvesting and post-harvesting procedures influence

on quality. Fish freshness and quality assessment for sea bass and sea bream. Cah
Options Mediterr 51, 55–61.
Castell, C.H. and Greenough, M.F. (1957). The action of Pseudomonas on fish muscle: 1.
Organisms responsible for odours produced during incipient spoilage of chilled fish
muscle. J Fish Board Canada 14, 617–25.
Cayhan, G. and Selli, S. (2011)). Characterization of the key aroma compounds in cooked
grey mullet (Mugil cephalus) by application of aroma extract dilution analysis.
J Agric Food Chem 59, 654–9.
Chiba, A., Hamaguchi, M., Kosaka, M., et al. (1991). Quality evaluation of fish meat by
31 phosphorus‐nuclear magnetic resonance. J Food Sci 56, 660–4.
Chinivasagam, H.N., Bremner, H.A., Wood, A.F., and Nottingham, S.M. (1998).
Volatile components associated with bacterial spoilage of tropical prawns. Int J
Food Microbiol 42, 45–55.
Chung, H.Y. and Cadwallader, K.R. (1994). Aroma extract dilution analysis of blue crab
claw meat volatiles. J Agric Food Chem 42, 2867–70.
Dainty, R.H. (1996). Chemical/biochemical detection of spoilage. Int J Food Microbiol
33, 19–33.
Dalgaard, P. (2000). Fresh and lightly preserved seafood. In Shelf Life Evaluation of
Foods. Aspen Publishers, pp. 110–39.
Decker, E.A. and Xu, Z. (1998). Minimizing rancidity in muscle foods. Trends Food Sci
Technol 9, 241.
Delbarre-Ladrat, C., Verrez-Bagnis, V., Noel, J., and Fleurence, J. (2004). Relative con-
tribution of calpain and cathepsins to protein degradation in muscle of sea bass
(Dicentrarchus labrax L.). Food Chem 88, 389–95.
Elliott, R.P. (1947). Evaluation of surface pH as a freshness index for fish fillets. J Food
Sci 12, 87–98.
German, J.B., Bruckner, G.G., and Kinsella, J.E. (1986). Lipoxygenase in trout gill tis-
sue acting on arachidonic, eicosapentaenoic and docosahexaenoic acids. Biochim
Biophys Acta (BBA)-Lipids Lipid Metab 875, 12–20.
German, J.B., Chen, S.E., and Kinsella, J.E. (1985). Lipid oxidation in fish tissue. Enzymic
initiation via lipoxygenase. J Agric Food Chem 33, 680–3.
Gram, L. and Dalgaard, P. (2002). Fish spoilage bacteria–problems and solutions. Curr
Opin Biotechnol 13, 262–6.
Grigorakis, K. (2007). Compositional and organoleptic quality of farmed and wild gil-
thead sea bream (Sparus aurata) and sea bass (Dicentrarchus labrax) and factors
affecting it: A review. Aquaculture 272, 55–75.
Gudmundsdottir, G. and Stefansson, G. (1997). Sensory and chemical changes in spice‐
salted herring as affected by handling. J Food Sci 62, 894–97.
Guillen, M.D., Errecalde, M.C., Salmeron, J., and Casas, C. (2006). Headspace vola-
tile components of smoked swordfish (Xiphias gladius) and cod (Gadus morhua)
detected by means of solid phase microextraction and gas chromatography–mass
spectrometry. Food Chem 94, 151–6.
Hall, G.M. (2012). Fish Processing Technology. Springer Science & Business Media.
Hall, G. and Andersson, J. (1983). Volatile fat oxidation products.: I. Determination
of odour thresholds and odour intensity functions by dynamic olfactometry. Leb
Technol 16, 354–61.
Herbert, R.A., Ellis, J.R., and Shewan, J.M. (1975). Isolation and identification of the
volatile sulphides produced during chill‐storage of North Sea cod (Gadus morhua).
J Sci Food Agric 26, 1187–94.
 Fish 539

Herbert, R.A., Hendrie, M.S., Gibson, D.M., and Shewan, J.M. (1971). Bacteria active
in the spoilage of certain sea foods. J Appl Bacteriol 34, 41–50.
Herbert, R.A. and Shewan, J.M. (1976). Roles played by bacterial and autolytic enzymes
in the production of volatile sulphides in spoiling North Sea cod (Gadus morhua). J
Sci Food Agric 27, 89–94.
Ho, C.T. and Hartman, T.G. (1994). Lipids in Food Flavors. ACS Publications.
Hodge, J.E. (1967). In Chemistry and Physiology of Flavors, edited by H.W. Shultz, E.A.
Day, and L.M. Libbey. AVI Publishing Co, Westport, CT, pp. 465.
Hognadottir, A. (2000). Flavor Perception and Volatile Compounds in Fish. Fisheries
Research Institute.
Hsieh, R.J., German, J.B., and Kinsella, J.E. (1988). Lipoxygenase in fish tissue: Some
properties of the 12-lipoxygenase from trout gill. J Agric Food Chem 36, 680–5.
Hultin, H.O. (1985). In Characteristics of Muscle Tissue, in Food Chemistry, 2nd ed.,
edited by O.R. Fennema. Marcel Dekker Inc., New York, NY.
Hultin, H.O. (1994). Oxidation of lipids in seafoods. In Seafoods: Chemistry, Processing
Technology and Quality. Springer, pp. 49–74.
Hultin, H.O. and Kelleher, S.D. (2000). Surimi processing from dark muscle fish. In Food
Science & Technology. Marcel Dekker, Inc., pp. 59–78.
Huss, H.H. (1995). Quality and Quality Changes in Fresh Fish. Food & Agriculture
Org.
Ikeda, S. (1980). Other organic components and inorganic components. In Advances in
Fish Science and Technology, edited by J. Connell. Fishing News Books, England,
pp. 111–24.
Joffraud, J.J., Leroi, F., Roy, C., and Berdague, J.L. (2001). Characterisation of volatile
compounds produced by bacteria isolated from the spoilage flora of cold-smoked
salmon. Int J Food Microbiol 66, 175–84.
Jonsdottir, R., Olafsdottir, G., Chanie, E., and Haugen, J.E. (2008). Volatile compounds
suitable for rapid detection as quality indicators of cold smoked salmon (Salmo
salar). Food Chem 109, 184–95.
Jonsdottir, R., Olafsdottir, G., Hauksson, S., and Einarsson, J.M. (2007). Flavorants
from seafood byproducts. In Handbook of Food Products Manufacturing. Wiley,
pp. 931–945.
Jonsdottir, R., Olafsdottir, G., Martinsdóttir, E., and Stefansson, G. (2004). Flavor char-
acterization of ripened cod roe by gas chromatography, sensory analysis, and elec-
tronic nose. J Agric Food Chem 52, 6250–6.
Jørgensen, B.R. and Huss, H.H. (1989). Growth and activity of Shewanella putrefaciens
isolated from spoiling fish. Int J Food Microbiol 9, 51–62.
Jørgensen, L.V., Huss, H.H., and Dalgaard, P. (2001). Significance of volatile com-
pounds produced by spoilage bacteria in vacuum-packed cold-smoked salmon
(Salmo salar) analyzed by GC-MS and multivariate regression. J Agric Food Chem
49, 2376–81.
Josephson, D. (1985). Enzymic generation of volatile aroma compounds from fish. In
Biogeneration of Aromas, ACS Symposium Series 317. American Chemical Society,
pp. 201–19.
Josephson, D.B., Lindsay, R.C., and Stuiber, D.A. (1983). Identification of compounds
characterizing the aroma of fresh whitefish (Coregonus clupeaformis). J Agric Food
Chem 31, 326–30.
Josephson, D.B., Lindsay, R.C., and Stuiber, D.A. (1984). Variations in the occurrences
of enzymically derived volatile aroma compounds in salt-and freshwater fish. J Agric
Food Chem 32, 1344–7.
540 Food Aroma Evolution

Kaewsrithong, J., Ushio, H., and Ohshima, T. (2001). Seasonal variation of phospha-
tidylcholine hydroperoxides in blood of sweet smelt Plecoglossus altivelis. Comp
Biochem Physiol Part B Biochem Mol Biol 130, 33–42.
Kawai, T. and Sakaguchi, M. (1996). Fish flavor. Crit Rev Food Sci Nutr 36, 257–98.
Koide, K., Yanagisawa, I., Ozawa, A., et al. (1992). Studies on the kerosene-like off-
flavour producing yeast in surimi minced fish meat-based products. Bull Jap Soc Sci
Fish 58, 1925–30.
Koutsoumanis, K. and Nychas, G.-J.E. (2000). Application of a systematic experimental
procedure to develop a microbial model for rapid fish shelf life predictions. Int J
Food Microbiol 60, 171–84.
Laing, D.G. and Jinks, A. (1996). Flavour perception mechanisms. Trends Food Sci
Technol 7, 387–9.
Lauritzsen, K. and Olsen, R.L. (2004). Effects of antioxidants on copper induced lipid
oxidation during salting of cod (Gadus morhua L.). J Food Lipids 11, 105–22.
Lauzon, H.L. (2000). Shelf-life and bacteriological spoilage of American plaice
(Hippoglossoides platessoides). In Seafood in Health and Nutrition, Transformation
in Fisheries and Aquaculture: Global Perspectives. Science Techbook, St. John’s,
Canada, pp. 327–54.
Leduc, F., Tournayre, P., Kondjoyan, N., et al. (2012). Evolution of volatile odorous com-
pounds during the storage of European seabass (Dicentrarchus labrax). Food Chem
131, 1304–11.
Lindsay, R. (1994). Flavour of fish. In: Seafoods Chem. Process. Technol. Qual. Springer,
pp. 75–84.
Lindsay, R.C. (1990). Fish flavors. Food Rev Int 6, 437–55.
Lindsay, R.C., Josephson, D.B., and Olafsdottir, G. (1987). Chemical and biochemi-
cal indices for assessing the quality of fish packaged in controlled atmospheres. In
Seafood Quality Determination, Proceedings of the International Symposium on
Seafood Quality Determination, edited by D.E. Kramer and J. Liston, Coordinated
by University of Alaska Sea Grant Collage Program, Anchorage, AK.
Lougovois, V.P. and Kyrana, V.R. (2005). Freshness quality and spoilage of chill-stored
fish. Food Pol Contr Res 1, 35–86.
Love, R.M. and Muslemuddin, M. (1972). Protein denaturation in frozen fish XII. The
pH effect and cell fragility determinations. J Sci Food Agric 23, 1229–38.
Maarse, H. (2017). Volatile compounds in foods and beverages. In Food Science and
Technology. CRC Press, pp. 784.
Martin, R. (1988). Contaminants in relation to the quality of seafoods. Food Technol
42, 104–8.
Miller, A., Scanlan, R.A., Lee, J.S., and Libbey, L.M. (1973). Identification of the volatile
compounds produced in sterile fish muscle (Sebastes melanops) by Pseudomonas
fragi. Appl Microbiol 25, 952–5.
Milo, C. and Grosch, W. (1993). Changes in the odorants of boiled trout (Salmo fario) as
affected by the storage of the raw material. J Agric Food Chem 41, 2076–81.
Milo, C. and Grosch, W. (1996). Changes in the odorants of boiled salmon and cod as
affected by the storage of the raw material. J Agric Food Chem 44, 2366–71.
Mohan, C.O., Ravishankar, C.N., and Srinivasagopal, T.K. (2016). Packaging interven-
tions in low temperature preservation of fish: A review. MOJ Food Proc Technol 2,
13–25.
Moore, J.E.A., Forrester, L.J., and Pelosi, P. (1976). Specific anosmia to isobutyralde-
hyde: The malty primary odor. Chem Senses 2, 17–25.
 Fish 541

Mottram, D.S. and Madruga, M.S. (1994). Important sulfur-containing aroma volatiles
in meat. In Sulfur Compounds in Foods, ACS Symposium Series. ACS Publications,
pp. 180–7.
Nollet, L.M.L. and Toldra, F. (2009). Handbook of Seafood and Seafood Products
Analysis. CRC Press.
Oehlenschlager, J. and Rehbein, H. (2009). Basic facts and figures. In Fishery Products:
Quality, Safety and Authenticity. Wiley-Blackwell, United Kingdom, pp. 1–18.
Olafsdottir, G., Hognadottir, A., and Martinsdóttir, E. (1998b). Application of gas
sensors to evaluate freshness and spoilage of various seafoods. In Methods to
Determine the Freshness of Fish in Research and Industry. International Institute
of Refrigeration, Paris, pp. 100–9.
Olafsdottir, G., Hognadottir, A., Martinsdóttir, E., and Jonsdottir, H. (2000). Application
of an electronic nose to predict total volatile bases in capelin (Mallotus villosus) for
fishmeal production. J Agric Food Chem 48, 2353–9.
Olafsdottir, G., Jonsdottir, R., Lauzon, H.L., et al. (2005). Characterization of vol-
atile compounds in chilled cod (Gadus morhua) fillets by gas chromatography
and detection of quality indicators by an electronic nose. J Agric Food Chem 53,
10140–7.
Olafsdottir, G., Li, X., Lauzon, H.L., and Jonsdottir, R. (2002). Precision and applica-
tion of electronic nose for freshness monitoring of whole redfish (Sebastes marinus)
stored in ice and modified atmosphere bulk storage. J Aquat Food Prod Technol 11,
229–49.
Olafsdottir, G., Martinsdóttir, E., and Jonsson, E.H. (1997a). Gas sensor and GC mea-
surements of volatile compounds in capelin (Mallotus villosus). Dev Food Sci 38,
507–20.
Olafsdottir, G., Martinsdóttir, E., Oehlenschlager, J., et al. (1997b). Methods to evaluate
fish freshness in research and industry. Trends Food Sci Technol 8, 258–65.
Olafsdottir, G., Verrez-Bagnis, V., Luten, J.B., et al. (1998a). The need for methods to
evaluate fish freshness. In Methods to Determine the Freshness of Fish in Research
and Industry. IIR, pp. 17–29.
Oliveira, H., Pedro, S., Nunes, M.L., et al. (2012). Processing of salted cod (Gadus spp.):
A review. Compr Rev Food Sci 11, 546–64.
Pan, B.S. and Kuo, J.M. (1994). Flavour of shellfish and kamaboko flavorants. In
Seafoods: Chemistry, Processing Technology and Quality. Springer, pp. 85–114.
Pigott, G.M. (1990). Flavors and acceptance of formulated seafood products. Food Rev
Int 6, 661–79.
Quaranta, H.O., Piccini, J.L., and Perez, S.S. (1984). Volatile acids content of irradiated
tuna loins as a criterion of freshness. Dtsch Lebens Rundsch 4, 106–7.
Ramezani, Z., Zarei, M., and Raminnejad N. (2015). Comparing the effectiveness of
chitosan and nanochitosan coatings on the quality of refrigerated silver carp fillets.
Food Contr 51, 43–8.
Rehbein, H. and Oehlenschlager, J. (2009). Fishery Products: Quality, Safety,
Authenticity. Wiley-Blackwell, UK.
Robertson, G.L. (2009). Food Packaging and Shelf Life: A Practical Guide. CRC Press.
Salum, P., Guclu, G., and Selli, S. (2017). Comparative evaluation of key aroma-active
compounds in raw and cooked Red Mullet (Mullus barbatus) by aroma extract dilu-
tion analysis. J Agric Food Chem 65, 8402–8.
Sampels, S. (2015). The effects of storage and preservation technologies on the quality of
fish products: A review. J Food Proc Preserv 39, 1206–15.
542 Food Aroma Evolution

Schultz, H.W. (1967). Symposium on Foods: The Chemistry and Physiology of Flavors.
Oregon State University.
Selli, S. and Cayhan, G.G. (2009). Analysis of volatile compounds of wild gilthead sea
bream (Sparus aurata) by simultaneous distillation–extraction (SDE) and GC–MS.
Microchem J 93, 232–5.
Selli, S., Prost, C., and Serot, T. (2009). Odour-active and off-odour components in rain-
bow trout (Oncorhynchus mykiss) extracts obtained by microwave assisted distilla-
tion–solvent extraction. Food Chem 114, 317–22.
Serot, T., Baron, R., Knockaert, C., and Vallet, J.L. (2004). Effect of smoking processes
on the contents of 10 major phenolic compounds in smoked fillets of herring (Cuplea
harengus). Food Chem 85, 111–20.
Shahidi, F. and Cadwallader, K.R. (1997). In Flavor and Lipid Chemistry of Seafoods,
ACS Symposium Series 674. American Chemical Society, Washington, DC, pp. 1–8.
Shahidi, F. and Han, X. (1993). Encapsulation of food ingredients. Crit Rev Food Sci
Nutr 33, 501–47.
Shewan, J.M., Mackintosh, R.G., Tucher, C.G., and Erhenberg, A.S.C. (1953). The devel-
opment of a numerical scoring system for the sensory assessment of the spoilage of
wet fish stored in ice. J Sci Food Agric 6, 183–98.
Sikorski, Z.E., Haard, N., Motohiro, T., and Pan, B.S. (1998). Quality. In Fish Drying
and Smoking. Production and Quality, edited by P.E. Doe. Lancaster, Technomic
Publishing Co. Inc., Basel, pp. 115.
Toldra, F. (1998). Proteolysis and lipolysis in flavour development of dry-cured meat
products. Meat Sci 49, S101–10.
Toldra, F., Aristoy, M.C., and Flores, M. (2000). Contribution of muscle aminopepti-
dases to flavor development in dry-cured ham. Food Res Int 33, 181–5.
Toldra, F. and Flores, M. (1998). The role of muscle proteases and lipases in flavor devel-
opment during the processing of dry-cured ham. Crit Rev Food Sci 38, 331–52.
Triqui, R. and Guth, H. (1997). Determination of potent odorants in ripened anchovy
(Engraulis encrasicholus L.) by aroma extract dilution analysis and by gas chro-
matography-olfactometry of headspace samples. In Flavor and Lipid Chemistry of
Seafoods. ACS Publications, pp. 31–8.
Undeland, I. (1997). Lipid oxidation in fish-causes, changes and measurements. In
Methods to Determine the Freshness of Fish in Research and Industry. International
Institute of Refrigeration, pp. 241–57.
Varlet, V., Knockaert, C., Prost, C., et al. (2006). Comparison of odour-active volatile
compounds of fresh and smoked salmon. J Agric Food Chem 54, 3391–401.
Varlet, V., Prost, C., and Serot, T. (2007). Volatile aldehydes in smoked fish: Analysis
methods, occurence and mechanisms of formation. Food Chem 105, 1536–56.
Whitfield, F.B. (1990). Flavour of prawns and lobsters. Food Rev Int 6, 505–19.
Whitfield, F.B. and Tindale, C.R. (1984). The role of microbial metabolites in food off-
flavours. Food Technol Aust 36, 204–13.
Withycombe, D.O.A. and Mussinan, C.Y.J. (1988). Identification of 2‐methyl‐3‐furan-
thiol in the steam distillate from canned tuna fish. J Food Sci 53, 658.
Zhang, C.H., Hirano, T., Suzuki, T., and Shirai, T. (1992). Enzymatically generated
specific volatile compounds in ayu (Plecoglossus altivelis) tissues. Bull Jap Soc Sci
Fish 58, 559–65.
 Chapter 26
Fruits and Vegetables
Rajnibhas Sukeaw Samakradhamrongthai

CONTENTS

26.1 The Perspective of Aroma Compounds from Fruits and Vegetables 544
26.2 Character Impact Compounds of Fruits and Vegetables 544
26.3 Aroma Compounds in Fruits and Vegetables 546
26.3.1 Biosynthesis of Volatile Compounds Responsible for Aroma in
Fruits and Vegetables 546
26.3.1.1 Amino Acid Pathway 547
26.3.1.2 Fatty Acid Pathway 547
26.3.1.3 Terpenoids Pathway 547
26.3.1.4 Carotenoid Pathway 548
26.3.2 Aroma Formation in Vegetables 548
26.3.3 Aroma Compound in Fruits and Vegetables and Its Occurrence 548
26.3.3.1 Aroma Compounds Containing Carbon, Hydrogen, and
Oxygen 548
26.3.3.2 Aroma Compounds Containing Oxygen Heterocycles and
Phenols 550
26.3.3.3 Aroma Compounds Containing Nitrogen 550
26.3.3.4 Aroma Compounds Containing Sulfur 551
26.4 Transitioning Factors on Aroma Compounds in Fruits and Vegetables 552
26.4.1 Effect of Maturity and Ripeness Stage 552
26.4.1.1 Preharvest Factors 552
26.4.1.2 Maturity and Ripeness Stage at Harvest 552
26.4.1.3 Postharvest Factors 552
26.4.2 Effects of Processing and Aging on Fruits and Vegetables Aromas 553
26.4.2.1 Non-Thermal Factors 553
26.4.2.2 Thermal Factors 554
26.4.3 Aging Factors 555
26.5 Matrix Structure of Fruits and Vegetables 556
26.6 The Interaction between Aroma Compounds and Fruit and Vegetable
Matrices 557
26.6.1 Interaction of Aroma Compounds and Carbohydrates 558
26.6.2 Interaction of Aroma Compounds and Proteins 558
26.6.3 Interaction of Aroma Compounds and Lipids 558
26.6.4 Interaction of Aroma Compounds and Other Compounds 559
26.7 Binding of Aroma Compounds in Fruits and Vegetables 559
26.8 Sensory Perception and Evaluation of Fruit and Vegetable Aromas 559
26.9 Conclusion 560
References 561

543
544 Food Aroma Evolution

26.1 THE PERSPECTIVE OF AROMA COMPOUNDS

FROM FRUITS AND VEGETABLES

There are several functions of aroma compounds in food, not only conveying the vital
character of the food and providing variety with regard to what we consume but also
alerting us to rancid and unsafe food, stimulating the appetite, and providing an emo-
tional connection to past experiences (Parker, 2015). Aroma compounds are highly
volatile, low-molecular weight compounds present at low levels. They can influence the
perception of five detectable basic tastes: sweet, salty, sour, bitter, and umami, and also
several potential other tastes including fatty (Chandrashekar et al., 2006; Meilgaard et
al., 2007; Mattes, 2009). Odor and taste are the typical characteristics of aromas. Owing
to the compounds’ volatility, these are perceived primarily in the nasal cavity, though
taste receptors exist in the mouth and are impacted when chewing. Color and appear-
ance may be the initial quality attributes that attract us to a fruit or vegetable product,
but aroma can deliver the largest impact on acceptability and the desire to repeatedly
consume a product. Different aroma compounds contribute to the final aroma via stimu-
lation of taste receptors in the mouth or by retronasal stimulation in the nasal cavity
during chewing (Goldberg et al., 2018). The complexity of interactions between many
compounds that contribute to the aroma of fresh produce makes it difficulties to arrive at
meaningful conclusions based solely on instrumental analysis. Sensory analysis provides
critical additional elements for evaluating the aroma of produce (Wilson and Stevenson,
2006; Le Berre et al., 2008). The aims of this chapter are to provide an overview of aroma
compounds typically identified in fruits and vegetables and to put these into context,
including the influences and interactions of aroma release through their matrices and
highlighting the character compounds that impact consumer perception.

26.2 CHARACTER IMPACT COMPOUNDS

OF FRUITS AND VEGETABLES

Aroma compounds from fruits and vegetables are mostly volatile compounds which can be
defined as esters, alcohols, aldehydes, and ketones. Numerous compounds from different
fruits and vegetables have been identified, but further investigation is needed to identify
how compounds contribute to the impact character of each commodity, their threshold
concentration, potency, and interaction within its matrices (Kader, 2018). Consequently,
all fruits and vegetables consist of potentially hundreds of volatile ­compounds emitting
different ratios and concentrations of these aroma molecules. A diverse array of com-
pounds contribute to the characteristics of fresh fruits and vegetables (Parker, 2015;
Maffei, 2016). As aroma is one of the most appreciated characteristics of fruits and veg-
etables, volatile compounds that provide aroma character are likely to play a key role
in determining the perception and acceptability of products by consumers. Identification
of key volatile aroma metabolites that carry the unique character of the natural fruit is
essential, as it provides the primary identification of fruit and vegetable characteristics and
their sensorial perception (Cheong et al., 2010). Aroma is very diverse and difficult to clas-
sify, but it can be split into many categories, for example, spicy, flowery, fruity, resinous
or balsamic, burned, and foul (Gould, 1983; Barrett, Beaulieu, and Shewfelt, 2010). It is
possible to characterize vegetables into four groups depending on their aroma compound
qualities. The first group has a single character impact compound: aroma is largely given
by one compound. Apples with ethyl butanoate (Dixon and Hewett, 2000), citrus fruit
 Fruits and Vegetables 545

with linalool (Berger, 2007), bananas with isoamyl acetate (Jayanty et al., 2002), are
examples of this group. The second group only involves a small number of limited volatile
compounds that provide a specific character to fruits and vegetables. Grapes with octanoic
acid and some volatile alcohols (Rosilllo et al., 1999), onions with characteristic sulfide
compounds (Husain, 2010), and celery with distinctive phthalides (Thappa et al., 2003)
are some of the fruits and vegetables in this group. The third group has combined char-
acter impact compounds where many compounds are required to reproduce the aroma.
There are many fruits and vegetables that exhibit a character influenced by combined
volatile compounds. Most are tropical fruits and vegetables such as mango, papaya, and
pineapple, along with others like garlic, parsley, and coriander (Jiang and Song, 2010;
Husain, 2010). The fourth group has specific character impact compounds whose aroma
cannot be reproduced even by the combination of many character compounds. Examples
in this group include snap beans, muskmelons, tomatoes, pear (trans-2, cis-4-decadieno-
ate), cucumber (trans-2, cis-6-nonadienal), green pepper (2-isobutyl-3-methoxypyrazine),
raw potato (2-isopropyl-3-methoxypyrazine), cooked mushrooms (1-octen-3-one), and
beetroot (geosmin) (Gormly, 1981; Barrett, Beaulieu, and Shewfelt, 2010; Husain, 2010).
Interestingly, the generic green notes of all fruits and vegetables are mostly highly volatile
carbon chains (C5 –C9), and species character is given by its derivative to generate character
impact compounds. It is also worth remarking, however, that not all fruits and vegetables
have character impact compounds; their unique character can be attributed to a combina-
tion of aroma compounds. Table 26.1 shows a list of character impact compounds in com-
mon with fruits and vegetables; their structures are discussed in the following sections. In
addition to hydroxy compounds, aldehydes, ketones, acids, esters, and sulfur-containing
compounds, oxygen heterocyclics have also been found to be important. Pyrazines are
important components in vegetables, especially those with a methoxyl group. In others,
apples, raspberries, tomatoes, and celery aroma is given by a character impact compound
plus several other contributory aroma compounds (Tamura et al., 2011; Parker, 2015).

TABLE 26.1 Character Impact Compounds of Various Fruits and Vegetables

Characteristic Produce Character Compound Reference
Fruit
Apple (Red) 2-Methyl butan-1-ol Dixon and Hewett, 2000
Apple (Green) Ethyl-2-methyl butanoate Dixon and Hewett, 2000
Banana Isoamyl acetate Ji and Srzednicki, 2015
Grape Methyl anthranilate Rosilllo et al., 1999
Mango 4-Hydroxy-2,5-dimethyl-3(2H)-furanone Pino and Mesa, 2006
Pineapple Allyl hexanoate Tokitomo et al., 2005
Peach γ-Decalacton Aubert and Milhet, 2007
Vegetable
Garlic S-alk(en)yl cysteine sulfoxides Edris and Fadel, 2002
Cucumber (E,Z)-2,6-nonadienal Buescher and Buescher,
2001
Onion 3-Mercapto-2-methyl pentane-1-ol Granvogl et al., 2004
Pumpkin Hexanal, (E)-2-hexenal, (Z)-3-hexen-1-ol, Maarse, 1991
and 2,3-butanedione
Tomato (Z)-3-hexenal and beta-ionone Hayata et al., 2002
Source: Adapted from Jiang and Song, 2010; Husain, 2010; Parker, 2015.
546 Food Aroma Evolution

26.3 AROMA COMPOUNDS IN FRUITS AND VEGETABLES

The strength and character of aroma are important parameters for consumers to dis-
tinguish freshness and maturity. There are over a thousand compounds that have been
identified in plant volatiles. The composition of volatile compounds, even in single plant
material, is complicated. For example, there are approximately 270 volatile compounds
detected in mango (Pino and Mesa, 2006), and more than 400 volatile compounds iden-
tified in the fruit of the tomato (Hayata et al., 2002; Maneerat et al., 2002). Since each
compound has a different odor characteristic and threshold level for recognition (Zhou,
Klee, and Sarnoski, 2018), the compositional differences of aroma compounds can help
to differentiate fruits and vegetables from different plant parts, from different species or
cultivars, from different harvested areas, and, occasionally, from different maturity/rip-
ening stages. Understanding the biosynthesis mechanism of each aroma compound may
result in the creation of more appreciated and high eminence plant materials. Therefore,
it is valuable to have a closer look at the metabolic pathways that provide an increase
in aroma formation in fruits and vegetables (Song and Forney, 2008). There are sev-
eral pathways involved in volatile compound formation starting from polysaccharides,
amino acids, lipids, terpenoids, and carotenoids. The initiated compound is able to pro-
duce different volatile compounds via these pathways, the multiplicity of the compound
is accomplished through additional modification reactions such as hydrolysis, degrada-
tion, acylation, methylation, oxidation/reduction, and cyclic ring closure (Figure 26.1).

26.3.1 Biosynthesis of Volatile Compounds Responsible

for Aroma in Fruits and Vegetables

As volatiles comprise at least five chemical classes, there are several pathways involved
in volatile biosynthesis. These have not been fully described but appear to be common

FIGURE 26.1 Schematic pathway of aroma compound formation from a natural carbon
pool. (Adapted from Schwab, Davidovich-Rikanati, and Lewinsohn, 2008.)
 Fruits and Vegetables 547

in different fruits and vegetables. Amino acids, membrane lipids, and carbohydrates are
highly important in the biosynthesis of volatile compounds, influencing aroma formation.
Though volatile compounds are synthesized via a few major biochemical pathways, vari-
ous forms of enzymatic modifications, such as hydroxylations, acetylations, and methyl-
ations, increase the diversity of the released volatiles by increasing their volatility in the
final step of their formation (Gang, 2005; Hadi et al., 2013). An important step in the
biosynthetic pathway of aroma compounds is the availability of primary precursor sub-
strates, including fatty acids and amino acids, which are highly regulated during fruit and
vegetable development in terms of amount and composition (Song and Bangerth, 2003).

26.3.1.1 Amino Acid Pathway

Amino acids can undergo initial deamination or transamination, leading to the formation
of the corresponding α-keto acid. Subsequent decarboxylation is followed by reductions,
oxidations, and/or esterification to aldehydes, acids, alcohols, and esters (Reineccius,
2006). The formation of volatile branched-chain compounds, alcohols, aldehydes, and
esters in fruits and vegetables such as banana, apple, strawberry, and tomato originate
from the branched-chain amino acids leucine, isoleucine, and valine (Perez et al., 2002;
Goff and Klee, 2006).

26.3.1.2 Fatty Acid Pathway

Fatty acids are major precursors of aroma compounds in most fruits and vegetables.
The straight-chain fatty acid-derived alcohols, aldehydes, ketones, acids, esters, and lac-
tones ranging from C1 to C20 are important character impact aroma compounds that are
responsible for fresh fruit aromas with high concentrations. They are basically formed
by three processes: α-oxidation, β-oxidation, and the lipoxygenase pathway (Sanz, Olias,
and Perez, 1997; Schwab and Schreier, 2002).

26.3.1.2.1 ß-Oxidation
α- and β-oxidation are major processes in the formation of aroma molecules in all organ-
isms undertaken by the degradation of straight-chain fatty acids. The specific pathways
in plants can be explained through amino acid metabolism and biosynthesis of fatty acid
catabolism but also through hormonal compounds. β-oxidation results in a successive
removal of C2 units (acetyl CoA) from the parental fatty acid (Baker et al., 2006).

26.3.1.2.2 Lipoxygenase
For polyunsaturated fatty acids, lipoxygenase (LOX) is a metabolism pathway which
happens during the ripening of fruit. Cell wall and membranes become more perme-
able; the LOX pathway becomes active without tissue disruption (Sanz, Olias, and Perez,
1997). The LOX biosynthetic pathway can be an alternative to the β-oxidation of fatty
acids when the produce is ripening to provide substrates for ester production and aroma
formation. It can avoid the oxidation that produces an undesirable aroma or off-aroma
(Hadi et al., 2013).

26.3.1.3 Terpenoids Pathway
Terpenoids are the main class of plant secondary metabolites with many volatile com-
plexes. Hemiterpenes (C5), monoterpenes (C10), sesquiterpenes (C15), homoterpenes (C11
and C16), and some diterpenes (C20) have a high vapor pressure allowing the aroma
release into the atmosphere. Terpenoids are derived from the universal C5 precursor as
isopentenyl diphosphate (IPP) and its allylic isomer dimethylallyl diphosphate (DMAPP),
548 Food Aroma Evolution

which in higher plants are generated from two independent pathways located in separate
intracellular compartments (Dudareva et al., 2005).

26.3.1.4 Carotenoid Pathway
The carotenoid metabolism pathway can create isoprenoid molecules which are impor-
tant for aroma formation in fruits and vegetables. The oxidation reaction of carotenoid
leads to the production of apocarotenoids, which are catalyzed by a family of carotenoid
cleavage dioxygenases (CCDs). These apocarotenoid volatiles are synthesized only at the
latest stage of ripening, even though the CCD enzymes are present throughout the devel-
opment of fruits, vegetables, and flowers. Nonetheless, the initial dioxygenase cleavage
step can create character compounds in some produce: Arabidopsis (Arabidopsis thali-
ana), tomato (Solanum lycopersicum), and petunia (Petunia hybridea) (Auldridge et al.,
2006; Rosati, Diretto, and Giuliano, 2009)

26.3.2 Aroma Formation in Vegetables

Vegetables do not have a ripening period as fruits do; thus, the aroma formation is also
different. Aroma in vegetables is primarily released during cellular disruption. It is a
consequence of a cutting prior to cooking or the mastication process. Cellular disrup-
tion allows enzymes to mix with substrates, separated within the vegetable cell, result-
ing in the evolution of volatile aroma substances. Meanwhile, fatty acid, carbohydrate,
and amino acid metabolism also serves to deliver precursors of the vegetable aroma
(Reineccius, 2006; Barrett, Beaulieu, and Shewfelt, 2010).

26.3.3 Aroma Compound in Fruits and Vegetables and Its Occurrence

The aroma of fruits and vegetables is considered to be a complex mixture. Four major
classes of compounds which contribute to the fruit and vegetable aroma are described
in the subsequent sections. The most important volatile materials of citrus fruit include
terpene hydrocarbons, carbonyl components, alcohols, esters, and volatile organic acids.
These compounds are found in the flavedo and oil sacs that are embedded in the juice
vesicles (Hadi et al., 2013). The structures of volatile compounds found in plants are
diverse. However, aroma compounds can be categorized by their structural similarities
and their relationships in biosynthesis pathways.

26.3.3.1 Aroma Compounds Containing Carbon, Hydrogen, and Oxygen

26.3.3.1.1 Aldehydes
Aldehydes are components that are comprised of a straight-chain carbon. The C3–C5
aldehydes (propanal, butanal, and pentanal) tend to have a rather chemical/malty/green
note that is hard to define. The branched-chain C5 aldehydes exhibit low thresholds of
aroma and are found in most fresh fruits and vegetables, whereas C6 aldehydes such as
hexanal become characteristically green (Engel et al., 2002).
Studies indicate that hexanal is the aroma most found in apples (Fuhrmann and
Grosch, 2002), tomatoes (Mayer et al., 2003), Pontianak oranges (Fischer et al., 2008),
and pomegranates (Cadwallader et al., 2010). The E- and Z-isomers which are consid-
ered as unsaturated aldehydes also tend to have lower odor thresholds and are often
character impact compounds, with the shorter chain analogs delivering green aromas,
 Fruits and Vegetables 549

such as cis-3-hexenal in tomatoes (Buttery et al., 1971). At greater than six carbon
atoms, the aroma starts to become fatty and tends to provide a fishy aroma, which
is undesirable in fruits and vegetables. However, there are higher carbons, such as
C9 –C14, that give fruit and vegetable notes such as (Z)-6-nonenal (cucumber note),
(E)-2-alkenals (coriander note), isopropyl-2-hexenal (woody, lavender), and 5-methyl-
2-phenyl-2-hexenal (chocolate aroma) (Parker, 2015). In addition, there are aromatic
ring aldehydes such as benzaldehyde (cherry, almond), phenylacetaldehyde (rose, honey),
and cinnamaldehyde (cinnamon) that are also important components of the aroma in
fruits and vegetables. The most universal of all aroma compounds can be indicated by
vanillin, which is an aldehyde with the chemical name 4-hydroxy-3-methoxybenzalde-
hyde (Elmore et al., 2014).

26.3.3.1.2 Alcohols
Alcohols are a part of the aroma that can be identified in fruits and vegetables, but their
contribution to aroma tends to be less than aldehydes. Straight-chain alcohols commonly
found in fruits increase with their maturity (Parker, 2015). A double-bound molecule
of alcohols provides characteristic point notes of aroma, for example, cis-3-hexen-1-ol
(a characteristic green leaf note) and 1-octen-3-ol (a characteristic of mushrooms).
Moreover, the bicyclic alcohols such as geosmin and 2-methyl isoborneol can convey
earthy and musty notes with very low odor thresholds that provide characteristic notes to
beetroot and baby corn (Murray et al., 1975; Mottram et al., 2011). Its presence is attrib-
uted to the action of actinomycetes in the soil in which the produce was grown.

26.3.3.1.3 Ketones
Examples of straight-chain methyl ketones, containing one carbonyl group are 2-heptanone
(fruity pear aroma), and 3-octanone (earthy, mushroom notes). The more structurally com-
plex ketones play a key role in the aroma of fruits and vegetables, such as filbertone (hazelnut
aroma), geranyl acetone (floral rose aroma), and raspberry ketone (sweet, and raspberry
aroma). In vegetables, the carotenoid-derived ketones like β-damascenone and β-ionone are
found to provide woody notes to red peppers (Jun and Kim, 2002) and carrots (Buttery and
Takeoka, 2013). Additionally, these ketones can provide a trace note in orchard fruits and
deep juicy notes, for example, in berries, tomatoes, and apples (Parker, 2015).

26.3.3.1.4 Esters
Esters give the fundamental aroma of most fruits and vegetables. Ethyl acetate can be
found in most ripe and ripening fruit, for example, melons, apples, pineapples, and straw-
berries (Gonzalez et al., 2009). Ethyl esters are major components of fruit aroma, par-
ticularly in ripe fruits where the production of ethanol has boosted their formation. Some
esters can be characteristic for specific fruits: 3-methyl butyl acetate is characteristic of
pear or pear drops, allyl hexanoate is typical for pineapple, cis-3-hexenyl butanoate con-
veys a leafy green aroma, and the higher (C9) esters are important for melon aroma. The
longer chain ethyl esters become soapy, cheesy, and waxy, possibly affecting the rejection
of fruits and vegetables by the consumer (Qian and Reineccius, 2003).

26.3.3.1.5 Lactones
Cyclic esters as lactones are potent aroma compounds formed from hydroxy acid.
Γ-lactones such as γ-octalactone and γ-decalactone convey peach, coconut, and creamy
aromas in peach and coconut milk. Subsequently, these are very widespread in the
tropical aroma and flavor industry. Besides, jasmine lactone provides a floral petal-like
550 Food Aroma Evolution

aroma to green tea. However, there are several lactones identified as sweet cream butter.
5-hydroxydec-2-enoic acid lactone is found in cocoa and used in the production of choco-
late (Katsuno et al., 2014; Parker, 2015).

26.3.3.1.6 Terpenes and Terpenoides

The major aroma compounds in essential oil are terpenes, terpenoids, and sesquiterpenes,
which are responsible for the characteristic aroma profile of many fruits (particularly cit-
rus), herbs, and spices. Units of isoprene (C5H10) are compiled and biosynthesized to cre-
ate terpenes, terpenoids, and sesquiterpenes. These can be structures in linear, cyclic, or
polycyclic form. However, those that are responsible for odor tend to contain two or three
isoprene units (monoterpenes and sesquiterpenes, respectively). Other terpenes are less
common in fruits and herbs such as sabinene (citrus) and α-thujene (woody). Sesquiterpenes
can provide more interesting aromas with α-valencene which conveys typical citrus notes
and both farnesene and humulene give a woody spicy note, whereas pinene, myrcene, and
ocimene are major volatile compounds with a basil aroma (Parker, 2015).

26.3.3.2 Aroma Compounds Containing Oxygen Heterocycles and Phenols

26.3.3.2.1 Furans and Furanoids
Furans can be found in fresh produce. Furanoid terpene, such as linalool oxide which
conveys a floral herby note, tends to appear during storage and is indicative of oxidation.
Generally, most furans result from thermal processing. These are a result of lipid oxida-
tion or the Maillard reaction. Another bicyclic furanoid, theaspirane, can be found in
tea, grapes, wine, and sherry with camphoraceous notes, blackcurrant notes, and naph-
thalene notes (Collin et al., 2011). Furaneol is another example of a furanoid that can
be detected in low concentrations. It can provide sweet caramel notes to strawberry,
pineapple, tomatoes, and tea (Parker, 2015). Furthermore, eucalyptol, which possesses a
camphor-like aroma, is the major aroma component of bay leaf, along with eugenol, cis-
3-hexenal, isoeugenol, and linalool (Hadi et al., 2013).

26.3.3.2.2 Phenols
Many phenols are particularly aroma-active compounds in fruits and vegetables. For
example, 4-methyl guaiacol is described as sweet, candy-like, vanilla, leathery, spicy, and
smoky. Vanillin is also the most popular aroma and the key constituent of vanilla (Vanilla
planifolia L.). It is a phenol with sweet vanilla aroma and smoky undertones. The most
aroma-active phenols are the chlorophenols and bromophenols with related anisoles that
are often implicated as taints. Additionally, terpene-derived phenols such as thymol and
eugenol are examples of phenols that provide character impact notes to thyme and cloves,
respectively (Parker, 2015).

26.3.3.3 Aroma Compounds Containing Nitrogen

Amine, pyrroles, pyridine, and pyrazines are nitrogen-containing aroma compounds that
are relatively rare in nature, but plentiful in cooked foods where the Maillard reaction is
responsible for generating a series of nitrogen heterocycles. In this section, there will be a
discussion on indoles and pyrazines as they convey the most aroma-related notes in fruits
and vegetables, such as floral and green notes.

26.3.3.3.1 Indole
Indole is a powerful nitrogen-containing compound that, to some individuals, conveys a
pleasant floral and green note in fruits and vegetables, such as green tea (Katsuno et al.,
 Fruits and Vegetables 551

2014) and pomelo (Cheong et al., 2011). However, it also conveys an unpleasant fecal
note that is often associated with boar taint. The related 3-methylindole skatole exists in
the aroma of lilies, which can be perceived as pleasant and floral for some people, but it
can be unpleasant for others.

26.3.3.3.2 Pyrazines
Pyrazines found in uncooked potatoes and other vegetables are methoxy-substituted and
powerful odorants. 2-methoxy-3-isobutyl pyrazine is the character impact compound
in green bell pepper and is identified as the most potent odorant in raw French beans.
The homologous 2-isopropyl-3-methoxypyrazine is known as bean pyrazine because it
conveys earthy, pea, and beany notes to soy milk, and earthy notes to potatoes. It is
also found to be aroma-active in parsley leaves and having gravies aroma in vegetables
(Parker, 2015; Siegmund, 2015).

26.3.3.4 Aroma Compounds Containing Sulfur

26.3.3.4.1 Sulfides
The simple sulfides (dimethyl sulfide, dimethyl disulfide, dimethyl trisulfide, and the par-
ent thiol methanethiol) make an important contribution to cooked aromas, even though
individually their aromas are objectionable and sulfurous. Dimethyl sulfide is important
for fruit aroma as well as for sweetcorn and asparagus aromas (Parker, 2015). Dimethyl
trisulfide is the major cause of off-flavors in overcooked Brassica vegetables. Dimethyl
trisulfide, dimethyl thiosulfinate, dimethyl thiosulfonate, and 2-methyl thiopyridine are
identified as aroma-active breakdown products arising from S-methyl cysteine sulfoxide
(Kubec et al., 1998).

26.3.3.4.2 Thiophenes
The aroma of simple thiophenes are often described as sulfurous and it is generally not
very potent. Coffee contains many sulfur compounds. Kahweofuran is a bicyclic thio-
phene found in coffee, conveying a roasted, smoky, and sulfurous aroma, whereas thio-
phenemethanol conveys a coffee-like aroma. The longer chain alkylthiophenes, such as
2-pentylthiophene and alkylthiazoles, have relatively little aroma and seem to act as a
sink for excess hydrogen sulfide (Elmore et al., 2000).

26.3.3.4.3 Thiazoles and Thiazolines

Thiazoles tend to give cooked, roasted, and toasty notes. Many substituted thiazoles
have been identified in potatoes, but the thiazole that appears most frequently is 2-acetyl-
thiazole. It conveys a nutty, roasted, popcorn aroma, and the related compound, 2-ace-
tyl-2-thiazoline, provides a lower threshold of freshly baked bread. 2-isobutylthiazole is
present in raw tomatoes and gives them their characteristic green note (Hongsoongnern
and Chambers, 2007).

26.3.3.4.4 Thioesters and Mercaptoesters

These esters are intense and objectionable in concentrated form, but when diluted suf-
ficiently, they give the characteristic fruity and tropical aromas to many tropical fruits.
S-Methyl 2-methyl-butane thioate, ethyl methylthio acetate, and ethyl 3-(methylthio)
propanoate are aroma-active in ripe Charentais melons (Lignou et al., 2014) and seem to
contribute to the ripe aroma of melon with 3-Mercaptohexyl acetate has also been indi-
cated to be important in guava fruit (Sinuco et al., 2010)
552 Food Aroma Evolution

26.3.4.4.5 Isothiocyanates
Isothiocyanates are hydrolysis products of glucosinolates and are found in high concen-
trations in Brassica vegetables and recognized by their pungent aromas (Ghawi et al.,
2012). Allyl isothiocyanate is found to be a key odorant, contributing pungent, black
mustard-like notes in cooked cauliflower (Engel et al., 2002). Correspondingly, methyl
thiocyanate, butyl isothiocyanate, 2-methyl butyl isothiocyanate, and sulfides have been
found to be important in broccoli aroma. In addition, 4-methyl thiobutyl and 5-methyl
thiopentanenitril are noted to be abundant aroma-active compounds in salad rocket
(Eruca sativa) (Jirovetz et al., 2002; Jacobsson et al., 2004).

26.4 TRANSITIONING FACTORS ON AROMA

COMPOUNDS IN FRUITS AND VEGETABLES

Aroma compounds are responsible for the distinctive aromas associated with fruits and
vegetables, and these compounds can be categorized into endogenous (naturally occur-
ring) and secondary compounds resulting from the enzymatic reaction. Several factors,
such as maturity, ripening, processing, and aging, affect the transitioning of aroma in
fruits and vegetables (Beaulieu and Baldwin, 2002; Forney, 2008)

26.4.1 Effect of Maturity and Ripeness Stage

26.4.1.1 Preharvest Factors
The influences of harvest maturity and storage time on compounds that serve as precur-
sors for aroma formation are critical factors that determine the ultimate levels of vola-
tiles. Climatic conditions (temperature, light, rain, and wind) and cultural applications
(planting density, tree pruning, fruit thinning, nutrient, and water quantities; control of
weeds, diseases, and insects), result in high yield but often produce a less than optimal
aroma quality (Hewett, 2006).

26.4.1.2 Maturity and Ripeness Stage at Harvest

The maturity stage at harvest is one of the most important factors influencing the aroma
quality of fruits and vegetables. Non-climacteric plants are better tasting when harvested
immature, while climacteric plants are the better tasting when harvested fully ripe.
However, harvesting fruits and vegetables before they reach optimal maturity is a com-
mon commercial practice. Another reason for harvesting climacteric plants before the
optimal maturity stage is reached is to sufficiently guarantee that the aroma withstands
handling procedures and to maximize their storage potential. However, some fruits such
as apples can be harvested at the early pre-climacteric stage and kept in either air or
controlled atmospheres for various durations before sale, never reaching a good eating
quality (Fellman et al., 2003; Kader, 2018).

26.4.1.3 Postharvest Factors
There is information about optimal harvesting and handling procedures based on reduc-
ing quantitative losses through the maintenance of appearance and textural quality of
fruits and vegetables (Kader, 2018). Changes in volatiles and aroma that occur during
marketing and storage present an additional challenge. The development of technology
can provide more accurate control over the postharvest conditions, including temperature,
 Fruits and Vegetables 553

humidity, and atmosphere composition (Forney, 2001). The longer the period between
harvesting and consuming, the greater the losses of characteristic aromas and the develop-
ment of off-aromas of tomatoes (Kader et al., 1978) and strawberries (Pelayo-Zaldivar et
al., 2005). Therefore, it is very important to identify optimal postharvest handling condi-
tions (time, temperature, relative humidity, and atmospheric composition) that maintain
the aroma quality of fruits, vegetables, and their value-added products. The postharvest
quality of fruits and vegetables should be based on aroma rather than appearance. The
loss of aroma can happen due to the losses of sugars, acids, and volatile compounds, espe-
cially esters, together with off-aromas from the fermentation metabolism. Off-aromas
can arise from environmental sources and from the aroma–matrix interactions, all of
which can unbalance the aroma profile (Sucan, 2004). The individual contributions of
aroma compounds and their interactions in terms of the overall aroma quality need to be
determined for fruits and vegetables. The effect of maturity, harvest, handling, storage
temperature, and shelf-life period needs to be evaluated for aroma quality. The effects
of using an ethylene inhibitor on the aroma quality of fruits and vegetables need to be
determined in order to evaluate the extension of postharvest aroma quality.
Furthermore, a lot of research has revealed the influence of maturity and ripening on
aroma volatiles and flavor in fruits and vegetables. The evolution of volatile compounds
can be classified and comprehended depending on whether there are decreasing, increasing,
or mixed responses of the concentrations in relation to the harvest and storage date (Table
26.2). This can provide a prediction for the harvesting and storage of fruits and vegetables
which can be of great advantage and encourage producers to harvest at partially ripe to fully
ripe stages by developing handling methods that protect the fruits from physical damage.

26.4.2 Effects of Processing and Aging on Fruits and Vegetables Aromas

26.4.2.1 Non-Thermal Factors
Chilled storage will generally reduce the rate of aroma loss. However, while chilled stor-
age maintains strawberries in good condition for days as far as appearance and texture are

TABLE 26.2 Volatile Compound Evolution of Pineapple and Avocado

during Harvesting and Storage
Increasing Mixed Response
Produces Decreasing Compounds Compounds Compounds
Pineapple Acetic acid, butyl ester Hexanoic Ethyl acetate
Butanoic acid, 2-methyl- 1-Butanol,
methyl ester 3-Methyl-
Butanoic acid, ethyl ester acetate
Octanoic acid, methyl ester 1-Butanol,
Octanoic acid, ethyl ester 2-Methyl-acetate
Avocado 1-Penten-3-one Acetaldehyde Limonene
Hexanal Methyl acetate
(E)-2-Hexanal Pentanal
2,4-Hexadienal β-Myrcene
Benzaldehyde 2,4-Heptadienal
Nonanal
Source: Adapted from Obenland et al., 2012; Thu et al., 2017.
554 Food Aroma Evolution

concerned, the loss of aroma can be noticeable. Certain forms of packaging, for example,
pre-packaging with synthetic films, can conserve aroma. Most produce yield significant
numbers of volatile compounds as indicators of its ripening. Many of these volatile com-
pounds are produced in trace amounts, which are below the thresholds of most analytical
instruments but can be detected by human olfaction (Goff and Klee, 2006). Volatiles can
be classified as primary or secondary compounds, indicating whether they were present
in the intact fruit tissue or produced as a result of tissue disruption (Jordan, Seelye, and
McGlone, 2001). Analysis of volatiles from either intact or disrupted fruit tissues will
influence the aroma profiles and final aroma interpretation. The volatile profiles of fruit
are complex and vary depending on the cultivar, ripeness, pre- and postharvest environ-
mental conditions, fruit sample (either intact fruit, slices, or homogenized samples), and
analytical methods utilized (Kader, 2018).

26.4.2.2 Thermal Factors
The initial thermal treatment of processed products can cause loss of water-soluble and
oxygen-labile nutrients (Rickman, Barrett, and Bruhn, 2007). There are water-soluble
compounds like phenolic compounds that change during processing and aging. This
appears to be highly variable in fruits and vegetables. This processing provides greater
convenience to the consumer and introduces higher diversity into their diet. These
changes are however accompanied by changes in the aroma quality of fruits and veg-
etables. Even though aroma can evolve during processing, cooking, and aging, the altera-
tion can be negligible when relative to year-round availability and convenience (Barrett,
Beaulieu, and Shewfelt, 2010). Aroma compounds are often only released by cell disrup-
tion (Vicente, Greve, and Labavitch, 2006). Some aroma compounds are bound to sugars
such as glycosides or glucosinolates. The glycosides in aroma compounds in fruits are
mainly O-β-D-glucosides and O-diglycosides, but triglycosides have also been identi-
fied (Vicente et al., 2007). The proportion of glycosidically bound volatiles is usually
greater than that of the free volatiles, making them an important potential source of
aroma compounds. The odorous aglycones may be released from the sugar moiety during
maturation, processing, and storage, or by the action of enzymes, acids, or heat (Bood
and Zabetakis, 2002). In Table 26.3, Utama-ang et al. (2018) studied the effect of garlic
preparation and its drying conditions through its physicochemical and sensory proper-
ties. Chopped garlic with higher drying temperatures suggested that aroma evolved dur-
ing the preparation and processing period which was caused by cell disturbance, and the

TABLE 26.3 Effect of Preparation Method and Drying Temperature on

Allium sativum L. toward Its Main Aroma
Garlic Main Aroma (mg/g Dry Basis)
Preparation Drying Allyl Methyl Dimethyl Dimethyl
Method Temperature (°C) Sulfide Difulfide Trisulfide
Chopped 40 1.71±0.05c 1.14±0.02b 0.85±0.02c
Chopped 60 6.17±0.01a 5.39±0.02a 4.18±0.01a
Sliced 40 0.42±0.01d 1.34±0.01b 0.26±0.01b
Sliced 60 2.77±0.01b 1.12±0.0.1b 0.84±0.01c
Source: Adapted from Utama-ang et al., 2018.
Note: The different letter in the same column indicates the significant differ-
ence (p<0.05) of all treatment.
 Fruits and Vegetables 555

increasing temperature also accelerated the reaction of the aroma formation pathway to
create the specific aroma of the fruits and vegetables. The appropriate processing condi-
tion can influence aroma, which potentially provides an indicator of how to control the
desirable volatile compounds in fruits and vegetables through the production. It is also
indicated that a suitable temperature and preparation method can provide maximum
aroma content with good sensory quality.

26.4.2.2.1 Processed Fruits and Vegetables

Blanching results in loss of aroma in fruits and vegetables and destroys enzymes which
are responsible for aroma development. In canned and dried vegetables, aroma is lost
during heating. This can imply that canned and dried products can sometimes provide
undesirable aromas (Mukherjee and Chattopadhyay, 2007). In canned fruits and veg-
etables, the aroma is diluted by the syrup or brine, respectively; in food, volatiles are
lost during drying, while in frozen foods a gradual loss of volatiles during storage occurs
(Devi et al., 2015).

26.4.2.2.2 Fruit and Vegetable Products

Vegetable powders, jams, fruit juices, fruit pulps, wine, and other items are included
under fruit and vegetable products. The aromas from those products are also of impor-
tance as they are used as ingredients that are largely based on the volatile components of
vegetable powders and chopped vegetables, especially onion (Forney, 2008). The impor-
tance of aroma in fruit and vegetable products is greater than realized as the soluble
solids and natural acidity can affect the color, taste, texture, nutritional value, and
aroma of the product with the addition of sucrose and citric acid (Barrett, Beaulieu, and
Shewfelt, 2010). In addition, the volatile compounds can both evolve and degrade during
the process (non-thermal and thermal). The volatile compound that corresponds to the
distinctive aroma can evolve and degrade through a maceration and pressing process.
The maceration process can increase the content of hexanal, trans-2-hexenal, and other
lipid oxidation products through stimulation of lipoxygenase activity (Boylston, 2010).
Some of the non-thermal processes can increase the yield of processed juice and increase
the content of esters and desirable volatile compounds to enhance the aroma in fruits
and vegetables. The heat treatment of low pH results in the hydrolysis of glycosides and
the release of aroma compounds. During the heating process, the glycosidic precur-
sors of several key aroma compounds, including raspberry ketone and damascenone
(Roberts and Acree, 1996), are degraded to release these potent aroma compounds and
increase the aroma intensity of the fruit. Pectinases processes are performed on fruits
and vegetables to improve the clarity of the product and may increase the content of the
volatile compounds released from the glycosidically bound precursors (Cheetam, 2002;
Boylston, 2010).

26.4.3 Aging Factors

Many specific aroma compounds have been identified in fruits and vegetables. Enzymes
and genes control their production. It is known that volatile compounds are mainly derived
from three biosynthetic pathways: lipases, hydroperoxide lyases, and cleavage enzymes on
lipid components, followed by the action of alcohol dehydrogenases (Oke and Paliyath,
2010). Further, outstanding aroma compounds such as eugenol, phenethyl alcohol, and
guaiacol are derived from the shikimic acid pathway and nor-isoprenoids, such as β-ionone
556 Food Aroma Evolution

and geranyl acetone, are degraded from long-chain terpenoids, like β-carotene and lyco-
pene. Monoterpenes such as linalool are formed directly from geranyl diphosphate via the
isoprenoid pathway (Oke and Paliyath, 2010). There are numbers of fruits and vegetables
that provide different aromas while aging through the ripening process after harvesting.
The fresh green characteristic of short-chained aldehydes decreases in ripening produce
such as guavas, apples, coriander, and strawberries. Furthermore, the green aroma com-
pounds only increase until the ripening period is reached and then decrease. The mono-
terpene profile also changes during the ripening of coriander and citrus fruits (Yang et al.,
2009; Asif, 2012). In some fruits and vegetables, there is a reduction of linalool as matura-
tion progresses, whereas the limonene content increases in citrus cultivars (Rodríguez et
al., 2011). In many fruits (apple, mango, strawberry, kiwi, papaya, and guava), there is an
increase of esters and lactones (Hadi et al., 2013). In tomatoes and peaches, apocarotenoid
and furanone-derived compounds are not emitted until relatively late in fruit ripening, and,
during this process, the amount of apocarotenoids increases by a factor of 40 (Mathieu
et al., 2009; Brandi et al., 2011). Interestingly, in strawberries and kiwi, sulfur volatiles
increased by as much as 100% when they mature (García et al., 1999; Du et al., 2011).

26.5 MATRIX STRUCTURE OF FRUITS AND VEGETABLES

There is complexity in food components consisting of polysaccharides, proteins, lipids,

vitamins, minerals, and other bioactive compounds. These interact with each other and
with the aroma compounds. The nature of the aroma compounds in the matrix is consid-
ered an important factor toward evolution effects from processing to aging. Aroma com-
pounds can be discharged, and this depends on their availability in the vapor phase and on
their affinity for the matrices. Fruit and vegetable matrices are comprised of many matrix
components (Husain, 2010). Plant tissues are multiphase systems with an intricate internal
microstructure formed by cells, intercellular spaces, capillaries, and pores. Four major types
of mature plant tissues are parenchyma (storage cells), vascular (conducting cells), support-
ing cells, and protecting cells (Alzamora et al., 2005) (Figure 26.2). Edible portions of most
fruits and vegetables are composed of fleshy parenchyma cells, which form the bulk of the

FIGURE 26.2 Schematic representation of a parenchyma cell in plants.

 Fruits and Vegetables 557

softer parts of plants. It is approximately 50 to 500 µm across and polyhedral or spherical

in shape. One or more vacuoles contain a watery solution of organic acids, salts, pigments,
and aroma compounds that are responsible for the osmotic potential of the cell (Alzamora
et al., 2017). In fruits and vegetables, the intercellular spaces rather than the wall pores
play the most important role. Intercellular air spaces are commonly found in parenchyma-
tous tissue and have been estimated to be 20–25% of the total volume in apples, 15% in
peaches, 37–45% in mushrooms, and 1% in potatoes. For instance, mature cells of apple
parenchyma tissues may be 50–500 µm in diameter with interconnecting air spaces ranging
from 210–350 µm across. These spaces are large enough that intercellular components can
pass through, permitting the free-bound aroma compounds to pass through spaces between
intercellular spaces of fruits and vegetables (Lapsley, Escher, and Hoehn, 1992; Ben-Arie,
Kislev, and Frenkel, 1979; Alzamora et al., 2017).
The physicochemical properties of the volatile compounds, when interacting with the
chemical nature and structure of the fruit and vegetable matrix, are powerful enough
to modify the concentration of the volatile compounds and therefore their perception
(Alzamora et al., 2017). A typical fruit or vegetable may have more than a few hundred
different volatile components, but in total, these comprise only a few parts per million of
the whole product (Husain, 2010). The buildup of the aroma compounds in fruits and veg-
etables is a complex, dynamic process during which the concentration of volatiles changes
both qualitatively and quantitatively. Moreover, the respiration rate for a climacteric fruit
is about twice the rate of a pre-climacteric fruit. The peak in apples, for instance, shows up
right after the fruit is ripe. Furthermore, commodities and/or cultivars with higher rates of
respiration (strawberry, blackberry, raspberry, cauliflower, lima bean, and avocado) tend
to have a shorter storage life than those with lower rates of respiration, such as apples,
citrus, grapes, and kiwifruits (Fagundes, Carciofi, and Monteiro, 2011). At any rate, all
postharvest operations and technological manufactures can affect the aroma of fruits and
vegetables, leading to the logical conclusion of reducing, as much as possible, the number
and the length of the postharvest operations and proceedings (Husain, 2010).

26.6 THE INTERACTION BETWEEN AROMA COMPOUNDS

AND FRUIT AND VEGETABLE MATRICES

Aroma–matrix interactions in fruits and vegetables influence aroma release and its per-
ception. The parameters that influence the release of aroma compounds can define and
provide useful information to control the aroma response of fruits and vegetables and
allow for the effective use of volatile compound materials (Schober and Peterson, 2004;
Paravisini and Guichard, 2017). Matrix components can bind, entrap, or encapsulate
volatile and nonvolatile compounds when the binding locations of matrix compounds
are still available. Consequently, the interactions decrease the rate of aroma release and
affect the aroma intensity and quality of fruits and vegetables. The mechanism of binding
between aroma compounds and matrices can be classified into three categories (Naknean
and Meenune, 2010; Shukla and Priyadashi, 2012):

1. Binding: The inclusion, adsorption, absorption, and retention of aroma com-

pounds onto nonvolatile substrates
2. Partitioning: The distribution of aroma compounds between phases such as the
oil, water, and gas phases
3. Release: The availability of aroma compounds from the bulk foods into the gas
phase for sensory perception
558 Food Aroma Evolution

The type of interaction depends on the physicochemical properties of aroma compounds

in the matrix. There are four main groups of aroma compound interactions in matrices
(Naknean and Meenune, 2010; Shukla and Priyadashi, 2012) which can be categorized
as follows:

1. Covalent bonding—The irreversible bonding such as the interaction between

aldehyde or ketone and the amino group of proteins
2. Hydrogen bonding—The occurrence between polar or volatile alcohol and het-
eroatom (N, S, O) of food components
3. Hydrophobic bonding—The weak and reversible bonding such as van der Waals
bond between nonpolar compounds and fat molecules
4. Physical binding—The occurrence between aroma compounds and starch or
starch derivatives, for example, the inclusion complexes

26.6.1 Interaction of Aroma Compounds and Carbohydrates

Carbohydrates can form inclusion complexes due to their helical structure, whereby
hydrophobic regions exist inside the polymer in which lipophilic aromas can be retained,
whether they are alcohols, aldehydes, acids, esters, terpenes, pyrazines, and other aroma
compounds. In dry systems, several aroma compounds were bound weakly to glucose,
saccharose, and lactose (McGorrin, 1996). Moreover, polysaccharides such as agar, algi-
nates, cellulose, guar gum, and methylcellulose, may bind at varying degrees of strength
to volatile aroma compounds (e.g., acetaldehyde, ethanol, diacetyl, ethyl acetate, and
2-hexanone) (McGorrin, 1996; Guichard, 2002)

26.6.2 Interaction of Aroma Compounds and Proteins

The effects of protein binding can be rather complicated since the binding of volatiles
is strongly dependent on the concentration and water content of the protein. Protein
also increases the binding capacity of aliphatics, aldehydes, and ketones, whereas the
alcohols decrease this. The binding of aroma substances to proteins depends on the
degree of denaturation, the temperature, and pH. However, aldehydes tend to react
chemically with protein amino groups, resulting in an irreversible binding (McGorrin,
1996). Free amino acids can bind with a series of volatile aroma compounds in aqueous
media. Ketones and alcohols are reversibly bound via hydrogen bonding to the amino or
carboxyl groups of amino acids, while as with proteins, some aldehydes react chemically
with amino groups to form Schiff bases. The reaction of aldehydes or ketones with cyste-
ine to form a thiazolidine is reversible with heating, particularly at low pH. (McGorrin,
1996; Guichard, 2002)

26.6.3 Interaction of Aroma Compounds and Lipids

The interactions of aromas with lipids are usually related to the relative amount of
aroma solubilized in the lipid and water phases. The main lipids in foods consist pre-
dominantly of di- and triglycerides. Interestingly, triglycerides can bind or solubilize sub-
stantial amounts of lipophilic and partlylipophilic aroma compounds. The quantity of
bound aroma compounds is also dependent on the fatty acid chain length and degree of
 Fruits and Vegetables 559

unsaturation in the triglyceride composition (McGorrin, 1996). In lipid–water mixtures

and emulsions, aroma compounds are distributed between the lipid and water phases as a
function of their structure, the temperature, and the type of lipid. The amount of aroma
bound by fat or oil is dependent on the chain length of the volatile compound within a
homologous series. This implies that the concentration in the gas phase reduces as the
chain length increases. Aroma–ingredient interactions can produce a different aroma
release behavior even with a low threshold (Guichard, 2002). Thus, the reduction of
the fat amount has the effect of raising the equilibrium vapor pressure of the aroma
compounds and changing its time-intensity release profile. Consequently, volatile aromas
cannot be retained in the matrix and are released immediately, resulting in a strong but
quickly dissipating aroma impression (McGorrin, 1996; Guichard, 2002)

26.6.4 Interaction of Aroma Compounds and Other Compounds

Aroma interactions have been described for individual classes of food components.
However, most foods are mixtures of several ingredient classes. Initial attempts have
been made to explore interactions with two or more components. If three components
are mixed, significantly more binding of the aroma is observed (McGorrin, 1996).
Primary evidence has been established for the potential use of a protein-stabilized W/
OAV emulsion for the retardation of aroma release (Naknean and Meenune, 2010). The
combination of protein or starch with monoglyceride emulsifiers was shown to bind
less to aroma than protein or starch in the absence of an emulsifier (McGorrin, 1996;
Guichard, 2002).

26.7 BINDING OF AROMA COMPOUNDS IN FRUITS AND VEGETABLES

Aroma compounds are bound in fruits and vegetables as glycosides. In this structure,
they are not volatile as such and, thus, they do not contribute to the aroma of fruits and
vegetables unless they are released from the glycoside (Hadi et al., 2013). The aroma-
active compound is released from the glycoside-binding side by maturation and ripening
(Lelel et al., 2003) but also during storage. From a technological point of view, the aroma
release from glycosides during processing (thermal and non-thermal) can exceed the num-
ber of free aroma compounds by a factor of up to 10:1 (Reineccius, 2006). Therefore, the
liberation of glycosidically bound aroma compounds can be challenging for the aroma
industry (Figure 26.3).
An example of aroma release is that of vanillin from the vanilla pod. The free vanil-
lin concentration in the green vanilla pod is very low, whereas the vanillin is present as
glucovanillin, the beta-D-glycoside of vanillin. The aglycone vanillin, as well as other
glycosidically bound aroma compounds, are released enzymatically upon ripening of the
vanilla pods (Figure 26.4) (Odoux et al., 2003; Perez Silva et al., 2011).

26.8 SENSORY PERCEPTION AND EVALUATION

OF FRUIT AND VEGETABLE AROMAS

Aroma and odor can be evaluated through instrumental methods and human evalua-
tion techniques. There are two subjective types of human evaluation on aroma, that is,
560 Food Aroma Evolution

FIGURE 26.3 Glycosidic bond formation of monosaccharide: (a) 1,4-glycosidic bond,

and (b) 1,6-glycosidic bond.

FIGURE 26.4 The enzymatic hydrolysis reaction scheme of vanillin creation. (Adapted
from Robert and Marjorie, 1977.)

difference test methods and consumer acceptance. The method of aroma or odor evalu-
ation depends on the product and its characteristics, the target market, and the aroma
or odor components of interest. Consumer acceptance tests are utilized to evaluate new
products, changes in manufacturing procedures, reformulations of existing products, or
for routine quality checking of the manufactured product vs. those of competitors. This
type of testing requires a large number of consumers representing a good cross-section
of the population. On the other hand, the difference method will use a small group of
individuals trained to act as an instrument in describing attributes of processed products.
Several numbers of product difference tests exist, including paired comparison, triangle,
dilution, ranking, numerical scoring, descriptive, and aroma or odor difference methods.
Sensory quality is a major criterion for making a purchasing decision by the consumer,
therefore, many factors, including variety, culture conditions, picking date, and posthar-
vest handling and storage methods should be able to inform the consumer of the quality
of the product, especially the factors that can affect flavor and aroma of fruits and veg-
etables (Kabir and Sindhu, 2012).

26.9 CONCLUSION

Fruits and vegetables play an important role in human nutrition and diet. Many com-
pounds are responsible for the aromas that have strong penetration odors with low
 Fruits and Vegetables 561

threshold values There are four significant types of volatile compounds which are con-
sidered as character impact compounds: terpene hydrocarbons, carbonyl components,
alcohols, esters, and volatile organic acids can be commonly found in fruit whereas sul-
fur-containing amino acids, fatty acids, glucosinolates, terpenoids, and phenolic com-
pounds are commonly found in vegetables. Moreover, the aroma compounds in fruits
and vegetables are found to be bound in matrices in the form of inclusion, adsorption,
and absorption. The binding of aroma and the matrices occurs by covalent bonding,
hydrogen bonding, hydrophobic bonding, and physical binding. Most of the aroma com-
pounds are bound to matrices components (carbohydrate, protein, and fat) by a glyco-
sidic bond. Consequently, the enzymatic and mechanic interactions can affect the release
of aroma. The physicochemical interactions that occur between aroma compounds
and other constituents of the matrices affect the retention of aroma compounds during
processing treatments. The identification of individual aroma compounds and matrix-
bounds is essential to inform and understand the phenomena involved in aroma release.
Therefore, the processing of fruits and vegetables should be operated appropriately to
cause the least fluctuation in the aroma release. Subsequently, many studies that char-
acterized aroma and flavor in fresh and processed fruits and vegetables have attempted
to assess the underlying aroma binding and release mechanisms. The understanding of
mechanisms and factors that affect aroma retention and aroma binding can lead to new
knowledge for the conservancy of aroma and flavor quality.

REFERENCES

Alzamora, S.M., A.B. Neito, M.A. Castro, P. Gómez, A.G. Loredo, J. Fava, and S. Vicente.
2017. Tissue structure and rheological and texture characteristics of minimally pro-
cessed fruits. In: Fresh-Cut Fruits and Vegetables: Technology, Physiology, and
Safety, ed. S. Pareek. Boca Raton, FL: Taylor & Francis Group.
Alzamora, S.M., D. Salvatori, M.S. Tapia, A. López-Malo, J. Welti-Chanes, and P. Fito.
2005. Novel functional food from vegetable matrices impregnated with biologically
active compounds. Journal of Food Engineering 67:205–14.
Asif, M. 2012. Physico-chemical properties and toxic effect of fruit-ripening agent cal-
cium carbide. Annual Tropical Medicine and Public Health 5:150–6.
Auldridge, M.E., A., Block, J.T., Vogel, C., Dabney-Smith, I., Mila, M., Bouzayen,
M., Magallanes-Lundback, D., DellaPenna, D.R., McCarty, and H.J. Klee. 2006.
Characterization of three members of the Arabidopsis carotenoid cleavage dioxy-
genase family demonstrates the divergent roles of this multifunctional enzyme fam-
ily. Plant Journal 45:982–93.
Baker, A., I.A. Graham, M. Hodsworth, and S.M. Smith. 2006. Chewing the fat:
β-Oxidation in signaling and development. Trends in Plant Science 11:124–32.
Barrett, D.M., J.C., Beaulieu, and R., Shewfelt. 2010. Color, flavor, texture, and nutri-
tional quality of fresh-cut fruits and vegetables: Desirable levels, instrumental and
sensory measurement, and the effects of processing. Critical Reviews in Food
Science and Nutrition 50(5):369–89.
Beaulieu, J.C. and E.A., Baldwin. 2002. Flavor and aroma of fresh-cut fruits and veg-
etables. In: Fresh-Cut Fruits and Vegetables: Science, Technology, and Market, ed.
O. Lamikanra. Boca Raton, FL: CRC Press.
Ben-Arie, R., N. Kislev, and C. Frenkel. 1979. Ultrastructural changes in the cell walls of
ripening apple and pear fruit. Plant Physiology 64:197–202.
562 Food Aroma Evolution

Berger, R.G. 2007. Flavours and Fragrances—Chemistry, Bioprocessing and

Sustainability. Berlin: Springer-Verlag.
Bood, K.G. and I. Zabetakis. 2002. The biosynthesis of strawberry flavor (II): Biosynthetic
and molecular biology studies. Journal of Food Science 67:2–8.
Boylston, T.D. 2010. Temperate fruit juice flavors: In Handbook of Fruits and Vegetables
Flavors, ed. Y.H., Hui. Hoboken, NJ: John Wiley & Sons.
Brandi, F., E., Bar, F., Mourgues, G., Horváth, E., Turcsi, G., Giuliano, A., Liverani, S.,
Tartarini, E., Lewinsohn, and C., Rosati. 2011. Study of “Redhaven” peach and
its white-fleshed mutant suggests a key role of CCD4 carotenoid dioxygenase in
carotenoid and norisoprenoid volatile metabolism. BMC Plant Biology 26:11–24.
Buescher, R.H. and R.W., Buescher. 2001. Effect of oxygen and free fatty acid on cucum-
ber flavour generation. Journal of Food Science 66:357–62.
Buttery, R.G. and G.R., Takeoka. 2013. Cooked carrot volatiles. AEDA and odor activ-
ity comparisons. identification of linden ether as an important aroma component.
Journal of Agricultural and Food Chemistry 61:9063–6.
Buttery, R.G., R.M., Seifert, D.G., Guadagni, and L.C., Ling. 1971. Characterization
of additional volatile components of tomato. Journal of Agricultural and Food
Chemistry 19:524–9.
Cadwallader, K.R., L.C., Tamamoto, and S.C., Sajuti. 2010. Aroma components of fresh
and stored pomegranate (Punica granatum L.) juice. ACS Symposium Series (ACS
Publication) 1036:93–101.
Chandrashekar, J., M.A., Hoon, N.J.P., Ryba, and C.S. Zuker. 2006. The receptors and
cells for mammalian taste. Nature 444(7117):761–9.
Cheetham, P.S.J. 2002. Plant-derived natural sources of flavor: In Food Flavour Technology,
ed. A.J., Taylor. Sheffield, UK: Sheffield Academic Press, pp. 105–52.
Cheong, K.W., C.P., Tan, H., Mirhosseini, N.S.A., Hamid, A., Osman, and M. Basri.
2010. Equilibrium headspace analysis of volatile flavour compounds extracted
from soursop (Anoan muricata) using solid phase microextraction. Food Research
International 43:1267–76.
Cheong, M.W., S.Q., Liu, J., Yeo, H.K., Chionh, K., Pramudya, P., Curran, and B., Yu.
2011. Identification of aroma-active compounds in Malaysian pomelo (Citrus gran-
dis (L.) Osbeck) peel by gas chromatography-olfactometry. Journal of Essential Oil
Research 23:34–42.
Collin, S., S., Nizet, T.B., Claeys, and P.-M., Despatures. 2011. Main odorants in Jura
florsherry wines. Relative contributions of sotolon, abhexon, and theaspirane-
derived compounds. Journal of Agricultural and Food Chemistry 60:380–7.
Devi, M.P., N., Bhowmick, and M.R., Bhanusree. 2015. Preparation of value-added
products through preservation: In Value Addition of Horticultural Crops: Recent
Trends and Future Directions, eds. A.B., Sharangi and S., Datta. Springer India.
Dixon, J. and E.W., Hewett. 2000. Factors affecting apple aroma/flavour volatile con-
centration: A review. New Zealand Journal of Crop and Horticultural Science
28:155–73.
Du, X., M., Song, and R., Rouseff. 2011. Identification of new strawberry sulfur vola-
tiles and changes during maturation. Journal of Agricultural and Food Chemistry
59:1293–300.
Dudareva, N., S., Andersson, I., Orlova, N., Gatto, M., Reichelt, and D., Rhodes. 2005.
The nonmevalonate pathway supports both monoterpene and sesquiterpene forma-
tion in snapdragon flowers. Proceeding of the National Academy of Science of the
United Stated of America 102:933–8.
 Fruits and Vegetables 563

Edris, A.E. and H.M., Fadel. 2002. Investigation of the volatile aroma components of
garlic leaves essential oil. Possibility of utilization to enrich garlic bulb oil. European
Food Research and Technology 214:105–7.
Elmore, A.T., J.S., Dodson, and D.S., Mottram. 2014. Reactions of propylene glycol with
the constituents of food flavourings: In Flavour Science, eds. V., Ferreira and R.,
Lopez. Amsterdam: Elsevier.
Elmore, J.S., D.S., Mottram, and M.M., Campos-Arribas. 2000. The reactions between
hydrogen sulfide and lipid-derived aldehydes in cooked meat. Czech Journal of Food
Science 18:8–9.
Engel, E., C., Baty, D., Le Corre, I., Souchon, and N., Martin. 2002. Flavor-active
compounds potentially implicated in cooked cauliflower acceptance. Journal of
Agricultural and Food Chemistry 50:6459–67.
Fagundes, C., B.A.M., Carciofi, and A.R., Monteiro. 2011. Estimate of respiration rate
and physicochemical changes of fresh-cut apples stored under different temperature.
Food Science and Technology 32(1):60–7.
Fellman, J.K., D.R., Rudell, T.S., Mattinson, and J.P., Mattheis. 2003. Relationship
of harvest maturity to flavor regeneration after CA storage of delicious apples.
Postharvest Biology and Technology 27:39–51.
Fischer, A., W., Grab, and P., Schieberle. 2008. Characterization of the most odor-active
compounds in a peel oil extract from Pontianak oranges (Citrus nobilis var. Lour.
Microcarpa Hassk.). European Food Research and Technology 227:735–44.
Forney, C.F. 2001. Horticultural and other factors affecting aroma volatile composition
of small fruit. HortTechnology 11:529–38.
Forney, C.F. 2008. Flavour loss during postharvest handling and marketing of fresh-cut
produce. Stewart Postharvest Review 4(3):5.1–5.10. doi: 10.2212/spr.2008.3.5.
Fuhrmann, E. and W., Grosch. 2002. Character impact odorants of the apple cultivars
elstar and cox orange. Nahrung/Food 46:187–93.
Gang, D.R. 2005. Evolution of flavors and scents. Annual Review of Plant Biology
56:301–25.
García, D., R., Zamora, J.M., Gómez, and J.A., Hódar. 1999. Bird rejection of unhealthy
fruits reinforces the mutualism between juniper and its avian dispersers. Oikos
85:536–44.
Ghawi, S.K., L., Methven, R.A., Rastall, and K., Niranjan. 2012. Thermal and high
hydrostatic pressure inactivation of myrosinase from green cabbage: A kinetic study.
Food Chemistry 131:1240–7.
Goff, S.A. and H.J. Klee. 2006. Plant volatile compounds: Sensory cues for health and
nutritional value. Science 311:815–9.
Goldberg, E.M., K., Wang, J., Goldberg, and M., Aliani. 2018. Factors affecting the
ortho- and retronasal perception of flavors: A review. Critical Reviews in Food
Science and Nutrition 58(6):913–23. doi: 10.1080/10408398.2016.1231167.
Gonzalez, M., C., Gaete-Eastman, M., Valdenegro, C.R., Figueroa, L., Fuentes, R.,
Herrera, and M.A., Moya-Leon. 2009. Aroma development during ripening of
fragaria chiloensis fruit and participation of an alcohol acyltransferase (FcAAT1)
gene. Journal of Agricultural and Food Chemistry 57:9123–32.
Gormly, T.R. 1981. Aroma in fruits and vegetables: In Quality in Stored and Processed
Vegetables and Fruit, eds. P.W., Goodenough and R.K., Atkin. New York, NY:
Academic Press.
Gould, W.A. 1983. Food Quality Assurance. Westport, CN: AVI Publishing Company,
pp. 314.
564 Food Aroma Evolution

Granvogl M., M., Christlbauer, and P., Schieberle. 2004. Quantitation of the intense
aroma compound 3-mercapto-2-methylpentan-1-ol in raw and processed onions
(Allium cepa) of different origins and in other allium varieties using a stable isotope
dilution assay. Journal of Agricultural and Food Chemistry 52:27–97.
Guichard, E. 2002. Interactions between flavor compounds and food ingredients and
their influence on flavor perception, Food Reviews International 18(1):49–70.
doi:10.1081/FRI-120003417.
Hadi, M.A.M., F.-J., Zhang, F.-F., Wu, C.-H., Zhou, and J. Tao. 2013. Advances in fruit
aroma volatile research. Molecules 18:8200–29.
Hayata, Y., C., Maneerat, H., Kozuka, K., Sakamoto, and Y., Ozajima. 2002. Flavor
volatile analysis of “House Momotaro” tomato fruit extracts at different ripen-
ing stages by Porapak Q column. Journal of Japan Society Horticultural Science
71:473–9.
Hewett, E.W. 2006. An overview of preharvest factors influencing postharvest quality
of horticultural products. International Journal of Postharvest Technology and
Innovation 1:4–15.
Husain, Q. 2010. Chemistry and biochemistry of some vegetable flavors: In Handbook
of Fruits and Vegetables Flavors, ed. Y.H., Hui. Hoboken, NJ: John Wiley & Sons.
Hongsoongnern, P. and E., Chambers, IV. 2007. A lexicon for texture and flavor
characteristics of fresh and processed tomatoes. Journal of Sensory Studies
23:583–99.
Jacobsson, A., T., Nielsen, and I., Sjoeholm. 2004. Influence of temperature, modi-
fied atmosphere packaging, and heat treatment on aroma compounds in broccoli.
Journal of Agricultural and Food Chemistry 52:1607–14.
Jayanty, S., J., Song, N., Rubinstein, A., Chong, and R.M., Beaudry. 2002. Temporal
relationship between ester biosynthesis and ripening events in bananas. Journal of
American Society for Horticultural Science 127:998–1005.
Ji, L. and G. Srzednicki. 2015. Extraction of aromatic compounds from banana peels.
Acta Horticulturae 1088:541–6. doi: 10.17660/ActaHortic.2015.1088.99.
Jiang, Y. and J. Song. 2010. Fruits and fruit flavor: Classification and biological charac-
terization: In Handbook of Fruits and Vegetables Flavors, ed. Y.H., Hui. Hoboken,
NJ: John Wiley & Sons.
Jirovetz, L., D., Smith, and G., Buchbauer. 2002. Aroma compound analysis of Eruca
sativa (Brassicaceae) SPME headspace leaf samples using GC, GC–MS, and olfac-
tometry. Journal of Agricultural and Food Chemistry 50:4643–6.
Jordon, R.B., R.J., Seelye, and V.A., McGlone. 2001. A sensory-based alternative to Brix/
acid ratio. Food Technology 55(6):35–44.
Jun, H.-R. and Y.-S., Kim. 2002. Comparison of volatile compounds in red pep-
per (Capsicum annuum L.) powders from different origins. Food Science and
Biotechnology 11:293–302.
Kabir, Y. and J.S., Sidhu. 2012. Flavor of fruit and fruit products and their sensory quali-
ties: In Handbook of Fruits and Fruit Processing, 2nd Edition, eds. K.S., Nirmal,
S.S., Jiwan, B., József, S.B.W., James, and M., Pilar Cano. John Wiley & Sons.
Kader, A.A. 2018. Perspective: Flavor quality of fruits and vegetables. Journal of the
Science of Food and Agriculture 88:1863–8.
Kader, A.A., L.L., Morris, M.A., Stevens, and M., Albright-Holton. 1978. Composition
and flavor quality of fresh market tomatoes as influenced by some postharvest han-
dling procedures. Journal of American Society for Horticultural Science 103:6–13.
 Fruits and Vegetables 565

Katsuno, T., H., Kasuga, Y., Kusano, Y., Yaguchi, M., Tomomura, J., Cui, Z., Yang,
S., Baldermann, Y., Nakamura, T., Ohnishi, N., Mase, and N., Watanabe. 2014.
Characterisation of odorant compounds and their biochemical formation in green
tea with a low temperature storage process. Food Chemistry 148:388–95.
Kubec, R., V., Drhova, and J., Velisek. 1998. Thermal degradation of S-methylcysteine
and its sulfoxide—Important flavor precursors of Brassica and Allium vegetables.
Journal of Agricultural and Food Chemistry 46:4334–40.
Lalel, H.J.D., Z., Singh, and S.C., Tan. 2003. Aroma volatiles production during fruit rip-
ening of “Kensington Pride” mango. Postharvest Biology and Technology 27:323–36.
Lapsley, K.G., F.E., Escher, and E. Hoehn. 1992. The cellular structure of selected apple
varieties. Food Structure 11:339–49.
Le Berre, E., T., Thomas-Danguin, N., Béno, G., Coureaud, P., Etiévant, and J., Prescott.
2008. Perceptual processing strategy and exposure influence the perception of odor
mixtures. Chemical sense 33(2):193–9. doi: 10.1093/chemse/bjm080.
Lignou, S., J.K., Parker, C., Baxter, and D.S., Mottram. 2014. Sensory and instrumen-
tal analysis of medium and long shelf-life Charentais cantaloupe melons (Cucumis
melo L.) harvested at different maturities. Food Chemistry 148:218–29.
Maarse, H. 1991. Volatile Compounds in Foods and Beverages, Vol. 21. New York, NY,
Basel, Hong Kong: Marcel Dekker, pp. 5678–91.
Maffei, M.E. 2016. Changes in biosynthesis of aroma volatile compounds during on-tree
maturation of “Pink Lady” apples. South African Journal of Botany 76:612–31.
Maneerat, C., Y., Hayata, H., Kozuka, K., Sakamoto, and Y., Osajima. 2002. Application
of the porapak q column extraction method for tomato flavor volatile analysis.
Journal of Agricultural and Food Chemistry 50:3401–4.
Mathieu, S., V.D., Cin, Z., Fei, H., Li, P., Bliss, M.G., Taylor, H.J., Klee, and D.M.,
Tieman. 2009. Flavour compounds in tomato fruits: Identification of loci and
potential pathways affecting volatile composition. Journal of Experimental Botany
60:325–37.
Mattes, R.D. 2009. Is there a fatty acid taste? Annual Review of Nutrition 29:305–27.
Mayer, F., G., Takeoka, R., Buttery, Y., Nam, M., Naim, Y., Bezman, and H.,
Rabinowitch. 2003. Aroma of fresh field tomatoes. ACS Symposium Series (ACS
Publication) 836:144–61.
McGorrin, R.J. 1996. Introduction: In Flavor-Food Interactions. ACS Symposium Series
633, eds. R.J., McGorrin and J.V., Leland. Washington, DC: American Chemical
Society.
Meilgaard, M., C.V., Civille, and T., Carr. 2007. Sensory Evaluation Techniques, 2nd
Edition. Boca Raton, FL: CRC Press.
Mottram, D.S., C.E., Mills, C., Wagstaff, and A.T., Dodson. 2011. Investigation of the
characteristic earthy flavor of baby corn (Zea Mays): In Advances and Challenges in
Flavor Chemistry & Biology, eds. T., Hofmann, W., Meyerhof, and P., Schieberle.
Freising, Germany: Deutsche Forschungsanstalt für Lebensmittelc.
Mukherjee, S. and P.K., Chattopadhyay. 2007. Whirling bed blanching of potato cubes
and its effects on product quality. Journal of Food Engineering 78:52–60.
Murray, K.E., P.A., Bannister, and R.G., Buttery. 1975. Geosmin: Important volatile
constituent of beetroot (Beta vulgaris). Chemistry & Industry (London) 22:973–4.
Naknean, P. and M. Meenune. 2010. Factor affecting retention and release of fla-
vour compounds in food carbohydrates. International Food Research Journal
17:23–34.
566 Food Aroma Evolution

Obenland, D., S., Collin, J. Sievert, F., Negm, and M.L. Arpaia. 2012. Influence of matu-
rity and ripening on aroma volatiles and flavor in “Hass” avocado. Postharvest
Biology and Technology 71:41–50.
Odoux, E., J. Escoute, J.L. Verdeil, and J.M., Brillouer. 2003. Localization of beta-D-
glycosidase activity and glucovanillin in vanilla bean (Vanilla planifolia Andrews).
Annals of Botany 92:437–44.
Oke, M. and G., Paliyath. 2010. Flavor from Transgenic Vegetables: In Handbook of
Fruits and Vegetables Flavors, ed. Y.H., Hui. Hoboken, NJ: John Wiley & Sons.
Paravisini, L. and E. Guichard. 2017. Interactions between aroma compounds and food
matrix: In Flavour: From Food to Perception, eds. E., Guichard, C., Salles, M.,
Mprzel, and A.-M., Le Bon. Hoboken, NJ: John Wiley & Sons.
Parker, J.K. 2015. Introduction to aroma compounds in foods: In Flavour Development,
Analysis and Perception in Food and Beverages, eds. J.K., Parker, S., Elmore, and
L., Methven. United Kingdom: Woodhead Publishing.
Pelayo-Zaldivar, C., S.E., Ebeler, and A.A., Kader. 2005. Cultivar and harvest date
effects on flavor and other quality attributes of California strawberries. Journal of
Food Quality 28:78–97.
Perez, A.G., R., Olias, P., Luaces, and C., Sanz. 2002. Biosynthesis of strawberry aroma
compounds through amino acid metabolism. Journal Agricultural Food Chemistry
50:4037–42.
Perez Silva, A., Z. Gunata, J.P. Lepoutre, and E. Odoux. 2011. New insight on the genesis
and fate of odor-active compounds in vanilla beans (Vanilla planifolia G. Jackson)
during traditional curing. Food Research International 44:2930–7.
Pino, J.A. and J., Mesa. 2006. Contribution of volatile compounds to mango (Mangifera
indica L.) aroma. Flavour and Fragrance Journal 21:207–13.
Qian, M. and G., Reineccius. 2003. Static headspace and aroma extract dilution analysis
of Parmigiano Reggiano cheese. Journal of Food Science 68:794–8.
Reineccius, G. 2006. Flavor Chemistry and Technology, 2nd Edition. Boca Raton, FL,
USA: CRC Press.
Rickman, J.C., D.M., Barrett, and Bruhn, C. 2007. Review Nutritional compari-
son of fresh, frozen and canned fruits and vegetables. Part 1. Vitamins C and
B and phenolic compounds. Journal of the Science of Food and Agriculture 87:
930–44.
Roberts, D.D. and T.E. Acree. 1996. Effects of heating and cream addition on fresh rasp-
berry aroma using a retronasal aroma simulator and gas chromatography olfactom-
etry. Journal of Agricultural and Food Chemistry 44:3919–25.
Robert, J.D. and C.C., Marjorie. 1977. Basic Principles of Organic Chemistry, 2nd
Edition. Menlo Park, CA: W. A. Benjamin, Inc.
Rodríguez, A., V., San Andrés, M., Cervera, A., Redondo, B., Alquézar, T., Shimada, …
L., Peña. 2011. The monoterpene limonene in orange peels attracts pests and micro-
organisms. Plant Signaling & Behavior 6(11):1820–3.
Rosati, C., G., Diretto, and G. Giuliano. 2009. Biosynthesis and engineering of carotenoids
and apocarotenoids in plants: State of the art and future prospects. Biotechnology
and Genetic Engineering Reviews 26:151–74.
Rosilllo, L., M.R., Salinas, J., Garijo, and G.L., Alonso. 1999. Study of volatiles in grapes
by dynamic headspace analysis Application to the differentiation of some Vitis vinif-
era varieties. Journal of Chromatography A 847:155–9.
 Fruits and Vegetables 567

Sanz, C., J.M., Olias, and A.G., Perez. 1997. Aroma biochemistry of fruits and veg-
etables: In Phytochemistry of Fruits and Vegetables. New York, NY, USA: Oxford
University Press Inc., pp. 125–55.
Schober, L.A. and G.D., Peterson. 2004. Flavor release and perception in hard candy:
Influence flavour compound-compound interactions. Journal of Agricultural and
Food Chemistry 52:2623–7.
Schwab, W., D.-R., Rachel, and L., Efraim. 2008. Biosynthesis of plant-derived flavor
compounds. The Plant Journal: For Cell and Molecular Biology 54(4):712–32.
Schwab, W. and P., Schreier. 2002. Enzymic formation of flavor volatiles from lipids:
In Lipid Biotechnology, eds. T.M., Kuo and H.W., Gardner. New York, NY, USA:
Marcel Dekker, pp. 293–18.
Shukla, A. and S.S., Priyadarshi. 2012. Factor affecting withholding and role of flavour
compounds in food vitamins. International Journal of Biology, Pharmacy, and
Allied Sciences 1(7):1012–9.
Siegmund, B. 2015. Biogenesis of aroma compounds: Flavour formation in fruits and veg-
etable. In Woodhead Publishing Series in Food Science, Technology and Nutrition,
Flavour Development, Analysis and Perception in Food and Beverages, eds. J.K.,
Parker, J.S., Elmore, and L., Methven. Cambridge, UK: Woodhead Publishing.
Sinuco, D.C., M., Steinhaus, P., Schieberle, and C., Osorio. 2010. Changes in odour-
active compounds of two varieties of Colombian guava (Psidium guajava L.) during
ripening. European Food Research and Technology 230:859–64.
Song, J. and C.F. Forney. 2008. Flavour volatile production and regulation in fruit.
Canadian Journal of Plant Science 88:537–50.
Song, J. and F., Bangerth. 2003. Fatty acids as precursors for aroma volatile biosynthesis
in pre-climacteric and climacteric apple fruit. Postharvest Biology and Technology
30:113–21.
Sucan, M.K. 2004. Identifying and preventing off-flavors. Food Technology 58(11):36–8.
Tamura, H., A., Fujita, M., Steinhaus, E., Takahisa, H., Watanabe, and P., Schieberle.
2011. Assessment of the aroma impact of major odor-active thiols in pan-roasted
white sesame seeds by calculation of odor activity values. Journal of Agricultural
and Food Chemistry 59:10211–8.
Thappa, R.K., A.K., Dhar, S.S., Balyan, S., Khan, P., Raina, P.L., Dhar, and D.K.,
Choudhary. 2003. Characterization of volatiles in rambutan fruit (Nephelium lap-
paceum L.). Journal of Food Science and Technology—Mysore 40:4264–431.
Thu, S.L., H.L.M., Tam, W. Tongdeesoontorn, and P. Suthiluk. 2017. Quality changes
and volatile compounds in fresh-cut “Phulae” pineapple during cold storage. Current
Applied Science and Technology 17(2):162–71.
Tokitomo, Y., M., Steinhaus, A., Büttner, and P., Schieberle. 2005. Odor—Active constit-
uents in fresh pineapple (Ananas comosus [L.] Merr.) by quantitative and sensory
evaluation. Bioscience, Biotechnology, and Biochemistry 69:1323–30.
Utama-ang, N., T., Cheewinworasak, N., Simawonthamgul, and R.S.
Samakradhamrongthai. 2018. Effect of drying condition of Thai garlic (Allium sati-
vum L.) on physicochemical and sensory properties. International Food Research
Journal 25(4):1365–72.
Vicente, A.R., C., Greve, and J.M. Labavitch. 2006. Recent findings in plant cell wall
structure and metabolism: Future challenges and potential implications for soften-
ing. Stewart Postharvest Review 2:1–8.
568 Food Aroma Evolution

Vicente, A.R., M., Saladie, J.K.C., Rose, and J.M., Labavitch. 2007. The linkage between
cell wall metabolism and fruit softening: Looking to the future. Journal of the
Science of Food and Agriculture 87:1435–48.
Wilson, D.A. and R.J. Stevenson. 2006. Learning to Smell: Olfactory Perception from
Neurobiology to Behavior. Baltimore, MD: Johns Hopkins University Press.
Yang, C., Y., Wang, Z., Liang, P., Fan, B., Wu, L., Yang, Y., Wang, and S., Li. 2009.
Volatiles of grape berries evaluated at the germplasm level by headspace-SPME with
GC–MS. Food Chemistry 114:1106–14.
Zhu, Y., H.J. Klee, and P.J. Sarnoski. 2018. Development and characterization of a high-
quality plum tomato essence. Food Chemistry 267:337–43.
 Chapter 27
Spices and Herbs
Alejandro Hernández, Emilio Aranda, Rocío
Casquete, Cristina Pereira, and Alberto Martín

CONTENTS

27.1 Introduction 569

27.2 Volatile Compounds in Spices and Herbs 571
27.2.1 Seeds, Nuts, Arils, and Kernels 571
27.2.2 Barks and Latex 575
27.2.3 Leaf Spices 576
27.2.4 Flower Buds and Stigma 578
27.2.5 Roots, Bulbs, Rhizomes, and Tubers 578
27.2.6 Fruits and Berries 580
27.3 Changes in the Composition of Volatiles Associated with Processing and
Commercial Presentation 583
References 584

27.1 INTRODUCTION

Spices and herbs comprise a diverse group of vegetal-origin products used as ingredients
for their flavoring, coloring, and preserving properties. The term “herb” refers to prod-
ucts from green parts of the plant, primarily the leaves. “Spice” refers to products from
other parts of plants, often dried, such as flowers, roots, bulbs, rhizomes, stems, barks,
and seeds.
Archaeological evidence shows that herbs and spices were used long before recorded
history to repel insects and mask human odors from animals during hunting, for example.
The culinary use of herbs and spices by humans occurred later, dating back 50,000 years
BC (https://www.astaspice.org/). In the Middle Ages, spices were some of the most valu-
able items of trade. The spice trade, which began in earnest in the Middle East around
3000 BC, stimulated the beginnings of economic globalization. Spices were highly desir-
able, yet expensive, so cheaper means of obtaining spices from the East were sought.
Navigations by Marco Polo, Vasco da Gama, and Christopher Columbus, for example,
who explored new trade routes, led to the Age of Exploration and discovery of the New
World. Over the following centuries, battles, conflicts, and wars broke out for control of
the spice trade (Parthasarathy et al., 2008). As new preservation techniques were devel-
oped, the value of spices decreased.
Spices and herbs have been historically appreciated for the aroma and flavor they
impart to food, preservative properties that allow extending the shelf life of food

569
570 Food Aroma Evolution

TABLE 27.1 World Production, Harvest, and Economic Data of the Main Spices
Item World Main Producer
Production Area Import
(×106 t) Harvested Value (USD Production
(×106 ha) Millions) Country (×106 t)
Star anise, 1.18 1.37 843.4 India 0.63
fennel,
coriander
Chilis and 3.92 1.80 1654.6 India 1.39
peppers, dry
Cinnamon 0.22 0.28 481.1 Indonesia 0.08
(canella)
Cloves 0.18 0.64 462.6 Indonesia 0.09
Garlic 26.57 1.47 3286.9 China 21.20
Ginger 3.27 0.41 758.4 India 1.11
Mustard seed 0.70 0.74 213.8 Canada 0.25
Nutmeg and 0.12 0.38 488.1 India 0.04
cardamoms
Pepper (Piper 0.55 0.53 2885.4 Vietnam 0.22
spp.)
Peppermint 0.11 <0.01 — Morocco 0.10
Sesame seed 6.11 10.58 2434.4 United 0.94
Republic of
Tanzania
Vanilla <0.01 0.09 816.5 Madagascar <0.01
Source: FAOSTAT, 2017.

commodities, and for their medicinal applications. These ingredients have also been
exploited to conceal food alteration and spoilage or the poor quality of raw materials.
The importance of spices and herbs across human history continues into the present.
Table 27.1 highlights the production data and economic values of the main spices and
herbs in 2017, provided by the database of the Food and Agriculture Organisation of the
United Nations (FAOSTAT) (http://www.fao.org/faostat/). Sesame (Sesamum indicum
L.) occupies the biggest area harvested among the spices (10.58 million hectares), fol-
lowed by chilies and peppers (1.87 million hectares). The most productive spice in 2017
was garlic (Allium sativum L.), with a yield of 26.57 million tons, despite a cultivation
area of only 1.47 million hectares. Spices represent a substantial economic value. In 2016,
the import values ranged from USD$462.6 million for cloves to USD$3286.9 million for
garlic. As can be observed from the data, the production of spices and herbs is polarized
toward the Asian continent, particularly, India, but also Indonesia and China.
Beyond their flavoring, coloring, and preserving properties, spices and herbs are valued
for their biological activities. Their antioxidant capability is one of the most recognized
benefits of using spices and herbs, which are rich in a wide diversity of phytochemicals,
including phenolic compounds, carotenoids, plant sterols, and glucosinolates (Embuscado,
2015; Srinivasan, 2014) that block the production of reactive oxygen species. For this
reason, spices and herbs or their derived compounds are employed to inhibit lipid per-
oxidation in different processed foods and as health-promoting substances, by defending
 Spices and Herbs 571

against oxidative stress and inflammatory processes (Kaefer and Milner, 2008; Rubió et
al., 2013). Besides their antioxidant efficacy, the phenolic compounds in spices and herbs
act as powerful antimicrobials (Shan et al., 2007; Weerakkody et al., 2010). Studies sug-
gest that Gram-positive bacteria are more sensitive than Gram-negative bacteria to phe-
nolic compounds from oregano (Origanum vulgare L.), cloves (Syzygium aromaticum or
Eugenia aromaticum or Eugenia caryophyllata Thunb.), and cinnamon (Cinnamomum
cassia Presl), among others (Shan et al., 2007). Owing to their multiple benefits, spices
and herbs are highly appreciated and are deeply studied. Nevertheless, their flavoring and
aromatic compounds are the main reason for their extended culinary use.
The next section approaches the key volatile compounds that characterize the fla-
vor and aroma of the main spices and herbs, which are grouped according to the parts
of the plant.

27.2 VOLATILE COMPOUNDS IN SPICES AND HERBS

Spices and herbs are frequently discussed regarding the parts of the plant used for flavor-
ing and colorant purposes. Below, the main volatile compounds of the most representa-
tive spices and herbs are detailed and summarized (Table 27.2), according to the part of
the plant used.

27.2.1 Seeds, Nuts, Arils, and Kernels

Vanilla (Vanilla planifolia) is derived from the orchids of the genus Vanilla, through a
traditional curing procedure of at least 6 months. During this process, the pods turn
black and biochemical and chemical transformations are carried out, mainly by native
β-glucosidase enzymes of the vanilla bean, hydrolyzing the glucosides and releasing the
aroma compounds. Vanillin (4-hydroxy-3-methoxybenzaldehyde) is the main constituent
(around 19,000 ppm) and the most characteristic flavor and aroma constituent found in
the cured vanilla bean, although unique notes in the vanilla aroma are provided by various
minor compounds in the cured pods, including p-hydroxybenzaldehyde, p-methoxybenz-
aldehyde (anisaldehyde), piperonal, p-hydroxybenzyl alcohol, vanillyl alcohol, and vanillic
acid (Pérez-Silva et al., 2006). Guaiacol, 4-methylguaiacol, acetovanillone, and p-cresol,
found at comparatively much lower concentrations in vanilla pods (2.6–13.7 ppm), also
have a considerable impact on the vanilla aroma (Silva et al., 2011). Anisaldehyde and guai-
acol are described as key components of the complex flavor of V. tahitensis (Brunschwig
et al., 2012) and have been correlated with positive and negative odors, respectively, in
extracts of V. planifolia (Hoffman et al., 2005), the main species harvested for vanilla.
Foeniculum vulgare Mill. (Apiaceae family) is commonly known as fennel, and its
aromatic seeds have been used as a culinary spice in many countries. Among more than
50 volatile compounds identified in fennel seed, the major constituents are limonene,
γ-terpinene, fenchone, estragole, trans-anethole, and 6,4-methoxy-benzaldehyde (Fang et
al., 2006). Estragole, trans-anethole, fenchone, and 1-octen-3-ol are the most intense odor
compounds detected in fennel fruit extracts. Estragole, trans-anethole, and p-anisalde-
hyde represent the characteristic anise, sweet, and licorice notes of this spice (Díaz-Maroto
et al., 2005). Other spices, such as anise, caraway, dill, and cumin, all also members of the
Apiaceae family, share the anise-like aroma of fennel. Although trans-anethole is the pre-
dominant component of the essential oil composition of anise fruits (Pimpinella anisum L.),
572 Food Aroma Evolution

TABLE 27.2 Summary of Main Volatile Compounds of Spices

Main Volatile
Part of Common Taxonomic Compound/s of
Plant Name Family Scientific Name Essential Oils Reference
Bean Vanilla Orchidaceae Vanilla Vanillin Pérez-Silva et
planifolia L. al. (2006)
Seed Fennel Apiaceae Foeniculum trans-Anethole, Díaz-Maroto
vulgare Mill. estragole et al. (2005)
Seed Dill seed Apiaceae Anethum D-carvone Blank and
graveolens L. Grosch
(1991)
Seed Caraway Apiaceae Carum carvi L. D-carvone Laribi et al.
(2010)
Seed Cumin Apiaceae Cuminum Cuminaldehyde Ravi et al.
cyminum L. (2013)
Seed Cardamom Zingiberaceae Elettaria α-Terpinyl acetate, Marongiu et
cardamomum L. 1,8 cineole al. (2004)
Seed Nutmeg Myristicaceae Myristica fragrans Sabinene Morsy (2016)
(aril) Houtt.
Seed Mustard Brassicaceae Brassica 2-Butyl- Sharma et al.
juncea L., isothiocyanate, (2018)
B. nigra L. phenyl ethyl
isothiocyanate,
3-isothiocyanato-1-
propene
Seed, Celery Apiaceae Anethum Phthalides Sellami et al.
leaf, graveolens var. (3-n-butylphthalide, (2012)
root dulce, A. sedanolide),
graveolens var. limonene
rapaceum, A.
graveolens var.
secalinum
Seed Coriander Umbelliferae Coriandrum Linalool Shahwar et al.
sativum L. (2012)
Leaf (E)-2-decenal Shahwar et al.
(2012)
Bark Cinnamon Lauraceae Cinnamomum sp. trans- Hammid et al.
Cinnamaldehyde (2016)
Latex Asafoetida Apiaceae Ferula Sulfur-containing Sahebkar and
asafoetida L. compounds Iranshahi
(2011)
Leaf Origanum Lamiaceae Origanum Thymol, carvacrol Bozin et al.
vulgare L. (2006),
Jerković et
al. (2001)
Leaf Parsley Apiaceae Petroselinum p-Mentha-1,3,8- Díaz-Maroto
crispum L. triene, myristicin et al. (2002)
(Continued )
 Spices and Herbs 573

TABLE 27.2 (Continued) Summary of Main Volatile Compounds of Spices

Main Volatile
Part of Common Taxonomic Compound/s of
Plant Name Family Scientific Name Essential Oils Reference
Leaf Bay leaf Lauraceae Laurus nobilis L. 1,8-Cineole Sellami et al.
(eucalyptol) (2011)
Leaf Basil Lamiaceae Ocimum Methyl chavicol Pirbalouti et
basilicum L. al. (2013)
Leaf Rosemary Lamiaceae Rosmarinus L. marinus Bozin et al.
officinalis L. officinalisl) (2007),
risticinmonenesothio Calín-
limonene Sánchez et al.
(2011)
Leaf Spearmint Lamiaceae Mentha spicata L. D-carvone Díaz-Maroto
et al. (2003)
Leaf Thyme Lamiaceae Thymus vulgaris Thymol Venskutonis
L. (1997)
Flower Cloves Myrtaceae Syzygium Eugenyl acetate Chaieb et al.
buds aromaticum L. (2007)
Flower Saffron Iridaceae Crocus sativus L. Safranal D’Archivio et
stigmas al. (2018)
Rhizome Ginger Zingiberaceae Zingiber α-Zingiberene, Huang et al.
officinale Roscoe 6-gingerol, 6-shogaol (2012)
Rhizome Turmeric Zingiberaceae Curcuma longa L. ar-Turmerone Khanam
(2018)
Bulb Garlic Alliaceae Allium sativum L. Allicin, (E)/(Z)-ajoene Martins et al.
(2016)
Bulb Onion Alliaceae Allium cepa L. Thiosulfinates, Choi et al.
thiosulfonates, (2017)
mono-, di- and
trisulfides
Root Horseradish Brassicaceae Armoracia Allyl isothiocyanate, Petrović et al.
rusticana L. 2-phenylethyl (2017)
isothiocyanate
Rhizome Wasabi Brassicaceae Wasabia japonica Allyl isothiocyanate Depree et al.
Matsum. (1998)
Fruit Black Piperaceae Piper nigrum L. α-Pinene, β-pinene, Kapoor et al.
pepper 3-carene, limonene, (2009)
caryophyllene
Fruit Paprika Solanaceae Capsicum Acetic acid Martín et al.
annuum L. (2017)
Fruit Chili pepper Solanaceae Capsicum α-Copaene, Patel et al.
annuum L., C. β-copaene, limonene, (2016)
baccatum L., C. o-cymene
chinense L., C.
frutescens L., C.
pubescens L.
(Continued )
574 Food Aroma Evolution

TABLE 27.2 (Continued) Summary of Main Volatile Compounds of Spices

Main Volatile
Part of Common Taxonomic Compound/s of
Plant Name Family Scientific Name Essential Oils Reference
Fruit Black velvet Fabaceae Dialium indum L. cis-Linalool oxide Pélissier et al.
tamarind (2001)
Fruit Star anise Magnoliaceae Illicium verum trans-Anethole Singh et al.
Hook. filius (2006)
Fruit Allspice Myrtaceae Pimenta Eugenol Tucker et al.
officinalis Lindl. (1991)

γ-himachalene, trans-pseudoisoeugenyl 2-methylbutyrate, p-anisaldehyde, and methyl

chavicol are also notable in the anise aroma profile, with differences according to the
country of origin (Orav et al., 2008). The compound trans-anethole has been described as
a good antibacterial agent against several foodborne pathogens (Diao et al., 2014).
In caraway (Carum carvi L.) and dill seeds (Anethum graveolens L.), the chemical
class characterization of their seed essential oil has evidenced the prevalence of D-carvone,
and its derivates trans-dihydrocarvone and cis-dihydrocarvone; besides other monoterpene
hydrocarbons, with limonene as the principal constituent (Blank and Grosch, 1991; Laribi
et al., 2010). Besides the use of D-carvone as a fragrance and flavor, this monoterpene is
attractive as a potato sprouting inhibitor, antimicrobial agent, and for its application in the
medical field (de Carvalho and da Fonseca, 2006). In the seeds of coriander (Coriandrum
sativum. L.), the main volatile compound of its essential oil is linalool (Gil et al., 2002;
Shahwar et al., 2012), which comprises more than half of the essential oil, although there
are remarkable differences among studies associated with the varieties, agronomic traits,
and region, for instance (Shahwar et al., 2012). Other compounds in coriander seed oil are
cis-dihydrocarvone, γ-terpinene, α-pinene, camphor, decanal, geranyl acetate, limonene,
geraniol, camphene, and D-limonene (Msaada et al., 2007). The essential oil and plant
extracts of this member of the Apiaceae family possess promising antibacterial, antifungal,
and antioxidative activities of various uses in foods (Mandal and Mandal, 2015).
Cumin (Cuminum cyminum L.) is a different aromatic plant included in the
Apiaceae family, and its seeds are used to flavor foods. The cumin odor is principally
due to the aldehydes, such as cuminaldehyde (p-isopropyl benzaldehyde, p-cuminalde-
hyde), p-menth-3-en-7-al, and p-menth-1,3-dien-7-al, which constitute the major volatile
compounds present in the volatile oil of cumin, together with monoterpenes, such as
β-pinene, p-cymene, γ-terpinene, and β-myrcene. The monoterpene profile of cumin is
associated with its regional location (Ravi et al., 2013).
Seed essential oils of various spices belonging to the family Zingiberaceae are well
known for their antifungal and antioxidant activities. Cardamom spice consists of the
whole or ground dried fruit of Elettaria cardamomum L. The volatile oil of cardamom
seeds obtained by both supercritical carbon dioxide extraction and solvent extraction,
respectively, are rich in α-terpinyl acetate, 1,8-cineole, and linalool (Marongiu et al.,
2004). Oxygenated monoterpenes, namely 1,8-cineole (syn. eucalyptol; has a fresh
mint-like smell) and α-terpineol, also constitute the major portion of the seed essential
oil of a large cardamom (Amomum subulatum Roxb.), alongside α- and β-pinene, and
allo-aromadendrene (Gurudutt et al., 1996). α-Pinene, β-pinene, sabinene, myrcene,
α-phellandrene, terpinene, terpinolene, linalool, linalyl acetate, terpinen-4-ol, γ-terpineol,
citronellol, geraniol, methyl eugenol, and trans-nerolidol, found at relatively much lower
 Spices and Herbs 575

concentrations in cardamom beans, also have a considerable impact on the cardamom

aroma (Baby and Ranganathan, 2016; Korikanthimath et al., 1997).
In the family Myristicaceae, nutmeg (Myristica fragrans Houtt.) and mace (aril of
the nutmeg) are among the most popular spices. The predominant volatile compounds of
nutmeg and mace oleoresins obtained by different extraction techniques are monoterpene
hydrocarbons (e.g., sabinene, α-pinene, β-pinene, and limonene), followed by aromatic
compounds, mainly myristicin, acids (myristic acid), monoterpene alcohols (terpinen-
4-ol), and esters (Abdurrasheed and Janardanan, 2009; Morsy, 2016). Myristicin, a
phenylpropane derivative, is the principal aromatic constituent of the volatile oil of both
nutmeg and mace, but it is also a psychotropic component of these spices, with a tradi-
tional use for several medical ailments (rheumatism, psychosis, digestive disorders, and
anxiety), in addition to its use as an aphrodisiac and abortifacient (Barceloux, 2008).
In vegetables of the mustard family (Brassicaceae), glucosinolates are broken down enzy-
matically by myrosinase, mainly into isothiocyanates, cyanides, and thiocyanates, many of
which are the primary aroma contributors. The most common types of mustard are Brassica
juncea L. (also known as Indian mustard or brown mustard) and Sinapis alba/Brassica alba
(also called yellow mustard, or white mustard). The major volatile constituents of Brassica
spp. seed extracts are sulfur compounds, products of glucosinolate degradation, which con-
tribute to intense seed-like, woody, and astringent sensory attributes. The main volatile com-
pounds of B. juncea L. seed extracts are 2-butyl isothiocyanate, phenyl ethyl isothiocyanate,
and 3-isothiocyanato-1-propene, which have a strong mustard aroma (Sharma et al., 2018).
In raw white mustard seeds, methanethiol, dimethyl trisulfide, 2-furanmethanethiol, 2-iso-
propyl-3-methoxypyrazine, 2-isobutyl-3-methoxypyrazine, 4-ethenyl-2-methoxyphenol,
phenylacetic acid, 2-aminoacetophenone, 1,8-cineole, and 4-hydroxy-3-methoxybenzalde-
hyde have been characterized as aroma-active compounds (Ortner et al., 2016).
The dry ripe seeds of celery (Apium spp.), belonging to the Apiaceae family, are light
brown to brown with a warm, bitter taste and characteristic aroma, and used as a spice
for flavoring various foods (Sowbhagya et al., 2007). Apium graveolens var. dulce (stalk
celery), A. graveolens var. rapaceum (celeriac or root celery) and A. graveolens var. seca-
linum Alef. (leaf celery) are some of the numerous varieties that are used in food (Roslon
et al., 2010). The distinctive aroma of celery is attributed to the presence of its phthalide-
rich essential oil, featuring mainly 3-n-butylphthalide and sedanolide (Sellami et al.,
2012). These phthalides (also sedanenolide) in conjunction with the monoterpenes (rep-
resented by limonene, α- and β-pinene, and myrcene), sesquiterpenes (mainly β-selinene)
are key contributors to the distinct celery flavor, although the amount of each of them
depends on the part of the plant, variety, and cultivated place. Although limonene is the
predominant constituent of the volatile oil of celery, it is the phthalide fraction, which
gives A. graveolens the characteristic spicy taste and aroma (Kurobayashi et al., 2006).
Also, due to its high nitrate content (around 3–4%), celery (also known as celery
powder) is used as a curing salt in meat products (Sebranek et al., 2012). Although the
consumption of celery is generally considered safe, the plant is a common cause of food
allergies (for instance, 30–40% of food allergic patients in Switzerland and France are
sensitized to celery [Kokotkiewicz and Łuczkiewicz, 2016]).

27.2.2 Barks and Latex

The bark of many trees has been used since ancient times as a spice, due to the aroma
provided by the presence of volatile compounds (Leela, 2008). Some species that emit
576 Food Aroma Evolution

these types of volatile compounds belong to the Lauraceae family, mainly of the genus
Cinnamomum, which have been studied extensively for their essential oil constituents
and are well known for their fragrance, medical value, and inhibitory properties against
microorganisms (molds, yeast, and bacteria) (Soliman and Badeaa, 2002). The bark of
several Cinnamomum species has been used mainly in cakes and confectionery, as well as
perfumery, but also for its health benefits (Hammid et al., 2016; Leela, 2008).
The functional classes of volatile compounds present in Cinnamomum species can
be broadly classified into monoterpenes, sesquiterpenes, and phenylpropenes (Hammid
et al., 2016; Leela, 2008; Wang et al., 2011). Some authors have identified a total of 57
kinds of volatile compounds in the bark of these species, including alcohols, aldehydes,
alkanes, alkenes, amines, carboxylic acids, ethers, esters, and ketones (Jayaprakasaha
et al., 2003; Wang et al., 2011). The main volatile compound imparting aroma to the
bark of the two species, Cinnamomum zeylanicum and C. cassia, is trans-cinnamalde-
hyde (Hammid et al., 2016; Leela, 2008; Ribeiro-Santos et al., 2017; Wang et al., 2011).
Cinnamomum zeylanicum also contains other key aroma compounds, such as camphor,
β-caryophyllene, p-cineole, camphene, α-pipene, hydrocinnamic aldehyde, α-terpineol,
terpinene 4-ol, linalool, and eugenol (Boughendjioua and Djeddi, 2018; Hammid et al.,
2016; Ribeiro-Santos et al., 2017). Cinnamomum cassia includes, among its major volatile
compounds, coumarin, benzaldehyde, cinnamyl acetate, β-caryophyllene, acetophenone,
methyl chavicol, 2,3-hexanediol, 1H-indole-3-acetamide, copaene, and glycerol-1-methyl
ether (Mehta and Kumar, 2017; Ribeiro-Santos et al., 2017; Wang et al., 2011).
Besides, the latex of some plant species also contains volatile compounds that
make them important spices. Many species of the Ferula genus, belonging to the family
Apiaceae, have historically been known as a rich source of aromatic resins and reputed
in traditional medicine as natural remedies, for a variety of disorders (Sahebkar and
Iranshahi, 2011). One of the characteristic species of this family is Ferula asafoetida,
which has a strong, tenacious, and sulfurous odor. It is an important spice used in Indian
food preparation. The essential oil of F. asafoetida is a complex mixture of compounds
with antimicrobial activity (Divya et al., 2014). Sulfur-containing compounds constitute
the dominant class of compounds in the oil, followed by monoterpene hydrocarbons,
sesquiterpene hydrocarbons, oxygenated sesquiterpenes, and oxygenated monoter-
penes, respectively (Sahebkar and Iranshahi, 2011). Several authors identified the major
volatile compounds of F. asafoetida as 1-propenyl sec-butyl disulfide, sec-butyl-(E,Z)-
1-propenyl disulfide, 1-(methylthio)propyl-(E,Z)-1-propenyl disulfide, germacrene B,
α-pinene, β-pinene, bis(1-methyl propenyl) disulfide, eudesmol (10-epi-γ), (E)-β-ocimene,
β-phellandrene, and limonene (Divya et al., 2014; Khajeh et al., 2005; Sahebkar and
Iranshahi, 2010; Takeoka, 2001).

27.2.3 Leaf Spices

Leaf spices are commonly termed herbs. Herbs comprise a limited number of botanical
families and genera, and both the fresh and dried leaves are used. Herbs are incorporated
to enhance the flavor of foods, such as salads, pasta, pizza, vegetables, meat, soups, and
seafood. Compounds from terpenoids and the monoterpenes family are significant and
most striking in the flavor of herbs, followed by sesquiterpenes.
The composition of the volatile fraction in herbs is influenced by cultural practices,
harvesting season, varieties, and the type of processing. The season of harvesting may
lead to serious fluctuations in the volatile components of herbs. As Jerković et al. (2001)
 Spices and Herbs 577

demonstrated, the yield of oregano (O. vulgare L. spp. hirtum; Lamiaceae) essential oil
ranged from 312.6 mg/100 g in April to 2331.0 mg/100 g in July, and many individual
volatile compounds were seasonally affected as well. The essential oil presents high con-
tents of oxygenated monoterpenoids, like thymol, carvacrol, p‐cymene, and γ‐terpinene.
Bozin et al. (2006) revealed carvacrol (61.3%), followed by thymol (13.9%) as the main
volatile compounds. Another herb rich in thymol is thyme (Thymus vulgaris L.), which,
in addition to thymol, has a predominance of oxygenated monoterpenoids, such as
α-terpineol, borneol, linalool, and 1,8-cineole (Lee et al., 2005; Venskutonis, 1997).
The varieties, cultivars or landraces (genotypes), significantly influence the compo-
sition of essential oils. Basil (Ocimum basilicum L.) is rich in methyl chavicol, as the
principal volatile compound. Interestingly, however, Pirbalouti et al. (2013) noted differ-
ences in the volatile compounds of two basil landraces, highlighting methyl chavicol, lin-
alool, and trans-ransrences in the volatile compounds of two basil landraces, highlighting
methyln landraces.
Coriander (C. sativum L.) has a double culinary function as a herb (leaves) and spice
(seed). Approximately 0.03–2.6% and 0.1–0.001% of the volatile compounds contained
in its essential oil are derived from its seeds and leaves, respectively (Shahwar et al.,
2012). (E)-2-Decenal is generally considered as the most abundant volatile compound
in the essential oil from the leaves (Shahwar et al., 2012); however, a recent analysis of
essential oils from mixed stem and leaves, conducted in Iran, found decanal, cis-phytol,
and 1-tetradecanol as more abundant than (E)-2-decenal (Pirbalouti et al., 2017). The
geographic origin and extraction methods are the possible reasons for this discrepancy.
The relative abundance of other compounds, for example, linalool, (E)-2-dodecenal,
(E)-2-tetradecenal, 2-decen-1-ol, (E)-2-undecenal, dodecanal, (E)-2-tridecenal, and
7-dodecenal, vary among studies (Macleod and Islam, 1976; Potter and Fagerson, 1990;
Shahwar et al., 2012).
Parsley (Petroselinum crispum L.) is another herb with extensive culinary uses, both
fresh and dried. Parsley seed is widely employed for essential oil extraction because it is
rich in volatile compounds, mainly apiole, β-phellandrene, p-mentha-1,3,8-triene, and
myristicin (Díaz-Maroto et al., 2002a), although the predominance changes depending
on the origin of the plant. Some major compounds, like p-mentha-1,3,8-triene and myris-
ticin, together with minor compounds, notably, 2-sec-butyl-3-methoxypyrazine, linalool,
(Z)-6-decenal, and (Z)-3-hexenal, are the main contributors to the characteristic aroma
of parsley (Masanetz and Grosch, 1998).
Among the volatile compounds that define the aroma of bay leaf (Laurus nobilis L.,
Lauraceae), 1,8-cineole (eucalyptol) is the most abundant one (comprises >50% of the
essential oil composition). Other relevant terpenes are linalool, methyl eugenol, terpinen-
4-ol, α-terpinyl acetate, β-pinene, and sabinene (Díaz-Maroto et al., 2002b; Sellami et al.,
2011). Minor benzene-derived compounds, like eugenol, methyl eugenol, and elemicin,
have been associated with the desirable spicy flavor of bay leaf (Pino et al., 1993).
Rosemary (Rosmarinus officinalis L.) shows a great heterogeneity in essential oil
composition. The volatile compounds mainly consist of monoterpenoids and monoter-
penes (89.9–98.0%). Among them, according to Calín-Sánchez et al. (2011), α-pinene is a
major component (19.8%) for Polish samples. Other studies found camphor and 1,8-cin-
eole as the principal constituents of essential oils of Spanish rosemary (Díaz-Maroto
et al. 2007), or limonene (22%) and camphor (22%) in Serbian samples (Bozin et al.
2007). These studies agreed that genotypes and geographical origin influence the bio-
logical activities and volatile compositions. Other volatile compounds found in relevant
concentrations are borneol, verbenone, and camphene.
578 Food Aroma Evolution

The essential oil of spearmint (Mentha spicata L.) is especially rich in oxygenated
monoterpenes. Carvone is the major component and the most important in the spearmint
flavor. Limonene and 1,8-cineole also predominate (Díaz-Maroto et al., 2003).

27.2.4 Flower Buds and Stigma

Cloves are the dried flower buds of S. aromaticum L., a tree of the family Myrtaceae. The
clove is its dried, unopened flower buds. Clove is used extensively for flavoring all kinds
of food products, such as meats, sausages, baked goods, confectionery, candies, table
sauces, pickles, and rice dishes, and is used extensively in curry powders and masalas.
Eugenol is usually considered as the major volatile component of cloves, although other
components, present in lesser amounts, such as benzyl alcohol, also play an important
role. Chaieb et al. (2007) identified 36 components in clove buds. High concentrations of
eugenyl acetate, β-caryophyllene, 2-heptanone, ethyl hexanoate, humulenol, α-humulene,
calacorene, calamenene, α- and β-humulene, benzaldehyde, β-ylangene, and chavicol have
been described (Lee and Shibamoto, 2001; Leela and Sapna, 2008). The minor constitu-
ents, like methyl amyl ketone and methyl salicylate, are responsible for the characteristic
pleasant odor of cloves (Leela and Sapna, 2008).
Saffron is a spice derived from the three dry stigmas of the pistil of the flower of
Crocus sativus L. (Iridaceae). It is considered the most expensive spice and is appreciated
worldwide for its unique aroma, taste, color, and medicinal properties. Saffron is used
almost exclusively for cooking purposes, to give color and flavor to food products. The
typical saffron flavor is developed during the drying process by the enzymatic and ther-
mal degradation of picrocrocin (Lage and Cantrell, 2009). Although aldehydes, alcohols,
acids, phenol, ketones, and furanone are present, safranal (2,6,​6 -tri​methy​l-1,3​- cycl​ohexa​
diene​-1-ca​rboxa​ldehy​de) is the major volatile compound responsible for the saffron aroma
(Amanpour et al., 2015; Lage and Cantrell, 2009; Urbani et al., 2015). Safranal and these
other hydrolysis products may undergo further degradation to yield additional volatile
compounds (D’Archivio et al., 2018). Along with safranal, other main compounds are
3,5,5-trimethyl-2-cyclohexen-1-one (isophorone), 2,6,6-trimethyl-2-cyclohexene-1,4-di-
one (4-ketoisophorone), and 2,2,6-trimethyl-1,4-cyclohexanedione. Among additional
compounds important in imparting the saffron flavor are 3,5,5-trimethyl-3-cyclohexen-
1-one, 2,6,6​-trim​ethyl​-1,4-​cyclo​hexad​iene-​1-car​boxal​dehyd​e, 4-hyd​roxy-​2 ,6,6​-trim​
ethyl​-1-cy​clohe​xen-1​- carb​oxald​ehyde​, and 2-hyd​roxy-​4,4,6​-trim​ethyl​-2,5-​cyclo​hexad​
ien-1​-one​(Amanpour et al., 2015; Maggi et al., 2009; Urbani et al., 2015).

27.2.5 Roots, Bulbs, Rhizomes, and Tubers

Some of the main root-, bulb-, rhizome-, and tuber-derived spices are described in this
section. Among them, ginger (Zingiber officinale Roscoe), a rhizome of the Zingiberaceae
family, is one of the most ancient spices in the world. It has been widely used as a spice or
a common condiment for a variety of food and beverages, conferring a specific flavor and
odor, and, like other spices, has recognized therapeutic effects against different diseases
(Oriani et al., 2018). Ginger and ginger oleoresin are used in the food industry because
both contain volatile compounds. The most pungent are the gingerols and shogaols, espe-
cially 6-gingerol and 6-shogaol, and volatile constituents (sesquiterpenes and monoter-
penes). The main volatile compounds in ginger oleoresin are the sesquiterpenes, notably,
 Spices and Herbs 579

α-zingiberene (26–37%), β-sesquiphellandrene (10–13%), β-phellandrene (7.4–13%),

α-curcumene (14%), and geranial (6.6–8.1%) (Huang et al., 2012; Oriani et al., 2018).
Depending on the type of dehydration or conservation technique, changes in these types
of compounds decrease its quality. For example, 6-gingerol, the most abundant com-
pound in ginger essential oil, can be converted into 6-shogaol (An et al., 2016; Huang et
al., 2012).
Turmeric (Curcuma longa L.) is a root of the Zingiberaceae family. Turmeric powder
(present in curry powder) or turmeric volatile oil is used as a spice and food colorant
and has recognized benefits in treating diabetes, human immunodeficiency virus, and
tumors, for example (Park et al., 2012). Turmeric oil consists of different sesquiterpenes
and curcuminoids. The main volatile compounds in turmeric oil are ar-turmerone (31–
67.1%), β-turmerone (2.0–37.9%), α-turmerone (0.0–21.3%), caryophyllene (0.0–6.8%),
β-cedrene (2.3–5.3%), and β-curcumene (0.0–6.1%), among others. These compounds
can be affected by the method used to extract the essential oil (Chang et al., 2006;
Khanam, 2018). In addition to its flavor and colorant uses, curcumin (yellow colorant
of turmeric) is notable for its potent antioxidant, antimutagenic, anti-inflammatory, and
antitumor properties (Srinivasan, 2017).
Garlic (A. sativum L.) is a bulb of the Alliaceae family. It has been cultivated since
ancient times and used as a spice for culinary purposes, owing to its potent flavor attri-
butes, being consumed either as a raw vegetable (fresh leaves or dried cloves) or after
processing in the form of oil. Apart from its characteristics as a spice, many cultures
have exploited the important therapeutic properties of garlic (Rivlin, 2001; Srinivasan,
2017). Organosulfur compounds, especially allicin, (E)/(Z)-ajoene, 2-vinyl-4H-1,3-
dithiin, diallyl sulfide, diallyl disulfide, and diallyl trisulfide are highly dominant,
but the concentrations are affected by pre-harvest (genotype and various cultivation
practices) and postharvest conditions (storage conditions and processing treatments)
(Martins et al., 2016). When garlic tissue is disrupted, the enzyme alliinase rapidly
lyses the cytosolic alkenyl-cysteine sulfoxides (mainly alliin) to form thiosulfinates
(mostly allicin), which are the most abundant constituents in fresh garlic. In lyophilized
garlic, allicin formation starts when water is added (de Diego et al., 2007). This com-
pound is chemically unstable, thermolabile and it can be transformed upon different
culinary processes, producing diverse organosulfur compound profiles, predominated
by ajoenes and vinyldithiins (Block, 2010; Locatelli et al., 2015). All these compounds
are responsible for the characteristic pungency of garlic (Locatelli et al., 2017). Torres-
Palazzolo (2018) showed that all organosulfur compounds were bioaccessible in raw
and cooked garlic.
Onion (Allium cepa) is a bulb belonging to the Alliaceae family, used in cooking and
as a food ingredient. Common to all Allium species (e.g., garlic), is the enzyme alliinase
that catalyzes the hydrolysis of S-alk(en)yl-L-cysteine sulfoxides to produce pyruvate,
ammonia, and sulfur-containing volatiles, including mono-, di-, and trisulfides (Choi et
al., 2017; Colina-Coca et al., 2013). The dominant flavor compounds are generated by
the spontaneous reactions undergone by the sulfenic acids among themselves and with
other compounds. The result is a mixture of sulfur-containing compounds, including
thiosulfinates, thiosulfonates, and mono-, di-, and trisulfides. Among the main sulfides
in onions, dimethyl trisulfide, propenyl propyl disulfide, dipropyl disulfide, propenyl
methyl disulfide, methyl propyl trisulfide, and dipropyl trisulfides can be mentioned.
Thiopropanal sulfoxide, the lachrymatory or tear factor, is formed from the S-trans-
prop-1-enyl cysteine sulfoxide (isoalliin) precursor, but quickly degrades into other com-
pounds, such as propanal, because of its instability.
580 Food Aroma Evolution

Colina-Coca et al. (2013) isolated 18 compounds in the volatile fraction of onion,

belonging mainly to di- and trisulfides, and aldehydes. However, the dehydration pro-
cess can drastically affect some volatile components. For example, dipropyl disulfide
accounted for 77.7% of the total volatile compounds in fresh onion, but this was dra-
matically reduced by drying (Choi et al., 2017).
Horseradish (Armoracia rusticana L.) is a root belonging to the Brassicaceae fam-
ily. The plants of this genus contain glucosinolates. Owing to their intense pungency and
piquant flavor, horseradish roots are used as a condiment for a range of foods (e.g., meat,
pickled vegetables, bakery products). The main volatile constituents of horseradish roots
and responsible for its sharp aroma and flavor, are the sulfur-containing substances,
isothiocyanates and glucosinolates, mostly sinigrin, followed by gluconasturtiin, in the
raw material, transforming into allyl isothiocyanate and 2-phenylethyl isothiocyanate,
respectively, after grinding (Petrović et al., 2017). Some other secondary metabolites
identified in the roots of horseradish are flavonoids (kaempferol derivatives) and caffeic
acid (Herz et al., 2017).
In comparison, the other Armoracia species have distinctly smaller roots and differ-
ent volatiles profile. Armoracia macrocarpa root volatile fractions were dominated by
methyl thioalkyl isothiocyanates, mostly berteroin (5-methylthiopentyl isothiocyanate;
55.0–59.0%) and lesquerellin (6-methylthiohexyl isothiocyanate; 34.1–36.4%) (Petrović
et al., 2017). These results were different from those in the tested A. rusticana. Although
2-phenylethyl isothiocyanate, the second most abundant volatile of A. rusticana, was
present in the A. macrocarpa volatile fraction, only low quantities were found (3.0–
3.5%). These differences in the volatile profiles are reflected in the different odor and
taste. Armoracia macrocarpa is less pungent when compared with A. rusticana because
it has lower concentrations of berteroin and lesquerellin (Uchida et al., 2012).
Wasabi (Wasabia japonica Matsum.) or Japanese horseradish belongs to the
Brassicaceae family. Although the rhizome is the most valuable part of the plant, other
parts have some of the pungent taste. The paste of its root, liberating allyl isothiocyanate
by hydrolysis with myrosinase, is used in the preparation of sauces and condiments. The
flavor of wasabi is similar to horseradish root (Depree et al., 1998).
The pungent taste of wasabi and other cruciferous plants derives from the degrada-
tion products of glucosinolates. Myrosinase converts the glucosinolates to glucose and
an unstable aglycone that is transformed into an isothiocyanate, specifically 2-propenyl
isothiocyanate, commonly referred to as allyl isothiocyanate. These volatile elements are
rapidly evaporated in the air, so wasabi paste must be prepared fresh for each meal.
The distinct wasabi flavor is provided by allyl isothiocyanate, together with three other
compounds, namely, 6-methylthiohexyl isothiocyanate (fresh wasabi flavor), 7-methy-
thioheptyl isothiocyanate (sweetish wasabi flavor) and 8-methylthiooctyl isothiocyanate
(weakly pungent wasabi flavor) (Depree et al., 1998).
The methyl sulfinylalkyl compounds are found in both wasabi and horseradish,
whereas the methyl thioalkyl compounds are found in wasabi alone (Depree et al., 1998).

27.2.6 Fruits and Berries

There are about 30 types of fruit that are commonly used as spices and for flavoring
purposes throughout the world (Wilson, 2016). One of these is black pepper (Piper
nigrum), which consists of the dry and immature fruit of the Piperaceae family. The
characteristic aroma and flavor of black pepper are contributed by the pepper essential
 Spices and Herbs 581

oil components, mainly, terpene hydrocarbons (89%), oxygenated terpenes, and aromatic
compounds (Kapoor et al., 2009). The main monoterpene hydrocarbons present in black
pepper oil are α- and β-pinenes, 3-carene, sabinene, and limonene while the dominant
sesquiterpene is caryophyllene (Jelén and Gracka, 2015; Kapoor et al., 2009; Menon and
Padmakumari, 2005). The compounds β-phellandrene, β-bisabolene, eugenol, terpinen-
4-ol, hedycaryol, β-eudesmol, α-humulene, zingiberene, elemicin, bulnesol, cubenol, and
caryophyllene oxide, found in relatively lower concentrations in black pepper beans, are
also considered as main odourants in black pepper (Jelén and Gracka, 2015; Kapoor et
al., 2009). Oils rich in caryophyllene possess sweet floral odors, whereas oils with high
pinene content give turpentine as an unpleasant odor.
Capsicum is composed of fruits of the genus Capsicum L. (Solanaceae). The spices of
these fruits are used commonly in the forms of fresh and dried products, and their odor
is characteristic, sternutatory but agreeable, and mildly to intensely pungent (Wilson,
2016). The volatiles profile of Capsicum is mainly affected by the variety (Pino et al.,
2011), ripening stage (Pino et al., 2006), and processing method (Cremer and Eichner,
2000; Mateo et al., 1997). Acids, aldehydes, alcohols, esters, ketones, and pyridines are
the main chemical classes comprising the volatiles in fresh peppers (Capsicum annuum
L.). Most of the acids found in peppers originate from the degradation of lipids, and these
compounds can significantly contribute to the odor (Jun and Kim, 2002). Tissue dis-
ruption increases the amount of volatile unsaturated C6 aldehydes and alcohols, which
might play an important role in determining the green bell and hot pepper flavors. Along
with 2-isobutyl-3-methoxypyrazine, other alkyl-methoxypyrazines are representative
compounds of the genus Capsicum. These volatile compounds, in addition to hexanal,
hexanol, cis‑2-hexanal, and cis‑2-hexenol, are the main compounds found in fresh pepper
(C. annuum L.) (Pino et al., 2006) and are responsible for the freshly cut grass or ground
green leaf odor. Kim et al. (2007) reported that the main compounds in fresh peppers
(C. annuum L.) were 3-hydroxypyridine, acetic acid, 2-furanmethanol, butyrolactone,
and 2-propanone. In contrast, Habanero chili pepper is rich in (E)-2-hexenal, hexyl-
3-methylbutanoate, (Z)-3-hexenyl 3-methylbutanoate, hexyl pentanoate, 3,3-dimethyl-
cyclohexanol, and hexadecanoic acid (Pino et al., 2006). However, drying bell peppers
causes quality changes associated with a decrease in compounds with “fresh” odor notes
and an increase in products coming from auto-oxidation of unsaturated fatty acids and
Strecker degradation, such as acetaldehyde, 2-methylpropanal, 2-methylbutanal, and
3-methylbutanal (Ríos et al., 2008).
Paprika comes from milling the dry fruits of different varieties of C. annuum L.
Three main types of paprika are marketed in Spain and differentiated by their particular
drying processes (smoked, sun-dried, and oven-dried) and the pepper variety employed.
These aspects affect the profile of the volatiles (Velázquez et al., 2014; Ziino et al., 2009).
Smoked paprika is richer in alcohols, phenols, pyrroles, and pyranones, whereas, the
oven-dried samples are characterized by their aldehydes and terpenes. Sun-dried paprika
has significantly lower amounts of odorant substances than their smoked and oven-dried
counterparts (Martín et al., 2017). Acetic acid, 3- and 2,3-butanediol, dihydro-2(3H)-
furanone, dihydro-5-methyl-2(3H)-furanone, 2-propanone, and 3-hydroxy-2-butanone
are the main volatile compounds found in all three types of paprika. However, com-
pounds, such as phenol, 2-methylphenol (o-cresol), 2-methoxyphenol (guaiacol), and
2,6-dimethoxyphenol (syringol) are present in substantial amounts in smoked paprika,
less abundant in sun-dried samples and absent from the oven-dried samples (Martín et
al., 2017). These compounds are associated with the thermal degradation of lignin and
are the main chemicals responsible for the smoky flavor (Guillén and Manzanos, 2002).
582 Food Aroma Evolution

Different to the smoked and oven-dried samples, pyridines, alcohols, and sulfur com-
pounds are exclusive to sun-dried paprika and mostly associated with thermal decompo-
sition of the amino acids (Maga, 1981), whereas, heptanal, 5-hexen-2-ol, 3-octen-1-ol,
and terpenes, such as L-β-pinene and trans‑caryophyllene, only exist in oven-dried
paprika. These volatile compounds can originate from the lipid metabolism of the pepper
or lipid oxidation.
The chili pepper (Capsicum) genus is categorized into 38 species, of which 5 are
domesticated; C. annuum (Eggink et al., 2012; Mazida et al., 2005), C. frutescens and
C. chinense (Rodríguez-Burruezo et al., 2010), as well as C. baccatum and C. pubescens
(Kollmannsberger et al., 2011). The others are classified as semi-domesticated or wild
(United States Department of Agriculture [USDA], 2016). One major reason for the inter-
est in their cultivation is due to their aroma, which is derived from a complex chemical
composition of volatiles that can present considerable modifications, according to the
variety being studied, the location where it is cultivated, the processing procedure, and
the degree of maturation. Terpenes, esters, and hydrocarbons are the dominant aromatic
compounds and are represented in all chili pepper types, whereas ketones are detected in
low concentrations in only some chili pepper samples (Garruti et al., 2013; Junior et al.,
2012; Kollmannsberger et al., 2011; Patel et al., 2016). α-Copaene, β-copaene, limonene,
o-cymene, n-hexyl hexanoate, 2-methyl tridecanoate, pentadecane, n-tetradecane, 2-hex-
enal, and 2-nonanone are the most abundant compounds found in chilies. However,
β-pinene, α-selinene, β-cadinene, allo-aromadendrene, and α-phellandrene are restricted
to C. baccatum and C. chinense (Patel et al., 2016).
Black velvet tamarind is a common name for several trees in the genus Dialium indum
(Fabaceae). The characteristic aroma of their fruit can be used to define the maturity stage
or distinguish different fruit types (Ong et al., 2008). Alcohols, alkanes, acids, and alde-
hydes are responsible for their aroma (Pélissier et al., 2001). cis-Linalool oxide (furanoid)
is the primary compound in black velvet tamarind species, but other major volatile con-
stituents of the tamarind aroma are 4-hydroxy-2,5-dimethyl-3(2H)-furanone, limonene,
geranyl acetone, (Z)-3-hexenal, linalool, nonanal, cinnamyl acetate, α-pinene, 3,5-d​ihydr​
oxy-6​-meth​yl-2,​3-dih​ydro-​4H-py​ran-4​-one,​ and (E)-2-dodecenal. Linalool, limonene,
4-hydroxy-2,5-dimethyl-3(2H)-furanone, nonanal, and (Z)-3-hexenal are described as
the compounds that show significant differences in the odor profiles between different
tamarinds (D. guineense, D. dinklagei, and D. pachyphyllum) (Lasekan and See, 2015).
Star anise is the ripe dried fruit of Illicium verum Hook. filius (Magnoliaceae). lllicium
verum is a rich source of lignans and seco-prezizaane-type sesquiterpenes (Lee et al., 2003;
Song et al., 2007), which occur exclusively in Illicium species (Chang et al., 2010). The
main component in the essential oil from star anise fruit is trans-anethole (1-methoxy-
4-(1-propenyl) benzene), which accounts for 85–90% of the total components (Singh et
al., 2006). The other constituents include terpenes, pinenes, β-phellandrene, α-diterpene,
limonene, estragole, safrol, and terpineol, among others (Tuan and Ilangantileket, 1997).
The closely related Japanese star anise (I. anisatum) is highly toxic due to the presence
of anistan, a poisonous sesquiterpene lactone. It also contains shikimin and sikimitoxin,
both responsible for severe inflammation of several bodily organs (Wichtl, 2004), and
safrole and eugenol, two compounds that do not exist in l. verum and are used to identify
its adulteration (Howes et al., 2009).
Allspice consists of the dried fruits of Pimenta officinalis Lindl. (Myrtaceae). Allspice
is aromatic and pungent and possesses the flavor and aroma of clove, nutmeg, cinnamon,
and black pepper. In allspice fruits, eugenol is the main constituent. Eugenol methyl ether
is found in minor amounts, and terpenes, such as myrcene, 1,8-cineol, and α-phellandrene,
 Spices and Herbs 583

exist in trace amounts. Limonene, terpinolene, β-caryophyllene, and β-selinene have been
identified in the essential oil of the berries (Tucker et al., 1991).

27.3 CHANGES IN THE COMPOSITION OF VOLATILES ASSOCIATED

WITH PROCESSING AND COMMERCIAL PRESENTATION

The impact of drying on the volatile compounds in herbs and spices depends mainly on
the drying parameters (Figure 27.1) but, also, the main constituents of the aromatic pro-
file of the plants and the initial moisture content. In general, for herbs, such as parsley,
coriander, thyme, and sage, air drying at ambient temperature results in few losses in vol-
atile compounds when compared with the fresh products (Díaz-Maroto et al., 2002a,b;
Venskutonis, 1997). Proportionately, under conditions of natural drying, greater losses of
the non-oxygenated monoterpene hydrocarbons rather than of the oxygenated constitu-
ents of these herbs occur. It is due to the differences in the volatility and the oxidation
of their precursors (Venskutonis, 1997). However, a drastic reduction in the quantity
of volatiles is found in herb extracts dried at high temperatures (45–60°C) as is shown

FIGURE 27.1 Percentage of variation on total volatile compounds (in light gray) or main
volatile compounds (in dark gray) of dried herbs influenced by drying method. Extraction
methods: SDE (solvent distillation extraction); HS (headspace); HD (hydro distillation);
MTR (microwave technique resin). (1) Venskutonis, 1997; (2) Díaz-Maroto et al., 2002b;
(3) Huopalahti et al., 1985; (4) Antal et al., 2011; (5) Figiel et al., 2010; (6) Calín-Sánchez
et al., 2012; (7) Calín-Sánchez et al., 2012; (8) Di Cesare et al., 2003; (9) Rao et al., 1998;
(10) Parmar et al., 2018; (11) Yousif et al., 2000.
584 Food Aroma Evolution

the Figure 27.1. Under these conditions, the severe loss of primary aroma compounds
involved in the “fresh” odor note is replaced by secondary aroma compounds related
to lipid autoxidation, products derived from the carbohydrate metabolism and Strecker
degradation (Huopalahti et al., 1985). In dry fruits, such as paprika, these secondary
aroma compounds constitute a relevant fraction of their flavor (Cremer and Eichner,
2000). During lyophilization, the vacuum conditions can cause substantial losses of rele-
vant aroma compounds, whereas, at high pressure, fewer losses of these compounds, and
even increases in the concentration of certain components have been described (Antal et
al., 2011; Díaz-Maroto et al., 2002a,b). Another alternative for spice drying is vacuum-
microwave. This energy combined with lowered pressures induces faster evaporation of
water from the vegetal material being efficiently dried. The vacuum-microwave system is
used for drying herbs such as oregano, rosemary, and sweet basil (Calín-Sánchez et al.,
2012; Szumny et al., 2010; Figiel et al., 2010).
Comminution is another operation applied to spices. In normal grinding, the product
quality, flavor, and aroma, is severely affected, with losses of about 30% of the volatile
oil due to an increase in the exposition surface and the high temperatures present dur-
ing this process. This fact can be partially palliated by strategies such as circulating cold
air or water around the grinder. The use of liquid nitrogen for the cryogenic grinding
of spices, such as coriander seed, assures any loss of volatiles is significantly reduced
(Balasubramanian et al., 2012; Saxena et al., 2015).

REFERENCES

Abdurrasheed, K. M., and C. Janardanan. 2009. “Chemical composition of nutmeg and

mace (Myristica fragrans Houtt.) from Tellicherry and Kannur regions, Kerala.”
Journal of Spices and Aromatic Crops, 18 (2): 108–10.
Amanpour, A., A. S. Sonmezdag, H. Kelebek, and S. Selli. 2015. “GC–MS–olfactometric
characterization of the most aroma-active components in a representative aromatic
extract from Iranian saffron (Crocus sativus L.).” Food Chemistry, 182: 251–6.
An, K., D. Zhao, Z. Wang, J. Wu, Y. Xu, and G. Xiao. 2016. “Comparison of dif-
ferent drying methods on Chinese ginger (Zingiber officinale Roscoe): Changes
in volatiles, chemical profile, antioxidant properties, and microstructure.” Food
Chemistry, 197B: 1292–300.
Antal, T., A. Figiel, B. Kerekes, and L. Sikolya. 2011. “Effect of drying methods on
the quality of the essential oil of spearmint leaves (Mentha spicata L.).” Drying
Technology, 29 (15): 1836–44.
Baby, K. C. and T. V. Ranganathan. 2016. “Effect of enzyme pre-treatment on extrac-
tion yield and quality of cardamom (Elettaria cardamomum Maton) volatile oil.”
Industrial Crops and Products, 89: 200–6.
Balasubramanian, S., M. K. Gupta, and K. K. Singh. 2012. “Cryogenics and its applica-
tion with reference to spice grinding: A review.” Critical Reviews in Food Science
and Nutrition, 52 (9): 781–94.
Barceloux, D. G. 2008. Medical Toxicology of Natural Substances: Foods, Fungi,
Medicinal Herbs, Plants, and Venomous Animals. Hoboken, NJ, USA: John Wiley
& Sons Inc.
Blank, I. and W. Grosch. 1991. “Evaluation of potent odorants in dill seed and dill
herb (Anethum graveolens L.) by aroma extract dilution analysis.” Journal of Food
Science, 56 (1): 63–7.
 Spices and Herbs 585

Block, E. 2010. Garlic and Other Alliums: The Lore and The Science. New York, NY,
USA: RSC Publishing.
Boughendjioua, H. and S. Djeddi. 2018. “Study of the organoleptic and physicochemi-
cal properties of cinnamon essential oil (Cinnamomum zeylanicum).” American
Journal of Life Science Researches, 6 (3): 123–30.
Bozin, B., N. Mimica-Dukic, I. Samojlik, and E. Jovin. 2007. “Antimicrobial and anti-
oxidant properties of rosemary and sage (Rosmarinus officinalis L. and Salvia offi-
cinalis L., Lamiaceae) essential oils.” Journal of Agricultural and Food Chemistry,
55 (19): 7879–85.
Bozin, B., N. Mimica-Dukic, N. Simin, and G. Anackov. 2006. “Characterization of the
volatile composition of essential oils of some Lamiaceae spices and the antimicro-
bial and antioxidant activities of the entire oils.” Journal of Agricultural and Food
Chemistry, 54 (5): 1822–8.
Brunschwig, C., P. Senger-Emonnot, M. L. Aubanel, A. Pierrat, G. George, S. Rochard,
and P. Raharivelomanana. 2012. “Odor-active compounds of Tahitian vanilla fla-
vor.” Food Research International, 46 (1): 148–57.
Calín-Sánchez, Á., A. Szumny, A. Figiel, K. Jałoszyński, M. Adamski, and Á. A.
Carbonell-Barrachina. 2011. “Effects of vacuum level and microwave power on
rosemary volatile composition during vacuum–microwave drying.” Journal of Food
Engineering, 103 (2): 219–27.
Calín-Sánchez, Á., K. Lech, A. Szumny, A. Figiel, and Á. A. Carbonell-Barrachina. 2012.
“Volatile composition of sweet basil essential oil (Ocimum basilicum L.) as affected
by drying method.” Food Research International, 48 (1): 217–25.
Chaieb, K., H. Hajlaoui, T. Zmantar, A. B. Kahla‐Nakbi, M. Rouabhia, K. Mahdouani,
and A. Bakhrouf. 2007. “The chemical composition and biological activity of clove
essential oil, Eugenia caryophyllata (Syzigium aromaticum L. Myrtaceae): A short
review.” Phytotherapy Research, 21 (6): 501–6.
Chang, L. H., T. T. Jong, H. S. Huang, Y. F. Nien, and C. M. J. Chang. 2006. “Supercritical
carbon dioxide extraction of turmeric oil from Curcuma longa Linn. and purifica-
tion of turmerones.” Separation and Purification Technology, 47 (3): 119–25.
Chang, J. Y., M. H. Abd El-Razek, Y. H. Chen, Y. B. Cheng, S. Y. Chen, C. T. Chien, Y.
H. Kuo, and Y. C. Shen. 2010. “Phytoquinoids and secoprezizaane-type sesquiter-
penes from Illicium arborescens.” Helvetica Chimica Acta, 93 (1): 123–32.
Choi, S. M., D. J. Lee, J. Y. Kim, and S. T. Lim. 2017. “Volatile composition and sensory
characteristics of onion powders prepared by convective drying.” Food Chemistry,
231: 386–92.
Colina-Coca, C., D. González-Peña, E. Vega, B. de Ancos, and C. Sánchez-Moreno. 2013.
“Novel approach for the determination of volatile compounds in processed onion
by headspace gas chromatography–mass spectrometry (HS GC–MS).” Talanta, 103:
137–44.
Cremer, D. R. and K. Eichner. 2000. “Formation of volatile compounds during heating
of spice paprika (Capsicum annuum) powder.” Journal of Agricultural and Food
Chemistry, 48 (6): 2454–60.
D’Archivio, A. A., L. Di Pietro, M. A. Maggi, and L. Rossi. 2018. “Optimization using
chemometrics of HS-SPME/GC–MS profiling of saffron aroma and identification
of geographical volatile markers.” European Food Research and Technology, 244
(9): 1605–13.
de Carvalho, C. C. R. and M. M. R. da Fonseca. 2006. “Carvone: Why and how should
one bother to produce this terpene.” Food Chemistry, 95 (3): 413–22.
586 Food Aroma Evolution

de Diego, M., M. Avello, S. Mennickent, M. Fernández, and P. Fernández. 2007.

“Validated liquid chromatographic method for quantitative determination of allicin
in garlic powder and tablets.” Journal of Separation Science, 30 (16): 2703–7.
Depree, J. A., T. M. Howard, and G. P. Savage. 1998. “Flavour and pharmaceutical
properties of the volatile sulphur compounds of wasabi (Wasabia japonica).” Food
Research International, 31 (5): 329–37.
Diao, W. R., Q. P. Hu, H. Zhang, and J. G. Xu. 2014. “Chemical composition, anti-
bacterial activity and mechanism of action of essential oil from seeds of fennel
(Foeniculum vulgare Mill.).” Food Control, 35 (1): 109–16.
Díaz-Maroto, M. C., I. J. Díaz-Maroto Hidalgo, E. Sánchez-Palomo, and M. S. Pérez-
Coello. 2005. “Volatile components and key odorants of fennel (Foeniculum vul-
gare Mill.) and thyme (Thymus vulgaris L.) oil extracts obtained by simultaneous
distillation–extraction and supercritical fluid extraction.” Journal of Agricultural
and Food Chemistry, 53 (13): 5385–9.
Díaz‐Maroto, M. C., M. S. Pérez‐Coello, E. Sánchez‐Palomo, and M. A. González Viñas.
2007. “Impact of drying and storage time on sensory characteristics of rosemary
(Rosmarinus officinalis L.).” Journal of Sensory Studies, 22 (1): 34–48.
Díaz-Maroto, M. C., M. S. Pérez-Coello, M. A. González Viñas, and M. D. Cabezudo.
2003. “Influence of drying on the flavor quality of spearmint (Mentha spicata L.).”
Journal of Agricultural and Food Chemistry, 51 (5): 1265–9.
Díaz-Maroto, M., M. Pérez-Coello, and M. Cabezudo. 2002a. “Effect of different dry-
ing methods on the volatile components of parsley (Petroselinum crispum L.).”
European Food Research and Technology, 215 (3): 227–30.
Díaz-Maroto, M. C., M. S. Pérez-Coello, and M. D. Cabezudo. 2002b. “Effect of drying
method on the volatiles in bay leaf (Laurus nobilis L.).” Journal of Agricultural and
Food Chemistry, 50 (16): 4520–4.
Di Cesare, L. F., E. Forni, D. Viscardi, and R. C. Nani. 2003. “Changes in the chemical
composition of basil caused by different drying procedures.” Journal of Agricultural
and Food Chemistry, 51 (12): 3575–81.
Divya, K., K. Ramalakshmi, P. S. Murthy, and L. J. M. Rao. 2014. “Volatile oils from
Ferula asafoetida varieties and their antimicrobial activity.” LWT-Food Science and
Technology, 59 (2): 774–9.
Eggink, P. M., C. Maliepaard, Y. Tikunov, J. P. W. Haanstra, A. G. Bovy, and R. G. F.
Visser. 2012. “A taste of sweet pepper: Volatile and non-volatile chemical composi-
tion of fresh sweet pepper (Capsicum annuum) in relation to sensory evaluation of
taste.” Food Chemistry, 132 (1): 301–10.
Embuscado, M. E. 2015. “Spices and herbs: Natural sources of antioxidants—A mini
review.” Journal of Functional Foods, 18: 811–9.
Fang, L., M. Qi, T. Li, Q. Shao, and R. Fu. 2006. “Headspace solvent microextraction-
gas chromatography–mass spectrometry for the analysis of volatile compounds from
Foeniculum vulgare Mill.” Journal of Pharmaceutical and Biomedical Analysis, 41
(3): 791–7.
FAOSTAT. 2017. http://www.fao.org/faostat/en/#home.
Figiel, A., A. Szumny, A. Gutiérrez-Ortíz, and Á. A. Carbonell-Barrachina. 2010.
“Composition of oregano essential oil (Origanum vulgare) as affected by drying
method.” Journal of Food Engineering, 98 (2), 240–7.
Garruti, D. S., N. O. F. Pinto, V. C. C. Alves, M. F. A. da Penha, E. C. Tobaruela,
and I. M. S. Araújo. 2013. “Volatile profile and sensory quality of new varieties of
Capsicum chinense pepper.” Ciência e Tecnologia de Alimentos, 33 (S1): 102–8.
 Spices and Herbs 587

Gil, A., E. B. de la Fuente, A. E. Lenardis, M. López Pereira, S. A. Suárez, A. Bandoni, C.

van Baren, P. D. L. Lira, and C. M. Ghersa. 2002. “Coriander essential oil composi-
tion from two genotypes grown in different environmental conditions.” Journal of
Agricultural and Food Chemistry, 50 (10): 2870–7.
Guillén, M. D. and M. J. Manzanos. 2002. “Study of the volatile composition of an aque-
ous oak smoke preparation.” Food Chemistry, 79 (3): 283–92.
Gurudutt, K. N., J. P. Naik, P. Srinivas, and B. Ravindranath. 1996. “Volatile constit-
uents of large cardamom (Amomum subulatum Roxb.).” Flavour and Fragrance
Journal, 11 (1): 7–9.
Hammid, S. A., Z. Assim, and F. Ahmad. 2016. “Chemical composition of Cinnamomum
species collected in Sarawak.” Sains Malaysiana, 45 (4): 627–32.
Herz, C., H. T. T. Tran, M. R. Márton, R. Maul, S. Baldermann, M. Schreiner, and E.
Lamy. 2017. “Evaluation of an aqueous extract from horseradish root (Armoracia
rusticana Radix) against lipopolysaccharide-induced cellular inflammation
reaction.” Evidence-Based Complementary and Alternative Medicine, 2017:
1950692.
Hoffman, P., A. Harmon, P. Ford, M. Zapf, A. Weber, S. King, R. Grypa, E. Philander,
L. González, and K. Lentz. 2005. “Analytical approaches to vanilla quality and
authentication.” In Vanilla: Proceedings of the First International Congress, pp.
41–9. Carol Stream, IL, USA: Allured Publishing.
Howes, M. J. R., G. C. Kite, and M. S. J. Simmonds. 2009. “Distinguishing Chinese star
anise from Japanese star anise using thermal desorption–gas chromatography–mass
spectrometry.” Journal of Agricultural and Food Chemistry, 57 (13): 5783–9.
Huang, B., G. Wang, Z. Chu, and L. Qin. 2012.” Effect of oven drying, microwave
drying, and silica gel drying methods on the volatile components of ginger
(Zingiber officinale Roscoe) by HS-SPME-GC-MS.” Drying Technology, 30 (3):
248–55.
Huopalahti, R., E. Kesälahti, and R. Linko. 1985. “Effect of hot air and freeze drying
on the volatile compounds of dill (Anethum graveolens L.) herb.” Agricultural and
Food Science, 57 (2): 133–8.
Jayaprakasha, G. K., L. J. M. Rao, and K. K. Sakariah. 2003. “Volatile constituents from
Cinnamomum zeylanicum fruit stalks and their antioxidant activities.” Journal of
Agricultural and Food Chemistry, 51 (15): 4344–8.
Jelén, H. H. and A. Gracka. 2015. “Analysis of black pepper volatiles by solid phase
microextraction–gas chromatography: A comparison of terpenes profiles with
hydrodistillation.” Journal of Chromatography A, 1418: 200–9.
Jerković, I., J. Mastelić, and M. Miloš. 2001. “The impact of both the season of col-
lection and drying on the volatile constituents of Origanum vulgare L. ssp. hirtum
grown wild in Croatia.” International Journal of Food Science and Technology, 36
(6): 649–54.
Jun, H. R. and Y. S. Kim. 2002. “Comparison of volatile compounds in red pep-
per (Capsicum annuum L.) powders from different origins.” Food Science and
Biotechnology, 11 (3): 293–302.
Junior, S. B., A. M. Tavares, J. T. Filho, C. A. Zini, and H. T. Godoy. 2012. “Analysis of
the volatile compounds of Brazilian chilli peppers (Capsicum spp.) at two stages of
maturity by solid phase micro-extraction and gas chromatography–mass spectrom-
etry.” Food Research International, 48 (1): 98–107.
Kaefer, C. M. and J. A. Milner. 2008. “The role of herbs and spices in cancer preven-
tion.” Journal of Nutritional Biochemistry, 19 (6): 347–61.
588 Food Aroma Evolution

Kapoor, I. P. S., B. Singh, G. Singh, C. S. De Heluani, M. P. De Lampasona, and C. A. N.

Catalan. 2009. “Chemistry and in vitro antioxidant activity of volatile oil and oleo-
resins of black pepper (Piper nigrum).” Journal of Agricultural Food Chemistry, 57
(12): 5358–64.
Khajeh, M., Y. Yamini, N. Bahramifar, F. Sefidkon, and M. Reza Pirmoradei. 2005.
“Comparison of essential oils compositions of Ferula assa-foetida obtained by
supercritical carbon dioxide extraction and hydrodistillation methods.” Food
Chemistry, 91 (4): 639–44.
Khanam, S. 2018. “Influence of operating parameters on supercritical fluid extraction
of essential oil from turmeric root.” Journal of Cleaner Production, 188: 816–24.
Kim, I. K., A. M. Abd El-Aty, H. C. Shin, H. B. Lee, I. S. Kim, and J. H. Shim. 2007.
“Analysis of volatile compounds in fresh healthy and diseased peppers (Capsicum
annuum L.) using solvent free solid injection coupled with gas chromatography–
flame ionization detector and confirmation with mass spectrometry.” Journal of
Pharmaceutical and Biomedical Analysis, 45 (3): 487–94.
Kokotkiewicz, A. and M. Łuczkiewicz. 2016. “Celery (Apium graveolens var. dulce
(Mill.) Pers.) oils.” In Essential Oils in Food Preservation, Flavor and Safety, edited
by V. E. Preedy, pp. 325–38. London, UK: Academic Press.
Kollmannsberger, H., A. Rodríguez-Burruezo, S. Nitz, and F. Nuez. 2011. “Volatile and
capsaicinoid composition of ají (Capsicum baccatum) and rocoto (Capsicum pube-
scens), two Andean species of chile peppers.” Journal of the Science of Food and
Agriculture, 91 (9): 1598–611.
Korikanthimath, V. S., R. Mulge, and T. J. Zachariah. 1997. “Variation in yield and
quality characters of cardamom clones.” Journal of Medicinal and Aromatic Plant
Sciences, 19 (4): 1024–7.
Kurobayashi, Y., E. Kouno, A. Fujita, Y. Morimitsu, and K. Kubota. 2006. “Potent odor-
ants characterize the aroma quality of leaves and stalks in raw and boiled celery.”
Bioscience, Biotechnology, and Biochemistry, 70 (4): 958–65.
Lage, M. and C. L. Cantrell. 2009. “Quantification of saffron (Crocus sativus L.) metabolites
crocins, picrocrocin and safranal for quality determination of the spice grown under dif-
ferent environmental Moroccan conditions.” Scientia Horticulturae, 121 (3): 366–73.
Laribi, B., K. Kouki, A. Mougou, and B. Marzouk. 2010. “Fatty acid and essential oil
composition of three Tunisian caraway (Carum carvi L.) seed ecotypes.” Journal of
the Science of Food and Agriculture, 90 (3): 391–6.
Lasekan, O. and N. S. See. 2015. “Key volatile aroma compounds of three black velvet
tamarind (Dialium) fruit species.” Food Chemistry, 168: 561–5.
Lee, K. G. and T. Shibamoto. 2001. “Antioxidant property of aroma extract isolated
from clove buds [Syzygium aromaticum (L.) Merr. et Perry].” Food Chemistry, 74
(4): 443–8.
Lee, S. J., K. Umano, T. Shibamoto, and K. G. Lee. 2005. “Identification of volatile com-
ponents in basil (Ocimum basilicum L.) and thyme leaves (Thymus vulgaris L.) and
their antioxidant properties.” Food Chemistry, 91 (1): 131–7.
Lee, S. W., G. Li, K. S. Lee, D. K. Song, and J. K. Son. 2003. “A new phenylpropanoid
glucoside from the fruits of Illicium verum.” Archives of Pharmacal Research, 26
(8): 591–3.
Leela, N. K. 2008. “Cinnamon and cassia.” In Chemistry of Spices, edited by V. A.
Parthasarathy, B. Chempakam, and T. J. Zachariah pp. 124–41. Oxford, UK: CAB
International.
 Spices and Herbs 589

Leela, N. K. and V. P. Sapna. 2008. “Clove.” In Chemistry of Spices, edited by V. A.

Parthasarathy, B. Chempakam, and T. J. Zachariah, pp. 147–64. Oxford, UK: CAB
International.
Locatelli, D. A., J. C. Altamirano, R. E. González, and A. B. Camargo. 2015. “Home-
cooked garlic remains a healthy food.” Journal of Functional Foods, 16: 1–8.
Locatelli, D. A., M. A. Nazareno, C. M. Fusari, and A. B. Camargo. 2017. “Cooked
garlic and antioxidant activity: Correlation with organosulfur compound composi-
tion.” Food Chemistry, 220: 219–24.
Macleod, A. J. and R. Islam. 1976. “Volatile flavour components of coriander leaf.”
Journal of the Science of Food and Agriculture, 27 (8): 721–5.
Maga, J. A. 1981. “Pyrroles in foods.” Journal of Agricultural and Food Chemistry, 29
(4): 691–4.
Maggi, L., M. Carmona, C. P. del Campo, C. D. Kanakis, E. Anastasaki, P. A. Tarantilis,
M. G. Polissiou, and G. L. Alonso. 2009. “Worldwide market screening of saf-
fron volatile composition.” Journal of the Science of Food and Agriculture, 89 (11):
1950–4.
Mandal, S. and M. Mandal. 2015. “Coriander (Coriandrum sativum L.) essential oil:
Chemistry and biological activity.” Asian Pacific Journal of Tropical Biomedicine,
5 (6): 421–8.
Marongiu, B., A. Piras, and S. Porcedda. 2004. “Comparative analysis of the oil and
supercritical CO2 extract of Elettaria cardamomum (L.) Maton.” Journal of
Agricultural and Food Chemistry, 52 (20): 6278–82.
Martín, A., A. Hernández, E. Aranda, R. Casquete, R. Velázquez, T. Bartolomé, and R.
Córdoba. 2017. “Impact of volatile composition on the sensorial attributes of dried
paprika.” Food Research International, 100: 691–7.
Martins, N., S. Petropoulos, and I. C. F. R. Ferreira. 2016. “Chemical composition and
bioactive compounds of garlic (Allium sativum L.) as affected by pre- and post-
harvest conditions: A review.” Food Chemistry, 211: 41–50.
Masanetz, C. and W. Grosch. 1998. “Key odorants of parsley leaves (Petroselinum
crispum [Mill.] Nym. ssp. crispum) by odour-activity values.” Flavour and Fragrance
Journal, 13 (2): 115–24.
Mateo, J., M. Aguirrezábal, C. Domínguez, and J. M. Zumalacárregui. 1997. “Volatile
compounds in Spanish paprika.” Journal of Food Composition and Analysis, 10
(3): 225–32.
Mazida, M. M., M. M. Salleh, and H. Osman. 2005. “Analysis of volatile aroma com-
pounds of fresh chilli (Capsicum annuum) during stages of maturity using solid
phase microextraction (SPME).” Journal of Food Composition and Analysis, 18
(5): 427–37.
Mehta, M. J. and A. Kumar. 2017. “Green and efficient processing of Cinnamomum
cassia bark by using ionic liquids: Extraction of essential oil and construction
of UV‐resistant composite films from residual biomass.” Chemistry—An Asian
Journal, 12 (24): 3150–5.
Menon, A. N. and K. P. Padmakumari. 2005. “Essential oil composition of four major
cultivars of black pepper (Piper nigrum L.)—IV.” Journal of Essential Oil Research,
17 (2): 206–8.
Morsy, N. F. S. 2016. “A comparative study of nutmeg (Myristica fragrans Houtt.) oleo-
resins obtained by conventional and green extraction techniques.” Journal of Food
Science and Technology, 53 (10): 3770–7.
590 Food Aroma Evolution

Msaada, K., K. Hosni, M. B. Taarit, T. Chahed, M. E. Kchouk, and B. Marzouk. 2007.

“Changes on essential oil composition of coriander (Coriandrum sativum L.) fruits
during three stages of maturity.” Food Chemistry, 102 (4): 1131–4.
Ong, B. T., S. A. H. Nazimah, C. P. Tan, H. Mirhosseini, A. Osman, D. Mat Hashim,
and G. Rusul. 2008. “Analysis of volatile compounds in five jackfruit (Artocarpus
heterophyllum L.) cultivars using solid-phase microextraction (SPME) and gas chro-
matography–time-of-flight mass spectrometry (GC–TOFMS).” Journal of Food
Composition and Analysis, 21 (5): 416–22.
Orav, A., A. Raal, and E. Arak. 2008. “Essential oil composition of Pimpinella anisum L.
fruits from various European countries.” Natural Product Research, 22 (3): 227–32.
Oriani, V. B., I. D. Alvim, B. N. Paulino, F. R. Procópio, G. M. Pastore, and M. D.
Hubinger. 2018. “The influence of the storage temperature on the stability of lipid
microparticles containing ginger oleoresin.” Food Research International, 109:
472–80.
Ortner, E., M. Granvogl, and P. Schieberle. 2016. “Elucidation of thermally induced
changes in key odorants of white mustard seeds (Sinapis alba L.) and rapeseeds
(Brassica napus L.) using molecular sensory science.” Journal of Agricultural and
Food Chemistry, 64 (43): 8179–90.
Park, S. Y., M. L. Jin, Y. H. Kim, Y. Kim, and S. J. Lee. 2012. “Anti-inflammatory effects
of aromatic-turmerone through blocking of NF-κB, JNK, and p38 MAPK signaling
pathways in amyloid β-stimulated microglia.” International Immunopharmacology,
14 (1): 13–20.
Parmar, M. R., V. B. Bhalodiya, and R. L. Rajput. 2018. “Effect of drying conditions on
phytochemicals in vacuum drying.” International Journal of Chemical Studies, 6
(5): 3278–84.
Parthasarathy, V. A., B. Chempakam, and T. J. Zachariah, eds. 2008. Chemistry of
Spices. Oxford, UK: CAB International.
Patel, K., C. Ruiz, R. Calderon, M. Marcelo, and R. Rojas. 2016. “Characterization
of volatile profiles in 50 native Peruvian chili pepper using solid phase microex-
traction–gas chromatography mass spectrometry (SPME–GCMS).” Food Research
International, 89: 471–5.
Pélissier, Y., C. Haddad, C. Marion, M. Milhau, and J. Bessière. 2001. “Volatile constitu-
ents of fruit pulp of Dialium guineense Wild. (Cesalpinaceae).” Journal of Essential
Oil Research, 13 (2): 103–4.
Pérez-Silva, A., E. Odoux, P. Brat, F. Ribeyre, G. Rodriguez-Jimenes, V. Robles-Olvera,
M. A. García-Alvarado, and Z. Günata. 2006. “GC-MS and GC-olfactometry
analysis of aroma compounds in a representative organic aroma extract from cured
vanilla (Vanilla planifolia G. Jackson) beans.” Food Chemistry, 99 (4): 728–35.
Petrović, S., M. Drobac, L. Ušjak, V. Filipović, M. Milenković, and M. Niketić. 2017.
“Volatiles of roots of wild-growing and cultivated Armoracia macrocarpa and
their antimicrobial activity, in comparison to horseradish, A. rusticana.” Industrial
Crops and Products, 109: 398–403.
Pino, J., E. Sauri-Duch, and R. Marbot. 2006. “Changes in volatile compounds of
Habanero chile pepper (Capsicum chinense Jack. cv. Habanero) at two ripening
stages.” Food Chemistry, 94 (3): 394–8.
Pino, J., P. Borges, and E. Roncal. 1993. “The chemical composition of laurel leaf oil
from various origins.” Molecular Nutrition and Food Research, 37 (6): 592–5.
Pino, J., V. Fuentes, and O. Barrios. 2011. “Volatile constituents of Cachucha peppers
(Capsicum chinense Jacq.) grown in Cuba.” Food Chemistry, 125 (3): 860–4.
 Spices and Herbs 591

Pirbalouti, A. G., E. Mahdad, and L. Craker. 2013. “Effects of drying methods on qual-
itative and quantitative properties of essential oil of two basil landraces.” Food
Chemistry, 141 (3): 2440–9.
Pirbalouti, A. G., S. Salehi, and L. Craker. 2017. “Effect of drying methods on qualita-
tive and quantitative properties of essential oil from the aerial parts of coriander.”
Journal of Applied Research on Medicinal and Aromatic Plants, 4: 35–40.
Potter, T. L. and I. S. Fagerson. 1990. “Composition of coriander leaf volatiles.” Journal
of Agricultural and Food Chemistry, 38 (11): 2054–6.
Rao, L. J., M. Singh, B. Raghavan, and K. O. Abraham. 1998. “Rosemary (Rosmarinus offici-
nalis L.): Impact of drying on its flavor quality.” Journal of Food Quality, 21 (2): 107–15.
Ravi, R., M. Prakash, and K. K. Bhat. 2013. “Characterization of aroma active com-
pounds of cumin (Cuminum cyminum L.) by GC–MS, e-Nose, and sensory tech-
niques.” International Journal of Food Properties, 16 (5): 1048–58.
Ribeiro-Santos, R., M. Andrade, N. R. de Melo, F. R. dos Santos, I. de Araújo Neves,
M. G. de Carvalho, and A. Sanches-Silva. 2017. “Biological activities and major
components determination in essential oils intended for a biodegradable food pack-
aging.” Industrial Crops and Products, 97: 201–10.
Ríos, J. J., E. Fernández-García, M. I. Mínguez-Mosquera, and A. Pérez-Gálvez. 2008.
“Description of volatile compounds generated by the degradation of carotenoids in
paprika, tomato and marigold oleoresins.” Food Chemistry, 106 (3): 1145–53.
Rivlin, R. S. 2001. “Historical perspective on the use of garlic.” Journal of Nutrition,
131 (3): 951S–54S.
Rodríguez-Burruezo, A., H. Kollmannsberger, M. C. González-Mas, S. Nitz, and N.
Fernando. 2010. “HS-SPME comparative analysis of genotypic diversity in the vola-
tile fraction and aroma-contributing compounds of Capsicum fruits from the ann-
uum–chinense–frutescens complex.” Journal of Agricultural and Food Chemistry, 58
(7): 4388–400.
Roslon, W., E. Osinska, and J. Gajc-Wolska. 2010. “The influence of raw material sta-
bilization on the quality of celery (Apium graveolens L.) leaves.” In Proceedings of
the VI International Postharvest Symposium, Turkey, 2009, pp. 201–8. The Hauge:
Secretariat of ISHS, International Society for Horticultural Science.
Rubió, L., M. J. Motilva, and M. P. Romero. 2013. “Recent advances in biologically
active compounds in herbs and spices: A review of the most effective antioxidant
and anti-inflammatory active principles.” Critical Reviews in Food Science and
Nutrition, 53 (9): 943–53.
Sahebkar, A. and M. Iranshahi. 2010. “Biological activities of essential oils from the
genus Ferula (Apiaceae).” Asian Biomedicine, 4 (6): 835–47.
Sahebkar, A. and M. Iranshahi. 2011. “Volatile constituents of the genus Ferula
(Apiaceae): A review.” Journal of Essential Oil Bearing Plants, 14 (5): 504–31.
Saxena, S. N., Y. K. Sharma, S. S. Rathore, K. K. Singh, P. Barnwal, R. Saxena, P.
Upadhyaya, and M. M. Anwer. 2015. “Effect of cryogenic grinding on volatile oil,
oleoresin content and anti-oxidant properties of coriander (Coriandrum sativum L.)
genotypes.” Journal of Food Science and Technology, 52 (1): 568–73.
Sebranek, J. G., A. L. Jackson-Davis, K. L. Myers, and N. A. Lavieri. 2012. “Beyond cel-
ery and starter culture: Advances in natural/organic curing processes in the United
States.” Meat Science, 92 (3): 267–73.
Sellami, I. H., I. Bettaieb, S. Bourgou, R. Dahmani, F. Limam, and B. Marzouk. 2012.
“Essential oil and aroma composition of leaves, stalks and roots of celery (Apium gra-
veolens var. dulce) from Tunisia.” Journal of Essential Oil Research, 24 (6): 513–21.
592 Food Aroma Evolution

Sellami, I. H., W. A. Wannes, I. Bettaieb, S. Berrima, T. Chahed, B. Marzouk, and F.

Limam. 2011. “Qualitative and quantitative changes in the essential oil of Laurus
nobilis L. leaves as affected by different drying methods.” Food Chemistry, 126 (2):
691–7.
Shahwar, M. K., A. H. El-Ghorab, F. M. Anjum, M. S. Butt, S. Hussain, and M. Nadeem.
2012. “Characterization of coriander (Coriandrum sativum L.) seeds and leaves:
Volatile and non volatile extracts.” International Journal of Food Properties, 15
(4):736–47.
Shan, B., Y. Z. Cai, J. D. Brooks, and H. Corke. 2007. “The in vitro antibacterial activ-
ity of dietary spice and medicinal herb extracts.” International Journal of Food
Microbiology, 117 (1): 112–9.
Sharma, A., P. K. Rai, and S. Prasad. 2018. “GC–MS detection and determination of
major volatile compounds in Brassica juncea L. leaves and seeds.” Microchemical
Journal, 138: 488–93.
Silva, A. P., Z. Günata, J. P. Lepoutre, and E. Odoux. 2011. “New insight on the genesis
and fate of odor-active compounds in vanilla beans (Vanilla planifolia G. Jackson)
during traditional curing.” Food Research International, 44 (9): 2930–7.
Singh, G., S. Maurya, M. P. de Lampasona, and C. Catalan. 2006. “Chemical con-
stituents, antimicrobial investigations and antioxidative potential of volatile oil and
acetone extract of star anise fruits.” Journal of the Science of Food and Agriculture,
86 (1): 111–21.
Soliman, K. M. and R. I. Badeaa. 2002. “Effect of oil extracted from some medicinal
plants on different mycotoxigenic fungi.” Food and Chemical Toxicology, 40 (11):
1669–75.
Song, W. Y., Y. B. Ma, X. Bai, X. M. Zhang, Q. Gu, Y. T. Zheng, J. Zhou, and J. J.
Chen. 2007. “Two new compounds and anti-HIV active constituents from Illicium
verum.” Planta Medica, 73 (4): 372–5.
Sowbhagya, H. B., S. R. Sampathu, and N. Krishnamurthy. 2007. “Evaluation of size
reduction on the yield and quality of celery seed oil.” Journal of Food Engineering,
80 (4): 1255–60.
Srinivasan, K. 2014. “Antioxidant potential of spices and their active constituents.”
Critical Reviews in Food Science and Nutrition, 54 (3): 352–72.
Srinivasan, K. 2017. “Antimutagenic and cancer preventive potential of culinary spices
and their bioactive compounds.” PharmaNutrition, 5 (3): 89–102.
Szumny, A., A. Figiel, A. Gutiérrez-Ortíz, and Á. A. Carbonell-Barrachina. 2010.
“Composition of rosemary essential oil (Rosmarinus officinalis) as affected by dry-
ing method.” Journal of Food Engineering, 97 (2), 253–60.
Takeoka, G. 2001. “Volatile constituents of asafoetida.” In Aroma Active Compounds
in Foods: Chemistry and Sensory Properties, edited by G. R. Takeoka, M. Güntert,
and K. H. Engel, pp. 33–44. Washington, DC: American Chemical Society.
Torres-Palazzolo, C., D. Ramirez, D. Locatelli, W. Manucha, C. Castro, and A. Camargo.
2018. “Bioaccessibility and permeability of bioactive compounds in raw and cooked
garlic.” Journal of Food Composition and Analysis, 70: 49–53.
Tuan, D. Q. and S. G. Ilangantileket. 1997. “Liquid CO2 extraction of essential oil from
star anise fruits (Illicium verum H.).” Journal of Food Engineering, 31 (1): 47–57.
Tucker, A. O., M. J. Maciarello, and L. R. Landrum. 1991. “Volatile leaf oils of Caribbean
Myrtaceae. II. Pimenta dioica (L.) Merr. of Jamaica.” Journal of Essential Oil
Research, 3 (3): 195–6.
 Spices and Herbs 593

Uchida, K., Y. Miura, M. Nagai, and M. Tominaga. 2012. “Isothiocyanates from Wasabia
japonica activate transient receptor potential ankyrin 1 channel.” Chemical Senses,
37 (9): 809–18.
Urbani, E., F. Blasi, C. Chiesi, A. Maurizi, and L. Cossignani. 2015. “Characterization
of volatile fraction of saffron from central Italy (Cascia, Umbria).” International
Journal of Food Properties, 18 (10): 2223–30.
USDA. 2016. ARS; National Genetic Resources Program. Germplasm Resources
Information Network (GRIN). Beltsville, MD: National Germplasm Resources
Laboratory. Accessed January 18, 2016.
Velázquez, R., A. Hernández, A. Martín, E. Aranda, G. Gallardo, T. Bartolomé, and
M. G. Córdoba. 2014. “Quality assessment of commercial paprikas.” International
Journal of Food Science and Technology, 49 (3): 830–9.
Venskutonis, P. R. 1997. “Effect of drying on the volatile constituents of thyme (Thymus
vulgaris L.) and sage (Salvia officinalis L.).” Food Chemistry, 59 (2): 219–27.
Wang, R., R. Wang, and B. Yang. 2011. “Comparison of volatile compound composition
of cinnamon (Cinnamomun cassia Presl) bark prepared by hydrodistillation and
headspace solid phase microextraction.” Journal of Food Process Engineering, 34
(1): 175–85.
Weerakkody, N. S., N. Caffin, M. S. Turner, and G. A. Dykes. 2010. “In vitro antimi-
crobial activity of less-utilized spice and herb extracts against selected food-borne
bacteria.” Food Control, 21 (10): 1408–14.
Wichtl, M. 2004. Herbal Drugs and Phytopharmaceuticals. Stuttgart, Germany:
MedPharm GmbH Scientific Publishers.
Wilson, L. 2016. “Spices and flavoring crops: Fruit and seeds.” In Encyclopedia of Food
and Health, edited by B. Caballero, P. M. Finglas, and F. Toldrá. Oxford, UK:
Academic Press.
Yousif, A. N., T. D. Durance, C. H. Scaman, and B. Girard. 2000. “Headspace volatiles
and physical characteristics of vacuum‐microwave, air, and freeze‐dried oregano
(Lippia berlandieri Schauer).” Journal of Food Science, 65 (6): 926–30.
Ziino, M., C. Condurso, V. Romeo, G. Tripodi, and A. Verzera. 2009. “Volatile com-
pounds and capsaicinoid content of fresh hot peppers (Capsicum annuum L.) of
different Calabrian varieties.” Journal of the Science of Food and Agriculture, 89
(5): 774–80.
 Chapter 28
Off-Flavors in Alcoholic
Beverages: An Overview
Rosa Perestrelo, Catarina Silva, and José S. Câmara

CONTENTS

28.1 Introduction 595

28.2 The Most Common Off-Flavors in Alcoholic Beverages 596
28.2.1 Fatty Acids 596
28.2.2 Alcohols 596
28.2.3 Aldehydes 605
28.2.4 Anisoles 606
28.2.5 Furans 606
28.2.6 Ketones 607
28.2.7 Pyrazines 607
28.2.8 Volatile Phenols 608
28.2.9 Sulfur Compounds 609
28.3 Prevention and Reduction of Off-Flavors in Alcoholic Beverages 610
28.3.1 Procedures for Preventing the Off-Flavor Formation 610
28.3.2 Procedures for Reducing or Removing Off-Flavors 612
28.4 Future Outlook 613
References 614

28.1 INTRODUCTION

Over the past decade, consumer complaints related to off-flavors of food and beverages
have become one of the most common problems confronting the industry. An off-flavor
is an atypical odor resulting from degradation of food components (e.g., lipid oxidation,
microbiological spoilage, the Maillard reaction), incidental contamination from the envi-
ronment (e.g., packaging materials), and loss of key odorants (Sakamaki et al., 2011;
Ridgway, Lalljie, and Smith, 2009). Typically, their presence in an alcoholic beverage does
not represent a safety risk to the consumer, but the perception of low quality and adverse
publicity can be tremendously costly to the industry (Ridgway, Lalljie, and Smith, 2009). In
this sense, a lot of research has been done to develop and implement preventive and remedi-
ation approaches for reducing and removing off-flavors from alcoholic beverages without
causing a negative impact on the sensorial profile. The preventive actions are focused on
their hygienic nature, namely sanitized front-end processing equipment, whereas the reme-
diation approach can be organized into two groups, namely those intended to decrease the
headspace off-flavor concentration and those directed to the remove of off-flavors from the

595
596 Food Aroma Evolution

alcoholic beverages. Most of the proposed remediation approaches are not yet authorized
by the International Organisation of Vine and Wine (OIV). Despite decades of research,
deterioration of flavor, including loss of fresh aromas, and the appearance of off-flavors,
is still probably the greatest challenge for breweries and wineries. As stated above, several
biochemical and chemical mechanisms are involved in the formation of off-flavors in alco-
holic beverages, and the most common are reported below.

28.2 THE MOST COMMON OFF-FLAVORS IN ALCOHOLIC BEVERAGES

28.2.1 Fatty Acids

Fatty acids constitute an important chemical family of aroma compounds that can con-
tribute to cheesy, rancid, and fatty odors (Ma et al., 2016). The production of nonvolatile
and volatile acids by yeast is predominantly associated with glycolysis (e.g., acetic acid), the
tricarboxylic acid cycle (e.g., malic acid and citric acid), amino acid metabolism (e.g., phenyl-
acetic acid), and fatty acid metabolism (e.g., butanoic acid, hexanoic acid, and octanoic acid)
(Liu, 2015). The volatile short-chain acids, such as pyruvic, acetic, lactic, citric, succinic,
and malic acid impart a bitter flavor to beer (Blanco, Andrés-Iglesias, and Montero, 2016).
These are primarily originated from wort and are undesirable for the taste of beer and foam
stability, while medium-chain fatty acids, such as hexanoic, octanoic, and decanoic acid
afforded off-flavors, characterized as rancid goaty, are often called caprylic flavor (Table
28.1). Additionally, acetic acid is the main volatile acid in alcoholic beverages (e.g., wine,
beer, and cider), causing a significant reduction in quality when its content is higher than the
odor threshold (OT). For this reason, the control of acetic acid production during winemak-
ing and brewing is crucial (Zhang, Jia, and Zhang, 2012). In this sense, Spanish regulation
established a maximum concentration of acetic acid in cider (Zuriarrain et al., 2015).

28.2.2 Alcohols

Alcohols are synthesized by yeast during fermentation through catabolic and anabolic
pathways (amino acid metabolism), as can be observed in Figure 28.1 (Styger, Prior, and
Bauer, 2011). However, some alcohols are already present in the grape and are sustained
throughout the fermentation process. The concentration of alcohols is determined by
the efficiency of the corresponding amino acid uptake and sugar utilization rate (Blanco,
Andrés-Iglesias, and Montero, 2016). It has been reported that concentrations of alcohols
below 300 mg/L contribute positively to the global aroma perception, whereas concentra-
tions higher than 400 mg/L can have a detrimental effect (Swiegers et al., 2005). Special
attention should be given to the presence of geosmin, 1-octen-3-ol, methylisoborneol,
fenchol, and trans-octenol since they are responsible for musty or moldy off-flavors, even
at very low concentrations (nanograms per liter), due to their low OT (Callejón et al.,
2016). Geosmin, methylisoborneol, and 1-octen-3-ol are produced as secondary metabo-
lites by specific microorganisms. These alcohols have been detected in grapes and wines,
and are associated with rotten grapes and grape juices contaminated by Botrytis cinerea
(Callejón et al., 2016; Lopez Pinar et al., 2017; Sadoughi et al., 2015). 1-Octen-3-ol off-
flavor had a higher contribution to the overall aroma composition of wines from Croatia
(Maslov et al., 2017). Fenchol and trans-octenol have also been reported in musts or
crushed grapes (Swiegers et al., 2005). In beer, propanol, butanol, and 3-methyl-1-butanol
TABLE 28.1 Main Off-Flavors Identified in Alcoholic Beverages
Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
Fatty acids
Acetic acid Vinegar-like Saccharomyces can produce acetic Beer 71–200 mg/L (Swiegers et al., 2005; Zhang, Jia, and
character, sour acid; excessive concentrations Wine 280 mg/L Zhang, 2012)
are produced by aerobic acetic
acid bacteria
Butanoic acid Cheese, rancid Formed by anabolic pathways in Beer 1899 (Campo et al., 2006; Petropulos et al.,
yeast; β-oxidation of higher fatty Wine 173 2014; Kishimoto et al., 2018)
Decanoic acid Fatty, acids Beer 10 mg/mL (Campo et al., 2006; Petropulos et al.,
unpleasant Wine 1000 2014; Ma et al., 2016)
Dodecanoic acid Dry, laurel oil Wine 1000 (Campo et al., 2006; Petropulos et al.,
flavor, metallic 2014)
Hexanoic acid Cheese, rancid Beer 5 mg/mL (Campo et al., 2006; Petropulos et al.,
Wine 420 2014; Ma et al., 2016)
3-Methylbutanoic Cheese, rancid Beer 1230 (Campo et al., 2006; Petropulos et al.,
acid Wine 33 2014; Kishimoto et al., 2018)
Octanoic acid Cheese, fatty Beer 10 mg/mL (Campo et al., 2006; Petropulos et al.,
acid harsh, Wine 500 2014; Ma et al., 2016)
rancid
Alcohols
3-Methyl-1-butanol Mold, fusel Formed during fermentation by Beer 17 mg/L (Campo et al., 2006)(Kishimoto et al.,
Off-Flavors in Alcoholic Beverages: An Overview

alcohol, harsh deamination and Wine 30 mg/L 2018)

decarboxylation reactions from
isoleucine
1-Octanol Mold, corky Contamination from the corked Beer 900 (Cheng et al., 2015; Tan and Siebert,
top Wine 120 2004)
(Continued )
597
TABLE 28.1 (Continued) Main Off-Flavors Identified in Alcoholic Beverages
598

Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
1-Hexanol Herbaceous Formed by Pichia fermenting; Beer 4000 (Swiegers et al., 2005; Cheng et al.,
Yeast activity Wine 8000 2015; Tan and Siebert, 2004)
1-Octen-3-ol Fungus, Fungal flora on the grape; Beer 31 (Boutou and Chatonnet, 2007; Callejón
mushroom Contamination from the corked Wine 40 ng/L et al., 2016)
top
2-Methylisoborneol Earthy, Fungal flora on the grape Wine 30–55 ng/L (Boutou and Chatonnet, 2007; Callejón
muddy-earthy, et al., 2016)
musty
Eosin Camphor, Fungal flora on the grape; Wine 50–90 ng/L (Boutou and Chatonnet, 2007; Callejón
muddy-earthy, Contamination from the corked et al., 2016)
moldy top
(+)-Fenchol Muddy Fungal flora on the grape Wine 50 (Boutou and Chatonnet, 2007; Callejón
et al., 2016)
Aldehydes
Acetaldehyde Aldehydic, Yeast activity, Ethanol oxidation, Beer 25 mg/L (Swiegers et al., 2005; Tan and Siebert,
cidery, fruity, Microbial contamination Wine 100 mg/L 2004)
Food Aroma Evolution

grassy, green,
sour and
musty
Acrolein Acrid Formed microbiologically Beer 15 mg/L (Ridgway, Lalljie, and Smith, 2010; Tan
and Siebert, 2004)
(E,E)-2,4-Decadienal Deep-fried, Deterioration of flavor and aroma Beer 0.3 (Tan and Siebert, 2004)
papery, during storage
cardboard
(Continued )
TABLE 28.1 (Continued) Main Off-Flavors Identified in Alcoholic Beverages
Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
3-Methylbutanal Malty, potato Strecker degradation Beer 9.6 (Mayr et al., 2015; Kishimoto et al.,
almond Wine 6 2018)
2-Methylpropanal Malty Beer 86–1000 (Mayr et al., 2015; Andrés-Iglesias et
Wine 4.6 al., 2016)
(E)-2-Nonenal Cardboard, Lipid oxidation Beer 0.10 (Mayr et al., 2015; Kishimoto et al.,
cucumber, Wine 0.17 2018)
papery
Phenylacetaldehyde Cooked potato, Strecker degradation Beer 39.7 (Rodrigues et al., 2011; Mayr et al.,
floral, honey Wine 1 2015; Kishimoto et al., 2018)
Anisoles
TCA Corky, mold, Bio-methylation of corresponding Beer 12 (Slabizki and Schmarr, 2013; Callejón et
musty phenol, source of pollution Wine 0.03–50 ng/L al., 2016)
TeCA Mold-dust, variable (e.g., wood Wine 5–35 ng/L (Boutou and Chatonnet, 2007; Slabizki
musty preservatives) and Schmarr, 2013; Callejón et al.,
2016)
TBA Corky, mold, Wine 2–7.9 ng/L (Slabizki and Schmarr, 2013; Callejón et
musty al., 2016)
PCA Musty-earthy, Wine 10 (Boutou and Chatonnet, 2007; Callejón
musty et al., 2016)
2,6-Dichloroanisole Corky, musty — (Boutou and Chatonnet, 2007)
Off-Flavors in Alcoholic Beverages: An Overview

2,6-Dichlorophenol Disinfectant, Contamination of yeast hulls — (Boutou and Chatonnet, 2007)

medicinal added during fermentation
Furans
2-Furfuryl ethyl Solvent, papery, Deterioration of flavor and aroma Beer 6 (Rodrigues et al., 2011)
ether cardboard during storage; Thermal
degradation
(Continued )
599
TABLE 28.1 (Continued) Main Off-Flavors Identified in Alcoholic Beverages
600

Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
2-Furanmethanol Burned, sweet Thermal degradation Wine 2000 (Herrero et al., 2016)
2-Furfural Bitter, burned, Deterioration of flavor and aroma Beer 15–50 mg/L (Rodrigues et al., 2011; Mayr et al.,
caramel, nutty, during storage; Thermal Wine 14 mg/L 2015)
winey degradation
HMF Bready, Deterioration of flavor and aroma Beer 36 mg/L (Rodrigues et al., 2011)
cardboard, during storage; Thermal Wine 100 mg/L
caramel, degradation
cucumber,
papery
5-Methylfurfural Bitter almond, Deterioration of flavor and aroma Wine 2000 (Tao et al., 2016; Herrero et al., 2016;
sweet during storage; Thermal Mayr et al., 2015)
degradation
Sotolon Burned sugar, Aldol condensation of Beer 0.54 (Pereira et al., 2018; Mayr et al., 2015;
curry, spice acetaldehyde and α-ketobutyric Wine 5–15 Kishimoto et al., 2018)
acid followed by lactonization;
Thermal degradation
Ketones
Food Aroma Evolution

Acetoin Butter, cream Yeast activity Wine 150 mg/L (Swiegers et al., 2005)
2,3-Butanedione Buttery Yeast activity Beer 100–200 (Swiegers et al., 2005; Andrés-Iglesias et
Wine 200 al., 2016)
(+)-Fenchone Earthy- Fungal flora on the grape Wine 110–500 (Boutou and Chatonnet, 2007)
camphor,
muddy
1-Octen-3-one Fungus, Fungal flora on the grape; Beer 0.003 (Boutou and Chatonnet, 2007; Callejón
mushroom Auto-oxidation of fats Wine 40–70 et al., 2016; Kishimoto et al., 2018)
(Z)-1,5-Octadien-3- Metallic, raw Auto-oxidation of fats (methyl Beer 0.0004 (Allamy, Darriet, and Pons, 2017;
one fish linolenate) Wine 0.009 Kishimoto et al., 2018)
(Continued )
TABLE 28.1 (Continued) Main Off-Flavors Identified in Alcoholic Beverages
Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
2,3-Pentanedione Buttery Yeast activity Beer 900–1000 (Tian, 2010; Andrés-Iglesias et al.,
2016)
Pyrazines
DMMP Fungal must Endogenous presence in the grape Wine 2 ng/L (Fontana, 2012; Callejón et al., 2016)
IPMP Muddy Endogenous presence in the grape Beer 6 (Boutou and Chatonnet, 2007; Callejón
Wine 2 ng/L et al., 2016)
IBMP Green pepper, Endogenous presence in the Wine 1–16 ng/L (Boutou and Chatonnet, 2007; Callejón
herbaceous, grape; Fungal flora on the grape et al., 2016)
vegetal
MDP Corky Bacterial or fungal contamination Wine 2.1 ng/L (Boutou and Chatonnet, 2007;
of the cork Simpson, Capone, and Sefton, 2004)
SBMP Bell pepper, ivy Endogenous presence in the Wine 1–2 ng/L (Alberts et al., 2009)
leaves, peas grape; Fungal flora on the grape
Phenols
4-Ethylphenol Phenolic, Formed by Brettanomyces Wine 430 (Boutou and Chatonnet, 2007; Swiegers
leather Dekkera yeasts et al., 2005; Buron et al., 2012)
4-Ethyl guaiacol Phenolic, spicy, Formed by Brettanomyces Wine 33 (Boutou and Chatonnet, 2007; Swiegers
pharmaceuti- Dekkera yeasts et al., 2005; Buron et al., 2012)
cal
4-Ethyl catechol Phenolic Formed by Brettanomyces Wine 50 (Boutou and Chatonnet, 2007; Buron et
Off-Flavors in Alcoholic Beverages: An Overview

Dekkera yeasts al., 2012; Buron et al., 2011)

4-Vinyl guaiacol Pharmaceutical, Formed by Saccharomyces Beer 98 (Boutou and Chatonnet, 2007; Swiegers
phenolic, cerevisiae yeasts Wine 380 et al., 2005; Kishimoto et al., 2018)
smoke, sweet
4-Vinylphenol Pharmaceutical, Formed by Saccharomyces Wine 1500 (Boutou and Chatonnet, 2007; Swiegers
gouache cerevisiae yeasts et al., 2005)
(Continued )
601
TABLE 28.1 (Continued) Main Off-Flavors Identified in Alcoholic Beverages
602

Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
Guaiacol Phenolic, Bacterial (or thermic) Beer 65 (Boutou and Chatonnet, 2007; Neto,
smoke, sweet transformation of the vanillin in Rocha, and Silvestre, 2007)
guaiacol; Contamination from
the corked top
CMP Plastic, Contamination of yeast hulls — (Capone et al., 2010)
disinfectant, added during fermentation
medicinal
Sulfur compounds
Benzenemethanethiol Struck flint Formed by the reaction of Wine 0.3 ng/L (Smith et al., 2015)
hydrogen sulfide with ethanol or
acetaldehyde
Dimethyl sulfide Asparagus, Formed by the reaction of Beer 30 (Swiegers et al., 2005; Smith et al.,
boiled hydrogen sulfide with ethanol or Wine 25 2015)
cabbage, acetaldehyde; Oxidation of the
asparagus, mercaptans
corn, truffle
Diethyl sulfide Garlic, rubber Formed by the reaction of Beer 0.4 (Smith et al., 2015)
Food Aroma Evolution

hydrogen sulfide with ethanol or Wine 0.9

acetaldehyde; Oxidation of the
mercaptans
Dimethyl disulfide Burned rubber, Formed by the reaction of Beer 7.5 (Swiegers et al., 2005; Smith et al.,
cooked hydrogen sulfide with ethanol or Wine 4.3 2015)
cabbage, acetaldehyde; Oxidation of the
onion mercaptans
Dimethyl trisulfide Cabbage, sulfur Formed by the reaction of Beer 0.00012–0.1 (Kishimoto et al., 2018)
hydrogen sulfide with ethanol or
acetaldehyde; Oxidation of the
mercaptans
(Continued )
TABLE 28.1 (Continued) Main Off-Flavors Identified in Alcoholic Beverages
Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
Ethanethiol Fecal, onion, Formed during fermentation by Beer 1.7 (Swiegers et al., 2005; Smith et al.,
putrefaction, the reaction of hydrogen sulfide Wine 1.1 2015)
rubber with ethanol or acetaldehyde
Ethylthio acetate Onion, Degradation pathways of Beer 10 (Smith et al., 2015)
sulfurous C-cysteinyl and S-glutathionyl
conjugates by yeast during
alcoholic fermentation
Hydrogen sulfide Rotten egg Yeast activity Beer 8 (Swiegers et al., 2005; Smith et al.,
Wine 1.1–1.6 2015)
Methanethiol Cooked Formed during fermentation by Beer 2 (Swiegers et al., 2005; Smith et al.,
cabbage, the reaction of hydrogen sulfide Wine 1.8–3.1 2015)
onion, with ethanol or acetaldehyde
putrefaction,
rubber
4MMP Box tree, cat Degradation pathways of Wine 3.3 ng/L (Iizuka-Furukawa et al., 2017; Smith et
urine C-cysteinyl and S-glutathionyl al., 2015)
conjugates by yeast during
alcoholic fermentation
Methylthio acetate Cheese, egg, Degradation pathways of Beer 50 (Smith et al., 2015)
sulfurous C-cysteinyl and S-glutathionyl
conjugates by yeast during
Off-Flavors in Alcoholic Beverages: An Overview

alcoholic fermentation
Methional Cooked Strecker degradation Beer 1.8–250 (Rodrigues et al., 2011; Mayr et al.,
potato-like, Wine 0.5 2015; Smith et al., 2015)
soy sauce
(Continued )
603
TABLE 28.1 (Continued) Main Off-Flavors Identified in Alcoholic Beverages
604

Odor
Off-Flavor Descriptions Origin Beverage OTs (µg/L) Reference
Methionol Cauliflower, Formed during fermentation by Beer 500–2000 (Mayr et al., 2015; Smith et al., 2015)
cooked deamination and Wine 1000
potato-like decarboxylation reactions from
methionine
2-Methyl-3- Nut, thiamine Degradation pathways of Beer 0.190 (Kishimoto et al., 2018)
furanthiol C-cysteinyl and S-glutathionyl
conjugates by yeast during
alcoholic fermentation
3-Methyl-2-butene- Roasted, Photolysis (illumination) of Beer 0.007–0.01 (Hill and Smith, 2000; Gros and Collin,
1-thiol skunky iso-α-acids from the hops and of 2012)
sulfur-containing amino acids
Legend: 
4MMP—4-mercapto-4-methyl-penta-2-one; CMP—2-chloro-6-methylphenol; DMMP—3,5-dimethyl-2-methoxypyrazine; HMF—5-
hydroymethylfurfural; IBMP—2-isobutyl-3-methoxypyrazine; IPMP—2-isopropyl-3-methoxypyrazine; MDP—2-methoxy-3,5-
dimethylpyrazine; OTs—Odor threshold; PCA—2,3,4,5,6-pentachloroanisole; Ref.—Reference; SBMP—3-sec-butyl-2-methoxypyrazine;
TBA—2,4,6-tribromoanisole; TCA—2,4,6-trichloroanisole; TeCA—2,3,4,6-tetrachloroanisole.
Food Aroma Evolution
 Off-Flavors in Alcoholic Beverages: An Overview 605

FIGURE 28.1 A schematic representation of the synthesis of major chemical families

produced by yeast during alcoholic fermentation. (Adapted from Swiegers et al., 2005;
Styger, Prior, and Bauer, 2011.)
have been associated with alcohol off-flavors, such as solvent and bitter odors (Spedding
and Aiken, 2015).

28.2.3 Aldehydes

Acetaldehyde is the main oxidation-related aldehyde identified in alcoholic beverages. This

aldehyde may be formed as an enzymatically derived by-product of yeast and acetic bacteria
metabolism, or as a non-enzymatic oxidation product of ethanol and phenolic compounds
(Ioannidou, Samouris, and Achilias, 2016; Sheridan and Elias, 2015). The concentration of
acetaldehyde depends on fermentation conditions, namely temperature, pH, yeast nutrient
availability, oxygen, and sulfur dioxide (SO2) levels (Ioannidou, Samouris, and Achilias,
2016). Although higher concentrations of acetaldehyde usually cause an unpleasant pun-
gent aroma, an appropriate acetaldehyde content can contribute a pleasant green apple
aroma (Liu et al., 2018). Its presence is highly reported in wines, beers, cider, and whiskey
(Ioannidou, Samouris, and Achilias, 2016). In addition, the off-flavor of alcoholic bever-
ages is often associated with the appearance of Strecker aldehydes. The Strecker aldehydes
can be formed by (i) Strecker degradation of parent amino acid with α-dicarbonyl or an
o-quinone; (ii) the direct oxidation of the corresponding alcohol with a peroxidation mech-
anism; and/or (iii) the release of these compounds adducts with SO2-hydroxyalkylsulfonic
acid (Monforte, Martins, and Silva Ferreira, 2017). These aldehydes contribute greatly to
the character of beer due to their low OTs and their sensitivity to aging (Table 28.1). They
include benzaldehyde, 2-methylpropanal, 2-methybutanal, 3-methylbutanal, and phenyl-
acetaldehyde. Flavors vary with compounds and concentrations but may be reminiscent
of varnish, green grass, potato, lilac, cheese, among others, and generally detract from
the perceived freshness of beer (Baert et al., 2012). Another off-flavor identified in beer
is (E)-2-nonenal that is responsible for the formation of a papery and cardboard-like fla-
vor in aged beers (Guido et al., 2004). This aldehyde can be formed by auto-oxidation or
enzymatic oxidation of linoleic and linolenic acids with lipoxygenases during mashing and
malting, and also formed by inappropriate storage (Andrés-Iglesias et al., 2015). Perhaps,
606 Food Aroma Evolution

the characteristic cardboard flavor in beer is essentially caused by (E)-2-nonenal, the effects
of (E,E)-2,4-decadienal and Strecker aldehydes could not be underestimated (Saison et al.,
2009). Hence, the addition of a mixture of Strecker aldehydes contributed to Maillard,
sulfury, and cardboard notes. Particularly, 3-methylbutanal, 2-methylpropanal, and phen-
ylacetaldehyde were found to be important on a lesser level. As noted for beers, wine aroma
changes dramatically during bottle aging due to oxidation reactions that promote the for-
mation of off-favors, mainly phenylacetaldehyde and (E)-2-nonenal. From these, methi-
onal and phenylacetaldehyde are among the oxidation-related aroma compounds that have
received attention due to their aroma impact and possible contribution to the aroma of red
and white wines (Ballester et al., 2018).

28.2.4 Anisoles

The moldy taint of wine also recognized as cork taint is due to the presence of chloro-
anisoles and bromoanisoles. Their presence is one of the main reasons for the rejection
of wine by consumers (Cravero et al., 2015). Conventionally, and incorrectly, the pres-
ence of chloroanisoles (specially 2,4,6-trichloroanisole or TCA) was exclusively associated
with the cork, but can also arise from all wood-containing materials (e.g., oak barrels).
TCA confers a very unpleasant fungal aroma to the wine, even at concentrations as low
as 2 to 4 ng/L (Cravero et al., 2015; Franc, David, and de Revel, 2009), as well as to
the beer (Kishimoto et al., 2018). The origin of TCA and its derivates is a methylation
and dehalogenation of pentachlorophenol fungicides, which are a product of the fungi
Penicillium, Aspergillus, Actinomyces, and Streptomyces spp. The products then migrate
from corks during the storage of bottled wines giving a dusty, moldy, and musty odor
to the wine (Silva et al., 2011). These molecules probably appear in wines as a conse-
quence of using fungicides, biocides, and herbicides in the vineyard, wood preservatives
in the wood used to make barrels, and corks or hypochlorite to bleach corks or wash bar-
rels (Özhan et al., 2009). This chemical family also includes 2,3,4,6-tetrachloroanisole
(TeCA), 2,3,4,5,6-pentachloroanisole (PCA), and 2,4,6-tribromoanisole (TBA). Pizarro
et al. (Pizarro, Pérez-del-Notario, and González-Sáiz, 2010) reported that TCA was pres-
ent in 62% of the analyzed wines, but confirmed that some degradation products such as
2,6-dichloroanisole (DCA) and brominated anisoles like TBA may also contribute to the
typical corky off-flavor (Pizarro, Pérez-del-Notario, and González-Sáiz, 2010). If the cork
stopper is in direct contact with the wine, volatile and nonvolatile compounds soluble in
ethanol/water can migrate to the wine, thus contributing to the wines sensory properties.
However, as a natural product, cork can be contaminated and attacked in several ways,
which lead to changes in its properties, and is potentially able to cause off-flavors in wines.
Although these compounds present low OTs, their presence even in very low concentra-
tions (down to ng/L) is enough to promote off-flavors in cork sealed beverages, particularly
in wines (Özhan et al., 2009). Other than TCA, it is known that several other compounds
can be related to off-flavors caused by cork, for example, 1-octen-3-ol, TBA, 3-methyl-
1-butanol, and 1-octanol (Neto, Rocha, and Silvestre, 2007; Slabizki and Schmarr, 2013).

28.2.5 Furans

The presence of sotolon (4,5-dimethyl-3-hydroxy-2(5H)-furanone) is known to impart

Madeira-oxidized, walnut, spicy, curry, and burned odors to many alcoholic beverages,
 Off-Flavors in Alcoholic Beverages: An Overview 607

depending on its concentration and enantiomeric distribution (Scholtes, Nizet, and Collin,
2015; Pereira et al., 2018). Sotolon can be formed through (i) aldol condensation of acet-
aldehyde and α-ketobutyric acid followed by lactonization, (ii) peroxidation of acetalde-
hyde; (iii) thermally produced form intermediates generated from the Maillard reaction,
and (iv) oxidative degradation of ascorbic acid in the presence of ethanol (König et al.,
1999; Pereira et al., 2018; Scholtes, Nizet, and Collin, 2015). Several studies have dem-
onstrated that sotolon plays an important role in the global aroma of fortified Madeira
wines (Câmara et al., 2004), aged Port wines (Ferreira, Barbe, and Bertrand, 2003),
sherry (Moreno et al., 2005), and Chinese Syrah wines (Zhao et al., 2017), at a level
generally above 10 μg/L. On the other hand, at concentrations higher than 600 μg/L,
sotolon could be responsible for the typical rancid odor found in some fortified French
wines. Moreover, the prevention of oxidation is required to decrease the Madeira off-
flavor brought to the beer by sotolon (Scholtes, Nizet, and Collin, 2015). Other furans
resulting from the Maillard reaction, namely 2-furfural, 2-furanmethanol, and 2-fur-
furyl ethyl ether are regarded as off-flavors in beers (Vanderhaegen et al., 2003). More
studies performed on beer showed that the furfural ethyl ether is an important aging
flavor, especially when present at concentrations higher than its OT (6 µg/L), resulting in
a solvent-like aging flavor (Vanderhaegen et al., 2004; Saison et al., 2009).

28.2.6 Ketones

From this chemical family, 2,3-butanedione (diacetyl), 2,3-pentanedione, 1-octen-

3-one, 3-octanone, and (Z)-1,5-octadien-3-one are the most common off-flavors
reported in alcoholic beverages (Ochando et al., 2018; Liu, 2015; Callejón et al., 2016).
2,3-Butanedione formation occurs through enzymatic and chemical processes involved
in food degradation, as Maillard reactions. In wine, it is frequently associated with the
malolactic fermentation and, to a slighter degree, with the alcoholic fermentation (Ramos
et al., 2017; Ochando et al., 2018). This chemical compound imparts butter, toasty, and
spoiled odors to wine and beer when present at concentrations higher than its OTs (Wang
et al., 2017). On the other hand, 2,3-pentanedione has a toffee-like aroma but is not as
potent as 2,3-butanedione and can also cause off-flavors in beer (Liu, 2015). 1-Octen-3-
one can be formed from the metabolism of many fungal species, such as Erysiphe necator
or Penicillium brevicompactum. This ketone, when reaching concentrations above its OT
in wines, from 40 to 70 ng/L, is responsible for the musty odors of alcoholic beverages.
Fortunately, 1-octen-3-one can be enzymatically reduced by Saccharomyces cerevisiae to
3-octanone and (Z)-5-octen-3-one during alcoholic fermentation, which is a much less
odorant compound with a higher OT (Liu, 2015).

28.2.7 Pyrazines

Pyrazines are heterocyclic naturally occurring compounds, and the classification is

dependent on their origin (Sidhu et al., 2015). In this context, alkyl methoxypyrazines
(MPs) have been identified in many foodstuffs of plant origin as relevant aroma com-
pounds of the wine, due to their low OT (1 to 10 ng/L), but can also be generated as
Maillard reaction products (Slabizki et al., 2014). The most common MPs found in wines
are 3-isopropyl-2-methoxypyrazine (IPMP), 3-isobutyl-2-methoxypyrazine (IBMP), and
3-sec-butyl-2-methoxypyrazine (SBMP). These MPs can have a positive impact on the
608 Food Aroma Evolution

aroma profile of certain wine varietals (e.g., Sauvignon blanc, Cabernet Sauvignon), with
concentrations in the range of approximately 8 to 15 ng/L, whereas concentrations higher
than 30 ng/L can be detrimental leading to unpleasant odors, such as overpowering
green, herbaceous, and unripe odors (Fontana and Bottini, 2016; Alberts et al., 2009).
The IBMP is the most abundant congener in Sauvignon blanc wine, representing approxi-
mately 80% of the total MPs content, whereas SBMP and IPMP represent around 10%
of the total MPs content (Alberts et al., 2009). Additionally, 2-methoxy-3,5-dimethylpyr-
azine is an off-flavor with an unpleasant, musty, moldy aroma, when present at concen-
trations higher than its OT (2.1 ng/L in white wine). It has been assessed by some wine
industry personnel as second regarding TCA as a cause of cork taint in Australian wine
(Simpson, Capone, and Sefton, 2004).

28.2.8 Volatile Phenols

Volatile phenols are composed of two groups, namely the vinylphenols (4-vinylphenol,
4-vinylguaiacol) and the more commonly discussed ethylphenols (e.g., 4-ethylphenol,
4-ethylguaiacol). The presence of volatile phenols is associated with aromas described
most commonly as medicinal, leather, barnyard, sweat, horsey, and animal, even at low
concentrations, due to their low OTs (Noestheden, Dennis, and Zandberg, 2018).
The volatile phenols biosynthesis during fermentation by Brettanomyces/Dekkera
bruxellensis leads to wine defect colloquially known as “Brett-taint,” as shown in Figure
28.2. B. bruxellensis yeasts usually do not require nutrition-rich environments for their
growth. It has been reported that this genus of yeast is the only microorganism that
can survive in wine after bottling, due to its ability to resist in the anaerobic conditions
(Renouf and Lonvaud-Funel, 2007). The hydroxycinnamate decarboxylase and vinylphe-
nol reductase activities of these yeasts enable them to produce 4-ethylphenol and 4-eth-
ylguaiacol from p-coumaric acid (Kheir et al., 2013). The hydroxycinnamic acids are
usually bonded in the form of tartaric acid esters being released by hydrolysis. Regarding
the metabolic activity of yeasts and bacteria, other factors such as oak maturation can
also increase the number of volatile phenols in wine. The off-flavor defects due to the
presence of these molecules are one of the main organoleptic problems occurring dur-
ing the elaboration of many fermented alcoholic beverages (e.g., wine, beer, or cider).
Although volatile phenols can contribute positively to the aroma of some wines, they
are better known for their contribution to off-flavors such as barnyard or stable, which
result from high concentrations of ethylphenols. In particular, 4-ethylguaiacol and 4-eth-
ylphenol concentrations showed a marked increase during oak maturation (Swiegers et
al., 2005). However, the volatile phenols precursor contents have been shown to be very
different in wines and ciders (Buron et al., 2011). According to the matrices, the off-flavor
thresholds can largely differ, being influenced by product complexity (Buron et al., 2012).
It was also previously shown that these compounds could be associated with animal or
spicy aromatic notes in cider derived products, such as the “Calvados” distilled bever-
age. In this cider type, 4-ethylphenol, 4-ethylguaiacol, and 4-vinylphenol were the most
concentrated phenols, but traces of 4-methyl, 4-propylguaiacol, 4-methyl, and 4-pro-
pylphenol were also detected. Recently, Buron et al. (Buron et al., 2012) showed that
4-ethylcatechol and 4-ethylphenol were the main volatile phenols found in French ciders
and phenolic off-flavor defects were found in ciders containing up to 12 and 3 mg/L,
respectively. In beers, the two main volatile phenols conferring flavor are 4-vinylguaia-
col and 4-vinylphenol. Vinylphenols, especially 4-vinylguaiacol and 4-vinylphenol, are
 Off-Flavors in Alcoholic Beverages: An Overview 609

FIGURE 28.2 A schematic representation of volatile phenols formation from hydroxi-

cinnamate precursors in wines by Bettranomyces bruxellensis/Dekkera. (Adapted from
Milheiro et al., 2017a.)

responsible for the pharmaceutical odor, particularly in white wines. In addition, 4-eth-
ylguaiacol and 4-ethylphenol can also appear in beers as a result of the reduction reaction
of their corresponding vinylphenols. The presence of these volatile phenols is appreciated
in certain beers, whereas in others, when present at high concentrations, it is considered
as an off-flavor that negatively affect their quality.

28.2.9 Sulfur Compounds

The sulfur compounds are an important chemical class for alcoholic beverage aroma due
to their extremely low OTs (in order μg/L) that can confer off-flavors such as putrefac-
tion, rotten egg, cabbage, onion, and garlic-like (Table 28.1). There are five groups of
sulfur compounds, namely heterocyclic compounds, polysulfides, thioesters, thiols, and
sulfides (Hirst and Richter, 2016).
Several sulfur compounds have been identified in alcoholic beverages, namely meth-
anethiol, ethanethiol, hydrogen sulfide (H 2S), dimethylsulfide (DMS), dimethyldisulfide
(DMDS), dimethyltrisulfide (DMTS), methional, methionol, and S-methylthioesters of
short-chain fatty acids (acetate, propanoate, butanoate) (Patrignani et al., 2016; Liu,
2015; Slaghenaufi et al., 2017). These can be the result of the enzymatic (e.g., products
610 Food Aroma Evolution

of fermentative pathways) and non-enzymatic (e.g., photochemical or thermal processes)

reactions (Fracassetti and Vigentini, 2018), and yeast-mediated conversion. For exam-
ple, the formation of sulfur compounds by yeasts include (i) the degradation of sulfur-
containing amino acids; (ii) the degradation of sulfur-containing pesticides; and (iii) the
release and/or the metabolism of grape-derived sulfur-containing precursors (Swiegers
et al., 2005). H 2S is a well-known off-flavor produced by yeast from cysteine or sulfate
via the sulfate reduction sequence, especially under nitrogen limitation. In addition, H 2S
can react with unsaturated aldehydes or ketones to generate several thiols that may be
desirable or undesirable for alcoholic beverages flavor according to their concentrations.
This includes 2-mercaptoethanol, 3-mercaptoethanol, 3-mercaptohexanol, 4-mercapto-
4-methyl-2-pentanone, and acetate esters such as 3-mercaptohexy acetate (Liu, 2015).
Polyfunctional thiols have been related to beer off-flavor, such as 3-methyl-2-buten-1-
thiol (3MBT), 4-mercapto-4-methyl-2-pentanone, and 3-mercapto-3-methylbutyl-for-
mate. 3MBT is a well-known skunky off-flavor in beer and is generated when sunlight
strikes a beer bottle (Noba et al., 2018), whereas 4-mercapto-4-methyl-2-pentanone
and 3-mercapto-3-methylbutyl-formate are responsible for a ribs and cats off-flavor.
Moreover, 2-mercapto-3-methyl-1-butanol (2M3MB) has been identified in beer and
has an onion-like off-flavor. 3-Mercapto-3-methyl-1-butanol (3M3MB) is an isomer of
2M3MB, imparts a similar flavor, and has been detected in beer and wine (Noba et al.,
2018, 2017). Noba et al. (2017) previously quantified the OT values for 2M3MB and
3M3MB in beer samples and showed that only 2M3MB contributes to the onion-like
off-flavor in beer. 3-Mercapto-2-methylbutanol and 2-mercapto-3-methylbutanol were
associated with onion-like odors in beer (Vichi, 2015). H 2S, methanethiol, and ethane-
thiol and propanethiol have been reported for putrefaction, garlic, onion, and rotten egg-
like odors in beers and wines (Vichi, 2015). In addition, methanethiol and ethanethiol,
described as onion or rubber, are highly reactive compounds with low OTs around 1.1
μg/L, affecting wines in clear bottles and exposed to light (Fracassetti and Vigentini,
2018; Swiegers et al., 2005; Grant-Preece et al., 2017). Another off-flavor, namely ben-
zenemethanethiol also known as benzylmercaptan has been identified in Chardonnay
and Sauvignon Blanc wines as having a struck flint/burned aroma and can be considered
part of the style rather than a defect when found at a higher concentration (Smith et al.,
2015). Generally, the concentration of sulfur compounds decreases during wine aging,
being the reduction of these compounds more accentuated in oak barrel aging stainless
steel tank aging (Ye et al., 2016).

28.3 PREVENTION AND REDUCTION OF OFF-

FLAVORS IN ALCOHOLIC BEVERAGES

One of the main challenges of the beverage industry is to obtain products that are of
high quality and competitive in the market. In this sense, several preventive procedures
to avoid microbial wine contamination, proliferation, and remedial treatments have been
proposed to improve the wine sensory impact by avoiding the off-flavors formation.

28.3.1 Procedures for Preventing the Off-Flavor Formation

Currently, emerging non-thermal technologies for alcoholic beverage preservation is a

subject of increasing importance due to the efficiency of these technologies on controlling
 Off-Flavors in Alcoholic Beverages: An Overview 611

pathogenic or spoilage microorganisms responsible for the off-flavor formation (Morata

et al., 2017). The main advantage of these technologies is the reduction of energy con-
sumption, improvement of process safety, preservation of nutritional properties, and
increasing global quality of the final product (Paniagua-Martínez et al., 2018). Several
studies have focused on implementing non-thermal technologies in wines and beers to
inactivate Brettanomyces or Dekkera bruxellensis (van Wyk and Silva, 2017; Morata
et al., 2017; van Wyk, Farid, and Silva, 2018) and Saccharomyces cerevisiae (Milani,
Alkhafaji, and Silva, 2015), respectively.
Brettanomyces or Dekkera bruxellensis is considered a major spoilage concern
for the wine industry worldwide, leading to detrimental economic loss worldwide.
Undesirable off-flavors described as medicinal, horsey, or barnyard-like can result from
the presence of as little as 104 CFU/mL Brettanomyces (van Wyk, Farid, and Silva, 2018).
Moreover, these yeasts can survive, proliferate, and contaminate the wine during sev-
eral steps of the winemaking process (Cravero et al., 2018). The addition of SO2 , an
antimicrobial and antioxidant additive, is the most common and efficient procedure to
prevent Brettanomyces contamination. However, due to its negative effects on consum-
ers including allergic reaction, headaches, asthma, and abdominal pain, new emerging
non-thermal technologies to destroy or strongly minimize the initial wild microbiota
allowing more hygienic winemaking process have been proposed (van Wyk and Silva,
2017; Morata et al., 2017; van Wyk, Farid, and Silva, 2018). Van Wyk and Silva (2017)
studied the effect of high-pressure processing (HPP) on the inactivation of B. bruxellen-
sis in red wines. The results showed that increasing treatment time and pressure leads to
increased yeast inactivation, and using 400 MPa for 5 s led to the complete inactivation
(˃6 log reduction) of B. bruxellensis. Cravero et al. (2018) investigated the potentiality
of electrolyzed water, aqueous, and gaseous ozone to control B. bruxellensis on wine
grapes. The antimicrobial potential against B. bruxellensis inocuon postharvest grapes
was demosntrated through these procedures. In another study performed by Cravero et
al. (2016), a decrease was observed of about 0.5 log CFU/mL of the total yeast popula-
tion on the grapes’ surface independently of the dose of electrolyzed water applied. The
introduction of this innovative antimicrobial agent allows the reduction of SO2 during
the winemaking process. Lustrato et al. (2015) presented an innovative approach based
on low electric current treatment (LEC) efficient to inhibit wine spoilage by D. bruxel-
lensis and to prevent the formation of off-flavors during red wine storage in oak barrels.
On the other hand, Durner et al. (2017) evaluated the potentiality of UV-C technology
to inactivate yeast and bacteria in must and wine. However, UV-C technology is known
for its ability to produce off-flavors, such as methional, but according to these authors the
application of UV-C at doses lower than 2 kJ/L is uncritical based on sensory analysis.
Fang, Hallam, and Bermúdez (2016) tested the performance of non-thermal plasma air
purifiers on its removal effectiveness of TCA and TBA in wine cellars, which are respon-
sible for unpleasant moldy and musty odors. According to these authors, the proposed
method was effective in removing TCA and TBA in a wine cellar after five days of the
operation of non-thermal plasma air purifiers.
Saccharomyces cerevisiae is used in the brewing and baking industries, and its unde-
sirable activity after fermentation produces off-flavors such as 2,3-butanedione, alde-
hydes, ketones, and sulfur compounds (Milani, Alkhafaji, and Silva, 2015; Milani and
Silva, 2016, 2017). Milani, Alkhafaji, and Silva (2015) investigated the feasibility of
a non-thermal continuous preservation technology, pulsed electric fields (PEF), for S.
cerevisiae inactivation, as well as the impact on beer sensorial properties. The results
obtained demonstrated that the inactivation of S. cerevisiae ascospores by PEF is higher
612 Food Aroma Evolution

in the beers with higher alcohol content, with a maximum of 2.2 log reduction for 7%
alc/vol beer (45 kV/cm, 70 μs treatment, T < 43°C). In another study, Milani and Silva
(2016) performed the non-thermal pasteurization of beer using HPP technology, and this
process at 600 MPa during 5 s resulted in a higher than 7 log reduction in the S. cerevisiae
ascospores. The extent of inactivation of S. cerevisiae ascospores by HPP depends on the
alcohol content. The same authors also investigated the efficiency of ultrasound-assisted
thermal pasteurization or thermosonication of beer in the inactivation of S. cerevisiae
ascospores. The TS method promoted the S. cerevisiae ascospores inactivation at 43, 50,
and 55°C in beer compared to room temperature ultrasound, with a maximum of 3.7
log reduction after 55°C and 20 min treatment (Milani and Silva, 2017). Moreover, pas-
teurization of beer by HPP, TS, and thermal pasteurization, were compared in terms of
the energy requirements and the S. cerevisiae ascospores inactivation by Milani, Ramsey,
and Silva (2016). HPP and TS can be alternatives to thermal pasteurization, since they
promote greater log reduction S. cerevisiae ascospores inactivation with shorter process-
ing time, or less energy in case of HPP. Therefore, according to these authors, HPP is a
promising technology for pasteurization in the beer industry.
The emerging non-thermal technologies that have been assessed to prevent the off-
flavor formation, such as HPP, PEF, LEC, and UV-C are very promising taking into
account their efficiency, but further investigation should be performed in order to evalu-
ate their impact more deeply on wine quality and the maintenance of its sensory proper-
ties until its consumption (Milheiro et al., 2017a).

28.3.2 Procedures for Reducing or Removing Off-Flavors

Several remedial procedures have been proposed to remove the already formed off-
flavors from alcoholic beverages as well as to reduce their negative impact on the over-
all beverage aroma, especially in wine aroma. Some studies have evaluated the impact
of these procedures on wine quality. However, most of the proposed procedures are
not yet authorized by the OIV (Milheiro et al., 2017a). In this context, Botezatu and
Pickering (2015) investigated the efficiency of a poly-lactic acid (PLA) biopolymer (sur-
face area 350 cm 2 /L) to remediate red wine with tainting odors. These odors arise from
elevated concentrations of isopropylmethoxypyrazine (IPMP), isobutylmethoxypyr-
azine (IBMP), 4-ethylphenol, and 4-ethylguaiacol. These off-flavors were reduced after
an 8 h treatment, IPMP (51%), IBMP (26%), 4-ethylphenol (2%), and 4-ethylguaiacol
(20%), with no or minimum effects on other volatiles profile according to gas chroma-
tography-mass spectrometry (GC-MS) analysis. In turn, Liang et al. (2018) evaluated
the practical usage of a putative imprinted magnetic polymer (PIMP) in winemaking
as a pre- and post-fermentative remedial treatment for elevated grape must IBMP con-
centration. Pre-fermentation addition of PIMP removed between 20 to 30% less IBMP
when compared to the post-fermentation addition, but also had a lesser effect on other
wine volatiles and color parameters. As mentioned above, the presence of 4-ethylphenol
and 4-ethylguaiacol is very detrimental to wine quality, since it contributes unpleas-
ant odors to wine, such as phenolic, animal, or horsey odors. In order to reduce its
negative sensory impact in red wines, Filipe-Ribeiro et al. (Filipe-Ribeiro, Cosme, and
Nunes, 2018a,b; Filipe-Ribeiro et al., 2017) tested chitins, chitosans, and activated car-
bon (AC) with different physicochemical characteristics. According to these authors,
chitosan with high deacetylation degrees, including fungal chitosan, which is already
 Off-Flavors in Alcoholic Beverages: An Overview 613

approved for use in wines, is an efficient method for reducing the negative sensory
impact of these off-flavors in red wines. Regarding AC, the superficial area and volume
of micropores are crucial to volatile phenol removal efficiency. Lisanti et al. (2017)
tested the efficiency of alternative fining agents, such as activated charcoal, polyvinyl
polypyrrolidone (PVPP), and zeolite to reduce the concentration of volatile phenols off-
flavors in red wines. The sensory outcome of AC and PVPP was a decrease in the inten-
sity of VPs off-odors, being more predominant with charcoal. Milheiro et al. (2017b)
performed a screening of eight fining agents (e.g., mineral, protein, and polysaccharide
based) for reducing the levels of these volatile phenols in red wines. The results obtained
by these authors showed that activated carbon was the most efficient fining agent in
removing 4-ethylphenol and 4-ethylguaiacol from red wines (57%) resulting in a 75%
decrease of headspace concentration of these off-flavors, whereas the lower reductions
were observed with egg albumin (19%) resulting in a 30% decrease in the headspace
concentration. Finally, Lisanti et al. (2014) evaluated the efficiency of seven treatments,
namely activated charcoal, bentonite, PVPP, yeast cell walls, potassium caseinate, zeo-
lite, and grape seed oil, in decreasing the concentration of geosmin, responsible for an
earthy off-flavor in wine. The sensory analysis confirmed the efficacy of the grape seed
oil as a remedial treatment since the concentration of geosmin decreased both in red
and in white wines by 83 and 81% of the initial concentration (360 ng/L), respectively.
All the effective treatments also caused a decrease of several aroma compounds, most
of them esters. So, the most efficient remedial procedure should be suitable to reduce or
remove off-flavors, without losses of positive aromatic compounds.

28.4 FUTURE OUTLOOK

Alcoholic beverages (e.g., wine and beer) have been the focus of exhaustive biotechno-
logical research in recent years to improve flavor and aroma. Regarding biotechnological
research, yeast genetic engineering approaches have been developed to improve the flavor of
alcoholic beverages. The main limitation of the application of strains under real winemak-
ing conditions is the absence of stability or cell vigor of these strains in industrial settings
(Carrau, Gaggero, and Aguilar, 2015). Other approaches or the generation of interspecific
wine yeast hydrides have positively improved wine flavor by reducing off-flavor production
and enhancing volatile thiols release in Saccharomyces (Dufour et al., 2013). In turn, the
increasing strain diversity will improve the aroma and flavors, which offer groundbreak-
ing opportunities in alcoholic beverages biotechnology (Carrau, Gaggero, and Aguilar,
2015). The application of self-cloning techniques and mutagenesis for improving flavor
has expanded, once it does not involve foreign DNA (Liu, 2015). An example of the appli-
cation of the self-cloning technique is the development of a self-cloning bottom-fermenting
yeast with high SSU1 expression. This constructed yeast contributes to the production of
greater quality beer with a high concentration of sulfite and low concentration of H 2S,
3-methyl-2-buten-1-thiol, and 2-mercapto-3-methyl-1-butanol when compared with the
parent strain (Iijima and Ogata, 2010).
On the other hand, preventive and remedial approaches have been proposed to
moderate or remove the presence of off-flavors in alcoholic beverages without causing a
negative impact on the global sensorial profile. Most of the proposed strategies are not
yet authorized by the OIV, and consequently are not allowed in winemaking (Milheiro
et al., 2017a). To bypass this situation, non-thermal approaches, such as high-pressure
614 Food Aroma Evolution

processing, pulsed electric field, ultraviolet C light have been assessed as able to remove
off-flavors in alcoholic beverages. Further research needs to be done to achieve more effi-
cient approaches for producing great quality alcoholic beverages.

REFERENCES

Alberts, Philippus, Maria A. Stander, Sylvia O. Paul, and Andre de Villiers. 2009. “Survey
of 3-Alkyl-2-Methoxypyrazine Content of South African Sauvignon Blanc Wines
Using a Novel LC−APCI-MS/MS Method.” Journal of Agricultural and Food
Chemistry 57 (20). American Chemical Society: 9347–55. doi:10.1021/jf9026475.
Allamy, Lucile, Philippe Darriet, and Alexandre Pons. 2017. “Identification and
Organoleptic Contribution of (Z)-1,5-Octadien-3-One to the Flavor of Vitis Vinifera
Cv. Merlot and Cabernet Sauvignon Musts.” doi:10.1021/acs.jafc.6b05293.
Andrés-Iglesias, Cristina, Jakub Nešpor, Marcel Karabín, Olimpio Montero, Carlos A.
Blanco, and Pavel Dostálek. 2016. “Comparison of Carbonyl Profiles from Czech
and Spanish Lagers: Traditional and Modern Technology.” LWT—Food Science and
Technology 66 (March). Academic Press: 390–7. doi:10.1016/J.LWT.2015.10.066.
Andrés-Iglesias, Cristina, Olimpio Montero, Daniel Sancho, and Carlos A. Blanco. 2015.
“New Trends in Beer Flavour Compound Analysis.” Journal of the Science of Food
and Agriculture 95 (8). John Wiley & Sons: 1571–6. doi:10.1002/jsfa.6905.
Baert, Jeroen J., Jessika De Clippeleer, Paul S. Hughes, Luc De Cooman, and Guido Aerts.
2012. “On the Origin of Free and Bound Staling Aldehydes in Beer.” Journal of
Agricultural and Food Chemistry 60 (46). American Chemical Society: 11449–72.
Ballester, Jordi, Mathilde Magne, Perrine Julien, Laurence Noret, Maria Nikolantonaki,
Christian Coelho, and Régis Gougeon. 2018. “Sensory Impact of Polyphenolic
Composition on the Oxidative Notes of Chardonnay Wines.” Beverages 4 (1): 19.
doi:10.3390/beverages4010019.
Blanco, Carlos A., Cristina Andrés-Iglesias, and Olimpio Montero. 2016. “Low-Alcohol
Beers: Flavor Compounds, Defects, and Improvement Strategies.” Critical Reviews
in Food Science and Nutrition 56 (8). Taylor & Francis: 1379–88. doi:10.1080/104
08398.2012.733979.
Botezatu, Andreea and Gary James Pickering. 2015. “Novel Applications for Biomaterials:
The Case of Remediation of Wine Taints Using Poly-Lactic Acid Polymer.” Applied
Mechanics and Materials 749 (April): 70–3. doi:1​0.402​8/www​.scie​ntifi​c.net​/AMM.​
749.7​0.
Boutou, Stéphane and Pascal Chatonnet. 2007. “Rapid Headspace Solid-Phase
Microextraction/Gas Chromatographic/Mass Spectrometric Assay for the
Quantitative Determination of Some of the Main Odorants Causing off-Flavours
in Wine.” Journal of Chromatography A 1141 (1). Elsevier: 1–9. doi:10.1016/J.
CHROMA.2006.11.106.
Buron, Nicolas, Monika Coton, Patrick Legendre, Jérôme Ledauphin, Valérie Kientz-
Bouchart, Hugues Guichard, Daniel Barillier, and Emmanuel Coton. 2012.
“Implications of Lactobacillus Collinoides and Brettanomyces/Dekkera Anomala in
Phenolic off-Flavour Defects of Ciders.” International Journal of Food Microbiology
153 (1–2). Elsevier: 159–65.
Buron, Nicolas, Hugues Guichard, Emmanuel Coton, Jérôme Ledauphin, and Daniel
Barillier. 2011. “Evidence of 4-Ethylcatechol as One of the Main Phenolic off-Fla-
vour Markers in French Ciders.” Food Chemistry 125 (2). Elsevier: 542–8.
 Off-Flavors in Alcoholic Beverages: An Overview 615

Callejón, R.M., C. Ubeda, R. Ríos-Reina, M.L. Morales, and A.M. Troncoso. 2016.
“Recent Developments in the Analysis of Musty Odour Compounds in Water and
Wine: A Review.” Journal of Chromatography A 1428 (January). Elsevier: 72–85.
doi:10.1016/J.CHROMA.2015.09.008.
Câmara, José Sousa, José C. Marques, María A. Alves, and Antonio C. Silva Ferreira.
2004. “3-Hydroxy-4,5-Dimethyl-2(5H)-Furanone Levels in Fortified Madeira
Wines: Relationship to Sugar Content.” Journal of Agricultural and Food Chemistry
52 (22): 6765–9. doi:10.1021/jf049547d.
Campo, Eva, Vicente Ferreira, Ana Escudero, José C. Marqués, and Juan Cacho. 2006.
“Quantitative Gas Chromatography-Olfactometry and Chemical Quantitative
Study of the Aroma of Four Madeira Wines.” Analytica Chimica Acta 563: 180–7.
Capone, D.L., K.A. van Leeuwen, K.H. Pardon, M.A. Daniel, G.M. Elsey, A.D. Coulter,
and M.A. Sefton. 2010. “Identification and Analysis of 2-Chloro-6-Methylphenol,
2,6-Dichlorophenol and Indole: Causes of Taints and Off-Flavours in Wines.”
Australian Journal of Grape and Wine Research 16 (1): 210–7.
Carrau, Francisco, Carina Gaggero, and Pablo S. Aguilar. 2015. “Yeast Diversity and
Native Vigor for Flavor Phenotypes.” Trends in Biotechnology 33 (3). Elsevier
Current Trends: 148–54. doi:10.1016/J.TIBTECH.2014.12.009.
Cheng, Guo, Ye Liu, Tai-Xin Yue, Zhen-Wen Zhang, Guo Cheng, Ye Liu, Tai-Xin
Yue, and Zhen-Wen Zhang. 2015. “Comparison between Aroma Compounds
in Wines from Four Vitis Vinifera Grape Varieties Grown in Different Shoot
Positions.” Food Science and Technology (Campinas) 35 (2). SBCTA: 237–46.
doi:10.1590/1678-457X.6438.
Cravero, Francesco, Vasileios Englezos, Fabrizio Torchio, Simone Giacosa, Susana Río
Segade, Vincenzo Gerbi, Kalliopi Rantsiou, Luca Rolle, and Luca Cocolin. 2016.
“Post-Harvest Control of Wine-Grape Mycobiota Using Electrolyzed Water.”
Innovative Food Science & Emerging Technologies 35 (June). Elsevier: 21–8.
doi:10.1016/J.IFSET.2016.03.010.
Cravero, Francesco, Vasileios Englezos, Kalliopi Rantsiou, Fabrizio Torchio, Simone
Giacosa, Susana Río Segade, Vincenzo Gerbi, Luca Rolle, and Luca Cocolin. 2018.
“Control of Brettanomyces Bruxellensis on Wine Grapes by Post-Harvest Treatments
with Electrolyzed Water, Ozonated Water and Gaseous Ozone.” Innovative Food
Science & Emerging Technologies 47 (June). Elsevier: 309–16. doi:10.1016/J.
IFSET.2018.03.017.
Cravero, Maria Carla, Federica Bonello, Maria Del Carmen, Pazo Alvarez, Christos
Tsolakis, and Daniela Borsa. 2015. “The Sensory Evaluation of 2,4,6-Trichloroanisole
in Wines.” Journal of the Institute of Brewing 121: 411–7.
Dufour, Matthieu, Adrien Zimmer, Cécile Thibon, and Philippe Marullo. 2013.
“Enhancement of Volatile Thiol Release of Saccharomyces cerevisiae Strains Using
Molecular Breeding.” Applied Microbiology and Biotechnology 97 (13). Springer-
Verlag: 5893–905. doi:10.1007/s00253-013-4739-7.
Durner, Dominik, Kathrin Diesler, Patricia Golombek, Lisa Kromm, Mario Stahl, Karlis
Briviba, Maren Scharfenberger-Schmeer, and Ulrich Fischer. 2017. “Inactivation
of Microorganisms by UV-Treatment of Must and Wine.” In Edited by Jean-
Marie Aurand. BIO Web of Conferences, Vol. 9 (July). EDP Sciences: pp. 02001.
doi:10.1051/bioconf/20170902001.
Fang, Lei, David Hallam, and Raúl Bermúdez. 2016. “Experimental Studies on Removal
of Airborne Haloanisoles by Non-Thermal Plasma Air Purifiers.” Energy and
Buildings 130 (October). Elsevier: 238–43. doi:10.1016/J.ENBUILD.2016.08.035.
616 Food Aroma Evolution

Ferreira, A.C. Silva, Jean-Christophe Barbe, and Alain Bertrand. 2003. “3-Hydroxy-4,5-
Dimethyl-2(5H)-Furanone: A Key Odorant of the Typical Aroma of Oxidative Aged
Port Wine.” American Chemical Society. doi:10.1021/JF0342932.
Filipe-Ribeiro, Luís, Fernanda Cosme, and Fernando M. Nunes. 2018a. “Reducing the
Negative Sensory Impact of Volatile Phenols in Red Wine with Different Chitosans:
Effect of Structure on Efficiency.” Food Chemistry 242 (March). Elsevier: 591–600.
doi:10.1016/J.FOODCHEM.2017.09.099.
Filipe-Ribeiro, Luís, Fernanda Cosme, and Fernando M. Nunes. 2018b. “Data on
Changes in Red Wine Phenolic Compounds and Headspace Aroma Compounds
after Treatment of Red Wines with Chitosans with Different Structures.” Data in
Brief 17 (April). Elsevier: 1201–17. doi:10.1016/J.DIB.2018.02.029.
Filipe-Ribeiro, Luís, Juliana Milheiro, Carlos C. Matos, Fernanda Cosme, and Fernando
M. Nunes. 2017. “Reduction of 4-Ethylphenol and 4-Ethylguaiacol in Red Wine
by Activated Carbons with Different Physicochemical Characteristics: Impact on
Wine Quality.” Food Chemistry 229 (August). Elsevier: 242–51. doi:10.1016/J.
FOODCHEM.2017.02.066.
Fontana, Ariel R. 2012. “Analytical Methods for Determination of Cork-Taint
Compounds in Wine.” TrAC Trends in Analytical Chemistry 37 (July). Elsevier:
135–47. doi:10.1016/J.TRAC.2012.03.012.
Fontana, Ariel R., and Rubén Bottini. 2016. “QuEChERS Method for the Determination
of 3-alkyl-2-methoxypyrazines in Wines by Gas Chromatography-Mass
Spectrometry.” Food Analytical Methods 9 (12). Springer US: 3352–9. doi:10.1007/
s12161-016-0532-4.
Fracassetti, Daniela and Ileana Vigentini. 2018. “Occurrence and Analysis of Sulfur
Compounds in Wine.” In Grapes and Wines—Advances in Production, Processing,
Analysis and Valorization. InTech.
Franc, Céline, Frank David, and Gilles de Revel. 2009. “Multi-Residue off-Flavour
Profiling in Wine Using Stir Bar Sorptive Extraction–Thermal Desorption–Gas
Chromatography–Mass Spectrometry.” Journal of Chromatography A 1216 (15).
Elsevier: 3318–27.
Grant-Preece, Paris, Celia Barril, Leigh M. Schmidtke, Geoffrey R. Scollary, and Andrew
C. Clark. 2017. “Light-Induced Changes in Bottled White Wine and Underlying
Photochemical Mechanisms.” Critical Reviews in Food Science and Nutrition 57
(4): 743–54. doi:10.1080/10408398.2014.919246.
Gros, Jacques and Sonia Collin. 2012. “Identification of a New Light-Struck off-Flavour
in ‘light-Stable’ Beers.” Cerevisia 37 (1): 10–4.
Guido, L.F., J.R. Carneiro, J.R. Santos, P.J. Almeida, J.A. Rodrigues, and A.A. Barros.
2004. “Simultaneous Determination of E-2-Nonenal and β-Damascenone in
Beer by Reversed-Phase Liquid Chromatography with UV Detection.” Journal of
Chromatography A 1032 (1–2). Elsevier: 17–22.
Herrero, Paula, María Pilar Sáenz-Navajas, José Miguel Avizcuri, Laura Culleré, Pedro Balda,
Elena C. Antón, Vicente Ferreira, and Ana Escudero. 2016. “Study of Chardonnay
and Sauvignon Blanc Wines from D.O.Ca Rioja (Spain) Aged in Different French Oak
Wood Barrels: Chemical and Aroma Quality Aspects.” Food Research International
89 (November). Elsevier: 227–36. doi:10.1016/J.FOODRES.2016.08.002.
Hill, Peter G. and Roger M. Smith. 2000. “Determination of Sulphur Compounds in
Beer Using Headspace Solid-Phase Microextraction and Gas Chromatographic
Analysis with Pulsed Flame Photometric Detection.” Journal of Chromatography A
872 (1–2). Elsevier: 203–13.
 Off-Flavors in Alcoholic Beverages: An Overview 617

Hirst, M.B. and C.L. Richter. 2016. “Review of Aroma Formation through Metabolic
Pathways of Saccharomyces cerevisiae in Beverage Fermentations.” American
Journal of Enology and Viticulture 67 (4). American Journal of Enology and
Viticulture: 361–70. doi:10.5344/ajev.2016.15098.
Iijima, K. and T. Ogata. 2010. “Construction and Evaluation of Self-Cloning Bottom-
Fermenting Yeast with High SSU1 Expression.” Journal of Applied Microbiology
109 (6): 1906–13. doi:10.1111/j.1365-2672.2010.04819.x.
Iizuka-Furukawa, Sachiko, Atsuko Isogai, Kazutaka Kusaka, Tsutomu Fujii, and
Yoshinori Wakai. 2017. “Identification of 4-Mercapto-4-Methylpentan-2-One
as the Characteristic Aroma of Sake Made from Low-Glutelin Rice.” Journal
of Bioscience and Bioengineering 123 (2). Elsevier: 209–15. doi:10.1016/J.
JBIOSC.2016.09.002.
Ioannidou, Maria D., George Samouris, and Dimitris S. Achilias. 2016. “Acetaldehyde
Contamination of Water, Alcoholic, and Non-Alcoholic Beverages Stored in Glass
or Plastic Bottles.” Toxicological & Environmental Chemistry 98 (10). Taylor &
Francis: 1183–90. doi:10.1080/02772248.2015.1115505.
Kheir, Joyce, Dominique Salameh, Pierre Strehaiano, Cédric Brandam, and Roger
Lteif. 2013. “Open Archive Toulouse Archive Ouverte (OATAO) Impact of
Volatile Phenols and Their Precursors on Wine Quality and Control Measures of
Brettanomyces/Dekkera Yeasts.” European Food Research and Technology 237
(5): 655–71.
Kishimoto, Toru, Shigekuni Noba, Nana Yako, Minoru Kobayashi, and Tetsuya
Watanabe. 2018. “Simulation of Pilsner-Type Beer Aroma Using 76 Odor-Active
Compounds.” Journal of Bioscience and Bioengineering 126 (3). Elsevier: 330–8.
doi:10.1016/J.JBIOSC.2018.03.015.
König, T., B. Gutsche, M. Hartl, R. Hübscher, and P. Schreier, and W. Schwab. 1999.
“3-Hydroxy-4,5-Dimethyl-2(5H)-Furanone (Sotolon) Causing an Off-Flavor:
Elucidation of Its Formation Pathways during Storage of Citrus Soft Drinks.”
American Chemical Society. doi:10.1021/JF981244U.
Liang, Chen, Renata Ristic, Vladimir Jiranek, and David W. Jeffery. 2018. “Chemical
and Sensory Evaluation of Magnetic Polymers as a Remedial Treatment for Elevated
Concentrations of 3-Isobutyl-2-Methoxypyrazine in Cabernet Sauvignon Grape
Must and Wine.” Journal of Agricultural and Food Chemistry 66 (27). American
Chemical Society: 7121–30. doi:10.1021/acs.jafc.8b01397.
Lisanti, Maria Tiziana, Angelita Gambuti, Alessandro Genovese, Paola Piombino, and
Luigi Moio. 2014. “Earthy off-Flavour in Wine: Evaluation of Remedial Treatments
for Geosmin Contamination.” Food Chemistry 154 (July). Elsevier: 171–8.
doi:10.1016/J.FOODCHEM.2013.12.100.
Lisanti, Maria Tiziana, Angelita Gambuti, Alessandro Genovese, Paola Piombino, and
Luigi Moio. 2017. “Treatment by Fining Agents of Red Wine Affected by Phenolic
off-Odour.” European Food Research and Technology 243 (3). Springer Berlin
Heidelberg: 501–10. doi:10.1007/s00217-016-2763-4.
Liu, Chunfeng, Qi Li, Chengtuo Niu, Feiyun Zheng, and Yun Zhao. 2018. “Simultaneous
Determination of Diethylacetal and Acetaldehyde during Beer Fermentation and
Storage Process.” Journal of the Science of Food and Agriculture 98 (12): 4733–41.
doi:10.1002/jsfa.9008.
Liu, S.-Q. 2015. “Impact of Yeast and Bacteria on Beer Appearance and Flavour.”
Brewing Microbiology (January). Woodhead Publishing: 357–74. doi:10.1016/
B978-1-78242-331-7.00017-4.
618 Food Aroma Evolution

Lopez Pinar, Angela, Rahil Ghadiriasli, Philippe Darriet, and Andrea Buettner.
2017. “Unexpected Impact of 2-Methylisoborneol as off-Odour Substance in
Aged Wines.” Food Chemistry 220 (April). Elsevier: 498–504. doi:10.1016/J.
FOODCHEM.2016.10.021.
Lustrato, Giuseppe, Gabriele Alfano, Antonella De Leonardis, Vincenzo Macciola, and
Giancarlo Ranalli. 2015. “Inactivation of Dekkera Bruxellensis Yeasts in Wine
Storage in Brand New Oak Barrels Using Low Electric Current Technology.”
Annals of Microbiology 65 (4). Springer, Berlin Heidelberg: 2091–8. doi:10.1007/
s13213-015-1047-8.
Ma, Chengye, Yuanyuan He, Yanfei Cao, Xingda Bai, and Hongjun Li. 2016.
“Analysis of Flavour Compounds in Beer with Extruded Sorghum as an Adjunct
Using Headspace Solid-Phase Micro-Extraction and Gas Chromatography-Mass
Spectrometry.” Journal of the Institute of Brewing 122 (2). John Wiley & Sons:
251–60. doi:10.1002/jib.330.
Maslov, Luna, Ivana Tomaz, Marin Mihaljević Žulj, and Ana Jeromel. 2017. “Aroma
Characterization of Predicate Wines from Croatia.” European Food Research
and Technology 243 (2). Springer, Berlin Heidelberg: 263–74. doi:10.1007/
s00217-016-2741-x.
Mayr, Christine M., Dimitra L. Capone, Kevin H. Pardon, Cory A. Black, Damian Pomeroy,
and I. Leigh Francis. 2015. “Quantitative Analysis by GC-MS/MS of 18 Aroma
Compounds Related to Oxidative Off-Flavor in Wines.” Journal of Agricultural and
Food Chemistry 63 (13): 3394–401. doi:10.1021/jf505803u.
Milani, Elham A. and Filipa V.M. Silva. 2016. “Nonthermal Pasteurization of Beer by
High Pressure Processing: Modelling the Inactivation of Saccharomyces cerevisiae
Ascospores in Different Alcohol Beers.” High Pressure Research 36 (4). Taylor &
Francis: 595–609. doi:10.1080/08957959.2016.1190354.
Milani, Elham A. and Filipa V.M. Silva. 2017. “Ultrasound Assisted Thermal
Pasteurization of Beers with Different Alcohol Levels: Inactivation of Saccharomyces
cerevisiae Ascospores.” Journal of Food Engineering 198 (April). Elsevier: 45–53.
doi:10.1016/J.JFOODENG.2016.11.015.
Milani, Elham A., John G. Ramsey, and Filipa V.M. Silva. 2016. “High Pressure
Processing and Thermosonication of Beer: Comparing the Energy Requirements and
Saccharomyces cerevisiae Ascospores Inactivation with Thermal Processing and
Modeling.” Journal of Food Engineering 181 (July). Elsevier: 35–41. doi:10.1016/J.
JFOODENG.2016.02.023.
Milani, Elham Alami, Sally Alkhafaji, and Filipa V.M. Silva. 2015. “Pulsed Electric Field
Continuous Pasteurization of Different Types of Beers.” Food Control 50 (April).
Elsevier: 223–9. doi:10.1016/J.FOODCONT.2014.08.033.
Milheiro, Juliana, Luís Filipe-Ribeiro, Alice Vilela, Fernanda Cosme, and Fernando
M. Nunes. 2017a. “4-Ethylphenol, 4-Ethylguaiacol and 4-Ethylcatechol in Red
Wines: Microbial Formation, Prevention, Remediation and Overview of Analytical
Approaches.” Critical Reviews in Food Science and Nutrition (December). Taylor
& Francis: 1–25. doi:10.1080/10408398.2017.1408563.
Milheiro, Juliana, Luís Filipe-Ribeiro, Fernanda Cosme, and Fernando M. Nunes.
2017b. “A Simple, Cheap and Reliable Method for Control of 4-Ethylphenol and
4-Ethylguaiacol in Red Wines. Screening of Fining Agents for Reducing Volatile
Phenols Levels in Red Wines.” Journal of Chromatography B 1041–2 (January):
183–90. doi:10.1016/j.jchromb.2016.10.036.
 Off-Flavors in Alcoholic Beverages: An Overview 619

Monforte, Ana Rita, Sara I.F.S. Martins, and Antonio C. Silva Ferreira. 2017. “Strecker
Aldehyde Formation in Wine: New Insights into the Role of Gallic Acid, Glucose,
and Metals in Phenylacetaldehyde Formation.” Journal of Agricultural and Food
Chemistry 66 (10): acs.jafc.7b00264. doi:10.1021/acs.jafc.7b00264.
Morata, Antonio, Iris Loira, Ricardo Vejarano, Carmen González, María Jesús Callejo,
and José Antonio Suárez-Lepe. 2017. “Emerging Preservation Technologies in
Grapes for Winemaking.” Trends in Food Science & Technology 67 (September).
Elsevier: 36–43. doi:10.1016/J.TIFS.2017.06.014.
Moreno, Jose A., Luis Zea, Lourdes Moyano, and Manuel Medina. 2005.
“Aroma Compounds as Markers of the Changes in Sherry Wines Subjected
to Biological Ageing.” Food Control 16 (4). Elsevier: 333–8. doi:10.1016/J.
FOODCONT.2004.03.013.
Neto, Paula Vieira, Silvia M. Rocha, and Armando J.D. Silvestre. 2007. “Simultaneous
Headspace Solid Phase Microextraction Analysis of off-Flavour Compounds from
Quercus Suber L. Cork.” Journal of the Science of Food and Agriculture 87 (4).
John Wiley & Sons: 632–40.
Noba, Shigekuni, Nana Yako, Hiroaki Sakai, Minoru Kobayashi, and Tetsuya Watanabe.
2018. “Identification of a Precursor of 2-Mercapto-3-Methyl-1-Butanol in Beer.”
Food Chemistry 255 (July). Elsevier: 282–9. http:​//www​.ncbi​.nlm.​nih.g​ov/pu​bmed/​
29571​478.
Noba, Shigekuni, Nana Yako, Minoru Kobayashi, Susumu Masuda, and Tetsuya
Watanabe. 2017. “Search for Compounds Contributing to Onion-like off-Flavor
in Beer and Investigation of the Cause of the Flavor.” Journal of Bioscience and
Bioengineering 124 (4). Elsevier: 419–24. doi:10.1016/J.JBIOSC.2017.05.009.
Noestheden, Matthew, Eric G. Dennis, and Wesley F. Zandberg. 2018. “Quantitating
Volatile Phenols in Cabernet Franc Berries and Wine after On-Vine Exposure to
Smoke from a Simulated Forest Fire.” Journal of Agricultural and Food Chemistry
66 (3). American Chemical Society: 695–703. doi:10.1021/acs.jafc.7b04946.
Ochando, Thomas, Jean-Roch Mouret, Anne Humbert-Goffard, Jean-Marie
Sablayrolles, and Vincent Farines. 2018. “Vicinal Diketones and Their Precursors in
Wine Alcoholic Fermentation: Quantification and Dynamics of Production.” Food
Research International 103. Elsevier: 192–9.
Özhan, Didehan, R. Ertan Anli, Nilufer Vural, and Mustafa Bayram. 2009. “Determination
of Chloroanisoles and Chlorophenols in Cork and Wine by Using HS-SPME and
GC-ECD Detection.” Journal of the Institute of Brewing 115: 71–7.
Paniagua-Martínez, Ingrid, Alejandra Ramírez-Martínez, Vinicio Serment-Moreno, Sueli
Rodrigues, and César Ozuna. 2018. “Non-Thermal Technologies as Alternative Methods
for Saccharomyces cerevisiae Inactivation in Liquid Media: A Review.” Food and
Bioprocess Technology 11 (3). Springer, US: 487–510. doi:10.1007/s11947-018-2066-9.
Patrignani, Francesca, Fabio Chinnici, Diana I. Serrazanetti, Pamela Vernocchi, Maurice
Ndagijimana, Claudio Riponi, and Rosalba Lanciotti. 2016. “Production of Volatile
and Sulfur Compounds by 10 Saccharomyces cerevisiae Strains Inoculated in
Trebbiano Must.” Frontiers in Microbiology 7 (March). Frontiers: 243. doi:10.3389/
fmicb.2016.00243.
Pereira, Vanda, João M. Leça, João M. Gaspar, Ana C. Pereira, and José C. Marques.
2018. “Rapid Determination of Sotolon in Fortified Wines Using a Miniaturized
Liquid-Liquid Extraction Followed by LC-MS/MS Analysis.” Journal of Analytical
Methods in Chemistry 2018 (December). Hindawi: 1–7. doi:10.1155/2018/4393040.
620 Food Aroma Evolution

Petropulos, Violeta Ivanova, Elena Bogeva, Trajče Stafilov, Marina Stefova, Barbara
Siegmund, Nicole Pabi, and Ernst Lankmayr. 2014. “Study of the Influence of
Maceration Time and Oenological Practices on the Aroma Profile of Vranec
Wines.” Food Chemistry 165 (December). Elsevier: 506–14. doi:10.1016/J.
FOODCHEM.2014.05.144.
Pizarro, C., N. Pérez-del-Notario, and J.M. González-Sáiz. 2010. “Optimisation of a
Simple and Reliable Method Based on Headspace Solid-Phase Microextraction for
the Determination of Volatile Phenols in Beer.” Journal of Chromatography A 1217
(39). Elsevier: 6013–21.
Ramos, Rui M., Luís M. Gonçalves, Vlastimil Vyskočil, and José A. Rodrigues. 2017.
“Voltammetric Determination of Trace Amounts of Diacetyl at a Mercury Meniscus
Modified Silver Solid Amalgam Electrode Following Gas-Diffusion Microextraction.”
Talanta 169 (July). Elsevier: 203–8. doi:10.1016/J.TALANTA.2017.03.077.
Renouf, Vincent and Aline Lonvaud-Funel. 2007. “Development of an Enrichment Medium
to Detect Dekkera/Brettanomyces Bruxellensis, a Spoilage Wine Yeast, on the Surface
of Grape Berries.” Microbiological Research 162 (2). Urban & Fischer: 154–67.
Ridgway, K., S.P.D. Lalljie, and R.M. Smith. 2009. “Food Additives and
Contaminants Analysis of Food Taints and Off-Flavours: A Review.”
doi:10.1080/19440040903296840.
Ridgway, K., S.P.D. Lalljie, and R.M. Smith. 2010. “Analysis of Food Taints and Off-
Flavours: A Review.” Food Additives & Contaminants: Part A 27 (2). Taylor &
Francis Group: 146–68. doi:10.1080/19440040903296840.
Rodrigues, João A., António S. Barros, Beatriz Carvalho, Tiago Brandão, Ana M. Gil,
and António C. Silva Ferreira. 2011. “Evaluation of Beer Deterioration by Gas
Chromatography–Mass Spectrometry/Multivariate Analysis: A Rapid Tool for
Assessing Beer Composition.” Journal of Chromatography A 1218 (7). Elsevier: 990–6.
Sadoughi, Navideh, Leigh M. Schmidtke, Guillaume Antalick, John W. Blackman, and
Christopher C. Steel. 2015. “Gas Chromatography–Mass Spectrometry Method
Optimized Using Response Surface Modeling for the Quantitation of Fungal Off-
Flavors in Grapes and Wine.” Journal of Agricultural and Food Chemistry 63 (11):
2877–85. doi:10.1021/jf505444r.
Saison, Daan, David P. De Schutter, Bregt Uyttenhove, Freddy R. Filip Delvaux, and
Freddy R. Filip Delvaux. 2009. “Contribution of Staling Compounds to the Aged
Flavour of Lager Beer by Studying Their Flavour Thresholds.” Food Chemistry 114
(4). Elsevier: 1206–15.
Sakamaki, Kensuke, Susumu Ishizaki, Yasutaka Ohkubo, Yoshiyuki Tateno, and Akira
Fujita. 2011. “Identification of the Medicinal Off-Flavor Compound Formed from
Ascorbic Acid and (E)-Hex-2-Enal.” Journal of Agricultural and Food Chemistry
59 (12): 6667–71. doi:10.1021/jf200846h.
Scholtes, Caroline, Sabrina Nizet, and Sonia Collin. 2015. “How Sotolon Can Impart a
Madeira Off-Flavor to Aged Beers.” Journal of Agricultural and Food Chemistry
63 (11): 2886–92.
Sheridan, Marlena K. and Ryan J. Elias. 2015. “Exogenous Acetaldehyde as a Tool for
Modulating Wine Color and Astringency during Fermentation.” Food Chemistry
177 (June): 17–22. doi:10.1016/j.foodchem.2014.12.077.
Sidhu, Davinder, Jensen Lund, Yorgos Kotseridis, and Cedric Saucier. 2015.
“Methoxypyrazine Analysis and Influence of Viticultural and Enological Procedures
on Their Levels in Grapes, Musts, and Wines.” Critical Reviews in Food Science
and Nutrition 55 (4). Taylor & Francis: 485–502.
 Off-Flavors in Alcoholic Beverages: An Overview 621

Silva, Maria A., Michel Julien, Michael Jourdes, and Pierre-Louis Teissedre. 2011.
“Impact of Closures on Wine Post-Bottling Development: A Review.” European
Food Research and Technology 233 (6). Springer-Verlag: 905–14.
Simpson, Robert F., Dimitra L. Capone, and Mark A. Sefton. 2004. “Isolation and
Identification of 2-Methoxy-3,5-Dimethylpyrazine, a Potent Musty Compound
from Wine Corks.” American Chemical Society. doi:10.1021/JF049484Z.
Slabizki, Petra, Charlotte Legrum, Reinhard Meusinger, and Hans-Georg Schmarr.
2014. “Characterization and Analysis of Structural Isomers of Dimethyl
Methoxypyrazines in Cork Stoppers and Ladybugs (Harmonia Axyridis and
Coccinella Septempunctata).” Analytical and Bioanalytical Chemistry 406 (25).
Springer Berlin Heidelberg: 6429–39. doi:10.1007/s00216-014-8049-4.
Slabizki, Petra and Hans-Georg Schmarr. 2013. “Analysis of Corky off-Flavour Compounds
at Ultra Trace Level with Multidimensional Gas Chromatography-Electron Capture
Detection.” Journal of Chromatography A 1271 (1). Elsevier: 181–4.
Slaghenaufi, Davide, Loris Tonidandel, Sergio Moser, Tomás Román Villegas, and
Roberto Larcher. 2017. “Rapid Analysis of 27 Volatile Sulfur Compounds in
Wine by Headspace Solid-Phase Microextraction Gas Chromatography Tandem
Mass Spectrometry.” Food Analytical Methods 10 (11). Springer US: 3706–15.
doi:10.1007/s12161-017-0930-2.
Smith, M.E., M.Z. Bekker, P.A. Smith, and E.N. Wilkes. 2015. “Sources of Volatile
Sulfur Compounds in Wine.” Australian Journal of Grape and Wine Research 21
(December). John Wiley & Sons (10.1111): 705–12. doi:10.1111/ajgw.12193.
Spedding, G. and T. Aiken. 2015. “Sensory Analysis as a Tool for Beer Quality
Assessment with an Emphasis on Its Use for Microbial Control in the Brewery.”
Brewing Microbiology (January). Woodhead Publishing: 375–404. doi:10.1016/
B978-1-78242-331-7.00018-6.
Styger, Gustav, Bernard Prior, and Florian F. Bauer. 2011. “Wine Flavor and Aroma.”
Journal of Industrial Microbiology & Biotechnology 38 (9). Springer-Verlag: 1145–
59. doi:10.1007/s10295-011-1018-4.
Swiegers, J.H., E.J. Bartowsky, P.A. Henschke, and I.S. Pretorius. 2005. “Yeast and
Bacterial Modulation of Wine Aroma and Flavour.” Australian Journal of Grape
and Wine Research 11 (2): 139–73.
Tan, Yongxi and Karl J. Siebert. 2004. “Quantitative Structure−Activity Relationship
Modeling of Alcohol, Ester, Aldehyde, and Ketone Flavor Thresholds in Beer from
Molecular Features.” American Chemical Society. doi:10.1021/JF035149J.
Tao, Yang, Da-Wen Sun, Adrian Górecki, Wioletta Błaszczak, Grzegorz Lamparski,
Ryszard Amarowicz, Józef Fornal, and Tomasz Jeliński. 2016. “A Preliminary
Study about the Influence of High Hydrostatic Pressure Processing in Parallel
with Oak Chip Maceration on the Physicochemical and Sensory Properties of a
Young Red Wine.” Food Chemistry 194 (March). Elsevier: 545–54. doi:10.1016/J.
FOODCHEM.2015.07.041.
Tian, Jiyuan. 2010. “Determination of Several Flavours in Beer with Headspace Sampling–
Gas Chromatography.” Food Chemistry 123 (4). Elsevier: 1318–21.
van Wyk, Sanelle and Filipa V.M. Silva. 2017. “High Pressure Inactivation of
Brettanomyces Bruxellensis in Red Wine.” Food Microbiology 63 (May). Academic
Press: 199–204. doi:10.1016/J.FM.2016.11.020.
van Wyk, Sanelle, Mohammed M. Farid, and Filipa V.M. Silva. 2018. “SO2, High
Pressure Processing and Pulsed Electric Field Treatments of Red Wine: Effect on
Sensory, Brettanomyces Inactivation and Other Quality Parameters during One
622 Food Aroma Evolution

Year Storage.” Innovative Food Science & Emerging Technologies 48 (August).

Elsevier: 204–11. doi:10.1016/J.IFSET.2018.06.016.
Vanderhaegen, Bart, Hedwig Neven, Kevin J. Verstrepen, Freddy R. Delvaux, Hubert
Verachtert, and Guy Derdelinckx. 2004. “Influence of the Brewing Process on
Furfuryl Ethyl Ether Formation during Beer Aging.” American Chemical Society.
doi:10.1021/JF0490854.
Vanderhaegen, Bart, Hedwig Neven, Stefan Coghe, Kevin J. Verstrepen, Hubert
Verachtert, and Guy Derdelinckx. 2003. “Evolution of Chemical and Sensory
Properties during Aging of Top-Fermented Beer.” American Chemical Society.
doi:10.1021/JF034631Z.
Vichi, Stefania, Nuria Cortés-Francisco, and Josep Caixach. 2015. “Analysis of Volatile
Thiols in Alcoholic Beverages by Simultaneous Derivatization/Extraction and
Liquid Chromatography–High Resolution Mass Spectrometry.” Food Chemistry
175. Elsevier: 401–8. doi:10.1016/J.FOODCHEM.2014.11.095.
Wang, Ji-Yu, Xin-Jie Wang, Xian Hui, Shui-Hong Hua, Heng Li, and Wen-Yun
Gao. 2017. “Determination of Diacetyl in Beer by a Precolumn Derivatization-
HPLC-UV Method Using 4-(2,​3 -Dim​ethyl​- 6-Qu​inoxa​linyl​)-1,2​-Benz​enedi​amine​
as a Derivatizing Reagent.” Journal of Agricultural and Food Chemistry 65 (12).
American Chemical Society: 2635–41. doi:10.1021/acs.jafc.7b00990.
Ye, Dong-Qing, Xiao-Tian Zheng, Xiao-Qing Xu, Yun-He Wang, Chang-Qing Duan,
and Yan-Lin Liu. 2016. “Evolutions of Volatile Sulfur Compounds of Cabernet
Sauvignon Wines during Aging in Different Oak Barrels.” Food Chemistry 202
(July). Elsevier: 236–46. doi:10.1016/J.FOODCHEM.2016.01.139.
Zhang, Yanqing, Shiru Jia, and Wujiu Zhang. 2012. “Predicting Acetic Acid Content in
the Final Beer Using Neural Networks and Support Vector Machine.” Journal of the
Institute of Brewing 118 (4). John Wiley & Sons: 361–7. doi:10.1002/jib.50.
Zhao, Pengtao, Jinxin Gao, Michael Qian, and Hua Li. 2017. “Characterization of
the Key Aroma Compounds in Chinese Syrah Wine by Gas Chromatography-
Olfactometry-Mass Spectrometry and Aroma Reconstitution Studies.” Molecules
22 (7): 1045. doi:10.3390/molecules22071045.
Zuriarrain, Andoni, Juan Zuriarrain, Ana Isabel Puertas, María Teresa Dueñas, and Iñaki
Berregi. 2015. “Quantitative Determination of Lactic and Acetic Acids in Cider by
1H NMR Spectrometry.” Food Control 52 (June). Elsevier: 49–53. doi:10.1016/J.
FOODCONT.2014.12.028.
 Section V
Influences on Flavor Perception
 Chapter 29
Interactions between the Food
Matrix and Aroma Compounds
in Relation to Perception
Elisabeth Guichard

CONTENTS

29.1 Introduction 626

29.2 Physicochemical Properties of Aroma Compounds Involved in the
Interactions 627
29.2.1 Thermodynamic and Kinetic Properties of Aroma Compounds in
Simple Systems 628
29.2.2 Molecular Properties of Aroma Compounds 628
29.3 Interactions between Aroma Compounds and Macromolecules in Simple
Systems 629
29.3.1 Interactions between Aroma Compounds and Proteins 630
29.3.1.1 Milk Proteins 630
29.3.1.2 Plant Proteins 631
29.3.2 Interactions between Aroma Compounds and Lipids 632
29.3.3 Interactions between Aroma Compounds and Carbohydrates 633
29.3.3.1 Simple Sugars: Mono- and Disaccharides 633
29.3.3.2 Oligo- and Polysaccharides 634
29.3.4 Interaction with Other Ingredients 636
29.3.4.1 Polyphenols 636
29.3.4.2 Maillard Melanoidins 636
29.3.4.3 Alcohol 637
29.3.4.4 Salts 637
29.4 Interactions between Aroma Compounds and Macromolecules in
Real Food, the Combined Effect of Different Nonvolatile Compounds 637
29.4.1 Combined Effect of Lipids and Proteins in Dairy Products 637
29.4.2 Combined Effects of Carbohydrates and Proteins 639
29.4.3 Carbohydrates as Fat Substitutes 639
29.4.4 Reformulation of Low-Sugar Products 640
29.4.5 Impact of the Structure and Texture 641
29.5 Conclusion and Future Trends 642
References 642

625
626 Food Aroma Evolution

29.1 INTRODUCTION

Food acceptability for the consumer is mainly driven by organoleptic properties and among
them flavor perception. Flavor can be defined as a multisensory perception involving olfac-
tory, gustatory, and trigeminal sensations during food consumption. Flavor perception results
from the integration at the central level of olfactory, gustatory, and trigeminal information
from the chemical compounds present in the food (Thomas-Danguin, 2009). Thus, flavor
perception is due to a wide range of stimuli: odorant compounds which activate the olfactory
receptors, taste compounds which activate the gustatory receptors, and trigeminal compounds
which activate the trigeminal nerve. In this chapter, we will only focus on the retention/
release mechanisms of odorant/aroma compounds as a function of food composition. Aroma
compounds activate olfactory receptors, via the orthonasal route, by smelling the food before
consumption, and via the retronasal route during the in-mouth process (Figure 29.1).
Orthonasal and retronasal olfaction differ in terms of odor quality and threshold,
and the two modes have different impacts on consumption behavior (Hummel and Seo,
2017). In order to be perceived at the receptor level, aroma compounds have first to be
released from the food matrix in the saliva during oral food processing, then transferred
into the gas phase in the oral cavity, and transported to the olfactory receptors in the
nasal cavity via the respiratory flow (Buettner et al., 2001, 2002, 2008).
Aroma compounds are small molecules with various chemical properties as described
in previous chapters of this book. Their release in the gas phase depends on their interac-
tions with the food matrix (Guichard, 2002). Food is a complex system containing mix-
tures of volatile compounds responsible for aroma perception and nonvolatile compounds
such as lipids, proteins, and carbohydrates which interact with aroma compounds (Seuvre
and Voilley, 2017) and thus impact their release during food consumption (Beauchamp
and Zardin, 2017). Food formulation answering both nutritional and environmental
requirements constitutes one of the main objectives for the food industry.

FIGURE 29.1 Transfer of aroma compounds from the food to the olfactory receptors
during food consumption.
 The Food Matrix and Aroma Compounds  627

Most developed and developing countries are confronted with a rising rate of
­utrition-related pathologies, notably obesity, cardiovascular diseases, and diabetes,
n
which are related to unbalanced diets characterized by excess consumption of fat, salt,
and sugar. In response to this situation, the food industry is making an effort to integrate
these nutritional criteria in the formulation of food products. However, these ingredients
play an important role in the structure of foods and also in their organoleptic properties,
mainly texture and flavor. Fat contributes to the texture, aroma, and mouthfeel percep-
tion of foods; it constitutes a solvent and a vector for aroma compounds and controls
their release (Druaux and Voilley, 1997). In dairy products, a decrease in fat content
needs the addition of fat substitutes or texturing agents to restore the texture and mouth-
feel, but these ingredients induce modifications in aroma release and aroma perception
(Lubbers et al., 2007). Salt and sugar are responsible for salty and sweetness perception
but also act on the ratio of aroma compounds present in the gas phase. Sugars also play
an important role in food texture, in the water activity, and shelf life. Their substitu-
tion by intensive sweeteners will result in modification of texture which can be restored
with the addition of other thickeners (soluble fibers, carbohydrates) and modifications of
aroma release and perception (King et al., 2006). Modification of a nonvolatile composi-
tion in a portion of food to answer nutritional recommendations (low salt, sugar, and fat
content) will not only modify the texture of the food but also the release of aroma com-
pounds by modifying the interactions between odorant compounds and macromolecules.
By 2050, the necessity of feeding more than 9 billion people will require an increase
in global protein production. This increase will only be possible by reformulating food
in more sustainable ways, for example, by means of a substantial substitution of animal
protein with plant protein. Indeed, the environmental impact of animal protein produc-
tion is particularly high compared to the production of plant protein. The substitution of
animal protein with plant protein would also be beneficial in terms of public health since
a high consumption of products containing animal protein is associated with increased
prevalence of diseases and mortality in the long term. There are only a few studies in the
literature comparing the effects of this substitution on the release of aroma compounds.
It is thus of interest to better understand the different mechanisms involved in the
retention/release behavior of aroma compounds in food according to its composition.
The aim of this review is to present the different types of interactions between aroma
compounds and nonvolatile ingredients present in the food matrix alone or in a mixture.
The first part will describe the main physicochemical properties of aroma compounds
involved in the interactions with food matrix ingredients, the second part will present the
different types of interactions in simple model systems, and the third part will look at the
interactions occurring in a real food system, which also take into account the interactions
between food ingredients and their impact on aroma release.

29.2 PHYSICOCHEMICAL PROPERTIES OF AROMA

COMPOUNDS INVOLVED IN THE INTERACTIONS

Aroma compounds are small molecules which are volatile at ambient temperatures
and able to reach the olfactory receptors so that they can be perceived. They are pres-
ent in very low quantities in food (less than 1%), have a low molecular weight (<400
Da), but present various chemical functions. They are able to interact with nonvolatile
compounds by covalent or non-covalent binding. In the case of irreversible covalent
binding, such as the formation of a Shiff base between aldehydes and proteins (Meynier
628 Food Aroma Evolution

et al., 2004), the aroma compounds cannot be released into the air and are thus not
able to reach the olfactory receptors. Different types of reversible non-covalent bind-
ing occur between aroma compounds and macromolecules including van der Waals
forces or dipole–dipole interactions, the hydrogen bond between electronegative atoms
and hydrophobic effects for the most apolar molecules (Seuvre and Voilley, 2017). The
nature and strength of the interaction depends on the nature of both the aroma com-
pound and the macromolecule.

29.2.1 Thermodynamic and Kinetic Properties of Aroma

Compounds in Simple Systems

The rate of aroma release from products is controlled by the volatility of the aroma
compounds in the product (thermodynamic factor) and the resistance to mass transfer
from product to air (kinetic factor) (Voilley, 2006). The retention/release of aroma com-
pounds by the food matrix can be measured with the air–matrix partition coefficients
(K) between the gas phase and the food matrix at thermodynamic equilibrium. These
coefficients are expressed as the proportion of the concentrations of aroma compounds
in the air and product phase under equilibrium conditions:

CG
K= (29.1)
CM

where CG is the concentration of the volatile compound in the gas phase and CM the
concentration of the volatile compound in the matrix. A partition coefficient (K) can be
expressed using the mass fraction (km), the molar fraction (Ki), or the molar concentra-
tion (ki). Therefore, in order to compare values obtained in different units, some conver-
sions are necessary, as detailed previously (Guichard, 2012).
As aroma release and perception are time-dependent phenomena, kinetic parameters
supply information that enables a clearer understanding of the behavior of volatile com-
pounds in food matrices. Diffusion is a spontaneous process by which matter is trans-
ported from one part of a system to another by random molecular movements, leading
to complete mixing. The molecules in solution move according to rotational and trans-
lational movements. Two main factors can impact the diffusion process: (i) obstacles or
entrapment effects due to the nature of macromolecules and their structural organiza-
tion, and (ii) the strength and nature of specific interactions (chemical or non-chemical
such as hydrogen bonding) between small solutes (including water molecules and ions)
and large food molecules (Tavel et al., 2008).

29.2.2 Molecular Properties of Aroma Compounds

Several physicochemical properties of aroma compounds are involved in their retention/

release from the food matrix, such as molecular dimension and shape, chemical function,
polarity, hydrophobicity, volatility, and diffusivity, as recently reviewed (Goubet et al.,
1998; Seuvre and Voilley, 2017).
The molecular dimension is a key parameter for explaining the inclusion of aroma
compounds in the amylose helix (Arvisenet et al., 2002) and in the cavity of cyclodex-
trins (Goubet et al., 2001). If the size of the aroma compound is too small in comparison
 The Food Matrix and Aroma Compounds  629

with the cavity, hydrophobic interactions dominate, but if the size is close to that of the
cavity, van der Waals interactions will predominate (Seuvre and Voilley, 2017). However
the shape of the guest molecule seems to also play an important role in the strength of
the binding as it will influence the contact area between the aroma and the polyoside
cavity, as demonstrated in the case of α-, β-, and γ-cyclodextrins (Inoue et al., 1993).
Considering the aroma compound, it was observed that molecular descriptors encod-
ing the relative negative charge area (Jurs-RNCS) and those describing the shape of the
aroma compound (PMI-Y, Shadow-XY, and RadOf-Gyration) were also involved in the
retention by ι–carrageenan: esters with a more globular form and with ramifications,
such as 3-methylbutyl acetate or 2-methylpropyl 3-methylbutanoate were less retained
than the linear ethyl esters (Chana et al., 2006).
The volatility of aroma compounds in solution is the ability of the aroma to be
volatilized in the gas phase. It can be estimated with the measurement of the air–water
partition coefficient, which is temperature-dependent according to the Vant’Hoff law.
A rise in temperature increases the air–water partition coefficients; this linear relation-
ship can then be used to predict values at other temperatures other than experimental
(Kopjar et al., 2010). This air–water partition coefficient depends on both the molecular
mass, physical properties such as enthalpy of vaporization, and also shape descriptors of
aroma compounds, such as molecular flexibility, electron properties, or charge distribu-
tion (Tromelin et al., 2010).
Aroma compounds are solubilized in the aqueous and/or lipid phase as a function of
their hydrophobicity (logP value) and/or polarity. Most aroma compounds are more sol-
uble in oil than in water (logP > 1) and thus considered as hydrophobic. In oil-containing
foods, in order to be perceived, aroma compounds need to be transferred from the lipid
phase to the water phase and then to the vapor phase, so the amount and nature of fat
exerts a strong influence on aroma release (de Roos, 1997). As many proteins and carbo-
hydrates possess a hydrophobic cavity, hydrophobic aroma compounds are thus able to
enter into this cavity and the strength of this hydrophobic effect increases, within a same
chemical class, with the hydrophobicity of the aroma compound, as has been shown for
β-lactoglobulin (Sostmann and Guichard, 1998a). Aroma compounds with logP values
below 1 are considered polar. The polarity of the aroma compounds is an important
parameter in the case of interactions with carbohydrates, as hydrogen bonding occurs
between polar groups such as hydroxyl or carboxyl groups (Boland et al., 2004; Lubbers
et al., 2007). Considering the molecular descriptors involved in binding with different
thickeners, it was observed that the elongated shape of the molecule and its capacity to
involve van der Waals interactions favored hydrophobic interactions. However, descrip-
tors encoding the charge distribution at the surface of the molecules were also involved
in the binding, suggesting that the retention/release properties of aroma compounds by
carbohydrates were more linked to specific chemical properties than to chemical class
(Lubbers et al., 2007).

29.3 INTERACTIONS BETWEEN AROMA COMPOUNDS

AND MACROMOLECULES IN SIMPLE SYSTEMS

Aroma compounds are able to bind to different food ingredients, which will decrease
their ability to be transferred from the liquid to the gas phase and to reach the olfactory
receptors in order to be perceived. Food is a complex system which contains different
types of macromolecules, which can be classified into proteins, lipids, and carbohydrates.
630 Food Aroma Evolution

This part will present the different types of interactions between aroma compounds and
macromolecules in simple model systems.

29.3.1 Interactions between Aroma Compounds and Proteins

Food proteins have little flavor of their own, but are known to bind and trap aroma com-
pounds, which reduces their release in the air phase (Tromelin et al., 2006a). Proteins
mainly interact with aroma compounds by means of molecular interactions, ionic bond-
ing, hydrogen bonding, and hydrophobic bonding (Lubbers et al., 1998; Guichard, 2012).
The nature and the strength of the interaction depend both on the protein structure
and on the molecular and physicochemical properties of the aroma compound. Globular
proteins are used as food ingredients for different applications such as gelation, thicken-
ing, emulsification, and foaming. The functional and binding properties of globular pro-
teins depend on their molecular structure under the environmental conditions. It is thus
important to consider not only the different types of proteins but also their structural
changes according to the medium or to the process.
Most of the studies related to aroma–protein interactions were realized with milk
proteins, and only a few with plant proteins.

29.3.1.1 Milk Proteins
The interactions between β-lactoglobulin, one of the main proteins found in dairy prod-
ucts, and aroma compounds have been widely studied (Lubbers et al., 1998; Seuvre et
al., 2001; Guichard, 2002; van Ruth and Villeneuve, 2002). For the same protein, the
strength of the interaction depends on the physicochemical properties of the aroma
compound. For instance, the strength of hydrophobic interactions with β-lactoglobulin
increases in line with the hydrophobicity (logP value) of the compounds (O’Neill and
Kinsella, 1987; Sostmann and Guichard, 1998b); for example, linear esters with a long
hydrophobic chain are more tightly retained in model systems with protein than with
short-chain esters (Jouenne and Crouzet, 2000a), and are thus less released in the gas
phase and less perceived (Andriot et al., 2000). This retention also induces a decrease
in diffusion coefficients (Jung et al., 2002) and mass transfer coefficients (Le Guen and
Vreeker, 2003). Indeed, three-dimensional quantitative structure activity relationship
(3-D QSAR) molecular modeling studies have shown the existence of two groups of
ligands, confirming the presence of at least two binding sites on β-lactoglobulin and put-
ting forward the role of hydrogen bonding (Tromelin and Guichard, 2003, 2004). The
binding within the central cavity occurs for aroma compounds that have an elongated
structure whereas the binding onto the protein surface in a site located between strand
β G, α helix, and strand β I of β-lactoglobulin occurs for aroma compounds that have or
adopt a compact structure (Tavel et al., 2008).
However, the temperature and ionic conditions of the matrix may also modify the
structure of the protein and thus the nature of the interaction (Boelrijk et al., 2006).
In response to a modification in the environment during processing (heating,
mechanical stress), β-lactoglobulin may shift from a “native” state to a “denatured” state,
which will change the nature of the interactions (O’Neill, 1996; Famelart et al., 2004).
As an example, interactions between two aroma compounds (β-ionone and guaiacol)
and β-lactoglobulin, adopting a molten globule state, were studied by two-dimensional
nuclear magnetic resonance (2-D NMR) spectroscopy. The less tightly packed structure
of the molten globule favored ligand binding, in particular within the central cavity.
 The Food Matrix and Aroma Compounds  631

The greater flexibility of the calyx entrance and the conformational change of loop EF
induced easier access to the central cavity after the thermal treatment (Tavel et al., 2010).
Modifications of protein conformation were also noticed when changing the pH. For
example, β-lactoglobulin exists as a monomer at pH 3 and forms a dimer at higher pH
values. Although the three-dimensional structure of β-lactoglobulin at an acidic pH is
very similar to that of a subunit within the dimer at higher pH, differences occur in the
orientation of two loops and of the flanking three-turn α-helix at the termini (Tromelin
et al., 2006b). Between pH 3 and pH 9, the flexibility of β-lactoglobulin was shown to
be modified, thus allowing improved accessibility to primary and secondary hydrophobic
binding sites and hence an increase in retention, whereas a reduction in retention was
observed between pH 9 and pH 11 due to the alkaline denaturation of protein (Jouenne
and Crouzet, 2000b).
Moreover, NaCl modifies the structure of proteins, as has been observed with
β-lactoglobulin which is mainly present in its dimeric form in the presence of NaCl
because of a modification of electrostatic forces (Sakurai et al., 2001). This modification
induced by NaCl results in an increase in the binding of octan-2-one by β-lactoglobulin
(Jouenne and Crouzet, 1997).
Interactions with other proteins (α-lactalbumin, bovine serum albumin, caseins) have
also been evidenced (Tromelin et al., 2006a,b; Ammari and Schroen, 2018). The retention
of aroma compounds by α-lactalbumin is generally lower than that by β-lactoglobulinin
(Jasinski and Kilara, 1985). Bovine serum albumin (BSA) is a globular non-glycoprotein,
which has been studied for its binding of fatty acids by hydrophobic interactions, but to
a lesser extent for its binding properties toward aroma compounds (Jasinski and Kilara,
1985). The binding affinity is influenced by the chain length, functional group, and struc-
tural state of the protein. Around 21 binding sites have been reported for this protein
of high molecular weight. BSA binds carbonyl compounds with a higher affinity than
the other milk proteins, inducing conformational changes to the protein. The structural
state of the binding sites determines how strongly the flavor will bind to the protein
(Damodaran and Kinsella, 1980).
Caseins, which are among the most abundant milk proteins, are unfolded proteins
but more flexible than the typical globular proteins. All caseins exhibit an amphipathic
character (Wong et al., 1996), they preferentially locate at hydrophobic interfaces but
are more hydrophilic than almost all globular proteins (Holt et al., 2013). There are two
fundamental functions of caseins: the effective transport of Ca 2+ and the self-associations
that lead to the colloid state (Farrell et al., 2002). The association of bovine casein with
small hydrophobic molecules involves an inclusion mechanism within the hydrophobic
interior of micelle-like protein associates. Both reversible and irreversible binding occurs
with caseins. Caseins interact with aldehydes by means of covalent and non-covalent
bonding (Fares et al., 1998), but other reversible binding was identified. The addition of
sodium caseinate in model wine induces a decrease in the activity coefficients of aroma
compounds in the following order: β-ionone > hexanol > ethyl hexanoate = isoamyl ace-
tate (Voilley et al., 1991). The authors explain the retention of hexanol by the formation
of hydrogen bonds and that of esters by hydrophobic effect. This retention effect was also
associated with a decrease in odor intensity.

29.3.1.2 Plant Proteins
Considering the increasing demand for plant proteins, proteins of leguminous plants are
one of the most promising materials for formulating new forms of food products. These
proteins are isolated from soy, peas, and beans and may be used as functional additives.
632 Food Aroma Evolution

They are composed of two main units, vicilin, and legumin. However, there are only
a few studies on their interactions with aroma compounds. It has been shown that the
native leguminous proteins possess a higher binding affinity than denaturated proteins
for hexyl acetate (Semenova et al., 2002). Denaturation of proteins by heat results in
intensive aggregation of basic protein chains and causes an increase in the number of
binding sites altogether with a decrease in the value of intrinsic affinity constant. Acid
denaturation (pH 3) dramatically alters the native protein structure, producing a loss of
hexyl acetate bound to the protein. Competitive binding of aroma compounds with 11S
glycinin (legumin class) suggested that some structures of aroma compounds are more
suitable than others for binding to the protein (Semenova et al., 2002). Soy protein is
mainly composed of β-conglycinin 7S (vicilin class) and glycinin 11S (legumin class), but
a preferential interaction with 2-nonanone was found with the 7S fraction (Damodaran
and Kinsella, 1980). It has also been shown that oxidation modifies the structural and
functional characteristics of the protein but also its binding properties. As an example,
the addition of malondialdehyde to soy protein induced, at low concentrations, a higher
binding of hydrophobic aroma compounds and, at higher concentration, protein denatur-
ation and a lower binding of hexanal and nonanal (Wang et al., 2018). Modifications of
the environmental conditions affect the interactions with soy proteins, and heat-induced
denaturation, which is the first step of tofu-making, is strongly affected by the presence
of salts (Shimizu et al., 2017).
Proteins are also major constituents in fruits and, for example, grape proteins, such
as thaumatin-like proteins (TLP), which are involved in haze formation during winemak-
ing, have been shown to interact with aroma compounds and mainly with esters (Di
Gaspero et al., 2017) with a higher binding activity observed for ethyl octanoate. This
binding of esters to TLC was responsible for the loss of perceived aroma after removing
haze formation in wines by using bentonite, due to the fact that esters mainly contribute
to the overall wine aroma.

29.3.2 Interactions between Aroma Compounds and Lipids

In food, lipids are among the ingredients that exert the most effect on the partitioning
of volatile compounds between the product and the vapor phase (de Roos, 2006). In fat-
containing foods, the retention/release behavior of aroma compounds mainly depends on
their solubility in the matrix. First, most of the aroma compounds are more soluble in oil
than in water, these compounds are considered as hydrophobic (logP > 1) and are thus
less released in the vapor phase from oil than from water (lower air–oil than air–water
partition coefficient). The less hydrophobic or more hydrophilic compounds (logP < 1),
such as diacetyl (or 2, 3-butanedione, logP = –1.34), present quite similar air–water and
air–oil partition coefficients (Guichard, 2002).
In systems containing lipids, aroma compounds are distributed between the lipid
and the aqueous phase following the physical laws of partition. However, the oil–water
partition coefficients of aroma compounds depend on the nature of the oil. For example,
in the case of hydrophobic compounds (ethyl hexanoate and 2-nonanone), sunflower oil–
water partition coefficients are higher than olive oil–water partition coefficients. This can
be explained by the higher amount of unsaturated fatty acids in sunflower oil (C18:2)
which leads to a higher solubility of aroma compounds (Guichard, 2002; Roudnitzky et
al., 2003). Aroma compounds are also more soluble in short-chain than in long-chain
saturated triglycerides (Maier, 1975). As aroma compounds are only soluble in liquid fat,
 The Food Matrix and Aroma Compounds  633

a high amount of solid fat reduces their solubility and thus increases their release in the
vapor phase (Roudnitzky et al., 2003).
The effect observed in model systems is the same in real foods. A decrease in the air–
matrix partition coefficient was observed after increasing the amount of lipids in emul-
sions; this effect increases in line with the hydrophobicity of aroma compounds (Doyen et
al., 2001). In model cheeses, an increase in the fat content was shown to induce a decrease
in both the air–matrix partition coefficient and the diffusion coefficients for heptan-
2-one and ethyl hexanoate, the most hydrophobic compounds (Lauverjat et al., 2009),
whereas no effect, or a reverse effect, was observed for diacetyl which is more soluble in
water than in oil (logP = –1.34). In yogurts, an increase in the fat level decreases both
the air–matrix partition coefficients (Nongonierma et al., 2006) and the diffusion coef-
ficients (Déléris et al., 2007) for the most hydrophobic compounds. In ice cream, it was
also observed that different fat types induce different aroma release behaviors. Fats with
a high solid fat content at the ice cream swallowing temperature (15°C) have a reduced
capacity to solubilize hydrophobic aroma compounds, which will increase their release
in the mouth in comparison with ice creams realized with liquid fat and thus increase
their perception (Relkin et al., 2004; Ayed et al., 2018). However, the effect of fat type on
aroma release and perception is lower than the effect of fat content.

29.3.3 Interactions between Aroma Compounds and Carbohydrates

Carbohydrate compounds are ubiquitous in food products and usually play a key role in
taste, especially sweetness and texture. Three categories of carbohydrates can be consid-
ered regarding their molecular weight: simple sugars such as glucose and sucrose (mono-
and disaccharides), oligosaccharides (2–10 monosaccharides) such as oligofructose and
polysaccharides (>10 monosaccharides) such as starch (Delarue and Giampaoli, 2000).
Their impact on aroma release is quite difficult to predict since they are able to induce
both retention and release effects, depending on the conditions and on the molecules. In
one way, they can induce small physicochemical (pH, ionic strength, water activity) and
textural changes in the matrix and thus indirectly modify aroma release. In another way,
carbohydrates can directly trap the aroma compounds and induce a retention effect. In
any way, sugar-induced effects are highly dependent on the intrinsic properties of the
food matrix such as the type of carbohydrates, their concentration (Nawar, 1971), and
also on the properties of aroma compounds such as steric hindrance, polarity, and rela-
tive volatility, as already reviewed (Goubet et al., 1998).

29.3.3.1 Simple Sugars: Mono- and Disaccharides

Among all simple sugars, sucrose is likely the most abundant. It occurs naturally as a
polymerization product of glucose and fructose by glycosidic linkage. In most cases,
the effect of sucrose on aroma release is dependent on the level of concentration added.
Indeed, a dynamic flavor release study from sucrose solutions showed that the rate-lim-
iting factor determining the initial flavor release was the hydration of sucrose which
depends on the molarity of sucrose (Rabe et al., 2003a). In simple model systems, the
addition of sucrose up to 40% induced only a slight increase in the headspace concentra-
tion of menthone and isoamyl acetate (Ebeler et al., 1988), whereas a significant increase
in the concentrations of acetone and 1-octanol in the vapor phase was observed with the
addition of increasing amounts of sugar (from 1 to 50%) (Voilley et al., 1977). In a soft
drink model system, an increase in the concentration of sucrose (from 20 to 60% w/w)
634 Food Aroma Evolution

was shown to significantly increase the release of tutti-frutti aroma compounds such
as esters and terpenes (Hansson et al., 2001). This phenomenon can be explained by
the so-called salting-out effect due to the hydration of the sucrose molecules by water,
which leads to a decrease in the water activity of the matrix. This usually tends to lower
the solubility of aroma compounds in water and thus to increase their concentration in
the gas phase. However, the addition of sucrose also makes the solvent character of the
solution more hydrophobic, which reduces or reverses the salting-out effect for the more
hydrophobic compounds. In viscous solutions, sucrose can also induce a retention effect
which may be explained with steric bulk linked with an increase in the viscosity or by the
existence of specific interactions between sucrose and aroma compounds (Roberts et al.,
1996). In aqueous solution with increasing amounts of sucrose, it was observed that the
release of aroma compounds with low retention times increases due to a modification of
the interactions between sugar molecules and water, whereas that of aroma compounds
with higher retention times decreases, due to an effect of increasing the medium viscosity,
which will affect the release of aroma compounds with higher molecular weight (Nahon
et al., 1998). In water solutions, an addition of 35% sucrose induced a decrease in self-
diffusion coefficients measured by DOSY NMR for ethyl butanoate, which was not asso-
ciated with differences in chemical shifts of aroma peaks in the different media, but only
to an increase in viscosity (Savary et al., 2006). Nahon et al. (2000) attempted to model
aroma release from aqueous solutions containing sucrose. At lower sucrose concentra-
tions, the partition coefficient primarily controlled aroma release, whereas, at higher
sucrose concentrations, the influence of the mass transfer coefficient was more important.

29.3.3.2 Oligo- and Polysaccharides

Polysaccharides are commonly used in food for their textural properties as thickeners.
Polysaccharides are also widely used as encapsulating agents to trap aroma compounds
and to prevent aroma loss during processing, for example, during thermal treatment. In
low-fat food, natural polysaccharides such as starch can be used as fat replacers (Jorgensen
et al., 2012). In its native state, starch is composed of a well-organized network of amylose
and amylopectin. Both are made up of 1,4-linked α-D-anhydroglucopyranoses, but they
differ in the degree of polymerization and ramification. The ratio between amylose and
amylopectin will determine the physical properties of starch. Amylose is an essentially
linear polymer, whereas amylopectin is much larger and is branched. In the presence of a
ligand, a conformational change occurs in the polymeric structure of amylose to complex
the ligand within the cavity of a single amylose helix (Immel and Lichtenthaler, 2000),
forming V-type amylose helices, as demonstrated for isopropanol and acetone (Buleon
et al., 1990). However, amylose is not able to form V-complexes with all aroma com-
pounds. Using X-ray analysis followed by differential scanning calorimetry (DSC), the
formation of V-type complex with amylose was verified for ethyl hexanoate and linalool
but not for isoamyl acetate (Arvisenet et al., 2002). In aqueous solution, hydrophobic
compounds such as ethyl hexanoate are less released in the presence of amylose whereas
the release in the air phase of isoamyl acetate measured by dynamic headspace analysis is
not affected by amylose (Langourieux and Crouzet, 1994). As shown in model systems,
amylose binds flavor compounds with the formation of inclusion complexes (Rutschmann
and Solms, 1990a,b,c; Jouquand et al., 2006). In the case of lactone, it has been demon-
strated that inclusion complexes were formed with linear chains of more than five car-
bons (γ-nonalactone to γ-dodecalactone) whereas a poor complexity occurred with shorter
chains, such as γ-heptalactone (Heinemann et al., 2001). The ability to form inclusion com-
plex can be used to stabilize labile aroma compounds in a specific matrix. For example, a
 The Food Matrix and Aroma Compounds  635

potent aroma compound from rice, 2-acetylpyridine (Maraval et al., 2008), was success-
fully stabilized after the formation of a V-complex with amylose (Hausch et al., 2018). The
type of complex between aroma and amylose can influence aroma release and thus aroma
perception (Heinemann et al., 2005; Pozo-Bayon et al., 2008). However, aroma retention
by starch is not always explained by complexation with amylose and could be due also
to interaction with amylopectin (Langourieux and Crouzet, 1994; Arvisenet et al., 2002)
resulting from adsorption involving hydrogen bonds.
Maltodextrins and cyclodextrins (CDs) are well-known agents obtained from the
degradation of starch. The interaction between these polysaccharides and aroma com-
pounds have been extensively studied (Kant et al., 2004; Jouquand et al., 2008). The most
common CDs are α-cyclodextrin (α-CD), β-cyclodextrin (β-CD), and γ-cyclodextrin (γ-
CD) composed of six, seven, and eight glucosyl units, respectively (Ciobanu et al., 2013).
CDs have a hydrophilic outer surface and a hydrophobic inner cavity able to trap the lipo-
philic compounds and to generate inclusion complex. A study of the interactions between
13 volatile aroma compounds and 6 CDs showed that stable 1:1 inclusion complexes are
formed in all cases. However, it appears that β-CDs are the most versatile CDs for the
studied compounds and showed the best complexation efficiency (Ciobanu et al., 2013).
Indeed, complexation is dependent both on the hydrophobicity of the aroma compound
and the molecular size and geometry of the cyclodextrin which will drive the strength of
the inclusion complex (Goubet et al., 1998; Astray et al., 2010). Extrinsic parameters,
such as temperature, can also impact the retention/release behavior in carbohydrate sys-
tems, and differ according to the polysaccharide studied. In a maltodextrin solution for
example, retention is favored by an increase of temperature from 60 to 80°C which sug-
gests an hydrophobic effect whereas no such temperature effect influenced the retention
by β-cyclodextrin due to the energetically favorable nonpolar–nonpolar interaction of
hydrophobic compounds with the inner cavity of β-cyclodextrin (Jouquand et al., 2004).
Pectin belongs to the family of complex polysaccharides composed of 1,4-linked α-D-
galactosyluronic acid. In nature, pectins occur in plant cells and are commonly used as
thickeners in food products. In gel model systems, both the rigidity of the gel and the nature
of hydrocolloid have a considerable effect on aroma release and perception (Boland et
al., 2006). These authors showed that when the rigidity of the gel increased (from 0.75 to
1% w/w of pectin), the air–gel partition coefficient decreased, and the flavor perception
decreased. However, in soft drink model systems, other authors showed that the addition of
pectin below 2.5% did not impact aroma release suggesting that viscosity had no effect on
aroma release (Hansson et al., 2001). In order to understand the retention/release behavior
in the pectin gels and to investigate the nature of the interactions involved, a statistical analy-
sis was performed on liquid–vapor partition coefficients of 51 aroma compounds (Ayed et
al., 2014). The retention/release balance of solvated hydrophobic molecules was explained
by the presence of electrostatic charges on the pectin, which can alter the network of water
molecules. Different release behaviors were observed as a function of the chemical func-
tionalities of aroma compounds and the structure of the carbon chains: alcohols exhibit
the highest retention whatever the matrix composition (pectin and/or starch) and within a
chemical class of aroma compounds the presence of a double bond induced higher retention.
Among the thickening agents, carrageenans, principally composed of highly sulfated,
alternating α(1→3) and β(1→4) linked galactase residues, are widely used in the food indus-
try, κ– and ι–carrageenan as gelling agents and λ–carrageenan as a thickening agent. The
impact of the addition of ι–carrageenan to water on aroma release is rather low (Chana
et al., 2006), whereas the addition of λ–carrageenan induced a significant decrease in
the air–liquid partition coefficient of the most hydrophobic compounds within a same
636 Food Aroma Evolution

chemical class (Bylaite et al., 2005). However, in the presence of salt (NaCl), a three-
dimensional network is formed between ι–carrageenan molecules leading to an increase
in gel hardness and a decrease in air–matrix partition coefficients (Juteau et al., 2004).

29.3.4 Interaction with Other Ingredients

29.3.4.1 Polyphenols
Other macromolecules which are able to bind aroma compounds are polyphenols, which
are important compounds in fruits and vegetables. Polyphenols contribute to the color,
astringency, and taste of such products. Even if the interactions between wine polyphenols
and proteins have been the subject of many studies (Siebert et al., 1996), their interactions
with aroma compounds are less understood. The elimination of polyphenols through
filtration or fining treatment and precipitation by polymerization during wine aging has
been suspected of producing flavor balance modifications. However, only a few studies
demonstrated the effect of monomeric phenols on the partitioning of odorants. The aque-
ous solubility of anisole, 2,3-diethylpyrazine, and ethyl benzoate increased in solutions
containing propyl gallate, chlorogenic acid, caffeine, or naringin (King and Solms, 1982).
Weak retention of different aroma compounds by catechin has also been observed by
measuring the dissociation constants (Kd) by 1H NMR (Dufour and Bayonove, 1999),
with the highest affinity for benzaldehyde (Kd = 0.187), the lowest for 3,5-dimethoxy-
phenol (Kd = 0.427), and intermediate for esters. A small retention of ethyl hexanoate
and hexanal by catechin, similar to that observed with β-lactoglobulin was also observed
(Jung and Ebeler, 2003); however, the retention of esters by catechin in model wine dif-
fered according to the volatile compounds and was higher for ethyl octanoate, which
was explained by the hydrophobic effect (Lorrain et al., 2013). Gallic acid significantly
decreased the volatility of 2-methylpyrazine, while naringin had less effect on the head-
space volatility of this aroma (Aronson and Ebeler, 2004). However, no effect of gallic
acid was observed on the release of esters from model wine (Lorrain et al., 2013). It seems
thus difficult to generalize the results obtained in simple systems due to the impact of
other ingredients on the interactions.

29.3.4.2 Maillard Melanoidins
Melanoidins are the nonvolatile compounds resulting from the non-enzymatic brown-
ing Maillard reaction and also from the reaction between ascorbic acid and amino acid.
They occur in most thermally treated food products. In a coffee beverage, for example,
they compose up to 30% of the nonvolatile fraction. Melanoidins are nitrogen polymers,
but their chemical structure is not yet fully elucidated. However, their interactions with
aroma compounds have been extensively studied (Ames, 1997; Charles-Bernard et al.,
2005; Hofmann and Schieberle, 2002; Lopez-Galilea et al., 2008; Obretenov et al., 2002).
Particularly, the interactions between melanoidins and thiols have been deeply studied due
to the typical sulfuric and roasted odor notes of thiols responsible for the typical aroma
of coffee. In a model system, melanoidins induced a significant decrease in the headspace
concentration of three important coffee odorants, 2-furfurylthiol (FFT), 3-methyl-2-bu-
tene-1-thiol, and 3-mercapto-3-methylbutyl formate (Hofmann et al., 2001). However,
aldehydes were not impacted by the addition of melanoidins in the system. Indeed, fur-
ther studies highlighted the existence of a covalent bond between melanoidins and thiols
through a nitrogen-containing intermediate (Hofmann and Schieberle, 2003). Another
study showed that interactions between coffee melanoidins and aroma compounds highly
 The Food Matrix and Aroma Compounds  637

depend on the method used for the extraction of melanoidins and on the properties of
aroma compounds (Andriot et al., 2004). These authors showed that melanoidins induced
the retention of methylketone and esters probably by means of hydrophobic interactions.
Moreover, it appeared that the lyophilization process, which is commonly used in order to
isolate melanoidins, modifies the retention effect suggesting denaturation of the polymers.

29.3.4.3 Alcohol
Ethanol represents the major volatile compound in alcoholic beverages and has been
shown to decrease the partition coefficient of various classes of volatile compounds by
increasing their solubility (Voilley et al., 1991). Thus, in systems which contain ethanol,
aroma release is changed, due partly to a modification in the air–liquid partition as a
function of aroma compound logP value, but also due to other physicochemical effects
such as micelle formation and surface tension effects. At a concentration of 12% ethanol,
the headspace concentration of aroma compounds was decreased as a function of their
hydrophobicity (Aznar et al., 2004).

29.3.4.4 Salts
The presence of salts (e.g., NaCl) induces an increase in the release of aroma compounds
due to a salting-out effect (Endo et al., 2012). Indeed, Na+ and Cl− ions are able to mobi-
lize water molecules for their hydration, so less water is available for the solubilization of
aroma compounds, which are therefore more released into the vapor phase with higher
air–matrix partition coefficients (Rabe et al., 2003b). However, the effect depends on
the nature and concentration of the salt. At a concentration of 0.5M, NaCl, and CaCl2
increased the release of ketones by a factor of 1.3, whereas the increase was up to 2.5 for
Na2SO4 (Wang and Arntfield, 2015). This salting-out effect is more marked with respect
to hydrophobic compounds, which are less soluble in water, and occurs not only in water
but also in model cheeses (Lauverjat et al., 2009).
In the presence of proteins, there is a combined effect of salts on aroma release and
on the protein structure. For example, the interactions between salt-extracted pea protein
isolates (PPIs) and ketones was modified in the presence of salts (Wang and Arntfield,
2015), varying in their nature and concentration (0.05 and 0.5M). A decrease in bind-
ing was observed with NaSCN at the highest concentration, a decrease was observed for
NaCl at the lowest concentration and an increase at the highest concentration, whereas
an increase with Na2SO4 was observed at both concentrations.

29.4 INTERACTIONS BETWEEN AROMA COMPOUNDS

AND MACROMOLECULES IN REAL FOOD, THE COMBINED
EFFECT OF DIFFERENT NONVOLATILE COMPOUNDS

In real food, aroma compounds are able to interact with the different nonvolatile compounds
present in the matrix (Figure 29.2), and their release in the air phase is the result of multiple
interactions, which are difficult to predict from single interactions in simple systems.

29.4.1 Combined Effect of Lipids and Proteins in Dairy Products

Dairy products are mainly composed of lipids and proteins, and the effect of these non-
volatile compounds is not the addition of the single effects. Lipids and proteins participate
638 Food Aroma Evolution

FIGURE 29.2 Different types of interactions occurring in real food.

in the structuration of the food and interact with each other. In dairy products, aroma
compounds are more strongly retained by caseins than by whey proteins (Saint-Eve et al.,
2006). In oil-in-water emulsions, sodium caseinate modifies the oil–water interface, induc-
ing a greater resistance to the mass transfer of aroma compounds, as has been observed
for ethyl esters (Landy et al., 1998). In fat-free dairy matrices, a high amount of caseins
increases the retention of limonene (logP = 4.3), while the less hydrophobic ethyl hexanoate
(logP = 2.4) and the hydrophilic diacetyl (logP = –1.3) are not affected (Heilig et al., 2011).
The effect of lipids is greater than that of protein. For example, adding just 0.5%
miglyol (triglyceride) to water induces a greater decrease in the volatility of nonan-2-one
than adding 3% protein (β-lactoglobulin) (Seuvre et al., 2001). The effect of protein also
depends on the fat content. In emulsions varying in fat and protein content, a greater reten-
tion was observed, at the highest protein content, for the more hydrophobic aroma com-
pounds, such as ethyl octanoate (logP = 3.8), whatever the fat level (3 and 9%), whereas
the effect of protein content on medium hydrophobic aroma compounds, such as butyl
propionate (logP = 2.3), was only significant at the lowest fat level (Ayed et al., 2018). For
the same amount of proteins, at a low-fat level, a small amount of proteins are located at
the oil–water interface. Thus, a greater amount of protein is present in the water phase and
able to interact with aroma compounds, explaining why not only the most hydrophobic
compounds, with higher binding activities, but also the less hydrophobic compounds, with
low binding activities, are bonded by proteins and less released in the gas phase.
The combined effects of salt, protein, and fat on aroma release have been studied in
model cheeses (Boisard et al., 2014). A reduction in fat content associated with a low salt
content induced a decrease in the rate of in vivo aroma release. The authors advanced the
hypothesis that model cheeses with a lower fat content and thus a higher protein content,
presented a thicker and stronger network with a more rigid microstructure, as was also
observed in cheddar cheeses (Bryant et al., 1995), which thus limited the diffusion of aroma
compounds in the network and thus the rate of release in the air phase. The effect of salt
content differed during the chewing process (Boisard et al., 2014). An increase in the salt
content triggered more rapid aroma release from the protein phase of the model cheese
to the oral cavity and hence to the nasal cavity, inducing a higher rate of aroma release.
Model cheeses with a higher salt content presented a larger droplet size which reduced the
transfer of hydrophobic aroma compounds from the fat to the aqueous phases; the release
of nonan-2-one, a more hydrophobic compound than ethyl butanoate therefore occurred
later after swallowing the cheese with higher salt content. This means that the direct effect
 The Food Matrix and Aroma Compounds  639

of cheese composition on in vivo aroma release is difficult to dissociate from the effect of
cheese microstructure and texture. In another study on model cheeses, it was observed that
the addition of salt modifies the texture by decreasing water activity, increasing firmness,
and thereby increasing the release of aroma compounds (Saint-Eve et al., 2009).

29.4.2 Combined Effects of Carbohydrates and Proteins

With the aim of protecting them from losses during processing, aroma compounds can
be encapsulated in bio-polymer based systems such as protein–polysaccharide complexes
(Santos et al., 2018). Complexes between proteins and biopolymers are formed mainly by
electrostatic interaction. They do not require a chemical reagent or elevated temperature.
For example, the release of 2-octanone bound by BSA/pectin complexes showed that weak
attractive interactions occurred between BSA and pectin at pH 6.4, leading to a preferential
binding by BSA while at pH 4.3, a strong interbiopolymer interaction occurred with bind-
ing on both polymers (Burova et al., 1999). This suggests a competitive binding of BSA and
2-octanone in the presence of pectin. Other combinations between proteins and polysaccha-
rides have already been used with success in food applications, including flavor encapsula-
tion (Weinbreck et al., 2004; Xiao et al., 2011; Koupantsis et al., 2014). Heat denaturated
β-lactoglobulin was found to form electrostatic complexes with pectin. These particles,
formulated with coacervates between β-lactoglobulin and different naturally occurring
polysaccharides, with mucoadhesive properties, such as alginate, pectin, Arabic, and acacia
gum, could serve as carriers for hydrophobic compounds in non-fat foods and clear bever-
ages and could also be used for masking unpleasant flavor in different types of formulations
(Diarrassouba et al., 2017). Soy proteins were used in a complex with a gum Arabic microen-
capsulation of sweet orange oil, and it was observed that the addition of sucrose significantly
increased the yield of encapsulation (Xiao et al., 2011). Complex coacervation between milk
protein and carboxymethylcellulose was used to encapsulate β-pinene (Koupantsis et al.,
2014), to protect this encapsulated aroma from losses and oxidation during food process.
Different types of beverages, such as wine or beer, contain both protein and polysaccha-
rides, which are able to interact with each other and thus to impact on aroma release differently
as when present alone. As an example, in beer realized with different levels of carbohydrates
and proteins, a decrease in the headspace concentration of myrcene, ethyl hexanoate, and
isoamyl acetate was observed with an increase in the concentration of both protein and car-
bohydrate (Castro and Ross, 2013). This decrease also induced a decrease in the associated
perceived aroma. The combined effects of proteins, polysaccharides, glycerol, and ethanol,
were also observed in wine, resulting in different sensory perceptions, which were explained
by the modulation of the retention/release behavior of both wine proteins and polysaccharides
as a function of the medium (Jones et al., 2008). In model wines realized with different con-
centrations of ethanol, tannins, and fructose, the higher retention of aroma compounds was
observed at the highest concentration of these different ingredients and tannins increased the
release of aroma compounds at the lowest ethanol concentrations (8–12%), whereas ethanol
and to a lesser extent fructose showed a retention effect (Villamor et al., 2013).

29.4.3 Carbohydrates as Fat Substitutes

Carbohydrates are often added to food as a thickening agent and more specifically in
low-fat products in order to restore the mouthfeel sensation highly appreciated in fat
640 Food Aroma Evolution

products. Even if the main effect of thickener agents is usually attributed to a modi-
fication of the viscosity, they also often induce a significant decrease in perceived fla-
vor above the critical concentration (c*) at which the carbohydrate network begins
(Godshall, 1997). In yogurts, the addition of low methoxyl pectin reduced aroma con-
centration in the headspace, but this effect was not significant (Lubbers and Guichard,
2003; Decourcelle et al., 2004). On the contrary, in fat-free dairy models, it has been
shown that an increase in pectin concentrations in gel from 0.04 to 0.15% caused a sig-
nificant increase in aroma release (Lubbers et al., 2007). These discrepant results empha-
size once again the importance of the matrix composition and the properties of aroma
compounds on aroma release.

29.4.4 Reformulation of Low-Sugar Products

With the aim to reduce sugar content in foods, there is a need to better understand the
interactions between the different molecules used as sweeteners and aroma compounds
in various sweet products and their effect on flavor perception.
Among monosaccharides, fructose and glucose are reducing sugars commonly used
as food ingredients themselves but also occur naturally from the degradation of sucrose.
Lactose is the main carbohydrate of milk and thus widely present in dairy products. In
coffee beverages, Piccone et al. (Piccone et al., 2012) showed that the addition of the
same sugar in different matrices containing no sugar (ready-to-drink or Espresso coffee)
could induce different effects on aroma release. In espresso coffee beverages, the addi-
tion of sucrose, fructose, or lactose caused a significantly higher release of some furanic
compounds and a lower release of pyrazines. No significant effect was observed between
the different sugars. On the contrary, in ready-to-drink beverages, the presence of sugars
induced either no change or a retention effect depending on the sugar type. Lactose for
example was the sugar that mostly caused high retention. However, while a significantly
higher release could be explained by a salting-out effect, the authors did not explain the
retention effect. In this case, the effects could also be the result of interactions between
other nonvolatile compounds and carbohydrates. For example, the nonvolatile matrix of
coffee contains up to 30% of brown polymers called melanoidins known to interact with
aroma compounds, as previously reviewed.
In solid systems such as cereal bars, the addition of polydextrose has been demon-
strated to favor the release of acetaldehyde, esters, menthol, and menthone, which was
attributed to a salting-out effect due to the reduction of free water (Heenan et al., 2012).
In sugar confectionery, glucose syrup and corn syrup are commonly used as sweet-
eners. They are derived from the hydrolysis of starch and contain a complex mixture of
mono- and polysaccharides such as polydextrose. The dextrose equivalent (DE) is usually
measured to evaluate the amount of reducing sugars and thus the degree of hydrolysis. In
model fruit pastilles, the addition of corn syrups DE40 and DE60 tends to reduce aroma
release in comparison to simple glucose–sucrose mixtures (Lubbers and Guichard, 2003).
In accordance, in low concentration solutions, the highly volatile compounds (benzalde-
hyde, ethyl butanoate, and butyl isovalerate) are retained more extensively in the presence
of polydextrose compared to a pure water solution whereas an increase in the release of
the less volatile compound cis-3-hexen-1-ol is observed (Siefarth et al., 2011). Authors
assumed that the diffusion of the higher molecular weight volatile compounds could be
affected by the meshes of the carbohydrate network. In contrast, compounds with lower-
molecular weight have a greater ability to diffuse through the matrix.
 The Food Matrix and Aroma Compounds  641

The interest for artificial sweeteners has been growing for years, which has led the
food industry to reformulate their products and thus scientists to study their impact on
aroma release in different media. However, as seen previously for carbohydrates, the
impact of artificial sweeteners is not yet predictable. In water, no effect of increasing
amounts of sodium cyclamate was observed on the release of a mixture of aroma com-
pounds from different chemical classes and molecular weights (Nahon et al., 1998). In
fat-free yogurts, an in vitro study has shown that the addition of aspartame and potas-
sium acesulfame did not change the aroma release behavior (Decourcelle et al., 2004).
However, other authors showed that during yogurt consumption, the release of ethyl
butanoate was decreased by about 20% in the presence of 400 ppm of aspartame (Mei
and Reineccius, 2007). The effects are also different depending on the food product.
In beverages, greater volatility of aroma compounds was found in products containing
aspartame or acesulfame K compared to sucrose, which was partly explained by their
lower content in soluble solids (King et al., 2006). In alcoholic beverages, the addition
of acesulfame K or sorbitol did not change the volatility of different aroma compounds
such as esters and terpenes, whereas the addition of aspartame decreased the volatility
of linalool and benzaldehyde (Da Porto et al., 2006). Authors attributed this effect to a
modification of the interactions between water and ethanol in the presence of aspartame.

29.4.5 Impact of the Structure and Texture

The effect of matrix structure and texture on the release properties of aroma compounds
has often been studied in model gels (Lubbers, 2006), although some groups have tried
to understand the effect of the texture and microstructure of dairy products on aroma
release (Gierczynski et al., 2011). In multiphase systems, the composition and nature of
proteins and lipids induce different microstructures which will exert different effects on
the retention and release properties of aroma compounds. It becomes thus difficult to
dissociate the effect of composition from the effect of texture. Different studies have tried
to demonstrate the effect of texture and/or microstructure at the same food composition.
By modifying the energy of emulsification, emulsions with different droplet sizes were
realized (Charles et al., 2000). With an increase in emulsion droplet size, an increase
in the release of hydrophilic compounds and a decrease in the release of hydrophobic
compounds was observed. This was explained by differences in aroma compounds mass
transfer due to the modification of the surface area and thus the exchanges between
the oil and water phase. In yogurts with the same protein concentration, a mechanical
treatment was applied to decrease the viscosity (Saint-Eve et al., 2006). It was observed
that this induced decrease in viscosity resulted in an increase in aroma release and thus
in the intensity of aroma perception. Moreover, in the same study, the nature of pro-
tein induced differences in the microstructure affecting aroma release parameters, and
caseinate enriched yogurts had a higher viscosity than whey protein–enriched yogurts
which decreased the release of aroma compounds in the air phase. In model cheeses, a
modification of the protein structure by acidification with chymosin was shown to induce
an increase in viscosity and as a consequence a greater and faster release of all aroma
compounds (Gierczynski et al., 2007). A similar observation was made with real cheeses
varying in firmness (Repoux et al., 2012). The release rate of ethyl propanoate and nonan-
2-one during the in-mouth breakdown increased with the firmness of the cheeses, due to
a greater amount of particles formed at the beginning of the chewing process in the case
of firm cheeses, which lead to an increase in the exchange area. After swallowing, the
642 Food Aroma Evolution

release of the most hydrophobic compound, nonan-2-one decreased more slowly than
that of ethyl propanoate due to its greater solubility in the lipid film remaining in the
mouth, leading to a higher in-mouth remanence and thus a longer sensory persistence
(Guichard et al., 2017).
NaCl also modifies the structure of the food matrix; for example, a higher NaCl con-
tent increases the strength of ι–carrageenan gels and the self-diffusion of ethyl butanoate
(Gostan et al., 2004), which can be explained by the greater size of the open space located
between the gel chains, resulting in fewer obstacles to the free diffusion of solutes.
Another interesting combined effect is that of phenolic compounds used in olive oil
emulsions stabilized by β-lactoglobulin (Genovese et al., 2015). It was also reported that
the addition of phenolic compounds modified the structure of the emulsion, improving
the dispersion degree, but also modified the final sensory perception of the product. The
authors thus demonstrated that the increase of aroma released by the addition of phenolic
compounds was due to interactions between polyphenols and whey proteins, which thus
reduced the binding between aroma and proteins by a competition effect. These results
are of great interest in the formulation of healthier foods with the addition of polyphenols
and providing a good aroma perception.

29.5 CONCLUSION AND FUTURE TRENDS

Food is a very complex system composed of different ingredients which impact its tex-
ture and also the release of the aroma compounds responsible for aroma perception.
These ingredients interact with each other in different ways, as described in this chapter.
However, it seems very difficult to be able to predict aroma release from a complex food
matrix from the individual molecular interactions between the aroma compounds pres-
ent in the food and the macromolecules. However, some general trends exist between the
effects observed in simple model systems and real food. A better understanding of the
different types of molecular interaction in simple and more complex systems could help
in the formulation of new food products answering nutritional guidelines, by substituting
traditional ingredients with healthier or more sustainable ingredients.

REFERENCES

Ammari, A. and K. Schroen (2018). “Flavor retention and release from beverages: A
kinetic and thermodynamic perspective.” Journal of Agricultural and Food
Chemistry 66(38): 9869–81.
Andriot, I., M. Harrison, N. Fournier, and E. Guichard (2000). “Interactions between
methyl ketones and beta-lactoglobulin: Sensory analysis, headspace analysis, and
mathematical modeling.” Journal of Agricultural and Food Chemistry 48(9):
4246–51.
Aronson, J. and S. E. Ebeler (2004). “Effect of Polyphenol compounds on the headspace
volatility of flavors.” American Journal of Enology and Viticulture 55(1): 13–21.
Arvisenet, G., P. Le Bail, A. Voilley, and N. Cayot (2002). “Influence of physicochemical
interactions between amylose and aroma compounds on the retention of aroma in
food-like matrices.” Journal of Agricultural and Food Chemistry 50(24): 7088–93.
 The Food Matrix and Aroma Compounds  643

Astray, G., J. C. Mejuto, J. Morales, R. Rial-Otero, and J. Simal-Gandara (2010).

“Factors controlling flavors binding constants to cyclodextrins and their applica-
tions in foods.” Food Research International 43(4): 1212–8.
Ayed, C., S. Lubbers, I. Andriot, Y. Merabtine, E. Guichard, and A. Tromelin (2014).
“Impact of structural features of odorant molecules on their retention/release behav-
iours in dairy and pectin gels.” Food Research International 62: 846–59.
Ayed, C., S. Martins, A.-M. Williamson, and E. Guichard (2018). “Understanding fat,
proteins and saliva impact on aroma release from flavoured ice creams.” Food
Chemistry 267(Special Issue): 132–9.
Aznar, M., M. Tsachaki, R. S. T. Linforth, V. Ferreira, and A. J. Taylor (2004). “Headspace
analysis of volatile organic compounds from ethanolic systems by direct APCI-MS.”
International Journal of Mass Spectrometry 239(1): 17–25.
Beauchamp, J. and E. Zardin (2017). “Odorant detection by on-line chemical ionization
mass spectrometry.” In Handbook of Odor. A. Buettner (Ed.), Springer: 355–408.
Boelrijk, A. E. M., G. Smit, K. G. C. Weel, and J. J. Burger (2006). “Flavour release
from liquid food products.” In Flavour in Food. A. Voilley and P. Etiévant (Eds.),
Cambridge, CB1 6AH, UK, Woodhead Publishing Limited and CRC Press LLC.
Part 3: 260–84.
Boisard, L., I. Andriot, C. Martin, C. Septier, V. Boissard, C. Salles, and E. Guichard
(2014). “The salt and lipid composition of model cheeses modifies in-mouth flavour
release and perception related to the free sodium ion content.” Food Chemistry
145(2014): 437–44.
Boland, A. B., C. M. Delahunty, and S. M. van Ruth (2006). “Influence of the texture
of gelatin gels and pectin gels on strawberry flavour release and perception.” Food
Chemistry 96(3): 452–60.
Boland, A. B., K. Buhr, P. Giannouli, and S. M. van Ruth (2004). “Influence of gelatin,
starch, pectin and artificial saliva on the release of 11 flavour compounds from
model gel systems.” Food Chemistry 86(3): 401–11.
Bryant, A., Z. Ustunol, and J. Steffe (1995). “Texture of cheddar cheese as influenced by
fat reduction.” Journal of Food Science 60(6): 1216.
Buettner, A., A. Beer, C. Hannig, and M. Settles (2001). “Observation of the swallowing
process by application of videofluoroscopy and real-time magnetic resonance imag-
ing-consequences for retronasal aroma stimulation.” Chemical Senses 26(9): 1211–9.
Buettner, A., A. Beer, C. Hannig, M. Settles, and P. Schieberle (2002). “Physiological and
analytical studies on flavor perception dynamics as induced by the eating and swal-
lowing process.” Food Quality and Preference 13(7–8): 497–504.
Buettner, A., S. Otto, A. Beer, M. Mestres, P. Schieberle, and T. Hummel (2008).
“Dynamics of retronasal aroma perception during consumption: Cross-linking on-
line breath analysis with medico-analytical tools to elucidate a complex process.”
Food Chemistry 108(4): 1234–46.
Buleon, A., M. M. Delage, J. Brisson, and H. Chanzy (1990). “Single-crystals of v-amy-
lose complexed with isopropanol and acetone.” International Journal of Biological
Macromolecules 12(1): 25–33.
Burova, T. V., N. V. Grinberg, I. A. Golubeva, A. Y. Mashkevich, V. Y. Grinberg, and V.
B. Tolstoguzov (1999). “Flavour release in model bovine serum albumin/pectin/2-
octanone systems.” Food Hydrocolloids 13(1): 7–14.
Bylaite, E., J. Adler-Nissen, and A. S. Meyer (2005). “Effect of xanthan on flavor release
from thickened viscous food model systems.” Journal of Agricultural and Food
Chemistry 53(9): 3577–83.
644 Food Aroma Evolution

Castro, L. F. and C. F. Ross (2013). “The effect of protein and carbohydrate levels on
the chemical and sensory properties of beer.” Journal of the American Society of
Brewing Chemists 71(4): 186–92.
Chana, A., A. Tromelin, I. Andriot, and E. Guichard (2006). “Flavor release from iota-
carrageenan matrix: A quantitative structure-property relationships approach.”
Journal of Agricultural and Food Chemistry 54(10): 3679–85.
Charles, M., V. Rosselin, L. Beck, F. Sauvageot, and E. Guichard (2000). “Flavor release
from salad dressings: Sensory and physicochemical approaches in relation with the
structure.” Journal of Agricultural and Food Chemistry 48(5): 1810–6.
Ciobanu, A., I. Mallard, D. Landy, G. Brabie, D. Nistor, and S. Fourmentin (2013).
“Retention of aroma compounds from Mentha piperita essential oil by cyclodex-
trins and crosslinked cyclodextrin polymers.” Food Chemistry 138(1): 291–7.
Da Porto, C., F. Cordaro, and N. Marcassa (2006). “Effects of carbohydrate and noncar-
bohydrate sweeteners on the orange spirit volatile compounds.” Lwt-Food Science
and Technology 39(2): 159–65.
Damodaran, S. and J. E. Kinsella (1980). “Flavor protein interactions. Binding of carbon-
yls to bovine serum albumin: Thermodynamic and conformational effects.” Journal
of Agricultural and Food Chemistry 28(3): 567–71.
de Roos, K. B. (1997). “How lipids influence food flavor.” Food Technology 51(1):
60–3.
de Roos, K. B. (2006). “How lipids influence flavor perception.” In Food Lipids:
Chemistry, Flavor, and Texture. F. Shahidi and H. Weenen (Eds.), Vol. 920, ACS:
145–58.
Decourcelle, N., S. Lubbers, N. Vallet, P. Rondeau, and E. Guichard (2004). “Effect
of thickeners and sweeteners on the release of blended aroma compounds in fat-
free stirred yoghurt during shear conditions (Journal Article).” International Dairy
Journal 14(9): 783–9.
Delarue, J. and P. Giampaoli (2000). “Study of interaction phenomena between aroma
compounds and carbohydrate matrixes by inverse gas chromatography.” Journal of
Agricultural and Food Chemistry 48(6): 2372–5.
Déléris, I., C. Lauverjat, I. C. Tréléa, and I. Souchon (2007). “Diffusion of aroma com-
pounds in stirred yogurts with different complex viscosities.” Journal of Agricultural
and Food Chemistry 55(21): 8681–7.
Di Gaspero, M., P. Ruzza, R. Hussain, S. Vincenzi, B. Biondi, D. Gazzola, G. Siligardi,
and A. Curioni (2017). “Spectroscopy reveals that ethyl esters interact with proteins
in wine.” Food Chemistry 217: 373–8.
Diarrassouba, F., G. Garrait, G. Remondetto, and M. Subirade (2017). “β-Lactoglobulin-
based nano and microparticulate systems for the protection and delivery of bioac-
tives.” Engineering Foods for Bioactives Stability and Delivery. Y. H. Roos and Y.
D. Livney (Eds.). New York, NY, Springer: 199–224.
Doyen, K., M. Carey, R. S. T. Linforth, M. Marin, and A. J. Taylor (2001). “Volatile
release from an emulsion: Headspace and In-Mouth studies.” Journal of Agricultural
and Food Chemistry 49(2): 804–10.
Druaux, C. and A. Voilley (1997). “Effect of food composition and microstructure
on volatile flavour release.” Trends in Food Science & Technology 8(November):
364–8.
Dufour, C. and C. L. Bayonove (1999). “Interactions between wine polyphenols and
aroma substances. An insight at the molecular level.” Journal of Agricultural and
Food Chemistry 47(2): 678–84.
 The Food Matrix and Aroma Compounds  645

Ebeler, S. E., R. M. Pangborn, and W. G. Jennings (1988). “Influence of dispersion

medium on aroma intensity and headspace concentration of menthone and isoamyl
acetate.” Journal of Agricultural and Food Chemistry 36(4): 791–6.
Endo, S., A. Pfennigsdorff, and K.-U. Goss (2012). “Salting-out effect in aqueous NaCl
solutions: Trends with size and polarity of solute molecules.” Environmental Science
& Technology 46(3): 1496–503.
Famelart, M. H., J. Tomazewski, M. Piot, and S. Pezennec (2004). “Comprehensive study
of acid gelation of heated milk with model protein systems.” International Dairy
Journal 14: 313–21.
Fares, K., P. Landy, R. Guilard, and A. Voilley (1998). “Physicochemical interactions
between aroma compounds and milk proteins: Effect of water and protein modifica-
tion.” Journal of Dairy Science 81(1): 82–91.
Genovese, A., N. Caporaso, L. De Luca, A. Paduano, and R. Sacchi (2015). “Influence of
olive oil phenolic compounds on headspace aroma release by interaction with whey
proteins.” Journal of Agricultural and Food Chemistry 63(15): 3838–50.
Gierczynski, I., E. Guichard, and H. Labouré (2011). “Aroma perception in dairy prod-
ucts: The roles of texture, aroma release and consumer physiology. A review.”
Flavour and Fragrance Journal 26(3): 141–52.
Gierczynski, I., H. Labouré, E. Sémon, and E. Guichard (2007). “Impact of hardness of
model cheese on aroma release: In vivo and in vitro study.” Journal of Agricultural
and Food Chemistry 55(8): 3066–73.
Gostan, T., C. Moreau, A. Juteau, E. Guichard, and M.-A. Delsuc (2004). “Measurement
of aroma compound self-diffusion in food models by DOSY.” Magnetic Resonance
in Chemistry 42(6): 496–9.
Goubet, I., C. Dahout, E. Sémon, E. Guichard, J. L. Le Quere, and A. Voilley (2001).
“Competitive binding of aroma compounds by beta-cyclodextrin.” Journal of
Agricultural and Food Chemistry 49(12): 5916–22.
Goubet, I., J.-L. Le Quere, and A. J. Voilley (1998). “Retention of aroma compounds by
carbohydrates; influence of their physicochemical characteristics and of their physi-
cal state. A review.” Journal of Agricultural and Food Chemistry 46(5): 1981–90.
Guichard, E. (2002). “Interactions between flavor compounds and food ingredients and
their influence on flavor perception.” Food Reviews International 18(1): 49–70.
Guichard, E. (2012). “Binding and release of flavor compounds.” In Food Flavors:
Chemical, Sensory and Technological Properties. H. Jelen (Ed.), Boca Raton, FL
(United States), CRC Press: 137–54.
Guichard, E., M. Repoux, E. M. Qannari, H. Labouré, and G. Feron (2017). “Model
cheese aroma perception is explained not only by in vivo aroma release but also
by salivary composition and oral processing parameters.” Food & Function 8(2):
615–28.
Hansson, A., J. Andersson, and A. Leufven (2001). “The effect of sugars and pectin
on flavour release from a soft drink-related model system.” Food Chemistry 72(3):
363–8.
Hausch, B. J., J. A. Little, J. A. Kenar, and K. R. Cadwallader (2018). “Starch–flavor
complexation applied to 2-acetyl-1-pyrroline.” Journal of Agricultural and Food
Chemistry 66(44): 11718–28.
Heenan, S., C. Soukoulis, P. Silcock, A. Fabris, E. Aprea, L. Cappellin, T. D. Mark, F.
Gasperi, and F. Biasioli (2012). “PTR-TOF-MS monitoring of in vitro and in vivo
flavour release in cereal bars with varying sugar composition.” Food Chemistry
131(2): 477–84.
646 Food Aroma Evolution

Heinemann, C., B. Conde-Petit, J. Nuessli, and F. Escher (2001). “Evidence of starch

inclusion complexation with lactones.” Journal of Agricultural and Food Chemistry
49(3): 1370–6.
Heinemann, C., M. Zinsli, A. Renggli, F. Escher, and B. Conde-Petit (2005). “Influence
of amylose-flavor complexation on build-up and breakdown of starch structures in
aqueous food model systems.” Lebensmittel-Wissenshaft und Technologie 38(8):
885–94.
Holt, C., J. A. Carver, H. Ecroyd, and D. C. Thorn (2013). “Invited review: Caseins and
the casein micelle: Their biological functions, structures, and behavior in foods.”
Journal of Dairy Science 96(10): 6127–46.
Hummel, T. and H.-S. Seo (2017). “Orthonasal and retronasal perception.” Flavour:
From Food to Perception. E. Guichard, C. Salles, M. Morzel, and A.-M. Le Bon
(Eds.). Chichester, UK, Wiley, J. & Sons: 310–8.
Immel, S. and F. W. Lichtenthaler (2000). “The hydrophobic topographies of amylose
and its blue iodine complex.” Starch-Starke 52(1): 1–8.
Inoue, Y., T. Hakushi, Y. Liu, L. H. Tong, B. J. Shen, and D. S. Jin (1993). “Thermodynamics
of molecular recognition by cyclodextrins .1. Calorimetric titration of inclusion com-
plexation of naphthalenesulfonates with alpha-cyclodextrin, beta-cyclodextrin, and
gamma-cyclodextrin – Enthalpy entropy compensation.” Journal of the American
Chemical Society 115(2): 475–81.
Jasinski, E. and A. Kilara (1985). “Flavor binding by whey proteins.” Milchwissenschaft
40(10): 596–9.
Jones, P. R., R. Gawel, I. L. Francis, and E. J. Waters (2008). “The influence of interac-
tions between major white wine components on the aroma, flavour and texture of
model white wine.” Food Quality and Preference 19(6): 596–607.
Jorgensen, A. D., S. L. Jensen, G. Ziegler, A. Pandeya, A. Buleon, B. Svensson, and A.
Blennow (2012). “Structural and physical effects of aroma compound binding to
native starch granules.” Starch-Starke 64(6): 461–9.
Jouenne, E. and J. Crouzet (1997). “Influence of sodium chloride and urea on volatile
compounds-β-lactoglobulin interactions.” In Interaction of Food Matrix with
Small Ligands Influencing Flavour and Texture, Gothenburg (Sweden), European
Communities.
Jouenne, E. and J. Crouzet (2000a). “Determination of apparent binding constants for
aroma compounds with beta-lactoglobulin by dynamic coupled column liquid chro-
matography.” Journal of Agricultural and Food Chemistry 48(11): 5396–400.
Jouenne, E. and J. Crouzet (2000b). “Effect of pH on retention of aroma compounds by
β-lactoglobulin.” Journal of Agricultural and Food Chemistry 48(4): 1273–7.
Jouquand, C., V. Ducruet, and P. Giampaoli (2004). “Partition coefficients of aroma
compounds in polysaccharide solutions by the phase ratio variation method.” Food
Chemistry 85(3): 467–74.
Jouquand, C., V. Ducruet, and P. Le Bail (2006). “Formation of amylose complexes with
C6-aroma compounds in starch dispersions and its impact on retention.” Food
Chemistry 96(3): 461–70.
Jouquand, C., Y. Aguni, C. Malhiac, and M. Grisel (2008). “Influence of chemical
composition of polysaccharides on aroma retention.” Food Hydrocolloids 22(6):
1097–104.
Jung, D. M., J. S. de Ropp, and S. E. Ebeler (2002). “Application of pulsed field gradi-
ent NMR techniques for investigating binding of flavor compounds to macromol-
ecules.” Journal of Agricultural and Food Chemistry 50(15): 4262–9.
 The Food Matrix and Aroma Compounds  647

Jung, D. M. and S. E. Ebeler (2003). “Investigation of binding behavior of alpha- and beta-
ionones to beta-lactoglobulin at different pH values using a diffusion-based NOE
pumping technique.” Journal of Agricultural and Food Chemistry 51(7): 1988–93.
Juteau, A., J. L. Doublier, and E. Guichard (2004). “Flavor release from iota-carrageenan matri-
ces: A kinetic approach.” Journal of Agricultural and Food Chemistry 52(6): 1621–9.
Kant, A., R. S. T. Linforth, J. Hort, and A. J. Taylor (2004). “Effect of beta-cyclodex-
trin on aroma release and flavor perception.” Journal of Agricultural and Food
Chemistry 52(7): 2028–35.
King, B. M. and J. Solms (1982). “Interactions of volatile flavor compounds with propyl
gallate and other phenols as compared with caffeine.” Journal of Agricultural and
Food Chemistry 30(5): 838–40.
King, B. M., P. Arents, N. Bouter, C. A. A. Duineveld, M. Meyners, S. I. Schroff, and S.
T. Soekhai (2006). “Sweetener/sweetness-induced changes in flavor perception and
flavor release of fruity and green character in beverages.” Journal of Agricultural
and Food Chemistry 54(7): 2671–7.
Kopjar, M., I. Andriot, A. Saint-Eve, I. Souchon, and E. Guichard (2010). “Retention
of aroma compounds: An interlaboratory study on the effect of the composition of
food matrices on thermodynamic parameters in comparison with water (Research
article).” Journal of the Science of Food and Agriculture 90(8): 1285–92.
Langourieux, S. and J. Crouzet (1994). “Study of aroma compounds polysaccharides
interactions by dynamic exponential dilution.” Food Science and Technology-
Lebensmittel-Wissenschaft & Technologie 27(6): 544–9.
Lauverjat, C., I. Deleris, C. I. Tréléa, C. Salles, and I. Souchon (2009). “Salt and aroma
compound release in model cheeses in relation to their mobility.” Journal of
Agricultural and Food Chemistry 57(21): 9878–87.
Le Guen, S. and R. Vreeker (2003). “Interactions between flavour compounds and milk
proteins under static and dynamic conditions.” In Flavour Research at the Dawn of
the Twenty-First Century, Editions Tec & Doc.
Lorrain, B., S. Tempere, N. Iturmendi, V. Moine, G. de Revel, and P. L. Teissedre (2013).
“Influence of phenolic compounds on the sensorial perception and volatility of red
wine esters in model solution: An insight at the molecular level.” Food Chemistry
140(1–2): 76–82.
Lubbers, S. (2006). “Texture-aroma interactions.” In Flavour in Food. A. Voilley, and
P. Etiévant (Eds.), Cambridge, CB1 6AH, UK, Woodhead Publishing Limited and
CRC Press LLC. Part 3: 327–44.
Lubbers, S. and E. Guichard (2003). “The effects of sugars and pectin on flavour release
from a fruit pastille model system.” Food Chemistry 81(2): 269–73.
Lubbers, S., N. Decourcelle, D. Martinez, E. Guichard, and A. Tromelin (2007). “Effect
of thickeners on aroma compound behavior in a model dairy gel.” Journal of
Agricultural and Food Chemistry 55(12): 4835–41.
Lubbers, S., P. Landy, and A. Voilley (1998). “Retention and release of aroma compounds
in foods containing proteins.” Food Technology 52(5): 68–214.
Maier, H. G. (1975). “Binding of volatile aroma substances to nutrients and foodstuffs.”
In International Symposium on Aroma Research, Zeist, the Netherlands, Pudoc,
Wageningen.
Maraval, I., C. Mestres, K. Pernin, F. Ribeyre, R. Boulanger, E. Guichard, and Z. Gunata
(2008). “Odor-active compounds in cooked rice cultivars from Camargue (France)
analyzed by GC-O and GC-MS.” Journal of Agricultural and Food Chemistry
56(13): 5291–8.
648 Food Aroma Evolution

Mei, J. and G. Reineccius (2007). “The influence of texture on aroma release and percep-
tion related to dairy products.” In Flavor of Dairy Products. K. R. Cadwallader, M.
Drake, and R. J. McGorrin (Eds.), Vol. 971, ACS: 93–110.
Meynier, A., V. Rampon, M. Dalgalarrondo, and C. Genot (2004). “Hexanal and
t-2-hexenal form covalent bonds with whey proteins and sodium caseinate in aque-
ous solution.” International Dairy Journal 14(8): 681–90.
Nahon, D. F., M. Harrison, and J. P. Roozen (2000). “Modeling flavor release from
aqueous sucrose solutions, using mass transfer and partition coefficients.” Journal
of Agricultural and Food Chemistry 48(4): 1278–84.
Nahon, D. F., P. Koren, J. P. Roozen, and M. A. Posthumus (1998). “Flavor release
from mixtures of sodium cyclamate, sucrose, and an orange aroma.” Journal of
Agricultural and Food Chemistry 46(12): 4963–8.
Nawar, W. W. (1971). “Some variables affecting composition of headspace aroma.”
Journal of Agricultural and Food Chemistry 19(6): 1057–9.
Nongonierma, A. B., M. Springett, J. L. Le Quere, P. Cayot, and A. Voilley (2006).
“Flavour release at gas/matrix interfaces of stirred yoghurt models.” International
Dairy Journal 16(2): 102–10.
O’Neill, T. E. (1996). “Flavor binding by food proteins: An overview.” In Flavor-Food
Interactions, Washington, DC, American Chemical Society.
O’Neill, T. E. and J. E. Kinsella (1987). “Binding of alkanone flavors to beta-lactoglob-
ulin: Effects of conformational and chemical modification.” Journal of Agricultural
and Food Chemistry 35(5): 770–4.
Piccone, P., V. Lonzarich, L. Navarini, G. Fusella, and P. Pittia (2012). “Effect of sugars
on liquid-vapour partition of volatile compounds in ready-to-drink coffee bever-
ages.” Journal of Mass Spectrometry 47(9): 1120–31.
Pozo-Bayon, M. A., B. Biais, V. Rampon, N. Cayot, and P. Le Bail (2008). “Influence
of complexation between amylose and a flavored model sponge cake on the degree
of aroma compound release.” Journal of Agricultural and Food Chemistry 56(15):
6640–7.
Rabe, S., U. Krings, and R. G. Berger (2003a). “Dynamic flavor release from sucrose
solutions.” Journal of Agricultural and Food Chemistry 51(17): 5058–66.
Rabe, S., U. Krings, and R. G. Berger (2003b). “Initial dynamic flavour release from
sodium chloride solutions.” European Food Research and Technology 218(1): 32–9.
Relkin, P., M. Fabre, and E. Guichard (2004). “Effect of fat nature and aroma com-
pound hydrophobicity on flavor release from complex food emulsions.” Journal of
Agricultural and Food Chemistry 52(20): 6257–63.
Repoux, M., H. Labouré, P. Courcoux, I. Andriot, E. Sémon, C. Yven, G. Feron, and E.
Guichard (2012). “Combined effect of cheese characteristics and food oral process-
ing on in vivo aroma release.” Flavour and Fragrance Journal 27(6): 414–23.
Roberts, D. D., J. S. Elmore, K. R. Langley, and J. Bakker (1996). “Effects of sucrose , guar
gum , and carboxymethycellulose on the release of volatile flavor compounds under
dynamic conditions.” Journal of Agricultural and Food Chemistry 44: 1321–6.
Roudnitzky, N., H. Irl, G. Roudaut, and E. Guichard (2003). “Influence of fat nature on
flavour release.” In Flavour Research at the Dawn of the Twenty-First Century. J.
L. Le Quere, and P. X. Etievant (Eds.), Paris (France), Lavoisier Tec & Doc: 136–9.
Rutschmann, M. A. and J. Solms (1990a). “Formation of inclusion complexes of starch
with different organic compounds. II. Study of ligand binding in binary model sys-
tems with decanal, 1-naphthol, monosteatate and monopalmitate.” Lebensmittel
Wissenschaft und Technologie 23: 70–9.
 The Food Matrix and Aroma Compounds  649

Rutschmann, M. A. and J. Solms (1990b). “Formation of inclusion complexes of starch

with different organic compounds. III. Study of ligand binding in binary model sys-
tems with (-)limonene.” Lebensmittel Wissenschaft und Technologie 23(1): 80–3.
Rutschmann, M. A. and J. Solms (1990c). “Formation of inclusion complexes of starch
with different organic compounds. IV. Ligand binding and variability in helical con-
formations of V amylose complexes.” Lebensmittel Wissenschaft und Technologie
23(1): 84–7.
Saint-Eve, A., A. Juteau, S. Atlan, N. Martin, and I. Souchon (2006). “Complex viscosity
induced by protein composition variation influences the aroma release of flavored
stirred yogurt.” Journal of Agricultural and Food Chemistry 54(11): 3997–4004.
Saint-Eve, A., C. Lauverjat, C. Magnan, I. Deleris, and I. Souchon (2009). “Reducing salt
and fat content: Impact of composition, texture and cognitive interactions on the
perception of flavoured model cheeses.” Food Chemistry 116(1): 167–75.
Sakurai, K., M. Oobatake, and Y. Goto (2001). “Salt-dependent monomer-dimer equilib-
rium of bovine beta-lactoglobulin at pH 3.” Protein Sci 10(11): 2325–35.
Semenova, M. G., A. S. Antipova, L. E. Belyakova, Y. N. Polikarpov, L. A. Wasserman,
T. A. Misharina, M. B. Terenina, and R. V. Golovnya (2002). “Binding of aroma
compounds with legumin. III. Thermodynamics of competitive binding of aroma
compounds with 11S globulin depending on the structure of aroma compounds.”
Food Hydrocolloids 16(6): 573–84.
Semenova, M. G., A. S. Antipova, T. A. Misharina, and R. V. Golovnya (2002). “Binding
of aroma compounds with legumin. I. Binding of hexyl acetate with 11S globulin
depending on the protein molecular state in aqueous medium.” Food Hydrocolloids
16(6): 557–64.
Seuvre, A. M. and A. Voilley (2017). Physico-Chemical Interactions in the Flavor-
Release Process. Springer.
Seuvre, A. M., M. A. E. Diaz, and A. Voilley (2001). “Retention of aroma compounds by
beta-lactoglobulin in different conditions.” Food Chemistry 77(4): 421–9.
Shimizu, S., R. Stenner, and N. Matubayasi (2017). “Gastrophysics: Statistical thermody-
namics of biomolecular denaturation and gelation from the Kirkwood-Buff theory
towards the understanding of tofu.” Food Hydrocolloids 62: 128–39.
Siebert, K. J., N. V. Troukhanova, and P. Y. Lynn (1996). “Nature of polyphenol-protein
interactions.” Journal of Agricultural and Food Chemistry 44(1): 80–5.
Siefarth, C., O. Tyapkova, J. Beauchamp, U. Schweiggert, A. Buettner, and S. Bader
(2011). “Mixture design approach as a tool to study in vitro flavor release and vis-
cosity interactions in sugar-free polyol and bulking agent solutions.” Food Research
International 44(10): 3202–11.
Sostmann, K. and E. Guichard (1998a). “Immobilized β-lactoglobulin on a HPLC-
column: A rapid way to determine protein-flavour interactions.” Food Chemistry
62(4): 509–13.
Sostmann, K. and E. Guichard (1998b). “Immobilized beta-lactoglobulin on a HPLC-
column: A rapid way to determine protein/flavour interactions.” Food Chemistry
62: 509–13.
Tavel, L., C. Moreau, S. Bouhallab, E. C. Y. Li-Chan, and E. Guichard (2010).
“Interactions between aroma compounds and beta-lactoglobulin in the heat-induced
molten globule state.” Food Chemistry 119(4): 1550–6.
Tavel, L., E. Guichard, and C. Moreau (2008). “Contribution of NMR spectroscopy to
flavour release and perception.” In Annual Reports on NMR Spectroscopy. G. A.
Webb (Ed.), Vol. 64, Amsterdam (NLD), Elsevier: 173–88.
650 Food Aroma Evolution

Tavel, L., I. Andriot, C. Moreau, and E. Guichard (2008). “Interactions between beta-
lactoglobulin and aroma compounds: Different binding behaviors as a function of
ligand structure.” Journal of Agricultural and Food Chemistry 56(21): 10208–17.
Thomas-Danguin, T. (2009). “Flavor.” In Encyclopedia of Neuroscience. M. D. Binder,
N. Hirokawa, and U. Windhorst (Eds.), Berlin Heidelberg (Deutschland), Springer-
Verlag GmbH, Version électronique: 4399.
Tromelin, A. and E. Guichard (2003). “Use of Catalyst in a 3D-QSAR study of the inter-
actions between flavor compounds and β-lactoglobulin.” Journal of Agricultural
and Food Chemistry 51(7): 1977–83.
Tromelin, A. and E. Guichard (2004). “2D-and 3D-QSAR models of interaction between
flavor compounds and beta-lactoglobulin using catalyst and Cerius2.” Qsar &
Combinatorial Science 23(4): 214–33.
Tromelin, A., I. Andriot, and E. Guichard (2006a). “Protein-flavour interactions.” In
Flavour in Food. A. Voilley and P. Etievant (Eds.), Cambridge, CB1 6AH (GBR),
Woodhead Publishing Limited. Part 2. Flavour retention and release from the food
matrix: 172–207.
Tromelin, A., I. Andriot, and E. Guichard (2006b). “Protein-flavour interactions.” In
Flavour in Food. A. Voilley and P. Etiévant (Eds.), Cambridge, CB1 6AH, UK,
Woodhead Publishing Limited and CRC Press LLC. Part 3: 172–207.
Tromelin, A., I. Andriot, M. Kopjar, and E. Guichard (2010). “Thermodynamic and
structure property study of liquid-vapor equilibrium for aroma compounds.”
Journal of Agricultural and Food Chemistry 58(7): 4372–87.
van Ruth, S. M., and E. Villeneuve (2002). “Influence of beta-lactoglobulin, pH and pres-
ence of other aroma compounds on the air/liquid partition coefficients of 20 aroma
compounds varying in functional group and chain length.” Food Chemistry 79(2):
157–64.
Villamor, R. R., M. A. Evans, D. S. Mattinson, and C. F. Ross (2013). “Effects of etha-
nol, tannin and fructose on the headspace concentration and potential sensory sig-
nificance of odorants in a model wine.” Food Research International 50(1): 38–45.
Voilley, A. (2006). “Flavour retention and release from the food matrix: An overview.”
In Flavour in Food. A. Voilley and P. Etiévant (Eds.), Cambridge, CB1 6AH, UK,
Woodhead Publishing Limited and CRC Press LLC. Part 3: 117–32.
Voilley, A., D. Simatos, and M. Loncin (1977). “Gas phase concentration of volatiles in
equilibrium with a liquid aqueous phase.” Lebensmittel -Wissenchaft & Technologie
10: 45–9.
Voilley, A., V. Beghin, C. Charpentier, and D. Peyron (1991). “Interactions between aroma
substances and macromolecules in a model wine.” Lebensmittel Wissenschaft und
Technologie 24(5): 469–72.
Wang, J., M. M. Zhao, C. Y. Qiu, and W. Z. Sun (2018). “Effect of malondialdehyde
modification on the binding of aroma compounds to soy protein isolates.” Food
Research International 105: 150–8.
Wang, K. and S. D. Arntfield (2015). “Effect of salts and pH on selected ketone fla-
vours binding to salt-extracted pea proteins: The role of non-covalent forces.” Food
Research International 77: 1–9.
Wong, D. W. S., W. M. Camirand, and A. E. Pavlath (1996). “Structures and function-
alities of milk proteins.” Critical Reviews in Food Science and Nutrition 36(8):
807–44.
 Chapter 30
Food Emulsions as Flavor
Delivery Systems
Like Mao, Yrjö H. Roos, Costas G. Biliaderis, and Song Miao

CONTENTS

30.1 Introduction 652

30.2 Physical Chemistry of Flavor Release 652
30.2.1 Flavor Release from Emulsions 653
30.2.1.1 Partition Equilibrium in Different Phases 653
30.2.1.2 Flavor Release from Real Food 654
30.2.2 Flavor Release during Eating 655
30.3 Emulsion Ingredients and Flavor Release 656
30.3.1 Oil Phase 656
30.3.1.1 Oil Property 656
30.3.1.2 Oil Content 657
30.3.2 Water Phase 658
30.3.2.1 Emulsifiers 658
30.3.2.2 Thickening Agents 659
30.3.2.3 Salts 660
30.4 Emulsion Properties and Flavor Release 660
30.4.1 Droplet Size 660
30.4.2 Rheological Behavior 661
30.4.3 pH 661
30.4.4 Other Factors 662
30.5 Effect of Mouth Conditions on Flavor Release and In Vitro Studies 663
30.5.1 Effect of Saliva on Flavor Release 663
30.5.2 Model Mouths and Throats 663
30.6 Novel Emulsions for Controlled Flavor Release 665
30.6.1 Multilayer Emulsion 665
30.6.2 Gelled Emulsion 666
30.6.3 Multiple Emulsion 667
30.6.4 Pickering Emulsion 667
30.6.5 Self-Assembly Structured Emulsion 668
30.7 Conclusion 668
References 669

651
652 Food Aroma Evolution

30.1 INTRODUCTION

There is a growing interest in functional foods that are low fat, low sugar, low salt, or that
are bioactive-enriched, to develop healthier diets for human wellbeing. In the meantime,
consumers ask for food with desirable organoleptic properties, particularly texture and
flavor profiles. It is a big challenge for academic and industrial researchers to design healthy
food products without sacrificing food flavor, as flavor release is not only influenced by
food ingredients but also food structures (Druaux and Voilley, 1997). Flavor release from
the food matrix affects flavor perception. Therefore, researchers have been working on
designing delivery systems for flavor compounds to better control their release when incor-
porated in foods, and subjected to different environmental stresses encountered during
product processing, storage, and mastication in the oral cavity.
Emulsions consist of two immiscible phases, one of which is dispersed in the other as
small droplets; these are known as the dispersed phase and continuous phase, respectively
(McClements, 2005). Functional food ingredients can be incorporated into the dispersed
droplets, and thus isolated from the external environment by the continuous phase. Many
studies have shown that emulsions are effective delivery systems for functional food ingredi-
ents (bioactive lipids, anti-oxidants, flavor, etc.) to protect them from degradation (mechani-
cal, chemical, and enzymatic), to disperse them in aqueous media, to control their release,
and finally to improve their bioavailability in the human gastrointestinal tract (Velikow
and Pelan, 2008; McClements, 2010). An important characteristic of an emulsion is that
the structures in the water phase, oil phase, and interface can be designed to meet spe-
cial requirements, and the release of the compounds incorporated can then be modified
(Mao and Miao, 2015; McClements and Li, 2010). Flavor release from emulsions includes
the partitioning and mass transfer of the flavor molecules between the oil phase, interface,
water phase, and finally headspace (Taylor, 1998). Changes in headspace concentration and
release rate could affect the overall flavor perception. The successful development of delivery
systems with controlled flavor release depends on a good understanding of the effects of
emulsion properties (e.g., droplet size and distribution, and viscosity), and of environmen-
tal stresses affecting flavor release, as well as the interactions between flavor compounds
and emulsion components. Recent work suggested the possibility to modulate flavor release,
including release rate and intensity, by modifying emulsion microstructures through food
engineering techniques (Ghosh et al., 2006; Benjamin et al., 2012b; Mao et al., 2014).
The objective of this article is to give an updated overview of recent advances in
­emulsion-based delivery systems for food flavors. A brief summary is first made on the
theories developed to describe flavor release from emulsions and our understanding of
flavor release in the mouth. More emphasis is made on factors affecting flavor release,
that is, different emulsion ingredients and emulsion properties, and how controlled flavor
release can be achieved by appropriate modification of all structural elements of the food
dispersion. In the last section, novel structured emulsions with the potential to better
deliver flavors in more complex systems are introduced. It should be noted, however, that
the flavors discussed in this review are smell-related compounds (volatiles), whereas taste-
related compounds are not included.

30.2 PHYSICAL CHEMISTRY OF FLAVOR RELEASE

Volatile flavors are perceived when they are in contact with receptors in the nose (olfac-
tory epithelium) either orthonasally by directly sniffing foods or retronasally via the
 Food Emulsions as Flavor Delivery Systems 653

delivery of volatiles during mastication and/or swallowing. Before flavor perception is

realized, flavor compounds have to move out from the food matrix, and experience immi-
gration in the oral cavity (and/or nasal cavity).

30.2.1 Flavor Release from Emulsions

Flavor release is mainly controlled by two factors, the volatility of flavor compounds
(thermodynamic factor) and the resistance to mass transfer from the emulsion to the air
phase (kinetic factor) (de Roos, 2000). The thermodynamic factor determines the reten-
tion or partition of flavors in the matrix at equilibrium, and the kinetic factor mainly
affects the release rate of flavors from food. Flavor release from an O/W emulsion can be
simplified into four steps: (1) flavor movement inside the oil droplets; (2) movement over
the oil–water interface; (3) movement within the aqueous phase; (4) movement across the
air–emulsion interface (Figure 30.1).
Theoretically, each step affects the release rate and contributes to the headspace con-
centration. In reality, only one or two steps can dominate the release.

30.2.1.1 Partition Equilibrium in Different Phases

Static headspace analysis is widely applied to evaluate flavor release, and it is based on
the theory of partition equilibrium. In a closed system, flavor compounds dissolved in

FIGURE 30.1 Schematic diagram of flavor release from O/W emulsion and affecting
factors.
654 Food Aroma Evolution

an emulsion can reach partition equilibrium in different phases, and the partition coef-
ficient is dependent on the affinity of flavor compounds for each phase. Therefore, for a
simple O/W emulsion, the overall distribution of flavor compounds between the emulsion
matrix and its headspace can be expressed as (Buttery et al., 1973):

1 f f
= D + C (30.1)
KGE KGD KGC

where KGE is the partition coefficient of a flavor between air (headspace) and emulsion;
ϕD is the mass fraction of dispersed phase; ϕC is the mass fraction of the continuous
phase (ϕDϕC = 1); KGC is the partition coefficient between air and the continuous phase
(aqueous emulsifier solution); and KGD is the partition coefficient between air and the
dispersed phase. Several studies showed that this model could give a good prediction of
the headspace–emulsion partition coefficient when the interfacial binding was insignifi-
cant (Guyot et al., 1996; Ghosh et al., 2007). On the other hand, the difference in the
calculated and experimental KGE values can be regarded as proof of interfacial binding
(Karaiskou et al., 2008; Meynier et al., 2005).
In many emulsions, particularly the ones stabilized by biopolymers, flavor binding to
interfacial components has been widely observed, and it plays a significant role in flavor
partition (Guichard, 2002). For reversible binding, a modified model of Equation 30.1
describing flavor partition was proposed by McClements (2005):

1 f f A K*
= D + C + S IC (30.2)
KGE KGD KGC KGC

where A S is the interfacial area per unit volume of an emulsion (A S =6ϕ/d 32, d 32 is the
v­ olume-surface mean droplet diameter); K*IC is the interfacial binding coefficient (K*IC =
ΓI /CC , ΓI is the mass of flavor compound per unit of interfacial area; CC is the flavor com-
pound concentration in the continuous phase). From Equation 30.2, it is observed that
KGE is droplet size dependent. When droplet size is large enough, A S will be infinitesimal.
Then interfacial binding can be neglected, and Equation 30.2 is equal to Equation 30.1.
For the irreversible binding, McClements (2005) proposed a different model:

1 æ CE ö æ fD fC ö
=ç ÷ç K + K ÷ (30.3)
e
KGE C
è E - A G
S I ø è GD GC ø

where CE is flavor concentration added to the emulsion, and ΓI is the mass of flavor com-
pounds irreversibly bound to the interface per unit of interfacial area. It should be noted
that Equation 30.2 and Equation 30.3 do not involve flavor binding in the continuous phase
or dispersed phase, for example, flavor solubilization in emulsifier micelles, gum binding.
As a result, the predicted values of KGE could be higher than the experimental values.

30.2.1.2 Flavor Release from Real Food

The eating processes usually finish in a short time, and flavor release hardly reaches
equilibrium. Therefore, the perception of volatile flavor mainly depends on the initial
dynamic release. The theory of dynamic release reveals more information about the
process of flavor release, as more parameters are considered, which on the other hand
results in the application of more complex models to describe the process. The following
 Food Emulsions as Flavor Delivery Systems 655

equation proposed by Harrison and Hills (1997) is an example of the models used to
predict headspace concentration at a given time (t):

Ctf (0)
C g (t ) =
1 + K b Cb æ v g ö é ìï Age hD æ 1 vg 1 ö üï ù (30.4)
+ ç ÷ ê1 - exp í - + tý ú
K ge è ve ø ê îï v g è K ge ve 1 + K bCb ÷ø þï ú
ç
ë û

where Ctf(0) is the initial flavor concentration (mg cm−3) in the emulsion, Kb is the flavor
binding affinity to biopolymers (including some emulsifiers) (M−1), Cb is the biopolymer
concentration in the emulsion (M), vg , and ve are the headspace and emulsion volumes
(cm3), and Age is the gas/emulsion surface (cm 2). The mass transfer coefficient hD (cm s−1)
and the gas–emulsion partition coefficient Kge can be derived from the above equation at
the initial release rate (t→0) and at the equilibrium headspace concentration (t→infinite),
respectively. This model follows the penetration theory, which was developed to describe
flavor release from liquid samples. In this theory, flavor transfer through the gas–liquid
interface is assumed to be rate-limiting and is driven by molecular diffusion and eddy
diffusion. The binding effect is taken into consideration in this model, and it has been
successfully applied to predict flavor release from different liquid systems (Perreault et al.,
2010; Benjamin et al., 2012b).
Lian et al. (2004) have developed another model which took two rate-limiting steps
into consideration: one was the flavor transfer from droplets to the continuous phase;
the other one was the flavor transfer from continuous phase to the headspace. Actually,
for emulsions with smaller droplet size, where flavor release is governed by the mass
transfer across the air–emulsion interface, this model is similar to the model developed
by Harrison and Hill (1997). Nevertheless, for emulsions with larger droplet size, Lian et
al.’s model suggested that the transfer across the droplet interface becomes rate-limiting,
as the larger droplets can offer a significant barrier to diffusion. In Lian et al.’s study on
gelled emulsions, the experimental data of four ketones and one ester fitted the model
well (Lian et al., 2004).

30.2.2 Flavor Release during Eating

Food in the mouth undergoes mastication, salivation, bolus formation, and finally swal-
lowing. Each step can have a big influence on flavor release and flavor perception. Liquid
foods normally stay in the mouth for several seconds before swallowing, so only one or
two of the above processes have a significant effect on flavor release. When liquids are
kept in the mouth, a tight closure formed by the soft palate and the base of the tongue
prevents flavor transfer from the oral cavity to the nasal cavity. There is also no flavor
transfer during swallowing, as the airway from the trachea to the nose is closed at that
stage (Bosman, 1980; Normand et al., 2004). As a result, flavor perception is only avail-
able after swallowing (Land, 1996). Studies suggested that the highest perception was
achieved in the first expiration immediately after swallowing (Linforth and Taylor, 2000).
These flavors originate from the air phase above liquid food in the mouth before swal-
lowing, which are later transported to the throat during swallowing, and finally trans-
ferred to the nose in the first expiration. This process is called “swallow breath,” which
finishes in a short time (de Roos and Wolswinkel, 1994; Land, 1996). There is another
process of flavor transfer after swallowing with a much lower flavor concentration, which
656 Food Aroma Evolution

contributes to the persistence of flavor perception. Specifically, a thin film of liquid food
will remain at the surface of the pharynx after swallowing, and the flavor in the film can
be released and transported to the nasal cavity during exhalation. As there is a gradi-
ent in flavor concentration between the thin film and the exhaled air, this release can
last for a short period of time (Buettner et al., 2002; Weel et al., 2004a; Boelrijk et al.,
2006). Moreover, the little amount of liquid food remaining in the mouth (adsorbed to
the mucosa) after the first swallowing may also contribute to the persistence of flavor
perception (Harrison, 1998; Salles et al., 2011).

30.3 EMULSION INGREDIENTS AND FLAVOR RELEASE

30.3.1 Oil Phase

Most food flavors are lipophilic, and oils play a more significant role in their release than
any other emulsion ingredient, for example, proteins (Innocente et al., 2014). Oils can act
as flavor precursors, as solvents for flavor compounds, and as flavor release modulators.
Change in oil property or oil content can lead to a significantly modified flavor release
profile (de Roos, 1997), which is difficult to recover by adding fat replacers. However,
consumers are highly cautious about the fat/oil content of food, as overconsumption of
fat/oil can increase the risks of many diseases, for example, obesity, cardiovascular dis-
eases, cancers (Gastaldelli and Basta, 2010). Therefore, knowledge about the effects of oil
on flavor release becomes essential for food scientists to properly design fat-reduced food.

30.3.1.1 Oil Property
Flavors in different oils generally have different release behaviors, due to their affinity
for different oils. The chain length of fatty acids determines the polarity of oils, which
is one of the main factors affecting the interactions between flavor compounds and oils.
Most flavor compounds are lipophilic, and they have a higher affinity for oils containing
longer-chain fatty acids (Harvey et al., 1995). However, there was a study showing that
some hydrophobic flavor compounds had a higher release from the emulsion containing
lipids with average carbon number (CN) of C14 or C16 than from the emulsion contain-
ing C9 lipids (Rabe et al., 2003a). Another study reported no difference in flavor release
when changing the lipid type from milk fat (C16 and C18) to MCT (C6 and C8) in emul-
sions (Roberts et al., 2003b). It seemed that lipophilicity of different oils was not the only
factor that influenced the affinity of flavors for the oils, or the affinity had already been
so high that change in lipophilicity of the oils did not show any effect on the mentioned
studies. Rabe et al. (2003a) suggested that the molarity of oil (droplet concentration)
in the system could give additional effects. They made emulsions containing the same
molarity of C16 and C9 but they differed in the mass fraction. These authors did not
observe any significant difference in flavor release between the two emulsions. Roberts
et al. (2003b) proposed that the saturation level of different oils could affect flavor parti-
tion: with stearine (a more saturated fat), flavor release was shown to be slower and of
lower intensity than the release from olein (a more unsaturated fat). This result was in
agreement with observations made by Welsh and Williams (1989), who found the oil–
water partition coefficients of many flavors were lower in olive oil than in sunflower oil.
However, Roudnitzky et al. (2003) found that the volatility of ethyl hexanoate in veg-
etable oils differing by their percentages of oleic (C18:1), linoleic (C18:2), and linolenic
(C18:3) decreased when the unsaturation level increased. The above inconsistent findings
 Food Emulsions as Flavor Delivery Systems 657

revealed that oils may affect flavor release through different mechanisms, depending on
oil nature (composition and physical state) and flavor characteristics.
Lipids in emulsions can indeed be present in a solid (i.e., fat), partial solid, or liquid
(i.e., oil) state, which can exhibit different effects on flavor release (Roberts et al., 2003b;
Relkin et al., 2004). The consensus is that flavor compounds can only partition into liq-
uid oil, and the formation of solid oil could inhibit the migration of flavor compounds
(Maier, 1975; McNulty and Karel, 1973). In an O/W emulsion, flavor compounds had
higher headspace concentrations above a solid droplet emulsion than those above a liquid
droplet emulsion (Relkin et al., 2004; Ghosh et al., 2006). Nevertheless, the initial sorp-
tion rates of flavor compounds were much higher for the solid lipid, which was attributed
to the adsorption of flavor compounds onto the surface of solid lipid particles (Ghosh et
al., 2006). In a milk-based liquid emulsion, Roberts et al. (2003b) found that an increase
in the content of solid palm fat resulted in accelerated flavor release. However, when the
melted palm fat was tested, there was no difference in flavor release from emulsions with
different oils (Roberts et al., 2003b). In a more complicated situation, flavor compounds
were dissolved in liquid oil before oil solidification, and flavor release was reported to
slow down as oil crystallization went on (increase in solid fat content). This finding could
be explained by the fact that flavor molecules were trapped within the solid particles, and
the movement of flavors across the droplet surface was inhibited (Roberts et al., 2003b).

30.3.1.2 Oil Content
The oil content of food has been shown to affect not only the perceived intensity but also
the temporal profile of flavors, as well as the release behavior throughout storage. Oil
reduction or even complete omission of an oil phase in food can lead to a drastic shift of
the overall flavor profile. Fat-free products therefore often show an undesirable transient
flavor burst, as the release is no longer mediated by a fat phase (Rabe et al., 2003a).
It has been well documented that a reduction in oil content will accelerate the release
of lipophilic flavor compounds, and the headspace concentration of flavors above an
emulsion with lower oil content is normally higher. In other words, to trigger the same
flavor release, flavor compounds added to emulsions with lower oil content could be at
a lower level than that added to emulsions with higher oil content (Bayarri et al., 2006).
Only a small portion of flavors (volatile) are hydrophilic compounds, and they have dif-
ferent release behaviors. Both in vivo and static headspace studies showed that the release
of hydrophilic flavor compounds was not or even positively affected by changing oil
content (Rabe et al., 2003a; Giroux et al., 2007; Frank et al., 2011). The effect of oil
content on flavor release is so dominant that sometimes it makes the contribution of other
ingredients undetectable. Roberts and Pollien (2000) found that the evidence of protein
or lactose binding with lipophilic compounds was seen as the concentration of skimmed
milk increased, but the binding effect was no longer detected after the addition of 1.3%
lipid. A similar study found that in the presence of ≥1% oil in an oil (MCT)–water
system, the bindings between flavor compounds and β-lactoglobulin were insignificant
(Seuvre et al., 2000). Giroux et al. (2007) compared the effect of oil and protein content
on flavor release from emulsions and found that increasing the oil content (from 1–8%)
reduced flavor release eight times more than when protein (from 0.1 to 3.16%) was added
to the system.
The oil content also plays an important role in flavor perception under oral condi-
tions, but the role is not as strong as that under static conditions (Weel et al., 2004b;
Roberts et al., 2003a; Doyen et al., 2001; Karaiskou et al., 2008). As some researchers
suggested, the perceived flavor is from the “swallow breath” and the thin liquid layer
658 Food Aroma Evolution

formed on the throat surface, and the role of oil as flavor reservoir is not as significant
as that in the bulk phase situation (Land, 1996; Linforth and Taylor, 2000). Moreover,
flavor perception could be affected by the presence of sweets and the viscosity of different
systems, and the role of flavor intensity may be weakened (Bayarri et al., 2006).
Finally, a change in oil nature or oil content may influence flavor release indirectly
by modifying the physicochemical properties of the emulsions concerned, for example,
droplet size, viscosity, and emulsion stability (discussed in Section 30.4).

30.3.2 Water Phase

The water phase generally makes up the majority fraction of an O/W emulsion, and it
may contain many ingredients, for example, emulsifiers, thickening agents, salts, and
minerals. Therefore, the water phase also contributes a lot to the overall perception of a
food emulsion. All the ingredients may interact with flavor compounds, thereby modify-
ing their release behaviors. However, as only a small part of volatile flavor compounds
are hydrophilic, the water phase does not influence flavor release as much as the oil phase.

30.3.2.1 Emulsifiers
Emulsifiers play critical roles in the formation of emulsions, and they have a strong impact
on most emulsion properties. When flavor compounds are incorporated in emulsions, the
emulsifiers can influence flavor release through different mechanisms. Among all of the
food emulsifiers, proteins are the most widely used and extensively investigated emulsi-
fiers regarding flavor release. It is generally acknowledged that proteins in emulsions can
slow flavor release, and the effects are dependent on protein and flavor types. Widder and
Fisher (1996) observed that sodium caseinate had the capacity to inhibit the release of
esters, and Maier (1970) found casein and egg albumin could bind acetone and acetalde-
hyde. Dubois et al. (1996) reported an increase in retention of diacetyl and diallyl sulfide
in a model cheese system with a higher interfacial concentration of calcium caseinate.
Some studies revealed that β-lactoglobulin in emulsions could modify the release behav-
iors of aldehydes, ketones, and esters (Guichard, 2002). Charles et al. (2000) indicated
that ethyl hexanoate exhibited significantly higher release from the emulsion containing
α-lactalbumin than from the emulsion containing β-lactoglobulin. However, once the
proteins were hydrolyzed by enzymes, the peptides had a reduced capacity to retain flavor
compounds (Wong et al., 2013).
The reduced flavor release in protein-stabilized emulsions is achieved mainly through
two different mechanisms. First, proteins are likely to interact with flavor compounds,
through reversible or irreversible interactions. In most cases, flavor compounds interact
with proteins through hydrophobic binding or hydrogen bonding, both of which are
reversible (Lubbers et al., 1998). Wu et al. (1999) proposed that the most probable bind-
ing site for these compounds was the hydrophobic pocket (the central calyx) within the
protein structure. In the case of aldehydes, irreversible covalent binding with proteins
was also reported (Gremli, 1974), and these compounds are likely to be bound at the
protein surface (Lübke et al., 2000). For a series of different milk proteins, the binding
capacity to 2-nonanone followed the order: bovine serum albumin > β-lactoglobulin >
α-lactalbumin > αs1-casein > β-casein, and the capacity of whey protein isolate (WPI)
was stronger than that of sodium caseinate (Kühn et al., 2007). According to Harrison
and Hills (1997), binding reduces the concentration of free flavors in the aqueous phase
and, consequently, the flavors released into the headspace. Second, proteins adsorbed
 Food Emulsions as Flavor Delivery Systems 659

at the interface can act as barriers to slow mass transfer of flavor molecules, leading to
reduced release rates (Harvey et al., 1995). For example, the presence of β-lactoglobulin
at a miglyol–water interface strengthened the resistance against the transfer of benz-
aldehyde across the lipid layer (Rogacheva et al., 1999). A similar result was found
by Guichard and Langourieux (2000), who reported that the presence of interfacial
β-lactoglobulin inhibited the transfer of hydrophobic flavor compounds from the oil
droplets to the water phase. Land (1978) further confirmed these findings, and it was
indicated that the inclusion of a small amount of emulsifier in a non-emulsified oil–water
system had no effect on the headspace concentration of dimethylsulfide, whereas in an
emulsified system the headspace concentration was significantly decreased. Due to the
large variations in physicochemical properties of the proteins, the interfaces formed by
different proteins may vary in thickness, porosity, compactness, and so on. (Wilde et
al., 2004; McClements, 2005), which could result in different barrier effect on flavor
release. Benjamin et al. (2014) studied the effect of legume proteins (soy, pea, and lupin)
on flavor release, in comparison with β-lactoglobulin. The findings suggested that the
ability of legume proteins to interact with flavors was comparable with β-lactoglobulin
for the affinity to more hydrophobic compounds, and different legume proteins showed
a similar effect on flavor release.
In terms of some small molecular weight surfactants (e.g., tweens, spans), only lim-
ited studies have been performed concerning their effects on flavor release. Although no
sufficient proof is available about the interactions between surfactants and flavor com-
pounds, there was evidence showing that changes in surfactant concentration and type
in the emulsion can significantly modify partition coefficients of many flavor compounds
(van Ruth et al., 2002a). Also, the volatility of flavor compounds is reduced in the pres-
ence of a surface-active compound than in water alone (Landy et al., 1996). On the other
hand, when surfactants formed micelles/reverse micelles in the emulsions, they were able
to incorporate flavor molecules and modify their release behaviors (Rabe et al., 2003a;
Benjamin et al., 2014).
Finally, it is important to point out that emulsifiers play significant roles on emulsion
properties, particularly on droplet size and emulsion stability, which may also show big
influences on flavor release (discussed in Section 30.4).

30.3.2.2 Thickening Agents
Thickening agents are normally added to increase emulsion viscosity, so as to get desired
emulsion stability and textural properties. Moreover, some thickeners can be used as fat
replacers to formulate fat-reduced food (Conforti et al., 1996). The addition of thicken-
ing agents can also modify flavor release. This is because the thickening agents can create
physical barriers inside emulsions against the mass transfer of flavor compounds. When
the thickeners formed a gel network, flavors could be trapped inside, and the release
was inhibited (Guichard et al., 1991). Interactions between thickening agents and flavor
compounds have also been observed, though the contribution to flavor retention was
rather small (Boland et al., 2004; Karaiskou et al., 2008). Pectin could interact with
flavor compounds through van der Waals attraction forces between the alkyl patch of
the flavor molecules and the hydrophobic domains of pectin molecules (Maier, 1970).
Moreover, hydrogen atoms in the undissociated carboxyl group of pectin could interact
with unshared electron pairs of heteroatoms and oxygen atoms of flavor molecules via
hydrogen bonding (Braudo et al., 2000). Hydroxypropyl methylcellulose (HPMC) was
shown to bind allyl disulfide (Cook et al., 2003), and it could result in lower flavor percep-
tion (Ferry et al., 2006). In an emulsified edible film matrix, κ-carrageenan was suggested
660 Food Aroma Evolution

to bind ketone with its –OH group (Marcuzzo et al., 2010). However, κ-carrageenan in
a whole milk system did not affect the headspace concentration nor the in vivo release of
the flavors (González-Tomás et al., 2007). Additionally, the texture of the food created by
a thickening agent can also affect flavor release (see Section 30.3.4.2).

30.3.2.3 Salts
People have long realized that the addition of salts can enhance flavor perception, and
the phenomenon is termed “salting-out.” For hydrophilic flavor compounds, “salting-out”
results from the reduction in the number of water molecules available to solubilize flavor
compounds (Nawar, 1971). For lipophilic flavor compounds, “salting-out” can lower their
concentration in the water phase and drive them further to the oil phase and out into the
gas phase. Therefore, higher salt concentration in emulsions can lead to a greater release
of flavor compounds (Poll and Flink, 1984; Rabe et al., 2003b; Benjamin et al., 2012b;
Bortnowska, 2012). A study on the effects of different salts in a protein-stabilized system
indicated that the addition of NaCl and KCl had a stronger salting-out effect for methyl-
butanal, hexanal, octanal, methional, and pentanone than the addition of MgCl2 and
CaCl2. It was proposed that salts affected the interactions between flavor compounds and
proteins and that NaCl and KCl can weaken the bindings to a higher level (Pérez-Juan et
al., 2006). However, when flavor compounds were incorporated into oil droplets, “salting-
out” only gave slight influence on their partition. The presence of salts in an emulsion also
fosters electrostatic screening effects, which can further modify the structure of the interfa-
cial film, and result in modified interactions between flavor compounds and the interface.
Some emulsions can even collapse (e.g., phase separation) after a certain period of storage
in salted systems, and flavor compounds could move more freely from the dispersed phase
to the continuous phase, leading to intensified flavor release. Salt-triggered flavor release
is generally concentration-dependent. However, in many cases, there is a critical concen-
tration point above which a higher salt concentration does not further affect the flavor
release process (Benjamin et al., 2012b).

30.4 EMULSION PROPERTIES AND FLAVOR RELEASE

30.4.1 Droplet Size

As discussed previously, droplet characteristics produce large effects on the physicochem-

ical properties of an emulsion. In an O/W emulsion, flavors have to move out from the
oil droplets to the water phase before transferring to the air phase, and the transfer rate
may be dependent on droplet size. Many studies have been carried out to understand the
effect of droplet size on flavor release, but no consistent conclusion can be made. Smaller
droplets have a larger interfacial area, and the travel path from the droplet center to the
interface is shorter, both of which can induce faster mass transfer of flavor molecules.
This hypothesis was confirmed by Benjamin et al. (2012b), who found that the release
rates of flavor compounds in an aggregated emulsion were lower than those in a finely
distributed emulsion. Other studies argued that smaller droplets with the larger interfa-
cial area could absorb more emulsifiers, which would result in slowed flavor release (van
Ruth et al., 2002b) and reduced air–liquid partition coefficients (Meynier et al., 2005;
van Ruth et al., 2002b).
However, the transfer of flavor compounds between the dispersed phase and con-
tinuous phase is generally thought to be very fast, especially when droplet size is reduced
 Food Emulsions as Flavor Delivery Systems 661

to micrometer or sub-micrometer range. Therefore, it may be difficult to find any sig-

nificant difference in flavor release from two emulsions with different droplet size but
within the same size range (Rabe et al., 2003a; Carey et al., 2002; Miettinen et al., 2002;
Matsumiya et al., 2015).

30.4.2 Rheological Behavior

There is evidence that flavor perception can be suppressed by increasing the viscosity of a
solution, though the magnitude of the suppression may vary among different flavors (Ferry
et al., 2006). The Stokes–Einstein equation reveals that diffusion is negatively propor-
tional to viscosity. Therefore, an increase in viscosity will reduce flavor diffusion in differ-
ent phases. A slower flavor release was observed in emulsion systems containing pectin or
gum Arabic (Karaiskou et al., 2008; Druaux and Voilley, 1997). Baines and Morris (1987)
reported that in order to produce a three-fold reduction in flavor perception using guar
gum, it was required to increase the viscosity of the system by at least two-fold. In a gel-
like viscous system, flavor release could be affected by the gel network structure created
by gelling agents. Studies showed that an increase in gel hardness could reduce the flavor
released into the headspace; that is, firm gels made from carrageenan and gelatin could
enhance flavor retention (Carr et al., 1996; Guniard and Marty, 1995). It has also been
observed that flavor release was slower in the gels with a higher gelatin concentration and
that the release rate was dependent on the rate at which the gels collapsed after melting and
chewing (Bakker et al., 1996). When different flavors were considered, hydrophobic flavor
compounds were more likely to be affected. For different gelling agents, pectin and starch
had a relatively weaker effect on flavor release than gelatin, probably because of their less
compact gel network structures (Boland et al., 2004). In a system containing Ca-alginate
gels, flavor release was highly dependent on the Ca2+-alginate ratio. With higher Ca2+
concentration, the gel strength was enhanced, and less flavor was released from the system
(Baines, Morris, 1987). Hollowood et al. (2002) revealed that the perceived strawberry
flavor in hydroxypropyl methylcellulose (HPMC) thickened solution was largely reduced
when HPMC concentration was above a critical value (known as “coil overlap concentra-
tion”), at which HPMC starts to form an entangled network.
However, other studies failed to demonstrate any difference in flavor release when
adjusting emulsion viscosity (Basaran et al., 1998; Siefarth et al., 2011). It was thus pro-
posed that the pores of the biopolymer network were much larger than the size of the
diffusing flavor molecules, and flavor molecules can travel through the entangled bio-
polymer chains without any barrier effect (Basaran et al., 1998). Another possible reason
might be that the concentration of the thickener was below the coil overlap concentra-
tion in these systems (Siefarth et al., 2011). It was also argued that, when the headspace
concentration of flavor is governed by the mass transfer at the air–liquid interface rather
than that at the oil–water interface, the viscosity of the emulsion system can only exhibit
a slight effect on flavor release (Roberts et al., 1996).

30.4.3 pH

Acids are common food ingredients, which besides other functions in composite food
systems are sometimes used as flavor enhancers. Acids in food not only influence taste
but also modify aroma release. In a soft drink system, a pH change from 5.0 to 3.0 or
662 Food Aroma Evolution

4.0 (using citric acid) resulted in a significant increase of the release of esters (isopentyl
acetate and ethyl hexanoate; Hansson et al., 2001). Nevertheless, when the pH was low-
ered to 2.0, with the further addition of citric acid did not show any effect on the release
of esters, menthone, and linalool. When the pH was regulated by a mixture of citric acid
and sodium hydroxide, there was no difference in the release of esters, menthone, and
linalool among samples at pH 3.0, 4.0, and 5.0. The authors have attributed the results
mainly to the chelating effect of the dissociated form of citric acid (RCOO −); the more
citric acid added to the solution, the greater the amount of dissociated citric acid would
be available to interact with flavors, leading to lower headspace flavor concentration in
the sample at pH 2.0. When both citric acid and sodium hydroxide were added, all the
samples (pH 2.0, 3.0, 4.0, 5.0) had a large amount of RCOO − (sample at pH 5.0 had the
highest), and therefore the difference in flavor release could not be detected (Hansson
et al., 2001).
A pH adjustment also affects the pK values of flavor compounds, which modifies the
partition and release of flavors (Bennett, 1992). Moreover, the properties of emulsifiers
(especially proteins) are highly dependent on pH. A change in pH can largely modify
the interactions between emulsifiers and flavor compounds. For example, when pH was
adjusted from 3.0 or 4.0 to 5.0, more than 30% of ethyl hexanoate, 2-decanone, and
1-octanol were released to the headspace above emulsions stabilized by β-lactoglobulin
(Benjamin et al., 2012b). In egg yolk or starch, sodium octenylsuccinate stabilized emul-
sions, an increase of pH from 3.0 to 9.0 resulted in enhanced retention of diacetyl,
which was attributed to the strengthened interactions between diacetyl and the sta-
bilizers through electrostatic attraction or hydrogen bonding at alkaline conditions
(Bortnowska, 2012).
Jouenne and Crouzet (2000) studied the effect of pH on flavor release from a
β-lactoglobulin solution and they found that the release of methyl ketones, ethyl esters,
limonene, and myrcene was significantly slowed by changing pH to 6.0 and then to 9.0,
but all the flavors were less retained when the pH value increased to 11.0. It was sug-
gested that there were enhanced interactions between flavors and β-lactoglobulin at pH
3.0–9.0, as the flexibility of the protein molecules, the surface exposure of residues, and
the unfolding of peripheric α-helix and β-sheet were likely to increase as pH increased
from 3.0 to 9.0. However, at pH 11.0, the tertiary structure of the protein was largely
modified due to alkaline denaturation (unfolding), and the flavor-protein interactions
were weakened.

30.4.4 Other Factors

Other factors, including emulsion stability and emulsion types, were poorly addressed
in the literature, but their effects on flavor release should not be neglected. For example,
creaming in emulsions resulted in more oil phase transferred to the liquid–air surface,
which then induced more flavor released into the headspace (Benjamin et al., 2012b).
Studies on flavor release from emulsions of different types showed that the transfer rates
of diacetyl toward the headspace were higher in an O/W emulsion than in a W/O emul-
sion stabilized with the same emulsifier (Salvador et al., 1994). The inclusion of a water
phase in a fat blend could reduce the headspace concentrations of butanoic acid and
hexanoic acid (Shiota et al., 2011). Nevertheless, a perception test failed to identify any
difference between O/W and W/O emulsions with the same compositions (Bakker and
Mela, 1996).
 Food Emulsions as Flavor Delivery Systems 663

30.5 EFFECT OF MOUTH CONDITIONS ON FLAVOR

RELEASE AND IN VITRO STUDIES

30.5.1 Effect of Saliva on Flavor Release

In the oral cavity, the release behavior of flavors from emulsions are mainly influenced
by saliva. Apparently, dilution of food with saliva will first disturb the partition and
mass transfer of flavors in the aqueous phase and oil phase, leading to different releasing
kinetics. Many studies conducted either in vitro or in vivo showed a significant decrease
in flavor release with an increase of saliva volume (Mehinagic et al., 2004; van Ruth and
Roozen, 2000), possibly because of the interaction between flavor compounds and the
ingredients in the saliva, especially proteins (van Ruth et al., 2001). However, there are
other flavor compounds which are released more into the headspace of the matrix–saliva
mixture with a high proportion of saliva, probably due to the salting-out effect (Deibler
et al., 2001). With the presence of many enzymes (e.g., α-amylase, lipase, and esterase)
in saliva, esters, thios, and aldehydes were reported to experience enzymatic conversion
to other flavors upon contact with saliva for some time (Buettner, 2002a,b), and losses
of some flavors may also occur (Buettner, 2002b). Several studies have reported that a
full saliva containing salts, mucin, enzymes, and so on, could enhance flavor release
(Benjamin et al., 2012b; Mao et al., 2013b). Moreover, the temperature change induced
by salivation should also be taken into account when studying flavor release in the mouth
(Benjamin et al., 2012b). On the other hand, many studies have reported that emulsion
properties can be greatly influenced by human saliva, which can then influence flavor
release. Studies conducted on emulsion stabilized either by proteins (e.g., β-lactoglobulin,
sodium caseinate, and lactoferrin) or surfactants (e.g., tween 20, SDS) showed salivas
could induce emulsion flocculation (bridge flocculation or depletion flocculation), and
finally phase separation (Vingerhoeds et al., 2005; Sigh and Sarkar, 2011). The presence
of different salts in human saliva can also modify the interfacial charge of droplets and
the pH of the emulsions (Sarkar et al., 2009). The effect of saliva on emulsion properties
have been thoroughly discussed in the review by van Aken et al. (2007).

30.5.2 Model Mouths and Throats

Due to the availability of human samples, and the large variability of the samples among
individuals, it is essential to develop mouth simulators (model mouths) to predict flavor
release. The advantage of in vitro studies is that each oral parameter can be controlled,
facilitating the understanding of its individual role on flavor release. Several model
mouths have been developed in the past two decades, each of which can mimic the oral
process to some extent. In this part of the review, we only introduce some model mouths/
throats suitable for emulsion studies. Comparison of more mouth models can be found in
the review of Salles et al. (2011).
The simplest apparatus for studying flavor release in the oral cavity was developed by
van Ruth et al. (1994) (Figure 30.2A). The model mouth contained a sample flask (70 mL)
which was surrounded by a water jacket with circulating water (37°C), and sample–saliva
mixture was added to the flask during experiments.
A constant flow of nitrogen (at the flow rate of 20 mL/min) drove flavor release from
the samples, and also flushed the headspace to a Tenax trap. The trapped flavors were later
thermally desorbed and analyzed by GC. A more delicate model mouth was proposed by
664 Food Aroma Evolution

FIGURE 30.2 Representative model mouths (A, B, C) and throat (D) for in vitro studies
of flavor release from emulsions. (A—van Ruth, et al., 1994; B—Elmore and Langley,
1996; C—Benjamin et al., 2012a; D—Weel et al., 2004a.)

Elmore and Langley (1996) (Figure 30.2B). The apparatus contained two taps on the
top, which allowed the bottom part of the system to be removed without disturbing the
gas flow to the MS detector. By turning the taps on and off, one can carefully monitor
the start and finish time of the experiment. The apparatus consisted of a magnetic stir-
rer, which facilitated the mixing of liquid foods in the samples vessel (~125 mL) and
simulated typical shear force in the mouth (~50 s−1). Benjamin et al. (2012a) designed a
novel model mouth to study the effect of tongue pressure on flavor release (Figure 30.2C).
The artificial tongue was made of elastomer rubber, and it could be driven up and down
vertically through the glass shaft (connected to an actuator), which simulated the tongue
flattening against the hard palate during eating. Different patterns of tongue movement
(ramp, pulse, or sine wave) could be applied, and the tongue pressure was monitored by a
sensitive pressure sensor attached to the bottom center of the mouth chamber.
As discussed previously, a thin layer of liquid food can remain on the surface of the
pharynx, which also contributes to flavor perception. Weel et al. (2004a) and Boelrijk
et al. (2006) attempted to develop a model throat to study flavor release from the throat
in vitro. The main part of the model throat is a vertical glass tube (surrounded by a water
jacket), with a 3 mm thick tube of rubber (with clamp) in the middle which can con-
trol the liquid–saliva mixture to flow down through the tube (Figure 30.2D). When the
 Food Emulsions as Flavor Delivery Systems 665

samples flow down and drain, a thin layer of the sample can form on the surface of the
tube. Then airflow (1 L/min) enters the tube from the bottom and moves upward, and the
flavor released from the thin layer flows out with the air into the MS detector.

30.6 NOVEL EMULSIONS FOR CONTROLLED FLAVOR RELEASE

Due to their simple structures and compositions, conventional emulsions are prone to
flocculation, coalescence, creaming, and separation. It is thus difficult to achieve con-
trolled or targeted release/delivery of active compounds incorporated in unstable emul-
sions. Therefore, novel emulsions with tailor-made structures in the water phase, oil
phase, and interface have been developed, and they can be used to deliver flavor com-
pounds in a more controlled manner.

30.6.1 Multilayer Emulsion

A multilayer emulsion is an emulsion in which the droplets are surrounded by two or

more layers (Figure 30.3), which is generally prepared through a layer-by-layer technique
(LBL). The LBL technique normally consists of two (or more) steps of layer adsorption:
a charged emulsifier is first deposited onto the surface of the droplets; then an oppositely
charged emulsifier or polymer is attracted by the previously adsorbed layer.
As the formation of a multilayer interface is mainly driven by electrostatic forces,
ways that can modify these forces, such as pH adjustment and ionic strength modifica-
tion, have been proven to alter the structures of the interfacial layer (e.g., layer thickness,
compactness, and charge density) (Guzey and McClements, 2006).
Emulsions containing oil droplets surrounded by a multilayered interface have been
reported to exhibit better stability against pH change, heating, freeze-thawing, high ionic
strength, and so on (Guzey and McClements, 2006), and they are more effective in protect-
ing nutrients from degradation and improving nutrient adsorption (Djordjevic et al., 2007;
Hou et al., 2011; Zhang and Zhong, 2015). More importantly, the multilayer emulsions

FIGURE 30.3 Different types of emulsions suitable for flavor delivery. In the self-assem-
bly structured emulsion, only the emulsion containing reverse micelles is illustrated.
666 Food Aroma Evolution

can be designed to have different responsiveness to environmental stresses, and thus allow
better-controlled delivery of the active compounds incorporated (Guzey and McClements,
2006; Mao and Miao, 2015). Mao et al. (2013b) prepared WPI-pectin multilayer emulsions
for the delivery of volatile flavor compounds, and they found that flavor was released at a
lower intensity in the multilayer emulsion than that in a WPI single layer emulsion. When
the pH of the emulsion was adjusted, the pectin could be detached from the interface,
leading to higher release rates and enhanced intensity of the flavors. In a lecithin-pectin
stabilized emulsion, the release of limonene was reduced by increasing the amount of pectin
and lecithin added (Yang et al., 2011). When multilayer emulsions were spray/freeze-dried,
greater flavor retention could be obtained (Kaasgaard and Keller, 2010; Gharsallaoui et
al., 2012). Under oral conditions, single layer (β-lactoglobulin) emulsions presented the fast
release of flavor compounds upon the addition of a small amount of saliva (due to emulsion
destabilization), whereas the multilayer (β-lactoglobulin-pectin) emulsion was only slightly
affected by saliva and presented almost the same flavor release profile as the primary emul-
sion without saliva (Benjamin et al., 2013).

30.6.2 Gelled Emulsion

In a gelled emulsion, oil droplets are trapped within gel particles (Figure 30.3), which
act as a barrier to slow the mass transfer and diffusion of the compounds incorporated.
The preparation of gelled emulsions usually involves two steps: first, a conventional O/W
emulsion is produced through homogenization; second, the emulsion is mixed with a
biopolymer solution (biopolymer can also be present in the water phase of the emul-
sion), which gels by adjusting pH, salt concentration, temperature, adding enzymes, or
applying mechanical forces (McClements, 2010). Lipophilic functional ingredients can be
dissolved in the oil droplets, and incorporated in the gel network. As the lipophilic ingre-
dients are isolated from the external environment, they usually have improved chemical
stability (e.g., oxidation). On the other hand, it is possible to get tailor-made structures
(e.g., composition, dimensions, hardness, and permeability) of the gel with different
responsiveness to environmental stresses, and thus allow controlled-target release of
the incorporated ingredients. Lee et al. (2006) prepared protein-stabilized emulsion gels
(induced by microbial transglutaminase) and found that flavor retention ranging from
60 to 100% could be obtained in emulsion gels, whereas only 5 to 25% was obtained in
ungelled emulsions when stored at 37°C for 72 h. Mao et al. (2014) explored the effect
of rheological properties of emulsion gels on flavor release. They reported that flavors
were released at lower rates and had lower air–gel partition coefficients in emulsion filled
protein gels than those in their ungelled counterparts. In gels with a higher WPI con-
tent which were more elastic, and the incorporated flavors had reduced release rates and
partition coefficients. An increase in the oil content resulted in more elastic but brittle
gels, showing however a minor influence on the release rate of the flavors. Malone and
Appelqvist (2003) designed biopolymer-gelled particles with encapsulated oil droplets
containing flavors. They found that the initial maximum flavor release in the mouth was
reduced due to the slower mass transfer of flavor molecules through the gel particles.
The release behavior can be modified by changing the droplet size or oil phase volume of
the emulsions. When the gel was broken down in a controlled manner by physiological
triggers, such as mechanical force, melting, and enzyme treatment, a controlled release
profile was obtained. For example, flavor compounds exhibited a faster release in a gela-
tinized amaranth amylose gelled emulsion than that in a gelatinized wheat starch gelled
 Food Emulsions as Flavor Delivery Systems 667

emulsion when the starch gel was broken down by α-amylase. Flavor release from the gel
was highly associated with the release of oil droplets during oral processing, and a change
in droplet size allows modified flavor release. Studies on oral processing suggested that
the release of oil droplets increased with droplet size (Guo et al., 2014), and smaller oil
droplets were more firmly entrapped into the gel matrix, even under strong mechanical
processing (Guo et al., 2013).

30.6.3 Multiple Emulsion

In a multiple emulsion, the droplets of the dispersed phase contain even smaller dispersed
droplets (Figure 30.3), such as water-in-oil-in-water (W/O/W) emulsion and oil-in-water-
in-oil (O/W/O) emulsion. A multiple emulsion has different layers deposited on both oil–
water interface and water–oil interface. Multiple emulsions are normally prepared using
a two-step homogenization method: the inner W/O (or O/W) emulsion is first produced,
which is then dispersed into the outer dispersed water (or oil) phase (Garti, 1997; Garti
and Bisperink, 1998).
A multiple emulsion contains two water (or oil) domains, which prolong the diffu-
sion length of compounds incorporated in the inner dispersed phase, leading to reduced
release rate and higher encapsulation efficiency of the core material. Moreover, it becomes
possible to effectively control the release of both hydrophilic and lipophilic ingredients
in a single system. W/O/W emulsion can be used in a fat-reduced system, as the oil con-
tent of the oil phase can be partially displaced by the inner water phase. Dickinson et
al. (1994) made a protein-stabilized W/O/W emulsion and found that the release rate of
butanol was reduced by a factor of two, in comparison to the release rate of the volatile
from an equivalent aqueous solution of butanol. Similar to the conventional emulsion,
flavor release from multiple emulsion is also affected by the emulsifier used. In an O/W/O
emulsion with gum Arabic as the inner droplet stabilizer, a combination of PGPR and
Span 80 on the outer interface can reduce flavor release to a greater extent than PGPR
alone (Cho and Park, 2003). When a multiple emulsion was spray-dried, the preparation
can be stored for a longer time with less flavor loss (Brückner et al., 2007).

30.6.4 Pickering Emulsion

Pickering emulsions are defined as emulsions, either of the O/W or W/O type, stabilized
by solid particles at the interface (Aveyard et al., 2003). Generally, they have similar
physicochemical properties as conventional emulsions, and they can be used to substitute
conventional emulsions in most cases, particularly in the case where a surfactant-free
system is essential. The interface of a Pickering emulsion is mechanically stronger, and
they can provide sufficient steric repulsion force to inhibit droplet coalescence. Different
types of silica are widely used to produce Pickering emulsions, and in the food industry,
cellulose nanocrystals (Kalashnikova et al., 2011), chitin nanocrystals (Tzoumaki et al.,
2011), starch particles (Yusoff and Murray, 2011), and flavonoid particles (Luo et al.,
2011) have proven to be suitable Pickering stabilizers.
With good physical stability, Pickering emulsions have been reported to better
deliver salts (Frasch-Melnik et al., 2010), curcumin (Wang et al., 2014), fatty acids
(Ruiz-Rodriguez et al., 2014), and retinol (Ghouchi Eskandar et al., 2009).Wang et al.
(2012) made CaCO3 particle-stabilized emulsions, and they tested the release behavior of
668 Food Aroma Evolution

limonene. The authors reported that non-encapsulated limonene released very fast and
most of the flavor released in less than 5 min, while limonene in the Pickering emulsion
exhibited slower release with a retention time over 40 min. It was possible to break up the
emulsion by dissolving CaCO3 particles in the acidic solution and thus triggered the faster
release of limonene. The Pickering emulsions, using biopolymer-based particles as stabi-
lizers, need to be studied further as potential vehicles for flavor delivery and controlled
release due to their exceptional stability properties.

30.6.5 Self-Assembly Structured Emulsion

Many emulsifiers, especially the small molecular surfactants, can not only form the inter-
facial film on the droplets, but also organize themselves into micelles (reverse micelles),
hexagonal, lamellar, and other self-assembled structures (Krog and Sparsø, 2004), which
then structure the emulsions (Figure 30.3).
Emulsifier micelles or reverse micelles can largely change partition coefficients of
flavor compounds, as they are capable of solubilizing them in their hydrophobic (hydro-
philic) domains (Rabe et al., 2003a). Van Ruth et al. (2002a) have studied the effect of
tween 20 on flavor release, and they found that tween 20 micelles can retain hydropho-
bic volatiles, and tween 20 reverse micelles can retain hydrophilic volatiles. In another
study, the formation of tween 80 micelles and PGPR (polyglycerol polyricinoleate) reverse
micelles was shown to protect citral flavor from degradation (Choi et al., 2010).
Monoglycerides (MGs) are oil-soluble emulsifiers and they can self-assemble into
lamellar, cubic, or hexagonal phases in contact with water or oil. Recent studies revealed
that the self-assembled structures can provide protection for sensitive ingredients
(Sagalowicz et al., 2006; Larsson, 2003). The application of an MG self-assembled struc-
ture to control flavor release has been reported recently. For MG structured W/O micro-
emulsions, Landy et al. (2007) reported that lipophilic volatile compounds are retained
at a greater extent in the MG structured emulsions, compared with the unstructured
W/O emulsions. In MG structured oil-in-water gel systems, Calligaris et al. (2010) dis-
covered that the equilibrium concentration of limonene in the headspace of MG gel was
significantly lower than that of a conventional emulsion. Phan et al. (2008) made MG
structured O/W emulsions with low oil content, in which delayed volatile release was
also observed. Mao et al. (2013a) found that flavors had lower headspace concentrations
and partition coefficients in MG structured emulsions than in conventional unstructured
emulsions, and that tween 20 stabilized emulsions (with MG) were more capable to slow
flavor release than WPI stabilized emulsions (with MG) (Mao et al., 2012). However, in
an oil-reduced system the role of monoglyceride crystalline structure is weakened and
lipophilic flavors are released at higher rates and of higher intensity (Mao et al., 2013a).

30.7 CONCLUSION

Extensive studies have been performed to understand flavor release from food emul-
sions and thereby to design effective colloidal dispersion systems with improved phys-
ical-chemical stability and desired flavor kinetic release profiles. Flavor release from
emulsions is affected by various factors, including ingredient composition and emulsion
structures; these factors can function either individually or synergistically. It is now
quite clear that oil plays a dominant role on flavor release, while emulsifiers and other
 Food Emulsions as Flavor Delivery Systems 669

ingredients are all likely to interact with flavor compounds and lead to altered flavor
release. Modification of emulsion structures has been regarded as a novel way to modu-
late flavor release, while maintaining the composition of the food emulsion to a large
extent. Structured emulsions, those which have been proved as effective delivery systems
for many functional food ingredients, could be also working well to deliver flavor com-
pounds. Therefore, emulsions have a great potential for applications in functional foods
(e.g., low fat, low salt, and low sugar products) with favorable flavor characteristics.
However, due to the complexity of food matrices, the diversity of flavor compounds
characteristics, and the miscellaneous responses of emulsion structures during process-
ing, transportation, and consumption, our understanding about flavor release from
food emulsions is rather limited, and the overall process is still very difficult to control.
Moreover, information obtained from studies on model systems does not fully reflect
flavor release behavior from a real food matrix. When flavor perception is concerned,
more well-designed models are required to understand the effect of oral conditions on
emulsion structures and flavor release, and to fully unravel the relationships between
texture and flavor perception.

REFERENCES

Aveyard, R., Binks, B.P., and Clint, J.H. (2003). Emulsions stabilized solely by solid col-
loidal particles. Adv. Colloid Interface Sci. 100–2: 503–46.
Baines, Z.V. and Morris, E.R. (1987). Flavour/taste perception in thickened systems: The
effect of guar gum above and below C*. Food Hydrocolloids 1: 197–205.
Bakker, J., Brown, W.E., Hills, B.M., Boudaud, N., Wilson, C.E., and Harrison, M.
(1996). Effect of the food matrix on flavour release and perception. In: Flavour
Science: Recent Developments, pp. 369–74. Taylor, A.J. and Mottram, D.S., Eds.,
Royal Society of Chemistry, Cambridge.
Bakker, J. and Mela, D.J. (1996). Effect of emulsion structure on flavour release and taste
perception. ACS Symp. Ser. 633: 36–47.
Basaran, T., Demetriades, K., and McClements, D.J. (1998). Ultrasonic imaging of grav-
itational separation in emulsions. Colloids Surf. A Physicochem. Eng. Asp. 136:
169–81.
Bayarri, S., Taylor, A.J., and Hort, J. (2006). The role of fat in flavour perception: Effect
of partition and viscosity in model emulsions. J. Agric. Food Chem. 54: 8862–8.
Benjamin, O., Silcock, P., Beauchamp, J., Buettner, A., and Everett, D.W. (2014).
Emulsifying properties of legume proteins compared to β-lactoglobulin and tween
20 and the volatile release from oil-in-water emulsions. J. Food Sci. 79: E2014–22.
Benjamin, O., Silcock, P., Beauchamp, J., and Everett, D.W. (2013). Volatile release and
structural stability of β-lactoglobulin primary and multilayer emulsions under simu-
lated oral conditions. Food Chem. 140: 124–34.
Benjamin, O., Silcock, P., Kieser, J.A., Waddell, J.N., and Everett, D.W. (2012a).
Development of a model mouth containing an artificial tongue to measure the
release of volatile compounds. Innov. Food Sci. Emerg. 15: 96–103.
Benjamin, O., Silcock, P., Leus, M., and Everett, D.W. (2012b). Multilayer emulsions as
delivery systems for controlled release of volatile compounds using pH and salt trig-
gers. Food Hydrocolloids 27: 109–18.
Bennett, C.J. (1992). Formulation low-fat food with good taste. Cereal Foods World 37:
429–32.
670 Food Aroma Evolution

Boelrijk, A.E.M, Smit, G., and Weel, K.G.C. (2006). Flavour release from liquid food
products. In: Flavour in Food, pp. 260–84. Etievant, P. and Voilley, A., Eds.,
Woodhead Limited Publishing, Cambridge.
Boland, A.B., Buhr, K., Giannouli, P., and van Ruth, S.M. (2004). Influence of gelatine,
starch, pectin and artificial saliva on the release of 11 flavour compounds from
model gel systems. Food Chem. 86: 401–11.
Bortnowska, G. (2012). Effects of pH and ionic strength of NaCl on the stability of diace-
tyl and (-)-α-pinene in oil-in-water emulsions formed with food-grade emulsifiers.
Food Chem. 135: 2021–28.
Bosman, J.F. (1980). Physiology of the mouth, pharynx, and oesophagus. In:
Otolaryngology, pp. 319–31. Paparella, M.M. and Shumrick, D.A., Eds., W.B.
Saunders, Philadelphia, PA.
Braudo, E.E., Plashchina, I.G., Kobak, V.V., Golovnya, R.V., Zhuravleva, I.L., and
Krikunova, N.I. (2000). Interactions of flavour compounds with pectic substances.
Nahrung—Food 44: 173–7.
Brückner, M., Bade, M., and Kunz, B. (2007). Investigation into the stabilization of a
volatile aroma compound using a combined emulsification and spray drying pro-
cess. Eur. Food Res. Technol. 226: 137–46.
Buettner, A. (2002a) Influence of human saliva on odorant concentrations. 2. Aldehydes,
alcohols, 3-alkyl-2-methoxypyrazines, methoxyphenols, and 3-hydroxy-4,
5-dimethyl-2(5H)-furanone. J. Agric. Food Chem. 50: 7105–10.
Buettner, A. (2002b) Influence of human salivary enzymes on odorant concentration
changes occurring in vivo. I. Esters and thiols. J. Agric. Food Chem. 50: 3283–9.
Buettner, A., Beer, A, Hanning, C., Settles, M., and Schieberle, P. (2002). Physiological
and analytical studies on flavour perception dynamics as induced by the eating and
swallowing process. Food Qual. Prefer. 13: 497–504.
Buttery, R.G., Guadagni, D.G., and Ling, L.C. (1973). Flavour compounds: Volatilities
in vegetable oil and oil-in-water mixtures. Estimation of odor threshold. J. Agric.
Food Chem. 21: 198–201.
Calligaris, S., Pieve, S.D., Arrighetti, G., and Barba, L. (2010). Effect of the structure of
monoglyceride–oil–water gels on aroma partition. Food Res. Int. 43: 671–7.
Carey, M.E., Asquith, T., Linforth, R.S.T., and Taylor, A. (2002). Modelling the partition of
volatile aroma compounds from a cloud emulsion. J. Agric. Food Chem. 50: 1985–90.
Carr, J., Baloga, D., Guinard, J.X., Lawter, L., Marty, C., and Cordelia, S. (1996). The
effect of gelling agent type and concentration on flavour release in model systems.
In: Flavour–Food Interactions, pp. 98–108. McGorrin, R.J. and Leland, J.V., Eds.,
American Chemical Society, Washington, DC.
Charles, M., Lambert, S., Brondeur, P., Courthaudon, J.L., and Guichard, E. (2000).
Influence of formulation and structure of an oil-in-water emulsion on the flavour
release. In: Flavour Release, pp. 342–54. Roberts, D.D. and Taylor, A.J., Eds.,
American Chemical Society, Washington, DC.
Cho, Y.H. and Park, J. (2003). Evaluation of process parameters in the O/W/O multiple
emulsion method for flavor encapsulation. J. Food Sci. 68: 534–8.
Choi, S.J., Decker, E.A., Henson, L. Popplewell, L.M., and McClements D.J. (2010).
Inhibition of citral degradation in model beverage emulsions using micelles and
reverse micells. Food Chem. 122: 111–6.
Conforti, F.D., Charles, S.A., and Dunkan, S.E. (1996). Sensory evaluation and con-
sumer acceptance of carbohydrate-based fat replacers in biscuits. J. Consum. Stud.
Home Econ. 20: 285–96.
 Food Emulsions as Flavor Delivery Systems 671

Cook, D.J., Linforth, R.S.T., and Taylor, A.J. (2003) Effects of hydrocolloid thickeners
on the perception of savory flavours. J. Agric. Food Chem. 51: 3067–72.
Deibler, K.D., Lavin, E.H., Linforth, R.S.T., Taylor, A.J., and Acree, T.E. (2001).
Verification of a mouth simulator by in vivo measurements. J. Agric. Food Chem.
49: 1388–93.
de Roos, K.B. (1997). How lipids influence food flavour. Food Technol. 51: 60–2.
de Roos, K.B. (2000). Physicochemical models of flavour release from foods. In: Flavour
Release, pp. 126–41. Roberts, D.D. and Taylor, A.J., Eds., American Chemical
Society, Washington, DC.
de Roos, K.B. and Wolswinkel, K. (1994). Non-equilibrium partition model for predict-
ing flavour release in the mouth. In: Trends in Flavour Research, pp. 15–32. Maarse,
H. and van der Heij, D.G., Eds., Elsevier Science, Amsterdam, the Netherlands.
Dickinson, E., Evison, J., Gramshaw, J.W., and Schwope, D. (1994). Flavor release from
a protein-stabilized water-in-oil-in-water emulsion. Food Hydrocolloids 8: 63–7.
Djordjevic, D., Cercaci, L., Alamed, J., McClements, D.J., and Decker, E.A. (2007).
Chemical and physical stability of citral and limonene in sodium dodecyl sulfate-
chitosan and gum Arabic-stabilized oil-in-water emulsions. J. Agric. Food Chem.
55: 3585–91.
Doyen, K., Carey, M., Linforth, R.S.T., Marin, M., and Taylor, A.J. (2001). Volatile
release from an emulsion: Headspace and in-mouth studies. J. Agric. Food Chem.
49: 804–10.
Druaux, C. and Voilley, A. (1997). Effect of food composition and microstructure on
volatile flavour release. Trends Food Sci. Techol. 8: 364–8.
Dubois, C., Sergent, M., and Voilley, A. (1996). Flavouring of complex media: A model
cheese sample. In: Flavour–Food Interactions, pp. 217–26. McGorrin, R. and
Ledand, J., Eds., American Chemical Society, Washington, DC.
Elmore, J.S. and Langley, R.K. (1996). Novel vessel for the measurement of dynamic fla-
vor release in real time from liquid foods. J. Agric. Food Chem. 44: 3560–3.
Ferry, A.L., Hort, J. Mitchell, J.R., Cook, D.J., Lagarrigue, S., and Pamies, B. (2006).
Viscosity and flavour perception: Why is starch different from hydrocolloids? Food
Hydrocolloids 20: 855–62.
Frank, D., Appelqvist, I., Piyasiri, U., Wooster, T.J., and Delahunty, C. (2011). Proton
transfer mass spectrometry and spectrometry and time intensity perceptual mea-
surement of flavour release from lipid emulsions using trained human subjects. J.
Agric. Food Chem. 59: 4891–903.
Frasch-Melnik, S., Norton, I.T., and Spyropoulos, F. (2010). Fat-crystal stabilised W/O
emulsions for controlled salt release. J. Food Eng. 98: 437–42.
Garti, N. (1997). Progress in stabilization and transport phenomena of double emulsions
in food applications. LWT—Food Sci. Technol. 30: 222–35.
Garti, N. and Bisperink, C. (1998). Double emulsions: Progress and applications. Current
Opin. Colloid Interface Sci. 3: 657–67.
Gastaldelli, A. and Basta, G. (2010). Ectopic fat and cardiovascular disease: What is the
link? Nutr. Metab. Cardiovasc. Dis. 20: 481–90.
Gharsallaoui, A., Roudaut, G., Beney, L., Chambin, O., Voilley, A., and Saurel, R.
(2012). Properties of spray-dried food flavors microencapsulation with two-lay-
ered membranes: Roles of interfacial interaction and water. Food Chem. 132:
1713–20.
Ghosh, S., Peterson, D., and Coupland, J.N. (2007). Aroma release from solid droplet
emulsions: Effect of lipid type. J. Am. Oil Chem. Soc. 84: 1001–14.
672 Food Aroma Evolution

Ghosh, S., Peterson, D.G., and Coupland, J.N. (2006). Effects of droplet crystallization
and melting on the aroma release properties of a model oil-in-water emulsion. J.
Agric. Food Chem. 54: 1829–37.
Ghouchi Eskandar, N., Simovic, S., and Prestidge, C.A. (2009). Chemical stability and
phase distribution of all-trans-retinol in nanoparticle coated emulsions. Int. J.
Pharm. 376: 186–94.
Giroux, H.J., Perreault, V., and Britten, M. (2007). Characterization of hydrophobic fla-
vour release profile in oil-in-water emulsion. J. Food Sci. 72: S125–9.
González-Tomás, L., Bayarri, S., Taylor, A.J., and Costell, E. (2007). Flavour release and
perception from model dairy custards. Food Res. Int. 40: 520–8.
Gremli, H.A. (1974). Interaction of flavour compounds with soy protein. J. Am. Oil
Chem. Soc. 51: 95A–97A.
Guichard, E. (2002). Interactions between flavour compounds and food ingredients and
their influence on flavour perception. Food Rev. Int. 18: 49–70.
Guichard, E., Issanchou, S., Descourvieres, A., and Etievant, P. (1991). Pectin concentra-
tion, molecular weight and degree of esterification: Influence on volatile composi-
tion and sensory characteristics of strawberry jam. J. Food Sci. 56: 1621–7.
Guichard E. and Langourieux, S. (2000). Interactions between β-lactoglobulin and fla-
vour compounds. Food Chem. 71: 301–8.
Guinard, J.X. and Marty, C. (1995). Time-intensity measurement of flavour release from
a model gel system: Effect of gelling agent type and concentration. J. Food Sci. 60:
727–30.
Guo, Q., Ye, A., Lad, M., Dalgleish, D., and Singh, H. (2013). The breakdown properties
of heat-set whey protein emulsion gels in the human mouth. Food Hydrocolloids
33: 215–24.
Guo, Q., Ye, A., Lad, M., Dalgleish, D., and Singh, H. (2014). Behaviour of whey protein
emulsion gel during oral and gastric digestion. Soft Matter 10: 4173–83.
Guyot, C., Bonnafont, C., Lesschaeve, I., Issanchou, S., Voilley, A., and Spinnler, H.E.
(1996). Effect of fat content on odor intensity of three aroma compounds in model
emulsions: δ-decalactone, diacetyl, and butyric acid. J. Agric. Food Chem. 44: 2341–8.
Guzey, D. and McClements, D.J. (2006). Formation, stability and properties of mul-
tilayer emulsions for application in the food industry. Adv. Colloid Interface Sci.
128–130: 227–48.
Hansson, A., Andersson, J., Leufvén, A., and Pehrson, K. (2001). Effect of changes in pH
on the release of flavour compounds from a soft drink-related model system. Food
Chem. 74: 429–35.
Harrison, M. (1998). Effect of breathing and saliva flow on flavour release from liquid
foods. J. Agric. Food Chem. 46: 2727–35.
Harrison M. and Hills, B.P. (1997). Mathematical model of flavour release from liquids
containing aroma-binding macromolecules. J. Agric. Food Chem. 45: 1883–90.
Harvey, B., Druaux, C., and Voilley, A. (1995). Effect of protein on the retention and
transfer of aroma compounds at the lipid–water interface. In: Food Macromolecules
and Colloids, pp. 154–63. Dickinson, E. and Lorient, D., Eds., The Royal Society
of Chemistry, Cambridge.
Hollowood, T.A., Linforth, R.S.T., and Taylor, A.J. (2002). The effect of viscosity on the
perception of flavour. Chem. Senses 27: 583–91.
Hou, Z., Gao, Y., Yuan, F., Liu, Y., Li, C., and Xu, D. (2011). Investigation into the phys-
icochemical stability and rheological properties of β-carotene emulsion stabilized by
soybean soluble polysaccharides and chitosan. J. Agric. Food Chem. 58: 8604–11.
 Food Emulsions as Flavor Delivery Systems 673

Innocente, N., Marchesini, G., and Biasutti, M. (2014). Effect of high-pressure homog-
enization on the retention of selected aroma compounds in model dairy emulsions.
Int. J. Food Sci. Techol. 49: 1992–2000.
Jouenne, E. and Crouzet, J. (2000). Effect of pH on retention of aroma compounds by
β-lactoglobulin. J. Agric. Food Chem. 48: 1273–7.
Kaasgaard, T. and Keller, D. (2010). Chitosan coating improves retention and redisper-
sity of freeze-dried flavor oil emulsions. J. Agric. Food Chem. 58: 2446–54.
Kalashnikova, I., Bizot, H., Cathala, B., and Capron, I. (2011). New Pickering emulsions
stabilized by bacterial cellulose nanocrystals. Langmuir 27: 7471–9.
Karaiskou, S., Blekas, G., and Paraskevopoulou, A. (2008). Aroma release from gum
Arabic or egg yolk-xanthan-stabilized oil-in-water emulsions. Food Res. Int. 41:
637–45.
Krog, N.J. and Sparsø, F.V. (2004). Food emulsifiers: Their chemical and physical prop-
erties. In: Food Emulsions, pp. 44–90. Friberg, S.E., Larsson, K. and Sjöblom, J.,
Eds., Marcel Dekker Inc., New York, NY.
Kühn, J., Zhu, X.Q., Considine, T., and Singh, H. (2007). Binding of 2-nonanone and
milk proteins in aqueous model systems. J. Agric. Food Chem. 55: 3599–604.
Land, D.G. (1978). Some factors influencing the perception of flavour contributing
substances in food in progress. In: Flavour Research, pp. 53–56. Land, D.G. and
Nursten, H., Eds., Applied Sciences Publishers, London.
Land, D.G. (1996). Perspectives on the effects of interactions on flavour perception: An
overview. In: Flavour–Food Interactions, pp. 2–11. McGorrin, R.J. and Leland, J.,
Eds., American Chemical Society, Washington, DC.
Landy, P., Courthaudon, J.L., Dubois, C., and Voilley, A. (1996). Effect of interface in
model food emulsions on the volatility of aroma compounds. J. Agric. Food Chem.
44: 526–30.
Landy, P., Pollien, P., Rytz, A., Leser, M.E., Sagalowicz, L., Blank, I., and Spadone, J.C.
(2007). Model studies on the release of aroma compounds from structured and non-
structured oil systems using proton-transfer reaction mass spectrometry. J. Agric.
Food Chem. 55: 1915–22.
Larsson, K. (2003). Molecular organization in lipids and emulsions. In: Food Emulsions
(4th ed.), pp. 93–106. Friberg, S., Larsson, K., and Sjöblom, J., Eds., Marcel Dekker
Inc., New York, NY.
Lee, H.A., Choi, S.J., and Moon, T.W. (2006). Characteristics of sodium caseinate- and
soy protein isolate-stabilized emulsion-gels formed by microbial transglutaminase.
J. Food Sci. 71: C352–7.
Lian, G.P., Malone, M.E., Homan, J.E., and Norton, I.T. (2004). A mathematical model
of volatile release in mouth from the dispersion of gelled emulsion particles. J.
Control Release 98: 139–55.
Linforth, R.S.T. and Taylor, A.J. (2000). Persistence of volatile compounds in the breath
after their consumption in aqueous solutions. J. Agric. Food Chem. 48: 5419–23.
Lubbers, S., Landy, P., and Voilley, A. (1998). Retention and release of aroma compounds
in foods containing proteins. Food Technol. 52: 68–74, 208–14.
Lübke, M., Guichard, E., and Le Quéré, J.L. (2000). Infrared spectroscopic study of
β-lactoglobulin interactions with flavour compounds. In: Flavour Release, pp. 282–92.
Roberts, D.D. and Taylor, A.J., Eds., American Chemical Society, Washington, DC.
Luo, Z., Murray B.S., Yusoff, A., Morgan, M.R.A., Povey, M.J.W., and Day, A.J. (2011)
Particle-stabilizing effects of flavonoids at the oil-water interface. J. Agric. Food
Chem. 59: 2636–45.
674 Food Aroma Evolution

Maier, H.G. (1970). Volatile flavouring substances in food stuffs. Angew. Chem. Int. Ed.
Engl. 9: 917–26.
Maier, H.G. (1975). Binding of volatile aroma substances to nutrients and foodstuffs.
In: Aroma Research, pp. 143–57. Maarse, H. and Groenen, P.J., Eds., Center for
Agricultural Publishing and Documentation, Wageningen, the Netherlands.
Malone, M. and Appelqvist, I.A.M. (2003). Gelled emulsion particles for the controlled
release of lipohilic volatiles during eating. J. Control Release 90: 227–41.
Mao, L. and Miao, S. (2015). Structuring food emulsions to improve nutrient delivery
during digestion. Food Eng. Rev., in press. doi:10.1007/s123903-015-9108-0.
Mao, L., O’Kennedy, B.T., Roos, Y.H., Hannon, J.A., and Miao, S. (2012). Effect of
monoglyceride self-assembled structure on emulsion properties and subsequent fla-
vor release. Food Res. Int. 48: 233–40.
Mao, L., Roos, Y.H., and Miao, S. (2013a). Volatile release from self-assembly structured
emulsions: Effect of monoglyceride content, oil content and oil type. J. Agric. Food
Chem. 61: 1427–34.
Mao, L., Roos, Y.H., and Miao, S. (2014). Study on the rheological properties and volatile
release of cold-set emulsion-filled protein gels. J. Agric. Food Chem. 62: 11420–8.
Mao, L., Roos, Y.H., O’Callaghan, D.J., and Miao, S. (2013b). Volatile release from
whey protein isolate-pectin multilayer stabilized emulsions: Effect of pH, salt, and
artificial salivas. J. Agric. Food Chem. 61: 6231–9.
Marcuzzo, E., Sensidoni, A., Debeaufort, F., and Voilley, A. (2010). Encapsulation of
aroma compounds in biopolymeric emulsion based edible films to control flavour
release. Carbohydr. Polym. 80: 984–8.
Matsumiya, K., Sasaki, M., Murakami, H., and Matsumura, Y. (2015). Oil droplet
coalescence not necessarily affects the flavor release from oil-in-water emulsions.
Colloids Surf. A Physicochem. Eng. Asp. 475: 19–26.
McClements, D.J. (2005). Food Emulsions: Principles, Practices and Techniques (2nd
ed.). CRC Press, Boca Raton, FL.
McClements, D.J. (2010). Emulsion design to improve the delivery of functional lipo-
philic components. Annu. Rev. Food Sci. Techol. 1: 241–69.
McClements, D.J. and Li, Y. (2010). Structured emulsion-based delivery systems:
Controlling the digestion and release of lipophilic food components. Adv. Colloid
Interface Sci. 159: 213–28.
McNulty, P.B. and Karel, M. (1973). Factors affecting flavour release and uptake in O/W
emulsions. I. Release und uptake models. J. Food Techol. 8: 309–18.
Mehinagic, E., Prost, C., and Demaimay, M. (2004). Optimization of extraction of apple
aroma by dynamic headspace and influence of saliva on extraction of volatiles. J.
Agric. Food Chem. 52: 5175–82.
Meynier, A., Lecoq, C., and Genot, C. (2005). Emulsification enhances the retention of
esters and aldehydes to a greater extent than changes in the droplet size distribution
of the emulsion. Food Chem. 93: 153–9.
Miettinen S.M., Tuorila, H., Piironen, V., Vehkalahti, K., and Hyvönen, L. (2002). Effect
of emulsion characteristics on the release of aroma as detected by sensory evalu-
ation, static headspace gas chromatography, and electronic nose. J. Agric. Food
Chem. 50: 4232–9.
Nawar, W.W. (1971). Variables affecting composition of headspace aroma. J. Agric. Food
Chem. 19: 1057–9.
Normand, V., Avison, S., and Parker, A. (2004). Modelling the kinetics of flavour release
during drinking. Chem. Senses 29: 235–45.
 Food Emulsions as Flavor Delivery Systems 675

Pérez-Juan, M., Flores, M., and Toldrá, F. (2006). Effect of ionic strength of different
salts on the binding of volatile compounds to porcine soluble protein extracts in
model systems. Food Res. Int. 40: 687–93.
Perreault, V., Britten, M., Turgeon, S.L., Seuvree, A.M., Cayot, P., and Voilley, A. (2010).
Effects of heat treatment and acid-induced gelation on aroma release from flavoured
skim milk. Food Chem. 118: 90–5.
Phan, V.A., Liao, Y.C., Antille, N., Sagalowicz, L., Robert, F., and Godinot, N. (2008).
Delayed volatile compound release properties of self-assembly structures in emul-
sions. J. Agric. Food Chem. 56: 1072–7.
Poll, L. and Flink, J.M. (1984). Aroma analysis of apple juice-influence of salt addition
on headspace volatile composition as measured by gas-chromatography and corre-
sponding sensory evaluation. Food Chem. 13: 193–207.
Rabe, S., Krings, U., and Berger, R.G. (2003a). Influence of oil-in-water emulsion charac-
teristics on initial dynamic flavour release. J. Sci. Food Agric. 83: 1124–33.
Rabe, S., Krings, U., and Berger, R.G. (2003b). Initial dynamic flavour release from
sodium chloride solutions. Eur. Food Res. Technol. 218: 32–9.
Relkin, P., Fabre, M., and Guichard, E. (2004). Effect of fat nature and aroma com-
pound hydrophobicity on flavour release from complex food emulsions. J. Agric.
Food Chem. 52: 6257–63.
Roberts, D.D, Elmore, J.S, Langley, K.R., and Bakker, J. (1996). Effects of sucrose, guar
gum and carboxymethylcellulose on the release of volatile flavour compounds under
dynamic conditions. J. Agric. Food Chem. 44: 1321–6.
Roberts, D.D. and Pollien, P. (2000). Relative influence of milk components on flavor
compound volatility. In: Flavour Release, pp. 321–32. Roberts, D.D. and Taylor,
A.J., Eds., American Chemical Society, Washington, DC.
Roberts, D.D., Pollien, P., Antille, N., Lindinger, C., and Yeretzian, C. (2003a).
Comparison of nosespace, headspace, and sensory intensity ratings for the evalua-
tion of flavour absorption by fat. J. Agric. Food Chem. 51: 3636–42.
Roberts, D.D., Pollien, P., and Watzke, B. (2003b). Experimental and modelling studies
showing the effect of lipid type and level on flavour release from milk-based liquid
emulsions. J. Agric. Food Chem. 51: 189–95.
Rogacheva, S., Espinosa-Diaz, M.A., and Voilley, A. (1999). Transfer of aroma com-
pounds in water–lipid systems: Binding tendency of β-lactoglobulin. J. Agric. Food
Chem. 47: 259–63.
Roudnitzky, N., Irl, H., Roudaut, G., and Guichard, E. (2003). Influence of fat nature on
flavour release. In: Flavour Research at the Dawn of the Twenty-First Century, 10th
Weurman Flavour Research Symposium, pp. 136–9. Le Quéré, J.L. and Etiévant,
P.X., Eds., Lavoisier, Paris, France.
Ruiz-Rodriguez, P., Meshulam, D., and Lesmes, U. (2014) Characterization of Pickering
O/W emulsions stabilized by silica nanoparticles and their responsiveness to in vitro
digestion conditions. Food Biophys. 9: 406–15.
Sagalowicz, L., Leser, M.E., Watzke, H.J., and Michel, M. (2006). Monoglyceride self-
assembly structures as delivery vehicles. Trends Food Sci. Techol. 17: 204–14.
Salles, C., Chagnon, M.C., Feron, G., Guichard, E., Laboure, H., Morzel, M., Semon, E.,
Tarrega, A., and Yven, C. (2011). In-mouth mechanisms leading to flavour release
and perception. Crit. Rev. Food Sci. Nutr. 51: 67–90.
Salvador, D., Bakker, J., Langley, K.R., Potjewijd, R., Martin, A., and Elmore, J.S.
(1994). Flavour release of diacetyl from water, sunflower oil and emulsions in model
systems. Food Qual. Prefer. 5: 103–7.
676 Food Aroma Evolution

Sarkar, A., Goh, K.K.T., and Singh, H. (2009). Colloidal stability and interactions of
milk-protein-stabilized emulsions in an artificial saliva. Food Hydrocolloids 23:
1270–8.
Seuvre, A.M., Espinosa-Díaz, M.A., and Voilley, A. (2000). Influence of the food matrix
structure on the retention of aroma compounds. J. Agric. Food Chem. 48: 4296–300.
Shiota, M., Isogai, T., Iwasawa, A., and Kotera, M. (2011). Model studies on volatile
release from different semisolid fat blends correlated with changes in sensory per-
ception. J. Agric. Food Chem. 59: 4904–12.
Siefarth, C., Tyapkova, O., Beauchamp, J., Schweiggert, U., and Buettner, A. (2011).
Influence of polyos and bulking agents on flavour release from low-viscosity solu-
tions. Food Chem. 129: 1462–8.
Sigh, H. and Sarkar, A. (2011). Behaviour of protein-stabilised emulsions under various
physiological conditions. Adv. Colloid Interface Sci. 165: 47–57.
Taylor, A.J. (1998). Physical chemistry of flavour. Int. J. Food Sci. Techol. 33: 53–62.
Tzoumaki, M.V., Moschakis, T., Kiosseoglou, V., and Biliaderis, C.G. (2011). Oil-in-
water emulsions stabilized by chitin nanocrystal particles. Food Hydrocolloids 25:
1521–9.
van Aken, G.A., Vingerhoeds, M.H., and de Hoog, E.H.A. (2007). Food colloids under
oral conditions. Curr. Opin. Colloid Interface Sci. 12: 251–62.
van Ruth, S.M., de Vries, G., Geary, M., and Giannouli, P. (2002a) Influence of composi-
tion and structure of oil-in-water emulsions on retention of aroma compounds. J.
Sci. Food Agric. 82: 1028–35.
van Ruth, S.M, Grossmann, I., Geary, M., and Delahunty, C.M. (2001). Interactions
between artificial saliva and 20 aroma compounds in water and oil model systems.
J. Agric. Food Chem. 49: 2409–13.
van Ruth, S.M., King, C., and Giannouli, P. (2002b). Influence of lipid fraction, emulsi-
fier fraction, and mean particle diameter of oil-in-water emulsions on the release of
20 aroma compounds. J. Agric. Food Chem. 50: 2365–71.
van Ruth, S.M. and Roozen, J.P. (2000). Influence of mastication and saliva on aroma
release in a model mouth system. Food Chem. 71: 339–45.
van Ruth, S.M., Roozen, J.P., and Cozijnsen, J.L. (1994). Comparison of dynamic head-
space mouth model systems for favour release from rehydrated bell pepper cuttings.
In: Trends in Favour Research, pp. 59–64. Maarse, H. and van der Heij, D.G., Eds.,
Elsevier, Amsterdam.
Velikow, K.P. and Pelan, E. (2008). Colloidal delivery systems for micronutrients and
nutraceuticals. Soft Matter 4: 1964–80.
Vingerhoeds, M.H., Blijdenstein, T.B.J., Zoet, F.D., and van Aken, G.A. (2005). Emulsion
flocculation induced by saliva and mucin. Food Hydrocolloids 19: 915–22.
Wang, M.S., Chaudhari, A., Pan, Y., Young, S., and Nitin, N. (2014). Controlled
release of natural polyphenols in oral cavity using starch Pickering emulsion. MRS
Proceedings, 1688, mrss14-1688-y08-11. doi:10.1557/opl.2014.482.
Wang X., Zhou W., Cao J., Liu W., and Zhu S. (2012). Preparation core-shell CaCO3
capsules via Pickering emulsion templates. J. Colloid Interf. Sci. 372: 24–31.
Weel, K.G.C, Boelrijk, A.E.M, Burger, J.J., Jacobs, M.A., Gruppen, H., Voragen, A.G.,
and Smit, G. (2004a). New device to simulate swallowing and in vivo aroma release
in the throat from liquid and semiliquid food systems. J. Agric. Food Chem. 52:
6564–71.
 Food Emulsions as Flavor Delivery Systems 677

Weel, K.G.C, Boelrijk, A.E.M, Burger, J.J., Jacobs, M.A., Gruppen, H., Voragen, A.G.,
and Smit, G. (2004b). Effect of emulsion properties on release of esters under static
headspace, in vivo, and artificial throat conditions in relation to sensory intensity. J.
Agric. Food Chem. 52: 6572–7.
Welsh, F.W. and Williams, R.E. (1989). The use of vegetable oils to recover compounds
from aqueous solutions. J. Chem. Techol. Biotechnol. 46: 169–78.
Widder, S. and Fisher, N. (1996). Measurement of the influence of food ingredients on
flavour release by headspace chromatography–olfactometry. In: Flavour Science.
Recent Developments, pp. 405–12. Taylor, A.J. and Mottram, D.S., Eds., the Royal
Society of Chemistry, Cambridge.
Wilde, P., Mackie, A., Husband, F., Gunning, P., and Morris, V. (2004). Proteins and
emulsifiers at liquid interfaces. Adv. Colloid Interface Sci. 20: 63–71.
Wong, R.C., Elias, R.J., Lambert, J.D., Relkin, P., and Coupland, J.N. (2013). Enzyme
triggered release of aroma molecules from oil-in-water emulsions. Colloids Surf. A
Physicochem. Eng. Asp. 42: 19–23.
Wu, S.Y., Pérez, M.D., Puyol, P., and Sawyer, L. (1999). Beta-lactoglobulin binds palmi-
tate within its central cavity. J. Biol. Chem. 274: 170–4.
Yang, T.S., Liu, T.T., and Hu, T.F. (2011). Effects of lecithin and pectin on riboflavin-
photosensitized oxidation of orange oil in a multilayered oil-in-water emulsion. J.
Agric. Food Chem. 59: 9344–50.
Yusoff, A. and Murray, B.S. (2011). Modified starch granules as particle stabilizers of
oil-in-water emulsions. Food Hydrocolloids 25: 42–55.
Zhang, Y. and Zhong, Q. (2015). Multiple-layered coatings on L-glutamine solid mic-
roparticles for the retention during storage and enteric delivery during in vitro diges-
tions. Food Hydrocolloids 43: 584–92.
 Chapter 31
Relationship between
Structure and Odor
Valentina Villalobos Coa, Vito Lubes, Johannes Polster,
Maria Monteiro de Araújo Silva, and Giuseppe Lubes

CONTENTS

31.1 Introduction 679

31.2 Theory on Aroma Perception 680
31.3 Structure–Odor Relationship 682
31.3.1 Influence of the Chain Length 683
31.3.2 Influence of the Functional Groups 685
31.3.3 Influence of the Unsaturation Degree and the Double Bonds Position 688
31.3.4 Influence of the Stereochemistry 689
31.4 Application of QSAR in Aroma Descriptors 689
31.5 Conclusion 690
References 691

31.1 INTRODUCTION

One of the key attributes for the acceptability of food products is its aroma. This prop-
erty can drastically influence manufacturing processes for further optimization pro-
cesses. This leads to the development of new products which aim to be more acceptable
to consumers (Lubes and Goodarzi, 2017). It consists of the emission of one particular
molecule or a defined combination of many molecules providing to the food, in this case,
its characteristic aroma. Volatile compounds responsible for odors and/or aroma are rep-
resented by a wide range of stable chemical compounds from different classes, such as,
alcohols, ketones, esters, phenols, lactones, pyrazines, and so on, with physical proper-
ties, for instance, low-molecular weight, low boiling point, and/or high vapor pressure in
natural conditions (Lubes and Goodarzi, 2018). So far, approximately 10,000 different
volatile compounds have been identified in foods and beverages. The complexity in the
aroma composition depends on the nature of the food. Complex foods, such as coffee,
for example, contain more than 1000 aroma compounds (Mottram and Elmore, 2003).
Nevertheless, not all of them contribute to the characteristic aroma, which will depend
on many factors, including odor character, concentration in the product, odor thresh-
old, vapor pressure, adsorption/absorption on the food matrix, chemical interaction with
other components in the matrix, synergism with other volatiles, and so on (Mottram and
Elmore, 2003).

679
680 Food Aroma Evolution

From an industrial point of view, the flavor and fragrance market is a billion-dollar
business with over USD$15 billion of sales per year with a compound annual growth rate
(CAGR) of ̴4 –6%, and for 2025 it is expected to reach USD$28.65 billion (Lubes and
Goodarzi, 2017). Many factors are responsible for boosting this market growth. One of
the most important factors is the change in consumer preferences for a healthier and sus-
tainable process where natural products are more available (Lubes and Goodarzi, 2017;
Mottram and Elmore, 2003). Fragrances and flavors, synthetically created or extracted
by companies from natural sources, are incorporated in our daily lives from toiletry to
laundry products up to food additives. Those chemical compounds extracted from plants,
for example, are built up via different biochemical pathways. One of the most important
examples are terpenoids, which are some of the largest and diverse natural chemicals
extracted from plants. Terpenoids comprise more than 30,000 chemical compounds
and are subclassified as monoterpenes (C10), sesquiterpenes (C15), and diterpenes (C20)
(Schall and Reiser, 2008). They are biosynthetically produced by two main pathways, the
mevalonate pathway and the 2-C-methyl-D-erythritol 4-phosphate/1-deoxy-D-xylulose
5-phosphate (MEP/DOXP) pathway which leads to the formation of isoprenyldiphos-
phate (IPP) and dimethylallyldiphosphate (DMAPP), precursors of terpenes and terpene-
derivatives (Berger, 2007). Terpenoids are the main components of essential oils, and
some of them are responsible for their characteristic aroma (Djilani and Dicko, 2012).
Most essential oils contain between 20–60 individual components, although there are
some with a more complex composition, such as jasmine, lemon, or cinnamon oils
(Djilani and Dicko, 2012). There are around 3000 known essential oils, but only 10%
are commercially traded in the world market, generating around USD$700 million per
year. Their uses extend from therapeutic treatment to fragrances and cosmetics (Djilani
and Dicko, 2012). Many of these isolated compounds are very well appreciated for their
particular aromas. Nevertheless, due to the low amount found naturally, vast amounts of
raw materials are required making them very expensive. A very well-known example is
“raspberry ketone” which provides a characteristic aroma to raspberries (Rubus ideaus)
(Larsen and Poll, 1990; Larsen and et al., 1991). It costs around USD$3000/kg when it
is extracted from natural sources. However, its synthetic version has an estimated cost of
USD$58/kg (Kutyna and Borneman, 2018).
Extraction from biological sources and chemical synthesis are the main methods of
production for aromas. However, new developments in the area of metabolomics, and in
synthetic biology, are going to be key research areas for the production of specific aroma
compounds (Kutyna and Borneman, 2018).
Despite the wide variety of chemical classes that could be present in the aroma com-
position of a determinate matrix, it is interesting to mention the fact that some aromas,
despite having different functional groups, could present similar odors, or conversely
chemical compounds with a similar structure could have a totally different aroma char-
acteristic. In order to understand this behavior, it is important to evaluate how aroma is
perceived in the nose and how the chemical structural differences from a molecule could
be quantified in order to predict its sensorial perception.

31.2 THEORY ON AROMA PERCEPTION

Chemoreception is defined as a physiological process whereby a living organism responds

to chemical stimuli (Ohloff, 1994). This property plays a significant role in the sur-
vival and reproduction of many different organisms. In insects, for example, their main
 Relationship between Structure and Odor 681

communication is dependent on chemical signals that trigger a specific behavior, and it

is used for locating food, mates, and other resources (Wilson, 1965; Lubes and Cabrera,
2018). In addition, this signal can be either inter- and/or intra-specific or even cast-spe-
cific and, in order to reach this condition, several attributes such as chemical class, chiral-
ity, and concentration are determinant aspects (Lubes and Cabrera, 2018). In humans,
although both anatomic and behavioral studies support the idea that they are able to
communicate biological information via odors (Doty, 1981), it is not vital in the process
of finding objects from the environment. Humans rely more on their sight. Nevertheless,
olfaction has a role in adding emotional attributes to certain events, especially in loving
or hating certain foods (Sarafoleanu et al., 2009).
Aroma perception is a result of a complex process. It starts with the molecular inter-
action of the odorant with the olfactory epithelium through two pathways: via the nose
(orthonasal) by sniffing or inhalation, and via the mouth (retronasal) during the inges-
tion of food or beverages (Negoias et al., 2008; Meierhenrich et al., 2004). These mole-
cules present some physicochemical properties such as moderate molecular weight (~400
Dalton), relatively low polarity, high vapor pressure, and lipophilicity (Ohloff, 1994). In
orthonasal perception, the chemical stimuli travel from an external environment through
the anterior nares to the mucosa during the inhalation process (Negoias et al., 2008;
Spors and Grinvald, 2002), while the retronasal process is more associated with food
and involves two mechanisms, taste and olfactory stimulation. During mastication, the
teeth break down the food structure to release volatiles, sometimes boosted by an enzy-
matic reaction from the saliva components and temperature of the mouth, allowing these
compounds to reach the nasal cavity during exhalation or swallowing (Negoias et al.,
2008; Hummel and Nordin, 2005). Figure 31.1 represents the orthonasal and retronasal
olfaction processes.
Odorant molecules, once they reach the epithelium, are activated by a subset of
around 1000 olfactory receptor types distributed across more than 6,000,000 recep-
tor cells (Doty, 2003). It has been estimated, on the basis of psychophysical testing,

FIGURE 31.1 Representation of the orthonasal and the retronasal olfaction processes.
In the orthonasal process, volatiles reach the nasal epithelium. In the retronasal pro-
cess, mastication breaks down the food structure releasing volatiles that reach the nasal
epithelium.
682 Food Aroma Evolution

that humans can discriminate at least 1 trillion (1.72 × 1012) olfactory stimuli or odors
(Bushdid et al., 2014). However, the intensity and pleasantness of a given odor differs
enormously between individuals, and this difference could be related to genetic variation,
as suggested by Keller et al. (2007), who, when studying a genotypic variation in a human
odorant receptor, OR7D4, observed a significant difference in the perception of andro-
stenone (5α-androst-16-en-3-one) and some other steroidal odors in terms of pleasant-
ness/unpleasantness and intensity (Keller et al., 2007). Both mechanisms of olfaction can
present differences in terms of odor threshold, intensity, and neural processing (Hummel
and Seo, 2016). It has been reported that orthonasal stimuli are less intense than retro-
nasal, meaning that orthonasal olfaction is more sensitive, and the odor identification is
even more efficient (Duffy et al., 1999; Burdach et al., 1984). In terms of brain activity,
both olfaction systems produce different patterns of brain activation. For example, it
was found that the activation of retronasal stimulation is at the base of the central sulcus
(Yamashita et al., 1999) producing a supra-additive response at the orbitofrontal cortex,
insula, and anterior cingulate cortex (Small et al., 2005).
The physiological mechanism involves the interaction of an odorant compound
with an olfactory receptor (OR) protein expressed at the surface of cilia of chemosen-
sory olfactory neurons in the epithelium (Man et al., 2004; Zozulya et al., 2001). At
the beginning of the 1990s, seven transmembrane ORs were identified (Buck and Axel,
1991). Mammalians ORs are G protein-coupled receptors belonging to Class I or A, more
specifically rhodopsin‐like GPCRs, integral membrane proteins with seven helical trans-
membrane (TM) domains and an extracellular N terminus (Man et al., 2004; Zozulya
et al., 2001).
Considering this, there is still no clarity on what chemical features are encoded by
olfactory receptors, which indicates that research focus in this domain has proven to be
difficult. In fact, of ~400 human odorant receptors, only 10% have a published ligand
(Mainland et al., 2015). Proteomic analyses through the liquid chromatography–tandem
mass spectrometry of human olfactory epithelial tissue are a key strategy for identifying
missing proteins. So far, a total of 1970 proteins were identified from the human nasal
epithelium, with two or more unique peptides detected in one or more samples (Hwang
et al., 2018). The Human Proteome Project (HPP) aims to discover high-stringency data
for all proteins encoded by the human genome in order to expand our knowledge on the
human proteome (Baker et al., 2017).

31.3 STRUCTURE–ODOR RELATIONSHIP

The relationship between the molecular structure of a chemical compound with its odor
quality or intensity is of considerable interest. The main problem in the application of
structure activity is the limited understanding of the olfactory mechanism and impreci-
sion of the odor measurement (related to odor description and intensity) (de Mello et al.,
2000). The first problem is related to the molecular recognition of the odorant by the
G-proteins. For that, two molecular features have been proposed as being important:
the molecular shape and the nature and disposition of its functional groups (Chastrette,
1997). The second problem is based on human perception, which could be highly subjec-
tive. On the one hand, the perception of odor is the result of physiological interactions
that depend on impression or association from the previous experience of the person.
People are limited in recognizing different odor descriptors due to the lack of experi-
ence or vocabulary to define the odor perceived. Dravnieks, by evaluating more than
 Relationship between Structure and Odor 683

140 odorants, proposed a list of 100 descriptors (Dravnieks, 1992). However, not all
descriptors could be associated with the same semantic level or properties of the olfac-
tion (Delasalle et al., 2014). On the other hand, a sensitivity problem associated with
intensity may vary from person to person and could even be completely different for two
enantiomers (Chastrette, 1997). One of the approaches used to overcome this difficulty
is the establishment of odor thresholds, which are defined as the concentration where the
odorant starts being perceived and depends, as expected, on many factors such as vapor
pressure, hydrophobicity, pKa, and so on. The implementation of a reference scale helps
to overcome this issue and consists of rating the intensity of an odorant with a reference
compound. For example, based on a determined concentration (in ppm or % vol/vol) of
n-butanol, the subject is asked to rate the intensity of a specific odorant.
Odor threshold (OT) has been widely used as an indicator of odor activity and can
be differentiated into the detection threshold, which is the lowest physical intensity where
a stimulus is perceptible, and the recognition threshold, defined as the lowest intensity at
which a stimulus can be identified (Jelen, 2011). OT depends on the partition coefficient
of a molecule in a determined medium due to the multiple physicochemical interactions of
this compound with its surroundings. In aqueous media, for example, it is greatly influ-
enced by the molecular chain length, unsaturation degree, double bonds position, stereo-
chemistry, and functional groups (Jelen and Gracka, 2016). Chain length impacts several
physicochemical parameters such as solubility, vapor pressure, viscosity, and even odor
perception. Most of the short-chain-length compounds are water soluble, and this prop-
erty starts decreasing with carbon chains greater than five. In addition, viscosity increases
in proportion with the chain length, as in the case of alcohols (Jelen and Gracka, 2016).
Functional groups can also influence OT due to differences in the solubility in a deter-
mined matrix. For example, a C5-alcohol is more water soluble than the alkane C5 due
to the possibility of forming hydrogen bridges. Van Gemert (2003) reported what could
be considered as the most comprehensive OT compilation with almost 18,000 threshold
values from about 3000 references (Van Gemert, 2003). In the following sections, the
influence of the structure in odorant systems is evaluated.

31.3.1 Influence of the Chain Length

As mentioned before, chain length affects the volatility and water solubility of a mol-
ecule. Studies performed in the homologous series, for example, alcohols (Schnabel et al.,
1998), showed a tendency for odor threshold to decrease with increment of the molecular
weight (Nagata and Takeuchi, 2003) (Figure 31.2).
This behavior can be predicted through non-linear regression approaches. For exam-
ple, Zarzo (2012) describes the OT variability of 114 aliphatic compounds based on
the alkyl chain length for different homologous series of carboxylic acids, aldehydes,
2-ketones, esters, 1-alcohols, amines, thiols, thioethers, and hydrocarbons. As a result,
the relatively high goodness-of-fit (R 2=0.90) suggested that OT values, of non-rigid mole-
cules, are a consequence of their functional group, molecular size, and also an interaction
between both factors. Some straight-chain compounds, such as alcohols, present fruity
notes from C4 to C9. Alcohols above C9, for example, present a waxier and unpleasant
note associated with an increment in the viscosity (Jelen, 2011). This could be due to the
aliphatic character of the odorant becoming more dominant. A series of studies performed
on alkanethiols, which are important flavor contributors of coffee, meat, wine, and so on,
showed that the chain length directly impacts the OT of primary, secondary, and even
684 Food Aroma Evolution

FIGURE 31.2 Odor threshold tendency due to molecular chain length in a series of
alkanols, alkan-2-ols, and alkan-3-ols. (Based on Schnabel et al., 1998.)

tertiary thiols (Polster and Schieberle, 2015). Compounds from C5 to C7 showed the low-
est OT, but by increasing chain length, the odor threshold increased as well (Figure 31.3)
(Polster and Schieberle, 2015).
Aroma descriptors of primary and secondary thiols were described as garlic-like
and/or burned notes until C4. C5 –C6 thiols were described as burned-roasted, and from
C9 on were described as soapy, such as in the case of alcohols (Polster and Schieberle,
2015). The same approach was followed on alka-1,5-dien-3-ones, alk-1-en-3-ones, alka-
1,5-dien-3-ols, and alk-1-en-3-ols (Lorber, Schieberle, and Buettner, 2014). On alka-1,5-
dien-3-ones and alka-1,5-dien-3-ols, the OT increases with the increment of the chain

FIGURE 31.3 Influence of the carbon chain length on the odor threshold of alkane-
1-thiols, alkane-2-thiols, and alkane-3-thiols. (Based on Polster and Schieberle, 2015.)
 Relationship between Structure and Odor 685

length, and the odor descriptor goes from geranium-like (C7) to citrus or green odor
quality (C14) depending on the isomer and the functional group evaluated. In general, the
most odor-active alkadienols were those with 7−10 carbon atoms with odor thresholds
ranging from 1 to 10 ng/L in air. The odor thresholds increased by a factor of 102 from
(Z)-deca-1,5-dien-3-ol to (Z)-dodeca-1,5-dien-3-ol (Lorber, Schieberle, and Buettner,
2014). Finally, the same conclusion was obtained on the evaluation of (E)-3-unsaturated
volatile acids, (E)-3-alkenols, and (E)-3-alkenals (Lorber and Buettner, 2015). On the
one hand, in the series of (E)-3-alkenoic acids, the odor quality changed successively
from sweaty to waxy. While, the odor qualities of (E)-3-alken-1-ols and (E)-3-alkenals
changed from grassy-green to an overall citrus-like, fresh, soapy, and coriander-like odor
with an increasing chain length. On the other hand, (E)-3-heptenoic acid, (E)-3-hexenoic
acid, and (E)-3-hexanal were those that presented the lowest thresholds in air (Lorber
and Buettner, 2015).

31.3.2 Influence of the Functional Groups

One of the key parameters in odor quality is the functional group of the molecule. This is
probably, one of the most complex aspects on structure–odor relationship due to the vast
pool of aroma–protein interactions that can be produced as a result of changes in the dipole
vector related to the disposition of the main functional group. The comparison here will be
limited to a small group of molecules. Table 31.1 shows the influence on the odor quality by
changing the functional group of a series of C6 –C9 compounds (Burdock, 2016).
Changing the functional group of compounds with similar structure affects their odor
threshold (Jelen and Gracka, 2016). In general, short-chain aromatic alcohols (until C9)
have a pleasant aroma; for example, isoamyl alcohol often contributes to a fruity aroma
(Myers, Issenberg, and Wick, 1970). An aliphatic or aromatic alcohol odor can change or
even disappear by changing the position or by removing the hydroxyl group (Reineccius,
2016). Examples are terpenes and terpenoids. Limonene presents a citrus, orange-like odor,
while its oxidized form, carveol, is described with a more minty scent (Burdock, 2016).
Short-chain aldehydes, saturated or unsaturated, are commonly found in food and pres-
ent a more green, fresh, or leaf-like aroma. In fact, some of them are produced by the lipid
oxidation process of linoleic or linolenic acid in contact with air (Lorber and Buettner,

TABLE 31.1 Influence of Changing the Functional Group on the Odor Quality of a Series of
Aliphatic Straight-Chain C6–C9 Compounds Based on Fenaroli’s Handbook
Alkane-1- Alkyl-1- Alkanoic Alkyl-1-
Compound Alkan-1-ols thiols Alkanals acetates acids amines
C6 Herbaceous, Burned, Green, Fruity Sweaty, Fishy
woody sulfury grassy rancid
C7 Woody, Sulfury, Fatty Fruity, Rancid, Ammonia-like
spicy onion pear sour
C8 Orange, rose Roasty, Fatty, Fruity Sour, Ammonia-like
sulfury citrus rancid
C9 Fatty, Sulfury, Fatty Floral, Fatty Pungent
orange-like fatty fruity
Source: Burdock, 2016.
686 Food Aroma Evolution

2015). Aldehydes, especially the unsaturated ones, can be used as an indicator of rancidity,
providing a “fishy” character to the food (Yu, Day, and Sinnhuber, 1961). As expected, due
to the effect of the chain length, the green-leafy odor of aldehydes starts vanishing with
an increasing number of carbon atoms toward a waxier smell. Aldehydes greater than C13
present a weak odor. However, it is not only aliphatic aldehydes that contribute significantly
to the aroma of some food, aromatic aldehydes, such as benzaldehyde or cinnamaldehyde,
are reported as important odorants presenting bitter almond and cinnamon-like aromas,
respectively (Jelen and Gracka, 2016). Studies on the volatility of aldehydes, ketones, and
esters in dilute water, showed that ketones presented the lowest air–water partition coef-
ficients at 25°C (Buttery, Ling, and Guadagni, 1969). At the same time, these compounds
are more volatile and more powerful as an odorant than the corresponding homologous
alcohols (Taylor and Linforth, 2009). Ketones present different odor attributes depending
on the chain length and the number of ketonic groups. Short-chain ketones such as 2-pro-
panone or 2-pentanone, are recognized as fruity or caramel-like (Berger, 2007), while the
presence of an unsaturation on 3-alkenones (from C5 to C8) brings a more ethereal smell
(Berger, 2007). Diketones, such as diacetyl (butane-2,3-dione) and acetoin, on the one
hand, are responsible for the buttery flavor in dairy products (Jelen and Gracka, 2016). On
the other hand, aromatic ketones, for instance, β-damascenone and β-ionone, contribute to
the floral smell of apple and raspberry, respectively (Berger, 2007).
Esters represent the most common group of compounds found in fruits. In apples,
for example, it is estimated that between 72–92% of the volatile composition is formed
by esters (Berger, 2007). The major biosynthetic route of esters is through the acyla-
tion of alcohols by the alcohol acyl-transferase (AAT) enzyme which can be expressed
on the peel or in the flesh of the fruit depending on the variety (Holland et al., 2005).
As described before, the fruity smell of esters starts vanishing when the carbon chain
increases and becomes soapier (Jelen and Gracka, 2016). Another factor that alters the
intensity or character of esters is the presence of double bonds. Unsaturated esters pres-
ent a higher OT than saturated ones, and the double bond position, as well as their con-
figuration, also affects their odor threshold (Jelen and Gracka, 2016). Acids, conversely
to esters, present a strong, pungent aroma. The most commonly recognized one is acetic
acid with its vinegar-like smell. The odor of C3 –C5 acids is reported as buttery-cheesy;
nevertheless, from C6 to C10 it becomes more unpleasant from rancid to fatty notes (Table
31.2) (Burdock, 2016). Longer chain acids are odorless but impart the acidity in food
systems (Jelen and Gracka, 2016).
Sulfur aliphatic compounds, such as thiols, thioesters, sulfides, polysulfides, or thia-
zoles are present in food in very low concentrations, but enough to impact its aromatic
properties. Thiols, for example, are molecules that contribute to the overall aroma of
food, including meat, coffee, and wine (Polster and Schieberle, 2015). This class of com-
pounds is among the most powerful odorants with odor threshold concentrations in the
range of picograms per liter. From the approximately 230 identified key food odorants,
30 of them are sulfur-containing molecules (Noe et al., 2017). It has been demonstrated
that changing the hydroxyl group in a molecule to a thiol group, in isosteric molecules,
causes a complete change of the odor quality (Wannagat et al., 1987). In contrast to thi-
ols, the odor thresholds of homologous alcohols decrease with an increasing carbon num-
ber from C3 to C8 (Polster and Schieberle, 2015). The addition of a second thiol group
in a molecule was evaluated by Polster and Schieberle (2015). It was found that these
molecules share some similarities with alkane-thiols; for instance, by increasing carbon
numbers, the thresholds of alkanedithiols increased exponentially. Additionally, alkan-
edithiols did not show a clear minimal OT with regard to the chain length, in contrast to
 Relationship between Structure and Odor 687

TABLE 31.2 Odor Quality of Some Aliphatic

Esters and Acids Based on Fenaroli’s
Handbook
Compound Fragrances
Esters
Ethyl acetate Solvent-like
Ethyl butanoate Fruity
Methylbutyl acetate Fruity; banana, pear
2-phenyl acetate Rose, honey-like
Ethyl hexanoate Apple, banana-like
Ethyl octanoate Pineapple, fatty
Ethyl decanoate Floral, fatty
Ethyl dodecanoate Fatty
Acids
Acetic acid Vinegar-like
Butyric acid Buttery
Hexanoic acid Rancid, fatty
Octanoic acid Oily, fatty
Decanoic acid Fatty, citrus-like
Source: Burdock, 2016.

what was found for alkane-1-thiols. It was concluded, therefore, that a second mercapto
group in the molecule does not result in a lower odor threshold (Polster and Schieberle,
2015). Studies on human odorant receptors (HOR) also showed that just 1 of the 391
HOR, OR2M3, responded exclusively to 3-mercapto-2-methylpentan-1-ol, 3-mercapto-
2-methylbutan-1-ol, and 3-mercapto-2-methylhexan-1-ol, producing the same onion-
and meat-like odor quality (Noe et al., 2017). Thiols can react with other components in
the matrix, leading to the formation of thioesters, which present a stronger aroma than
their homolog thiol. This strength, however, decreases from C2 to C6 (Jelen and Gracka,
2016). Sulfides, either symmetrical or asymmetrical, on the other hand, present a low
odor threshold with a strong sulfurous aroma that is sometimes described as boiled cab-
bage, asparagus, and garlic-like (Burdock, 2016).
From the group of nitrogen-containing molecules, amines can be considered as
the most important odorants. Nevertheless, amino acids react with reducing sugars in
Maillard reactions leading to the formation of more interesting aromatic compounds
that are key odorants in the majority of foods. Aliphatic amines have been described as
smelling ammonia-like (mainly for short-chain compounds) or fishy (for higher molecular
weights) (Burdock, 2016). The fishy aroma of amines is more intense for secondary and
tertiary amines, but the odor disappears by increasing the molecular weight, and they
become odorless compounds (Jelen and Gracka, 2016).
The majority of heterocycle compounds are formed through the thermal interaction
of reducing sugars and amino acids. Some other synthetic sources, such as hydrolytic and
pyrolytic degradation, as well as lipid oxidation, are other contributors of heterocycle
compounds in foodstuffs (Ho and Carlin, 1989). Due to the extension of this group, here
we will only be focused on pyrazines and thiazoles. Alkylpyrazines are considered as
some of the most important odorants in food with more than 200 compounds identified
and present, in general, a roasted nut-like flavor (Jelen and Gracka, 2016; Ho and Carlin,
688 Food Aroma Evolution

1989). The most potent odor of alkylmethoxypyrazines is a result of an alkyl group occu-
pying an ortho position to the methoxy group (Mihara and Masuda, 1988). In general, the
manipulation of methyl and methoxy groups on the pyrazine ring alters the odor thresh-
old but keeps its odor character (Teranishi, Buttery, and Guadagni, 1974). Additionally,
by increasing the chain of 2-methoxy-3-alkyl pyrazine, the odor threshold is reduced,
but the odor quality is increased (Teranishi, Buttery, and Guadagni, 1974). Mihara and
Matsuda (1988) reported that the odor threshold in dialkylpyrazines is mostly influenced
by the kind of substituent rather than the position. In this study, the odor threshold of
dialkylpyrazines increased with the following substitution pattern: 2,5-p​ositi​on < ​2,6-p​
ositi​on < ​2,3-p​ositi​on. While for dimethylpyrazines containing a polar group, such as
methoxy, ethoxy, methylthio, or acethyl, their odor threshold increased in reverse order
of substitution pattern, which means 2,3-p​ositi​on< ​2 ,6-p​ositi​on< ​2 ,5-p​ositi​on (Mihara
and Masuda, 1988).
Thiazoles are compounds possessing a five-membered ring with sulfur and nitro-
gen atoms in position 1 and 3, respectively. Their odor threshold ranges from 1 to
1000 µg/L. The unsubstituted ring is the one with the highest threshold, and this value
decreases by increasing the number of other groups in the ring (Jelen and Gracka,
2016; Teranishi, Buttery, and Guadagni, 1974). Their character can vary from bready
or roasted to green leaf with little changes in the structure. These compounds are
typically found in meat, especially in beef and lamb, and in roasted food (Jelen and
Gracka, 2016; Teranishi, Buttery, and Guadagni, 1974). They are formed from the
reaction of α-dicarbonyl with ammonia and hydrogen sulfide (Jelen and Gracka,
2016). Their odor behavior is closer to alkylpirazines and opposite to methoxypira-
zines (Teranishi, Buttery, and Guadagni, 1974). Thiazolines, which are the reduced
form of thiazoles, have also been reported in many foods but mainly in cooked beef.
For example, 2,4-dimethyl-3-thiazoline found in beef has been reported to have a
nutty, roasted, and vegetable aroma (Ho and Carlin, 1989).

31.3.3 Influence of the Unsaturation Degree and the Double Bonds Position

Double bonds play an important role in the intensity and odor character of a molecule.
For example, the addition of a double bond on 2-acetyl-2-thiazoline to form 2-acetylthia-
zole results in an odor sensation going from roasty-popcorn to sulfury-roasted, respec-
tively (Burdock, 2016). The odor threshold, in this case, increases from 0.12–1.3 µg/L
to 3–10 µg/L, respectively (Jelen and Gracka, 2016). However, in long-chain molecules,
this change only affects their odor threshold (Jelen and Gracka, 2016). In the case of
alcohols, unsaturated alcohol has a bigger impact on food flavor than the saturated one,
and double bonds near to the hydroxyl group in short-chain alcohols are reported to pro-
vide an unpleasant harsh aroma (Reineccius, 2016). In molecules with carbonyl groups,
the conjugation of double bonds, increases the polarity of the molecule making them
more soluble in water, affecting other physical properties as well (Teranishi, Buttery, and
Guadagni, 1974). The experimental evaluation of 3-alkenols, 3-alkenals, and 3-alkenoic
acids showed that the main odor quality in the series of (Z)-3-alken-1-ols was grassy-
green for C3 –C7 (3-alkenols), passing from fatty (C9 –C10) to citrus-like (for C11–C12).
(Z)-3-alkenals, on the other hand, revealed very similar main odor qualities and the smell
changes with a comparable pattern by increasing the chain length: starting with grassy,
to fatty and finally reaching citrus-like, soapy impressions for C11–C12 . In addition, the
(Z)-3-alkenoic acids series, were associated with odor qualities that are typical for small
 Relationship between Structure and Odor 689

fatty acids: starting with a sweaty odor impression to a plastic-like note developed by (Z)-
3-octenoic acid and (Z)-3-nonenoic acid, while C10 –C12 (3-alkenoic acids) were described
as waxy and acidic, respectively (Lorber and Buettner, 2015).
Adding an additional double bond to 2-alkenals, in order to convert these in 2,4-alka-
dienals, increases the length of conjugation and the polarity of the molecule (Teranishi,
Buttery, and Guadagni, 1974). Unsaturated aldehydes with double bonds in positions
2, 3, and 4 and the (E) or (Z) configuration have an odor threshold 1000 times lower
than their counterpart alcohols (Hatanaka, 1996). Finally, the investigation of steric hin-
drance of unsaturated esters suggested the following correlations: (1) unsaturation close
to the ester function reduces the fruity notes and increases the green ones, (2) intense
green notes are associated with cis olefins, and (3) the fruity character of esters dimin-
ishes with the increment on the steric bulk around the ester functions (Rossiter, 1996).

31.3.4 Influence of the Stereochemistry

The influence of the stereochemistry on a molecule can be very diverse. A good review of
this topic was published by Bentley in 2006 (Bentley, 2006). It is believed that the odor
of a molecule is not only related to its shape but also to its dipole vector (Chastrette et
al., 1992). Considering that the odor receptor is also a molecule, a good fit between the
corresponding parts of the odorant and the receptor is essential; therefore, geometrical
requirements are key factors (Chastrette et al., 1992). Nevertheless, so far, there are no
molecular rules about the odor or intensity of enantiomeric molecules. They could have
the same odor or intensity, the same odor quality but different thresholds or finally com-
pletely different notes. A good example of the influence of the stereochemistry is the pair
(R)- and (S)-carvon (Figure 31.4).
Each enantiomer can be found in nature, while (R)-carvon is perceived as caraway-
like; its enantiomeric form, (S)-carvon, is perceived as peppermint-like. Table 31.3 shows
a group of enantiomeric terpenes with different aromas.

31.4 APPLICATION OF QSAR IN AROMA DESCRIPTORS

The development and improvement of computational capacity led to an increase in QSAR

studies. This topic, applied in odorant molecules, has been widely discussed in the works
of Rossiter (1996), Chastrette (1997), and more recently of Tromelin (2016) (Rossiter,
1996; Chastrette, 1997; Tromelin, 2016).

FIGURE 31.4 Influence on the stereochemistry of (S)-carvon, (caraway-like odor; left)

and (R)-carvon (peppermint-like odor; right).
690 Food Aroma Evolution

TABLE 31.3 Enantiomeric Terpenes with

Their Respective Aroma Based on Fenaroli’s
Handbook
Monoterpene Enantiomer Fragrance
Carvone (R)-(–) Spearmint
(S)-(+) Caraway
Limonene (R)-(+) Orange
(S)-(–) Turpentine
α-Pinene (1R,5R)-(+) Slightly minty
(1S,5S)-(–) Pine tree
Menthol (1R,3R,4S)-(–) Minty
(1R,3R,4S)-(+) Phenolic
Source: Burdock, 2016.

QSAR studies are based on the hypothesis that changes in molecular structure pro-
duce proportional changes in an observed response or biological activity. A large number
of molecular descriptors have been created with this purpose. Among them, constitu-
tional, topological, geometrical, electrostatical, quantum-chemical, and thermodynami-
cal descriptors are the most important.
From this list, connectivity indices, which belong to a topological descriptor, are very
helpful to explain a large number of physicochemical properties (Galvez et al., 2011).
Tromelin (2016) recommends some good practices for the implementation of QSAR stud-
ies in order to avoid some of the pitfalls, pointing out the applicability domain and the
validation procedures (Tromelin, 2016). The choice of a molecular descriptor is essential
for delimiting the applicability domain and, furthermore, to avoid collinearity between
variables. Genetic algorithms allow, for example, to reduce the number of variables and
to select from a pool of possibilities those that are the most pertinent in the QSAR model
(Roger and Hopfinger, 1994). Finally, the validation process must be performed to make
sure that the model, in fact, can potentially predict a phenomenon. Among those meth-
ods, cross-validation and the Y-randomized test can be applied (Tropsha, Gramatica, and
Gombar, 2003; Rücker, Rücker, and Meringer, 2007).
Such QSAR approaches have been proven successful for predicting many properties,
for example, the odor threshold of some volatile organic compounds (Hau and Connell,
1998), the odor strength of alkylpyrazines unveiling the specific role of the two nitrogen
atoms in the molecule (Tsantili-Kakoulidou and Kier, 1992), and to simulate the molecular
mechanism of the human odor receptor with smelling compounds (Ahmed et al., 2018).

31.5 CONCLUSION

Aroma perception is a complex mechanism that depends on many factors. Among these
factors, molecular structure, concentration, release media, synergic or antagonist inter-
action, mechanism of olfaction, and genetical differences in odorant-receptor expression
are the most important to trigger a physiological response to a determined odorant. New
advances in computation, as well as in proteomics, will help to unveil even more molecu-
lar and physiological mechanisms of different molecules.
 Relationship between Structure and Odor 691

REFERENCES

Ahmed, L., et al. 2018. Molecular mechanism of activation of human musk receptors
OR5AN1 and OR1A1 by R-muscone and diverse other musk-smelling compounds.
Proceedings of the National Academy of Sciences 11517: E3950–8.
Baker, M. S., et al. 2017. Accelerating the search for the missing proteins in the human
proteome. Nature Communications 8: 14271.
Bentley, R. 2006. The nose as a stereochemist. Enantiomers and odor. Chemical Reviews
1069: 4099–112.
Berger, R. G. 2007. Flavours and Fragrances: Chemistry, Bioprocessing and Sustainability.
Springer Science and Business Media.
Buck, L. and Axel, R. 1991. A novel multigene family may encode odorant receptors: A
molecular basis for odor recognition. Cell 651: 175–87.
Burdach, K. J., Kroeze, J. H., and Köster, E. P. 1984. Nasal, retronasal, and gustatory per-
ception: An experimental comparison. Perception and Psychophysics 363: 205–8.
Burdock, G. A. 2016. Fenaroli’s Handbook of Flavor Ingredients. CRC Press.
Bushdid, C., Magnasco, M. O., Vosshall, L. B., and Keller, A. 2014. Humans can dis-
criminate more than 1 trillion olfactory stimuli. Science 3436177: 1370–2.
Buttery, R. G., Ling, L., and Guadagni, D. G. 1969. Food volatiles. Volatilities of alde-
hydes, ketones, and esters in dilute water solution. Journal of Agricultural and Food
Chemistry 172: 385–9.
Chastrette, M. 1997. Trends in structure-odor relationship. SAR and QSAR in
Environmental Research 63–64: 215–54.
Chastrette, M., Rognon, C., Sauvegrain, P., and Amouroux, R. 1992. On the role of chi-
rality in structure-odor relationships. Chemical Senses 175: 555–72.
de Mello Castanho Amboni, R. D., da Silva Junkes, B., Yunes, R. A., and Heinzen, V.
E. F. 2000. Quantitative structure–odor relationships of aliphatic esters using topo-
logical indices. Journal of Agricultural and Food Chemistry 488: 3517–21.
Delasalle, C., de March, C. A., Meierhenrich, U. J., Brevard, H., Golebiowski, J., and
Baldovini, N. 2014. Structure odor relationships of semisynthetic β-santalol ana-
logs. Chemistry and Biodiversity 1111: 1843–60.
Djilani, A. and Dicko, A. 2012. The therapeutic benefits of essential oils. In Nutrition,
Well-Being and Health. Bouayed, J. and Bohn, T. (Eds.). InTechOpen.
Doty, R. L. 1981. Olfactory communication in humans. Chemical Senses 64: 351–76.
Doty, R. L. 2003. Psychophysical measurement of human olfactory function, including
odorant mixture assessment. In Handbook of Olfaction and Gustation. Doty, R.
L. (Ed.). CRC Press.
Dravnieks, A. 1992. Atlas of odor character profiles. In Atlas of Odor Character Profiles.
Dravnieks, A. (Ed.). ASTM.
Duffy, V. B., Cain, W. S., and Ferris, A. M. 1999. Measurement of sensitivity to olfactory
flavor: Application in a study of aging and dentures. Chemical Senses 246: 671.
Galvez, J., Villar, V. M., Galvez-Llompart, M., and Amigo, J. M. 2011. Chemistry
explained by topology: An alternative approach. Combinatorial Chemistry and
High Throughput Screening 144: 279–83.
Hatanaka, A. 1996. The fresh green odor emitted by plants. Food Reviews International
123: 303–50.
Hau, K. M. and Connell, D. W. 1998. Quantitative structure-activity relationships
QSARs for odor thresholds of volatile organic compounds VOCs. Indoor Air 81:
23–33.
692 Food Aroma Evolution

Ho, C. T. and Carlin, J. T. 1989. Formation and aroma characteristics of heterocyclic

compounds in foods. In ACS Symposium Series. USA: American Chemical Society.
Holland, D., et al. 2005. Developmental and varietal differences in volatile ester forma-
tion and acetyl-CoA: Alcohol acetyl transferase activities in apple Malus domestica
Borkh. fruit. Journal of Agricultural and Food Chemistry 5318: 7198–203.
Hummel, T. and Nordin, S. 2005. Olfactory disorders and their consequences for quality
of life. Acta Oto-laryngologica 1252: 116–21.
Hummel, T. and Seo, H. S. 2016. Orthonasal and retronasal perception. In Flavour:
From Food to Perception. Guichard, E., Salles, C., Morzel, M., and Le Bon, A. M.
(Eds.). John Wiley & Sons.
Hwang, H., et al. 2018. Identification of missing proteins in human olfactory epithelial
tissue by liquid chromatography–tandem mass spectrometry. Journal of Proteome
Research. Article ASAP. doi:10.1021/acs.jproteome.8b00408.
Jelen, H. 2011. Food Flavors: Chemical, Sensory and Technological Properties. CRC Press.
Jelen, H. and Gracka, A. 2016. Characterization of aroma compounds: Structure,
physico-chemical and sensory properties. In Flavour: From Food to Perception.
Guichard, E., Salles, C., Morzel, M., and Le Bon, A. M. (Eds.). John Wiley & Sons.
Keller, A., Zhuang, H., Chi, Q., Vosshall, L. B., and Matsunami, H. 2007. Genetic varia-
tion in a human odorant receptor alters odour perception. Nature 449: 468–72.
Kutyna, D. and Borneman, A. 2018. Heterologous production of flavour and aroma com-
pounds in saccharomyces cerevisiae. Genes 97: 326.
Larsen, M. and Poll, L. 1990. Odour thresholds of some important aroma compounds
in raspberries. Zeitschrift für Lebensmittel-Untersuchung und -Forschung 191:
129–31.
Larsen, M., Poll, L., Callesen, O., and Lewis, M. 1991. Relations between the content
of aroma compounds and the sensory evaluation of 10 raspberry varieties Rubus
idaeus L. Acta Agriculturae Scandinavica 41: 447–54.
Lorber, K. and Buettner, A. 2015. Structure–odor relationships of E-3-alkenoic acids,
E-3-alken-1-ols, and E-3-alkenals. Journal of Agricultural and Food Chemistry
6330: 6681–8.
Lorber, K., Schieberle, P., and Buettner, A. 2014. Influence of the chemical structure on
odor qualities and odor thresholds in homologous series of Alka-1, 5-dien-3-ones,
Alk-1-en-3-ones, Alka-1, 5-dien-3-ols, and Alk-1-en-3-ols. Journal of Agricultural
and Food Chemistry 625: 1025–31.
Lubes, G. and Cabrera, A. 2018. Identification and evaluation of 3Z, 6Z, 8E-dodeca-3,
6, 8-trien-1-ol as the trail following pheromone on Microcerotermes exiguous
Isoptera: Termitidae. Revista de Biología Tropical 661: 303–11.
Lubes, G. and Goodarzi, M. 2017. Analysis of volatile compounds by advanced analyti-
cal techniques and multivariate chemometrics. Chemical Reviews 1179: 6399–422.
Lubes, G. and Goodarzi, M. 2018. GC–MS based metabolomics used for the identifica-
tion of cancer volatile organic compounds as biomarkers. Journal of Pharmaceutical
and Biomedical Analysis 147: 313–22.
Mainland, J. D., Li, Y. R., Zhou, T., Liu, W. L. L., and Matsunami, H. 2015. Human
olfactory receptor responses to odorants. Scientific Data 2: 150002: 1–9.
Man, O., Gilad, Y., and Lancet, D. 2004. Prediction of the odorant binding site of olfac-
tory receptor proteins by human–mouse comparisons. Protein Science 131: 240–54.
Meierhenrich, U. J., Golebiowski, J., Fernandez, X., and Cabrol-Bass, D. 2004. The
molecular basis of olfactory chemoreception. Angewandte Chemie International
Edition 4347: 6410–2.
 Relationship between Structure and Odor 693

Mihara, S. and Masuda, H. 1988. Structure-odor relationships for disubstituted pyr-

azines. Journal of Agricultural and Food Chemistry 366: 1242–7.
Mottram, D. S. and Elmore, J. S. 2003. Sensory evaluation: Aroma. In Encyclopaedia
of Food Sciences and Nutrition. Caballero, B., Trugo, L., and Finglas, P. M. (Eds.).
London: Academic Press.
Myers, M. J., Issenberg, P., and Wick, E. L. 1970. L-leucine as a precursor of isoamyl
alcohol and isoamyl acetate, volatile aroma constituents of banana fruit discs.
Phytochemistry 98: 1693–700.
Nagata, Y. and Takeuchi, N. 2003. Measurement of odor threshold by triangle odor bag
method. Odor Measurement Review 118: 118–27.
Negoias, S., Visschers, R., Boelrijk, A., and Hummel, T. 2008. New ways to understand
aroma perception. Food Chemistry 1084: 1247–54.
Noe, F., Polster, J., Geithe, C., Kotthoff, M., Schieberle, P., and Krautwurst, D. 2017.
OR2M3: A highly specific and narrowly tuned human odorant receptor for the
sensitive detection of onion key food odorant 3-mercapto-2-methylpentan-1-ol.
Chemical Senses 423: 195–210.
Ohloff, G. 1994. Scent and fragrances. In The Fascination of Odors and their Chemical
Perspectives. Springer-Verlag.
Polster, J. and Schieberle, P. 2015. Structure–odor correlations in homologous series of
alkanethiols and attempts to predict odor thresholds by 3D-QSAR studies. Journal
of Agricultural and Food Chemistry 635: 1419–32.
Reineccius, G. 2016. Flavor Chemistry and Technology. CRC Press.
Rogers, D. and Hopfinger, A. J. 1994. Application of genetic function approximation
to quantitative structure-activity relationships and quantitative structure-prop-
erty relationships. Journal of Chemical Information and Computer Sciences 344:
854–66.
Rossiter, K. J. 1996. Structure–odor relationships. Chemical Reviews 968: 3201–40.
Rücker, C., Rücker, G., and Meringer, M. 2007. y-Randomization and its variants in
QSPR/QSAR. Journal of Chemical Information and Modeling 476: 2345–57.
Sarafoleanu, C., Mella, C., Georgescu, M., and Perederco, C. 2009. The importance of
the olfactory sense in the human behavior and evolution. Journal of Medicine and
Life 22: 196–8.
Schall, A. and Reiser, O. 2008. Synthesis of biologically active guaianolides with a
trans-annulated lactone moiety. European Journal of Organic Chemistry 200814:
2353–64.
Schnabel, K. O., Belitz, H. D., and Von Ranson, C. 1998. Investigations on the structure-
activity relation of odorous substances. I. Detection thresholds and odor qualities
of aliphatic and alicyclic compounds containing oxygen functions. Zeitschrift für
Lebensmittel-Untersuchung und -Forschung 187: 215–23.
Small, D. M., Gerber, J. C., Mak, Y. E., and Hummel, T. 2005. Differential neural
responses evoked by orthonasal versus retronasal odorant perception in humans.
Neuron 474: 593–605.
Spors, H. and Grinvald, A. 2002. Spatio-temporal dynamics of odor representations in
the mammalian olfactory bulb. Neuron 342: 301–15.
Taylor, A. J. and Linforth, R. S. 2009. Food Flavour Technology. John Wiley & Sons.
Teranishi, R., Buttery, R. G., and Guadagni, D. G. 1974. Odor quality and chemi-
cal structure in fruit and vegetable flavors. Annals of the New York Academy of
Sciences 2371: 209–16.
694 Food Aroma Evolution

Tromelin, A. 2016. Prediction of perception using structure–activity models. In Flavor:

From Food to Behaviors, Wellbeing and Health. Etiévant, P., Guichard, E., Salles,
C., and Voilley, A. (Eds.). Woodhead Publishing.
Tropsha, A., Gramatica, P., and Gombar, V. K. 2003. The importance of being earnest:
Validation is the absolute essential for successful application and interpretation of
QSPR models. QSAR and Combinatorial Science 221: 69–77.
Tsantili-Kakoulidou, A. and Kier, L. B. 1992. A quantitative structure-activity relation-
ship QSAR study of alkylpyrazine odor modalities. Pharmaceutical Research 910:
1321–3.
Van Gemert, L. J. 2003. Compilations of Odour Threshold Values in Air, Water and
Other Media. Zeist: Oliemans, Punter & Partners BV.
Wannagat, U., Damrath, V., Schliephake, A., and Harder, U. 1987. Silasubstituted per-
fumes and isosteric compounds of perfumes. XI. Comparison of carbinols and silan-
ols with thiocarbinols and silanethiols. Monatshefte Für Chemie 118: 779–88.
Wilson, E. O. 1965. Chemical communication in the social insects. Science 1493688:
1064–71.
Yamashita, H., et al. 1999. Magnetic sensory cortical responses evoked by tactile stim-
ulations of the human face, oral cavity and flap reconstructions of the tongue.
European Archives of Otorhinolaryngology 2561: S42–6.
Yu, T. C., Day, E. A., and Sinnhuber, R. O. 1961. Autoxidation of fish oils. I. Identification
of volatile monocarbonyl compounds from autoxidized Salmon oil a, b, c. Journal
of Food Science 262: 192–7.
Zarzo, M. 2012. Effect of functional group and carbon chain length on the odor detec-
tion threshold of aliphatic compounds. Sensors 12(4): 4105–12.
Zozulya, S., Echeverri, F., and Nguyen, T. 2001. The human olfactory receptor reper-
toire. Genome Biology 26: research0018-1.
 Chapter 32
Bioactive Potential of
Sesquiterpenes
Iramaia Néri-Numa, Kele A.C. Vespermann, Carlos H. Carvalho,
Bruno Nicolau Paulino, Maria J. Macedo, Gláucia M. Pastore,
Juliano Lemos Bicas, and Gustavo Molina

CONTENTS

32.1 Introduction 695

32.2 Antioxidant Potential of Sesquiterpenes 696
32.3 Anticancer Potential of Sesquiterpene-Derived Aroma Compounds 701
32.4 Antimicrobial Potential of Sesquiterpenes 703
32.5 Anti-Inflammatory Effects Associated with Sesquiterpenes 705
32.6 Concluding Remarks 708
References 708

32.1 INTRODUCTION

Terpenes or isoprenoids constitute a class of chemical compounds with widely diverse

structures that are synthesized by living organisms. These chemical compounds con-
sist of isoprene (C5H8)n units and are classified as monoterpene (C10), sesquiterpene
(C15), diterpene (C20), triterpene (C30), and tetraterpene (C40) (Degenhardt, Köllner, and
Gershenzon, 2009; Gonzalez-Burgos and Gomez-Serranillos, 2012). Terpenes are the
most abundant group in nature, responsible for the characteristic odors of essential oils
(Janssens et al., 1992). Moreover, terpenes are secondary metabolites in the self-defense
mechanism of plants against pests and insects, also serving as an attractive scent for pol-
linators in general (Felipe, Oliveira, and Bicas, 2017; Feltran, 2014). These compounds
can also be found in mammals acting on cellular membrane stabilization, taking paths in
metabolic routes, and also serving as regulators in enzymatic reactions (Pimentel, 2012;
Vespermann et al., 2017). Meanwhile, sesquiterpenoids are a large class of naturally
occurring compounds with biological functions and desirable properties. Particularly,
synthesis of terpenoids in plants results in assorted metabolites (alcohols, acids, and alde-
hydes, or a combination of these) with diversified biochemical functions, such as phyto-
hormones, protein-modification reagents, and antioxidants, among others (Kumari et al.,
2013; Pichersky and Raguso, 2018). Among this group, the sesquiterpenes are one of the
largest classes of terpenoid compounds and common constituents of plant essential oils
(Sowden et al., 2005). This diverse group of terpenes, including linear and cyclic structures
with a broad distribution in Bryophyta and in the fungi kingdom (Duarte et al., 2017;

695
696 Food Aroma Evolution

Ludwiczuk, Skalicka-Wó Zniak, and Georgiev, 2016). Many sesquiterpenes, and their
alcohol, aldehyde, and ketone derivatives, are biologically active or precursors to metab-
olites with biological functions, while others have desirable fragrance, flavoring, and
medicinal properties (Fraga, 2004). Several sesquiterpenes are recognized for their poten-
tial as an aroma compound with pleasant and commercial characteristics and have also
been studied in the last few years regarding their biological potential. Figure 32.1 shows
the structure of the main sesquiterpenes discussed in this study and with broad bioactive
potential, including β-caryophyllene, patchoulene, farnesene, farnesol, valencene, noot-
katone, humulene, santalol, vernomelitensin, among others. Valencene, for example, is
found in several citrus fruits, such as in orange oil, peel, or juice, as well as in lemon peel
oil, mango, grapefruit juice, and oil, and so on. Santalols are found among the constitu-
ents of various sandalwood species (Santalum album L., S. spicatum, and S. autrocale-
donicum) with reported uses in baked goods, soft candy, and others.
Caryophyllene is a major component of the aroma of a cotton field, and present in
several oil compositions, such as those from angelica seed, black pepper, and the complex
rosemary oil. Meanwhile, nootkatone is recognized as a sesquiterpenoid with notewor-
thy odor characteristics, valuable for both food and cosmetics industries due to a pleasant
grapefruit-like note (Fraatz et al., 2009). The abundance of sesquiterpenes in nature and
their wide range of potential make research essential for future elucidations and com-
mercial exploitation. Thus, the objective of this chapter will be to address the bioactive
potential of sesquiterpenes and their derivatives, assessing their antioxidant, antican-
cer, antimicrobial, and anti-inflammatory potential, as well as to elucidate in vitro and
in vivo research conducted to identify the potential of these natural compounds.

32.2 ANTIOXIDANT POTENTIAL OF SESQUITERPENES

Recent data associated with the antioxidant behavior of terpenes have shown that these
compounds exert protective effects under oxidative stress conditions in several diseases
as well as aging processes, besides being employed as food preservatives in order to avoid
fat oxidation (Gonzalez-Burgos and Gomez-Serranillos, 2012; Graßmann, 2005). By

FIGURE 32.1 Structures of some sesquiterpenes presenting bioactive potential.

 Bioactive Potential of Sesquiterpenes 697

definition, an antioxidant is a substance that can delay, inhibit, or remove the oxida-
tive damage to a target substrate even when employed in a lower concentration than the
oxidized substrate (Gutteridge and Halliwell, 2010; Halliwell, 2007). Our body has a
complex antioxidant defense mechanism formed by enzymatic and non-enzymatic anti-
oxidants to protect cells against oxidative injuries and allowing homeostasis maintenance
(Figure 32.2) (Powers et al., 2004).
The antioxidant mechanisms that fit this definition include the scavenging of reac-
tive species (ROS, radical oxygen species, or RNS, radical nitrogen species), inactivation
of peroxides and other oxygen species, chelation of metals, and quenching of secondary
lipid oxidation products (Aruoma, 1998; Matkowski, 2008; Poprac et al., 2017).

FIGURE 32.2 An overview of an oxidative stress and antioxidant mechanism system.

The reactive species result from cellular metabolism, mainly by mitochondrial respira-
tory chain. Under normal physiological conditions, a balance between ROS/RNS and
the antioxidant mechanism system exists. However, exogenous factors such as pollution,
smoking, metabolism of xenobiotics, and natural aging progression, among others, can
increase the levels of reactive species disrupting the redox homeostasis and leading to
oxidative stress, which in turn is related to several chronic diseases when not neutralized
properly. Thus, the antioxidant defense mechanisms comprising non-enzymatic as well
as enzymatic antioxidants are responsible for scavenging overproduced ROS/RBS aiming
to keep the cellular equilibrium and avoid biomolecules lesions. Enzymatically, SOD is
responsible for neutralizing O2●– (the starting point of cascade reactions of ROS/RNS
production), turning it into H 2O2 , followed by CAT and GPx which in turn converts
H 2O2 into H 2O and O2. In addition, dietary antioxidants (phenolic compounds, carot-
enoids, vitamins C and E) play an important role in prevention and reduction of oxida-
tive stress, acting on both aqueous and lipid phases as well as in cell membranes and
lipoproteins. SOD, superoxide dismutase; CAT, catalase; GPx, glutathione peroxidase.
(From Marcadenti, 2015; Pham-Huy, He, and Pham-Huy, 2008; Rutkowski et al., 2007;
Prior, 2015.)
698 Food Aroma Evolution

However, to understand the mechanism of the action of antioxidants, it is neces-

sary to keep in mind how the reactive species are generated as well as the damage their
reactions cause. Conceptually, both ROS and RNS play a dual role and may present
either deleterious or beneficial effects on living systems (Figure 32.2) (Pham-Huy, He,
and Pham-Huy, 2008; Valko et al., 2007). Therefore, in the biological context, ROS/
RNS arise as byproducts of the metabolism or are produced “on purpose” at low to
moderate concentrations (Valko et al., 2007; Vieceli Dalla Sega et al., 2017). Once that
is done, ROS/RNS products play a crucial role in the regulation of many biological pro-
cesses such as cell migration, proliferation, gene expression, and angiogenesis (Incalza et
al., 2018; Prior, 2015; Rutkowski et al., 2007; Valko et al., 2007; Vieceli Dalla Sega et
al., 2017). On the other hand, at higher concentrations, ROS or RNS in “oxidative” or
“nitrosative” stress can disrupt equilibrium redox leading to cellular dysfunction and cel-
lular damage, since impaired regulation of ROS/RNS production is related to diabetes,
coronary diseases, neurodegeneration, and cancer (Fransen et al., 2012; Nimse and Pal,
2015; Poprac et al., 2017). Epidemiological studies have consistently shown that a high
dietary intake of fruits and vegetables, as well as whole grains, is strongly associated with
reduced risk of degenerative diseases (Liu, 2004; Russo, 2007). The rationale behind its
protective effects is the presence of antioxidant molecules that postpone autoxidation by
inhibiting ROS or RNS generation or by interrupting propagation of the reactive species
through the following mechanisms: (1) scavenging reactive species; (2) chelating metal
ions; (3) preventing the formation of peroxides; (4) breaking autoxidation chain reaction;
(5) reducing localized O2 concentrations (Brewer, 2011; Russo, 2007; Uttara et al., 2009).
Moreover, there is a growing interest in naturally occurring antioxidants for use in
food, cosmetic, and pharmaceutical products to replace synthetic ones which are being
restricted due to their carcinogenic effects (Shahidi, 2000; Tepe et al., 2005). Hence, the
development and evaluation of novel natural sources of antioxidants are important and
necessary. Due to the biological properties, not only polyphenols and carotenoids but
also tocopherols, ascorbic acid, erythorbic acid as well as its salts or derivatives have
been subject of studies aiming to understand their mechanistic aspects (Liu et al., 2018;
Lorenzo et al., 2018; Min et al., 2018; Quiroga, Nepote, and Baumgartner, 2019; Shahidi
and Zhong, 2010). With respect to the terpene classes, the pure essential oils obtained
from citric fruits, herbs, and spices include mainly monoterpenes and sesquiterpenes,
since their role in human health is directly linked to their chemical structures (Breitmaier,
2006; Graßmann, 2005; Zuzarte et al., 2015). Regarding the scavenging role of sesqui-
terpenes, the oxygenated molecules, particularly allylic alcohols, exhibit good antioxi-
dant capacity, just as much as terpenes (Ruberto and Baratta, 2000). The antioxidant
capacity and related metabolic effects of some sesquiterpenes are listed in Table 32.1.
For example, farnesol ([2E,​6E]-3​,7,11​-trim​ethyl​dodec​a-2,6​,10-t​rien-​1-ol)​ an aliphatic
sesquiterpene widely found in fruits, vegetables, herbs, as well as in the essential oils of
ambrette seeds and citronella, has the potential for alleviating oxidative stress (Eslahi,
Fahimi, and Sardarian, 2017; Khan and Sultana, 2011; Ku and Lin, 2015). Farnesol
also exerts a critical role in preventing neurodegenerative events as well as the modula-
tion of inflammatory mediators related to the oxidative stress by regulation key inflam-
matory cascade and also reducing the expression of Bax and antagonized LPS-induced
decreasing in anti-apoptotic Bcl-2 (Santhanasabapathy et al., 2015; Santhanasabapathy
and Sudhandiran, 2015).
Similarly, β-caryophyllene (tran​s-4,1​1,11-​t rime​thyl-​8 -meh​t ylen​ebicy​clo[7​, 2,0]​
undec​ - 4-4e​
ne), found in various plants, is a natural bicyclic sesquiterpene which
appears to pursue its neuroprotective role (by antioxidative action) inhibiting the
 Bioactive Potential of Sesquiterpenes 699

TABLE 32.1 The Antioxidant Capacity and Related Metabolic Effects of Sesquiterpenes
Antioxidant Capacity
Name of
Compound Model Biological Effect Reference
Acyclic
Sesquiterpene
Farnesol In vivo ↑Detoxifying enzymes GSH-Px, (Jahangir et
Activity of antioxidant CAT, GSH, and GR al., 2005);
enzymes biomarkers of ↓ LPO and XO; (Szűcs et al.,
protein oxidation; ↓ Lipid peroxidation; 2013);
TBARS; ↓ MDA level; (Lateef et
3-nitrotyrosine level. ↓ Micronuclei formation and al., 2013)
chromosomal aberrations
induced by cadmium;
Protective effect against (CdCl2)-
induced renal oxidative stress;
↑ geranylgeranylation of cardiac
proteins;
Decrease of infarct size in the rat
heart following ischemia/
reperfusion;
Protective effect against cigarette
smoke extract-induced oxidative
stress in prostate.
Monocyclic
Sesqiterpene
α-Zingiberene In vitro Scavenge superoxide and (Kuttan,
Superoxide radical by hydroxyl radicals; Jeena, and
NTB reduction ↑ Detoxifying enzymes SOD, Liju, 2013)
Hydroxyl radical by GSH-Px, CAT, and GSH;
Fe3+/ascorbate/EDTA/ ↓ lipid peroxidation;
H2O2 system Strong antinociceptive activity
DPPHIC50 and the mode of action might
ABTS involve the inhibition of
FRAP synthesis of arachidonic acid
TBARS metabolite mediated by COX
In vivo inhibition.
Activity of antioxidant
enzymes biomarkers of
protein oxidation;
PMA-induced
superoxide radical
generation.
Bicyclic
Sesqiterpene
β-Caryophyllene In vitro ↓ S. aureus; (Dahham et
DPPHIC50 ↓ clonogenicity, migration, al., 2015);
FRAP invasion and spheroid formation (Calleja et
Fe2+/ascorbate in HCT-116 cells; al., 2013)
(Continued )
700 Food Aroma Evolution

TABLE 32.1 (Continued) The Antioxidant Capacity and Related Metabolic Effects of
Sesquiterpenes
Antioxidant Capacity
Name of
Compound Model Biological Effect Reference
In vivo Selective anti-proliferative effects
Activity of antioxidant against HCT-116 cells (IC50 19
enzymes biomarkers of µM);
protein oxidation; Induces apoptosis via nuclear
TBARS. condensation and fragmentation
pathways including disruption of
mitochondrial membrane
potential;
↓ lipid peroxidation;
↓ LPO;
↓ Fibrosis and the expression of
Col1a1, Tgfb1, and Timp1
genes;
Protective effects of liver fibrosis
by reducing hepatic stellate cell
activation.
DPPH, 2,2-diphenyl-1-picrylhydrazyl; TEAC, trolox equivalent antioxidant capacity; FRAP, ferric-
reducing antioxidant power; NTB, 2-nitro-5-thiobenzoate; ORAC, oxygen radical absorbance
capacity; PMA, phorbol-12-myristate-3-acetate; GSH-Px, glutathione peroxidase, CAT, catalase,
GSH, glutathione, POX, peroxidase; XO, xanthine oxidase; LPO, 5-lipoxygenase; MDA, malondial-
dehyde; TBARS, thiobarbituric acid reactive substances; TCDD, 2,3,7,8-tetrachlorodibenzo-p-
dioxin; STZ, streptozotocin; HCT-116, colorectal cancer cells.

mRNA expression of inducible nitric oxide synthetase, interleukin (IL)-1b, IL-6, and
cyclooxygenase 2 in C6 microglial cells, and to reduce both nitric oxide and prostaglan-
din levels (Chang et al., 2013). Likewise, the chemotype-oils of Eugenia uniflora from
the Amazon which are rich in oxygenated sesquiterpenes and sesquiterpene hydro-
carbons such as curzerene, selina-1,3,7(11)-trien-2-one, selina-1,3,7(11)-trien-2-one
epoxide, germacrene B, caryophyllene oxide, and (E)-caryophyllene are able to scav-
enge DPPH (30.3–45.1%) and TEAC (217.0 to 228.3 mg TE/mL) radicals as well as to
inhibit a β-carotene/linoleic acid system in a range between 10.8 to 75.3%. Also, these
results were associated with strong cytotoxic activity against lung, colon, stomach, and
melanoma human cancer cell lines (Figueiredo et al., 2019). The essential oils of the
Baccharis species composed mostly of δ-elemene, β-caryophyllene, ɣ-muurolene, bicy-
clogermacrene, β-germacrene, germacrene D, spathulenol, τ-muurolol, and α-cadinol
display high antioxidant activity (DPPH= 0.328 mg AAE/g and ORAC FL >500.0 μmol
TE g−1) once these results are linked to both the antidematogenic activity and myeloper-
oxidase’s inhibition (Ascari et al., 2019; Zuccolotto et al., 2019). Although the health
effects of natural antioxidants in humans are generally considered promising, there
are definite challenges and limitations of the current data in better understanding the
molecular mechanisms responsible for its effects, together with the possible interactions
between different compounds and other dietary constituents. Another crucial point is
the low bioavailability of terpenes (due to restricted intestinal digestive and absorptive
dynamics, metabolism by the intestinal microflora and after absorption). Thus, further
 Bioactive Potential of Sesquiterpenes 701

in-depth research and prospective as well as both mechanistic and human trials remain
crucial for the progress of this field.

32.3 ANTICANCER POTENTIAL OF SESQUITERPENE-

DERIVED AROMA COMPOUNDS

Cancer is a severe multifactorial disease characterized by genetic mutations and altera-

tions where the cell growth and proliferation occur uncontrollably and is related as one
principal cause of mortality around the world (Braicu et al., 2017; Iqbal et al., 2017; Li
et al., 2018; Babaei et al., 2018). Some authors are alarmed at the increase in the number
of the cases of cancer in the world and have estimated that the number of deaths because
of this disease in 2020 will be approximately 12 million, suggesting a great demand for
better treatment strategies (Torre et al., 2015; Kanavos, 2006; Galadari et al., 2017).
Today, the traditional approaches applied for cancer treatment involves chemotherapy,
immunotherapy, radiotherapy, and surgical resection (Galadari et al., 2017; Oprea et al.,
2017; Sharma et al., 2019). However, these current methods exhibit many disadvantages
associated with side effects and multidrug resistance of cancer cells affecting the success
of cancer treatment (Breen et al., 2017; Oprea et al., 2017; Cui et al., 2018; Johnson et al.,
2019). According to Mazumder et al. (2018), between 2010 and 2017, 97 molecules were
approved by the Food and Drug Administration (FDA) as novel agents against different
types of cancer. In addition, many authors reported the anticancer potential of several
natural compounds from plants and microorganisms, showing new possibilities for the
pharmaceutical and biotechnological companies to explore for the development of novel
compounds for chemotherapy (Li et al., 2018, 2016; Fidyt et al., 2016; Epifano et al.,
2014; Nuutinen, 2018). In terms of natural products, the phytochemicals are considered
promising due to several biological activities, including antitumoral potential (Galadari
et al., 2017). In this context, some terpenes have been reported as potential anticancer
agents (e.g., betulic acid, celastrol, ochraceolides, and others) and the volatile sesquiter-
penes related to the aroma of many food products have been considered promising due
to anticancer properties shown in several essential oils obtained from medicinal plants,
fruits, and other vegetal tissues used as food by humans (Mazumder et al., 2018; Wiart,
2013; Babaei et al., 2018; Kapoor, 2013; Paduch et al., 2016). In the last few years, dif-
ferent essential oils constituted by sesquiterpenes as major components were reported
as cytotoxic against different tumor cell lines suggesting the role of these compounds
in anticancer activity and their potential for pharmaceutical and medical applications.
Regarding the anticancer activity, the number of recent studies associated with sesqui-
terpenes is lower than those available for monoterpenes, for example, data are basically
limited to essential oils containing compounds such as β-caryophyllene and its oxide,
humulene, santalol, farnesene, nootkatone and its metabolites, sesquiterpene lactones,
and other few related compounds (Pavithra et al., 2018a,b; Fidyt et al., 2016; Bommareddy
et al., 2016; Gliszczyńska et al., 2011; Babaei et al., 2018). β-Caryophyllene is a natural
bicyclic sesquiterpene naturally found in essential oils from different plants, for example
Origanum vulgare L., Pamburus missionis, Cinnamomum spp., Eugenia caryophyllata
L., Cannabis sativa L., Zingiber nimmonii, and Piper nigrum L. (Abbas et al., 2013;
Fidyt et al., 2016; Pavithra et al., 2018a). The essential oil obtained from the leaves of
Pamburus missionis showed high amounts of sesquiterpenes, mainly β-caryophyllene
(25.4%), aromadendrene oxide-(2) (14.01%), 4(14),11-eudesmadiene (7.17%), and the
diterpene phytol (6.88%) (Pavithra et al., 2018a). This essential oil showed IC50 values
702 Food Aroma Evolution

ranging from 50 to 400 μg/mL for various human cancer cell lines (HaCaT, MCF-7,
K562, A431, HL-60, MOLT-4, DLD1, HepG2) after 72 h of treatment.
Recently, the cytotoxic effects of the oxygenated derivative β-caryophyllene oxide were
reported against breast adenocarcinoma cells (MCF7 and MDA-MB-231) (Hanušová et
al., 2017). The results showed IC50 values of 69 μg/mL for MDA-MB-231 cells, while that
for MCF7 cells, the IC50 value was around 24 μg/mL, all results obtained after 72 h of
exposition. The anticancer activity of aromadendrene oxide 2 against human epidermoid
cancer (A431) and precancerous HaCaT cells was described by Pavithra et al. (2018b).
The results suggested that this oxygenated sesquiterpene exhibited a concentration and
time-dependent cytotoxic effect with IC50 values of 50 μM for A431 cells and 76 μM
for HaCaT cells after 72 h of exposure. Moreover, the combination of β-caryophyllene
and aromadendrene oxide 2 was able to reduce cell viability and to inhibit the colony
formation, and it was also verified that these compounds in combination induced cell
cycle arrest and apoptosis in HaCaT and A431 cells (Pavithra et al., 2018c). Previously,
Langhasova et al. (2014) reported the antiproliferative activity of the essential oil from
Myrica rubra leaves against human colon and ileocecal adenocarcinoma cell lines (HCT8,
SW620, SW480, HT29, and Caco2). The chemical composition of this essential oil was
mainly of sesquiterpenes, including β-caryophyllene (43.2%), α-humulene (21.6%), and
valencene (6.0%). The results of antiproliferative assays showed different efficiencies of
the essential oil on cancer cell lines, with IC50 values ranging from 30 to 49 μg/mL for
Caco2, HCT8, and HT29 cells, while for SW480 and SW620 cells the IC50 values were
from 116 and 132 μg/mL, respectively. Similarly, humulene also exhibited a cytotoxic
effect against colorectal cancer cells (HT29) with a with an IC50 value of 54 μM, while
one of its derivatives obtained from Asteriscuc vogelii was cytotoxic against lymphoma
(P-338), lung carcinoma (A-549), colon carcinoma (HT-29), and melanoma (MEL-28)
cells with IC50 values ranging from 1 μM to 2 μM (Lan et al., 2011; Rauter et al., 2001).
α-santalol, the major volatile compound of sandalwood oil, is another sesquiterpene
exhibiting anticancer activity. Studies indicated that this compound presented in vitro
and in vivo antitumoral effects against prostate cancer cells, epidermoid carcinoma cells,
and breast cancer cells (Bommareddy et al., 2012, 2016; Zhang et al., 2010). In addi-
tion, Ortiz et al. (2016) reported the cytotoxic effect of commercial sandalwood essential
oil, composed by α-santalol (25.34%), (Z)-nuciferol (18.34%), β-santalol (10.97%), and
(E)-nuciferol (10.46%), against MCF-7 and MCF-10A cell lines. The results showed IC50
values of 8.03 μg/mL for the MCF-7 cell line, while for the MCF-10A cell line the IC50
was of 12.3 μg/mL. In this context, Lee et al. (2015) studied the cytotoxic activity of
East Indian sandalwood oil and its major constituents (α-santalol and β-santalol) against
seven cancer cell lines (SCC-4, CAL 27, HSC-3, SCC-9, SCC-25, HN5, and HSC-2). In
summary, the IC50 values exhibited by the essential oil ranged between 0.8 and 1.6 μg/
mL, while for sesquiterpenes it was from 3.4 to 5.5 μM for α-santalol and 4.2 to 7.3
μM for β-santalol, respectively. Moreover, the essential oil obtained from Cedrelopsis
grevei leaves in which the major compounds were (E)-β-farnesene (27.61%), δ-cadinene
(14.48%), α-copaene (7.65%), and β-elemene (6.96%) showed cytotoxic effects against
human breast cancer cells (MCF-7) with an IC50 value of 21.5 μg/mL, suggesting the
role of sesquiterpenes in anticancer activity (Afoulous et al., 2013). Based on the infor-
mation on the anticancer activity of volatile terpenes, it is possible to verify that many
advances have been achieved for different terpenes with promising results in clinical stud-
ies. However, more studies are necessary to prove the effectiveness of the sesquiterpenic
aromas both in in vitro and in vivo studies, increasing the possibilities of applying ter-
penes as anticancer agents and for development of new drugs.
 Bioactive Potential of Sesquiterpenes 703

32.4 ANTIMICROBIAL POTENTIAL OF SESQUITERPENES

Antimicrobials are natural (antibiotic) or synthetic (chemotherapeutic) substances that

act on microorganisms by inhibiting their growth or causing their destruction (Diemer,
2016). Antibiotics are classified according to molecular structure and their antimicrobial
mechanisms. These mechanism’s actions consist of interrupting the synthesis of struc-
tural components or altering metabolic functions unique to microbial cells (Becker, 2013).
Antimicrobial metabolites from bioprocesses are an important alternative to overcome
increasing levels of drug resistance by plant and human pathogens. Antimicrobial com-
pounds can be used not only as drugs but also as food preservatives in the control of
deterioration (Bhatti et al., 2014). This section discusses antimicrobial sesquiterpenic com-
pounds, such as sesquiterpene lactones, which disrupt the cell walls of fungi and bacteria,
due to the polar groups in these compounds that interrupt phospholipid membrane (Figure
32.3) (Chadwick et al., 2013). Regarding the potential of sesquiterpenes, Picman (1984)
tested the antifungal activity of 45 sesquiterpene lactones against Microsporum cookie,
Trichophyton mentagrophytes, and Fusarium sp. The authors concluded that approxi-
mately 29% of the tested sesquiterpene lactones showed a considerably strong antifungal
activity, while 33% partially inhibited fungal growth when compared to assays without
lactones. Also, around 38% did not show any antimicrobial activity. Among the lactone
classes tested, germacranolides, guaianolides, pseudoguaianolides, and eudesmanolides
presented at least one sesquiterpene strongly capable of inhibiting fungal growth; the lat-
ter two being the ones with the highest numbers (four and five, respectively). They suggest
that the antimicrobial activity of these lactones have multiple causes, one of them being
the reactions between lactones and thiol groups on microbial enzymes.
Rugosal A, a novel antimicrobial sesquiterpene extracted from a wild rose species
called Rosa rugosa, was identified by Hashidoko, Tahara, and Mizutani (1989). This nat-
ural antimicrobial compound caused the considerable growth inhibition of Cladosporium
herbarum at concentrations of 0.9 µg/mm², while at concentrations higher than 3.75 µg/
mm² growth was completely compromised. The methyl ester of rugosic acid A presented
activity compared to that of rugosal A, while the free acid was capable of strongly inhib-
iting growth only at concentrations higher than 100 µg/mm². Antimicrobial activity of
sesquiterpenes was also found by Al-dabbas et al. (2005), when selina-4,11(13)-dien-
3-on-12-oic acid, an antibacterial sesquiterpene isolated from Varthemia iphionoides,
showed potent antimicrobial activity against Staphylococcus aureus, Bacillus subti-
lis, Micrococcus luteus, Escherichia coli, Bacillus cereus, and Salmonella enteritides.
The highest value of the minimum inhibitory concentrations (MICs) was 500 µg/mL
from Staphylococcus aureus, while the minimal observed value was 250 µg/mL. From a
novel sesquiterpene lactone isolated from Centaurea pullata, Djeddi et al. (2008) found

FIGURE 32.3 Effects of sesquiterpene lactones in the cell and cytoplasmic membrane.
704 Food Aroma Evolution

out that α-methylene-γ-lactone group was a potential antifungal group, being stronger
than miconazole, a commercial fungicide. Alarif et al. (2012) isolated three new lau-
rene-type sesquiterpenes from red alga Laurencia obtuse, identifying that 12-hydroxy
isolaurene and isolauraldehyde exhibited potential antimicrobial activity against B.
subtilis and S. aureus, the latter being also effective against Candida albicans. The
antimicrobial potential of this sesquiterpene is allegedly related to its molecular acces-
sibility and lipophilicity, as well as a conjugated carbonyl group in the molecule side
chain. Two inositol derivatives and four 3,4-seco-pseudoguaianolides were isolated by
Fortuna et al. (2011) from plant Hymenoxys robusta, a promising antimicrobial ses-
quiterpenes source. Out of the isolates, only vermeerin, a 3,4-seco-pseudoguaianolide,
showed antimicrobial activity against S. aureus, with a MIC of 10 µg/mL and show-
ing no toxicity against human macrophages. Amaya et al. (2012) were able to isolate
six sesquiterpenes lactones from Centratherum punctatum, which compromised biofilm
formation and elastase activity of Pseudomonas aeruginosa ATCC 27853 despite not
being able to completely inactivate the microorganism. Out of the compounds tested,
1-oxo-3,10-epoxy-8-methacryloyloxy-15-hydroxygermacra-2,4,11(13)-trien-6,12-olide,
1-oxo-3,10-epoxy-8-epoxymethacryloyloxy-15-hydroxygermacra-2,4,11(13)-trien-
6,12-olide, and 1-oxi-3,10-epoxy-5-hydroxy-8-tigloyloxy-germacra-2,4(15),11(13)-
trien-6,12-olide showed biological activity against pathogenic parts of P. aeruginosa.
In another study, AO et al. (2012) found two new sesquiterpenes, guai-9-en-4β-ol and
14,15-dionorguai-1,11-dien-9,10-dione, isolated from Syringa pinnatifolia Hemsl., the
first being effective against Bacillus coagulans (zone of inhibition of 15.34 mm) while
the latter showed antimicrobial activity against Proteus vulgaris (14.96 mm) and E.
coli (15.34 mm). Sesquiterpene 14,15-dionorguai-1,11-dien-9,10-dione was also highly
active against Fusarium oxysporum (15.32 mm). Eleven sesquiterpenes isolated from
the medicinal plant Chloranthus angustifolius were studied by Yang et al. (2014) on
their antimicrobial activities. Potent results when the MIC values of six of the sesqui-
terpenes were analyzed in tests against Candida albicans: henriol A, spicachloratin A,
tianmushanol, 8-O-methyltian-mushanol (all with a MIC value of 8 µg/mL), chloram-
ultilide A, and shizukaol B (with MIC value of 4 µg/mL). A novel sesquiterpene from
Microdiplodia sp. TT-12 and 13-hydroxylmacrophorin A, among three others, was iso-
lated by Shiono et al. (2015), the first being the only one that showed antimicrobial activ-
ity, weakly inhibiting Raffaelea quercivora JCM 11526, with 12 mm of inhibited area.
Also, Bunbamrung et al. (2017) isolated 13 sesquiterpenes from mushroom Gloestereum
incarnatum BCC41461, and obtained antimalarial and antimicrobial activity against
Bacillus cereus from sesquiterpenes incarnatin A (IC50 9.80 µg/mL) and incarnolactone
C (MIC 25 µg/mL), respectively. The interaction between the microorganisms tested and
the sesquiterpenes is yet to be elucidated, so the viability of this novel sesquiterpene
for antimicrobial purposes can be evaluated. Duan et al. (2018) worked with sesqui-
terpenes combined with α-amino acids, labeled as stereumamides A-D, isolated from
Stereum hirsutum FP-91666, a wildly spread mushroom. From these, steuramides A and
D showed antimicrobial activity against E. coli, S. aureus, and Salmonella typhimurium
(MIC 12.5, 12.5, and 25.0 µg/mL, respectively). It was also the first report of naturally
occurring quaternary ammonium compound (QAC), those being common antibacterial
agents used in the food industry. Also, Zhao et al. (2018) isolated six novel dihydro-β-
agarofuran sesquiterpenes from Monimopetalum chinense Rehd., a common medicinal
plant from China. It was concluded that sesquiterpene 5, named monichinine E, was a
potential compound for developing fungicide against Botrytis cinerea, presenting EC50
value of 9.33 µg/mL. Sesquiterpenes have been studied for their antimicrobial activities in
 Bioactive Potential of Sesquiterpenes 705

several areas, emerging as alternative compounds in that process due to their abundance
in nature and prominent results. However, more studies still need to be performed to
elucidate their mechanism of action, given that details on how sesquiterpenes affect cell
viability are scarce in literature reports.

32.5 ANTI-INFLAMMATORY EFFECTS

ASSOCIATED WITH SESQUITERPENES

Inflammation is a biological response to harmful stimuli, such as pathogens, physical

injuries, or cell damage (Ghasemian, Owlia, and Owlia, 2016), generating a combination
of heat, redness, swelling, and pain in a local area as physical symptoms (Prakash, 2017).
This could be an acute or chronical process, taking place as a coordinate activation
of numerous biochemical events, including the local vascular system, the immune system,
different cell types found in the injured tissue, and the expression of pro and anti-inflam-
matory mediators, including chemokines, cytokines, vasoactive amines, eicosanoids, and
proteolytic cascade products (de Cássia da Silveira e Sá, Andrade, and de Sousa, 2013;
Heras and Hortelano, 2009). A chronic inflammation occurs when this process is pro-
longed and deregulated, causing a mass cell destruction. This is associated with chronic
diseases, such as asthma, atherosclerosis, type 2 diabetes, Alzheimer’s disease, cancer,
and autoimmune diseases (de Cássia da Silveira e Sá, Andrade, and de Sousa, 2013; Heras
and Hortelano, 2009). Conventionally, clinical treatment for inflammatory diseases can
be performed with the use of nonsteroidal anti-inflammatory drugs (NSAIDs), glucocor-
ticoids, and disease-modifying agents of rheumatoid diseases (DMARDs), the latter
being able to reduce or prevent epithelial damage caused by the inflammatory process
(Tabas and Glass, 2013). On the other hand, NSAIDs act by reducing inflammation and
the pain caused, blocking arachidonic acid metabolism by cyclooxygenase enzyme
(COX), preventing prostaglandins production (Nonato et al., 2012). The NSAIDs drugs
are the most commonly used drugs to treat inflammatory diseases, and as analgesics as
well. However, their use can also cause gastrointestinal, renal and cardiovascular toxicity
(de Santana Souza et al., 2014; Salakhutdinov, Volcho, and Yarovaya, 2017). The use of
steroidal drugs as anti-inflammatory agents is also a controversial subject, given their
multiple side effects (Nonato et al., 2012). In this way, the use of alternative therapies
capable of efficiently treating inflammatory diseases while causing less aggressive side
effects have been widely studied lately. Among these, studies based on medicinal plant’s
essential oils and their isolated compounds (such as terpenes, for example) can be high-
lighted (de Santana Souza et al., 2014). By in vitro and in vivo assays, the therapeutic
potential of a series of terpenoids, like α-pinene, myrcene, p-cymene, linalool, myrtenol,
and borneol, have been investigated (Salakhutdinov, Volcho, and Yarovaya, 2017). Most
investigations concerning the anti-inflammatory potential of agents are undertaken in
vivo assays with the use of different test models, such as the carrageenan induced paw
edema (de Cássia da Silveira e Sá, Andrade, and de Sousa, 2013). Also, in vitro assays
with lipopolysaccharide (LPS)-stimulated RAW264.7 cells are widely used for evaluating
the anti-inflammatory effects of natural products, given the crucial role of macrophages
in inflammatory process (Bi et al., 2017). Some molecules that play an important role in
the regulation of immune and inflammatory responses are nuclear factor-kappa B (NF-
κB), interleukins (IL), tumor necrosis factor-α (TNF-α), and the expression of several
enzymes (oxygenases, nitric oxide synthases (NOS), and peroxidases), the reduction of
the expression of these molecules being the main goal of potential anti-inflammatory
706 Food Aroma Evolution

compounds studies (Prakash, 2017). Heme oxygenase-1 (HO-1) provides cyto-protective,

anti-apoptotic, and immunomodulatory effects in mammalian tissues, given the fact that
her synthesis is up-regulated in response to stress conditions, such as vascular injury,
ischemia, oxidative stress, and immune response (Tsoyi et al., 2011). Among sesquiter-
penes, nootkatone and valencene increased the relative HO-1 levels more than four-fold
in comparison to the control and significantly inhibited iNOS/NO production, and as a
consequence attenuated high mobility group box1 (HMGB1) release in LPS-activated
RAW264.7 cells (Tsoyi et al., 2011). In another study, the anti-inflammatory effect was
compared between β-patchoulene epoxide and β-patchoulene in LPS-stimulated
RAW264.7 macrophages, and it was demonstrated that β-patchoulene epoxide was a bet-
ter option to reduce the TNF-α, IL-12, IL-1β, and monocyte chemotactic protein-1
(MCP-1) levels, the production of NO and prostaglandin E2 (PGE2), and iNOs mRNA
expression than β-patchoulene (Wu et al., 2018). Deregulation of NF-κB levels is linked
to a number of pathologies in immune, cardiovascular, central nervous system (CNS),
muscular and renal systems, and also in inflammatory response. Therefore, there is an
interest in the search for biomolecules capable of inhibiting NF-κB activation (Heras and
Hortelano, 2009). In this context, vernomelitensin showed an excellent NF-κB inhibitory
activity, with a half maximal inhibitory concentration (IC50) of 3.6±0.4 µM, and also
inhibited STAT3 in an in vitro assay with NIH-3T3-KBF-Luc, HeLa-STAT3-luc and
HaCaT-ARE-Luc cell line, respectively. Other sesquiterpenes (8-(4′-hydroxymethacryloyl)-
dehydromelitensin, elemacarmanin, onopordopicrin, and carmanin) were also evaluated,
and presented lower inhibitory activity (22.5 to >50 µM) than vernomelitensin
(Formisano et al., 2017). The anti-inflammatory potential of seven new sesquiterpene
lactones isolated from Elephantopus mollis (8-O-methacryloylelephanpane, 2,4-
bis-O-methyl-8-O-methacryloylelephanpane, 4-O-ethyl-8-O-methacryloylelephanpane,
8-O-methacryloylisoelephanpane, 2-O-demethyltomenphantopin C, molephantin A,
and molephantin B) were evaluated on LPS-activated RAW264.7 cells. The new com-
pounds strongly reduced the nitric oxide production, with an IC50 between 0.57±0.17
and 14.34±1.61 µM (Wu et al., 2017). Since an excessive production of NO is involved in
the triggering of some inflammatory diseases, such as arthritis, ulcerative colitis, and
Crohn’s disease, the discovery of inhibitors of NO synthesis represents an important
therapeutic advance in the management of these inflammatory diseases (Wu et al., 2018).
In vitro studies help to study the cellular response in a closed system, but some responses
of inflammatory mechanisms are only evaluated in in vivo assays, such as tissue injury,
edema formation, and leukocyte infiltration (Nile and Park, 2013). The administration of
nootkatone and valencene significantly increased survival rates in cecal ligation and
puncture (CLP)-induced sepsis in mice. This can be a consequence of the induction of
HO-1 and reduction of HMGB1 induced by these sesquiterpenes (Tsoyi et al., 2011).
Nootkatone also prevented the diesel exhaust particles (DEP)-induced increase in airway
resistance, lung inflammation, and apoptosis in DEP-induced lung toxicity in mice. This
could be associated with an inhibition of NF-κB activation. Other factors were also
noticed, such as a decreased neutrophil infiltration in bronchoalveolar lavage fluid and
also reduction of TNF-α levels (Nemmar et al., 2018). The anti-inflammatory activity of
the sesquiterpene fraction from Eupatorium lindleyanum was evaluated using xylene-
induced ear edema in mice. The sesquiterpene fraction, composed by eupalinolide L,
eupalinolide, eupalinolide A, eupalinolide B, eupalinolide C, 3β-acetoxy-8β-(4′-
hydroxytigloyloxy)-14-hydroxylcostunolide, eupalinolide K, 2α-hydroxyeupatolide, and
eupalinolide H, inhibited the edema formation 21.53% (Wang et al., 2017). The sesqui-
terpenoid patchoulene epoxide reduced the levels of TNF-α and IL-1β, PGE2, and NO
 Bioactive Potential of Sesquiterpenes 707

levels and inhibited NF-κB activation in the ethanol-stimulated gastric tissues of mice
(Liang et al., 2018). The sesquiterpenes discussed above presented anti-inflammatory
effects both in in vivo and in vitro assays through the reduction of leukocyte migration,
reduction of interleukins, TNF-α, PG2, NO, MCP-1, NF-κβ levels, also through the
iNOS expression and production of HMGB-1 and HO-1 (Table 32.2). Sesquiterpenes
have been shown to significantly inhibit the development of edemas in in vivo models, as
well as affect several mechanisms associated with inflammatory responses in both in vivo
and in vitro assays. Given their presence in a wide number of essential oils commonly
used by the population, without harming effects, these compounds could be used as a
potential alternative to the traditional treatments of inflammatory diseases with steroidal
drugs and NSAIDs.

TABLE 32.2 Mechanism the of Anti-Inflammatory Action of Sesquiterpenes

Sesquiterpenes That Present Some Anti-
Inflammatory Effects
Mechanism of
Action In Vitro Assay In Vivo Assay Reference
Decrease of PGE2 Patchoulene epoxyde Patchoulene (Liang et al., 2018)
production epoxyde
Decrease the NO Nootkatone, valencene, Patchoulene (Tsoyi et al., 2011);
production patchoulene epoxyde, epoxyde (Wu et al., 2018,
β-patchoulene, and some 2017); (Liang et al.,
sesquiterpene lactones 2018)
HMGB-1 and Nootkatone and valencene Nootkatone and (Tsoyi et al., 2011)
HO-1 valencene
production
Inhibits the Patchoulene epoxyde and Patchoulene (Wu et al., 2018);
expression of β-patchoulene epoxyde and (Liang et al., 2018);
TNF-α nootkatone (Nemmar et al.,
2018)
Interleukins Patchoulene epoxyde and Patchoulene (Wu et al., 2018);
inhibition β-patchoulene epoxyde and (Liang et al., 2018)
β-patchoulene
Neutrophil Nootkatone (Nemmar et al., 2018)
migration
Reduction of Nootkatone, valencene, Patchoulene (Tsoyi et al., 2011);
iNOS expression patchoulene epoxyde, epoxyde (Wu et al., 2018)
β-patchoulene, and some
sesquiterpene lactones
Reduction of Patchoulene epoxyde and (Wu et al., 2018)
MCP-1 levels β-patchoulene
Reduction of Sesquiterpene lactones Patchoulene (Formisano et al.,
NF-κβ levels epoxyde and 2017); (Liang et al.,
nootkatone 2018); (Nemmar et
al., 2018)
HMGB, high mobility group box1; HO-1, heme oxygenase-1; MCP-1, monocyte chemotactic pro-
tein-1; NF-κβ, nuclear factor-kappa β; NO, nitric oxide; iNOS, nitric oxide synthase; PGE2, prosta-
glandin E2; TNF-α, tumor necrosis factor-α.
708 Food Aroma Evolution

32.6 CONCLUDING REMARKS

Based on the information about the bioactive potential of sesquiterpenes, specifically

for antioxidant, anticancer, antimicrobial, and anti-inflammatory assessment, it is pos-
sible to verify that many advances have been achieved for different sesquiterpenes with
promising results in several studies, both in vitro and/or in vivo. However, more research
is required to prove their efficacy, as well as their mechanism of action, optimal recom-
mended doses, and correct treatment; thus, developing safe and effective commercial
products for industrial applications in the food, cosmetic, and pharmaceuticals indus-
tries, as well as for the treatment of a wide and complex class of diseases that have drasti-
cally affected the population over the years.

REFERENCES

Abbas, Manal A., Mutasem O. Taha, Malek A. Zihlif, and Ahmad M. Disi. 2013.
“β-Caryophyllene Causes Regression of Endometrial Implants in a Rat Model of
Endometriosis Without Affecting Fertility.” European Journal of Pharmacology
702 (1–3) (February): 12–9. doi:10.1016/j.ejphar.2013.01.011.
Afoulous, Samia, Hicham Ferhout, Emmanuel Guy Raoelison, Alexis Valentin, Béatrice
Moukarzel, François Couderc, and Jalloul Bouajila. 2013. “Chemical Composition
and Anticancer, Antiinflammatory, Antioxidant and Antimalarial Activities of
Leaves Essential Oil of Cedrelopsis Grevei.” Food and Chemical Toxicology 56
(June): 352–62. doi:10.1016/j.fct.2013.02.008.
Alarif, Walied M., Sultan S. Al-lihaibi, Seif-eldin N. Ayyad, Mohamed H. Abdel-rhman,
and Farid A. Badria. 2012. “European Journal of Medicinal Chemistry Laurene-
Type Sesquiterpenes from the Red Sea Red Alga Laurencia Obtusa as Potential
Antitumor e Antimicrobial Agents.” European Journal of Medicinal Chemistry 55.
Elsevier Masson SAS: 462–6. doi:10.1016/j.ejmech.2012.06.060.
Al-dabbas, Maher M., Fumio Hashinaga, Samir A. M. Abdelgaleil, Toshihiko Suganuma,
Kohki Akiyama, and Hideo Hayashi. 2005. “Antibacterial Activity of an Eudesmane
Sesquiterpene Isolated from Common Varthemia, Varthemia Iphionoides.” 97:
237–40. doi:10.1016/j.jep.2004.11.007.
Amaya, Susana, José A. Pereira, Susana A. Borkosky, Juan C. Valdez, Alicia Bardón, and
Mario E. Arena. 2012. “Phytomedicine Inhibition of Quorum Sensing in Pseudomonas
Aeruginosa by Sesquiterpene Lactones.” European Journal of Integrative Medicine
19 (13). Elsevier GmbH.: 1173–7. doi:10.1016/j.phymed.2012.07.003.
AO, Wu-Li-Ji, Qing-Hu Wang, Si-Qin, Mu-Dan, Sa-Ren-Tu-Ya, Na-Yin-Tai Dai, and
Du-Ri-Si-Ha-La-Tu. 2012. “The Structural Elucidation and Antimicrobial Activities
of Two New Sesquiterpenes from Syringa pinnatifolia Hemsl.” Chinese Journal of
Natural Medicines 10 (6): 475–8. doi:10.1016/S1875-5364(12)60090-9.
Aruoma, Okezie I. 1998. “Free Radicals, Oxidative Stress, and Antioxidants in Human
Health and Disease.” Journal of the American Oil Chemists’ Society 75 (2). John
Wiley & Sons: 199–212. https://doi.org​/10.1​0 07/s​11746​-998-​0 032-​9.
Ascari, Jociani, Murilo Silva de Oliveira, Domingos Sávio Nunes, Daniel Granato,
Dilamara Riva Scharf, Edésio Simionatto, Michel Otuki, Bruna Soley, and Gustavo
Heiden. 2019. “Chemical Composition, Antioxidant and Anti-Inflammatory
Activities of the Essential Oils from Male and Female Specimens of Baccharis
punctulata (Asteraceae).” Journal of Ethnopharmacology 234 (April). Elsevier: 1–7.
https​://do​i.org​/10.1​016/J​.JEP.​2019.​01.00​5.
 Bioactive Potential of Sesquiterpenes 709

Babaei, Ghader, Azadeh Aliarab, Sina Abroon, Yusof Rasmi, and Shiva Gholizadeh-
Ghaleh Aziz. 2018. “Application of Sesquiterpene Lactone: A New Promising Way
for Cancer Therapy Based on Anticancer Activity.” Biomedicine & Pharmacotherapy
106 (October): 239–46. doi:10.1016/j.biopha.2018.06.131.
Becker, Daniel E. 2013. “Antimicrobial Drugs.” Anesthesia Progress 60: 111–23.
Bhatti, Haq Nawaz, Saleha Suleman Khan, Ajmal Khan, Mubeen Rani, Viqar Uddin
Ahmad, and Muhammad Iqbal Choudhary. 2014. “Biotransformation of
Monoterpenoids and Their Antimicrobial Activities.” Phytomedicine 21. Elsevier
GmbH.: 1597–626. doi:10.1016/j.phymed.2014.05.011.
Bi, Xiaoxu, Li Han, Tiange Qu, Yu Mu, Peipei Guan, Xiaodan Qu, Zhanyou Wang, and
Xueshi Huang. 2017. “Anti-Inflammatory Effects, SAR, and Action Mechanism of
Monoterpenoids from Radix Paeoniae Alba on LPS-Stimulated RAW 264.7 Cells.”
Molecules 22 (5): 715. doi:10.3390/molecules22050715.
Bommareddy, Ajay, Brittny Rule, Adam L. VanWert, Sreevidya Santha, and Chandradhar
Dwivedi. 2012. “α-Santalol, a Derivative of Sandalwood Oil, Induces Apoptosis in
Human Prostate Cancer Cells by Causing Caspase-3 Activation.” Phytomedicine 19
(8–9) (June): 804–11. doi:10.1016/j.phymed.2012.04.003.
Bommareddy, Ajay, Karryn Crisamore, Sarah Fillman, Sarah Brozena, James Steigerwalt,
Terra Landis, Adam L. Vanwert, and Chandradhar Dwivedi. 2016. “Abstract 4608:
Survivin Downregulation by α-Santalol Is Not Mediated through PI3K-Akt Pathway
in Human Breast Cancer Cells.” Cancer Research 76 (Suppl. 14) (July 15): 4608 .
doi:10.1158/1538-7445.am2016-4608.
Braicu, Cornelia, Nikolay Mehterov, Boyan Vladimirov, Victoria Sarafian, Seyed
Mohammad Nabavi, Atanas G. Atanasov, and Ioana Berindan-Neagoe. 2017.
“Nutrigenomics in Cancer: Revisiting the Effects of Natural Compounds.” Seminars
in Cancer Biology 46 (October): 84–106. doi:10.1016/j.semcancer.2017.06.011.
Breen, Sibilah, Sarah Kofoed, David Ritchie, Tracey Dryden, Roma Maguire, Nora
Kearney, and Sanchia Aranda. 2017. “Remote Real-Time Monitoring for
Chemotherapy Side-Effects in Patients with Blood Cancers.” Collegian 24 (6)
(December): 541–9. doi:10.1016/j.colegn.2016.10.009.
Breitmaier, Eberhard. 2006. Terpenes. Weinheim, Germany: Wiley-VCH Verlag GmbH
& Co. KGaA. https://doi.org/10.1002/9783527609949.
Brewer, M. S. 2011. “Natural Antioxidants: Sources, Compounds, Mechanisms of
Action, and Potential Applications.” Comprehensive Reviews in Food Science and
Food Safety 10 (4). John Wiley & Sons (10.1111): 221–47. https​://do​i.org​/10.1​111/j​
.1541​- 4337​. 2011​.0015​6.x.
Bunbamrung, Nantiya, Chakapong Intaraudom, Aibrohim Dramae, Nattawut Boonyuen,
Sukitaya Veeranondha, Pranee Rachtawee, and Pattama Pittayakhajonwut. 2017.
“Phytochemistry Letters Antimicrobial Activity of Illudalane and Alliacane
Sesquiterpenes from the Mushroom Gloeostereum Incarnatum BCC41461.”
Phytochemistry Letters 20 (May). Elsevier: 274–81. doi:10.1016/j.phytol.2017.05.017.
Calleja, Miguel Angel, Jose María Vieites, Trinidad Montero-Meterdez, María Isabel
Torres, María José Faus, Angel Gil, and Antonio Suárez. 2013. “The Antioxidant
Effect of β-Caryophyllene Protects Rat Liver from Carbon Tetrachloride-Induced
Fibrosis by Inhibiting Hepatic Stellate Cell Activation.” British Journal of Nutrition
109 (03). Cambridge University Press: 394–401. https​://do​i.org​/10.1​017/S​00071​14512​
00129​8.
Chadwick, Martin, Harriet Trewin, Frances Gawthrop, and Carol Wagstaff. 2013.
“Sesquiterpenoids Lactones: Benefits to Plants and People.” International Journal
of Molecular Sciences 14 (6): 12780–805. doi:10.3390/ijms140612780.
710 Food Aroma Evolution

Chang, Hyun-Joo, Ji-Myung Kim, Jae-Chul Lee, Won-Ki Kim, and Hyang Sook Chun.
2013. “Protective Effect of β-Caryophyllene, a Natural Bicyclic Sesquiterpene,
Against Cerebral Ischemic Injury.” Journal of Medicinal Food 16 (6): 471–80.
https://doi.org/10.1089/jmf.2012.2283.
Cui, Qingbin, Jing-Quan Wang, Yehuda G. Assaraf, Liang Ren, Pranav Gupta, Liuya
Wei, Charles R. Ashby, Dong-Hua Yang, and Zhe-Sheng Chen. 2018. “Modulating
ROS to Overcome Multidrug Resistance in Cancer.” Drug Resistance Updates 41
(November): 1–25. doi:10.1016/j.drup.2018.11.001.
Dahham, Saad, Yasser Tabana, Muhammad Iqbal, Mohamed Ahamed, Mohammed
Ezzat, Aman Majid, and Amin Majid. 2015. “The Anticancer, Antioxidant and
Antimicrobial Properties of the Sesquiterpene β-Caryophyllene from the Essential
Oil of Aquilaria Crassna.” Molecules 20 (7). Multidisciplinary Digital Publishing
Institute: 11808–29. https​://do​i.org ​/10.3​390/m​olecu​les20​07118​08.
de Cássia da Silveira e Sá, Rita, Luciana Andrade, and Damião de Sousa. 2013. “A
Review on Anti-Inflammatory Activity of Monoterpenes.” Molecules 18 (1): 1227–
54. doi:10.3390/molecules18011227.
Degenhardt, Jörg, Tobias G. Köllner, and Jonathan Gershenzon. 2009. “Monoterpene
and Sesquiterpene Synthases and the Origin of Terpene Skeletal Diversity in Plants.”
Phytochemistry 70 (15–6). Pergamon: 1621–37. https​://do​i.org​/10.1​016/J​.PHYT​
OCHEM​. 2009​.07.0​30.
de Oliveira Felipe, Lorena, Ana Maria de Oliveira, and Juliano Lemos Bicas. 2017.
“Bioaromas: Perspectives for Sustainable Development.” Trends in Food Science
and Technology, 62 (April): 141–53. https://doi.org/10.1016/j.tifs.2017.02.005.
de Santana Souza, Marilia Trindade, Jackson Roberto Guedes da Silva Almeida, Adriano
Antunes de Souza Araujo, Marcelo Cavalcante Duarte, Daniel Pens Gelain, José
Cláudio Fonseca Moreira, Marcio Roberto Viana dos Santos, and Lucindo José
Quintans-Júnior. 2014. “Structure-Activity Relationship of Terpenes with Anti-
Inflammatory Profile—A Systematic Review.” Basic & Clinical Pharmacology &
Toxicology 115 (3): 244–56. doi:10.1111/bcpt.12221.
Diemer, Andrea Wolf. 2016. AÇÃO ANTIMICROBIANA DE Rosmarinus Officinalis
E Zingiber Officinale FRENTE A Escherichia Coli E Staphylococcus Aureus EM
AÇÃO ANTIMICROBIANA DE Rosmarinus officinalis E Zingiber officinale
FRENTE A Escherichia Coli E Staphylococcus Aureus EM. UNIVATES.
Djeddi, Samah, Anastasia Karioti, Marina Sokovic, and Helen Skaltsa. 2008. “A Novel
Sesquiterpene Lactone from Centaurea Pullata : Structure Elucidation, Antimicrobial
Activity, and Prediction of Pharmacokinetic Properties.” Bioorganic & Medicinal
Chemistry 16: 3725–31. doi:10.1016/j.bmc.2008.01.056.
Duan, Yuan-chang, Jun Feng, Na Bai, Guo-hong Li, Ke-qin Zhang, and Pei-ji Zhao.
2018. “Fitoterapia Four Novel Antibacterial Sesquiterpene-α-Amino Acid
Quaternary Ammonium Hybrids from the Mycelium of Mushroom Stereum hir-
sutum.” Fitoterapia 128 (May). Elsevier: 213–7. doi:10.1016/j.fitote.2018.05.026.
Duarte, M. C. T., R. M. T. Duarte, R. A. F. Rodrigues, and M. V. N. Rodrigues. 2017.
“Essential Oils and Their Characteristics.” In Essential Oils in Food Processing, pp.
1–19. Wiley. https​://do​i.org​/10.1​0 02/9​78111​91493​92.ch​1.
Epifano, Francesco, Salvatore Genovese, Serena Fiorito, Véronique Mathieu, and
Robert Kiss. 2014. “Lapachol and Its Congeners as Anticancer Agents: A Review.”
Phytochemistry Reviews 13 (1) (March): 37–49. doi:10.1007/s11101-013-9289-1.
 Bioactive Potential of Sesquiterpenes 711

Eslahi, Hassan, Nafiseh Fahimi, and Ali Reza Sardarian. 2017. “Chemical Composition
of Essential Oils.” In Essential Oils in Food Processing, pp. 119–71. Wiley. https​://
do​i.org​/10.1​0 02/9​78111​91493​92.ch​4.
Feltran, Gabriel Primini. 2014. “Estudo Comparativo da Complexação de Monoterpenos
em Ciclodextrina: Preparação, Caracterização Química, Desenvolvimento
Tecnológico e Avaliação Biológica.” Master’s dissertation, Universidade Estadual de
Campinas, Sao Paolo, Brazil.
Fidyt, Klaudyna, Anna Fiedorowicz, Leon Strządała, and Antoni Szumny. 2016.
“β-Caryophyllene and β-Caryophyllene Oxide-Natural Compounds of Anticancer
and Analgesic Properties.” Cancer Medicine 5 (10) (September 30): 3007–17.
doi:10.1002/cam4.816.
Figueiredo, Pablo Luis B., Laine C. Pinto, Jamile S. da Costa, Alberto Ray C. da Silva, Rosa
Helena V. Mourão, Raquel C. Montenegro, Joyce Kelly R. da Silva, and José Guilherme
S. Maia. 2019. “Composition, Antioxidant Capacity and Cytotoxic Activity of Eugenia
Uniflora L. Chemotype-Oils from the Amazon.” Journal of Ethnopharmacology 232
(March). Elsevier: 30–8. https​://do​i.org​/10.1​016/J​.JEP.​2018.​12.01​1.
Formisano, Carmen, Cinzia Sanna, Mauro Ballero, Giuseppina Chianese, Carmina
Sirignano, Daniela Rigano, Estrella Millán, Eduardo Muñoz, and Orazio
Taglialatela-Scafati. 2017. “Anti-Inflammatory Sesquiterpene Lactones from
Onopordum Illyricum L. (Asteraceae), an Italian Medicinal Plant.” Fitoterapia 116
(January): 61–5. doi:10.1016/j.fitote.2016.11.006.
Fortuna, Antonio M, Zaida N Juárez, Horacio Bach, Anouf Nematallah, Yossef Av-gay,
Eugenio Sánchez-arreola, C A N Catalán, S Turbay, and Luis R Hernández. 2011.
“Phytochemistry Antimicrobial Activities of Sesquiterpene Lactones and Inositol
Derivatives from Hymenoxys Robusta.” Phytochemistry 72 (18). Elsevier: 2413–8.
doi:10.1016/j.phytochem.2011.09.001.
Fraatz, M., R. Berger, and H. Zorn. 2009. “Nootkatone—A Biotechnological Challenge.”
Applied Microbiology and Biotechnology 83: 35–41.
Fransen, Marc, Marcus Nordgren, Bo Wang, and Oksana Apanasets. 2012. “Role
of Peroxisomes in ROS/RNS-Metabolism: Implications for Human Disease.”
Biochimica et Biophysica Acta (BBA)—Molecular Basis of Disease 1822 (9).
Elsevier: 1363–73. https​://do​i.org​/10.1​016/J​.BBAD​IS.20​11.12​.001.​
Galadari, Sehamuddin, Anees Rahman, Siraj Pallichankandy, and Faisal Thayyullathil.
2017. “Molecular Targets and Anticancer Potential of Sanguinarine—A
Benzophenanthridine Alkaloid.” Phytomedicine 34 (October): 143–53. doi:10.1016/j.
phymed.2017.08.006.
Ghasemian, Mona, Sina Owlia, and Mohammad Bagher Owlia. 2016. “Review of Anti-
Inflammatory Herbal Medicines.” Advances in Pharmacological Sciences 2016:
1–11. doi:10.1155/2016/9130979.
Gliszczyńska, Anna, Agnieszka Łysek, Tomasz Janeczko, Marta Świtalska, Joanna
Wietrzyk, and Czesław Wawrzeńczyk. 2011. “Microbial Transformation of
(+)-Nootkatone and the Antiproliferative Activity of Its Metabolites.” Bioorganic &
Medicinal Chemistry 19 (7) (April): 2464–9. doi:10.1016/j.bmc.2011.01.062.
Gonzalez-Burgos, E. and M. P. Gomez-Serranillos. 2012. “Terpene Compounds in
Nature: A Review of Their Potential Antioxidant Activity.” Current Medicinal
Chemistry 19 (31): 5319–41. https​://do​i.org​/10.2​174/0​92986​71280​38333​35.
Graßmann, J. 2005. “Terpenoids as Plant Antioxidants.” Vitamins & Hormones 72
(January). Academic Press: 505–35. https​://do​i.org​/10.1​016/S​0083-​6729(​05)72​015-X​.
712 Food Aroma Evolution

Gutteridge, John M. C., and Barry Halliwell. 2010. “Antioxidants: Molecules, Medicines,
and Myths.” Biochemical and Biophysical Research Communications 393: 561–4.
https​://do​i.org​/10.1​016/j​.bbrc​. 2010​.02.0​71.
Halliwell, B. 2007. “Biochemistry of Oxidative Stress.” Biochemical Society Transactions
35 (Pt 5). Portland Press: 1147–50. https://doi.org/10.1042/BST0351147.
Hanušová, Veronika, Kateřina Caltová, Hana Svobodová, Martin Ambrož, Adam
Skarka, Natálie Murínová, Věra Králová, Pavel Tomšík, and Lenka Skálová. 2017.
“The Effects of β-Caryophyllene Oxide and Trans-Nerolidol on the Efficacy of
Doxorubicin in Breast Cancer Cells and Breast Tumor-Bearing Mice.” Biomedicine
& Pharmacotherapy 95 (November): 828–36. doi:10.1016/j.biopha.2017.09.008.
Hashidoko, Yasuyuki, Satoshi Tahara, and Junya Mizutani. 1989. “Antimicrobial from
Damaged Rosa Rugosa Leaves.” Phytochemistry 28 (2): 2–7.
Heras, B. and Sonsoles Hortelano. 2009. “Molecular Basis of the Anti-Inflammatory
Effects of Terpenoids.” Inflammation & Allergy-Drug Targets 8 (1): 28–39.
doi:10.2174/187152809787582534.
Incalza, Maria Angela, Rossella D’Oria, Annalisa Natalicchio, Sebastio Perrini, Luigi
Laviola, and Francesco Giorgino. 2018. “Oxidative Stress and Reactive Oxygen
Species in Endothelial Dysfunction Associated with Cardiovascular and Metabolic
Diseases.” Vascular Pharmacology 100 (January). Elsevier: 1–19. https​://do​i.org​
/10.1​016/J​.VPH.​2017.​05.00​5.
Iqbal, Javed, Banzeer Ahsan Abbasi, Tariq Mahmood, Sobia Kanwal, Barkat Ali, Sayed
Afzal Shah, and Ali Talha Khalil. 2017. “Plant-Derived Anticancer Agents: A
Green Anticancer Approach.” Asian Pacific Journal of Tropical Biomedicine 7 (12)
(December): 1129–50. doi:10.1016/j.apjtb.2017.10.016.
Jahangir, Tamanna, Tajdar Husain Khan, Lakshmi Prasad, and Sarwat Sultana. 2005.
“Alleviation of Free Radical Mediated Oxidative and Genotoxic Effects of Cadmium
by Farnesol in Swiss Albino Mice.” Redox Report 10 (6). Taylor & Francis: 303–10.
https://doi.org/10.1179/135100005X83671.
Janssens, L., H. L. De Pooter, N. M. Schamp, and E. J. Vandamme. 1992. “Production
of Flavours by Microorganisms.” Process Biochemistry 27: 195–215.
Johnson, K., J. Chang-Claude, A.-M. Critchley, C. Kyriacou, S. Lavers, T. Rattay, P.
Seibold, et al. 2019. “Genetic Variants Predict Optimal Timing of Radiotherapy to
Reduce Side-Effects in Breast Cancer Patients.” Clinical Oncology 31 (1) (January):
9–16. doi:10.1016/j.clon.2018.10.001.
Kanavos, P. 2006. “The Rising Burden of Cancer in the Developing World.” Annals of
Oncology 17 (Suppl. 8) (July 1): viii15–viii23. doi:10.1093/annonc/mdl983.
Kapoor, S. 2013. “d-Limonene.” Human & Experimental Toxicology 32 (11) (May 28):
1228 . doi:10.1177/0960327113489053.
Khan, Rehan and Sarwat Sultana. 2011. “Farnesol Attenuates 1,2-Dimethylhydrazine
Induced Oxidative Stress, Inflammation and Apoptotic Responses in the Colon of
Wistar Rats.” Chemico-Biological Interactions 192 (3). Elsevier: 193–200. https​://
do​i.org​/10.1​016/J​.CBI.​2011.​03.00​9.
Ku, Chi-Mei and Jin-Yuarn Lin. 2015. “Farnesol, a Sesquiterpene Alcohol in Herbal
Plants, Exerts Anti-Inflammatory and Antiallergic Effects on Ovalbumin-
Sensitized and -Challenged Asthmatic Mice.” Evidence-Based Complementary
and Alternative Medicine : ECAM 2015 (April). Hindawi: 387357. https://doi.
org/10.1155/2015/387357.
 Bioactive Potential of Sesquiterpenes 713

Kumari, Sangita, Piyush Priya, Gopal Misra, and Gitanjali Yadav. 2013. “Structural and
Biochemical Perspectives in Plant Isoprenoid Biosynthesis.” Phytochemistry Reviews
12 (2). Springer Netherlands: 255–91. https​://do​i.org​/10.1​0 07/s​11101​- 013-​9284-​6.
Kuttan, Ramadasan, Kottarapat Jeena, and Vijayastelter B. Liju. 2013. “Antioxidant,
Anti-Inflammatory and Antinociceptive Activities of Essential Oil from Ginger.”
Indian Journal of Physiology and Pharmacology 57. http:​//nat​urali​ngred​ient.​org/w​
p/wp-​conte​nt/up​loads​/51-6​2 .pdf​.
Lan, Yu-Hsuan, Yang-Chang Wu, Kai-Wei Wu, Jing-Gung Chung, Chi-Cheng Lu, Yuan-
Liang Chen, Tian-Shung Wu, and Jai-Sing Yang. 2011. “Death Receptor 5-Mediated
TNFR Family Signaling Pathways Modulate γ-Humulene-Induced Apoptosis in
Human Colorectal Cancer HT29 Cells.” Oncology Reports 25 (2) (February 1).
doi:10.3892/or.2010.1087.
Langhasova, Lenka, Veronika Hanušová, Jan Rezek, Barbora Stohanslova, Martin
Ambrož, Věra Králová, Tomas Vanek, et al. 2014. “Essential Oil from Myrica rubra
Leaves Inhibits Cancer Cell Proliferation and Induces Apoptosis in Several Human
Intestinal Lines.” Industrial Crops and Products 59 (August): 20–6. doi:10.1016/j.
indcrop.2014.04.018.
Lateef, Abdul, Muneeb U. Rehman, Mir Tahir, Rehan Khan, Abdul Quaiyoom Khan,
Wajhul Qamar, and Sarwat Sultana. 2013. Article in Toxicology International.
https​://do​i.org ​/10.4​103/0​971-6​580.1​11563​.
Lee, Brigette, Jonathan Bohmann, Tony Reeves, Corey Levenson, and April L. Risinger.
2015. “α- and β-Santalols Directly Interact with Tubulin and Cause Mitotic Arrest
and Cytotoxicity in Oral Cancer Cells.” Journal of Natural Products 78 (6) (May
20): 1357–62. doi:10.1021/acs.jnatprod.5b00207.
Li, Hui, Wei Guo, Xiao-jing Ma, Jia-shan Li, and Xin Song. 2016. “In Vitro and in
Vivo Anticancer Activity of Sophorolipids to Human Cervical Cancer.” Applied
Biochemistry and Biotechnology 181 (4) (October 29): 1372–87. doi:10.1007/
s12010-016-2290-6.
Li, Shou-Jie, Xuan Zhang, Xiang-Hua Wang, and Chang-Qi Zhao. 2018. “Novel Natural
Compounds from Endophytic Fungi with Anticancer Activity.” European Journal
of Medicinal Chemistry 156 (August): 316–43. doi:10.1016/j.ejmech.2018.07.015.
Liang, Jiali, Yaoxing Dou, Xue Wu, Huilin Li, Jiazhen Wu, Qionghui Huang, Dandan
Luo, et al. 2018. “Prophylactic Efficacy of Patchoulene Epoxide against Ethanol-
Induced Gastric Ulcer in Rats: Influence on Oxidative Stress, Inflammation and
Apoptosis.” Chemico-Biological Interactions 283 (January): 30–7. doi:10.1016/j.
cbi.2018.01.014.
Liu, Rui Hai. 2004. “Potential Synergy of Phytochemicals in Cancer Prevention:
Mechanism of Action.” Journal of Nutrition 134 (12). Oxford University Press:
3479S–85S. https://doi.org/10.1093/jn/134.12.3479S.
Liu, Zewen, Zhangpin Ren, Jun Zhang, Chia-Chen Chuang, Eswar Kandaswamy,
Tingyang Zhou, and Li Zuo. 2018. “Role of ROS and Nutritional Antioxidants in
Human Diseases.” Frontiers in Physiology 9 (May). Frontiers Media SA: 477. https​
://do​i.org​/10.3​389/f​phys.​2018.​0 0477​.
Lorenzo, Jose M., Paulo E. S. Munekata, Belen Gómez, Francisco J. Barba, Leticia Mora,
Cristina Pérez-Santaescolástica, and Fidel Toldrá. 2018. “Bioactive Peptides as
Natural Antioxidants in Food Products—A Review.” Trends in Food Science &
Technology 79 (September). Elsevier: 136–47. https​://do​i.org​/10.1​016/J​.TIFS​. 2018​
.07.0​03.
714 Food Aroma Evolution

Ludwiczuk, A., K. Skalicka-Wó Zniak, and M. I. Georgiev. 2016. “Terpenoids Learning

Objectives G.” Pharmacognosy, 233–66. https​://do​i.org​/10.1​016/B​978-0​-12-8​
02104​- 0.00​011-1​.
Marcadenti, Aline. 2015. “Dietary Antioxidant and Oxidative Stress: Interaction between
Vitamins and Genetics.” Journal of Nutritional Health & Food Science 3 (1). https​
://do​i.org​/10.1​5226/​jnhfs​. 2015​.0013​8.
Matkowski, Adam. 2008. “Plant in Vitro Culture for the Production of Antioxidants—A
Review.” Biotechnology Advances 26 (6). Elsevier: 548–60. https​://do​i.org​/10.1​
016/J​.BIOT​ECHAD​V.200​8.07.​0 01.
Mazumder, Aloran, Claudia Cerella, and Marc Diederich. 2018. “Natural Scaffolds
in Anticancer Therapy and Precision Medicine.” Biotechnology Advances 36 (6)
(November): 1563–85. doi:10.1016/j.biotechadv.2018.04.009.
Min, Y. N., Z. Y. Niu, T. T. Sun, Z. P. Wang, P. X. Jiao, B. B. Zi, P. P. Chen, D. L.
Tian, and F. Z. Liu. 2018. “Vitamin E and Vitamin C Supplementation Improves
Antioxidant Status and Immune Function in Oxidative-Stressed Breeder Roosters
by up-Regulating Expression of GSH-Px Gene.” Poultry Science 97 (4). Oxford
University Press: 1238–44. https://doi.org/10.3382/ps/pex417.
Nemmar, Abderrahim, Suhail Al-Salam, Sumaya Beegam, Priya Yuvaraju, Naserddine
Hamadi, and Badreldin H. Ali. 2018. “In Vivo Protective Effects of Nootkatone
against Particles-Induced Lung Injury Caused by Diesel Exhaust Is Mediated via the
NF-κB Pathway.” Nutrients 10 (263). doi:10.3390/nu10030263.
Nile, Shivraj Hariram and Se Won Park. 2013. “Optimized Methods for In Vitro and In
Vivo Anti-Inflammatory Assays and Its Applications in Herbal and Synthetic Drug
Analysis Optimized Methods for In Vitro and In Vivo Anti-Inflammatory Assays and
Its Applications in Herbal and Synthetic Drug Analysis.” Mini-Reviews in Medicinal
Chemistry 13 (95): 95–100. doi:10.2174/138955713804484712.
Nimse, Satish Balasaheb and Dilipkumar Pal. 2015. “Free Radicals, Natural Antioxidants,
and Their Reaction Mechanisms.” RSC Advances 5 (35). Royal Society of Chemistry:
27986. https://doi.org/10.1039/C4RA13315C.
Nonato, Fabiana Regina, Danielle Gomes Santana, Flavielle Martins de Melo, Gisele
Graça Leite dos Santos, Danielle Brustolim, Enilton A. Camargo, Damião P. de
Sousa, Milena Botelho Pereira Soares, and Cristiane Flora Villarreal. 2012. “Anti-
Inflammatory Properties of Rose Oxide.” International Immunopharmacology 14
(4): 779–84. doi:10.1016/j.intimp.2012.10.015.
Nuutinen, Tarmo. 2018. “Medicinal Properties of Terpenes Found in Cannabis Sativa and
Humulus Lupulus.” European Journal of Medicinal Chemistry 157 (September):
198–228. doi:10.1016/j.ejmech.2018.07.076.
Oprea, Adriana D., Raymond R. Russell, Kerry S. Russell, and Maysa Abu-Khalaf.
2017. “Chemotherapy Agents with Known Cardiovascular Side Effects and Their
Anesthetic Implications.” Journal of Cardiothoracic and Vascular Anesthesia 31 (6)
(December): 2206–26. doi:10.1053/j.jvca.2015.06.020.
Ortiz, Carmen, Luisa Morales, Miguel Sastre, William E. Haskins, and Jaime Matta.
2016. “Cytotoxicity and Genotoxicity Assessment of Sandalwood Essential Oil in
Human Breast Cell Lines MCF-7 and MCF-10A.” Evidence-Based Complementary
and Alternative Medicine 2016: 1–13. doi:10.1155/2016/3696232.
Paduch, Roman, Mariusz Trytek, Sylwia K. Król, Joanna Kud, Maciej Frant, Martyna
Kandefer-Szerszeń, and Jan Fiedurek. 2016. “Biological Activity of Terpene
Compounds Produced by Biotechnological Methods.” Pharmaceutical Biology 54
(6) (January 25): 1096–107. doi:10.3109/13880209.2015.1103753.
 Bioactive Potential of Sesquiterpenes 715

Pavithra, P. S., Alka Mehta, and Rama S. Verma. 2018a. “Aromadendrene Oxide 2,
Induces Apoptosis in Skin Epidermoid Cancer Cells through ROS Mediated
Mitochondrial Pathway.” Life Sciences 197: 19–29. doi:10.1016/j.lfs.2018.
01.029.
Pavithra, P. S., Alka Mehta, and Rama S. Verma. 2018b. “Induction of Apoptosis by
Essential Oil from P. Missionis in Skin Epidermoid Cancer Cells.” Phytomedicine
50: 184–95. doi:10.1016/j.phymed.2017.11.004.
Pavithra, P. S., Alka Mehta, and Rama S. Verma. 2018c. “Synergistic Interaction of
β-Caryophyllene with Aromadendrene Oxide 2 and Phytol Induces Apoptosis on
Skin Epidermoid Cancer Cells.” Phytomedicine 47 (August): 121–34. doi:10.1016/j.
phymed.2018.05.001.
Pham-Huy, Lien Ai, Hua He, and Chuong Pham-Huy. 2008. “Free Radicals, Antioxidants
in Disease and Health.” International Journal of Biomedical Science : IJBS 4 (2).
Master Publishing Group: 89–96. http:​//www​.ncbi​.nlm.​nih.g​ov/pu​bmed/​23675​073.
Pichersky, Eran and Robert A. Raguso. 2018. “Why Do Plants Produce so Many
Terpenoid Compounds?” New Phytologist 220 (3). John Wiley & Sons: 692–702.
https://doi.org/10.1111/nph.14178.
Picman, Anna K. 1984. “Antifungal Activity of Sesquiterpene Lactones.” Biochemical
Systematics and Ecology 12 (1): 13–8.
Pimentel, Mariana Recco. 2012. “Produção de Compostos de Aroma a Partir da
Biotransformação de Monoterpenos por Pseudomonas.” Master’s dissertation,
Universidade Estadual de Campinas, Sao Paolo, Brazil.
Poprac, Patrik, Klaudia Jomova, Miriama Simunkova, Vojtech Kollar, Christopher J.
Rhodes, and Marian Valko. 2017. “Targeting Free Radicals in Oxidative Stress-
Related Human Diseases.” Trends in Pharmacological Sciences 38 (7). Elsevier
Current Trends: 592–607. https​://do​i.org ​/10.1​016/J​.TIPS​. 2017​.04.0​05.
Powers, Scott K., Keith C. Deruisseau, John Quindry, and Karyn L. Hamilton. 2004.
“Dietary Antioxidants and Exercise.” Journal of Sports Sciences 22 (1): 81–94.
https​://do​i.org​/10.1​080/0​26404​10310​0 0140​563.
Prakash, Ved. 2017. “Terpenoids as Source of Anti-Inflammatory Compounds.”
Asian Journal of Pharmaceutical and Clinical Research 10 (3): 68. doi:10.22159/
ajpcr.2017.v10i3.16435.
Prior, Ronald L. 2015. “Oxygen Radical Absorbance Capacity (ORAC): New Horizons
in Relating Dietary Antioxidants/Bioactives and Health Benefits.” Journal of
Functional Foods 18 (October). Elsevier: 797–810. https​://do​i.org​/10.1​016/J​.JFF.​
2014.​12.01​8.
Quiroga, Patricia R., Valeria Nepote, and María T. Baumgartner. 2019. “Contribution of
Organic Acids to α-Terpinene Antioxidant Activity.” Food Chemistry 277 (March).
Elsevier: 267–72. https​://do​i.org​/10.1​016/J​.FOOD​CHEM.​2018.​10.10​0.
Rauter, Amélia P., Isabel Branco, Jaime Bermejo, Antonio G. González, Maria D.
Garcı ́a-Grávalos, and Arturo San Feliciano. 2001. “Bioactive Humulene Derivatives
from Asteriscus Vogelii.” Phytochemistry 56 (2) (January): 167–71. doi:10.1016/
s0031-9422(00)00304-6.
Ruberto, Giuseppe and Maria T. Baratta. 2000. “Antioxidant Activity of Selected
Essential Oil Components in Two Lipid Model Systems.” Food Chemistry 69 (2).
Elsevier: 167–74. https​://do​i.org​/10.1​016/S​0308-​8146(​99)00​247-2​.
Russo, Gian Luigi. 2007. “Ins and Outs of Dietary Phytochemicals in Cancer
Chemoprevention.” Biochemical Pharmacology. https​://do​i.org​/10.1​016/j​.bcp.​
2007.​02.01​4.
716 Food Aroma Evolution

Rutkowski, Ryszard, Sławomir A. Pancewicz, Krzysztof Rutkowski, and Joanna

Rutkowska. 2007. “Reactive Oxygen and Nitrogen Species in Inflammatory
Process.” Polski Merkuriusz Lekarski : Organ Polskiego Towarzystwa Lekarskiego
23 (134): 131–6. http:​//www​.ncbi​.nlm.​nih.g​ov/pu​bmed/​18044​345.
Salakhutdinov, Nariman F., Konstantin P. Volcho, and Olga I. Yarovaya. 2017.
“Monoterpenes as a Renewable Source of Biologically Active Compounds.” Pure
and Applied Chemistry 89 (8). doi:10.1515/pac-2017-0109.
Santhanasabapathy, R., S. Vasudevan, K. Anupriya, R. Pabitha, and G. Sudhandiran.
2015. “Farnesol Qu ells Oxidative Stress, Reactive Gliosis and Inflammation dur-
ing Acrylamide-Induced Neurotoxicity: Behavioral and Biochemical Evidence.”
Neuroscience 308 (November). Pergamon: 212–27. https​://do​i.org​/10.1​016/j​.neur​
oscie​nce.2​015.0​8.067​.
Santhanasabapathy, Rajasekaran and Ganapasam Sudhandiran. 2015. “Farnesol
Attenuates Lipopolysaccharide-Induced Neurodegeneration in Swiss Albino Mice
by Regulating Intrinsic Apoptotic Cascade.” Brain Research 1620 (September).
Elsevier: 42–56. https​://do​i.org​/10.1​016/J​.BRAI​N RES.​2015.​04.04​3.
Shahidi, Fereidoon. 2000. “Antioxidants in Food and Food Antioxidants.” Nahrung/
Food 44 (3). John Wiley & Sons: 158–63. https​ ://do​
i.org​
/10.1​
0 02/1​
521-3​
803(2​
00005​01)44​:3<15​8:AID​-FOOD​158>3​.0.CO​;2-L.​
Shahidi, Fereidoon and Ying Zhong. 2010. “Novel Antioxidants in Food Quality
Preservation and Health Promotion.” European Journal of Lipid Science and
Technology 112 (9). John Wiley & Sons: 930–40. https://doi.org/10.1002/ejlt.2010
00044.
Sharma, Anu, Matthew Campbell, Cassian Yee, Sangeeta Goswami, and Padmanee
Sharma. 2019. “Immunotherapy of Cancer.” In Clinical Immunology—Principles
and Practice, edited by Robert R. Rich, Thomas A. Fleisher, William T. Shearer,
Harry W. Schroeder, Jr., Anthony J. Frew, and Cornelia M. Weyand, pp. 1033–48.
e1. Amsterdam: Elsevier.
Shiono, Yoshihito, Hiromasa Koyama, Tetsuya Murayama, and Takuya Koseki. 2015.
“Phytochemistry Letters New Sesquiterpenes from the Endophyte Microdiplodia
Sp. T T-12 and Their Antimicrobial Activity.” Phytochemistry Letters 14.
Phytochemical Society of Europe: 143–7. doi:10.1016/j.phytol.2015.10.004.
Sowden, R. J., S. Yasmin, N. H. Rees, S. G. Bell, and L. L. Wong. 2005. “Biotransformation
of the Sesquiterpene (+)-Valencene by Cytochrome P450 Cam and P450 BM-3.”
Organic & Biomolecular Chemistry 3 (1): 57–64.
Szűcs, Gergő, Zsolt Murlasits, Szilvia Török, Gabriella F. Kocsis, János Pálóczi, Anikó
Görbe, Tamás Csont, Csaba Csonka, and Péter Ferdinandy. 2013. “Cardioprotection
by Farnesol: Role of the Mevalonate Pathway.” Cardiovascular Drugs and Therapy
27 (4): 269–77. https​://do​i.org​/10.1​0 07/s​10557​- 013-​6460-​2 .
Tabas, I. and C. K. Glass. 2013. “Anti-Inflammatory Therapy in Chronic Disease:
Challenges and Opportunities.” Science 339 (6116): 166–72. doi:10.1126/
science.1230720.
Tepe, Bektas, Munevver Sokmen, H. Askin Akpulat, Dimitra Daferera, Moschos
Polissiou, and Atalay Sokmen. 2005. “Antioxidative Activity of the Essential Oils
of Thymus Sipyleus Subsp. Sipyleus Var. Sipyleus and Thymus Sipyleus Subsp.
Sipyleus Var. Rosulans.” Journal of Food Engineering 66 (4): 447–54. https​://do​
i.org​/10.1​016/J​.JFOO​DENG.​2004.​04.01​5.
 Bioactive Potential of Sesquiterpenes 717

Torre, Lindsey A., Freddie Bray, Rebecca L. Siegel, Jacques Ferlay, Joannie Lortet-Tieulent,
and Ahmedin Jemal. 2015. “Global Cancer Statistics, 2012.” CA: A Cancer Journal
for Clinicians 65 (2) (February 4): 87–108. doi:10.3322/caac.21262.
Tsoyi, Konstantin, Hwa Jin Jang, Young Soo Lee, Young Min Kim, Hye Jung Kim,
Han Geuk Seo, Jae Heun Lee, Jong Hwan Kwak, Dong-ung Lee, and Ki Churl
Chang. 2011. “(+)-Nootkatone and (+)-Valencene from Rhizomes of Cyperus rotun-
dus Increase Survival Rates in Septic Mice Due to Heme Oxygenase-1 Induction.”
Journal of Ethnopharmacology 137 (3): 1311–7. doi:10.1016/j.jep.2011.07.062.
Uttara, Bayani, Ajay V. Singh, Paolo Zamboni, and R. T. Mahajan. 2009. “Oxidative
Stress and Neurodegenerative Diseases: A Review of Upstream and Downstream
Antioxidant Therapeutic Options.” Current Neuropharmacology 7 (65). Bentham
Science Publishers: 74. https​://ww ​w.ing​entac​onnec​t.com ​/cont​ent/b​en/cn ​/2009​
/0000​0 007/​0 0000​0 01/a​rt000​06#.
Valko, Marian, Dieter Leibfritz, Jan Moncol, Mark T. D. Cronin, Milan Mazur, and
Joshua Telser. 2007. “Free Radicals and Antioxidants in Normal Physiological
Functions and Human Disease.” The International Journal of Biochemistry & Cell
Biology 39 (1). Pergamon: 44–84. https​://do​i.org​/10.1​016/J​.BIOC​EL.20​06.07​.001.​
Vespermann, Kele A. C., Bruno N. Paulino, Mayara C. S. Barcelos, Marina G. Pessôa,
Glaucia M. Pastore, and Gustavo Molina. 2017. “Biotransformation of α- and
β-Pinene into Flavor Compounds.” Applied Microbiology and Biotechnology, 101
(5) (March): 1805–17.
Vieceli Dalla Sega, Francesco, Giorgio Aquila, Francesca Fortini, Mauro Vaccarezza, Paola
Secchiero, Paola Rizzo, and Gianluca Campo. 2017. “Context-Dependent Function of
ROS in the Vascular Endothelium: The Role of the Notch Pathway and Shear Stress.”
BioFactors 43 (4). Wiley-Blackwell: 475–85. https://doi.org/10.1002/biof.1359.
Wang, Fang, Huanhuan Zhong, Shiqi Fang, Yunfeng Zheng, Cunyu Li, Guoping Peng,
and Xinchun Shen. 2017. “Potential Anti-Inflammatory Sesquiterpene Lactones
from Eupatorium lindleyanum.” Planta Medicia 84 (2): 123–8. doi:10.1055/s-0043-
117742.
Wiart, Christophe. 2013. “Terpenes.” Lead Compounds from Medicinal Plants for the
Treatment of Cancer: 97–265. doi:10.1016/b978-0-12-398371-8.00002-7.
Wu, Xue, Jia-Li Liang, Yu-Hong Liu, Jia-Zhen Wu, Qiong-Hui Huang, Yu-Cui Li,
and Qing-Feng Xie. 2018. “Comparison of Anti-Inflammatory Effect between
β-Patchoulene Epoxide and β-Patchoulene in LPS-Stimulated RAW264.7
Macrophages.” European Journal of Inflammation 16 (55):205873921878507.
doi:10.1177/2058739218785075.
Wu, Zhong-Nan, Yu-Bo Zhang, Neng-Hua Chen, Mo-Jiao Li, Man-Mei Li, Wei Tang,
Ling Zhuang, Yao-Lan Li, and Guo-Cai Wang. 2017. “Sesquiterpene Lactones from
Elephantopus mollis and Their Anti-Inflammatory Activities.” Phytochemistry 137
(May): 81–6. doi:10.1016/j.phytochem.2017.01.020.
Yang, Xinzhou, Chao Wang, Jing Yang, Dingrong Wan, Qinxiong Lin, Guangzhong
Yang, Zhinan Mei, and Yunjiang Feng. 2014. “Antimicrobial Sesquiterpenes from
the Chinese Medicinal Plant, Chloranthus Angustifolius.” Tetrahedron Letters 55
(41). Elsevier: 5632–4. doi:10.1016/j.tetlet.2014.08.052.
Zhang, Xiaoying, Wei Chen, Ruth Guillermo, Gudiseva Chandrasekher, Radhey S.
Kaushik, Alan Young, Hesham Fahmy, and Chandradhar Dwivedi. 2010. “Alpha-
Santalol, a Chemopreventive Agent Against Skin Cancer, Causes G2/M Cell Cycle
718 Food Aroma Evolution

Arrest in Both P53-Mutated Human Epidermoid Carcinoma A431 Cells and P53
Wild-Type Human Melanoma UACC-62 Cells.” BMC Research Notes 3 (1) (August
3). doi:10.1186/1756-0500-3-220.
Zhao, Xingzeng, Shu Xu, Min Yin, Xiangyun Wang, Qizhi Wang, and Yu Shan. 2018.
“Phytochemistry Letters Six New Dihydro-β-Agarofuran Sesquiterpenes from the
Stems and Leaves of Monimopetalum chinense and Their Antimicrobial Activities.”
Phytochemistry Letters 27 (1). Elsevier: 160–6. doi:10.1016/j.phytol.2018.07.024.
Zuccolotto, Tatiana, Jaqueline Bressan, Allan Vinicius Félix Lourenço, Estevan
Bruginski, Andressa Veiga, Jane Marinho, Paola Aparecida Raeski, et al. 2019.
“Chemical, Antioxidant, and Antimicrobial Evaluation of Essential Oils and
an Anatomical Study of the Aerial Parts from Baccharis Species (Asteraceae).”
Chemistry & Biodiversity (February). John Wiley & Sons, cbdv.201800547. https://
doi.org/10.1002/cbdv.201800547.
Zuzarte, M., L. Salgueiro, Mónica Zuzarte, and Lígia Salgueiro. 2015. “Essential
Oils Chemistry: What Are Essential Oils?” https​://do​i.org​/10.1​0 07/9​78-3-​319-1​
9144-​7_2.
 Index
A Apocarotenoid, 80–81
Acids, 75–76, 395–396, 472–473 Aristotle’s perception, 15
Advanced glycation end products (AGEs), Aroma
288–290 additives (bioaromas), 298–299
AGEs, see Advanced glycation end products molecular point of view, 5–9
Alcoholic beverages, off-flavors in odor of perfume, 4–5
alcohols, 596–605 smell, 3–4
aldehydes, 605–606 taste, 3–4
anisoles, 606 vectors, 5–9
fatty acids, 596 Aroma compounds
furans, 606–607 biosynthesis
ketones, 607 amino acid derivates, 75–78
procedures carbohydrate-derived flavor compounds,
for preventing, 610–612 78–81
for reducing, 612–614 fatty acid-derived and lipophilic
pyrazines, 607–608 compounds, 70–74
sulfur compounds, 609–610 chemistry and organoleptic properties
volatile phenols, 608–609 alcohols, 68
Alcoholic fermentation, 297–298 aldehydes, 66–67
Alcohols, 68, 75–76, 388–391, 476–477, esters, 62–63
596–605, 637 furans, 68–69
Aldehydes, 66–67, 75–76, 391–393, 473–475, ketones, 67
605–606 lactones, 63–65
Alkalization process, 404 phenylpropenes, 67–68
α- and β-oxidation, straight-chain fatty acids pyrazines, 69
by, 73–74 sulfur compounds, 69–70
Amino acid derivates, 75–78 terpenoids, 65–66
acids, 75–76 volatile compounds, 61–62
alcohols, 75–76 flavor, concept of, 58
aldehydes, 75–76 natural aromas, 59–61
esters, 75–76 Atmospheric pressure chemical ionization
lactones, 75–76 mass spectrometry (APCI-MS), 161
N- and S-containing flavor molecules, 75–76 Atmospheric pressure ionization-mass
phenylpropenes and, 76–78 spectrometry (API MS), 164
Anisoles, 606
Anticancer potential, sesquiterpenes, 701–702 B
Anti-inflammatory effects, sesquiterpenes, Bakery
705–707 baked goods, 416–418
Antimicrobial potential, sesquiterpenes, biscuit and cracker elaboration, 420–421
703–705 bread and toast elaboration, 418
Antioxidant potential, sesquiterpenes, cake elaboration, 418–420
696–701 baking market, 415–416
APCI-MS, see Atmospheric pressure chemical biological point of view, 421–422
ionization mass spectrometry caramelization, 424–426
API MS, see Atmospheric pressure ionization- fermentation process, 422–423
mass spectrometry lipid oxidation process, 423–424

719
720 Index

Maillard reaction, 424–426 non-aqueous capillary electrophoresis

volatile compounds, during processing and (NACE), 198
storage, 426–427 theoretical principles, 197
aroma quality, 431–432 Capillary zone electrophoresis (CZE), 197
raw materials, 427–430 Caprylic flavor, 596
toasting of bread, 430–431 Caramelization, 272–274, 377, 424–426
Beer aroma Carbohydrate-derived flavor compounds,
brewing process, 357–358 78–81
esters, 358–359 apocarotenoid, 80–81
higher alcohols (fusel alcohols), 358 furanones, 78–79
diacetyl, 359–361 pyrones, 78–79
dimethyl sulfide (DMS), 359 terpenoid, 79–80
hop essential oils, 354–355 Carbohydrates, 633–636
alcohols, 355–356 Carvone (2-methyl-5-(1-methylethenyl)-
aldehydes, 356 2‑cyclohexen-1-one), 5
hydrocarbon fraction, 355 CE, see Capillary electrophoresis
4-hydroxi-4-methyl-2-pentenoic acid, Cheese and yogurt, 470–472
356–357 Chemical volatile compounds (CVC), 159
3-methylbutanoic acid, 356–357 Chemosensory system, flavor perception
4-methyl-3-pentenoic acid, 356–357 gustatory system
2-methylpropanoic acid, 356–357 anatomy and physiology, 27–29
Bimodal congruency, 40 sense of taste, 25–27
Binary taste interactions, 26 olfactory system
Bioaromas, 294 anatomy and physiology, 30–32
Biotech aromas, 294 orthonasal and retronasal olfaction, 30
Biscuit and cracker elaboration, 420–421 sense of smell, 29–30
Bovine milk, 469–470 oral somatosensory system
Bread and toast elaboration, 418 anatomy and physiology, 32–34
Brewing process, 357–358 somesthesis/chemesthesis, 32
esters, 358–359 Chiral capillary electrophoresis (CCE), 198
higher alcohols (fusel alcohols), 358 Chocolate production
Buffers, 195–196 post-harvest fermentation and drying
drying phase, 403
C fermentation of beans, 400–402
Cake elaboration, 418–420 primary processing phases, 399–403
Capillary electrochromatography (CEC), 198 secondary processing phases
Capillary electrophoresis (CE) alkalization process, 404
advantages and limitations, 199 conching phase, 408–409
applications grinding and refining phases, 406–408
enzymatic reactions, 203–210 roasting process, 404–406
non-enzymatic reactions, 199–203 starting material
instrumentation cocoa, geographical origins of, 399
buffers, 195–196 cocoa varieties, 397–398
capillaries, 194–195 Criollo, 398–399
detectors, 196–197 Forastero, 399
injection and separation, 196 Nacional, 398–399
system, 194 Trinitario, 398–399
pros and cons of different CE-modes, 199 volatile organic compound (VOC),
separation modes 384–385
capillary electrochromatography acids, 395–396
(CEC), 198 alcohols, 388–391
chiral capillary electrophoresis (CCE), 198 aldehydes, 391–393
micellar electrokinetic chromatography esters, 386–388
(MEKC), 197–198 furanones, 393–395
 Index 721

ketones, 391–393 E
phenols, 388–391 Elaboration
pyrazines, 385–386 biscuit and cracker, 420–421
pyrones, 393–395 bread and toast, 418
Cocoa, geographical origins of, 399 cake, 418–420
Cocoa varieties, 397–398 of processed cheese, 262–266
Coffee flavor Electroencephalography (EEG), 160
coffee production, fruit, 366–368 Electronic noses, 178–179
drying, 377–378 advantages and disadvantages of, 162
fermentation and roasting, 373–374 application, 183–186
influence of fermentation, 374–376 detection system (sensor array), 180–182
influence of roasting, 376 gas sensor response mechanism,
Maillard reaction, 377 182–183
Rhizopus oligosporus, modulation of signal processing system, 183
coffee aroma, 376 standard recognition methods, 183
thermal degradation of precursors, 377 sampling system, 179–180
flavor precursor formation, 368–373 Electronic tongues, 168–169; see also
raw bean composition, 368–373 Electronic noses
storage processes, 378 Electroosmotic flow (EOF), 194
Coffee, principal component analysis, 38 Enantiomeric odorants, 7
Comprehensive two-dimensional gas Enzymatic reactions, 203–210
chromatography (GC×GC), carbonyl compounds, 203–205
145–150 phenols, 206–210
Conching phase, 408–409 S-alk(en)yl-L-cysteine-sulfoxides, 206
Congruent taste-enhanced odor sensitivity, 36 terpenes, 206
Conventional methods, 130 Enzyme-derived aroma compounds
Cooking and aging, see Food processing aroma additives (bioaromas), 301–302
Cranial nerves (CNs), 28 during processing, 300
Criollo, 398–399 Enzyme–substrate interaction, 15
Crispness level, potato chips, 38 Esters, 62–63, 75–76, 358–359, 386–388, 477
Cross-fiber patterning theory, 17 Extraction methods
Cyclodextrins (CDs), 198 for liquid or solid samples
dynamic headspace extraction (DHE),
D 127–128
Dairy, see Milk/dairy solid-phase microextraction, 129
Dendrites, 9 static headspace extraction, 126–127
Diacetyl, 359–361 for liquid samples
Dimethyl sulfide (DMS), 359 liquid–liquid extraction (LLE), 124
Direct-extraction device (DED), 163 liquid–solid extractions (LSE), 125
Distillates solid-phase extraction (SPE), 125–126
aromas derived for solid samples
from carbohydrates, 342 conventional methods, 130
from cereals, 342–343 microwave-assisted extraction
from fruits, 343–344 (MAE), 132
aroma volatiles in, 338–340 supercritical and pressurized fluid
distillation, 346–348 extraction (SFE), 133–134
downstream modification, 348–350 ultrasound-assisted extraction (UAE),
fermentation, 344–346 130–132
processing, 340–342
qualitative mapping of, 338 F
raw materials, 340–342 Facial (CN VII), 28
undesirable aromas (taints), 350–351 Fat taste, 25
Dynamic headspace extraction (DHE), Fatty acid-derived compounds, 70–74
127–128 Fatty acids, 596
722 Index

Fermentation of beans, 400–402 physical chemistry of

Fermentation process, 422–423 during eating, 655–656
Fish aroma, 521–522 from emulsions, 653–655
environmental conditions and, 525 saliva on, 663
fat/oil amount on, 526–527 Flavor type, potato chips, 38
fish species on, 523–525 Food matrix
fresh fish aroma, 522 aroma compounds, physicochemical
pH levels on, 526 properties of
post-mortem alterations on, 522–523 vs. alcohol, 637
processing on, 533–534 vs. carbohydrates, 633–636
cooking process on, 536–537 vs. lipids, 632–633
ripening process on, 534–535 vs. Maillard melanoidins, 636–637
smoking process on, 535–536 molecular properties of, 628–629
storage conditions on, 527–528 vs. mono- and disaccharides, 633–634
lipid oxidation on, 533 vs. oligo- and polysaccharides,
microbial degradation on, 528–532 634–636
Flavor vs. polyphenols, 636
current knowledge of, 17–21 vs. proteins, 630–632
definition, 24 vs. salts, 637
Flavor chemistry methodologies, 467–468 thermodynamic and kinetic
head space-solid-phase microextraction properties, 628
(HS-SPME), 468 nonvolatile compounds
solvent-assisted flavor extraction carbohydrates and proteins, 639
(SAFE), 468 carbohydrates as fat substitutes,
thermal desorption (TD), 468–469 639–640
Flavor delivery systems lipids and proteins in dairy products,
mouth conditions 637–639
model mouths and throats, 663–665 low-sugar products, 640–641
novel emulsions for structure and texture, 641–642
gelled emulsion, 666–667 olfactory receptors, 626
multilayer emulsion, 665–666 Food processing
multiple emulsion, 667 flavor modifications, 274–277
pickering emulsion, 667–668 processed cheese
self-assembly structured emulsion, 668 caramelization reactions, 272–274
Flavor perception cooking of, 268–270
chemosensory system elaboration of, 262–266
gustatory system, 25–29 lipid degradation, 270–272
olfactory system, 29–32 Maillard reaction, 272
oral somatosensory system, 32–34 process-making for, 266–268
multisensory Forastero, 399
between chemosensory cues, 34–37 Free-solution capillary electrophoresis
stimulus medium, tactile or temperature (FSCE), 197
cues of, 37–39 Fruits and vegetables
visual or auditory cues on, 40–41 aroma compounds
Flavor release binding of, 559
emulsion ingredients and vs. lipids, 558–559
oil phase, 656–658 vs. proteins, 558
water phase, 658–660 vs. starch, 559
emulsion properties and aroma compounds from, 544
droplet size, 660–661 aroma compounds in
emulsion stability and emulsion types, aroma formation in, 548–552
662 biosynthesis of, 546–548
pH, 661–662 character impact compounds of, 544–545
rheological behavior, 661 matrix structure of, 556–557
 Index 723

sensory perception and evaluation of, I

559–560 Infrared (IR) heating, 314–315
transitioning factors on Ion mobility spectrometry-mass spectrometry
aging factors, 555–556 (IMS-MS), 167
maturity and ripeness stage, 552–553 Isothiocyanates (ITCs), 206
processing and aging, 553–555 Isotope effect, 248
functional magnetic resonance imaging
(fMRI), 10 K
Furanones, 78–79, 393–395 Ketones, 67, 391–393, 475–476, 607
Furans, 68–69, 606–607
Fusel alcohols, 358 L
Lactic fermentation, 295–297
G Lactones, 63–65, 75–76, 479
Gas chromatography L-glutamate, 25
principles and potentialities Lipid degradation, 270–272
comprehensive two-dimensional gas Lipid oxidation process, 423–424
chromatography (GC×GC), 145–150 Lipids, 558–559, 632–633
olfactometry (GC-O), 150–152 Lipophilic compounds, 70–74
one-dimensional gas chromatography Lipoxygenase pathway (in chain oxidation),
(1D-GC), 143–145 71–73
sample preparation, 142–143 Liquid chromatography (LC), 198
Generally recognized as safe (GRAS), 133 Liquid–liquid extraction (LLE), 124
Glossopharyngeal (CN IX), 28 Liquid–solid extractions (LSE), 125
Glycosyltransferases, 442 Lobry de Bruyn–Alberda van Ekenstein’s
G-protein coupled receptors (GPCRs), 9, 27 rearrangement, 273
Grape and wine volatile composition Low temperature long time (LTLT), 166
aging aromas, 451–455
fermentative aromas, 445–451 M
pre-fermentative aromas, 440–445 Magnetic resonance imaging (MRI), 160
varietal aromas, 440–445 Maillard melanoidins, 636–637
“Green leaf” volatiles (GLVs), 69 Maillard reaction, 272, 282, 424–426
Grinding and refining phases, 406–408 advanced glycation end products (AGEs),
Gustatory system 288–290
anatomy and physiology, 27–29 initial stage, 282–284
sense of taste, 25–27 intermediate stage, 285–286
late reactions, 286–287
H pool theory, 287–288
Headspace solid-phase microextraction Mass spectrometry (MS), 166
(HS-SPME), 468 Meat
Herbs, see Spices and herbs aroma analysis, 488
Hierarchical cluster analysis (HCA), 166 aroma compound analysis
High-pressure (HP) technology, 320–324 extraction techniques, 489–490
High temperature short time (HTST), 166 hierarchy of aroma compounds, 494
Hop essential oils, 354–355 odorant identification, 491
alcohols, 355–356 overall aroma perception, 495
aldehydes, 356 quantification and comparative studies,
hydrocarbon fraction, 355 491–494
4-hydroxi-4-methyl-2-pentenoic acid, sensory-directed location, 490–491
356–357 factors influencing meat aroma
3-methylbutanoic acid, 356–357 age at slaughter, 500–501
4-methyl-3-pentenoic acid, 356–357 aging, 503
2-methylpropanoic acid, 356–357 animal feeding, 501–502
Human olfaction, 97–98 animal species, 497–499
Hydro-distillation (HD), 130 bacterial spoilage, 502
724 Index

breed, 499–500 interactions between chemosensory cues

cooking, 504 crossmodal correspondence, 34
freezing and thawing, 504 intensity and pleasantness, 35–37
packaging systems, 503 lateralization/localization, 34–35
processing factors, 502–505 stimulus medium, tactile or temperature
production factors, 499–502 cues, 37–39
sex, 500 visual or auditory cues on, 40–41
storage of cooked meat, 505
relevant aroma compounds in, 495–497 N
Melanoproteins, 287 Nacional, 398–399
Metal oxide semiconductor (MOS) sensor N- and S-containing flavor molecules, 75–76
system, 165 Natural aromas, 59–61
Micellar electrokinetic chromatography Non-aqueous capillary electrophoresis
(MEKC), 197–198 (NACE), 198
Microbial-derived aroma compounds Non-enzymatic reactions, 199–203
alcoholic fermentation, 297–298 Non-enzymatic reactions, capillary
aroma additives (bioaromas), 298–299 electrophoresis (CE)
lactic fermentation, 295–297 amines, 200
stages of biotechnological production, 294 amino acids, 202–203
Microwave-assisted extraction (MAE), furanones, 200
130, 132 phenols, 200–201
Microwave heating, 310–314 pyranones, 201
Mid infrared spectroscopy (MIR), 163 pyrazines, 201–202
Milk/dairy thiamine, 203
bovine milk, 469–470 thiols, thioethers, di-, and trisulfides, 202
cheese and yogurt, 470–472 Novel non-thermal technologies
flavor chemistry methodologies, 467–468 applications, 317–318
head space-solid-phase microextraction definition, 317–318
(HS-SPME), 468 high-pressure (HP) technology, 320–324
solvent-assisted flavor extraction principles, 317–318
(SAFE), 468 pulsed electric field (PEF), 318–320
thermal desorption (TD), 468–469 ultrasound technology (US), 324–327
glycolysis, 471 Novel thermal processing techniques
lipolysis, 471 applications, 308–309
ovine and caprine milk, 470 definition, 308–309
proteolysis, 472 Ohmic heating
sensory analysis advantages and disadvantages of,
affective studies, 466 309–310
descriptive methods, 467 infrared (IR) heating, 314–315
difference testing, 466–467 microwave heating, 310–314
volatile compounds radio frequency (RF) heating, 315–317
acids, 472–473 principles, 308–309
alcohols, 476–477
aldehydes, 473–475 O
esters, 477 Oak lactones, 350
ketones, 475–476 Obitofrontal cortex (OFC), 11
lactones, 479 Odor; see also Structure–odor relationship
miscellaneous volatile compounds, descriptions, 8
479–480 properties, 6
sulfur compounds, 477–479 temporal and spatial determinants of, 18
Molecular structure of chemical compound, Odor identification, “Sniffin’ Sticks” smell
see Structure–odor relationship test, 99
Monosodium glutamate (MSG), 41 Odorous stimuli, 10
Multisensory flavor perception Odor perception threshold, 7
 Index 725

Ohmic heating Proton-transfer-reaction mass spectrometry

advantages and disadvantages of, 309–310 (PTR-MS)
infrared (IR) heating, 314–315 application of
microwave heating, 310–314 real-time VOC evolution analysis, 233
radio frequency (RF) heating, 315–317 screening VOC fingerprinting analysis,
Olfaction 230–233
human olfaction, 97–98 challenges and future perspectives, 233–234
route for, 98 direct injection mass spectrometry analysis,
Olfactometry (GC-O), 150–152 217–219
Olfactory bulb, 31 innovation
Olfactory disorders, 29, 30 compound identification, 229–230
Olfactory perception sensibility, 226–227
factors influencing throughputness, 227–229
age, 107–108 technology, 219–221
gender, 106–107 drift tube, 221
genetic factors, 108–109 switchable reagent ion, 222–226
hormonal levels, 110 PTR-MS, see Proton-transfer-reaction mass
olfactory disorders, 110–111 spectrometry
oral environment (saliva and Pulsed electric field (PEF), 318–320
microbiota), 109 Pyrazines, 69, 385–386, 607–608
Olfactory receptors, 626 Pyrones, 78–79, 393–395
Olfactory sensory pathway, 11
Olfactory system Q
anatomy and physiology, 30–32 Quadrupole mass spectrometer (QMS), 223
orthonasal and retronasal olfaction, 30
sense of smell, 29–30 R
One-dimensional gas chromatography (1D- Radio frequency (RF) heating, 315–317
GC), 143–145 Rapid analytical methods
Oral somatosensory system data integration and analysis, 163
anatomy and physiology, 32–34 in food systems
somesthesis/chemesthesis, 32 electronic noses, 163–168
Oral temperature effects, 38 electronic tongues, 168–169
Orthonasal food odor perception, 105 mass spectrometry-based systems,
Orthonasal olfaction, 30 163–168
methods to investigate, 99–102 sensors, 160–163
and retronasal olfaction, 98–99 Retronasal food odor perception, 105–106
vs retronasal olfaction Retronasal olfaction
neural level, 103–104 in food perception, 104–106
peripheral level, 103 method, 102
psychophysical level, 103 and orthonasal, 100–101
Roasting process, 404–406
P Rosa rugosa, 703
Papillae, 18
Perception, 15, 16 S
Phenols, 388–391 Salting-out effect, 634
Phenylpropenes, 67–68, 76–78 Salts, 637
Polyphenols, 636 Salty taste, 25
Pool theory, 287–288 Sensory perceptions, 16
Positron emission tomography (PET), 10 Sesquiterpenes
Potato chips, 38 anticancer potential of, 701–702
Pressurized fluid extraction (PLE), 130 anti-inflammatory effects, 705–707
Proteins, 558, 630–632 antimicrobial potential of, 703–705
Proton transfer reaction mass antioxidant potential of, 696–701
spectrometry, 161 Simultaneous distillation extraction (SDE), 130
726 Index

Smells Stir bar sorptive extraction (SBSE), 125

historical aspects, 3–4 Structure–odor relationship
perception of, 9–11 chain length, 683–685
social aspects of, 4–5 double bonds position, 688–689
“Sniffin’ Sticks” smell test, 101 functional groups, 685–688
age-related changes of, 107 QSAR studies, 689–690
odor identification, 101 stereochemistry, 689
Solid-phase extraction (SPE), 125–126 unsaturation degree, 688–689
Solid-phase microextraction (SPME), 125, Sulfur compounds, 69–70, 477–479,
129, 163 609–610
Solid-phase micro extraction-gas Supercritical and pressurized fluid extraction
chromatography mass spectrometry (SFE), 133–134
(SPME-GC-MS), 164 Supercritical fluid extraction (SFE), 130
Solvent-assisted flavor extraction (SAFE), 468 Surface acoustic wave (SAW) sensor
Solvent-free microwave extraction arrays, 164
(SFME), 132 Sweet taste, 25
Sour tastes, 25 Switchable reagent ion, PTR-MS
Soxhlet extraction, 130 data analysis, 224–226
Spices and herbs mass analyzers, 223
economic data, 570 quadrupole mass spectrometer (QMS), 223
harvest, 570 time-of-flight mass spectrometer (ToF-MS),
volatile compounds in 223–224
aril, 571–575
bark, 575–576 T
berries, 580–583 Taste
bulb, 578–580 historical aspects, 3–4
commercial presentation, 583–584 temporal and spatial determinants of, 18
flower bud, 578 Taste buds, 18, 27
fruits, 580–583 Taste quality
kernel, 571–575 current consensus on, 25
latex, 575–576 Taste quality names, 16
leaf spices, 576–578 Taste sense
nut, 571–575 current knowledge of, 17–21
processing, 583–584 historical aspects of, 15–17
rhizome, 578–580 Terpenoids, 5, 65–66, 79–80
root, 578–580 Theory on aroma perception, 668–682
seed, 571–575 Thermal desorption (TD), 468–469
stigma, 578 Thermodynamic and kinetic properties, 628
tuber, 578–580 Time-of-flight mass spectrometer (ToF-MS),
world production, 570 223–224
Stable isotope dilution assay (SIDA) Tongue temperature, 38
applications Trinitario, 398–399
H/C MDGC with SIDA-based
quantification, 249–253 U
non-MS SIDA applications, 248–249 Ultrasound-assisted extraction (UAE),
definitions, 241–242 130–132
historical development, 241–242 Ultrasound technology (US), 324–327
principle of, 242–243 Umami taste, 25
requirements, 243–247
Stable isotope dilution assays, 242 V
Starch, 559 Vagus (CN X), 28
Static headspace extraction, 126–127 Vegetables, see Fruits and vegetables
equilibrium, 126–127 Vicinal diketones (VDK), 359
pressurization, 127 Volatile organic compounds (VOCs), 123
sample transference, 127 Volatile phenols, 608–609