Preliminary identification of phytochemicals, green-synthesis, characterization and applications of silver-nanoparticles-using Asparagus-recemosus.

AbstractThe1 Present study aimed to identify the phytochemicals present in Asparagus recemosus and green synthesis, characterization and applications of silver nanoparticles. Three types of leaf and root aqueous extract namely fresh, sun dried and oven dried samples have been used for the phytochemical studies and further green synthesis, characterization and application studies. Phytochemicals such as Alkaloid, carbohydrate, tannin,flavanoids, saponin and steroid have been found to be present in all the three types of samples of root and tannin and flavonoids were found to be present only in sundried leaf sample and it was absent in both fresh and hot air oven dried leaf sample. Saponin was found to be present in both fresh and hot air oven dried samples but was absent in sun dried sample. On the other hand steroids were present only in fresh leaf sample and was absent in both sun dried and hot air oven dried samples. Green synthesis was identified after 24 hours of incubation in the mixed solution and it was indicated as color change from yellow to brown of mixed solution. UV analysis of 24 hrs. Of incubated solution showed peaks at 452nm, 454nm and 460 nm for the fresh, sun dried and hot air oven dried samples respectively confirmed the green synthesis of silver nanoparticles by reduction process. Further X Ray diffraction analysis confirmed the presence of silver in the nanoparticles as the Braggs peak obtained in plane (1, 1, 1) at 2ø scale 27.808⁰, 27.849⁰ and 27.905 for fresh, sun dried and hot air oven dried sample respectively. HR-SEM analysis results showed the formation of elliptical and spherical shaped silver nanoparticles. FT-IR spectroscopic study confirmed that the presence of Amide C=O stretch group (peak value at 1631.78cm-1), Carboxylic acid C=O stretch (peak value at 1728.22cm-1,) and Alcohol/Phenol O-H stretch (peak value, 3460.30cm-1).In antibacterial study, the zone of inhibition was found to be increased with increasing concentration of silver nanoparticle form 50µl to 150µl against both the pathogens E.coli and Staphylococcus aureus. Key words: Asparagus racemosus, Silver nanoparticles, phytochemicals and E.coli.IntroductionGreen organic synthesis of metal nanoparticles using medicinal plants is in the light of booming field of research in science. (Leela and Vivekanathan, 2008, Raveendran et al., 2006 and Piling, 2000). Light metal nanoparticle synthesis using photoperiodic plants leads wide spread area of applicability in nano medicine and medicinal chemistry (Magudapathy et al., 2001, Joerger et al., /2000 and Panacek et al., 2006). Small particle size (1-100nm) with crystal nature promotes nano particles penetrating capacity and drug delivery through cell. Scientists follows different areas of research in green synthesis for treatment of cancer, epilepsy (McNeil, 2005), antibiotic delivery (Ma gudapathy et al., 2001) and protective coating materialistic research against toxic microbes (Li Y et al., 2006). Coupling of light metal nanoparticles with the cancer specific ligands clinically motivates targeted anti-cancer drug delivery (Shankar et al., 2004). Organic and inorganic synthesis of silver nanoparticles and anti-microbial activity tests were extremely beneficial to the human immune system. (Mukherjee et al., 2001, Vigneswaran et al., 2007, Leela and Vivekanthan, 2008 and Vaseashta and Dimova-Malinovska, 2005). Synthesized nano particles with coated sample broth extracts having antimicrobial effects were observed clearly though microscopy (Morones et al., 2005). In-vitro synthesized and characterized light metal nanoparticles and subsequent results depends upon chemical reductions on aqueous sample solutions(Joerger et al., 2000), natural and artificial thermal variable parameters against inorganic solvents(Vaseashta and Dimo va-Malinovska,2005), photo sensitivity and reduction capability of nanoparticles (Roy et al., 1999).Wide spread applications of Nano technology in various areas focused on imaging(Wu J et al., 2009), sensing (Chan and Nie ,1998), targeted drug delivery (Joerger et al., 2000),gene delivery system (Sachlos et al.,2006) and artificial implants (Stoimenov et al., 2002 and Shirashi and Toshiba,2000). Different studies were carried out for focusing the effect of the medicinal plants synthesized metal nanoparticles (Azardicta indica , Cardiospermum ) against variable parameters. The Coating effect of nano particles found in studies were used as antimicrobial agent (Pileni, 2000 and Panacek et al., 2006) . Toxic effect of silver nano particle visually identified by the blue-grey discoloration on skin called argyria (Hadrup and Lam, 2014). Silver nano particles used in socks causes chances to reduce foot odor being released in the washing with possible negative consequences (Wu J et al., 2009).Present study focused on the green synthesis of silver metal nanoparticle using medicinal plant Asparagus racemosus using leaf and root aqueous extracts and its antibacterial activity. Experimentaldetails Materials Reagents all used in this experiment were obtained from Sigma Aldrich chemicals. Double distilled water was used throughout the experiments for the purpose of washing and extract preparation. Filtration was established by using Whatman No.1 filter papers. Glassware used in the experiments were washed well, rinsed with double distilled water and dried in a hot air oven. Preparation of aqueous extract of Asparagus racemosus The healthy, fresh roots from Asparagus racemosus plants were collected from VIT University nursery, Tamil Nadu, India. The roots were thoroughly washed with running tap water followed by distilled water to remove dirt and other impurities. The cleaned roots were shade dried for two weeks so as to remove the moisture completely and then root pieces were made into powder by using mixer grinder. The 5g of powdered roots were taken into a beaker along with 100 ml of distilled water and allowed to boil at 60˚C for 30 minutes. Then the cooled solution was filtered through Whatmann No.1 Filter paper to get the aqueous root extract. The obtained extract was stored in a refrigerator at 4 °C for further use [10, 12-13] Phytochemical analysis The powered root extract of Asparagus racemosus was used for phytochemical analysis. Standard phytochemical tests were adopted to screen the different types of phytochemicals present in the root extract. (12, 14-15).Biosynthesis of silver nanoparticles For the synthesis of silver nanoparticles (AgNPs) 25 ml of root extract of Asparagus racemosus was added to 75 ml of 0.1M AgNO3 aqueous silver nitrate solution. The mixed solution was stirred well and kept for 24 incubation for the color change.Characterization of silver nanoparticles Preliminary confirmation of green synthesis of silver nanoparticle after 24hrs.of incubation of mixed solution was done by UV-Visible spectrophotometer in the range between 300-700 nm. FTIR spectrum analysis was carried out for the aqueous root extract so as to find out the reducing biomolecules present in the extract. The structure of silver nanoparticles were determined by X–Ray diffraction analysis was done to further confirm or identify the presence of silver metal, its associated forms and crystal structure of green synthesized silver nanoparticles. Nanoparticle morphology such as size, shape and mean particle size were determined by HR-SEM analysis [10, 17)Application studiesAntibacterial activity: The antibacterial study of AgNPs was evaluated against two bacterial pathogens such as E. coli and S. aureus by disc diffusion method. The root extract and the synthesized Cu NPs solution were added from three level of concentration of 50ml, 100ml and 150ml respectively to the sterilized disc.These discs were carefully dispen