Harish Kumar, Renu Rani & Rajkumar Salar

Abstract— Rod shaped Nickel nanoparticles of size 20 to 25 nanometer were synthesized in the micellar cavity by reverse micelle method. X-Ray diffraction, TEM and FT-IR spectroscopic techniques were used to characterize Nickel nanoparticles. Antibacterial activity of Nickel nanoparticles was tested on six different bacteria i.e. Escherichia Coli, Lactobacillus, Stephilococcus Aureus, Pseudomonas Aeruginosa and Bacillus Subtitles. Minimum inhibitory concentration (MIC) of Nickel nanoparticles was around 300 ppm for different bacteria. Antibacterial activity of Ni nanoparticles (50 to 1200 ppm) were compared with seven well known antibiotics i.e. Ampicillin (10mcg), Penicillin G (2unit), Streptomycin (25mcg), Gentamicine (30mcg), Tetracycline (30mcg), fluconazole (25mcg) and Clotrimazole (10mcg). A significant decrease in the numbers of the Colony Forming Unit (CFU) has been observed with the increase in concentration of Nickel nanoparticles. It was observed that Nickel nanoparticles, due to their smaller size, have penetrated inside the bacterial cell and believed to have caused damage to the bacterial cell by interacting with phosphorous and sulphur containing compounds such as DNA etc. causing the bacterial cell death. Keywords— Nickel nanoparticles, nanotechnology, antibacterial study, reverse micellar synthesis.

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I. INTRODUCTION Research on nanomaterial is a step towards miniaturization of technology. Miniaturization is a general aim of the technology development that is taking place to produce smaller, faster, lighter and cheaper devices with greater functionality, while using less raw materials and consuming less energy. Research on nanomaterial will contribute significantly towards a sustainable usage of raw material and energy [1]. Properties of nanomaterials are different from their bulk form and are determined by the chemical composition, grain size, atomic structure, crystallographic orientation, coordination number and dimensionality. Nanoparticles due to their smaller size and large surface to volume ratio, exhibit interesting novel properties which include nonlinear optical behavior, increased mechanical strength, enhanced diffusivity, high specific heat, magnetic behavior and electric resistivity, etc. [2]. There are wide range of scientific applications of metal nanoparticles such as in the field of biotechnology [3], sensors [4], medical diagnostics [5], catalysis [6], high performance engineering materials, magnetic recording media, optics and conducting adhesives [7]-[9]. Physical vapor deposition, chemical vapor

Manuscript received November 17, 2010. This work was supported in part by the Dr. K. C. Bhardwaj, Vice Chancellor, Ch. Devi Lal University, Sirsa (India).

Harish Kumar, Assistant Professor, Dept. of Chemistry, Ch. Devi Lal University, Sirsa, Haryana 125055 (INDIA) (Phone: (+91)9466077870; Fax: +(91)01666-248123; e-mail: harimoudgil1@yahoo.co.in).

Renu Rani, Research Scholar, Dept. of Chemistry, Ch. Devi Lal University, Sirsa, Haryana 125055 (INDIA)

Raj Kumar Salar, Associate Professor, Department of Biotechnology, Ch. Devi Lal University, Sirsa, Haryana 125055 (INDIA).

deposition, aerosol processing, Sol-Gel process, reverse micelle method, mechanical alloying/ milling, are some of the commonly used methods by which nanoparticles can be synthesized.

Compare to other methods, the reverse micelle method is one of

the most promising wet chemistry synthesis approach. This method provides a favorable microenvironment for controlling the chemical reaction. As such the reaction rate can be easily controlled, and it is possible to obtain a narrow nanoparticle size distribution [10]. It is also called a bottom-up method for the synthesis of nanomaterials which should restrict the growth of the nuclei, prevents the agglomeration of particles and maintains the size distribution of these materials. This method does not require specialized or expensive equipments as is required by several other physical methods. The reaction takes place in the aqueous cores of the reverse micelles, which are dispersed in an organic solvent and are stabilized by a surfactant. The products are obtained after the reaction is homogeneous. The size of the core of the reverse micelles can also be controlled by changing water to surfactant ratio [11]. Reverse micelle solutions are a transparent, isotropic, thermodynamically stable, water-in-oil type microemulsion in which the aqueous phase is dispersed as nano-sized droplets surrounded by a monolayer of surfactant molecules in a continuous polar organic phase [12]-[13].

Main objective of the present study was to synthesize Nickel

nanoparticles in the micellar cavity by reverse micelle method in a favorable microenvironment with different W/S ratio and to characterize nickel nanoparticles using XRD, TEM and FT-IR techniques. An attempt will be made to explore possible applications of Nickel nanoparticles as antibacterial agent and to compare the antibacterial activity of Nickel nanoparticles at different concentrations with different well known antibiotics. The ultimate objective was to study the interaction between bacteria and Nickel nanoparticles to understand the bacterial inhibition mechanism.

2. MATERIALS AND METHODS

2.1 Materials required for Nickel nanoparticles Synthesis: Microemulsions were prepared from Cyclohexane, Triton X-100 as a surfactant, Hydrazine Hydrate and Tetrahydrofuran (THF) (Sigma-Aldrich, USA), Poly Vinyl Pyrolidone K-90 (PVP) (S. D. Fine Chem. Pvt. Ltd. INDIA), and aqueous solution of Nickel Chloride hexahydrate (SRL Sisco Chem. Pvt. Ltd. Bombay (India). 2.2 Material for Anti-bacterial Study: Beef extract and Agar-Agar Type-1 (Hi-Media Pvt. Ltd., Bombay, India), Peptone (Qualigen Chemicals Pvt. Ltd., India). Microorganisms strains were obtained from Microbial Type Culture Collection (MTCC), Institute of Microbial Technology, Chandigarh. E. Coli (MTCC 294), Pseudomonas Aeruginosa (MTCC 424), Staphylococcus Aureus (MTCC 3160), Lacto Bacillus and Bacillus Subtilis (MTCC 1133), were used in the present investigation as test organisms and seven well known antibiotics i.e. Ampicillin (10mcg), Penicillin G (2unit), Streptomycin (25mcg), Gentamicine (30mcg), Tetracycline (30mcg), fluconazole (25mcg) and Clotrimazole (10mcg) were obtained from Hi-Media Pvt. Ltd. Bombay.

Reverse Micellar Synthesis, Characterization & Antibacterial Study of Nickel Nanoparticles

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2.3 Method of Synthesis of Nickel nanoparticles: The microemulsion was prepared by using 1.0% of PVP, 225ml of Cyclohaxane, 50ml of 0.5M of NiCl4.6H2O solution. The average size of the synthesized Nickel nanoparticles powder was controlled by the W/S ratio. The microemulsion was mixed rapidly with continuous stirring, and after half an hour of equilibration, 10ml of 2.0M Hydrazine hydrate solution was added with continuous stirring which reduced nickel ions (Ni4+) to metallic Nickel inside the reverse micellar core. The reverse micelles were broken by adding THF to microemulsion solution. The Nickel metal nanoparticles thus released were separated and washed with triply distilled water to remove residual PVP in the sample. Same procedure was repeated by using different W/S ratio.

3. RESULTS AND DISCUSSION

It was observed that after reduction of nickel ions by Hydrazine hydrate, Nickel nanoparticles turned brown in colour. The density and bulk density increases with the increase in W/S ratio from 5 to 10 but decreases when W/S ratio is further increased to 15. This is due to the decrease in size of Nickel nanoparticles as W/S ratio increases from 5 to 10. The possible reason for this may be due to the decrease in size of Nickel nanoparticles inside the core of micelle leads to increase in number of Nickel nanoparticles inside the micelle core due to which the mass of Nickel nanoparticles increases and volume decreases and surface to volume ratio increases, due to this density increases. But, with further increase of W/S ratio to 15, the size of Nickel nanoparticles increases, due to which the surface to volume ratio decreases and as a result density decreases. 3.1 X-Ray Diffraction Analysis : X-ray diffraction analysis is a widely used method to find out the mean size and structure of nanoparticles. Ni Kα radiation used for characterization of Nickel nanoparticles during XRD measurement was 1.5419 Å at 298K. The Nickel diffraction peaks appeared at 2θ angles, in the sample prepared in W/S ratio of 5, 10 and 15 were shown in Fig. 1. 100% relative intensity is observed at the angles i.e. 15.42, 15.38 and 15.360 in all the three samples of Ni nanoparticles. XRD data were used to determine the mean size of Ni nanoparticles by using Scherrer equation.

0.9dCos

λβ θ

= (1)

Where, d is the mean diameter of the nanoparticles, λ is the wavelength of X-Ray radiation source, β is the angular full width at half maximum (FWHM) of the X-Ray diffraction peak at the diffraction angle,θ . Size of Nickel nanoparticles calculated using Scherrer equation corresponding to 100% intensity peak at W/S ratio of 5, 10 and 15 was 85, 60 and 43nm respectively. 3.2 Transmission Electron Microscopy: TEM (Model: Hitachi -H-7500, Resolution: 2Å) technique was used to determine the size and shape of Ni nanoparticles. It is clear from the TEM images of Ni nanoparticles (Fig. 2) that the Nickel nanoparticles formed are of different sizes in different W/S ratio. This may be due to nucleation and seedling processes occurring inside the micellar core. The shape of Nickel nanoparticles is rod shape and their average size distribution is 20 - 25nm.

3.3 Fourier Transform Infra-Red (FT-IR) Spectroscopy : The influence of chemical environment, technological and synthetic conditions [14]-[19] on the preparation of Ni nanoparticles and creating of chemical bonds was investigated by FT-IR spectroscopy (Model: Spectrum BX Series-79048). FT-IR spectra of Ni nanoparticles were taken to prove the creation of chemical bonds between atoms in surface groups and molecules. FT-IR spectrum in Figure 3 (a, b and c) gives information about the synthesis conditions of Ni nanoparticles. FT-IR spectrum shown in Figure 3 (a, b and c) is typical for an organic compound. In the wavelength range of 3500-1500cm-1 the bands are characteristic for stretching antisymmetric nas(N-H) and symmetric ns(N-H) mode of vibrations in NH2 and NH groups as well as for bending b(N-H) mode of vibrations of N-H bonds NH2 and NH groups. The bands in the wavelength range 1500-800 cm-1 are characteristics stretching vibrations n(CN) mode of C-N bonds, 1000-650 cm-1 wavelength range are characteristics of bending mode of C-H bonds in alkenes, wavelength range of 3150-3050 cm-1 is the characteristics due to stretching mode of C-H bonds in aromatics, and wavelength range of 3650- 3200 cm-1 are characteristics due to the O-H bonds in O-H groups of alcohols and phenols in surface groups. The bands in the FT-IR spectra of Ni nanoparticles proves the creation of chemical bonds between N-H, C-N, C-H and O-H atoms in different surface groups and molecules.

4. ANTI-BACTERIAL STUDY The antibacterial effect of metal nanoparticles has been

attributed to their small size and high surface to volume ratio, which allows them to attach closely with microbial membranes and is not merely due to the release of metal ions in solution [20]. Metal nanoparticles with bactericidal activity can be immobilized and coated on the surfaces, which may find application in various fields, i.e., medical instruments and devices, water treatment, food processing etc. Metal nanoparticles may be combined with polymers to form composites for better utilization of their antimicrobial activity. Metal nanoparticles are also finding application in various other fields, i.e., catalysis and sensors [21]-[23]. Only a few studies have reported the antibacterial properties of Ni nanoparticles having a significant promise as bactericidal agent [24]. Yoon [25] reported the antibacterial effects of Ag and Ni nanoparticles using single representative strains of E. Coli and B. Subtilis, where the Ni nanoparticles demonstrated superior antibacterial activity compared to the silver nanoparticles. Ni nanoparticles supported on various suitable materials, such as Carbon, Polyurethane foam, polymers and sepiolite have also been effectively used for bactericidal applications [26]-[30]. While various hypotheses have been proposed to explain the mechanism of antimicrobial activity of Ni nanoparticles, it is widely believed that Ni nanoparticles are incorporated in the cell membrane, which causes leakage of intracellular substances and eventually causes cell death. Some of the Ni nanoparticles also penetrate into the cells.

Antibacterial activity of the Nickel nanoparticles was determined, using the agar well diffusion assay method [31]. Approximately, 25ml of molten and cooled nutrient agar media were poured in the sterilized petri dishes. The plates were left over night at room temperature to check for any contamination to appear. The bacterial test organism E. Coli, Lactobacillus, Staphilococcus Aureus, Pseudomonas Aeruginosa and Bacillus Subtilis were grown in nutrient broth for 24hr. A 100µl nutrient broth culture of each bacterial organism was used to prepared bacterial lawns. Agar wells were prepared with the help of a sterilized stainless steel cork borer. The wells in each plate were loaded with 100 µl of different concentration of Ni nanoparticles. Various antibiotics used as positive control for each bacteria to compare the inhibition of growth of bacteria with Ni nanoparticles.

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The plates containing the bacteria and solutions of Nickel nanoparticles were incubated at 30°C. All the tests were repeated in quadruplicates. The antibacterial activity was taken on the basis of diameter of Zone of Inhibition which was measured at cross-angles after 24hr of incubation and the mean of four readings is shown in Table 1. Figure 4, illustrates the inhibition of the bacterial growth on the agar plates by using different concentrations of Ni nanoparticles (A = 50, B = 100, C = 200, D = 400, E = 600, F = 800, G = 1000, and H = 1200). Fig. 5 shows Zone of inhibition of Nickel nanoparticles at different investigated concentrations for different bacteria. The Zone of Inhibition in case of E. Coli was observed at 400ppm, in case of Lacto Bacillus was at 600ppm, in case of Staphilococcus Aureu was at 400ppm, in case of Pseudomonas Aeruginosa, was at 600ppm and in case of B. Subtilis, it was at 600ppm of Ni nanoparticle. On the comparison of the values of Zone of Inhibition with six different well known antibiotics, i.e. Ampicillin (10mcg), Chloramphenicol (25mcg), Penicillin G (1unit), Streptomycin (10mcg), Sulphatriad (300mcg), Tetracycline (25mcg) (Fig. 6), it was found that Nickel nanoparticles are good inhibitor of bacterial growth and shows almost similar antibacterial activity as that of six investigated antibiotics towards E. Coli and B. Subtilis at 100 ppm concentration (Table 2). It is also observed that Zone of Inhibition does not increase significantly on increasing concentration of Ni nanoparticles from 400 to 1200 ppm. So, 400 ppm is the optimum concentration of Ni nanoparticles for inhibiting growth of bacterial test organism. Nickel nanoparticles were dispersed in the autoclaved deionized water by ultrasonication so that soft agglomerates were broken down into smaller nanoparticles. Aqueous dispersions of these nanoparticles at desired concentrations were made. Number of bacteria cell (104 cells/ml) added in each flask containing 50ml of solution was considered closer to the real life situations. Shaking during incubation provided bacteria aeration and homogeneity. Control flasks containing all the initial reaction components except the Nickel nanoparticles showed no antibacterial activity. Bacterial cells grow by a process of binary fission in which one cell doubles in size and splits into halves to produce two identical daughter cells. Bacterial cell growth enhances the turbidity of the liquid nutrient medium, as a result optical absorption increases in the solution Transmission electron microscopy determined the distribution and location of the Nickel nanoparticles, as well as the change in morphology of the bacteria after treatment with Ni nanoparticles. Nickel has been found adhered to the bacterial cells. Bacteria have different membrane structures on the basis of which these are classified as Gram-negative or Gram-positive. The structural difference lies in the organization of peptidoglycan, which is the key component of membrane structure. Gram-negative bacteria exhibit a thin layer of peptidiglycan ( ≈ 2-3nm) between the cytoplasmic membrane and the outer cell wall. Outer membrane of E. Coli cells is predominantly constructed from tightly packed lipopolysacharide (LPS) molecules, which provide an effective permeability barrier [32]. The overall charge of bacterial cell at biological pH values is negative because of excess number of carboxylic groups, which upon dissociation make the cell surface negative. The opposite charges of bacteria and nanoparticles are attributed for their adhesion and bioactivity due to the electrostatic forces. It is logical to state that binding of the nanoparticles to the bacteria depends on the surface area available for interaction. Nanoparticles have large surface area available for interactions which enhances bactericidal effect than the large sized particles; hence they impart cytotoxicity to the microorganisms [33]. The mechanism by which the nanoparticles penetrates into bacteria is not understood completely, but studies suggest that when E. Coli was treated with Nickel, changes took place in its membrane morphology

and produced a significant increase in its permeability affecting proper transport through the plasma membrane, leaving the bacterial cells incapable of properly regulating transport through the plasma membrane, resulting into cell death. It was observed that Nickel nanoparticles have penetrated inside the bacteria and believed have caused damage by interacting with phosphorous and sulphur containing compounds such as DNA [34]. Nickel tends to have a high affinity to react with such compounds. In our study, it is considered that DNA may have lost its replication ability and cellular proteins become inactive after treatment with Nickel nanoparticles. Another reason would be the release of Ni ions from nanoparticles, which will have an additional contribution to the bactericidal efficiancy of Nickel nanoparticles. Heavy metals are toxic and reactive with proteins therefore, they bind protein molecules; as a result cellular metabolism is inhibited causing death of microorganism. It is believed that Nickel nanoparticles after penetration into bacteria inactive their enzymes, generate hydrogen peroxide and cause bacterial cell death [35]. Experimental observations have explained significantly the antibacterial behaviour of Nickel nanoparticles. It is observed in the micrograph, that bacterial cells shows critical changes and damage in the membrane structure. When E. Coli was treated with highly reactive metal oxide nanoparticles, an inhibitory effect has taken place [36]. Copper nanoparticles after adherence to the surface of the cell membrane distributed its respiration as Ni2+ interact with enzymes of the respiratory chains of bacteria [37]. It has been observed in our study that a degradation of the membrane structure of E. Coli has taken place. Metal depletion caused formation of irregular shaped pits in the outer membrane of bacteria which is caused by progressive release of LPS molecules and membrane proteins [38]. In addition, it is believed that Nickel binds to functional groups of proteins, resulting in protein denaturation [39]. As observed, bacterial growth log phase has a delayed trend at lower concentrations of Nickel nanoparticles loadings. Complete bacterial inhibition depends upon the concentrations of Nickel nanoparticles and on the number of bacterial cells. It reflects that Nickel nanoparticles have significant biocide effect in reducing bacterial growth for practical applications.

5. CONCLUSION

Average size of the Nickel nanoparticles determined from X-ray diffraction and TEM technique was found to be 20 to 25 nm. Nickel nanoparticles synthesized were of simple cubic structure confirmed by X-ray diffraction study. The FT-IR absorption bands are characteristic for stretching symmetric and antisymmetric vibrations, as well as for banding vibrations of chemical bonds in surface atom groups and molecules such as N-H in NH2 and NH groups, C-N, O-H in OH groups and H2O molecules. Minimum inhibitory concentration of Nickel nanoparticles for E. Coli was observed at 400ppm, in case of Lacto Bacillus was at 600ppm, in case of Staphilococcus Aureu was at 400ppm, in case of Pseudomonas Aeruginosa, was at 600ppm and in case of B. Subtilis, it was at 600ppm of Ni nanoparticle. Further, antibacterial property of Nickel nanoparticles (50 to 1200 ppm) were compared with seven well known antibiotics i.e. Ampicillin (10mcg), Penicillin G (2unit), Streptomycin (25mcg), Gentamicine (30mcg), Tetracycline (30mcg), fluconazole (25mcg) and Clotrimazole (10mcg) and it was observed that Nickel nanoparticles shows almost similar Zone of Inhibition as compared to the seven investigated antibiotics. 100µl is the optimum concentration of Nickel nanoparticles for inhibiting growth of bacterial test organism. The numbers of CFU have been observed to reduce significantly with the increasing Nickel nanoparticles loadings. It was observed that Nickel nanoparticles have penetrated inside the bacteria and believed to have caused damage by interacting with phosphorous and sulphur containing compounds such as DNA and cause bacterial cell death.

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6. FUTURE ASPECTS

Experimental investigations have shown encouraging results about the activity of different drugs and antimicrobial formulation in the form of Nickel nanoparticles. Nickel is nontoxic, safe inorganic antibacterial agent and has been described as being oligodynamic that is its ions are capable of causing a bacterostatic (growth inhibition) or even a bactericidal (antibacterial) impact. Therefore, it has the ability to exert a bactericidal effect at a very low concentration. It has a significant potential for a wide range of biological applications such as antibacterial agents for antibiotic resistant bacteria, preventing infections, healing wounds and anti-inflammatory. Therefore, Nickel ions being antibacterial component, can be employed in formulation of dental resin composites, bone cement, ion exchange fibres, coating for devices, to prevent bacterial growth in drinking water, sterilization, burn care etc. References [1] G. Schmid, M. Decker, H. Emst and H. Fuchs. Miniaturization and Material properties, Europaische Akademie’s, 2003. [2] H. Gleiter, Acta Materialia vol. 48, no. 1, 2000. [3] C. A. Mirkin, R. L. Letsinger, R. C. Mucic, and J. J. Storhoff, Nature, vol. 382, p. 607, 1996. [4] T. A. Taton, C. A. Mirkin and R. L. Letsinger, Science, vol. 289, p. 1757, 2000. [5] J. J. Storhoff, R. Elghanian, R. C. Mucic, C. A. Mirkin, and R. L. Letsinger, J. Am. Chem. Soc., vol. 120, p. 1959, 1998. [6] L. Guezi, D. Horvath, Z. Paszti, L.Joth, Z. E. Norvath, A. Karacs, and G. Peta, J. Phys. Chem., vol. B104, no. 14, p. 3183, 2000. [7] R. Dagani, Chem. Eng. News, vol. 77, p. 54, 1999. [8] J. F. Hamilton and R. C. Baetzold, Science, vol. 205, p. 1213, 1979. [9] G. Schmid, Chem. Rev., vol. 92, p. 1709, 1992. [10] Dong-Sik Bae, Eun-Jungkim, Jae-Hee Bang, Sang-Woo Kim, Kyong-Sop Han, Jong-Kyulee, Byung-Ik Kim, and James H. Adair, J. Met. Mater.-Int., vol. 4, pp. 291-294, 2005. [11] Ashok K. Ganguli, Sonalika Vaidya, and Tokeer Ahmad, Bull. Mater. Sci., Indian Academy of Sci., vol. 31, no. 3 pp. 415-419, June 2008. [12] D. S. Bae, S. W. Park , K. S. Han, and J. H. Adair, Met. Mater.- Int., vol. 7, p. 399, 2001. [13] M. P. Pileni, Structure and Reactivity in Reverse Micelles, Elsevier, Amsterdam 1989. [14] K. Nakamoto, Infrared and Raman Spectra in Inorganic and Coordination Compounds, 3rd Ed., John Wiley and Sons, New York, 1965. [15] I. Markova- Deneva, K. Alexandrova, G. Ivanova, I. Dragieva, “IR Spectroscopy Investigation of Metal Amorphous Nanoparticles”, J. Univ. Chem. Technol. Met. (Sofia), vol. 37, no. 4, pp. 19-26, 2002. [16] Markova- Deneva, “IR Spectroscopy as a method for investigation of nanostructures surface state”, Nanoscience and Nanotechnology Heron Press Science Series, vol. 7, pp. 269- 273, 2006. [17] I. Markova, I. Deneva, I. Dragieva, K. Alexadrova, “IR Spectroscopy investigations of nanoparticles and nanowire obtained by borohydride reduction method”, Nanoscience and Nanotechnology, Heron press Science Series, vol. 7, 2006, pp. 283-286,. [18] I. Markova, I. Dragierva, K. Alexandrova, “FT-IR Spectroscopy investigations of nanostructured materials”, 9th National Workshop Nanoscience and Nanotechnology, vol. 10, no. B9, Nov. 28-30, Sofia Poster session (topic B), (2007). [19] I. Denev, I. Markova, I. Dragieva, K. Alexandrova, “ FT-IR Spectroscopic investigations of nanosized materials”, International Scientific Conference, 60th Anniversary of the Department of Physical Chemistry, UCTM-Sofia, III. P04, Nov.

23, (2007), Sect. III “Metallurgy, Materials Science, Physico- Mathematical and Technical Sciences”, Poster session. [20] J. R . Morones, J. L.Elechiguerra, A. Camacho , K. Holt, J. B. Kouri, J. T. Ramirez , et al., The batericidal effect of silver nano particles, Nanotechnology, vol. 16, pp. 2346-2353, 2005. [21] A. Vaseashta, D. Dimova-Malinovska, Nanostructured and nanoscale devices, sensors and detectors, Sci. Technol. Adv. Mater., vol. 6, pp. 312-318, 2005. [22] E. Comini, Metal oxide nano- crystals for gas sensing, Anal. Chim. Acta., vol. 568, pp. 28-40, 2006. [23] A. Raveh, I. Zukerman, R. Shneck, R. Avni, I. Fried, Thermal stability of nano structured superhard coating: A Review, Surf Coat Technol., vol. 201, pp. 6136-6142, 2007. [24] N. Cioffi, et al., Copper nanoparticles/polymer composites with antifungal and bacteriostatic properties. Chem. Mater., vol. 17, pp. 5255-5262, 2005. [25] K. Yoon, J. H. Byeon, J. Park, J. Hwang, Susceptibility constants of Escherichia coli and Bacillus Subtilis to silver and copper nano particles, Sci. Total Environ., vol. 373, pp. 572-575, 2007. [26] V. Siva Kumar, B. M. Nagaraja, V. Shashikala, A. H. Padmasri, S. S. Madhavendra, B. D. Raju, et al., Highly efficient Ag/C Catalyst prepared by electro-chemical deposition method in controlling microorganisms in Water, J. Mol. Catal, A. Chem., vol. 223, pp. 313-319, 2004. [27] P. Jain, T. Pradeep, Potential of silver nanoparticles-coated polyurethane foam as an antibacterial water filter, Biotechnol., Bioeng., vol. 90, pp. 59-63, 2005. [28] Z. Li, D.Lee, X. Sheng, R. E. Cohen, M. F. Rubner, Two-level antibacterial coating with both release-killing contact-killing capabilities, Langmuir, vol. 22, pp. 9820-9823, 2006. [29] W. K. Son, J. H. Youk, W. H. Park, Antimicrobial cellulose acetate nanofibres containing silver nanoparticles, Carbohydr. Polym., vol. 65, pp. 430-434, 2006. [30] A. Esteban-Cubillo, C. Pecharroman, E. Aguilar, J. Santaren, J. S. Moya, Antibacterial activity of Copper monodispersed nanoparticles in to sepiolite, J. Mater. Sci., vol. 41, pp. 5208- 5212, 2006. [31] Raj K. Salal, and Suchitra, African J. of Microbiology Research, vol. 3, pp. 097-100, March 2009. [32] Sondi I. and Sondi B. S., J. Coll. Interf. Sci., vol. 275, p. 177, 2004. [33] C. Baker and A. Pradhan, J. Nanosci. Nanotechn., vol. 5, no. 1, 2004. [34] J. R. Morones and J. L Elechiguerra., Nanotechnology, vol. 16, p. 2346, 2005. [35] M. Kokkoris and C. C.Trapalis, Nuclear Instruments and Methods in Physics Research B, vol. 188, p. 67, 2002. [36] P. K. Stoimenov and R. L.Klinger, Langmuir, vol. 18, p. 6679, 2002. [37] K. B. Holt and A. J. Bard, Biotechnology, vol. 44, p. 13214, 2005. [38] N. A. Amro and L. P. Kotra, Langmuir, vol. 16, p. 2789, 2000. [39] J. A. Spadaro and T. J. Berger, Antimicrobial Agents and Chemotherapy, vol. 6, p. 637, 1974.

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Fig. 1 (a.) XRD image of Nickel nanoparticles formed in W/S ratio 5.

Fig. 1 (b.) XRD image of Nickel nanoparticles formed in W/S ratio 10.

Fig. 1 (c.) XRD image of Nickel nanoparticles formed in W/S ratio 15.

Fig. 2 (a) TEM images of rod shape Nickel nanoparticles formed in W/S ratio 5.

Fig. 2 (b) TEM images of rod shape Nickel nanoparticles formed in W/S ratio 10.

Fig. 2 (c) TEM images of rod shape Nickel nanoparticles formed in W/S ratio 15.

Fig. 3 (a) FT-IR Absorption spectra of Nickel nanoparticles synthesized by reverse micelle method (W/S = 5).

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Fig.3 (b) FT-IR Absorption spectra of Nickel nanoparticles synthesized by reverse micelle method (W/S = 10).

Fig.3 (c) FT-IR Absorption spectra of Nickel nanoparticles synthesized by reverse micelle method (W/S = 15).

Fig. 4 Images of Agar plates containing different concentration of Nickel nanoparticles in E. coli (A, B, C, D, E, F, G, and H) shows various concentrations of nickel nanoparticle solution.

Fig. 4 Images of Agar plates containing different concentration of Nickel nanoparticles in Lactobacillus (A, B, C, D, E, F, G, and H) shows various concentrations of nickel nanoparticle solution.

Fig. 4 Images of Agar plates containing different concentration of Nickel nanoparticles in Staphylococcus Aureus (A, B, C, D, E, F, G and H) shows various concentrations of nickel nanoparticle solution.

Fig.4 Images of Agar plates containing different concentration of Nickel nanoparticles in Pseudomonas Aeruginosa (A, B, C, D, E, F, G and H) shows various concentrations of nickel nanoparticle solution.

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Fig. 4 Images of Agar plates containing different concentration of Nickel nanoparticles in Bacillus Subtilis (A, B, C, D, E, F, G and H) shows various concentrations of nickel nanoparticle solution.

Fig. 5. Zone of inhibition shown by Nickel nanoparticles at different concentration in presence of bacterial culture.

Fig. 6. Zone of inhibition in mm shown by different standard antibiotics in presence of different bacterial culture.

Harish Kumar Allahabad, 30th Sept. 1975. M.Sc. Physical Chemistry, Ph.D. in Electrochemistry, Maharashi Dayanand University, Rohtak, Haryana (India) 2005. Nanotechnology, Electrochemistry, Biosensors and thermodynamics. Dr. Harish has more than eight years of teaching and research experience with more than 20 publications in his credit. ASSISTANT PROFESSOR, Dept. of Chemistry, Ch. Devi Lal University, Sirsa (India) Published three books. Text of Physical Chemistry (New Delhi : PHI Learning Pvt. Ltd., New Delhi, 2010 ). Dr. Kumar is member of different educational societies and member of editorial board of Asian Journal of Experimental chemistry, India.

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Penicillin‐ G

Gentamicin

Advances in Control, Chemical Engineering, Civil Engineering and Mechanical Engineering

ISBN: 978-960-474-251-6 94