Industrial Crops and Products 60 (2014) 212–216

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Industrial Crops and Products

journal homepage: www.elsevier.com/locate/indcrop

Coriandrum sativum seed extract assisted in situ green synthesis of

silver nanoparticle and its anti-microbial activity
G.M. Nazeruddin a , N.R. Prasad a,b,∗ , S.R. Prasad d , Y.I. Shaikh a ,
S.R. Waghmare b , Parag Adhyapak c
a
Poona College of Arts, Commerce & Science, Pune, India
b
Solid State Research Laboratory, Department of Chemistry, Shivaji University, Kolhapur, Maharashtra 416004, India
c
Material Research Laboratory, Centre for Material for Electronic Technology, Pune, India
d
DKTE Society’s Textile and Engineering Institute, Ichalkaranji, India

a r t i c l e i n f o a b s t r a c t

Article history: Extracellular in situ synthesis of metal nanoparticles using extracts of plants like Azadirachta indica
Received 17 December 2013 (Neem), and Zingiber officinale (Ginger) has been successfully carried out earlier. In this study we have
Received in revised form 13 May 2014 developed a novel method to synthesize silver nanoparticles by the process of bio-chemical reaction
Accepted 26 May 2014
between silver salt solution with seed extract of Coriandrum sativum without using any surfactant or
external energy. By this method physiologically stable, bio-compatible Ag nanoparticles (NPs) were
Keywords:
formed. These functionalized AgNPs could be used for targeted drug delivery with enhanced therapeutic
AgNPs
efficacy and minimal side effects. It is well known that on treating the metallic salt solution with some
Green synthesis
C. Sativum
plant extracts, a rapid reduction occur leading to the formation of highly stable metal nanoparticles. This
SEM method resulted in rapid synthesis of nanoparticles, i.e. reaction time was 1–2 h as compared to 2–4 days
Antimicrobial required by microorganisms. These nanoparticles were analysed by various characterization techniques
to reveal their morphology and chemical composition. TEM image of these NPs indicated the formation
of spherical, non-uniform, poly dispersed nanoparticles of mean size of 13.09 nm. A detailed study of
anti-microbial activity of nanoparticles and its synergetic effectiveness with common narrow spectrum
antibiotic has been successfully carried out.
© 2014 Published by Elsevier B.V.

1. Introduction
Nanomaterials have a long list of applicability in improving
human life and its environment. It has been found that 5000 years
old Indian System of Medicine Ayurveda had some knowledge of
nano-scale fabrication used for medicinal purposes (Prasad and
Elumalai, 2011). The process of formation of Bhasma (a special
kind of ayurvedic medicine) is well described in ancient book Ras-
Ratnakar by Nagarjuna, 50 BC. A bulk material has constant physical
properties regardless of its size, but at the nano-scale often this
is not true. Several well-characterized bulk materials have been
found to possess most interesting properties when studies in the
nanoscale. There are many reasons for this including the fact that
nanoparticles possess a very high aspect ratio, i.e. surface to volume
ratio. Nanoparticles are of great scientific interest as they bridge
∗ Corresponding author at: Department of Nanoscience and Technology, Shivaji
University, Kolhapur, India. Tel.: +91 9404265521.
E-mail address: neeraj prasad21@rediffmail.com (N.R. Prasad).
the gap between bulk materials and atomic and molecular level
(Thakkar et al., 2010). Large spectrum of research has been focused
to synthesize nanomaterials with complex morphologies and to
control the size and shape which is crucial in tuning their properties
(Khan et al., 2012; Vijaymohan et al., 2012). Several approaches are
in practice to generate the metallic nanoparticles such as electro-
chemical, sono-chemical and microwave assisted process, but most
of these strategies suffer from utilization of high energy, hazardous
chemicals and difficulty in purification (Vijaykumar et al., 2013). To
switchover these technical hitches biological principles have been
established recently (Castro et al., 2011).
DNA can be used as an ideal template for the synthesis of
nanoparticles due to some key features such as inherent nanoscale
dimensions, high specificity exerted towards inorganic materials
and structural flexibility to build NPs of different size and shape
(Aland, 2006).
Silver nanoparticles can be produced either intra or extra-
cellularly by using biological routes such as algae, fungi, bacteria
and plants, etc. There are some species of algae which are
used as bio-factory for the synthesis of metallic nanoparticles.

http://dx.doi.org/10.1016/j.indcrop.2014.05.040
0926-6690/© 2014 Published by Elsevier B.V.
 G.M. Nazeruddin et al. / Industrial Crops and Products 60 (2014) 212–216
While micro-organisms continue to be investigated for reduc-
tion of metals to synthesize nanoparticles, the use of plant
extracts is an exciting possibility and is relatively unexplored and
under-exploited (Mittal et al., 2013).
Plants have been known to bio-mineralize calcium carbonate,
silica and even magnetite internally. Certain plants are known to
hyper-accumulate these heavy metals or can even be induced to
hyper-accumulate within different parts of plants. The internal
accumulation of metal in plants can occur both via complexation of
the metal ion with a suitable bio-ligand in its native oxidation state
or after its reduction to a lower oxidation state. The possibility of
reduction of metal ions by plants and the presence of metal com-
plexing agents in them entices a materials scientist to use plants for
the goal of synthesizing nanoparticles and controlling their size and
shape and to experiment on the likelihood of forming nanoparti-
cles of low reduction potential metals (Anilkumar, 2007). Solution
temperature, concentration of metal salt and reducing agent and
reaction time all influence the shape and size of particle. The possi-
bility of synthesizing nanoparticles of different compositions using
plants would offer an environmentally friendly alternative to the
existing potentially toxic chemical and physical methods of prepa-
rations. This would also help stave off the growing apprehensions
related to environmental degradation and biological hazards apart
from being a cost effective process with potential to scale up for
large-scale synthesis.
Shiv Shankar has reported the synthesis of silver and gold
nanoparticles using decoction solution of Azadirachata indica. How-
ever, the nanoparticles synthesized by him are in the range of
50–100 nm and are not well separated from each other and tend to
form agglomerated structures. By reducing silver nitrate in solution
of tea extract or epicatechin of varying concentrations, spherical
silver nanoparticles were formed that had controllable size dis-
tribution depending upon the concentration of tea extract and
epicatechin in the samples (Kharisov et al., 2013).
In this research work an attempt has been made to extend these
green chemistry approaches for synthesizing nano-materials from
lower class of organisms to higher forms of plants. It leads to green
chemistry route which is more advantageous because it does not
require elaborate process such as intra cellular synthesis, multi-
ple purification steps, and maintenance of microbial cell culture.
In green chemistry approach of synthesis the reaction is normally
carried out at room temperature although in some processes a lit-
tle heating below 100 ◦ C was applied. High amount of phenolic
compounds in the plant extract are generally supposed to influ-
ence the reduction process and stabilize nanoparticles preventing
agglomeration. These nanoparticles are studied for their antibac-
terial properties and future work can be extended to study their
antioxidant, and anti-tumour properties (Rao et al., 2013).
Coriandrum sativum is widely available plant in tropical coun-
try like India. In the present research work C. sativum seed extract
have been used as a reducing agent for AgNPs biosynthesis. The
same extract also acts as a capping agent. This plant is described to
possess diuretic, purgative and appetizer property in Ayurveda. It is
also used as a medicine against griping pain, piles, leucorrhoea and
in eye diseases. It is reported in the literature that high consump-
tion of C. sativum leads to impotency in males and suppression of
menstrual cycles in female.
1.1. Silver nanoparticles
AgNPs can be used in antimicrobial agents, textile indus-
tries, water treatment, sunscreen lotions, etc. Silver nanoparticles
exhibit tremendous applications in various areas such as biomed-
ical field, spectral selective coatings for solar energy absorption,
optical receptors, bio-labelling, intercalation materials for elec-
trical batteries, filters, anti-microbial agents and sensors, etc.
213
(Shankar, 2005). Due to the development of antibiotic resistance in
pathogenic bacteria the pharmaceutical companies and researchers
are searching for new antibacterial agents and the AgNPs are
promising candidate for the same. The reduction of Ag+ ions leads
to the formation of silver atoms (Ag) which is followed by agglom-
eration into oligomeric clusters. These clusters eventually lead to
the formation of colloidal AgNPs. When the colloidal particles are
much smaller than the wavelength of visible light, the solution
possess characteristic yellow colour with an intense band in the
380–430 nm range and other less intense bands at longer wave-
length in the UV–visible absorption spectrum.
2. Materials and methods
Nanoparticles of Ag were prepared by using C. sativum seed
extract. C. sativum seeds have been purchased from local market
Jaysingpur, India. 50 g of seeds have been taken for the prepara-
tion of extract. The obtained seeds were sieved and thoroughly
washed in running water and finally rinsed with deionized water
until no foreign material remained behind. After washing the col-
lected seeds were kept in 100 ml of deionized water over night and
macerated. Then it was finely crushed in grinder mixer and filtered
by using cotton and collected in stoppered glass bottle. The extract
is diluted to 250 ml using deionized water and the solution was
kept in refrigerator at 4 ◦ C. The AgNO3 used in reaction was analyt-
ical grade chemical obtained from Sigma Aldrich and used without
further purification.
100 ml of 0.1 N AgNO3 solution was taken in 500 ml beaker.
Then 50 ml of C. sativum seed extract solution was taken in burette
and drop wise added at room temperature with constant stir-
ring. As soon as the seed extract solution was added dark brown
coloured precipitate appeared in the solution. After complete addi-
tion of 50 ml of solution a sufficient amount of the precipitate
was observed and it was separated by high speed centrifugation
at about 8000 rpm. The formation of AgNPs requires about 60 min
of time and that is determined using spectro-photometer. The sep-
arated solid mass was washed with alcohol for few times to remove
organic impurities soluble in alcohol. After complete washing the
solid mass was kept in oven for drying. The complete drying of
this solid mass resulted in a black coloured material which was
powdered in mortar and sampled for characterization purpose.
3. Experimental results and discussion
Biosynthesis of AgNPs was indicated by colour developed in
solution during the course of reaction. The intensity of colour
depends upon degree of bio-reduction of silver particles. Forma-
tion of AgNPs was confirmed by using different characterization
techniques.
3.1. UV spectroscopic graph showing absorbance of silver
nanoparticles
The blackish brown coloured sample powder was dissolved in
deionized water and sonicated. Then this solution was taken in
cuvette and exposed to UV–visible radiation and the absorbance
of the solution was recorded. Because of the surface plasmon reso-
nance phenomena resonant peak occurs at different wavelength for
different nanoparticles solution and as per the theory of resonance
maximum wavelength is absorbed at resonant wavelength. It is
reported in the literature that a typical AgNPs shows the character-
istic SPR at the wavelength in the range of 400–480 nm. Fig. 1 shows
SPR for the sample solution to occur at the wavelength of 421 nm
which confirms the presence of silver nanoparticles in the pre-
pared solution. The SPR absorbance is sensitive to the nature, size
214 G.M. Nazeruddin et al. / Industrial Crops and Products 60 (2014) 212–216
Fig. 1. UV spectroscopic graph showing absorbance of silver nanoparticles.
and shape of particles present in the solution and also it depends
upon their inner particle distance and the surrounding media. The
MATLAB analysis of this UV image shows that the image format is
JPEG with bit-depth of 24 bits and image resolution is found to be
492 × 492 and the file-size is 13.7 kB. Also it is seen that the image
class is uint8. The UV characterization has been done at Department
of Chemistry, Shivaji University, Kolhapur.
3.2. XRD graph showing silver nanoparticles
XRD patterns of synthesized AgNPs are shown in Fig. 2 which
is an absorbance vs 2␪ graph. The graph shows eight major peaks
and the peak position explains about the translational symmetry
namely size and shape of the unit cell, whereas the peak intensities
give details about the electron density inside the unit cell. The syn-
thesized nanoparticles are crystalline in nature. These peaks are
of cubic structure which is in agreement with the JCPDS file No.
00-004-0783.
3.3. Particle size distribution
Particle size distribution histogram from Fig. 3 indicates that
10.07% particles size is around 113 nm with a standard deviation
of 16.48% size while the remaining particles are around 633.5 nm
with a standard deviation of 20.58%. The difference in size as
observed from TEM and PDS may be due to the presence of bio-
active molecules of C. sativum on AgNPs surface. Prathna et al.
Fig. 2. XRD graph showing silver nanoparticles.
Fig. 3. Graph showing particle size distribution for Ag nanoparticles.
found the different particle size of AgNPs by changing the char-
acterization techniques and the particle size order is reported to
be DLS > AFM > TEM > XRD and our results are consistent with this
finding. The MATLAB analysis of this particle size distribution image
shows the bit-depth of 24 bits and image resolution of 1024 × 768.
Also the image file-size was 124.9 kB and image format JPEG with
image class as uint8. The particle size distribution graph is taken
from Dr D Y Patil University, Kolhapur, India.
3.4. SEM image of silver nanoparticles
The SEM image from Fig. 4 reveals the formation of cluster of
spherical beadlike structure of silver nanoparticles with nonuni-
form distribution. The MATLAB analysis gives the pixel depth of the
image equal to 8 bits and the image resolution of 1280 × 960. Also
the image file-size was found to be 1.23 MB and the image format
as TIFF. The SEM image has been taken with JSM-6360 instrument
which uses accelerating voltage of 20 kV. The SEM Image has been
taken at Department of Physics, Shivaji University, Kolhapur India.
3.5. TEM image of silver nanoparticles
TEM images from Fig. 5 reveal that there was poly-disperse
spherical particles with nonuniform distribution of nanoparticles
in the prepared sample. For TEM measurements, a drop of solution
containing the particle was deposited on a copper grid covered with
amorphous carbon. After allowing the film to stand for 2 min the
Fig. 4. SEM image of silver nanoparticles.
 G.M. Nazeruddin et al. / Industrial Crops and Products 60 (2014) 212–216
Fig. 5. TEM image for silver showing size of nanoparticles.
extract solution was removed by means of blotting paper and the
grid allowed drying before the measurement. It was observed that
the nanoparticles formed were of different sizes and particle size
was found to be 9.94, 13.94, 15.91 and 12.59 nm and the mean size
of about 13.09 nm which lies in the nano range. The TEM measure-
ment was done with JEOL model 1200Ex instrument operated at
an accelerating voltage of 80 kV. The TEM image was taken with
very high resolution and MATLAB analysis gives the pixel depth of
the image equal to 24 bits and the image format as JPEG. The TEM
images have been taken from National Chemical Laboratory, Pune,
India.
3.6. Antimicrobial activity of silver nanoparticles
The antimicrobial activity of AgNPs was evaluated against a
Gram-positive and a Gram-negative pathogenic bacteria as per
below mentioned agar diffusion method. In this method first the
bacterial cultures were developed on nutrient agar (NA) slants
which contained different ingredients such as peptones (5.0 g), agar
(15.0 g), meat extract (q.s.), sodium chloride (q.s.), and yeast extract
(q.s.) per litre of distilled water. After solidification of prepared cul-
ture medium the stainless steel cylinder were placed on the surface
and holes were punched in the medium. Freshly prepared AgNPs
were added into the wells that were made in the culture medium
plates. The samples were initially incubated for 15 min at 4 ◦ C tem-
perature and later were incubated for 24 h at 37 ◦ C temperature.
Positive test results were scored when a zone of inhibition was
observed around the well after the incubation process.
The antimicrobial activity has been tested against Gram-positive
and Gram-negative bacteria. A Gram-positive bacterium used for
experimentation was Bacillus subtilis NCIM 2635. The antimicro-
bial study given in Fig. 6 showed the well size to be 8 mm and
zone of inhibition of about 13 mm. Also the antimicrobial activity
of common narrow spectrum drug Penicillin is seen and is found
to be 13 mm, but the synergetic study with 1:1 mixture of AgNPs
and Penicillin shows a significant increase in antimicrobial activ-
ity and the zone of inhibition is found to be 17 mm. Thus it can
be claimed that these AgNPs are exhibiting antibacterial property
against Bacillus subtilis NCIM 2635 also the antimicrobial activity
of Penicillin has increased in the presence of AgNPs. Thus AgNPs
215
Fig. 6. Image of culture plate showing antimicrobial activity of silver nanoparticles.
can be used in antimicrobial coatings in medical instruments and
in textiles. The image was captured by Sony Cybershot camera and
shown in the picture. The MATLAB analysis of this antimicrobial
culture image shows that the image format is JPEG with bit-depth
of 24 bits, image class is uint8 and image resolution 4000 × 3000
and the file-size is 1.55 MB.
3.7. Possible mode of action of silver nanoparticles as
antibacterial agent
There are different theories behind the mode of action of AgNPs
as antibacterial agent. One of the theory says that silver is a soft
acid, and there is a natural tendency of an acid to react with a
base, in that case soft acid reacts with soft base. The cells are
mainly made up of sulfur and phosphorus which are soft bases.
Thus the reaction between nanoparticles (soft acid) and cells (soft
base) takes place which results in the formation of salt, i.e. degen-
eration and death of cells takes place. The interaction of the silver
nanoparticles with the sulfur and phosphorus of DNA can lead to
problems in the DNA replications of bacteria and thus terminate
the microbes.
However as per another theory positive charge on silver ion
may be responsible for its antimicrobial activity. It is possible
that the antibacterial activity is derived through the electrostatic
attraction between negatively charged cell membrane of micro-
organism and positively charged nanoparticles. In the present
scenario, AgNPs as antimicrobial agent have come up as promis-
ing candidate in the medical field. But still the exact mechanism
of the antibacterial effect of silver ions is not clearly under-
stood.
4. Discussion
It is well known that few medicinal plants exhibit anti-oxidant
property. Thus they can act as biological source of reducing agent.
On this belief choice of plants for this purpose were those carrying
medicinal and aromatic properties.
216 G.M. Nazeruddin et al. / Industrial Crops and Products 60 (2014) 212–216
Fig. 7. Possible reaction mechanism for reduction of metal ions using plant.
A thorough study on literature of C. sativum reveals that the
major components of the plant are coriandrol-C10 H18 O, maleic acid
(about 0.25%), volatile oil, sucrose, tannin, fatty acids and minerals,
etc. The aqueous seed extract contains protein which may act as a
bio-ligand. It has been reported by Lin et al. that the carbonyl groups
of proteins found in plants have a strong affinity to bind metals. So
the plant seed extract can act as capping agent and thus protect the
nanoparticles from agglomeration (Chunekar, 2010).
In addition, C. sativum plants possess traces of several antioxi-
dants, polyphenols, including flavonoids. The phenolics and other
phytochemicals within the plant not only result in the effective
reduction of silver salt to NPs but their chemical framework is also
effective at wrapping around nanoparticles to provide excellent
robustness against agglomeration.
Phenolic compounds possess hydroxyl and carboxyl groups
which are able to bind metals. The roots of many plants contain
high level of phenolic compounds which may inactivate ions by
chelating. Luteolin is a common flavones found in aerial part of
some plants. It is a known antioxidant, a free radical scavenger, an
agent in prevention of inflammation and a promoter carbohydrate
metabolism. The formation of enol form of the luteolin freely liber-
ates reactive hydrogen which is responsible for the conversion of
Ag+ to Ag0 and this reaction is illustrated in Fig. 7. The antioxidant
effectiveness of many polyphenols is essentially a result of ease
with which hydroxyl group is donated to a free radical and the abil-
ity of the aromatic structure to support an unpaired electron. The
exact mechanism of formation of these nanoparticles in biological
media is unknown. Presumably biosynthetic products or reduced
cofactors play an important role in the reduction of respective salts
to nanoparticles. It is quite probable that the phenols play an impor-
tant part in the reduction of ions to AgNPs as the concept of antiox-
idant action of phenol compound is not new (Ahmad et al., 2010).
5. Conclusion
The present green synthetic method is an easy, economical,
rapid and eco-friendly way to synthesize metallic nanoparticles
and capable of synthesizing AgNPs at room temperature. There
is no need of any external capping agent. This method produced
poly-dispersed, spherical nanoparticles which are well separated
from each other and no aggregation was observed. The method
discussed here involves a single pot synthesis. Based on the exper-
imentation, it can be concluded that this approach can be used
for industrial scale fabrication of nanomaterials. The experimenta-
tion showed that AgNPs possess significant antimicrobial activity
against Gram-positive bacteria Bacillus subtilus. The synergistic
study of AgNPs and common narrow spectrum antibiotics indicates
that the potency of common antibiotics can be enhanced in the
presence of Ag nanoparticles. Thus there is a future platform for
preparing nano-medicines, and targeted drug delivery, etc. Lastly,
it can be concluded that the green synthesis of nanoparticles is
an emerging and exciting area of nanotechnology and may have
significant impact on further advances in biotechnology.
Acknowledgements
The authors are thankful to Assistant Professor Hasan Osman
for all kind support in the progress of research work. They are
thankful to Dr. Prakash Sane for helping in TEM images and Vish-
wajeet Khot for helping in particle size distribution They are also
grateful to Dr. Sonavane, Department of Biotechnology, Ms. Sonali
Kalake, Ms. Dhanshree Mali, and Mr. Jaykumar Khot, Department
of Nanoscience and Technology, Shivaji University, Kolhapur, India
for analysis of antimicrobial activity.
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