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Green synthesis of silver nanoparticles using green

tea leaf extract, characterization and evaluation of
Open Access Article. Published on 18 January 2022. Downloaded on 4/3/2025 12:15:56 AM.

Cite this: Nanoscale Adv., 2022, 4, 911

antimicrobial activity
Hiba Abbas Widatalla, *a Layla Fathi Yassin,b Ayat Ahmed Alrasheid,c
Shimaa Abdel Rahman Ahmed,b Marvit Osman Widdatallah,a Sahar Hussein Eltiliba
and Alaa Abdulmoneim Mohamedd

The use of nanoparticles in medicine, nanomedicine, is very important to diagnose and treat diseases;
among the various metallic nanoparticles, silver nanoparticles (AgNPs) are very popular due to their
physical, chemical, and biological properties, encompassing a range of activities such as antiviral,
antifungal, anti-inflammatory, and anticancer activities. In this study, the synthesis of AgNPs was
conducted by the use of a nontoxic, ecofriendly method. Green tea (GT) leaf extract was used as
a reducing agent to convert silver ions into free AgNPs. The UV-vis spectrum showed a peak at 410 nm,
confirming the presence of AgNPs. A Fourier-transform infrared (FTIR) analysis of the GT extract and GT
AgNPs display spectra that is identical to those of polyphenols, polysaccharides, and proteins. All the
vibrational peaks in the GT extract spectrum were shifted in the AgNP spectrum, becoming narrower
after the encapsulation of nanoparticles. The scanning electron microscopy (SEM) images confirm the
presence of AgNPs with different sizes, ranging from 15 to 33 nm. Furthermore, the antibacterial activity
Received 29th June 2021
Accepted 17th December 2021
of the synthesized AgNPs in three different concentrations (10, 20, and 50 mg ml 1) showed appreciable
inhibition of bacterial growth against Staphylococcus aureus and Klebsiella sp. From the above findings,
DOI: 10.1039/d1na00509j
we can recommend the use of AgNPs from GT leaf extracts as an antimicrobial agent to treat chronic
rsc.li/nanoscale-advances infections.

important toward the diagnosis and treatment of diseases; in

Introduction addition, it is important to determine the efficacy and safety of
Nanotechnology is a rapidly developing branch of science that nanoparticles within biological systems.4 Among the most
incorporates many disciplines such as biology, chemistry, important types of nanoparticles, metallic nanoparticles have
physics, food, medicine, electronics, aerospace, and medicine a wide range of applications.5
and it examines the design, manufacture, assembly, and char- The therapeutic efficacy of metallic nanoparticles is due to
acterization of materials that are smaller than 1–100 nm in their optical property demonstrated by localized surface plas-
size.1 Nanoparticles are being applied in many elds of science mon resonance.6 Among the various metal nanoparticles, silver
such as medical, cosmetic, biomedical, drug/gene delivery, nanoparticles (AgNPs) are very popular and have many uses in
environmental, energy science, photoelectrochemical, and industry and biomedicine due to their physical and chemical
optical applications.2 The wide range of uses of nanoparticles is properties in addition to their biological properties, which
due to their unique properties such as high surface-to-volume encompass a range of activities such as antiviral, antifungal,
ratio, high surface energy, and unique mechanical, thermal, anti-inammatory, and anticancer activities.7 Surface chem-
electrical, magnetic, and optical behaviors.3 The use of nano- istry, size, size distribution, shape, particle morphology,
particles in medicine, also referred to as nanomedicine, is very particle composition, coating agglomeration, and dissolution
rate are factors that affect the effectiveness of AgNPs inside
biological systems.8
a
Department of Pharmacology, Faculty of Pharmacy, University of Medical Sciences Silver is a nontoxic, safe inorganic antibacterial agent that is
and Technology, Khartoum, Sudan. E-mail: [email protected] capable of killing about 650 types of disease-causing microor-
b
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Medical ganisms.5,9 However, the emergence of bacterial strains resis-
Sciences and Technology, Khartoum, Sudan
c
tant to metal ions has led to the search for new antibacterials.10
Department of Pharmacognosy, Faculty of Pharmacy, University of Medical Sciences
In 2015, the World Health Organization (WHO) published
and Technology, Khartoum, Sudan
d
Department of Clinical Pharmacy, Faculty of Pharmacy, University of Medical
a report on a global antimicrobial resistance surveillance
Sciences and Technology, Khartoum, Sudan system concluding that it is very common for bacteria such as

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Escherichia coli, Staphylococcus aureus, Shigella sp., Klebsiella was imaged by scanning electron microscopy (SEM). The anti-
pneumoniae, and Salmonella sp. to show resistance toward one bacterial activity of the synthesized GT AgNPs was evaluated.
or several antibiotics as they are commonly present in
communities and hospitals. The development of resistant
Materials and methods
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machineries by bacteria has led to an increase in the rates of

antibiotic resistance. All this motivates the search for a new Materials
alternative to conventional antimicrobials and antibiotics.10–12 GT sample. Dried, ground GT leaves (1 g) were obtained from
Different methods have been developed for the synthesis of a Sudanese tea company in the form of teabags (Coea). The
nanoparticles, and each method has its own advantages and teabags were purchased from a store in Khartoum, Sudan.
disadvantages.13 There are physical and chemical methods for Reagents and analytical solutions. Deionized water, silver
Open Access Article. Published on 18 January 2022. Downloaded on 4/3/2025 12:15:56 AM.

the synthesis of metallic nanoparticles. Although these nitrate, sodium hydroxide, dimethyl sulfoxide (DMSO), acetone,
methods produce particles of the desired characteristics, they Mueller–Hinton agar.
usually involve high cost, require effort, and are potentially
dangerous and toxic to the environment and living organisms.
To overcome the shortcomings of these physical and chemical Methods
methods, the biological approach has been used to synthesize GT extraction. About 30 ml water were added to 1 g dried
nanoparticles. Several biological resources such as plant ground GT leaves. The mixture was heated for 30 min at 50  C
extracts, microorganisms, milk, oilcake, and panchagavya have under magnetic stirring, then cooled and ltered.
been used as alternatives to synthesize metal nanoparticles.14,15 Synthesis of GT AgNPs. Different concentrations of aqueous
Among the various green sources explored, microalgae provide (GT) extracts were used to prepare the AgNPs: the concentration
worthwhile benets with respect to their ease of growth and that afforded the best stability and yield was selected. For the
their ability to survive in extreme conditions (namely, pH and preparation, 10 ml silver nitrate solution (0.01 M) were added to
temperature).16 Numerous advantages of the biological 20% aqueous GT extract solution in a dropwise manner, and
synthesis of AgNPs have been acknowledged in recent years. 2.5 ml NaOH (0.1 M) were added to the mixture. The resulting
Various plant materials and microorganisms have been identi- solution was then heated for 30 min at 50  C in order to increase
ed as potential candidates for AgNPs synthesis. It has been the yield of AgNPs. Finally, the GT AgNPs were separated by
suggested that the presence of specic proteins in plants and centrifugation (6000 rpm) and washed with 50% acetone and
microorganisms can cause the reduction of Ag+ ions. The allowed to dry at room temperature.
probable role of NADH-dependent nitrate reductase in the Characterization of GT AgNPs
reduction of Ag+ ions has been suggested.17 UV spectroscopy. The UV spectra were used to ascertain the
Nevertheless, the synthesis of AgNPs using plant extracts is formation of AgNPs. The AgNP solution was scanned over the
potentially advantageous over microorganisms because of its range of 200–400 nm by using a Shimadzu UV spectrophotom-
simple scale up.18 The use of microorganisms presents several eter. The dilution of GT AgNP solution was required; a ratio of
challenges such as high cost, mass cultivation of microorgan- 1 : 9 was used. The same dilution ratio was used for the GT
isms, maintenance of an aseptic environment, purication, and extract, which acted as the reference solution.
quantum of production.17 On the other hand, the ‘green’ Fourier-transform infrared (FTIR). The IR spectra of GT leaf
method that involves the use of extracts obtained from plants is aqueous extract and the centrifuged GT AgNPs sample were
cost-effective, efficient and environment-friendly.19 Several used to identify the possible chemical constituents involved in
studies have been performed in recent years to synthesize the synthesis and capping of GT AgNPs. The samples were
nanoparticles using various plant extracts such as Alternanthera analyzed by IRTracer-100 (Shimadzu). A pure KBr pellet was
dentata,20 Carica papaya,21 Coffea arabica,22 Azadirachta indica,23 used for the background. The spectra were recorded from 400 to
and Olea europaea.24 Furthermore, plant extracts derived from 4000 cm 1.
various species are regarded as a benecial system for nano- Scanning electron microscopy (SEM). SEM was used to char-
particle synthesis due to their tremendous capability to produce acterize the morphology and particle size of AgNPs. A thin lm
a variety of phytochemicals with a profound reducing poten- of oven-dried GT AgNP sample was prepared and used over
tial.25 Plant metabolites like sugars, terpenoids, polyphenols, a carbon-coated copper grid via a TESCAN MIRA-3 instrument
alkaloids, phenolic acids, and proteins play an important role in operated at an accelerated voltage of 20 kV.
reducing metal ions into nanoparticles. These phytochemicals Antibacterial activity of GT AgNPs. The inhibition of bacterial
are also responsible for stabilizing the synthesized growth of GTAgNPs was tested by using a disc-diffusion assay,
nanoparticles.26 as described by Sharma et al.28 with certain modications.
In this study, the synthesis of AgNPs was conducted by the The antimicrobial effect of GT AgNPs was tested against
use of green tea (GT) leaf extracts. GT leaves are known for their Gram-positive bacteria (Staphylococcus aureus) and Gram-
high content of various phytochemical constituents that are negative bacteria (Klebsiella spp). About 20 ml Mueller–
involved in the reduction of metallic ions into nanoparticles.27 Hinton agar were poured in sterile Petri dishes to prepare
The synthesized AgNPs were characterized by using UV-vis the base plates. About 0.1 ml standard bacterial stock
spectrometry and FTIR. The morphology of the nanoparticles suspensions (108–109 cfu ml 1) was streaked onto the base
plates using a sterile cotton swab. A stock solution of 20 mg

912 | Nanoscale Adv., 2022, 4, 911–915 © 2022 The Author(s). Published by the Royal Society of Chemistry
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ml 1 GT AgNPs solution was prepared with DMSO; three

dilutions of 5, 10, and 20 mg ml 1 were used. Further, 6 mm-
diameter sterilized lter paper discs were immersed in the GT
AgNP solutions and then placed on the surface of the test
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bacteria plates. Aer the plates were incubated for 24 h, the

diameters of the inhibition zones were measured.

Results and discussion

UV spectroscopy
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The UV-vis spectrometer is a spectral device that is widely used

to conrm the formation of AgNPs in a colloidal solution via the
surface plasmon resonance phenomena of metallic nano-
particles. This optical property is sensitive to size, shape,
concentration, and agglomeration state of the nanoparticles
produced.29 Fig. 1 shows the observable change of yellowish GT
into a brown color, which indicates the formation of GT AgNPs.
The UV scan of the GT AgNP solution showed a distinct Fig. 3 IR spectra of GT extract and GT AgNPs.
Gaussian-shaped peak at 410 nm. According to Grand et al.,30
the band corresponds to absorption by AgNPs is in the region of
400–450 nm, which is due to the excitation of surface plasmon 2850.7, 1653.9, 1384, 1107, 1053, and 997.2 cm 1. The broad
vibration. Fig. 2 shows the UV spectrum of GT AgNPs. band at 3566.3 cm 1 is due to the O–H stretching of alcohol in
polyphenols and N–H stretching in amines. The bands repre-
FTIR analysis senting C–H stretching in alkanes and O–H stretching in
FTIR spectra of dried GT AgNPs and GT extracts (Fig. 3) were carboxylic acids appear at 2918 and 2850.7 cm 1, respectively.
interpreted through the correlation of the absorption bands to The strong bands at 1653.9 and 1384 cm 1 correspond to the
the corresponding compounds for studying the phytochemical C]C aromatic group and primary amide group of proteins
constituents involved in the reduction and capping of AgNPs. (–CO–NH2), respectively; the 1107 and 1053 cm 1 peaks repre-
The GT extract showed distinguishing peaks at 3566.3, 2918, sent C–O–C stretching, and the strong band at 997.2 cm 1
represents C]C bending. A phytochemical analysis of GT
aqueous extract indicates the presence of polyphenols such as
gallic acid (GA), gallocatechin (GC), catechin (CE), epi-
gallocatechin, protein, avonoid, saponin, and glycosides.31
The most abundant constituent in GT is polyphenols,
accounting for 24–36% in dry weight in the form of catechins.32
The obtained IR results are identical to those of polyphenols,
polysaccharides, and proteins. The FTIR spectra of GT AgNPs
reveal clear peaks at 3689.8, 2924.09, 1634, 1238.9, 1701, 1043,
and 761.8 cm 1. The peaks at 3689.8, 2924.09, and 1634 cm 1
correspond to (O–H, N–H), C–H stretching, and N–H bending,
respectively. The other peaks at 1238.9, 1043, and 1701 cm 1
Fig. 1 Preliminary conformation of GT AgNP synthesis. GT extract (A) can be attributed to aromatic amine stretching, C–O stretching,
GT AgNPs (B). and C]O ketone stretching, respectively. The nal peak at
761.8 cm 1 corresponds to C–H bending. Changes in the
chemical constituents of GT and GT AgNPs can be identied by
comparing their FTIR spectra. All the vibrational peaks in the
GT spectrum were shied in the GT AgNP spectrum, becoming
narrower aer encapsulation of the nanoparticles. The struc-
ture–activity relationship of avonoids plays a critical role in GT
capacity to reduce several species; the abundance of the
hydroxyl (HO) group makes epigallocatechin gallate a powerful
antioxidant and a strong reducing agent for AgNP synthesis.33

SEM analysis
Fig. 4 shows the SEM images of GT AgNPs with different sizes
Fig. 2 UV spectrum of AgNP synthesized from GT extract. below 50 nm (ranging from 15 to 33 nm), which are

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Fig. 6 STD against Klebsiella spp. (A), GT AgNPs against Klebsiella spp.
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(B).

Fig. 4 SEM image of the synthesized nanoparticles.

Table 1 Antimicrobial activity of GT AgNPs

predominantly dispersed in the form of aggregates. The nano- Zone of Inhibition in mm

particles were not in direct contact with each other within the Klebsiella spp. S. aureus
formed aggregates, which can be explained by the stabilizing
action of capping agents present in the extract. These phyto- GT AgNPs concentration (mg ml 1)
chemicals are known to play an active role in reducing and 20 10 11
10 8 8
stabilizing metal nanoparticles.34
5 9 10

Antimicrobial agent (STD)

Antibacterial activity Ciprooxacin (5 mcg) 26 32
The antimicrobial activity of GT AgNPs has been reported in
several studies.35–38 In this study, different concentrations (5,
10, and 20 mg ml 1) of GT AgNPs were tested against Staphy-
were insignicant when compared to standard drug. Results
lococcus aureus and Klebsiella sp. As shown in Fig. 5 and 6 GT
shown in Table 1.
AgNPs possessed moderate activity against the tested bacteria
with inhibition zone ranging between 8–11 mm. A number of
theories for antimicrobial actions of silver nanoparticles Conclusion
solution have been proposed. Some of the proposed mecha-
nisms include changing the permeability of the cell The synthesized silver nanoparticles were characterized using
membrane, which enables the interaction with DNA, proteins, UV, FTIR and SEM. The morphology of GT AgNPs appear to be
and other phosphorus- and sulfur-containing cell constituents irregular and in the form of aggregates. GT AgNPs revealed
that are harmful to bacteria. Another mechanism is generating antimicrobial activity against selected bacterial species. The
free radicals responsible for the damage of membrane and difference in the FTIR spectra between the GT extract and GT
dissipation of the proton motive force, resulting in the AgNPs was used to indicate the functional groups involved in
disruption of membrane potential;39 however, the exact the reduction of Ag+ ions into nanoparticles.
mechanism has not been fully understood. The three
concentrations showed similar effectiveness in combating the
tested species with the highest concentration having the Author contributions
highest activity. The inhibition zones obtained by GT AgNPs
Hiba Abbas Widatallah was responsible for conceptualization,
investigation, methodology, project administration and writing
the original dra.
Layla Fathi Yassin was responsible for conceptualization,
investigation, methodology, project administration and writing
the original dra.
Ayat Ahmed Alrasheid was responsible for conceptualiza-
tion, investigation and supervision.
Shimaa Abdel Rahman Ahmed was responsible for concep-
tualization and investigation.
Marvit Osman Widdatallah was responsible for conceptual-
ization and investigation.
Sahar Hussein Eltilib was responsible for conceptualization
Fig. 5 STD against S. aureus (A), GT AgNPs against S. aureus (B). and investigation.

914 | Nanoscale Adv., 2022, 4, 911–915 © 2022 The Author(s). Published by the Royal Society of Chemistry
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Alaa Abdulmoneim Mohamed was responsible for concep- 19 V. V. Makarov, A. J. Love, O. V. Sinitsyna, S. S. Makarova,
tualization and investigation. I. V. Yaminsky, M. E. Taliansky and N. O. Kalinina,
“Green” nanotechnologies: synthesis of metal
Conflicts of interest nanoparticles using plants, Acta Naturae, 2014, 6(1), 35–44.
This article is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported Licence.

20 D. A. Kumar, V. Palanichamy and S. M. Roopan, Spectrochim.

There are no conicts to declare. Acta, Part A, 2014, 127, 168–171.
21 R. Sankar, P. Manikandan, V. Malarvizhi, T. Fathima,
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