ZnO Nanoparticles: Synthesis and Adsorption Study

doi:10.4236/ns.2009.12016

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Vol.1, No.2, 129-135 (2009) Natural Science

http://dx.doi.org/10.4236/ns.2009.12016

ZnO Nanoparticles: Synthesis and Adsorption Study

K. Prasad1, Anal K. Jha2

1University Department of Physics, T.M. Bhagalpur University, Bhagalpur - 812 007, India; *[email protected]

2University Department of Chemistry, T.M. Bhagalpur University, Bhagalpur - 812 007, India

Received 21 July 2009; revised 27 July 2009; accepted 30 July 2009.

ABSTRACT

A low-cost, green and reproducible probiotic

microbe (Lactobacillus sporogens) mediated

biosynthesis of ZnO nanoparticles is reported.

The synthesis is performed akin to room tem-

perature in five replicate samples. X-ray and

transmission electron microscopy analyses are

performed to ascertain the formation of ZnO

nanoparticles. Rietveld analysis to the X-ray

data indicated that ZnO nanoparticles have

hexagonal unit cell structure. Individual

nanoparticles having the size of 5-15 nm are

found. A possible involved mechanism for the

synthesis of ZnO nanoparticles has been pro-

posed. The H2S adsorption characteristic of ZnO

nanoparticles has also been assayed.

Keywords: ZnO Nanoparticle; Biosynthesis;

Nanobiotechnology; Eco-friendly; H2S Adsorption

1. INTRODUCTION

Nature by dint of its diversity provides exponential pos-

sibilities in terms of endearing adaptability of its con-

stituent cohorts. Both bacteria and fungi make such an

exciting category of microorganisms having naturally

bestowed property of reducing/oxidizing metal ions into

metallic/oxide nanoparticles thereby functioning as

‘mini’ nano-factories. [1,2] It is indeed their chemical

constitutions (or metabolic status) which provide them

strength to withstand such environmentally diverse

habitats. The non-pathogenic, gram positive, mesophilic

facultative anaerobe Lactobacillus, commonly used for

curdling of milk forms part of the beneficial community

of microbes present in the human intestinal tract.

Zinc oxide (ZnO) is considered to be a technologically

prodigious material having a wide spectrum of applica-

tions such as that of a semiconductor (Eg = 3.37 eV),

magnetic material, electroluminescent material, UV-abs-

orber, piezoelectric sensor and actuator, nanostructure

varistor, field emission displaying material, thermoelec-

tric material, gas sensor, constituent of cosmetics etc.

[3-9] There are several synthesis procedures for the

preparation of ultrafine oxide nanoparticles such as sol-

gel, hydrothermal, solvothermal, flame combustion,

emulsion precipitation, fungus mediated biosynthesis,

etc. [10-16] Each method has its own merits and demer-

its. They are time consuming, capital intensive and re-

quire trained manpower. Besides, the development of

eco-friendly, 'green' synthesis protocols is in line with

the recent RoHS and WEEE legislation stipulated by the

EU. Therefore, an urge to develop green synthesis pro-

tocols which goes in consonance with the above men-

tioned stipulations is need of the hour. Microbes exhibit

a natural capability to adapt to changes in their environ-

ment. Recent research devoted towards the study of in-

teraction between inorganic substances and biological

systems has highlighted its potential application for the

production of nanomaterials with interesting techno-

logical properties. [1,2,17-20] Numerous recent publica-

tions have highlighted the potential for microbes, par-

ticularly bacteria (including thermophilies) and fungi, to

synthesize metallic and/or oxide nanoparticles. [21-35]

The facultative nature of Lactobacilli, offers the poten-

tial to produce nanoparticles under both oxidizing and

reducing conditions. [2,18,35]

No work to the best of author’s acquaintance has so

far been reported regarding the synthesis of ZnO nano-

particles employing Lactobacilli. Lactobacilli strain,

cultured from spores an effort has been taken for synthe-

sizing ZnO nanoparticles (ZnO NPs) in the present work.

We have tried to explore a cost effective, green and read-

ily reproducible approach for the purpose of scaling up

and subsequent downstream processing. An effort to

understand the nano-transformation mechanism of bio-

synthesis has also been made. It is well established that

Hydrogen sulfide (H2S) is a colorless, corrosive and

highly toxic gas, a low concentration of which in air,

brings smell of rotten eggs and it substantially contrib-

utes towards air pollution. [36,37] The potential of ZnO

NPs towards H2S adsorption has also been assayed in the

present study.

K. Prasad et al. / Natural Science 1 (2009) 129-135

130

2. MATERIALS AND METHODS

2.1. Biosynthesis of ZnO Nanoparticles

Pharmaceutical grade Lactic acid Bacillus spore tablets

(SporeLac DS, Sanyko Pharmaceuticals, Japan) were

procured and two tablets were dissolved in 50 mL sterile

distilled water containing standard carbon and nitrogen

source. As per specification, each tablet was capable of

producing 120 million spores of the bacterium. The cul-

ture solution was allowed to incubate on room tempera-

ture overnight. Next day, the presence of Lactobacillus

was confirmed under an optical microscope. The pH of

this source culture solution was observed to be equal to 3.

Now, 10 mL of this source culture was doubled in vol-

ume by mixing equal volume of sterile distilled water

containing nutrients in five different hard glass test tubes.

In yet another tube instead of adding the source culture

solution, sterile distilled water containing nutrients was

pooled and this was treated as control. All these culture

tubes were gently heated on a steam bath and were al-

lowed to incubate overnight in laboratory ambience for

another 24 hours on orbital shaker. Next day, the pH was

taken and found to be in the range of 4-5 in case of cul-

ture solution and 7 in case of control. Small quantity of

NaHCO3 was added in culture solution until it attains pH

6. It was brought to this pH as a lower value delays the

process of transformation. [2] Similarly, a small volume

of distilled water along with carbon and nitrogen source

and NaHCO3 were pipetted in the control tube and the

pH = 8.5 were recorded. Analytical reagent grade Zinc

Chloride (ZnCl2) was taken into use for preparing a so-

lution of 0.25(M) strength at room temperature. Control

solution was prepared by adding 100 mL sterile distilled

water, carbon and nitrogen containing nutrients and the

mild base in known quantitative ratio (5:1:1). To each of

these tubes, 20 mL of Zinc Chloride solution was added.

The pH of the control tube was noted to be 8-9 in 5 dif-

ferent set of experiments. Culture solution containing

tubes including control tube were heated on the steam

bath up to 80°C for 5 to 10 minutes. An appearance of

starch like haziness in solution and white deposition at

the bottom of the tube was perceived as an indication of

commencement of transformation. No such deposition or

haziness was observed in control tube. The tubes were

allowed to incubate in the laboratory ambience for an-

other 9 hours, after which distinctly markable coalescent

white clusters deposited at the bottom of all the tubes

except in control. A remarkable change in pH was ob-

served at this stage (6.0 to 7.5) excluding control (8 to 9).

The chemical reactions which proceed in the culture

medium may be as follows:

(Lact ate) (Pyruvate)

).(.

336126COOHOHCHCHCOOHCCHOHC 

(Glu cose)

 33HCONaNaHCO

23 COOHHCO  

  ClZnZnCl 2

2)(2 OHZnOHZn 

OHZnOOHZn 22

)(

2.2. Characterization

The formation of ZnO NPs was checked by X-ray dif-

fraction (XRD) technique using an X-ray diffractometer

(XPERT-PRO, Pan Analytical) with CuK radiation ( =

1.5406Å) over a wide range of Bragg angles (10  2





80). The XY (2θ vs. intensity) data obtained from this

experiment were plotted with the WinPLOTR program

and the angular positions of the peaks were obtained

with the same program. [38] The dimensions of the unit

cell, hkl values and space group of ZnO NPs were ob-

tained using the DICVOL program in the FullProf 2000

software package and then refinement was carried out

through the profile matching routine of FullProf. [39]

The Bragg peaks were modeled with pseudo-Voigt func-

tion and the background was estimated by linear inter-

polation between selected background points. The crys-

tallite size (D) and the lattice strain of ZnO NPs were

estimated by analyzing the broadening of X-ray diffrac-

tion peaks, using Williamson-Hall approach. [40]











sin)/(2)/(cos 



DK (1)

where



is diffraction peak width at half intensity

(FWHM) and





is the lattice strain and K is the

Scherrer constant (0.89). The term Kλ/D represents the

Scherrer particle size distribution. TEM micrograph of

ZnO NPs was obtained using Hitachi H-7500 transmis-

sion electron microscope. The specimen was suspended

in distilled water, dispersed ultrasonically to separate

individual particles, and two drops of the suspension

deposited onto holey-carbon coated copper grids.

3. RESULTS

3.1. Structural and Microstructural Studies

Rietveld refinements on the X-ray (XRD) data were

done on ZnO NPs, selecting the space group P6/mmm.

Figure 1 depicts the observed, calculated and difference

XRD profiles for ZnO NPs after final cycle of refine-

ment. It can be seen that the profiles for observed and

calculated one are perfectly matching. The value of χ2

comes out to be equal to 3.16, which may be considered

to be very good for estimations. The profile fitting pro-

cedure adopted was minimizing the χ2 function. [41] The

XRD analyses indicated that ZnO NPs has a hexagonal

unit cell. The crystal data and refinement factors of ZnO

K. Prasad et al. / Natural Science 1 (2009) 129-135

131

Figure 1. Rietveld refined pattern of ZnO NPs in the space group P6/mmm. Symbols repre-

sent the observed data points and the solid lines their Rietveld fit. Inset: Williamson-Hall plot

for ZnO NPs.

Table 1. The crystal data and refinement factors of ZnO NPs obtained from X-ray powder diffraction data.

Parameters Results Description of parameters

Crystal System

Space group

a (Å)

b (Å)

c (Å)

α (°)

β (°)

γ (°)

V (Å3)

Rwp

Rexp

χ2

Hexagonal

P6/mmm

3.2524

5.2120

90.000

120.000

47.7463

25.2

23.8

13.4

0.175E-3

0.133E-3

3.16

0.6844

1.9059

1.776

Rp (profile factor) = 100[Σ|yi-yic|/Σ|yi|], where yi is the observed intensity and yic is the

calculated intensity at the ith step.

Rwp (weighted profile factor) = 100[Σωi|yi-yic|2/Σωi(yi)2]1/2, where 2

/1 ii



 and 2



is variance of the observation.

Rexp (expected weighted profile factor) = 100[(n-p)/Σωi(yi)2]1/2, where n and p are the

number of profile points and refined parameters, respectively.

RB (Bragg factor) = 100[Σ|Iobs-Icalc|/Σ|Iobs|], where Iobs is the observed integrated inten-

sity and Ical c is the calculated integrated intensity.

RF (crystallographic RF factor) = 100[Σ|Fobs-Fcalc|/Σ|Fobs|], where F is the structure

factor, F = √(I/L), where L is Lorentz polarization factor.

χ2 = Σωi(yi-yic)2.

d (Durbin–Watson statistics) = Σ{[ωi(yi-yic)-ωi-1(yi-1-yic-1)]2}/Σ[ωi(yi-yic)]2.

QD = expected d.

S (goodness of fit) = (Rwp/Rexp).

NPs obtained from XRD data are depicted in Table 1.

The lattice parameter as obtained for ZnO NPs is in good

agreement with the literature report (PCPDF No.

#89-0510). Inset Figure 1 illustrates the William-

son-Hall plot for ZnO NPs. A linear least square fitting



cosθ–sinθ data yielded the values of average crys-

tallite size and lattice strain respectively to be 11 nm and

0.0035. The low value of lattice strain might be due to

the fact that the procedure adopted in the synthesis of

nanoparticles is natural (biosynthetic) one.

K. Prasad et al. / Natural Science 1 (2009) 129-135

132

Figure 2. TEM photograph of ZnO NPs. Inset: ZnO NPs.

Figure 2 shows the TEM micrograph of ZnO NPs

(inset Figure 2) being formed using Lactobacillus strain.

The micrograph clearly illustrates the nanoparticles with

tubules and other irregular forms having the sizes of

5-15 nm. The measurement of size was carried along the

diameter of the particles. The difference in particle size

is possibly due to the fact that the nanoparticles are be-

ing formed at different times. It is found that the size of

the ZnO NPs estimated using TEM analysis to be in

fairly good agreement with the size estimated by the

Williamson-Hall approach.

3.2. Adsorption Study

Figure 3 shows the experimental setup to assess the ad-

sorption capacity of synthesized ZnO NPs as well as

bulk ZnO. Freshly prepared H2S was allowed to pass

through the equal quantities of bulk ZnO and ZnO NPs

(5 gm each) for a fixed span of time (30 min.) and flow

of gas was suitably regulated. The degree of absorption

was assessed directly through the change in colour of

lead acetate solution (from clear solution to black). It

was observed that the presence of bulk ZnO blackens the

solution (due to formation of lead sulfide) within 5 min-

utes, while ZnO NPs does the same in 25 min. The ex-

periment was pursued as five replicates and each gave

approximately the same result. This happens due to the

fact that nanoparticles have large surface to volume ratio

and hence a high surface activity and these features

might have led to better degree of adsorption of H2S in

comparison to its bulk counterpart. H2S absorption by

ZnO proceeds according to the reaction: ZnO + H2S →

ZnS + H2O that results into formation of inert Zinc sul-

fide.

4. DISCUSSION

Lactobacilli cells are prokaryotes in terms of cellular

organization. They are gram positive (a thick pepti-

doglycan cell wall) bacteria showing facultative anaero-

bic properties, which probably make them suitable can-

didate microorganism for biosynthesis of metal as well

as oxide nanoparticle. Like most of the bacteria, they

have a negative electro-kinetic potential; which readily

attracts the cations and this step probably acts as a trig-

ger of the procedure of biosynthesis. Earlier, such a pos-

sibility of biosorption and bioreduction had been re-

ported in case of silver iodide by the Lactobacillus sp.

A09*. [42] The mesophilic, non-pathogenic and faculta-

tively anaerobic microbe like Lactobacillus has robust

metabolic capabilities. Addition of simple carbohydrates

into the culture medium tends to lower the value of oxi-

dation-reduction potential (or the Eh value). The oxida-

tion-reduction potential expresses the quantitative char-

acter of degree of aerobiosis having a designated unit

expressed as rH2 (the negative logarithm of the partial

pressure of gaseous hydrogen). By controlling rH2 of the

nutrient medium, conditions can be engineered for the

growth of anaerobes in the presence of oxygen by low-

ering the rH2, and also by cultivating the aerobes in an-

aerobic conditions by increasing the rH2 of the medium.

Figure 3. Experimental set up to study adsorption of H2S by ZnO NPs.

K. Prasad et al. / Natural Science 1 (2009) 129-135

133

Zn(OH)

ZnO + H

(In culture solution )

pH - Dependent

membrane bound

Oxido-reductases

[O] [O]

(Lactobacillus Cell)

Partial pressure of gaseous H

(r-H

)i.e. Eh - dependent upon

available carbon source

(in Culture Solution)

Low pH

Oxidase activity

Low r-H

i.e. HighEh

High Oxidation potential

(In culture solution )

Figure 4. Schematic showing the mechanism for the biosynthesis of ZnO NPs.

Composition of nutrient media, therefore; plays a piv-

otal role in biosynthesis of metallic and/or oxide

nanoparticles which is done in the present investigation.

Energy yielding material – suitable carbohydrate (which

controls the value of rH2), the ionic status of the medium

pH and overall oxidation-reduction potential (Eh) of the

culture medium, all these factors cumulatively negotiate

the synthesis of ZnO nanoparticles in the presence of

Lactobacillus strain. Taking use of the above mentioned

facts, our group had earlier reported synthesis of metallic

cadmium [18], silver [17,34,43] as well as antimony

oxide [1,44] and titanium dioxide. [2] A mildly acidic

pH also activates the membrane bound oxidoreductases

and makes the requisite ambience for an oxide nanopar-

ticle synthesis as illustrated in Figure 4. Therefore, com-

pared to other techniques, the present procedure is less

expensive more reproducible, emphatically non-toxic

and a truly green approach.

A large quantity of hydrogen sulfide is liberated in gas

and petroleum industries and has been considered as a

major pollutant. Besides, according to the international

environmental regulations, H2S contained in the acid

gases should be effectively removed before its release to

atmosphere. Its characteristic odor could easily be per-

ceived in a dilution of 0.002 mg /L in ambient air. Intake

of higher concentrations, could lead to the collapse from

respiratory failure. For the purpose of protection, the

concentration should be reduced to less than 15 ppm. [45]

Pollution of underground aquifers has been a prevalent

problem in the areas adjoining oil and gas reserves,

which miserably affects the health of nearby inhabitants.

Use of ZnO NPs produced using present green and low

cost protocol based devices could prove to be an effec-

tive step towards mitigation of the menace.

5. CONCLUSIONS

The present biosynthesis method is a green low cost ap-

proach, capable of producing ZnO NPs nearby room

temperature. The synthesis of ZnO NPs might have re-

sulted due to variation in the level of rH2 or pH, which

activates the pH sensitive oxido-reductases. ZnO

nanoparticles could be effective in controlling the pollu-

tion generated due to H2S in air as well as underground

aquifers. However, bright possibility exists with regard

to the development of different products/devices in order

to get rid of the menace of different forms of air and/or

water pollution such as face masks, water filters,

de-odorizing cakes and screens. Cosmetic industries can

bank upon this product in order to synthesize sunscreen

lotions etc., which would be done in the immediate fu-

ture.

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