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Biological synthesis of gold nanocubes from Bacillus licheniformis
Bioresource Technology 100 (2009) 5356–5358
Contents lists available at ScienceDirect
Bioresource Technology journal homepage: www.elsevier.com/locate/biortech
Short Communication
Biological synthesis of gold nanocubes from Bacillus licheniformis Kalimuthu Kalishwaralal, Venkataraman Deepak, Sureshbabu Ram Kumar Pandian, Sangiliyandi Gurunathan * Department of Biotechnology, Kalasalingam University, Anand Nagar, Krishnankoil-626190, Tamil Nadu, India
a r t i c l e
i n f o
Article history: Received 4 February 2009 Received in revised form 21 May 2009 Accepted 21 May 2009 Available online 1 July 2009 Keywords: Gold nanocubes Bacillus licheniformis Enzyme Eco-friendly
a b s t r a c t Microorganisms play an important role in the eco-friendly synthesis of metal nanoparticles. This study illustrates the synthesis of gold nanocubes using the bacterium Bacillus licheniformis after 48 h of incubation at room temperature. The morphology of the samples was analyzed using scanning electron microscopy (SEM) and the particles formed were characterized to be nanocubes. The size of gold nanocubes in aqueous solution has been calculated using UV–Vis spectroscopy, XRD and SEM measurements. The nanoparticles are found to be polydisperse nanocubes in the size range 10–100 nm. Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction The area of nanotechnology encompasses the synthesis of nanoscale materials, the understanding and the utilization of their physicochemical and optoelectronic properties, and the organization of nanoscale structures into predefined superstructures. Thus, nanotechnology promises to play an increasingly important role in many key technologies of the new millennium (Schmid, 1994). Usually the chemical methods followed are carried out under very high temperatures to form gold nanoparticles and also result in environmental pollution due to toxicity of the reagents (reducing agents like borohydrates and acetylene) used (Wang et al., 2007). Consequently, researchers in the field of nanoparticle preparation have been looking at biological systems (Merroun et al., 2007) such as those that allow a commercially viable and environmentally clean synthesis of highly stabilized gold particles (Mukherjee et al., 2008). Beveridge and Murray (1980) first demonstrated that the exposure of Bacillus subtilis treated with gold chloride (AuCl4) resulted in the synthesis of gold nanoparticles. Biosynthesis of gold nanoparticles has been carried out using Rhodopseudomonas capsulata (He et al., 2007), Fusarium oxysporum (Mukherjee et al., 2002), Sargassum wightii (Singaravelu et al., 2007), Lactobacillus sp. (Binoj and Pradeep, 2002) and Helminthosporum solani (Kumar et al., 2008). Advantages of biological methods include tightly controlled, highly reproducible syntheses: biocompatible particles: and the avoidance of toxic surfactants or organic solvents. Moreover, bacteria are easy to handle and can be manipulated genetically. * Corresponding author. Tel.: +91 4563 289042; fax: +91 4563 289322. E-mail address: [email protected] (S. Gurunathan). 0960-8524/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.biortech.2009.05.051
Considering these advantages, a bacterial system could prove to be an excellent alternative for the synthesis of gold nanoparticles (Sweeney et al., 2006). The advent of nanotechnology has bestowed its use in the fields of drug delivery and energy science. The use of gold and gold compounds, as well as their potential therapeutic applications, in ancient and contemporary medicine has been reviewed periodically over the years (Tiekink, 2002). Such particles are being explored as gold-based pharmaceuticals (Evans et al., 2000; Okada et al., 1993; Tiekink, 2002) and as agents in biohydrogen production (Zhang and Shen, 2007). We report here, the synthesis of stable gold nanocubes by the reduction of aqueous AuCl4 by Bacillus licheniformis, at room temperature. This is a single-step process without the requirement of toxic chemicals and stringent conditions. The spectroscopic analysis and the results of XRD confirmed the crystalline nature of gold nanoparticles. Moreover, SEM analysis confirmed the synthesis of nanocubes by B. licheniformis. The size of the particles was calculated using equations derived earlier for UV–Vis spectra and XRD pattern line. The mechanism by which gold nanocubes are formed by bacteria is still unknown. 2. Methods 2.1. Preparation of gold nanoparticles B. licheniformis was grown in nitrate medium; yeast extract – 5 g/l; peptone – 5 g/l; potassium nitrate – 1 g/l. The cultures were incubated at 200 rpm for 24 h at room temperature. After 24 h, the cultures were centrifuged at 15,000 rpm for 15 min. The pellets were collected to obtain about 1 g of wet cells and re-suspended
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K. Kalishwaralal et al. / Bioresource Technology 100 (2009) 5356–5358
in 100 ml of a 1 mM aqueous AuCl4 solution and incubated for 48 h at 200 rpm followed by sonication and centrifugation (3000g; 30 min) (Kalimuthu et al., 2008). The gold nanoparticles suspended in supernatant were separated from solution by slow evaporation of water at 50–55 °C. The dried powder was collected and subjected to XRD and SEM measurements. 2.2. Characterization of Au nanoparticles Characterization of synthesized gold nanoparticles was carried out according to methods described previously (Kalimuthu et al., 2008; Kalishwaralal et al., 2008). UV/Vis spectroscopy measurements of gold nanoparticles samples were carried out on a Shimadzu (model 9200) UV–Vis spectrophotometer operating at a resolution of 0.72 nm. The dried powder was subjected to field emission scanning electron microscopy JEOL 6701 and to a XDL 3000 powder X-ray diffractometer. The SEM micrograph and the corresponding EDX spectrum were recorded by focusing on clusters of particles. Lattice parameters were calculated using high-angle reflections of XRD. The full widths at half-maximum (FWHM) values of X-ray diffractions were used to calculate particle sizes using the Debye–Scherrer formula. 3. Results and discussion The aqueous chloroaurate ions were reduced to metallic gold (Au0) on exposure to the bacterial biomass. The color of the reaction solution turned from pale yellow to dark purple which indicated the formation of gold nanoparticles (Kumar et al., 2008). The UV–Vis spectra results indicated that the reaction solution has an absorption maximum at about 540 nm attributable to the surface plasmon resonance band (SPR) of the gold nanoparticles
Fig. 1. SEM image of the AuCl4 treated bacterial cells. SEM micrograph of the 1 mM gold chloride treated B. licheniformis. High resolution (scale bar at 100 nm) image of the gold chloride treated sample.
(data not shown). Observation of this peak, assigned to a surface plasmon, is well documented for various metal nanoparticles with sizes ranging from 2 to 100 nm (Sastry et al., 1998). Size of the nanoparticle can be calculated using Eq. (1). With the use of the fit parameters determined from the theoretical values for d > 25 nm (k0 – 512; L1 – 6.53; L2 – 0.0216) the average of the absolute error in calculating the experimentally observed particle diameters is only 3%, and hence Eq. (1) allows a precise determination of d in the range of 5–100 nm (Haiss et al., 2007)
ln d¼
kspr k0 L1
ð1Þ
L2
X-ray diffraction was used to confirm the crystalline nature of the particle. The diffraction peaks at 2h = 38.31°, 44.46°, 64.67° and 77.45° obtained are identical with those reported for the standard gold metal (Au0) (Joint Committee on Powder Diffraction Standards-JCPDS, USA) (data not shown). No other diffraction peaks were observed, thus confirming the synthesis of pure gold nanoparticles (Kumar et al., 2008). In order to confirm the size and shape of the synthesized gold nanoparticles, the samples were analyzed under the scanning electron microscopy. The image revealed the synthesized nanoparticles are in the form of nanocubes (Fig. 1). Gold nanocubes were formed in several different sizes, ranging from polydisperse small nanocubes to large nanocubes. The particles were in the range of 10– 100 nm in size. XRD patterns were analyzed to determine the peak characteristics (intensity, position and width). Full width at half-maximum (FWHM) data was used with the Scherrer’s formula to determine mean particle size which is given by
d¼
0:9k b cos h
ð2Þ
where d – is the mean diameter of the nanoparticles, k – is wavelength of X-ray radiation source, b is the angular FWHM of the XRD peak at the diffraction angle h (Kalishwaralal et al., 2008). The particle size obtained from XRD line (Eq. (2)) and UV spectrum (Eq. (1)) broadening relatively matched with those obtained from the SEM results (Table 1). From these the average particle size was found to range from 10 to 100 nm. Metabolic products secreted by the bacteria, such as proteins, organic acids and polysaccharides, are expected to interact with the crystal faces, thereby changing the surface energies of the latter in due course. In this process, the bacteria functions as a cellular efflux pumping system and a periplasmic protein that binds gold specifically at the cell surface (Beveridge and Murray, 1980; Matias and Beveridge, 2008) bringing about alteration of solubility and toxicity via reduction, bio-sorption, bioaccumulation and lack of specific metal transport systems. The mechanism of reduction of Au nanoparticles in the bacterium is still not well understood. Previous reports have claimed that an enzyme belonging to the NADH reductase family is involved (He et al., 2007; Kumar et al., 2008). The reduction AuCl4 ions to Au0 ions facilitate the synthesis of nanocubes. The primarily formed gold nanoparticles are thermodynamically unstable in aqueous solution because of insufficient capping agent. At this point, they would tend to form linear assemblies
Table 1 FWHM values and particle sizes of gold nanoparticles from X-ray diffraction, UV–Vis spectrum and SEM measurements. 2h
FWHM (dh)
Particle size from XRD (nm)
Particles size from UV spectrum absorbance 540 nm
Particle size range from SEM (nm)
38.3147 46.4240 64.6727 77.457
0.5441 0.3212 1.0595 1.4338
25.48 nm
67 nm
10–100 (scale bar – 100 nm)
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K. Kalishwaralal et al. / Bioresource Technology 100 (2009) 5356–5358
driven by Brownian motion and short-range interaction. It is probable that proteins acted as the major bio-molecules involved in the bio-reduction and facilitated the synthesis of gold nanoparticles. It was shown that concentration of gold ions (AuCl4 ) played an important role in forming and stabilizing the shape of gold nanocubes (Delapierre et al., 2008; He et al., 2008). Even other anisotropic metal nanostructures are expected using the biosynthetic method. 4. Conclusion We report a green chemistry approach using B. licheniformis in the synthesis of gold nanocubes at room temperature without using any harmful reducing agents. The average particle size of nanocubes was found to be 10–100 nm. This single-step greener approach is general and cost effective. The flexibility of gold nanocubes could find applications in drug delivery and recently, gold nanocubes have extended its application to fields such as cancer diagnosis and treatment (Skrabalak et al., 2007). References Beveridge, T.J., Murray, R.G.E., 1980. Site of metal deposition in the cell wall of Bacillus subtilis. J. Bacteriol. 141, 876–887. Binoj, N., Pradeep, T., 2002. Coalescence of nanoclusters and formation of submicron crystallites assisted by Lactobacillus strains. Cryst. Growth Des. 2, 293–298. Delapierre, T.M., Majimel, J., Mornet, S., Duguet, E., Ravaine, S., 2008. Synthesis of non-spherical gold nanoparticles. Gold Bull. 41, 195–207. Evans, D.J., Cullinan, P., Geddes, D.M., Walters, E.H., Milan, S.J., Jones, P., 2000. Gold as an oral corticosteroid sparing agent in stable asthma. Cochrane Database Syst. Rev. 4, 1–17. Haiss, W., Thanh, N.T.K., Aveyard, J., Fernig, D.G., 2007. Determination of size and concentration of gold nanoparticles from UV–Vis spectra. Anal. Chem. 79, 4215– 4221. He, S., Guo, Z., Zhang, Y., Zhang, S., Wang, J., Gu, N., 2007. Biosynthesis of gold nanoparticles using the bacteria Rhodopseudomonas capsulate. Mater. Lett. 61, 3984–3987.
He, S., Zhang, Y., Guo, Z., Gu, N., 2008. Biological synthesis of gold nanowires using extract of Rhodopseudomonas capsulate. Biotechnol. Progr. 24, 476–480. Kalimuthu, K., Suresh Babu, R., Venkataraman, D., Mohd, Bilal, Gurunathan, S., 2008. Biosynthesis of silver nanocrystals by Bacillus licheniformis. Colloids Surf. B Biointerfaces 65, 150–153. Kalishwaralal, K., Deepak, V., Ramkumarpandian, S., Nellaiah, H., Sangiliyandi, G., 2008. Extracellular biosynthesis of silver nanoparticles by the culture supernatant of Bacillus licheniformis. Mater. Lett. 62, 4411–4413. Kumar, S.A., Peter, Y.A., Nadeau, J.L., 2008. Facile biosynthesis, separation and conjugation of gold nanoparticles to doxorubicin. Nanotechnology 19, 495101. Matias, V.R.F., Beveridge, T.J., 2008. Lipoteichoic acid is a major component of the Bacillus subtilis periplasm. J. Bacteriol. 190, 7414–7418. Merroun, M., Rossberg, A., Hennig, C., Scheinost, A.C., Selenska-Pobell, S., 2007. Spectroscopic characterization of gold nanoparticles formed by cells and S-layer protein of Bacillus sphaericus JG-A12. Mater. Sci. Eng. C 27, 188–192. Mukherjee, P., Senapati, S., Mandal, D., Ahmad, A., Khan, M.I., Kumar, R., Sastry, M., 2002. Extracellular synthesis of gold nanoparticles by the fungus Fusarium oxysporum. ChemBioChem 3, 461–463. Mukherjee, P., Roy, M., Mandal, B.P., Dey, G.K., Mukherjee, P.K., Ghatak, J., Tyagi, A.K., Kale, S.P., 2008. Green synthesis of highly stabilized nanocrystalline silver particles by a non-pathogenic and agriculturally important fungus T. asperellum. Nanotechnology 19, 075103. Okada, T.P.B.K., Shui, Q.Y., Gurney, M.E., 1993. Aurothiolates inhibit HIV-1 infectivity by gold(I) ligand exchange with a component of the virion surface. Virology 192, 631–642. Sastry, M., Patil, V., Sainkar, S.R.J., 1998. Electrostatically controlled diffusion of carboxylic acid derivatized silver colloidal particles in thermally evaporated fatty amine films. Phys. Chem. B 102, 1404–1410. Schmid, G. (Eds.), 1994. Clusters and Colloids. From Theory to Applications. VCH Verlagsgesellschaft, Weinheim, p. 546. Singaravelu, G., Arockiamary, J.S., Ganesh Kumar, V., Govindaraju, K., 2007. A novel extracellular synthesis of monodisperse gold nanoparticles using marine alga, Sargassum wightii Greville. Colloids Surf. B Biointerfaces 57, 97–101. Skrabalak, S.E., Chen, J., Au, L., Lu, X., 2007. Gold nanocages for biomedical applications. Adv. Mater. Deerfield 19, 3177–3184. Sweeney, S.F., Woehrle, G.H., Hutchison, J.E., 2006. Rapid purification and size separation of gold nanoparticles via diafiltration. J. Am. Chem. Soc. 128, 3190– 3197. Tiekink, E.R.T., 2002. Gold derivatives for the treatment of cancer. Crit. Rev. Oncol. Hematol. 42, 225–248. Wang, Z., Chen, J., Yang, P., Yang, W., 2007. Biomimetic synthesis of gold nanoparticles and their aggregates using a polypeptide sequence. Appl. Organometal. Chem. 21, 645–651. Zhang, Y., Shen, J., 2007. Enhancement effect of gold nanoparticles on biohydrogen production from artificial wastewater. Int. J. Hydrogen Energy 32, 17–23.
Contents lists available at ScienceDirect
Bioresource Technology journal homepage: www.elsevier.com/locate/biortech
Short Communication
Biological synthesis of gold nanocubes from Bacillus licheniformis Kalimuthu Kalishwaralal, Venkataraman Deepak, Sureshbabu Ram Kumar Pandian, Sangiliyandi Gurunathan * Department of Biotechnology, Kalasalingam University, Anand Nagar, Krishnankoil-626190, Tamil Nadu, India
a r t i c l e
i n f o
Article history: Received 4 February 2009 Received in revised form 21 May 2009 Accepted 21 May 2009 Available online 1 July 2009 Keywords: Gold nanocubes Bacillus licheniformis Enzyme Eco-friendly
a b s t r a c t Microorganisms play an important role in the eco-friendly synthesis of metal nanoparticles. This study illustrates the synthesis of gold nanocubes using the bacterium Bacillus licheniformis after 48 h of incubation at room temperature. The morphology of the samples was analyzed using scanning electron microscopy (SEM) and the particles formed were characterized to be nanocubes. The size of gold nanocubes in aqueous solution has been calculated using UV–Vis spectroscopy, XRD and SEM measurements. The nanoparticles are found to be polydisperse nanocubes in the size range 10–100 nm. Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction The area of nanotechnology encompasses the synthesis of nanoscale materials, the understanding and the utilization of their physicochemical and optoelectronic properties, and the organization of nanoscale structures into predefined superstructures. Thus, nanotechnology promises to play an increasingly important role in many key technologies of the new millennium (Schmid, 1994). Usually the chemical methods followed are carried out under very high temperatures to form gold nanoparticles and also result in environmental pollution due to toxicity of the reagents (reducing agents like borohydrates and acetylene) used (Wang et al., 2007). Consequently, researchers in the field of nanoparticle preparation have been looking at biological systems (Merroun et al., 2007) such as those that allow a commercially viable and environmentally clean synthesis of highly stabilized gold particles (Mukherjee et al., 2008). Beveridge and Murray (1980) first demonstrated that the exposure of Bacillus subtilis treated with gold chloride (AuCl4) resulted in the synthesis of gold nanoparticles. Biosynthesis of gold nanoparticles has been carried out using Rhodopseudomonas capsulata (He et al., 2007), Fusarium oxysporum (Mukherjee et al., 2002), Sargassum wightii (Singaravelu et al., 2007), Lactobacillus sp. (Binoj and Pradeep, 2002) and Helminthosporum solani (Kumar et al., 2008). Advantages of biological methods include tightly controlled, highly reproducible syntheses: biocompatible particles: and the avoidance of toxic surfactants or organic solvents. Moreover, bacteria are easy to handle and can be manipulated genetically. * Corresponding author. Tel.: +91 4563 289042; fax: +91 4563 289322. E-mail address: [email protected] (S. Gurunathan). 0960-8524/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.biortech.2009.05.051
Considering these advantages, a bacterial system could prove to be an excellent alternative for the synthesis of gold nanoparticles (Sweeney et al., 2006). The advent of nanotechnology has bestowed its use in the fields of drug delivery and energy science. The use of gold and gold compounds, as well as their potential therapeutic applications, in ancient and contemporary medicine has been reviewed periodically over the years (Tiekink, 2002). Such particles are being explored as gold-based pharmaceuticals (Evans et al., 2000; Okada et al., 1993; Tiekink, 2002) and as agents in biohydrogen production (Zhang and Shen, 2007). We report here, the synthesis of stable gold nanocubes by the reduction of aqueous AuCl4 by Bacillus licheniformis, at room temperature. This is a single-step process without the requirement of toxic chemicals and stringent conditions. The spectroscopic analysis and the results of XRD confirmed the crystalline nature of gold nanoparticles. Moreover, SEM analysis confirmed the synthesis of nanocubes by B. licheniformis. The size of the particles was calculated using equations derived earlier for UV–Vis spectra and XRD pattern line. The mechanism by which gold nanocubes are formed by bacteria is still unknown. 2. Methods 2.1. Preparation of gold nanoparticles B. licheniformis was grown in nitrate medium; yeast extract – 5 g/l; peptone – 5 g/l; potassium nitrate – 1 g/l. The cultures were incubated at 200 rpm for 24 h at room temperature. After 24 h, the cultures were centrifuged at 15,000 rpm for 15 min. The pellets were collected to obtain about 1 g of wet cells and re-suspended
5357
K. Kalishwaralal et al. / Bioresource Technology 100 (2009) 5356–5358
in 100 ml of a 1 mM aqueous AuCl4 solution and incubated for 48 h at 200 rpm followed by sonication and centrifugation (3000g; 30 min) (Kalimuthu et al., 2008). The gold nanoparticles suspended in supernatant were separated from solution by slow evaporation of water at 50–55 °C. The dried powder was collected and subjected to XRD and SEM measurements. 2.2. Characterization of Au nanoparticles Characterization of synthesized gold nanoparticles was carried out according to methods described previously (Kalimuthu et al., 2008; Kalishwaralal et al., 2008). UV/Vis spectroscopy measurements of gold nanoparticles samples were carried out on a Shimadzu (model 9200) UV–Vis spectrophotometer operating at a resolution of 0.72 nm. The dried powder was subjected to field emission scanning electron microscopy JEOL 6701 and to a XDL 3000 powder X-ray diffractometer. The SEM micrograph and the corresponding EDX spectrum were recorded by focusing on clusters of particles. Lattice parameters were calculated using high-angle reflections of XRD. The full widths at half-maximum (FWHM) values of X-ray diffractions were used to calculate particle sizes using the Debye–Scherrer formula. 3. Results and discussion The aqueous chloroaurate ions were reduced to metallic gold (Au0) on exposure to the bacterial biomass. The color of the reaction solution turned from pale yellow to dark purple which indicated the formation of gold nanoparticles (Kumar et al., 2008). The UV–Vis spectra results indicated that the reaction solution has an absorption maximum at about 540 nm attributable to the surface plasmon resonance band (SPR) of the gold nanoparticles
Fig. 1. SEM image of the AuCl4 treated bacterial cells. SEM micrograph of the 1 mM gold chloride treated B. licheniformis. High resolution (scale bar at 100 nm) image of the gold chloride treated sample.
(data not shown). Observation of this peak, assigned to a surface plasmon, is well documented for various metal nanoparticles with sizes ranging from 2 to 100 nm (Sastry et al., 1998). Size of the nanoparticle can be calculated using Eq. (1). With the use of the fit parameters determined from the theoretical values for d > 25 nm (k0 – 512; L1 – 6.53; L2 – 0.0216) the average of the absolute error in calculating the experimentally observed particle diameters is only 3%, and hence Eq. (1) allows a precise determination of d in the range of 5–100 nm (Haiss et al., 2007)
ln d¼
kspr k0 L1
ð1Þ
L2
X-ray diffraction was used to confirm the crystalline nature of the particle. The diffraction peaks at 2h = 38.31°, 44.46°, 64.67° and 77.45° obtained are identical with those reported for the standard gold metal (Au0) (Joint Committee on Powder Diffraction Standards-JCPDS, USA) (data not shown). No other diffraction peaks were observed, thus confirming the synthesis of pure gold nanoparticles (Kumar et al., 2008). In order to confirm the size and shape of the synthesized gold nanoparticles, the samples were analyzed under the scanning electron microscopy. The image revealed the synthesized nanoparticles are in the form of nanocubes (Fig. 1). Gold nanocubes were formed in several different sizes, ranging from polydisperse small nanocubes to large nanocubes. The particles were in the range of 10– 100 nm in size. XRD patterns were analyzed to determine the peak characteristics (intensity, position and width). Full width at half-maximum (FWHM) data was used with the Scherrer’s formula to determine mean particle size which is given by
d¼
0:9k b cos h
ð2Þ
where d – is the mean diameter of the nanoparticles, k – is wavelength of X-ray radiation source, b is the angular FWHM of the XRD peak at the diffraction angle h (Kalishwaralal et al., 2008). The particle size obtained from XRD line (Eq. (2)) and UV spectrum (Eq. (1)) broadening relatively matched with those obtained from the SEM results (Table 1). From these the average particle size was found to range from 10 to 100 nm. Metabolic products secreted by the bacteria, such as proteins, organic acids and polysaccharides, are expected to interact with the crystal faces, thereby changing the surface energies of the latter in due course. In this process, the bacteria functions as a cellular efflux pumping system and a periplasmic protein that binds gold specifically at the cell surface (Beveridge and Murray, 1980; Matias and Beveridge, 2008) bringing about alteration of solubility and toxicity via reduction, bio-sorption, bioaccumulation and lack of specific metal transport systems. The mechanism of reduction of Au nanoparticles in the bacterium is still not well understood. Previous reports have claimed that an enzyme belonging to the NADH reductase family is involved (He et al., 2007; Kumar et al., 2008). The reduction AuCl4 ions to Au0 ions facilitate the synthesis of nanocubes. The primarily formed gold nanoparticles are thermodynamically unstable in aqueous solution because of insufficient capping agent. At this point, they would tend to form linear assemblies
Table 1 FWHM values and particle sizes of gold nanoparticles from X-ray diffraction, UV–Vis spectrum and SEM measurements. 2h
FWHM (dh)
Particle size from XRD (nm)
Particles size from UV spectrum absorbance 540 nm
Particle size range from SEM (nm)
38.3147 46.4240 64.6727 77.457
0.5441 0.3212 1.0595 1.4338
25.48 nm
67 nm
10–100 (scale bar – 100 nm)
5358
K. Kalishwaralal et al. / Bioresource Technology 100 (2009) 5356–5358
driven by Brownian motion and short-range interaction. It is probable that proteins acted as the major bio-molecules involved in the bio-reduction and facilitated the synthesis of gold nanoparticles. It was shown that concentration of gold ions (AuCl4 ) played an important role in forming and stabilizing the shape of gold nanocubes (Delapierre et al., 2008; He et al., 2008). Even other anisotropic metal nanostructures are expected using the biosynthetic method. 4. Conclusion We report a green chemistry approach using B. licheniformis in the synthesis of gold nanocubes at room temperature without using any harmful reducing agents. The average particle size of nanocubes was found to be 10–100 nm. This single-step greener approach is general and cost effective. The flexibility of gold nanocubes could find applications in drug delivery and recently, gold nanocubes have extended its application to fields such as cancer diagnosis and treatment (Skrabalak et al., 2007). References Beveridge, T.J., Murray, R.G.E., 1980. Site of metal deposition in the cell wall of Bacillus subtilis. J. Bacteriol. 141, 876–887. Binoj, N., Pradeep, T., 2002. Coalescence of nanoclusters and formation of submicron crystallites assisted by Lactobacillus strains. Cryst. Growth Des. 2, 293–298. Delapierre, T.M., Majimel, J., Mornet, S., Duguet, E., Ravaine, S., 2008. Synthesis of non-spherical gold nanoparticles. Gold Bull. 41, 195–207. Evans, D.J., Cullinan, P., Geddes, D.M., Walters, E.H., Milan, S.J., Jones, P., 2000. Gold as an oral corticosteroid sparing agent in stable asthma. Cochrane Database Syst. Rev. 4, 1–17. Haiss, W., Thanh, N.T.K., Aveyard, J., Fernig, D.G., 2007. Determination of size and concentration of gold nanoparticles from UV–Vis spectra. Anal. Chem. 79, 4215– 4221. He, S., Guo, Z., Zhang, Y., Zhang, S., Wang, J., Gu, N., 2007. Biosynthesis of gold nanoparticles using the bacteria Rhodopseudomonas capsulate. Mater. Lett. 61, 3984–3987.
He, S., Zhang, Y., Guo, Z., Gu, N., 2008. Biological synthesis of gold nanowires using extract of Rhodopseudomonas capsulate. Biotechnol. Progr. 24, 476–480. Kalimuthu, K., Suresh Babu, R., Venkataraman, D., Mohd, Bilal, Gurunathan, S., 2008. Biosynthesis of silver nanocrystals by Bacillus licheniformis. Colloids Surf. B Biointerfaces 65, 150–153. Kalishwaralal, K., Deepak, V., Ramkumarpandian, S., Nellaiah, H., Sangiliyandi, G., 2008. Extracellular biosynthesis of silver nanoparticles by the culture supernatant of Bacillus licheniformis. Mater. Lett. 62, 4411–4413. Kumar, S.A., Peter, Y.A., Nadeau, J.L., 2008. Facile biosynthesis, separation and conjugation of gold nanoparticles to doxorubicin. Nanotechnology 19, 495101. Matias, V.R.F., Beveridge, T.J., 2008. Lipoteichoic acid is a major component of the Bacillus subtilis periplasm. J. Bacteriol. 190, 7414–7418. Merroun, M., Rossberg, A., Hennig, C., Scheinost, A.C., Selenska-Pobell, S., 2007. Spectroscopic characterization of gold nanoparticles formed by cells and S-layer protein of Bacillus sphaericus JG-A12. Mater. Sci. Eng. C 27, 188–192. Mukherjee, P., Senapati, S., Mandal, D., Ahmad, A., Khan, M.I., Kumar, R., Sastry, M., 2002. Extracellular synthesis of gold nanoparticles by the fungus Fusarium oxysporum. ChemBioChem 3, 461–463. Mukherjee, P., Roy, M., Mandal, B.P., Dey, G.K., Mukherjee, P.K., Ghatak, J., Tyagi, A.K., Kale, S.P., 2008. Green synthesis of highly stabilized nanocrystalline silver particles by a non-pathogenic and agriculturally important fungus T. asperellum. Nanotechnology 19, 075103. Okada, T.P.B.K., Shui, Q.Y., Gurney, M.E., 1993. Aurothiolates inhibit HIV-1 infectivity by gold(I) ligand exchange with a component of the virion surface. Virology 192, 631–642. Sastry, M., Patil, V., Sainkar, S.R.J., 1998. Electrostatically controlled diffusion of carboxylic acid derivatized silver colloidal particles in thermally evaporated fatty amine films. Phys. Chem. B 102, 1404–1410. Schmid, G. (Eds.), 1994. Clusters and Colloids. From Theory to Applications. VCH Verlagsgesellschaft, Weinheim, p. 546. Singaravelu, G., Arockiamary, J.S., Ganesh Kumar, V., Govindaraju, K., 2007. A novel extracellular synthesis of monodisperse gold nanoparticles using marine alga, Sargassum wightii Greville. Colloids Surf. B Biointerfaces 57, 97–101. Skrabalak, S.E., Chen, J., Au, L., Lu, X., 2007. Gold nanocages for biomedical applications. Adv. Mater. Deerfield 19, 3177–3184. Sweeney, S.F., Woehrle, G.H., Hutchison, J.E., 2006. Rapid purification and size separation of gold nanoparticles via diafiltration. J. Am. Chem. Soc. 128, 3190– 3197. Tiekink, E.R.T., 2002. Gold derivatives for the treatment of cancer. Crit. Rev. Oncol. Hematol. 42, 225–248. Wang, Z., Chen, J., Yang, P., Yang, W., 2007. Biomimetic synthesis of gold nanoparticles and their aggregates using a polypeptide sequence. Appl. Organometal. Chem. 21, 645–651. Zhang, Y., Shen, J., 2007. Enhancement effect of gold nanoparticles on biohydrogen production from artificial wastewater. Int. J. Hydrogen Energy 32, 17–23.