J Food Sci Technol (April 2021) 58(4):1349–1357

https://doi.org/10.1007/s13197-020-04646-5

ORIGINAL ARTICLE

Lentinula edodes mushroom as an ingredient to enhance

the nutritional and functional properties of cereal bars
Sara Rosicler Vieira Spim1 • Nathalia Roberta Cardoso Mendes Castanho1 •
Ana Maria Holtz Pistila1 • Angela Faustino Jozala2 • José Martins Oliveira Júnior3 •

Denise Grotto1

Revised: 29 June 2020 / Accepted: 8 July 2020 / Published online: 16 July 2020
Ó Association of Food Scientists & Technologists (India) 2020

Abstract Lentinula edodes (shiitake) is the second most 180 days of shelf life, neither at 25 °C nor at 37 °C, and
cultivated edible mushroom in the world; it has low lipid they followed the standards determined by National Health
contents, high protein and it is source of vitamins and Surveillance Agency. Sweet bars are an easy marketing
minerals. This study aimed to develop and to evaluate two alternative due to their stability, low-cost of production and
sweet and two salty food bars containing shiitake. The good acceptance, as well as flexibility to add other func-
binder elements were heated and then the dried elements tional ingredients beneficial to health, such as shiitake.
were added. The bars were shaped, and the sensorial test
was accomplished with hedonic scale of 9 points for Keywords Lentinula edodes
Functional bars
Food
analysis of texture, aroma, taste and appearance, and a development
Nutritional composition
Chemical
5-point scale for buying intention. The centesimal com- elements
position included percentages of moisture content, ashes,
lipids, proteins and carbohydrate contents. Chemical ele-
ments of shiitake were quantified by Energy Dispersive Introduction
X-ray Fluorescence. Glucans were determined using a
commercial kit. Phenolic compounds were determined with There are thousands of species of mushrooms in the world;
the Folin-Ciocalteu reagent. The shelf life was evaluated however, only about 25 varieties are cultivated with culi-
by microbiological control, up to 180 days, at temperatures nary characteristics and medicinal properties (Ng and Tan
of 25 °C and 37 °C. The sweet bar 1 (SwB1) had better 2017). Among those varieties, is the Lentinula edodes
sensory analysis and buying intention. Shiitake showed (popularly known as shiitake). Shiitake is the second most
high concentrations of calcium, iron, phosphorus, potas- cultivated edible mushroom in the world, accounting for
sium, zinc, manganese, phenolic compounds and glucans. 25% of the production. Shiitake is a complete nutritional
SwB1-bar maintained shiitake nutritional characteristics. food type because it has low lipid content, high protein
SwB1-bars did not present microorganisms for up to content, vitamins, minerals and fibers (Akesowan 2016).
Shiitake presents a variety of bioactive compounds with
great therapeutic application, such as polysaccharides,
& Denise Grotto glycoproteins and phenolic compounds, namely protocat-
[email protected] echuic, p-hydroxybenzoic, p-coumaric and cinnamic acids
1
(Roncero-Ramos et al. 2017; Rathore et al. 2017).
LAPETOX – Laboratory of Toxicological Research,
University of Sorocaba, Rodovia Raposo Tavares, Km 92.5,
According to Rincão et al. (2012), aqueous and methanolic
Sorocaba, São Paulo 18023-000, Brazil extracts, and polysaccharide isolated from shiitake, were
2 able to inhibit the replication of herpes virus and polio type
LaMInFE – Laboratory of Industrial Microbiology and
Fermentation Process, University of Sorocaba, Sorocaba, SP, 1 in vitro. At an appropriate dose, the mushroom presents
Brazil immunomodulatory action, anticancer (Pinya et al. 2019),
3
LAFINAU - Laboratory of Applied Nuclear, University of antiviral (Rincão et al. 2012), hipoglicemic and other
Sorocaba, Sorocaba, SP, Brazil health promoting properties (Spim et al. 2017).

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According to Dietary Reference Intake (DRI 2005), shiitake/kg/day (corresponding to 7 g of dried shiitake for a
dietary fiber helps in the prevention and supportive treat- 70 kg individual) has demonstrated hypocholesterolemia
ment of heart disease, with a recommendation of 20–38 g and hypoglycemic effects in pre-clinical investigation; and
of daily fiber intake. The impact of shiitake fibers on rat no toxicity (Grotto et al. 2015).
health was assessed by Anwar et al. (2019). Since intestinal Two functional Sweet bars (SwB) and two functional
microbiome can be modulated by diet, that author evalu- Salty bars (SaB) were designed and developed for com-
ated Wistar rats submitted to a control diet, a hyperc- parative purposes in the acceptability test. To develop the
holester, olemic diet and a hypercholesterolemic diet with functional bars, assuming the individual would have to eat
5% shiitake. Triglycerides, total cholesterol and low-den- two functional bars, the amount of dried shiitake used in a
sity lipoprotein (LDL) concentrations were lower and high- 25 g bar was 3.5 g.
density lipoprotein (HDL) concentrations were higher after The ingredients used in SwB were dried shiitake, oats,
the shiitake supplementation. Intestinal microbiome of the quinoa, chia, prune, flaxseed, and chestnut, peanut and
group supplemented with shiitake presented greater species sugars. The ingredients used in SaB were dried shiitake,
richness, characterized by the abundance of Clostridium oats, quinoa, rice flakes, flaxseed and sesame seeds, dried
and Bacteroides spp. tomatoes, peanut butter, parmesan cheese and condiments
b-Glucans present in shiitake can also act on lipid parsley, garlic, onion, oregano, thyme, bay leaf, pepper and
metabolism, reducing total cholesterol levels (Spim et al. salt. The general process of producing the functional bars is
2017). The polysaccharides can be considered functional shown in Fig. 1.
food. They can be used as prebiotics, especially by regu- The binder elements were heated under stirring at
lating the intestinal microbiota and the health of the indi- 115 ± 3 °C, until reaching Brix Grade 85–89. The other
vidual (Anwar et al. 2019). ingredients were added to the hot syrup and homogenized.
Based on their nutritional complexity, mushrooms can The blends were molded in a bar mold of 25 ± 1 g. After
be used for the development of functional food. Functional cooling, the functional bars were packed in a polypropy-
bars, for example, are an easy alternative to be marketed lene plastic vacuum package.
due to their stability, easy transportation, low-cost of pro-
duction and good acceptance (Hadi et al. 2018), besides the Sensory analysis and purchase intention
flexibility of other functional ingredients to be added to the
bar. The use of shiitake in biotechnology and in the pre- After defining the formulations of the functional bars
vention or treatment of chronic diseases, especially supplemented with shiitake, the bars were evaluated by 82
hypercholesterolemia and diabetes, has grown in recent untrained tasters, of both genders. The study is approved by
decades. For this reason, the present study developed,
characterized and evaluated the acceptance of the func-
tional bars supplemented with shiitake.

Materials and methods

Acquisition and processing of shiitake

Raw shiitake mushrooms were provided by the Yuri

Cogumelos company located in Sorocaba (São Paulo State,
Brazil). Raw mushrooms were shredded in a multiproces-
sor food (PhilcoÒ), weighed, and dried at 50 ± 2 °C for
48 h. After drying, the dried mushrooms were weighed
again and showed a moisture content from 87 to 92%,
meaning that 100 g of raw mushrooms produced about
10 g of dried mushrooms. Afterwards, they were stored in a
dry and airy place, protected from light, until the time of
making the functional bars.

Development of functional bars supplemented with Shiitake

The amount of dried shiitake added to the bars was selected Fig. 1 Flowchart of the production process of functional bars
based on Spim et al. (2017) study, in which 100 mg dried supplemented with Shiitake

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the Ethics Committee of the University of Sorocaba (ap- glacial acetic acid, 5 ml concentrated nitric acid, 2 g tri-
proval number 2.016.736/CAAE 65105416.8.0000). Each chloroacetic acid and 5 g diatomaceous sand by 45 min on
consumer received two copies of the Informed Consent heating. Subsequently, the samples were manually filtered,
Form (Silva et al. 2014) and they participated with com- washed with boiling water, 20 ml 70% alcohol and 20 ml
plete freedom to express discomfort or dissatisfaction. ethyl ether. Subsequently, the sample was dried, weighed
The sensory analysis was performed maintaining the and incinerated at 550 8C to ashes. The difference between
anonymity of the participants and the confidentiality of the the initial weight of the sample and the ash weight repre-
results. Samples of all four formulations were supplied sents the total fibers.
individually, with water. The test was carried out at the
Laboratory of Sensorial Evaluation of Experimental Food Concentration of chemical elements
Cooking (University of Sorocaba). The bars were analyzed
for texture, aroma, taste and appearance by a sensorial Chemical elements from shiitake in natura were assessed
acceptance test. For the quantification of the results, a he- using the technique of Energy Dispersive X-ray Fluores-
donic 9-point scale was used: I liked extremely ‘‘9’’, I liked cence (ED-XRF) (AmptekÒ). ED-XRF is a nuclear ana-
it very much ‘‘8’’, I liked it moderately ‘‘7’’, I liked it lytical technique used for the analysis of solid samples to
slightly ‘‘6’’, indifferent ‘‘5’’, slightly displeased ‘‘4’’, I determine the concentration of several elements, so that a
moderately disliked ‘‘3’’, I disliked ‘‘2’’ and I greatly dis- previous chemical treatment will not be necessary (Nasci-
liked ‘‘1’’ (Praseptiangga et al. 2019; Samuel and Peerkhan mento Filho, 1999). Essential and non-essential chemical
2020). The acceptance range was from 6 to 9 points. elements were quantified: calcium (Ca), iron (Fe), titanium
And a hedonic 5-point scale was used for the purchase (Ti), phosphorous (P), sulfur (S), chlorine (Cl), potassium
intention survey: would certainly buy ‘‘5’’, possibly buy (K), manganese (Mn), zinc (Zn), rubidium (Rb) and cad-
‘‘4’’, maybe buy ‘‘3’’, possibly would not buy ‘‘2’’ and mium (Cd).
certainly would not buy ‘‘1’’ (Silva et al. 2014).
Determination of glucans
Nutritional composition
The determination of total glucans was carried out in shi-
The centesimal composition was assessed by the percent- itake in natura and in the selected functional bar, using the
ages of moisture, ashes, lipids, proteins and carbohydrates, commercial Yeast & Mushroom-MegazymeÒ kit. For this
in shiitake in natura and in the functional bar supplemented purpose, 100 mg of samples and 1.5 mL of 37% HCl were
with shiitake selected in sensory analysis. incubated for 45 min at 30 °C. The volume was adjusted to
For the moisture content, 3 g of raw shiitake was dried 10 mL with deionized water and incubated for 2 h. Later,
at 105 °C until constant weight. To determine the ashes, 10 mL of 2 M KOH was added and the contents completed
3 g of the samples were charred and incinerated in Mufla to 100 mL with 200 mM sodium acetate buffer (pH 5.0)
(Quimis Enila) at 550 °C. For the lipid extraction, 100 mL and centrifuged at 1500 rpm for 10 min. The supernatant
of ethyl ether solvent was used for each sample for 6 h in (0.1 mL) was mixed to 0.1 ml of the exo-b-1,3-glucanase
the Soxhlet extractor (Samuel and Peerkhan 2020). and b-glycosidase solution and incubated at 40 °C for
Proteins were quantified using the micro Kjeldahl 60 min. Then, 1.5 mL of glucose oxidase/peroxidase was
method (Zhou et al. 2015), with some modifications. The added and incubated for an additional 20 min. at 40 °C.
method consisted of three phases: digestion, distillation and The absorbance was read at 510 nm.
titration. The digestion was based on the oxidation of For a-glucans, 2 mL of KOH was added to 100 mg of
70 mg of the sample at 380 °C with 5 mL concentrated the samples. After 20 min of stirring, 0.2 mL of amy-
sulfuric acid, 100 mg potassium sulfate and 100 mg copper loglucosidase and invertase were added, incubated at 40 °C
sulfate for six hours at 380 °C. In the distiller, nitrogen for 30 min. The tubes were centrifuged at 1500 rpm for
from proteins was reduced in ammonium sulfate. Upon 10 min. To a 0.1 mL of the supernatant, 0.1 mL of sodium
contact with 5 mL 5% boric acid, the ammonium sulfate acetate buffer and 3.0 mL of enzyme reagent were added,
was transformed in ammonium borate, titrated with 37% and incubated for 20 min at 40 °C. The absorbance was
hydrochloric acid. The centesimal composition of the measured at 510 nm (Lambda 35, PerkinElmer, Waltham,
mushroom was determined by adding the percentages of MA, USA). For the quantification of b-glucan, a-glucan
moisture, ashes, lipids, proteins and subtracting of 100%, was subtracted from total glucan. All analyses were per-
the difference was the percentage of carbohydrates. formed in triplicate.
The fibers were quantified using Mccleary et al. (2010)
and filtration manual techniques. For this, 2 g of dry lipid-
free samples were digested with 50 mL concentrated

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Determination of total phenolic compounds Results and discussion

Phenolic compounds were determined by weighing 2 g of Development of the functional bars

the shiitake in natura or the selected functional bar, in
triplicate, in volumetric flasks with methanol 80%. The Pilot tests were performed with different concentrations of
flasks were shaken every ten minutes for one hour, after binder ingredients. Six sweet functional bars supplemented
being filtered and centrifuged at 3000 rpm for 30 min to with shiitake were designed with sugar ranging from 40 to
obtain the supernatant. The supernatant (2 mL) was 52%. Two of the tastiest formulations moved forward to
transferred to a 25-mL flask; 1 mL of Folin-Ciocalteu the sensory and purchase tests. A sugar concentration of
reagent was added, and the volume was quenched with 42% was defined for the sweet bars, presenting satisfactory
20% sodium carbonate. The reading was carried out in a consistency, appearance, flavor and aroma. All the ingre-
spectrophotometer at 760 nm (Lapornik et al. 2005). A dients used in the functional SwB are described in Table 1.
calibration curve of gallic acid was done at concentrations The differences between the bars were the crystallized
12.5; 25.0; 37.5; 50.0; 62.5; 75 and 100 lL/mL. The results ginger in SwB2 and the peanut in SwB1, modifying the
were expressed as mg gallic acid equivalent (EAG)/g flavor.
sample. For the functional salty bars, pilot tests were performed
with different concentrations of wheat flour, soy lecithin,
Storage and shelf life condiments and xanthan gum. Four salty bars were devel-
oped and two of the tastiest bars (presented in Table 1)
Samples of the selected functional bar supplemented with moved forward to the sensory and purchase tests. The
shiitake were ground and diluted in saline (0.9%), followed differences between the salty bars were the peanut in the
by serial 1:10 dilutions. The diluted solutions were inoc- SaB1 and the parmesan cheese in the SaB2.
ulated in petri dishes, with the culture medium. The culture
media used were Plate Count Agar (PCA), indicated for Sensorial test and purchase intention survey
total count of microorganisms, and Potato Dextrose Agar
(PDA), used for counting molds and yeasts. The selective The sensorial analysis evaluates the flavor and the prefer-
culture media used were MacConKey Agar (used to Gram- ences of the consumers, as well as the acceptance of a new
negative like Escherichia coli), Mannitol-Egg Yolk Poly- product in the market. Through sensory analysis, it was
myxin Agar (recommended for the isolation of Bacillus possible to evaluate the bars supplemented with shiitake for
cereus), Salt Mannitol Agar (used to identify Staphylo- texture, flavor, aroma and appearance (Fig. 2a).
coccus aureus) and Bismuth Sulfite Agar (for isolation of No differences in appearance were observed comparing
Salmonella sp). all functional bars. There were no differences between the
The inoculation of the samples was performed under SwB1 and 2. There were no differences between the SaB1
aseptic conditions. After the incubation time at 37 °C for and 2. However, a significant decrease in the acceptance
bacteria and 25 °C for molds and yeasts, the colonies were rate of SaB was observed compared to SwB, which means
counted manually and expressed in colony forming units. the sweet flavor had greater acceptability than the salty one
The shelf life of the functional bar supplemented with did.
shiitake was accompanied by the growth kinetics of bac- Considering the 9-point hedonic scale as 100%, the
teria, molds and yeasts within 1, 7, 15, 30, 60, 90 and acceptability in terms of flavor were 78, 67, 44 and 44% for
180 days. The results were reported by microbial evalua- SwB1, SwB2, SaB1 and SaB2, respectively. Flavor
tion, meeting the specifications of the Codex Alimentarius acceptability was 11% higher for SwB1 when compared to
(FAO 1980) and Brazilian Food Microbiological Standards SwB2, and 34% higher for SwB1 when compared to both
(Brasil, 2001). salty bars. Aroma is also an important parameter for
pleasantness, and the percentages of acceptability were 78,
Statistical analysis 67, 56 and 56% for SwB1, SwB2, SaB1 and SaB2,
respectively.
Data were expressed as mean ± standard deviation. Data The purchase intention survey is presented in Fig. 2b.
were submitted to analysis of variance (ANOVA) followed Both salty bars did not have good acceptability among the
by the Tukey test (p \ 0.05) and the graphs were produced volunteers, and the intent-to-buy scales were predomi-
in GraphPad PrismÒ 6. nantly ‘‘certainly not to buy’’ and ‘‘possibly would not
buy’’. On the other hand, SwB1 and 2 presented the pref-
erence of the volunteers. SwB1 presented 70% of the
preference with ‘‘possibly would buy’’ and ‘‘certainly

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Table 1 Ingredients used in the

Ingredients SwB1 SwB2 Ingredients SaB1 SaB2
25 g functional Sweet Bars
SwB SaB
(SwB) and Salty Bars (SaB) % g % g % g % g

Dried Shiitake 14.0 3.50 14.0 3.50 Dried Shiitake 14.0 3.50 14.0 3.50
Flakes oat 12.0 3,00 12.0 3.00 Flakes oat 12.0 3.00 12.0 3.00
Brown sugar 8.3 2.10 8.3 2,10 Brazil nut 12.0 3.00 12.0 3.00
White sugar 30.0 7.50 30.0 7.50 Quinoa grains 10.0 2.50 10.0 2.50
Dry plum 9.0 2.25 9.0 2.25 Sesame seed 10.0 2.50 10.0 2.50
Brazilian nut 7.0 1.75 7.0 1.75 Flaxseed 9.0 2.25 8.0 2.00
Flaxseed 3.2 0.80 3.2 0.80 Dried tomato 8.0 2.00 8.0 2.00
Glucose 1.6 0.40 1.6 0.40 Peanut 14.0 3.50 – –
Soy lecithin 0.4 0.10 0.4 0.10 Peanut butter 3.0 0.75 3.0 0.75
Quinoa grains 3.5 0.86 3.5 0.86 Parmesan cheese – – 15.0 3.75
Crystallized ginger – – 8.0 2.00 White flour 3.0 0.75 3.0 0.75
Peanut 8.0 2.00 – – Whole wheat flour 3.0 0.75 3.0 0.75
Chia seed 2.0 0.50 2.0 0.50 Xanthan gum 1.0 0.25 1.0 0.25
Food glycerin 0.5 0.12 0.5 0.12 Soy lecithin 0.6 0.15 0.6 0.15
Coconut oil 0.5 0.12 0.5 0.12 Spice 0.4 0.10 0.4 0.10
Total 100 25.0 100 25.0 Total 100 25.0 100 25.0

Fig. 2 Sensory analysis and purchase intention of functional Sweet mean ± standard deviation of hedonic 9-points scale. *Different from
bars 1 (SwB1) and 2 (SwB2), and Salty bars 1 (SaB1) and 2 (SaB2) in SaB1 (p \ 0.05); #Different from SaB2 (p \ 0.05)
texture, flavor, aroma and appearance. Results are presented as

would buy’’ whereas SwB2 presented 56% of the Centesimal composition, glucans and total phenolic
preference. compounds
Therefore, the functional SwB1, with peanut flavor, was
chosen as the tastiest bar, and the other analyses (centesi- The results of the centesimal composition are shown in
mal composition, microbiological, glucans) were done in Table 2.
that bar. SwB1 had the percentage of moisture within the limit
In another study, cookies containing shiitake were pro- established by the national commission on food stan-
duced and analyzed sensorially (Toan and Thu 2018). dards—cereals and products derived from cereals cannot
Wheat flour was replaced with shiitake flour, at concen- have more than 15% moisture (Brasil 1978). SwB1 also
trations of 5%, 10% and 15%. The authors achieved a had lipids possibly due to the Brazilian nut and peanut.
significant increase in the concentration of fibers and pro- Comparatively, gluten-free cereal bars of amaranth and
teins in the cookie with shiitake flour compared to wheat flaxseed were developed, and the authors found 9.12%
flour. The cookie with 5% shiitake was the one that pleased moisture, 7.12% proteins and 7.87% lipids, differences
the participants the most. presented according to the ingredients (Pagamunici et al.
2014).

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Table 2 Centesimal composition of shiitake in natura and functional effective than the high-purity b-glucans in lowering gly-
SwB1 supplemented with shiitake cemic response. Therefore, the results depend on the
Shiitake SwB1 molecular weight, level of purity and preparation methods
of b-glucans.
Moisture (%) 90.3 ± 0.3 14.50 ± 0.23
High total phenolic compounds were found (176 mg
Ashes (%) 0.62 ± 0.01 1.45 ± 0.12 GAE/g dry basis) in shiitake and (82 mg GAE/g dry basis)
Lipids (%) 0.40 ± 0.16 14.12 ± 4.04 in the functional sweet bar, bioactive substances important
Proteins (%) 2.27 ± 0.23 11.29 ± 0.22 for cell health. Secondary plant metabolites such as phe-
Carbohydrates (%) 5.76 ± 0.41 61.99 ± 4.08 nolic acids, simple phenols, flavonoids, tannins, lignins,
Fibers (%) 9.99 ± 2.00 5.54 ± 1.50 tocopherols, are synthesized during their development and
a-glucans (%) 0.30 ± 0.01 16.46 ± 0.01 many of them have beneficial effects, such as the antioxi-
b-glucans (%) 26.5 ± 0.01 15.96 ± 0.01 dant effect (Ng and Tan 2017). Comparatively, total phe-
Total Phenolic Compounds * 176 ± 29 82 ± 5 nols were evaluated in 17 species of Mexican mushrooms,
*Data expressed as gallic acid equivalents (mg GAE/g) dry sample mostly edible. The authors found from 30.31 to 307.01 mg
GAE/100 g in mushroom samples on dry basis (Yahia et al.
2017).

Carbohydrate concentration in SwB1 increased sub- Concentration of chemical elements

stantially compared to shiitake in natura due to its com-
position in sugars, fibers and glucans. In the study of Silva The concentrations of the chemical elements—essential
et al. (2014), cereal bars were developed with 12.5% of and non-essential—are presented in Fig. 3. Shiitake pre-
pumpkin seed flour. The bars presented 11.45% humidity, sented high concentrations of K and P (24.5 and 7.9 mg/g,
63.34% carbohydrates, 12% protein, 5.51% lipids and respectively), considered macro-elements. The other ele-
6.17% insoluble fibers, differing from this study only in the ments were found in the order of lg/g, that is,
lipid concentration. microelements.
Protein concentration in shiitake was low in comparison Kolundžić et al. (2018) analyzed macro and microele-
to the study of Khaskheli et al. (2018), which found 12.1% ments from three species of mushrooms, including shiitake.
of proteins in shiitake. The differences can be influenced The most abundant elements were Ca, K and P, followed
by the substrate where the mushroom is cultivated, climate, by lithium (Li), selenium (Se) and Zn, similar to our
temperature and other care in the mushroom’s cultivation findings. On the other hand, some toxic elements such as
(Siwulski et al. 2019). Bach et al. (2018) analyzed the lead (Pb), arsenic (As) and Cd were detected at higher
chemical composition of the shiitake produced by two levels than those allowed.
different cultivation methods, in axenic substrates (pre- The variability in the chemical composition of the
pared with eucalyptus sawdust, wheat bran, maize germ substrates may interfere both in the nutritional yield and in
and limestone) and (wood logs sawtooth oak) and showed the levels of toxic elements. The findings of Siwulski et al.
that the physical–chemical composition influences the (2019) corroborated these results, highlighting the care for
moisture, protein, lipids, minerals and fibers. the substrates used in the mushrooms cultivation, ensuring
The percentage of a-glucan in shiitake in natura was consumers’ safety.
lower than in SwB1. The percentage was possibly higher in
the bar due to the presence of oats. The b-glucan presented
in shiitake in natura was not maintained in the SwB1 even
though 16% is a high concentration. In relation to shiitake
in natura, the b—glucan concentration was similar to that
found in a previous study from our group (34.5%) (Spim
et al., 2017) and from Kolundžić et al. (2018) (33,98%).
Bak et al. (2014) evaluated the stems and mycelia of ten
shiitake mushroom cultivars for b-glucan and found a
variation ranging from 20.06 to 44.21% in the mycelia and
from 29.74 to 56.47% in stems.
Thondre and Henry (2011) investigated the effect of
high-purity b-glucans in vitro digestion and in glycemic
response in individuals that consumed chapattis, a typical Fig. 3 Essential and non-essential chemical elements of shiitake in
bread in India. The low-purity b-glucans proved to be more natura by energy dispersive X-ray fluorescence

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In a shiitake from China, K and P concentrations were two bars that, after 30 days, showed growth of molds and
similar to this study (27.28 and 8.91 mg/g respectively) yeasts. The presence of molds and yeasts is attributed to the
whereas Ca (0.174 lg/g), iron (Fe) (36.29 lg/g), Zn high humidity content found in the samples (Ferreira et al.
(7.21 lg/g) and manganese (Mn) (1.74 lg/g) presented 2015).
significantly lower values (Li et al. 2018). According to Nerı́n et al. (2016), food contamination
can occur along the production process. Some contami-
Shelf life by microbiological control nants may already be present in the raw materials, but
others may be incorporated during handling, packaging,
SwB1 showed an initial total count of microorganisms of transportation, storage, preheating, disinfection, cleaning
9 9 10 CFU/g. After 180 days, the total count of and sterilization. These facts highlight the importance of
microorganisms was 7 9 10 CFU/g. These results were conducting chemical and microbiological analyses during
lower than the maximum limit allowed by National Health food processing, ensuring safety for consumers (Sharma
Surveillance Agency (ANVISA), evidencing the quality of et al. 2013).
raw materials and the sanitary control in the preparation of Microbiological criteria and standards for food should
the bars. Microbial food safety is an essential component of be observed, applying, whenever possible, the Hazard
food quality to the consumer (Nerı́n et al. 2016). Analysis System and Critical Control Points (HACCP) and
When samples of SwB1 were inoculated in a selective the microbiological quality of food products (Brasil 2001).
culture medium, few units (\ 102 CFU/g) of S. aureus and Some microorganisms are highly adherent to the grain and
B. cereus were found (Table 3). There was no growth for cannot be removed by simply washing in liquid medium.
E. coli and Salmonella sp (Table 3). Our results were However, they can be reduced by removing the surface of
within the limits described in European Union Commission the grain using an abrasive grinder. This microbial load
Recommendation (CE N8 1441/2007) and ANVISA. reduction is even more important in the case of fungi, as
In a comparative study, breakfast cereals with flakes of they can potentially produce mycotoxins (Laca et al. 2006).
barley, oats, rye and wheat were evaluated for bacterial
count, packed in three different materials. Total bacterial Conclusion
count and shelf life were evaluated for up to 6 months. The
initial count was 5.5 9 10 CFU/g for all samples, and after Both functional sweet bars pleased the volunteers more
1 month of storage, the samples remained stable (Kince than the salty ones did, with approval in the sensory
et al. 2017). analysis and purchase intention. Sweet bar 1 obtained the
Similarly, a bar rich in isoflavones and soy protein was highest score in the preference of volunteers, with peanut
developed to control dyslipidemia. The shelf life of the flavor.
product was assessed for 6 months, and there were no SwB1 maintained the shiitake concentrations of pro-
significant changes in the microbiological parameters in teins, fibers, glucans and phenolic compounds. The devel-
coliforms, S. aureus, B. cereus and Salmonella sp., pre- opment of functional food has brought health benefits and
senting less than 10 colonies of microorganisms in each the SwB1 is an easy-to-market alternative due to their
sample (Lobato et al. 2011). Cookies and cereal bars were stability, easy transportation, low-cost of production and
produced with fruit and vegetable residues, and microbio- good acceptance, as well as the flexibility to add
logical stability was tested, as well as shelf life for 0, 30 nutraceuticals such as shiitake.
and 90 days. The authors observed that the samples met the
standard established by the Brazilian rules, except for the

Table 3 Analysis of the shelf

Counting of microorganisms in Sweet bar 1 supplemented with shiitake
life of the functional SwB1
supplemented with shiitake, in Samples E. coli (CFU/g) S. aureus (CFU/g) B. cereus (CFU/g) Salmonella sp (CFU/g)
selective culture medias
24 °C 37 °C 24 °C 37 °C 24 °C 37 °C 24 °C 37 °C

7 days 0 0 2 9 10 0 0 0 0 0
180 days 0 0 2 9 10 0 1 9 10 1 9 10 0 0
ANVISA 5 9 10 103 5 9 102 0
ANVISA (Brazilian National Health Surveillance Agency)

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Acknowledgments The authors gratefully thank Valéria de Campos Kolundžić M, Radović J, Tačić A, Nikolić V, Kundaković T (2018)
Orsi for technical support, Yuri Cogumelos Company, Cefar Diag- Elemental composition and nutritional value of three edible
nostic Laboratory Ltda and J.P. Pereira. mushrooms from serbia. Zastita Materijala 59:45–50
Laca A, Mousia Z, Dıaz M, Webb C, Pandiella SS (2006) Distribution
Compliance with ethical standards of microbial contamination within cereal grains. J Food Eng
72:332–338
Conflict of interest The authors declare they have no conflict of Lapornik B, Prosek M, Wondra AG (2005) Comparison of extracts
interest. prepared from plant by-products using different solvents and
extraction. J Food Eng 71:214–222
Li S, Wang A, Liu L, Tian G, Wei S, Xu F (2018) Evaluation of
nutritional values of shiitake mushroom (Lentinus edodes) stipes.
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production of flour and the made biscuits from Shitake
jurisdictional claims in published maps and institutional affiliations.
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