FACETIOUS GREEN SYNTHESIS OF SILVER NANO PARTICLES USING LEAF EXTRACTS OF ALANGIUM SALVIFOLIUM [LAM.]

* Mita R. Patel 1 , Dr. Rasmikant A. Patel 2 , N. A. Pithawala 3 and Dr. Kespi A. Pithawala 3 . 1. Department of Chemistry, Gujarat Arts and Science College, Ahmedabad, Gujarat, India, 380006. 2. Department of Chemistry, Municipal Arts and Urban Science College, Mehsana, Gujarat, India. 3. Department of Biology, Gujarat Arts and Science College, Ahmedabad, Gujarat, India, 380006. ...................................................................................................................... Manuscript Info Abstract ......................... ........................................................................ Manuscript History

The current research in nanotechnology is chiefly aimed at easy, environment friendly and cost effective methods of green synthesis of nanoparticles. Here aqueous leaf extracts of Alangium salvifolium [Lam.] which act as reducing as well as capping agents were used for the formation of silver nanoparticles (SNP) from 0.1mM AgNO 3 solution. The SNP were formed within two hours and the formation was stable for weeks. The formation, characteristics, size and conformation of silver nanoparticles formed were carried out using UV-VIS spectroscopy, FTIR, Dynamic light scattering (DLS), SEM, TEM and EDX. Also the SNP formed this way showed fair antimicrobial activity against gram positive and gram negative bacteria. On the basis of result obtained it can be said that the easy production of silver nanoparticle using green chemistry can be effectively utilized in various fields in biomedical-nanotechnology as well as reducing the harmful microbial flora. . Of all these approaches the most simple, easy and eco-friendly technique is through green route and today it is the most well-known strategy for nanoparticle amalgamation. [Joseph and Mathew 2014, Mehmood et al. , 2013, Rajeshkumar et al., 2013. The metals and the salts most popularly used are the coinage metals as Au, Ag and Cu however Fe, Zn,Pdand Pt have also been similarly used [Umer et al., 2012, Bonet et al., 1999, Najagi et al. , 2011, Vilchis-Nestor et al., 2008, Zheng and Wang 2013, Shankar et al. , 2004. SNP are extensively put to use in therapeutics and biomedical research and hence needed to be fused by green course utilizing characteristic plant materials which is naturally saviour-faire as well as expands incorporation of conditions that are not harsh or leading to loss of vitality and instead does not have to utilize high molecular weight, and harmful chemicals which render it eco-friendly and biocompatible , Esumi et al., 1990, Anilkumar et al., 2007, Bali et al., 2006]. Bio-enthused formation of nanoparticles gives headway over harsh and physical strategies as it is quick and particular in their objective towards the applications where they can be used for their antimicrobial action [Sivaranjani and Meenakshi Sundaram 2013]. Medicinal plants are chiefly exploited as a part of extensive extents these days in view of the long term change against disorders after ordinary treatment [Patilet al., 2012]. In this investigation silver nano particles have been formed utilizing the green course i.e. from the Corresponding Author:-Mita R. Patel. Address:-Department of Chemistry, Gujarat Arts and Science College, Ahmedabad, Gujarat, India, 380006.
292 aqueous leaf extracts of Alangium salvifolium regularly called 'Ankol' tree. This tree is of rare distribution in south Gujarat and the fruits of it are consumed by the locals and wood is exquisitely used as timber. The seeds have oils that is put to various medicinal usages as treatment of leprosy and skin disorders [Ratra andGupta 2015, Xavier et al., 2005] Materials and Method:-Healthy leaves of Alangium salvifolium were hand plucked from the tree. Silver nitrate was bought from Hi-Media and all chemicals were of logical review with 98-100% immaculateness measure as analytical reagents and chemical grade. Deionized and double distilled glass water was utilized throughout the experiment.

Preparation of Leaf Extracts:-
Fresh leaves of Alangium salvifolium were washed with deionized water and after that with refined water to free them of any dirt material. These were then subjected to dry in shaded space for around 7 days and then pounded into fine powder utilizing a clean sanitized kitchen grinder (mechanized, stainless steel cutting edges).This powder was stored in airtight glass bottles and was utilized for planning fluid concentrates. This was set up by including 1 g of leaf powder to 50 ml of double distilled water and left overnight in refrigerator. The concentrate was filtered through Whatmann's filter paper No. 1 and the extract was utilized for formation of nanoparticles.

Synthesis of Ag nanoparticles:-
To 30 ml of 0.1mM AgNO 3 10 ml of leaf extract was added and at interval of every 15 minutes colorimetric readings were taken at 410nm to find the presence of formation of nano particles the colour change was an indication of formation of nano particles. These started forming within 15 minutes and continued up to 2 hours after that these particles were stable for about one month.

Characterization techniques:-
The biosynthesized silver nanoparticles were characterized by the following methods: Visual Observation:-A change of colour from pale yellow to reddish brown was observed in the solution after visible irradiation.

UV Spectrophotometric analysis:-
The characterization technique involved ultra-violet and visible spectroscopy. UV-Vis absorption spectra were measured using Systronic UV-117 spectrometer from 300nm to 700nm continuously and the leaf powder extract was used as the reference for the baseline correction.
Fourier Transform Infrared Spectroscopy Analysis:-FTIR analysis was carried out to determine the functional groups present in leaf extract and their possible involvement in the formation of silver nanoparticles. FTIR analysis were carried using a FTIR SHIMADZU 8300 instrument where the samples were incorporated with KBr pellets to acquire the spectra. The results were compared for shift in functional peaks. A FTIR graph can be useful for preliminary investigation of surface chemistry of biogenic nanoparticles (i.e. those chemicals that contain carbon). This technique is widely used for identification of chemical residues such as amine, carbonyl and hydroxyl functional groups in a molecule [Angela et.al. 2016]. The FTIR analysis was performed with reduced silver nanoparticles. The synthesized AgNPs sample was mixed with KBr to make a pellet in the ratio of 1:100. The FTIR instrument with diffuse reflectance mode attachment. All measurements were carried out in the range of 400-4,000 cm-1 at a resolution of 4 cm-1. For this fresh sample were sent for FTIR Analysis at Gujarat Laboratory, Ahmedabad. Samples with total of volume 1-2 ml were given in aqueous form formed by producing SNPs using the reduction reaction of 9 ml of 0.1 mM Silver Nitrate solution through 3 ml of plant extract.

Dynamic Light Scattering:-
These studies were carried out to get to know the particle size distribution in the solution. The particle size comes out to be 60 nm and hence this can be further verified from EM analysis.

SEM & EDX Analysis:-
The surface morphology of silver nanoparticles was examined using a scanning electron microscopy (6010 LA, Jeol).The elemental composition of the synthesized silver nanoparticles was analyzed using Energy Dispersive X-Ray Spectrometer. TEM Analysis:-Since the particle sizes were small TEM studies were carried out to get to know the exact shape of the particles this was done using transmission electron microscope (JEM1400 Plus, Jeol). The particle sizes of about 30nm to 60nm can be seen clearly.

Determination of antimicrobial activity:-Microorganisms:-
The bacterial pathogens namely Staphylococcus aureus, Streptococcus pneumonia, Escherchia coli, Salmonella typhi, obtained from the Department of Microbiology, Gujarat Arts and Science College, Ahmedabad. These human pathogens were used to study the antibacterial activity. The nutrient broth, nutrient agar were used growing the test bacterial strains and were maintained on corresponding agar slants at 4°C.

Preparation of inoculums:-
The bacterial pathogens were inoculated into sterile nutrient broth and incubated at 37°C for 24 hours until the culture attained a turbidity of 0.5 McFarland units. The final inoculum was standardized to 105 CFU/ml by diluting fresh cultures with sterile distilled water. Colonies were suspended in 5 ml of sterile 0.85% saline. The resulting suspension was vortex and the turbidity was adjusted to yield 2×10 6 cells/ml (0.5 McFarland standards).
Antibacterial activity:-Antibacterial activity of AgNPs was determined by the agar disc diffusion method [Balouiri et al., 2016]. Plates of Nutrient agar were evenly streaked across the complete surface throughout the petri plate so as to get a loan growth of the inoculums with the help of spread plate technique with a known volume of 0.01 ml of active young culture with approx. Microbial count as 105 CFU/ml. Sterile filter paper discs (5 mm diameter) were immersed in the 50 μl of synthesized AgNPs (10, 20, 30, 40, 50 μg/ml) and allowed to dry at room temperature and it was placed over the Nutrient agar plates. Streptomycin 10 mcg/disc was used as positive control and the disc immersed in distilled water was used as negative control. The plates were incubated overnight at 37 ºC and the zone of inhibition around each disc was measured. Experiments were done in triplicate and mean values of zone diameter were taken.

Results And Discussion:-
An easy, cheap and a practical approach for eco-friendly synthesis of silver nanoparticles using aqueous leaf extract of Alangium salvifolium as both reducing and stabilizing agent, under the prescribed condition of room temperature has been used without the use of harsh inducers or hazardous chemical additives and/or severe chemical reactions. The formation of SNP was predicted visually from a color change and confirmed using UV-visible spectroscopy (410 nm), Figure-