International Research Journal of Environment Sciences________________________________ ISSN 2319–1414Vol. 3(10), 75-78, October (2014) Int. Res. J. Environment Sci. International Science Congress Association 75 Antibacterial Properties of Various Medicinal Plants Extracts against Klebsiella Sp.Sneh Lata, Geetika Sharma and Harmanjot Kaur SandhuDepartment of Biotechnology, University Institute of Pharma Science, Chandigarh University, Gharuan, SAS Nagar, Punjab, INDIAAvailable online at: www.isca.in, www.isca.me Received 30th July 2014, revised 21st August 2014, accepted 30th September 2014 AbstractThe present study involves the checking of antibacterial properties of various medicinal plants like lemon grass, stevia, jatropha against Klebsiella sp. as these bacteria can cause many diseases. Firstly culture was authenticated by Gram staining and also biochemical characterization was done which confirms the strain of bacteria. After that extracts of these plants like methanol extract, chloroform extract and hot water extract were prepared by using respective chemical .These extracts were prepared from leaves of respective plant. Zone of inhibition of each plant extract against Klebsiella sp. was measured and antibacterial activity of each extract of plant was compared for the results. Keywords: Klebsiella sp., Lemon Grass, Stevia, Jatropha, Chloramphenicol. Introduction Klebsiella is a gram negative, rod shape bacteria. It leads to a range of diseases like pneumonia, urinary tract infection and soft tissue infection etc. Many methods are now discovered to avoid these diseases. But naturally some plants are there which show antibacterial properties to this bacterium. Stevia is a herb used in many parts of the world as a non caloric sweetener . Stevia can also be used in bakery, beverage and soft drink industries. Stevia produces decrease in blood pressure L emon grass belongs to Poaceae family which is also a herb has antifungal properties The plant is a native herb from india and is cultivated in other tropical and subtropical countries Lemon grassis used in different parts of the world in the treatment of digestive disorders, fevers, menstrual disorder, rheumatism and other joint pains. Jatropha is perennial shrub belongs to Euphorbiaceae family same as rubber tree and cassava. Oil content in the seed is about 30-40%7-8. All these medicinal plants are tested for present research. Material and Methods Authentication of culture was done by gram staining and by biochemically characterising the culture. Biochemical characterization was done with the help of primary biochemical tests which included: indole test, urease, oxidase, catalase, nitrate and citrate tests. For biochemical tests standard procedures were used . Culture Condition: The microorganism was cultured in an incubator maintained at temperature of 37\rC. The culture was hand shaken once a day to ensure proper distribution of nutrient to the cell for better growth and was microscopically examined. The bacterial culture was maintained by transferring it to the autoclaved flasks containing nutrient broth. Table-1 Preparation of NB (Nutrient Broth) Chemicals Quantity (per 100 ml) Peptone 0.5g Beef Extract 0.3g NaCl 0.8g Growth Curve: Growth of culture was checked by preparing100 ml of nutrient broth and inoculating with Klebsiella sp. Readings were taken on spectrophotometer after different time intervals and O.D of the bacterial culture was plotted against time intervals.Preparation of Different Plant Extract, Preparation of Methanol Extract: Methanol extract was prepared by crushing leaves with methanol in a pestle mortar and then filtered the extract by using the filter paper and was stored in a flask. Preparation of Hot Water Extract: Hot water extract was prepared by crushing leaves with distilled water in a flask and place them at100\rC and then filtered the extract by using filter paper and stored in a flask. Preparation of Chloroform Extract: Chloroform extract was prepared by crushing leaves with chloroform in a pestle mortar and then filtered the extract by using filter paper and stored in a flask. Preparation of Acetone Extract: Acetone extract was prepared by crushing leaves in acetone with pestle mortar and then filtered the extract by using filter paper and stored in flask. International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(10), 75-78, October (2014) Int. Res. J. Environment Sci. International Science Congress Association 76 Table-2 Preparation of NAM (Nutrient Agar Medium) S.No. Chemicals Quantity(per 100 ml) 1. Peptone 0.5g 2. Beef Extract 0.3g 3. NaCl 0.8g 4. Agar 2.0g Antibiotic Sensitivity: Discs of whatmann filter paper were prepared. Nutrient agar medium was poured in three different petri plates and allowed to solidify for 15-20 minutes. The antibiotic solution (chloramphenicol) was prepared in three different dilutions i.e. 50 mg/ml, 5mg/ml and 0.5mg/ml and was poured in the above three solidified petri plates. Dipped two discs in each of these three petri plates and waited for 1-2 minutes for absorbance of antibiotic solution. The 0.5ml of the bacterial culture was poured in each of the three petri plates. With the help of swabbing stick swabbed the culture on them from first petri plate in which 50mg/ml antibiotic solution was added. Take two discs one by one and set in the second petri plate. After 24hrs a zone of inhibition was measured. Antimicrobial Assay Using Different Plant Extract: The antimicrobial activity of different extract of lemon grass, stevia and jatropha was tested on microorganism using the paper disc diffusion method 10. Results and DiscussionGram Staining: In the present study Gram staining of the culture showed Gram negative, rod shaped bacteria. Table-3 showing results of gram staining below. Table-3 Results of Gram staining of the culture S.No Colour Shape Type 1. Pink Rod Shaped Gram -ve 2. Pink Rod Shaped Gram -ve 3. Pink Rod Shaped Gram -ve Biochemical Characterization of Isolates: After gram staining, biochemical characterization of culture was done. Results for biochemical characterization are depicted in table-4 showing the positive results for catalase, citrate, urease and nitrate tests while negative results were observed for oxidase and indole tests . Table-4 Results of Biochemical Characterization Culture Catalase Test Oxidase Test Indole Test Citrate Test Urease Test Nitrate Test Strains 1. +Ve -Ve -Ve +Ve +Ve +Ve Klebsiella 2. +Ve -Ve -Ve +Ve +Ve +Ve Klebsiella Table-5Growth Curve Readings of Klebsiella sp.S. No. Time O.D. 1. 0 hr 0.015 2. 30 min 0.018 3. 60 min 0.018 4. 90 min 0.012 5. 120 min 0.027 Figure- 1Growth curve of Klebsiella sp. 0306090120Absorbance at 660 nmTime (in hours) International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(10), 75-78, October (2014) Int. Res. J. Environment Sci. International Science Congress Association 77 Results of antimicrobial assay of different extracts against Klebsiella: Measurement of zone of inhibition of chloramphenicol against Klebsiella sp. was done. Results are depicted in table-6 Table-6Zone of inhibition of chloramphenicol against Klebsiella. S. No. Concentration of Chloramphenicol Zone of Inhibition 1. 0.5mg 10mm 2. 5mg 15mm 3. 50mg 20mm Antibacterial activity of Jatropha: Highest antibacterial activity of jatropha against Klebsiella was exhibited by hot water extract (10mm zone of inhibition) followed by chloroform extract, methanol extract and acetone extract.Table-7 showing results. Table -7 Zone of inhibition of different extract of Jatropha against Klebsiella S. No. Extract of Jatropha Zone of Inhibition 1. Hot Water 10mm 2. Methanol 7mm 3. Chloroform 5mm 4. Acetone 7mm Antibacterial activity of Stevia: Highest antibacterial activity of stevia against Klebsiella was exhibited by methanol extract (10mm zone of inhibition) followed by acetone extract and chloroform extract and hot water extract. Results are depicted in table-8 Table-8 Zone of inhibition of different extract of stevia against Klebsiella S.No. Extracts of Stevia Zone of Inhibition 1. Hot Water 6mm 2. Methanol 10mm 3. Chloroform 7mm 4. Acetone 9mm Antibacterial activity of Lemon grass: Highest antibacterial activity of lemon grass against Klebsiella was exhibited by chloroform extract (9mm zone of inhibition) followed by hot water extract, methanol extract and acetone extract. Table -9Zone of inhibition of different extract of lemon grass against KlebsiellaS.No. Extracts of Lemon Grass Zone of Inhibition 1. Hot Water 9mm 2. Methanol 7mm 3. Chloroform 10mm 4. Acetone 6mm Figure-2Effect of concentration of chloramphenicol against Klebsiella Concentration of chloramphenicol (mg) zone of inhibition (mm) Effect of concentration of Chloramphenicol International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(10), 75-78, October (2014) Int. Res. J. Environment Sci. International Science Congress Association 78 Figure-3 Zone of inhibition of different plant extracts against KlebsiellaConclusion On observing the overall antibacterial activity of different extracts it was found that hot water extract of jatropha, methanol extract of stevia and chloroform extract of lemon grass showed the highest antibacterial activity (10mm) against Klebsiella sp. References 1.Cardello HM, Da Silva MA and Damasio MH., Measurement of relative sweetness Of stevia extract, aspartame and Cyclamate/Saccharin blend as compared to sucrose at different concentrations, Plant Foods Hum Nutr, 54, 119-130 (1995) 2.Melis MS., Chronic administration of aqueous extract of Stevia Rabaudiana in rats: renal effects, J Ethnopharmacol, 47,129-134 (1995) 3.Shadab Q., Hanif M. and Chaudhary F.M., Antifungal activity by Lemongrass essential oils, Pak. J. Sci. Ind. Res 35, 246-249 (1992)4.Figueirinha A., Paranhos A., Perez-Alonso J.J., Santos-Buelga C., and Batista M.T. 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(eds), Methods in Plant Biochemistry, Academic Press, London, 24, 47-69 (1991)  \n \r           Zone of inhibition (mm)Plant Extract in different solventsZone of inhibition of different extracts against Klebsiella Jatropha Extract Stevia extract Lemon grass Extract