Journal of Radiation Research and Applied Sciences xxx (2015) 1e7 HOSTED BY Available online at www.sciencedirect.com ScienceDirect Journal of Radiation Research and Applied Sciences journal homepage: http://www.elsevier.com/locate/jrras Controllable synthesis of silver nanoparticles using Neem leaves and their antimicrobial activity Aparajita Verma, Mohan Singh Mehata * Laser-Spectroscopy Laboratory, Department of Applied Physics, Delhi Technological University, Bawana Road, Delhi 110042, India article info Article history: Received 15 September 2015 Received in revised form 8 October 2015 Accepted 4 November 2015 Available online xxx Keywords: Silver nanoparticles Azadirachta indica Absorption Photoluminescence Green synthesis abstract Silver nanoparticles (AgNPs) were synthesized using aqueous extract of Neem (Azadirachta indica) leaves and silver salt. XRD, SEM, FTIR, optical absorption and photoluminescence (PL) were measured and analysed. The synthesized AgNPs exhibits lowest energy absorption band at 400 nm. The effects of various parameters i.e., extract concentration, reaction ph, reactants ratio, temperature and interaction time on the synthesis of AgNPs were studied. It was found that the formation of AgNPs enhanced with time at higher temperature and alkaline ph. The AgNPs formed were found to have enhanced antimicrobial properties and showed zone of inhibition against isolated bacteria (Escherichia coli) from garden soil sample. Based on the results obtained, it can be concluded that the resources obtained from plants can be efficiently used in the production of AgNPs and could be utilized in various fields such as biomedical, nanotechnology etc. Copyright 2015, The Egyptian Society of Radiation Sciences and Applications. Production and hosting by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). 1. Introduction The field of nanotechnology has proved to be one of the most active areas of research (Moore, 2006; Sergeev & Shabatina, 2008). Synthesis of nanoparticles is increasing exponentially because of its wide range of applications in the field of optoelectronics, biosensors, bio-nanotechnology, biomedicine etc. (Bharali, Sahoo, Mozumdar, & Maitra, 2003; Mehata, Majumder, Mallik, & Ohta, 2010; Mehata, 2012, 2015; Ratnesh & Mehata, 2015; Saxena, Mozumdar, & Johri, 2006; Subbiah, Veerapandian, & Yun, 2010). Various physical and chemical methods have been formulated for the synthesis of nanopartilces of desired shape and size. However these methods are not economically feasible and environment friendly. Therefore, green synthesis has been considered as one of the promising method for synthesis of nanopartilces because of their biocompatibility, low toxicity and eco-friendly nature (Malik, Shankar, Malik, Sharma, & Mukherjee, 2014). Various microorganism and plants have proved to be a source of inspiration for nanomaterial synthesis. Some well-known examples of nanoparticles synthesized by microorganisms either intracellularly (Weiner & Dove, 2003)or extracellularly (Bansal, Bharde, Ramanathan, & Bhargava, 2012) are: synthesis of magnetite by magnetotactic bacteria (Dickson, 1999; Lovley, Stolz, Nord, & Phillips, 1987; Philipse & Maas, 2002) and synthesis of siliceous material by radiolarians and diatoms (Kr oger, Deutzmann, & Sumper, 1999; Mann, 1993; Oliver, Kuperman, Coombs, Lough, & Ozin, 1995). A mixture of curiosity and unshakable belief that mother earth * Corresponding author. E-mail addresses: msmehata@gmail.com, mohan.phy@dce.edu (M.S. Mehata). Peer review under responsibility of The Egyptian Society of Radiation Sciences and Applications. http://dx.doi.org/10.1016/j.jrras.2015.11.001 1687-8507/Copyright 2015, The Egyptian Society of Radiation Sciences and Applications. Production and hosting by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
2 Journal of Radiation Research and Applied Sciences xxx (2015) 1e7 has developed the best method for the synthesis of nano range of materials have led to a new and exciting field of research which involves microorganism and plants for the synthesis of nanomaterials. The green synthesis methods include synthesis of nanoparticles using microorganisms like bacteria, fungus, yeasts (Narayanan & Sakthivel, 2010), plants (Jha, Prasad, Prasad, & Kulkarni, 2009; Makarov et al., 2014; Mittal, Chisti, & Banerjee, 2013) and DNA (Sohn, Kwon, Jin, & Jo, 2011). Multiple species of bacteria and fungi have been investigated for the growth of nanoparticles of different composition and size, for example, synthesis of gold by Verticilliumsp (Narayanan & Sakthivel, 2010), synthesis of CdS quantum dots using fungi etc. (Ahmad et al., 2002). Besides microbes, use of part of plants like stem, leaves, roots etc. (Jha et al., 2009) for the synthesis of nanoparticles is yet another exciting possibility that is relatively unexplored. Advantage of using plants over microorganism is the elimination of the elaborate process of cell culture. Moreover, nanoparticles synthesized using biological methods are more compatible for medical use as compared to chemical and physical methods where toxic material may adsorb on the surface of the nanoparticles that may have adverse effect when used for medicinal purpose. The biosynthesis method employing plant extracts of Pelargonium graveolens, Medicagosativa, Azadirachta indica, Lemongrass, Aploevera, Cinnamomum Camphor (Jha et al., 2009; Makarov et al., 2014; Mittal et al., 2013; Shankar, Rai, Ahmad, & Sastry, 2004) have drawn great attention as an alternative to conventional methods, because plants are found in abundance in nature. In recent years, increasing antibiotic resistance by microbes is imposing serious threat to the health sector. Nanoparticles have proved to be a likely candidate for antimicrobial agent since their large surface to volume ratio ensures a broad range of attack on bacterial surface. One of the most promising nanoparticle which acts as a highly effective antimicrobial agent is silver. Various investigations on silver nanoparticles have been done to study its antimicrobial activity. AgNPs exhibited significant antibacterial activity against Escherichia coli, Staphylococcus aureus and antifungal activity against Trichophyton, Trichosporon beigelii and Candida albicans (Gajbhiye, Kesharwani, Ingle, Gade, & Rai, 2009). Considering the advantages of green synthesis over other methods, this study aims at the synthesis of AgNPs using aqueous Neem (Azadirachta indica) leaves extract. It focuses on the study of the effects of various physicoechemical parameters on AgNps. We also attempt to investigate about the antimicrobial effect of the synthesized nanoparticles. Azadirachta indica, which is a common plant known as Neem is found abundantly in India and in nearby Indian subcontinents. It belongs to Meliaceae family and is known for its various applications especially its medicinal property (Subapriya & Nagini, 2005). Azadirachta indica leaf extract is used in the synthesis of various nanoparticles like gold, zinc oxide, silver etc. The phytochemicals present in Neem are namely terpenoids and flavanones, which act as reducing as well as capping agent and helping in stabilizing the nanoparticles. When silver salt is treated with Neem leaf extract, the silver salt is reduced to AgNPs. The synthesized nanoparticles, which are capped with neem extract also exhibit enhanced antibacterial activity. 2. Materials and methods Silver nitrate was obtained from Sigma-Aldrich chemical Co. All the glassware were washed with distilled water and dried in oven. The petri-plates and agar were autoclaved before use. 20 g of finely cut Neem leaves were boiled in 100 ml water for 10 min and filtered to obtain Neem leaves extract. The extract of Neem leaves (5 ml) were mixed with 45 ml of 1 mm silver nitrate (AgNO 3 ) and colour change was observed indicating the formation of AgNPs (Shankar et al., 2004). The effects of various physicoechemical parameters were examined by varying the reactant concentration, ph, temperature and reaction time. Reduction of Ag þ ions was monitored after diluting a small amount of sample 20 times. Absorption spectra were recorded with UV/VIS/NIR spectrometer (Perkin Elmer Lambda 750) and photoluminescence (PL) spectra were recorded with Fluorolog-3 spectrofluorometer (Horiba Jobin Yyon) equipped with double-grating at excitation and emission monochromators (1200 grooves/mm) and an R928P photomultiplier tube (PMT). The excitation source was a 450 Watt CW xenon lamp. Effect of time was studied by measuring the absorption spectra of the solution at the time interval of 5, 15, 25, 35 and 45 min. Effect of ph was studied by varying the ph of both Neem broth and silver salt solution. 0.1 N KOH and 0.1 N HCL was added to adjust the ph of the solution. The ph variation was observed from ph 8e12 with an accuracy of ±0.2. Effect of temperature was measured by varying the temperature between 10 and 50 C with an accuracy of ±3 C. Neem broth containing AgNPs were centrifuged at 10,000 rpm for 15 min and the precipitate was thoroughly washed with sterile distilled water to get rid of any unwanted impurities. The purified pellet was then dried at 60 C and the sample was characterized using scanning electron microscope (SEM, Hitachi S7000N) and X-ray diffractometry (XRD, Bruker D8 advanced). Biomolecules responsible for the reduction of silver salt were studied using Fourier transform infrared (FTIR) spectrometer (Thermoscientific Nicolet 380). The synthesized AgNPs were then tested for their antibacterial property against bacteria obtained from garden soil samples. The bacteria were grown on 1.8% agar plates then a small amount of AgNPs were added for the study of antibacterial property. 3. Results and discussion 3.1. Effect of reaction time and concentration on the formation of AgNPs When silver salt (AgNO 3 ) is added to aqueous Neem leaf extract it results into a colour change from pale yellow to yellowish brown and finally to dark brown colour, as shown in Fig. 1. The change in colour of the solution is due to the presence of silver nanoparticles formed by the reduction of silver salt. The reduction of silver salt to silver ions is due to the presence of reducing agents. It was suggested that compounds like caffeine and theophylline act as reducing agent when Acalypha indica leaf extract was used (Krishnaraj et al., 2010). However, in Neem leaves extract natural reducing
Journal of Radiation Research and Applied Sciences xxx (2015) 1e7 3 Fig. 1 e Change in colour of the solution with time when silver salt was added to Neem leaf broth. agent like terpenoids and flavanones are present, which are responsible for reduction of silver salt to AgNPs. AgNPs show yellowish brown colour in aqueous solution due to the excitation of surface plasmon vibrations (Shankar et al., 2004). It is noticed that the complete colour change took after about 30 min, thereafter no further colour of the reaction mixture changed. This indicates that silver salt present in the reaction mixture has been reduced completely. Then the formation of silver nanoparticles was examined and confirmed by obtaining the respective absorption spectra. The absorption spectrum originated due to the strong surface plasmon resonance (SPR), i.e., due to resonant absorption of photons by AgNPs. The observed absorption band is size dependent, since SPR band depends on size and the refractive index of the solution (Amendola, Bakr, & Stellacci, 2010). Fig. 2 shows the absorption spectra of AgNPs obtained from the reaction of Neem leave extract and AgNO 3 recorded in the range of 250e700 nm at different reaction time. Absorption maximum is observed at 400 nm, which is at higher energy as that obtained by Shankar et al. (2004) and with olive leaf (Khalil, Ismail, El-Baghdady, & Mohamed, 2013), indicating that the prepared AgNPs are smaller and has uniform size distribution. It is observed that there is an increase in the absorbance with the passage of time (inset Fig. 2), indicating an enhancement in the formation of AgNPs. Change in colour was observed initially after 5 min of adding the salt solution to the Neem leaf broth. After 30 min, the colour of the solution becomes nearly constant, indicating that no silver salt was left for further reaction. The results are in complete correlation with that of reported by Shankar et al. (2004). With the passage of time, the intensity of SPR band increased without any shift in peak wavelength. According to Mie theory (Rout, Lakkakula, Kolekar, Mendhulkar, & Kashid, 2009), absorption spectra of spherical AgNPs exhibit single SPR band, and as the anisotropy increases the number of peaks increases. In the present study, SPR band suggests that the synthesized nanoparticles are spherical in shape which is further confirmed by SEM study. The size of the Ag Nanoparticles is calculated using modified Mie theory as given: gðrþ ¼g 0 þðay F Þ=R (1) Fig. 2 e Absorption spectra of AgNPs observed at five different reaction times of 10 3 M aqueous solution of silver nitrate with Neem leaf broth. Inset figure shows an increase of absorption intensity as a function of reaction time. where g(r) is resonance broadening, A is scattering process (3/ 4 in case of Ag), g o is the velocity of bulk scattering (5 10 12 s 1 for Ag) and v f is the Fermi velocity. From equation (1), the size of the AgNPs (obtained when silver salt was mixed with Neem leaf broth and left for 45 min) comes out to be approximately 4.0 nm. Fig. 3 shows the absorption spectra of AgNPs obtained with changing the concentration of Neem broth and silver salt. When the concentration of Neem leaf broth to silver salt is in the ratio 1:16, a relatively weak absorption band is observed. With increase in concentration of reaction mixture (1:02), the peak intensity increases abruptly, indicating an enhancement in the production of AgNPs. The absorption intensity increases monotonically with increasing concentration of Neem
4 Journal of Radiation Research and Applied Sciences xxx (2015) 1e7 Fig. 3 e Absorption spectra of silver nanoparticles at various concentrations of Neem leaf broth and silver salt (1:02, 1:04, 1:08, 1:12, 1:16). Inset figure shows an increase of absorption intensity as a function of concentration. broth (as shown in inset Fig. 3). Moreover, on increasing the concentration of Neem leaf broth the absorption spectrum shifted towards red (from 408 to 421 nm), indicating an increase in the size of AgNPs. 3.2. Effect of ph on the formation of AgNPs Another important parameter which affects the formation of nanoparticles is the ph of the solution. Change in ph affects the shape and size of the particles, as ph has the ability to alter the charge of biomolecules, which might affect their capping as well as stabilizing abilities. Fig. 4 shows change in peak absorption wavelength and intensity on varying the ph of the solution. As the ph increases from 9 to 13, the absorption maximum shifts from 383 to 415 nm. In addition to the spectral shift, the absorption intensity increases with increasing ph. This indicates that ph 13 is the most favourable ph for the synthesis of AgNPs using Neem leaf extract. Further, it was observed that the ph enhances the rate of reduction reaction, the colour change was observed very fast when AgNO 3 mixed with aqueous Neem leaf extract, i.e. within few minutes the colour of the sample changed to dark brown. The shift in the peak wavelength indicates that the size of the particles increases with increasing ph of the solution. As the diameter of the particles get larger, the energy required for excitation of surface plasmon electrons decreases, as a result the absorption maximum shifted towards the longer wavelength region. Moreover, it was observed that at acidic ph i.e. ph < 7, the formation of nanoparticles is suppressed. At high ph, the bioavailability of functional groups in Neem leaf extract promoted the synthesis of nanoparticles. However, at very high ph i.e. ph ~ 13, the particles became unstable and agglomerated, when kept for overnight. A linear relation is observed between absorption maximum and ph (inset Fig. 4). It has been mentioned (Khalil et al., 2013) that the absorbance of the AgNPs obtained from olive leave extract increases with increasing ph of the solution from 2 to 8. Upon further increasing the ph of the solution, the absorbance decreases. However, in the present study upon increasing the ph from 9 to 13 the absorbance increases monotonically, indicating that the alkaline ph is more favourable for the synthesis of AgNPs (Vanaja et al., 2013). 3.3. Effect of temperature on the formation of AgNPs Temperature is yet another important factor that affects the synthesis of nanoparticles significantly. Fig. 5 shows the absorption spectra of AgNPs at different temperatures in the range of 10e50 C. With the increase in temperature, the reduction of silver salt is enhanced, as indicated by rapid change in the colour of the solution. The peak absorption wavelength shifted toward blue from 433 to 397 nm, as temperature varies from 10 to 50 C. The shift in the band maximum is due to localization of surface plasmon resonance of the AgNPs. This indicates that the size of the synthesized nanoparticles decreases with increasing temperature, which may be due to the faster reaction rate at higher temperature. At high temperature, the kinetic energy of the molecules increases and silver ions gets consumed faster, thus leaving less possibility for particle size growth. Thus, smaller particles of uniform size distribution are formed at higher temperature. The present study is in complete correlation with the work reported based Fig. 4 e Absorption spectra of silver nanoparticles at different ph values (ph ¼ 9, 10, 11, 12, 13) of the reaction mixture. Inset figure shows a linear relation between the absorption maximum and ph. Fig. 5 e Absorption spectra of silver nanoparticles obtained at different reaction temperature of 10,20,30,40 and 50 C. Inset figure shows nearly a linear relation between the absorption maximum and temperature.
Journal of Radiation Research and Applied Sciences xxx (2015) 1e7 5 on the banana peel extract (Ibrahim, 2015), where the intensity of peak maximum increases with increasing temperature along with the blue shift in peak wavelength. 3.4. Surface morphology of AgNPs XRD pattern of AgNps was recorded in the range of 30e70 at 2q angles and analysed (as shown in supporting information Fig. S 1 ). The XRD pattern shows that the AgNps obtained by the reduction of silver salt using Neem leaf extract are crystalline in nature. The high intensity peaks at around 37,44 and 64 corresponding to three diffraction faces of silver. The XRD peak at around 37 represent to the Bragg reflection corresponding to the (111) plane. SEM images of the synthesized AgNPs obtained from Azadirachta indica leaf extract is given in supporting information as Figs. S 2 and Fig. S 3. The obtained nanoparticles are spherical in shape and crystalline in nature. The samples used to examine the morphology of AgNPs were kept for longer time, which may have been agglomerated, therefore showing larger particle size than that obtained from optical absorption spectra. Furthermore, to identify the possible biomolecules present in Neem leaf broth which are responsible for capping and stabilizing the metal nanoparticles FTIR spectra were measured. The FTIR spectra were measured for pure Neem leaf broth and purified AgNPs obtained after centrifugation of sample at 10,000 rpm (Fig. S 4 ). The FTIR spectrum of Neem leaf broth shows peaks at 3389, 1635 and 1390 cm 1. These peaks correspond to the groups present in the sample and are indicating to the OeH stretching (around 3389 cm 1 ), C]C group (around 1635 cm 1 ) and geminal methyl group (around 1380 cm 1 ), which is in good correlation with that of the other reports (Tripathy, Raichur, Chandrasekaran, Prathna, & Mukherjee, 2010). These bands suggest the presence of terpenoids in Neem leaf. It can be inferred that terpenoids present in Neem leaf extract acts as stabilizing as well as capping agents. Besides terpenoids, presence of flavanones are also possible. Terpenoids and flavanones interact through carbonyl groups and are adsorbed on the surface of the metal ions. Terpenoids reduces metal ions by oxidation of aldehydic groups in the molecules to carboxylic acid (Smitha, Philip, & Gopchandran, 2009). Fig. 6 e Photoluminescence spectra of AgNPs recorded at five different reaction times of 10 3 M aqueous solution of silver nitrate with Neem leaf broth. Excitation wavelength was 380 nm. Inset figure shows an increase of PL intensity as a function of reaction time. the agar plates containing bacterial colony. Zone of clearance is observed maximum at 12 mg ml 1 of AgNPs. The antibacterial activity of AgNPs can be explained due to the change in the cell membrane permeability or degradation of enzymes in bacteria. The zone of clearance observed at 12 mg ml 1 of AgNPs is 6 mm (see Fig. S 5 ). 3.6. Photoluminescence of synthesized AgNPs at different reaction conditions Fig. 6 shows photoluminescence (PL) spectra of AgNPs obtained with 380 nm excitation at various reaction time, the similar one used for the absorption spectra (cf. Fig. 2). The PL spectra show a strong and well defined peak at around 450 nm. With the passage of time, the peak intensity increases gradually, indicating an enhancement in nanoparticles synthesis. Such an enhancement in the PL intensity is also observed at higher concentration of silver salt with respect to Neem broth (Fig. 7), at alkaline ph and at higher temperature (see supporting 3.5. Antimicrobial activity of AgNPs Garden soil samples are taken and serially diluted on 1.8% agar plates, and a pour plating technique is used to culture the microorganisms present in different dilutions. The plates are marked for each dilution. Bacterial colony is observed after incubating the agar plates overnight at 37 C. Bacterial colonies are obtained from the plate which is serially diluted to 10 6 (dilution factor). Agar plates are four quadrants streaked to isolate different colonies and to obtain pure culture. Biochemical analysis is further done to determine the strain of bacteria. Colonies of E. coli and S. aureus were obtained. In the present study E. coli strain is used to study the antibacterial property of silver nanoparticles. The antibacterial activity of the synthesized AgNPs which is tested against bacterial colony obtained from soil sample is shown in supporting information (seefig. S 5 ). Different quantities of AgNPs (0, 2, 4, 8, 10 and 12 mg ml 1 ) are added to Fig. 7 e Photoluminescence spectra of AgNPs at various concentrations of Neem leaf broth and silver salt (1:02, 1:04, 1:08, 1:12, 1:16). Excitation wavelength was 350 nm. Inset figure shows an increase of PL intensity as a function of concentration.
6 Journal of Radiation Research and Applied Sciences xxx (2015) 1e7 Scheme 1 e Schematic representation of synthesis of AgNps. Neem leaves extract reacts with silver salt and formed AgNPs, which exhibit strong absorption, photoluminescence and antibacterial activity. information Figs. S 6 and S 7 ). The increase in intensity with increasing concentration of Neem broth can be explained by the presence of large number of functional groups that react with silver salt and increases the production of AgNPs. With the change in ph from acidic to alkaline, the intensity of PL band increases, indicating that the rate of reduction increases at higher ph. Higher temperature also support the rate of formation of AgNPs. It is known that the AgNPs emits light between 400 and 700 nm (Vigneshwaran et al., 2007), which occurs due to relaxation from the electronic motion of surface plasmon, when surface plasmon electrons absorbs light at resonant frequency a part of the absorption energy transferred into heat energy and part of it radiates as PL, and a recombination of sp electrons with holes in the d band (Parang et al., 2012). Acknowledgement A.V. thanks to all the lab members especially to Mr. R.K. Ratnesh for his constant help in the measurements. This work is financially supported by the DAE-BRNS, Govt. of India (Grant No. 2012/37P/20/BRNS/765). Appendix A. Supplementary data Supplementary data related to this article can be found at http://dx.doi.org/10.1016/j.jrras.2015.11.001. references 4. Conclusions AgNPs have been successfully synthesized using a well-known medicinal plant Neem leaf extract (Scheme 1). The synthesized AgNps are crystalline in nature, polydispersed and exhibit high energy SPR band at around 400 nm and a strong PL at around 450 nm (Scheme 1), depending on controllable parameters. The synthesis is found to be efficient in terms of reaction time as well as stability of the AgNPs. The rate of synthesize is faster in case of Neem as compared to the other biological methods microbes, DNA etc. Thus, the rate of reaction of biological synthesis is comparable to that of the chemical methods. The synthesis process, i.e., formation of AgNPs critically depends on the ph, temperature, reactant concentration and reaction time. By changing these environmental parameters, the size and shape of the synthesized nanoparticles can be altered. Synthesis of AgNPs is enhanced with time at higher temperature and alkaline ph. Green synthesized AgNPs are found to have enhanced antibacterial activity against bacterial colony isolated from soil sample. Due to the enhanced antimicrobial activity of AgNPs, it is effectively used in the field of medicine as well as in food and cosmetic industries. Ahmad, A., Mukherjee, P., Mandal, D., Senapati, S., Khan, M. I., Kumar, R., et al. (2002). Enzyme mediated extracellular synthesis of CdS nanoparticles by the fungus, Fusarium oxysporum. Journal of the American Chemical Society, 124(41), 12108e12109. http://dx.doi.org/10.1021/ja027296o. Amendola, V., Bakr, O. M., & Stellacci, F. (2010). A study of the surface plasmon resonance of silver nanoparticles by the discrete dipole approximation method: effect of shape, size, structure, and assembly. Plasmonics, 5(1), 85e97. http:// dx.doi.org/10.1007/s11468-009-9120-4. Bansal, V., Bharde, A., Ramanathan, R., & Bhargava, S. K. (2012). Inorganic materials using unusual microorganisms. Advances in Colloid and Interface Science, 179, 150e168. http://dx.doi.org/ 10.1016/j.cis.2012.06.013. Bharali, D. J., Sahoo, S. K., Mozumdar, S., & Maitra, A. (2003). Cross-linked polyvinylpyrrolidone nanoparticles: a potential carrier for hydrophilic drugs. Journal of Colloid and Interface Science, 258(2), 415e423. http://dx.doi.org/10.1016/s0021-9797(02)00099-1. Dickson, D. P. (1999). Nanostructured magnetism in living systems. Journal of Magnetism and Magnetic Materials, 203(1), 46e49. http://dx.doi.org/10.1016/s0304-8853(99)00178-x. Gajbhiye, M., Kesharwani, J., Ingle, A., Gade, A., & Rai, M. (2009). Fungus-mediated synthesis of AgNPs and their activity
Journal of Radiation Research and Applied Sciences xxx (2015) 1e7 7 against pathogenic fungi in combination with fluconazole. Nanomedicine: Nanotechnology Biology and Medicine, 5, 382e386. http://dx.doi.org/10.1016/j.nano.2009.06.005. Ibrahim, H. M. (2015). Green synthesis and characterization of silver nanoparticles using banana peel extract and their antimicrobial activity against representative microorganisms. Journal of Radiation Research and Applied Sciences. http:// dx.doi.org/10.1016/j.jrras.2015.01.007. Jha, A. K., Prasad, K., Prasad, K., & Kulkarni, A. R. (2009). Plant system: nature's nanofactory. Colloids and Surfaces B: Biointerfaces, 73(2), 219e223. http://dx.doi.org/10.1016/ j.colsurfb.2009.05.018. Khalil, M. M. H., Ismail, E. H., El-Baghdady, K. Z., & Mohamed, D. (2013). Green synthesis of silver nanoparticles using olive leaf extract and its antibacterial activity. Arabian Journal of Chemistry, 7, 1131e1139. http://dx.doi.org/10.1016/ j.arabjc.2013.04.007. Krishnaraj, C., Jagan, E. G., Rajasekar, S., Selvakumar, P., Kalaichelvan, P. T., & Mohan, N. (2010). Synthesis of silver nanoparticles using Acalypha indica leaf extracts and its antibacterial activity against water borne pathogens. Colloids and Surfaces B: Biointerfaces, 76(1), 50e56. http://dx.doi.org/ 10.1016/j.colsurfb.2009.10.008. Kr oger, N., Deutzmann, R., & Sumper, M. (1999). Polycationic peptides from diatom biosilica that direct silica nanosphere formation. Science, 286(5442), 1129e1132. http://dx.doi.org/ 10.1126/science.286.5442.1129. Lovley, D. R., Stolz, J. F., Nord, G. L., & Phillips, E. J. (1987). Anaerobic production of magnetite by a dissimilatory ironreducing microorganism. Nature, 330(6145), 252e254. http:// dx.doi.org/10.1038/330252a0. Makarov, V. V., Love, A. J., Sinitsyna, O. V., Makarova, S. S., Yaminsky, I. V., Taliansky, M. E., et al. (2014). Green nanotechnologies: synthesis of metal nanoparticles using plants. Acta Naturae, 6(1), 35. Malik, P., Shankar, R., Malik, V., Sharma, N., & Mukherjee, T. K. (2014). Green chemistry based benign routes for nanoparticle synthesis. Journal of Nanoparticles, 2014. http://dx.doi.org/ 10.1155/2014/302429. Mann, S. (1993). Molecular tectonics in biomineralization and biomimetic materials chemistry. Nature, 365(6446), 499e505. http://dx.doi.org/10.1038/365499a0. Mehata, M. S. (2012). Stark shifts and exciton dissociation in CdSe nanoparticle grafted conjugated polymer. Applied Physics Letters, 100(15), 151908. http://dx.doi.org/10.1063/ 1.3701719. Mehata, M. S. (2015). Enhancement of charge transfer and quenching of photoluminescence of capped CdS quantum dots. Scientific Reports, 5. http://dx.doi.org/10.1038/srep12056. Mehata, M. S., Majumder, M., Mallik, B., & Ohta, N. (2010). External electric field effects on optical property and excitation dynamics of capped CdS quantum dots embedded in a polymer film. Journal of Physical Chemistry C, 114(37), 15594e15601. http://dx.doi.org/10.1021/jp104124z. Mittal, A. K., Chisti, Y., & Banerjee, U. C. (2013). Synthesis of metallic nanoparticles using plant extracts. Biotechnology Advances, 31(2), 346e356. http://dx.doi.org/10.1016/ j.biotechadv.2013.01.003. Moore, M. N. (2006). Do nanoparticles present ecotoxicological risks for the health of the aquatic environment? Environment International, 32(8), 967e976. http://dx.doi.org/10.1016/ j.envint.2006.06.014. Narayanan, K. B., & Sakthivel, N. (2010). Biological synthesis of metal nanoparticles by microbes. Advances in Colloid and Interface Science, 156(1), 1e13. http://dx.doi.org/10.1016/ j.cis.2010.02.001. Oliver, S., Kuperman, A., Coombs, N., Lough, A., & Ozin, G. A. (1995). Lamellar aluminophosphates with surface patterns that mimic diatom and radiolarian microskeletons. Nature, 378(6552), 47e50. http://dx.doi.org/10.1038/378047a0. Parang, Z., Keshavarz, A., Farahi, S., Elahi, S. M., Ghoranneviss, M., & Parhoodeh, S. (2012). Fluorescence emission spectra of silver and silver/cobalt nanoparticles. Scientia Iranica F, 19(3), 943e947. http://dx.doi.org/10.1016/j.scient.2012.02.026. Philipse, A. P., & Maas, D. (2002). Magnetic colloids from magnetotactic bacteria: chain formation and colloidal stability. Langmuir, 18(25), 9977e9984. http://dx.doi.org/ 10.1021/la0205811. Ratnesh, R. K., & Mehata, M. S. (2015). Controlled synthesis and optical properties of tunable CdSe quantum dots and effect of ph. AIP Advances, 5, 097114. http://dx.doi.org/10.1063/1.4930586. Rout, R. W., Lakkakula, J. R., Kolekar, N. S., Mendhulkar, V. D., & Kashid, S. B. (2009). Phytosynthesis of silver nanoparticle using Gliricidia sepium (Jacq.). Current Nanoscience, 5, 117e122. http://dx.doi.org/10.2174/157341309787314674. Saxena, A., Mozumdar, S., & Johri, A. K. (2006). Ultra-low sized cross-linked polyvinylpyrrolidone nanoparticles as non-viral vectors for in vivo gene delivery. Biomaterials, 27(32), 5596e5602. http://dx.doi.org/10.1016/j.biomaterials.2006.06.029. Sergeev, G. B., & Shabatina, T. I. (2008). Cryochemistry of nanometals. Colloids and Surfaces A: Physicochemical and Engineering Aspects, 313, 18e22. http://dx.doi.org/10.1016/ j.colsurfa.2007.04.064. Shankar, S. S., Rai, A., Ahmad, A., & Sastry, M. (2004). Rapid synthesis of Au, Ag, and bimetallic Au coreeag shell nanoparticles using Neem (Azadirachta indica) leaf broth. Journal of Colloid and Interface Science, 275(2), 496e502. http:// dx.doi.org/10.1016/j.jcis.2004.03.003. Smitha, S. L., Philip, D., & Gopchandran, K. G. (2009). Green synthesis of gold nanoparticles using Cinnamomum zeylanicum leaf broth. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 74(3), 735e739. http://dx.doi.org/10.1016/ j.saa.2009.08.007. Sohn, J. S., Kwon, Y. W., Jin, J. I., & Jo, B. W. (2011). DNA-templated preparation of gold nanoparticles. Molecules, 16(10), 8143e8151. http://dx.doi.org/10.3390/molecules16108143. Subapriya, R., & Nagini, S. (2005). Medicinal properties of neem leaves: a review. Current Medicinal Chemistry-Anti-Cancer Agents, 5(2), 149e156. http://dx.doi.org/10.2174/ 1568011053174828. Subbiah, R., Veerapandian, M., & Yun, K. S. (2010). Nanoparticles: functionalization and multifunctional applications in biomedical sciences. Current Medicinal Chemistry, 17(36), 4559e4577. http://dx.doi.org/10.2174/092986710794183024. Tripathy, A., Raichur, A. M., Chandrasekaran, N., Prathna, T. C., & Mukherjee, A. (2010). Process variables in biomimetic synthesis of silver nanoparticles by aqueous extract of Azadirachta indica (Neem) leaves. Journal of Nanoparticle Research, 12(1), 237e246. http://dx.doi.org/10.1007/s11051-009-9602-5. Vanaja, M., Rajeshkumar, S., Paulkumar, K., Gnanajobitha, G., Malarkodi, C., & Annadurai, G. (2013). Kinetic study on green synthesis of silver nanoparticles using Coleus aromaticus leaf extract. Advances in Applied Science Research, 4(3), 50e55. Vigneshwaran, N., Ashtaputre, N. M., Varadarajan, P. V., Nachane, R. P., Paralikar, K. M., & Balasubramanya, R. H. (2007). Biological synthesis of silver nanoparticles using the fungus Aspergillus flavus. Materials Letters, 61, 1413e1418. http://dx.doi.org/10.1016/j.matlet.2006.07.042. Weiner, S., & Dove, P. M. (2003). An overview of biomineralization processes and the problem of the vital effect. Reviews in Mineralogy and Geochemistry, 54(1), 1e29. http://dx.doi.org/ 10.2113/0540001.