Characterization and Antibacterial activity of Green Synthesized
ZnO Nanoparticles from Ocimum basilicum Leaf Extract G.Parthasarathy1, M.Saroja2, M.Venkatachalam2, V.K. Evanjelene3 1 Ph.DResearch Scholar, Department of Electronics, Erode Arts and Science College, Erode. 2Associate Professor, Department of Electronics, Erode Arts and Science College, Erode. 3R and D Manager, Alpha Omega Hi Tech Bio Research Centre, Salem.
ABSTRACT The Zinc Oxide nanoparticles are mostly used in the field of pharmaceutical sciences and it has more number of applications which plays an important role on antibacterial activity. In modern era, the nanoparticle synthesized by biological methods using micro organisms, enzymes, and plant extracts has been suggested as ecofriendly to environment. The nanostructured and highly stable zinc oxide nanoparticles are produced by Zinc nitrate and Ocimum Basilicum leaf Extract. Morphological, Structural and antimicrobial properties of synthesized nanoparticles are characterized by using XRD analysis, SEM, UV-Vis Spectrophotometer, FTIR and Disc diffusion Method (Antimicrobial activity). SEM and XRD analysis shows that synthesized nanoparticles are found to be predominantly hexagonal in shape and the average particle size range from 9 to 18nm. While increasing the concentration of the leaf extract of Ocimum Basilicum, the ZnO nanoparticles size can be increased. Keywords: Zinc Oxide, Ocimum basilicum, Antimicrobial activity, SEM, XRD, UV, FTIR
Received 24.01.2017 Revised 10.02.2017 Accepted 22.04.2017
How to cite this article: G.Parthasarathy, M.Saroja, M.Venkatachalam, V.K. Evanjelene. Characterization and Antibacterial activity of Green Synthesized ZnO Nanoparticles from Ocimum basilicum Leaf Extract. Adv. Biores., Vol 8 [3] May 2017.29-35
INTRODUCTION Nanoparticles attracted tremendous interest that have been extensively used in biological applications. Nanomaterials are classified into three different groups as ‘0’, ‘1’ and ‘2’ dimensional nanostructures [3]. These Nanoparticles are having noticeable performance in bioelectronics and biomedical applications. The Engineering technology and scientific field of nanosystems are one of the most quickly and exigent developing discipline of nanotechnology [3]. Zinc Oxide is an inorganic metal oxide and has a characteristic of exhibiting a wide range of nanostructures. The photo oxidizing and photocatalytic properties are used against biological and chemical species, which are used to characterize bio-synthesized metal oxides. In this work we used environmentally benign leaf extract of Ocimum Basilicum which has more medicinal properties [9]. While it is a surface stabilizing agent it acts as a biotemplate for the synthesis of Zinc Oxide nanoparticles. The Morphological, structural and antibacterial properties of zinc oxide nanoparticles are also evaluated.
MATERIALS AND METHODS
Synthesis of Zinc Oxide Nanoparticles using Ocimum Basilicum leaf Extract The Ocimum Basilicum leaves are collected from the foot hills of Yercaud, Salem, TamilNadu, India. The fresh plant leaves were washed with normal tap water and should be shadow dried. The leaves are crushed and coarsely powdered by mortar and pestle. The Coarsed powders of 25g were subjected to successive extraction in 250 ml of methanol (solvent) with the help of Soxhlet apparatus. Finally, the obtained leaf extract solution was stored and used for the further research. Further 50 ml of Ocimum Basilicum methanol extract has taken from prepared stock solution (stored already) and boiled at 600 to 800C by using stirrer-heater. When temperature reached 600C, 5g of Zinc nitrate hexahydrate was added. The mixture was boiled until the formation of deep yellow coloured suspension. The obtained paste was
transferred to ceramic crucible and annealed at 4000C for 2 hours. Finally, the obtained light white coloured powder was used for antibacterial activity, Structural and for other characterization. Antimicrobial assay Inoculums are prepared by, that the stock cultures are stored at 40C on slope of nutrient agar. Active cultures of experiment was prepared by shifting a loopful of cells from the stock cultures to the test tube of Muller-Hinton broth(MHB), that are incubated without agitation for 24 hours at 370C and 250C. Then the cultures are diluted with the fresh Muller-Hinton broth to gain optical densities approximately to 2.0 x 106 CFU/ ml for bacteria. Here the disc diffusion method (Bauer et al., 1966) was used to screen the antibacterial activity. In vitro, the microbial activity has been screened by using Muller-Hinton Agar (MHA) from Hi-media, Mumbai. The MHA plates are prepared using 15 ml of molten media into sterile petri plates. The plates are dried for 5 minutes and 0.1 %, inoculums suspension was swabbed throughout the plate and dried for 5 minutes. The concentration of extracts is 4 mg/disc was loaded on 6 mm sterile disc. The loaded disc was placed on the surface of the medium and extract was allowed to diffuse for 5 minutes and the plates are kept in incubator for 24 hours at 370C. As a result, the incubation zones on the disc were measured with the help of transparent ruler. The inhibition zones are obtained in the range of millimeter.
RESULTS AND DISCUSSION
Macroscopic characteristics The Ocimum Basilicum leaf is a compound and opposite bipinnateparipinnate with oblong shape mucoranate apex and has an average length of 15–20 cm. Single leaflet is 2–3 cm long and 10–15 mm breadth, oblong, linear, mucronate, setaceous deciduous. On average, in a mature compound leaf, there are 8– 10 paired leaflets. Physicochemical analysis The Physicochemical analysis of Ocimum Basilicum is shown in the Table 1. It shows the soluble and insoluble ash in percentage (%). Table 1: Physicochemical Properties S.No Parameter Values obtained (%) 1 Total Ash (%) 11.31 2 Water Soluble ash (%) 14.6 3 Water Insoluble ash (%) 10.3 4 Acid Soluble ash (%) 7.6 5 Acid Insoluble ash (%) 0.47 6 Sulphated ash (%) 17.3 Phytochemical analysis The qualitative phytochemical analysis of the Ocimum Basilicum leaf is shown in the Table 2. It shows the presence of alkaloids, flavonoids, Terpenoids, phenols and carbohydrates and hence steroids, anthroquinone, saponins, tannin, oils and resins were found to be absent in the extract [9].
Table 2: The qualitative phytochemical analysis
Phytochemicals Observations Sample B Alkaloids Cream color + Mayer’s test Reddish brown solution/ precipitate + Wagner’s test Flavonoids Yellow orange + Lead acetate test Reddish brown / Orange colour precipitate + H2SO4 test Steroids Violet to blue or Green color formation - Liebermann-Burchard test Terpenoids Reddish brown precipitate + Salkowski test Anthroquinone Pink color - Borntrager’s test Phenols Deep blue to Black colour formation + Ferric chloride test White precipitate + Lead acetate test Saponin Stable persistent - Tannin Brownish green / Blue black - Carbohydrates Yellow / brownish / blue / green color + Oil and Resin Filter paper test - (+) - Presence, (-) - Absence
X-Ray Diffraction By scattering the X-ray beam on the sample, we get the information about the crystallographic structure, chemical and physical properties of the ZnO nanoparticles. The 2θ values 32.75, 34.66, 37.20, 57.41, 62.85, 66.14, and 68.03 is corresponding to the plane of (100), (002), (101), (110), (103), (112), and (201) respectively according to JCPDS No. 036 1451 shown in Figure 1. The particle sizes are obtained in the range of 13.86, 9.25 and 18.38 nm. The nanoparticle sizes are calculated by using Debye Scherrer's formula. λ D= Å β θ
Figure 1: XRD Pattern
UV-Vis Spectrum The obtained UV-Vis Spectrum result was shown in Figure 2. The result shows that the maximum absorption has takes place at 305.01nm.
Figure 2: UV-Vis Spectrum
SEM imaging This analysis was performed by using Hitachi S-4500 Scanning Electron Microscope. SEM image Figure 3 shows that the ZnO nanoparticles are hexagonal in shape and the particle sizes are < 50nm in range.
FTIR Spectrum The FTIR Spectrum of synthesized ZnO nanoparticles has shown in Figure 4. Form this result we analyzed that C-O stretching (R-O-R, H3C-OH, etc.) at 1065.42 cm-1, Amine group (NH2) at 3400 cm-1, Scissoring, N-H bending at 1615 cm-1, Keto group (C=O) at 1742 cm-1, Presence of carbonyl group stretching (C=O) at 1427.94 cm-1 and Carbon-Carbon stretching in ring (medium) in Aromatic ring occurs at 1586.63 cm-1. C-H stretching in aromatic ring (Ar-H) occurs at 3043.17 cm-1. 721.58 cm-1 peak showed that it is finger print region. Hence these variations are occurred due to the presence of metal oxides.
Figure 4: FTIR Spectrum
Antibacterial Activity The antibacterial activity of synthesized ZnO nanoparticles shows better effect against S.typhi, S.aureus, B.subtilis, E.coli and P.aeruginosa shown in Figure 5. The maximum zone of inhibition obtained for 50µl concentration was observed that E. coli (16 mm), S.aureus (14 mm), S.typhiand B.subtilis (13 mm) shown in Table 3. The methanol extracts has reported that it is more effective than aqueous and ethanol extracts against for all the organisms [17].
Figure 5: Antibacterial activity of ZnO Nanoparticles
Table 3: Antibacterial activity
Zone Of Inhibition (mm) S.N Concentration Concentration of Concentration of Concentration of Organisms O. Control of Sample Sample Sample Sample 20µl 30µl 40µl 50µl 1 S.typhi 23mm 00mm 07mm 11mm 13mm 2 S.aureus 22mm 00mm 00mm 09mm 14mm 3 B.subtilis 22mm 00mm 07mm 10mm 13mm 4 E.coli 23mm 00mm 00mm 11mm 16mm P.aeruginos 5 24mm 00mm 00mm 09mm 10mm a
CONCLUSION Green Synthesis of ZnO nanoparticles are safe and eco-friendly to environment while comparing to chemical synthesis. The Morphological studies and Characterizations are confirmed that the presence of ZnO nanoparticles and also the particle size are obtained in the range of 13.86nm, 9.25nm and 18.38nm. The antibacterial activity against S.typhi, S.aureus, B.subtilis, E.coli and P.aeruginosa has exhibited good results. Hence the ZnO nanoparticles are synthesized using the plant extract will overcome the chemical methods [23].
ACKNOWLEDGEMENTS The author is sincere thanks also to Dr.M.Saroja, Associate Professor and Dr.M.Venkatachalam, Head and Associate Professor, Department of Electronics, Erode Arts and Science College, Erode for their vital support and submission of the manuscript. The author extends his thanks to Dr.V.K.Evanjelene, Managing
Director of Alpha Omega Hi-tech Bio Research Centre, Salem for her vital support and providing the facility to work in the lab.
Conflict of Interest There are no conflicts of interest.
Financial support and sponsorship
Nil
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Rajiv, 2013. Bio-Fabrication of Zinc Oxide Nanoparticles Using Leaf Extract of Parthenium Hysterophorus L. and Its Size-Dependent Antifungal Activity Against Plant Fungal Pathogens
Rajiv, 2013. Bio-Fabrication of Zinc Oxide Nanoparticles Using Leaf Extract of Parthenium Hysterophorus L. and Its Size-Dependent Antifungal Activity Against Plant Fungal Pathogens
