Honey-Based and Ultrasonic-Assisted Synthesis of Silver Nanoparticles and Their Antibacterial Activities

Abstract

Food materials and biomolecular hosting the preparation of nanomaterials has achieved considerable
attention in recent years. This work demonstrates a facile, rapid, low cost, and environmentally
preparation of colloidal silver nanoparticles (Ag-NPs) under ultrasonic irradiation temperature using
natural honey as a capping and reducing agent. The effect of the Ag+ concentration, reducing agent,
and ultrasonic time in synthesis of Ag-NPs has been investigated. Well-defined spherical Ag-NPs
with a mean particle size of about 11.8 nm have been prepared due to the ultrasonic process.
The use of honey as a food capping/stabilizing and reducing agent provides green attributes to
this study. Ag-NPs capped by honey biomolecules exhibited antibacterial activity against pathogenic
bacteria. This suggested method is general and can be extended to other noble metals, such as
gold, palladium, and copper and may possibly find various additional application from medicine to
industry.

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Article
Journal of
Nanoscience and Nanotechnology
Vol. 16, 7989–7993, 2016
www.aspbs.com/jnn
Honey-Based and Ultrasonic-Assisted Synthesis of Silver
Nanoparticles and Their Antibacterial Activities
Reza Kazemi Oskuee1, Azhar Banikamali23, Bibi Sedigheh Fazly Bazzaz2,
Hasan Ali Hosseini4, and Majid Darroudi56∗
1Targeted Drug Delivery Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
2Biotechnology Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
3Neurogenic Inflammation Research Centre, Mashhad University of Medical Sciences,
Mashhad 9177948564, Iran
4Chemistry Department, Payame Noor University, 19395-4697 Tehran, Iran
5Nuclear Medicine Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
6Department of Modern Sciences and Technologies, Mashhad University of Medical Sciences, Mashhad, Iran
Food materials and biomolecular hosting the preparation of nanomaterials has achieved consider-
able attention in recent years. This work demonstrates a facile, rapid, low cost, and environmentally
preparation of colloidal silver nanoparticles (Ag-NPs) under ultrasonic irradiation temperature using
natural honey as a capping and reducing agent. The effect of the Ag+concentration, reducing agent,
and ultrasonic time in synthesis of Ag-NPs has been investigated. Well-defined spherical Ag-NPs
with a mean particle size of about 11.8 nm have been prepared due to the ultrasonic process.
The use of honey as a food capping/stabilizing and reducing agent provides green attributes to
this study. Ag-NPs capped by honey biomolecules exhibited antibacterial activity against pathogenic
bacteria. This suggested method is general and can be extended to other noble metals, such as
gold, palladium, and copper and may possibly find various additional application from medicine to
industry.
Keywords: Silver Nanoparticles, Honey, Ultrasonic Synthesis, Antibacterial.
1. INTRODUCTION
Food and biomolecules templating the synthesis of
nanoparticles has achieved considerable attention in recent
years due to their availability, biocompatibility, eco-
friendly, low cost, and low toxicity.1–5 Silver nanoparticles
(Ag-NPs) are one of the main metal and commercialized
nanoparticles. Ag-NPs have attracted much attention dur-
ing the past decays due to their potential applications
in medicine and industry.6–13 Many methods for fab-
ricating of Ag-NPs have been reported such as wet
chemical reduction,14 biological reduction,15 photochem-
ical reduction,16 UV-irradiation,1718 electrochemical,19
-ray irradiation,20 microwave,21 and laser ablation
technique,22–25 Therefore, investigating a simple method for
low-cost, eco-friendly, large-scale, and controlled growth
∗Author to whom correspondence should be addressed.
of Ag-NPs at normal physical and chemical conditions
is essential. Recently, honey has been used in green
synthesis of different nanomaterials as reducing, stabiliz-
ing, and capping agent and it is a suitable natural mate-
rial to apply green chemistry rules in nanoscience and
nanotechnology world.226–28 Honey is a sweet viscous
fluid which is fabricated by honeybees and it is basi-
cally consist of carbohydrates, enzymes, vitamins, minerals
and antioxidants.29 Honey-mediated and ultrasonic-assisted
synthesis of Ag-NPs has many advantages over other
biosynthesis methods, including avoidance of elaborate
processes e.g., drying and extraction of plant materials
and/or the maintenance of cell cultures. In this work, we
reported a simple method for the synthesis of well-defined
Ag-NPs that has the advantages of being a facile and
green process, food-directed, and of low cost. Silver nitrate
was used as the silver ions source at different ultrasonic
J. Nanosci. Nanotechnol. 2016, Vol. 16, No. 8 1533-4880/2016/16/7989/005 doi:10.1166/jnn.2016.13031 7989

Honey-Based and Ultrasonic-Assisted Synthesis of Silver Nanoparticles and Their Antibacterial Activities Oskuee et al.
irradiation times, silver cations concentration, and honey
contents. The obtained samples were characterized through
UV-vis spectroscopy, PSA, and TEM and their antibacterial
effects were then examined by MIC method.
2. MATERIALS AND METHOD
2.1. Materials and Reagents
Chemicals were analytical grade and used as received
without any further purification. AgNO3(99.98%, Merck
Figure 1. UV-vis spectra of Ag-NPs synthesized under different conditions; silver concentrations (A), honey concentrations (B), and ultrasonic
irradiation times (C).
KGaA, Darmstadt, Germany) and honey (Soltani, Iran)
were used as silver precursor and capping/reducing agent,
respectively. All glassware used in this work was cleaned
with a fresh solution of HNO3/HCl (3:1, v/v) and
washed thoroughly with double distilled water and dried
before use.
2.2. Synthesis of Ag-NPs
A modified method of Majid’s30 was adopted to synthe-
size Ag-NPs using natural honey. Colloidal Ag-NPs were
7990 J. Nanosci. Nanotechnol. 16, 7989–7993,2016

Oskuee et al. Honey-Based and Ultrasonic-Assisted Synthesis of Silver Nanoparticles and Their Antibacterial Activities
Figure 2. Particle size distribution of Ag-NPs synthesized in optimum
conditions; silver concentrations (0.3 M), honey concentrations (20 wt%),
and ultrasonic irradiation times (30 min).
prepared by reducing silver nitrate with ultrasonic waves
in the presence of natural honey. Briefly, 20 ml of sil-
ver nitrate solution (0.1 M) containing honey (20 wt%)
was exposed to high-intensity ultrasound irradiation under
ambient conditions for 20 min. Ultrasound irradiation was
carried out with a ultrasonic liquid processors (Hielscher
UP400S Sonicator, Germany, 24 kHz) immersed directly
into the reaction solution. For investigation of the role
of silver nitrate and honey concentrations in synthesis of
Ag-NPs, the experiment was also applied at different con-
centrations of silver nitrate and honey.
2.3. Characterization
The optical absorption properties of synthesized Ag-NPs
were characterized using a UV-vis spectrophotome-
ter (Lambda 35, PerkinElmer, USA) over the range
of 300–800 nm. The synthesized Ag-NPs were also
characterized using transmission electron microscopy
(TEM, LEO 12AB, Germany). The particle size distri-
butions of nanoparticles were determined using a par-
ticle size analyzer (PSA, Malvern, UK). The particle
size of nanoparticles was determined using the UTH-
SCSA Image Tool Version 3.00 program and their
Figure 3. TEM image (A) and its particle size distribution (B) of Ag-NPs synthesized in optimum conditions.
particle size distribution was calculated by SPSS software
Version 18.
2.4. Measurement of Minimum Inhibitory
Concentration (MIC)
MIC means minimum concentration of test material that
inhibits the growth of bacteria. MIC was determined
in 3 different bacteria including Staphylococcus aurous,
Pseudomonas aeruginosa, E. coli. Fifty micro-liters per
wells from each bacterium culture (106CFU/ml) were
poured in a sterile 96-well plate. Then serial dilutions of
nanoparticle solution were prepared in the media from
stock solution and added to the 96-well plates and incu-
bated at 37 C overnight. MIC was calculated using the
optical density of each well after 24 hrs.
3. RESULTS AND DISCUSSION
Five aqueous samples containing different concentrations
of silver nitrate (0.05, 0.1, 0.2, 0.3, and 0.4 M) and 20 wt%
honey are irradiated for 20 min. Depending on the silver
ions concentrations, the reaction solutions display a spec-
trum of yellow to dark brown colors. The UV-vis spec-
troscopy is a suitable route for displaying the presence
of metal NPs.3132 The optical observations illustrated that
these values varied at different experimental parameters
from 464 to 472 nm. Presence of one surface Plasmon res-
onance (SPR) bands in this region confirmed a spherical
shape of the Ag-NPs.33 While, there is a weak characteris-
tic UV-vis absorption peak of Ag-NPs after sonication for
Ag+concentration of 0.05 M (Fig. 1(A(a))), the SPR peak
at 464 nm displays the formation of Ag-NPs. Gradually
higher Ag+concentrations, from 0.1 to 0.4 M, increase
the corresponding peak intensities (Fig. 1(A(b–e))), with
concomitant red shifts from 468 to 472 nm. In order to
study the effect of honey concentration on synthesis of
Ag-NPs, samples containing 10 to 40 wt% of honey with
J. Nanosci. Nanotechnol. 16, 7989–7993, 2016 7991

Honey-Based and Ultrasonic-Assisted Synthesis of Silver Nanoparticles and Their Antibacterial Activities Oskuee et al.
constant concentration of silver nitrate (0.3 M) are synthe-
sized under ultrasonic irradiation for 20 min. Colloidal Ag-
NPs synthesized at lowest honey concentration (10 wt%)
appear yellow, showing a SPR band with max at 468 nm
(Fig. 1(B(a))).
As the concentration of honey is increased to 40 wt%,
the color of the solution gradually changes to dark brown,
displaying a red shift to 472 nm with continuous increase
in its intensity (Fig. 1(B(b–d))). As shown in these
results, when the initial Ag+and honey concentrations are
increased, Ag-NPs with smaller sizes and higher yields
are fabricated. To study the effects of reaction time on
Ag-NPs formation, samples with various times of ultra-
sonic irradiation e.g., 10, 20, 30, and 40 min are synthe-
sized. Depending on the ultrasonic irradiation times, the
reaction solutions display a spectrum of yellow to dark
brown colors. As the reaction time, the positions (max )
of the characteristic SPR peaks do not change signifi-
cantly (Fig. 1(C(a–d))). Upon increasing the irradiation
times, the SPR band intensities are remarkably increased,
rendering higher yields of Ag-NPs. It was subsequently
found that, in the honey solution, ultrasonic irradiation
played a key role in formation of Ag-NPs. Therefore, it
is possible to control the size and quantity of the Ag-NPs
by varying the ultrasonic irradiation parameter such as
irradiation time applied to the Ag+solutions. Formation
of small nanoparticles is also verified by optical proper-
ties of Ag-NPs due to the presence of SPR bands with
amax at about 450 nm. Thus, it can be assumed that
we have obtained colloidal silver solution having a parti-
cle size in nanoscale range (as determined by TEM). The
particle size distribution of prepared Ag-NPs in aqueous
honey solution was estimated by dynamic light scattering
(DLS) to evaluate the variation in particle size distribu-
tion. Figure 2 displays that the particle size of the Ag-NPs
ranged between 50 and 250 nm. Figure 3 illustrates a typ-
ical TEM image and its particle size distribution of the
synthesized spherical Ag-NPs and indicates the dominant
size of Ag-NPs about 118±490 nm with PDI =0439.
This is attributed to the high ‘surface area to volume’ ratio
and the quantum confinement effect caused by extremely
reduced size (i.e., electron confinement in a small
area).3034
The MIC values of prepared Ag-NPs against tested
microorganism were shown in Table I. The MIC value of
E. coli and Staphylococcus aureus was 19.46 ppm which
means better antibacterial activity of Ag NPs comparing
to Pseudomonas aeruginosa.
Tab l e I . The MIC for different bacteria.
Bacteria MIC (ppm)
Pseudomonas aeruginosa 40
Escherichia coli 19.46
Staphylococcus aureus 19.46
4. CONCLUSION
In ultrasonic assisted chemically route, colloidal Ag-NPs
can be synthesized by “green” chemistry method using
silver nitrate and honey as silver precursor and as a
reducing/stabilizing agent, respectively. Sono-chemically
reduction method revealed a suitable potential for prepar-
ing desired particle size and concentrations of colloidal
Ag-NPs. The concentrations of silver nitrate and honey
and ultrasonic irradiation time are the main effective
parameters in synthesis of Ag-NPs. As ultrasonic irra-
diation time increase, Ag-NPs become smaller and their
concentrations are enhanced. The results of this work indi-
cated a potential application of Ag-NPs in the inhibition
of oral microorganism infections and using in surgical
devices, instruments and wound healing bandages.
Acknowledgment: This study is the result of an
M.Sc.thesis(Mr.A.Banikamali) and has been finan-
cially supported by the Vice Chancellery of Research of
Mashhad University of Medical Sciences.
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Received: 17 November 2015. Accepted: 29 March 2016.
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