ESTIMATION OF BIO ACTIVITY OF ARIAL PARTS OF WITHANIA SOMNIFERA AGAINST THE BACTERIAL AND FUNGAL MICROBES Research Article DR. PREMLATA SINGARIYA* 1 , DR. PADMA KUMAR 1 AND DR. KRISHAN K. MOURYA 2 1 Laboratory of Tissue Culture and Secondary Metabolites, Department of Botany, University of Rajasthan, Jaipur 302004, 2 I st Gr. Govt. Veterinary Hospital, Pahari (Bharatpur) Rajasthan. Email: premlatasingariya@gmail.com, mourya07@gmail.com Received: 3 Feb 2012, Revised and Accepted: 26 March 2012 ABSTRACT We evaluated the bio-activity (antibacterial and antifungal) of Ethanol, acetone, Iso propyl alcohol, toluene and hexane extract of different arial parts (leaf and flower) of Withania somnifera. The dried and powdered parts were successively extracted using soxhlet assembly then antibacterial and antifungal activities were investigated with by both disc diffusion and serial dilution methods. The extracts of W. somnifera were evaluated or significantly inhibited six important bacteria (two Gram +ve and four Gram-ve bacteria) and two fungi Staphylococcus aureus (Gram +ve), Bacillus Subtilis (Gram +ve), Escherichia coli (Gram-ve), Raoultella planticola (Gram -ve), Pseudomonas aeruginosa (Gram-ve), Enterobactor aerogens (Gram- ve), Candida albicans and Aspergillus flavus to varying degrees. Leaf extracts of W. somnifera in different polar solvents showed highest activity in terms of inhibition zone, activity index, MIC, MBC/MFC and total activity. The inhibitory effect is very identical in magnitude and comparable with that of standard antibiotics. Gentamycin, the standard antibacterial drug used was effective in inhibiting these bacteria. The effect on S. aureus and B. subtilis were comparable to that of gentamycin. Ketoconazole, the standard antifungal used was effective against the fungi (C. albicans and A. flavus). Keywords: Withania somnifera, Raoultella planticola, Escherichia coli, Staphylococcus aureus and Candida albicans. INTRODUCTION Withania somnifera used in significant increase hemoglobin concentration, red blood cell count, white blood cell count, platelet count and body weight as compared to controls, as well as increased hemolytic antibody responses towards human erythrocytes 1 , Anti- inflammatory effect, analgesic effect, osteoarthritis 2 , immuno- potentiating and myeloprotective effect 3 , increased phagocytic activity and prolonged survival time 4 , antifungal activity of Withania has been confirmed elsewhere, attributed to the withanolides. Major causative agent of nosocomial infections is S. aureus 5 , E. aerogens along with E. coli. Raoultella planticola has been determined to cause severe pancreatitis in one case 6 . C. albicans is notorious for causing candidiasis, it can affect the esophagus with the potential of becoming systemic, causing a much more serious condition, afungemia called candidemia 7,8 . P. aeruginosa is involved in respiratory tract, urinary tract 9 , bloodstream, and central nervous system infections of nosocomial origin 10 and this pathogen is becoming resistant against gentamycin, ciprofloxacin 11 , tetracycline, chloramphenicol, and norfloxacin 12 . Bacillus Subtilis can contaminate food; however, they seldom result in food poisoning. E. aerogens is a nosocomial and pathogenic bacteria that causes opportunistic infections incl-ding most types of infections. MATERIAL AND METHODS Experimental design Crude extracts of different arial parts (leaf and flower) of W. somnifera (RUBL-20668) were prepared with a series of non polar to polar solvents by hot extraction method 13 in soxhlet assembly. Different extracts were then screened for antimicrobial activity by disc diffusion Assay 14 against a few medically important bacteria, fungi and fungi. The fraction showing best activity was then used for determining of minimum inhibitory concentration (MIC) by tube dilution method 15 and minimum bactericidal/fungicidal concentration (MBC/MFC). Collection of plant material Different parts of W. somnifera (RUBL-20668) were collected in the month of January from Jaipur district of Rajasthan. Plants samples were identified and deposited in the herbarium, department of botany, university of Rajasthan, Jaipur. The collected plant materials were separately shade dried for one week. Each shade dried plant part was powdered with the help of grinder. Fine powder of each sample was stored in clean container to be used for Soxhlet extraction following the method of Subramanian and Nagarjan 16 , in different polar solvents selected. Extraction procedure Each plant part (10 gm) was sequentially extracted with different solvents (250 ml) according to their increasing polarity by using Soxhlet apparatus for 18 hours at a temperature not exceeding the boiling point of the respective solvent. The obtained extracts were filtered by using Whatman No. 1 filter paper and then concentrated at 40 0 C by using an evaporator. The residual extracts were stored in refrigerator at 4 0 C in small and sterile glass bottles. Drugs and chemicals used Drugs: Gentamycin and Ketoconazole as standard antibiotics for bacteria and fungi respectively. Chemicals: Ethanol, acetone, iso propyl alcohol, toluene, hexane, Nutrient Agar (for bacteria), Sabouraud Dextrose Agar (for fungi). Micro-organisms: Test pathogenic microorganisms were procured from Microbial Type Culture Collection, IMTECH, Chandigarh, India. (table 1) Table 1: Name of the tested pathogens (bacteria, yeast and fungi) S. No. pathogens Name of Pathogens G+ve/G-ve Specimen no. 1. Bacteria Escherichia coli G-ve MTCC-46 2. Staphylococcus aureus G+ve MTCC-3160 3. Raoultella planticola G-ve MTCC-530 4. Pseudomonas aeruginosa G-ve MTCC-1934 5. Bacillus subtilis G+ve MTCC-121 6. Enterobactor aerogens G-ve MTCC-111 7. Fungi Candida albicans - MTCC-183 8. Aspergillus flavus - MTCC-277 International Journal of Pharmacy and Pharmaceutical Sciences ISSN- 0975-1491 Vol 4, Suppl 3, 2012 A A c c a a d d e e m mi i c c S S c c i i e e n n c c e e s s