chapter 3 synthesis and characterization of silver …€¦ · chapter 3 synthesis and...

Chapter 3

SYNTHESIS AND CHARACTERIZATION OF SILVER

NANOPARTICLES

3.1 Introduction

The important properties of nanoparticles that could affect nanoparticle behaviour

and toxicity comprises of particle size, shape, surface properties, aggregation

state, solubility, structure and chemical make-up. Therefore, when analysing

nanoparticles in different matrices, it is not only the composition and

concentration that will need to be determined but also the physical and chemical

properties of the engineered nanoparticles within the sample and the chemical

characteristics of any capping/functional layer on the particle surface. Methods

are available that have been developed for natural nanomaterials or engineered

nanomaterials in simple matrices which could be optimized to provide the

necessary information. These include microscopy, chromatography,

spectroscopy, centrifugation as well as filtration and related techniques. A

combination of these is often required (Tiede et al., 2008).

This chapter describes the synthesis of silver nanoparticles in yeast growth media

(YM media). The verification of the synthesis process and particle

characterization was done by the techniques of UV-Vis absorption spectroscopy,

transmission electron microscopy (TEM) and X-ray diffraction (XRD). A general

discussion of the experimental findings, in view of previous reports, concludes

the chapter.

A wide range of spectroscopic methods is available for nanoparticle analysis and

characterization. UV-Vis and infrared spectroscopy offer the possibility to

characterise nanoparticles, especially quantum dots and organic based

nanoaprticles like fullerenes and carbon nanotubes. Fourier transformation

infrared (FTIR) and UV-Vis spectroscopy have been used to compare aqueous

Synthesis and Characterization of Silver Nanoparticles 53

colloidal suspensions of C60 (Andrievsky et al., 2002). Pesika et al. (2003) also

used UV spectroscopy to study the relationship between absorbance spectra and

particle size distributions for quantum-sized nanocrystals. Particularly in the case

of silver nanoparticles, the UV-Vis absorption spectra have proved to be quite

sensitive as the plasmon peak and full width at half maximum (fwhm) depends on

the extent of colloid aggregation (Yamamoto et al., 2001).

X-ray diffraction is a non-destructive technique and can reveal information about

the crystallographic structure, size and elemental composition of natural and

manufactured materials. It has been widely applied in the characterization of

nanoparticles including those synthesized using biological agents. The diffraction

spectra can be conveniently compared with powder diffraction files (PDF)

available from the International Centre for Diffraction Data, USA for verification

of nanoparticle composition.

Microscopic techniques can be applied not only to visualize nanoparticles but

also to generate useful data on the size, size distribution and other measurable

properties (Rabinski and Thomas, 2004). Electron microscopy is one of the

techniques that have played a major role on studding nanoparticles. Since the

early conventional bright field images and the intermediate resolution dark field

techniques, to the high-resolution atomic images of nanoparticles the results have

shown that indeed the nanoparticles, in the range of a few nanometers, can have

well-defined crystal structures (Jose-Yacaman et al., 2001).

3.2 Materials and Methods

A list of all buffers and reagents used in the following protocols are described in

Appendix I.

3.2.1 Synthesis of Silver Nanoparticles

Silver nanoparticles used throughout this study were synthesized in the lab by the

Creighton method which involves reduction of silver nitrate (AgNO3) by sodium


borohydride (NaBH4). Reactions were carried out in de-ionized water as well as

in liquid Yeast Malt (YM) growth media.

Requirements

§ AgNO3 solution: 1.0 x 10-3

M

§ NaBH4 solution: 2.0 x 10-3

M

§ Magnetic stirrer & bar

§ Burette and glasswares

Procedure

(i) In a 100 ml Erlenmeyer flask 0.84 g of Yeast Malt media (HiMedia Labs)

was dissolved in 15 ml of de-ionized water.

(ii) 15 ml of 2.0 x 10-3

M NaBH4 (Merck) was then added and the flask was

cooled at 4ºC for 30 min.

(iii) A stir bar was placed in the Erlenmeyer and the assembly was centered on a

magnetic stir plate.

(iv) Under constant stirring, 10 ml of 1 x 10-3

M AgNO3 (SRL) was added drop-

wise using a burette supported with a clamp. The stirring was stopped

immediately after all of the silver nitrate solution had been added.

Note: All glasswares were first cleaned with chromic acid and then with

detergent. A final rinse was done in alcoholic KOH and then in de-ionized water

before use.

If stirring is continued once all the silver nitrate has been added, aggregation is

likely to occur.

The method described above was used for the synthesis of silver nanoparticles of

concentration 26.95 µg/ml in YM media. Other concentrations of Ag NPs were

also synthesized which are detailed in Table 3.1.


Stock Solutions

(I) Silver Nitrate Solution (AgNO3)

(a) 2.0 x 10-3

M (b) 2.6 x 10-3

M

(II) Sodium Borohydride Solution (NaBH4)

(c) 4.0 x 10-3

M (d) 4.0 x 10-2

M (e) 5.2 x 10-2

M

Table 3.1: Cocktail for nanoparticle synthesis

CONCENTRATION

(µg/ml)

AgNO3 (M) * NaBH4 (M) #

YM media (g)

5.39

2.0x10-4

(1ml stock a)

4.0x10-4

(3ml stock c)

0.84

16.17

6.0x10-4

(3ml stock a)

1.2x10-3

(9ml stock c)

0.84

26.95

1.0x10-3

(5ml stock a)

2.0x10-3

(15ml stock c)

0.84

37.73

1.4x10-3

(7ml stock a)

2.8x10-2

(21ml stock d)

0.84

48.51

1.8x10-3

(9ml stock a)

3.6x10-2

(27ml stock c)

0.84

53.90

2.0x10-3

(10ml stock a)

4.0x10-2

(30ml stock c)

0.84

70.07

2.6x10-3

(10ml stock b)

5.2x10-2

(30ml stock e)

0.84

* Final volume made up to 10 ml with de-ionized water. # Final volume made up

to 30 ml with de-ionized water.


3.2.2 Characterization of Nanoparticles

The synthesized particles were characterized by absorption spectroscopy, TEM

and XRD. This section describes how the analyses were made.

3.2.2.1 Analysis by Absorption Spectroscopy

The absorbance spectrum of the nanoparticle solutions – both in de-ionized water

and in YM media – were recorded immediately (within 1h) after the synthesis

was done. Before recording the absorbance spectrum a reference of de-ionized

water or pure YM media was recorded, as appropriate.

To assess the stability of the nanoparticle solutions, the absorbance spectrum was

recorded at 1 day, 15 days and 30 days after synthesis.

The spectrophotometer used was a Chemito UV Scan 2600 (Thermo Fisher) and

the software was Spectrum™ Version 6.87. Absorbance spectra were recorded

over the range of 300 – 700 nm. Wavelength of peak absorbance, λmax was noted

and the fwhm (full width at half-maximum) was calculated according to He et al.

(2001).

3.2.2.2 Analysis by Transmission Electron Microscopy

Particle size and shape were determined with a JEOL JSM 100 CX II

transmission electron microscope operating at a maximum of 100 kV.

100 µl of nanoparticle solutions were deposited on carbon coated copper grids

(400 mesh) and dried at 30ºC before image capture.

Particle size distributions were determined from TEM images and results were

plotted using GraphPad Prism®

Version 5.03.


3.2.2.3 Analysis by X-Ray Diffractometer

The crystal structure and also the particle size were studied by X-ray Diffraction.

1 ml of the nanoparticle solutions were spread on a glass slide and dried at 40ºC

in an oven. The process was repeated 3-4 times to obtain a thin film.

The spectra were recorded in a Phillips Xpert Pro Diffractometer (Cu Kα

radiation, λ1 = 1.54056; λ2 = 1.54439) running at 40 kV and 30 mA. The

diffracted intensities were recorded from 35.01 degrees to 79.99 degrees 2θ

angles.

The crystalline size was calculated from the half-height width of the diffraction

peak of XRD pattern using the Debye-Scherrer equation:

Where,

D = crystalline size, Ǻ

K = crystalline-shape factor

λ = X-ray wavelength

θ = observed peak angle, degree

β = X-ray diffraction broadening, radian

The crystallinity of the particles was evaluated through a comparison of

crystallite size from XRD and TEM particle size determination by the following

equation:

Where,

Icry is the crystallinity index

Dp is the particle size (obtained from either TEM or SEM morphological

analysis)

Dcry is the particle size (calculated from the Scherrer equation)


3.3 Experimental Findings

3.3.1 Particle synthesis and concentrations

Nanoparticles were synthesized in de-ionized water as well as in YM media

without the use of any stabilizing agent.

Figure 3.1 Silver nanoparticles synthesized in YM media at various

concentrations. (a) YM media (w/o Ag NPs) and YM media with 5.39, 26.95,

43.12, 48.51 and 53.90 µg/ml Ag NPs (left – right) and (b) Pure YM media (w/o

Ag NPs) and YM media with 58.29, 64.68 and 70.07 µg/ml Ag NPs (left – right).

The colour of the colloidal solutions varied from light yellow to dark brown

(Figure 3.1) depending on the concentration of nanoparticles. There was no

obvious aggregation of particles as the colour remained stable even after


autoclaving at 15 psi for 20 min. Aggregation is generally visualized by a change

in colour as the yellow darkens, turns violet then grayish as the particles settle

out. Finally a colourless solution with black precipitates of silver is seen. The

formation process and the optical properties of the silver nanoparticles can also

be identified from both the colour change and the UV-Vis spectra of the solutions

(He et al., 2002).

An important aspect of the synthesis process was the determination of particle

concentration, as the effect on cells would likely be dose dependent. We followed

a theoretical method based on the assumption that there is a compete reduction of

the AgNO3 used in the synthesis process. This is likely to be the case as the

reducing agent, NaBH4, was used in excess (up to 2 x 102 times) to that of the

silver salt. The UV-vis spectra of the synthesized nanoparticle solutions described

in Section 3.3.2 indicate that indeed there is total reduction of the silver salt as

absorption peaks attributable to silver ions (Ag+) were not detected.

In a typical synthesis reaction the silver salt was reduced to metallic silver

according to the following reaction:

The concentration of silver particles in the solution would then depend on the

molarity of the initial silver nitrate solution and the total reaction volume.

For example, in the synthesis of 26.95 µg/ml nanoparticle concentration:

10 ml of 10-3

M AgNO3 was reduced with 30 ml of 2 x 10-3

M NaBH4. Total

silver (At. wt. 107.8) content of the reaction volume would be 1.078 x 10-3

g in

40 ml which is equivalent to 26.95 µg/ml.

Likewise, nanoparticle solutions of different concentrations were prepared by

varying the strength of the starting silver nitrate solution albeit with concomitant

changes in the NaBH4 concentration to ensure complete reduction. A similar

method to calculate nanoparticle concentration was described by Xu et al. (2004).


3.3.2 Optical properties

The optical properties of nanoparticle colloidal solutions of various

concentrations were determined in a UV-Vis spectrophotometer. The spectra

recorded are presented below:

Figure 3.2 UV-Vis absorption spectra of silver nanoparticle solutions of

concentrations 5.39 µg/ml (I) and 16.17 µg/ml (II).



concentrations 26.95 µg/ml (III) and 37.73 µg/ml (IV).



concentrations 48.51 µg/ml (V) and 53.90 µg/ml (VI).



concentrations 64.68 µg/ml (VII) and 70.07 µg/ml (VIII).


Figure 3.6: Super-imposed UV-Vis spectra of the various nanoparticles

concentrations presented in Figures 3.2 – 3.5 above. Peak absorption can be

observed between 380 nm to 420 nm. Time: 1 h after synthesis.

Table 3.2: Optical characteristics of synthesized nanoparticle solutions: Peak

absorbance (λmax) & full width at half maximum (FWHM).

S. No Nanoparticle Concentration

(µg/ml)

λmax

(nm)

FWHM

(nm)

1 5.39 416.5 77.50

2 16.17 416.5 97.50

3 26.95 407.0 92.50

4 37.73 396.5 96.25

5 48.51 392.0 108.75

6 53.90 391.0 82.50

7 64.68 390.5 93.75

8 70.07 393.5 85.00


Figure 3.7 UV-Vis spectra of representative silver nanoparticle solution recorded

at different time intervals: 1 day (a); 15 days (b) and 30 days (c).

Table 3.3: Optical characteristics of a representative nanoparticle solution

recorded at different time intervals.

Time after Synthesis

(days)

λmax

(nm)

FWHM

(nm)

1 398.5 97.50

15 398.2 97.75

30 408.2 101.25

The absorption spectrum of the samples (Table 3.2) show a well-defined plasmon

band between 416 – 390 nm with a fwhm range of 77 – 109 nm, which are

characteristic of nanosized silver. Theoretical and experimental studies

(Chakraborty, 1998; Otter, 1961), in which the optical properties of silver

particles have been discussed report the appearance in the electronic absorption

spectrum of a band located at 396 nm, associated with the presence of small

spherical silver nanoparticles. If the particles would not be spherical (or

equiaxial), the absorption band would appear at longer wavelengths and would


gradually shift to shorter wavelengths as the particles become more spherical

(Chakraborty, 1998; Baia et al., 2006). Because the position of the electronic

absorption bands shown in Figure 3.6 and Table 3.2 are close to the above-

mentioned value we assume that the particles synthesized in YM media have a

roughly spherical shape. Particle size and shape were further confirmed by

electron microscopy as described below in Section 3.3.4.

The individual UV-VIS absorption spectra of the Ag NP samples, with particle

concentrations of 5.39 µg/ml to 70.07 µg/ml, are displayed in Figures 3.2 to 3.5.

Absorption bands are smooth with a single pronounced plasmon resonance that

appear around 400 nm. Lu et al. (2005) have reported that the electronic

transitions involving the Ag+ ion give rise to absorption bands located between

200 and 230 nm, whereas the electronic transitions of metallic Ag0 appear in the

250-330 nm spectral range. The UV-VIS spectra of all samples analyzed in this

study did not show distinct absorption signals around 230 nm due to the

electronic transitions involving Ag+ ions. The presence of silver ions in the

synthesized nanoparticle solutions can so be safely assumed to be either non-

existent or of infinitesimal concentration. As has been mentioned in Section

3.3.1, a large excess of the reducing agent was used during the synthesis process,

possibly leading to a complete reduction of the silver salt.

The stability of synthesized silver nanoparticle solutions was assessed by

recording the UV-vis spectra at intervals of 1, 15 and 30 days after storage at

ambient temperature. The evolution of UV-VIS spectra is shown in Figure 3.7 (b

and c curves). There was no obvious change in peak position for two weeks,

except for the increase of absorbance. Increase of absorption indicates that

amount of silver nanoparticles increases. The stable position of absorbance peak

indicates that new particles do not aggregate. After the fourth week (curve c) the

fwhm of the spectrum starts to become wider than before, and the position of the

peak has a slight red shift, implying the onset of nanoparticle aggregation. These

spectra demonstrate that the silver nanoparticle colloidal solution can remain

stable for about 1 month.


3.3.3 Crystalline structure

The X-ray diffraction spectrum of the synthesized nanoparticles is shown in

Figure 3.8.

Figure 3.8 XRD spectra of representative nanoparticle concentration (26.95

µg/ml): Peak indices and 2θ positions.

A number of Bragg reflections in the (111), (200), (220) and (311) set of lattice

planes were observed. The high intensity for fcc materials is generally (111)

reflection, which is observed in the samples. This confirmed the lattice structure

to be fcc (face centered cubic). The crystalline size was calculated from the half-

height width of diffraction peaks using the Debye-Scherrer equation. Data are

presented in Table 3.4.

Table 3.4: Particle size derived from XRD spectra.

Particle

concentration

2θ

(degree)

FWHM

(radian)

Particle size

(nm)

Crystal

lattice

26.95 µg/ml 38.31 0.014 10.3 fcc

70.07 µg/ml 38.35 0.013 10.9 fcc


Particle size of around 10 nm was derived from the XRD spectra. These values

represent a rough estimate as the total broadening of the diffraction peak is due to

the sample and the instrument.

Table 3.5: Crystallinity index of silver nanoparticles.

Sample Dp (nm) Dcry (nm) Icry Particle Type

Silver Nanoparticles 8.6 10.3 0.83 Monocrystalline

The crystallinity index (Icry) was calculated by comparision of the particle sizes

determined from the XRD and TEM data. The equation for Icry is described in

Section 3.2.2.4. The data above shows that the crsytallinity index of synthesized

nanoparticles is 0.83, that the silver metal was monocrystalline and fcc phase

structure was well indexed.

To sum up, in the X-ray diffraction pattern of silver nanoparticles four peaks at

2θ values of 38.3º, 44.5º, 64.6º and 77.1º corresponding respectively to (111),

(200), (211) and (300) planes of silver were observed. The results indicate that

the resultant particles are fcc silver nanoparticles with a predicted size of ~ 10

nm. The particle type is described to be monocrystalline from the crystallinity

index. These observations are discussed further in Section 3.4.

3.3.4 Morphology and Particle Size

The electron micrographs of particles synthesized in YM media and in de-ionized

water are presented below.


Figure 3.9 Transmission electron micrographs of silver nanoparticles synthesized

in yeast malt media. Time: 12 h after synthesis.

Figure 3.10 Transmission electron micrographs of silver nanoparticles

synthesized in de-ionized water. Time: 12 h after synthesis.

It was observed that particles synthesized were more or less spherical in shape.

To determine the size distribution, a total of ~ 600 particles (in YM media) and ~

150 particles (in de-ionized water) were measured. Mean particle size was

smaller (8.6 nm) for particles synthesized in YM media compared to 12.6 nm for


particles synthesized in de-ionized water. Also, the particles were well dispersed

in YM media whereas clusters were observed in de-ionized water suggesting

aggregation of nanoparticles. Results are summarized in Figure 3.11 and

Table 3.6.

Figure 3.11 Size distribution of nanoparticles obtained from TEM picture

analysis. a) particles synthesized in YM media and b) in de-ionized water.

A t- Test was performed based on the data provided in Table 3.6 to evaluate the

difference in size of nanoparticles synthesized in the two solutions.

Table 3.6: Data for t Test of size distribution.

Parameter YM media De-ionized water

Mean 8.603 12.630

SD 8.577 14.760

N 627 155

P value and statistical significance:

The two tailed P value is less than 0.0001. By conventional criteria, this

difference is considered to be extremely statistically significant.

It is evident that nanoparticles synthesized in YM media were significantly

smaller in size compared with those synthesized in de-ionized water.


3.4. Discussion

Silver nanoparticles were readily synthesized in the yeast malt media without the

use of any additional stabilizing agents. As mentioned elsewhere, the use of the

reducing agent, NaBH4, in large excess to that of the silver salt, AgNO3,

eliminated the possibility of the presence of any residual silver ions. Additionally,

constituents of the growth media likely stabilized the nanoparticles as no

precipitation was observed after the synthesis process. The synthesis of silver

nanoparticles by chemical reduction of silver salts with a reducing agent has been

widely used in microbial studies (Kim et al., 2007; Xu et al., 2004). Gogoi et al.

(2006) had previously reported the synthesis of silver nanoparticles in LB (Luria

Bertnii) broth in their study on E. coli bacteria. This process circumvents the

stringent conditions often employed for nanoparticle synthesis and also

simultaneously minimizes the ‘artifact effect’ which chemical agents might have

on the test organism. We propose that direct synthesis in growth media is an

efficient way of studying nanoparticle-cell interactions as the particles are smaller

in size and poly-dispersed leading to a more robust contact. The preparation of

uniform nanosized particles with specific requirements in terms of size, shape,

and physical and chemical properties is of great interest (Sondi and Salopek-

Sondi, 2004) and in the present study we were able to achieve the synthesis of

stable and well dispersed silver nanoparticles of fairly uniform size distribution.

UV-Vis absorption spectra have been proved to be quite sensitive to the

formation of silver colloids because silver nanoparticles exhibit an intense

absorption peak due to the surface plasmon (it describes the collective excitation

of conduction electrons in a metal) excitation (Gao et al., 2005). We observed the

spectra of silver colloids of different concentrations in the range of 300 to 700

nm. Well defined plasmon bands around 400 nm (minimum at 390.5 nm to a

maximum at 416.5 nm) were seen. Sols with a single visible extinction band near

400 nm are characteristic of silver particles substantially smaller than the

wavelength of light (Creighton et al., 1979). Besides the pronounced silver

plasmon resonances that appear between 400 and 500 nm, the electronic


transitions involving the Ag+ ion give rise to absorption bands located between

200 and 230 nm, whereas the electronic transitions of metallic Ag0 appear in the

250-330 nm spectral range (Lu et al., 2005).

It is known that the color of metal particles is caused by the sum of the effects of

absorption and scattering of visible light (Kapoor, 1998). According to Mie’s

theory, small spherical nanocrystals should exhibit a single surface plasmon

band, whereas anisotropic particles should exhibit two or three bands, depending

on their shape. Absorption spectra of larger metal colloidal dispersions can

exhibit broad or additional bands in the UV-Vis range due to the excitation of

plasma resonances or quadrupole and higher multipole plasmon excitation (He et

al., 2002). The UV-Vis spectra of the different nanoparticle concentrations in our

study show only one symmetric absorption peak. The good symmetric absorption

peaks with a nearly unchanged width imply that the size of the nanoparticles is

very uniform, which does not depend on the silver concentration (He et al.,

2001).

A comparison of the fwhm values obtained for the different particle solutions

(Table 3.2) confirms the formation of nanoparticles of uniform size distribution.

When the solution system is monodisperse (narrow size distribution) the peak

shape is symmetric and the value of the fwhm is small. When the system is

polydisperse, the peak shape is asymmetric, which suggests that the peak actually

consists of two or more absorption peaks (Brown et al., 2000). We find that the

peak shapes are symmetric, and the corresponding fwhm values ranged from 108

to 77 nm. Those results mean that the size distribution becomes narrower, and the

colloid system is monodisperse. This phenomenon can also be seen in the

following TEM results discussed below.

To confirm the stability of silver nanoparticle solutions, we measured the

absorption spectra of one of the colloid systems at different time intervals. As

shown in Figure 3.7, in the initial three-week period there is no obvious change in

the peak position, except for slight increase in absorbance intensity. The stable

position of the absorbance peak indicates that the particles do not aggregate


(Sileikaite et al., 2006). After the fourth week, the fwhm of the spectrum starts to

become wider than before, and the position of the peak has a slight red shift,

implying the onset of nanoparticle aggregation. These spectra demonstrate that

the silver nanoparticle colloidal solution can remain stable for about 1 month.

The X-ray diffraction pattern of the synthesized nanoparticles (Figure 3.8) shows

diffraction peaks at 2θ = 38.3º, 44.5º, 64.6º and 77.1º, which can be respectively

indexed to (111), (200), (220) and (311) planes of pure silver. This data compares

favourably with the powder diffraction file (PDF) no. 04-0783 pertaining to pure

silver available from the Joint Committee on Powder Diffraction Standards

(JCPDS). The results clearly indicate the presence of silver in the form of highly

crystalline face-centred cubic (fcc) structures of silver nanoparticles (Wang et al.,

2005; Lanje et al., 2010). It may be noted that an additional peak at 2θ = 81.5º

(approximate) corresponding to (222) lattice plane was observed in other studies

(Liu et al., 2007; Pan et al., 2010). In the current study this peak index is missing

as the diffracted intensities were recorded only up to 79.9º 2θ angles.

The particle size calculated from the high intensity (111) peak was 10.3 nm and

the corresponding crystallinity index (Icry) was 0.83. If crystallinity (Icry) is close

to 1, then it is assumed that the crystallite size represents monocrystalline units

whereas a polycrystalline population would have a much larger crystallinity

index (Pan et al., 2010). The particle size estimated was higher than that observed

in electron micrographs. In the Debye-Scherrer equation used to calculate particle

size, the width of the diffraction is taken into account. In general, this broadening

is due to particle size and strain, but for particles less than 100 nm appreciable

broadening of the XRD lines will occur and the total broadening of the diffraction

peaks is due to both the sample and the instrument (Theivasanthi and Alagar,

2011).

Microscopic techniques are employed not only to visualize nanoparticles but can

also generate useful data on the size, size distribution and other measurable

properties (Rabinski and Thomas, 2004). However, it needs to be recognized that

the image analysis of the microscope outputs is as crucial as imaging itself. Only


small amounts of samples can be analyzed by microscopic techniques and this

has an impact on the statistical significance of the results. The average particle

size is a number average and size distribution obtained by image analysis

depends on the number of particles measured. Since there are often fewer larger

particles it is important to count and measure enough particles to obtain good

counting statistics on these size fractions (Tiede et al., 2008).

The analysis of electron micrographs of the synthesized nanoparticles largely

confirmed the results obtained from UV-Vis spectra. The particles exhibit a

smooth, spherical morphology and are well dispersed (Figure 3.9). Size

distribution is narrow ranging from 2.5 nm to >50 nm (Figure 3.11). In the case

of particles synthesized in YM media the mean particle size was 8.6 nm while

particles synthesized in de-ionized water were somewhat larger being 12.6 nm on

an average. This difference was found to be extremely statistically significant.

Clustering of particles was apparently more pronounced in de-ionized water

(Figure 3.10) this may be due to the fact that no additional stabilizing agent was

applied during synthesis whereas in YM media the constituents of the media

likely stabilized the particles in solution.

The only other similar synthesis process described by Gogoi et al. (2006)

reported particle sizes ranging from 2 to 5 nm. Kim et al. (2007) reported

synthesis of highly monodisperse particles with an average diameter of 13.5 nm.

In their study the synthesis was accomplished in triple distilled water without

additional stabilizers. With the use of Daxad 19 (sodium salt of a high molecular

weight naphthalene sulfonate formaldehyde condensate) as stabilizing agent and

ascorbic acid as the reducing agent, Sondi and Salopek-Sondi (2004) reported

mean particle size of 12.3 nm. Apart from the use of stabilizers, size is also

influenced by the choice and concentration of the reducing agent used for

synthesis. The antimicrobial properties of silver nanoparticles are size dependent

(Rai et al., 2009; Panacek et al., 2006) as smaller sized particles possessed larger

surface area to volume ratio. In our study the particle size obtained was similar to


previous reports and could be expected to influence the growth and cellular

activities of the test organism.

The experiments on particle characterization confirmed that the synthesis of

silver nanoparticles in the growth media was successful and had also provided us

with estimates about the particle size and morphology.

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