| Summary: | 碩士 === 國立高雄大學 === 生物科技研究所 === 98 === An antibiotic-secreting bacterium was isolatetd from boxing glove and named as Y-1. Based on the morphology of the bacterium on PDA plate and Gram staining, Y-1 was identified as a Gram-negative bacterium. From previous research, there are rare Gram-negative bacteria that secret antibacterial substances in natural products. After experimental research, the best fermentation media of Y-1 was 1% glucose, 0.7% soybean flour and 0.3% yeast extract, incubating at 30℃, 150r.p.m for 24 hours, then there would be highest antibacterial substance produced by Y-1. In isolation and purification part, first we used high speed centrifugation to exclude bacteria and high molecular weight substances so we can collect suspension part of crude fermentation liquid. After previous step, we can exclude proteins and most glutinous substances by precipitation of calcium chloride and acetone. Based on polarity of chemical solutions, we used Al2O3 and silica column chromatography to exclude most impurities. Finally, based on molecular weight to process Sephadex LH-20 gel filtration then we can get transparent and purified substances. By thin layer chromatography analysis and production of crystal, it showed that this antibacterial substance has high purity. The final recovery rate is 4.3%. Y-1 has high inhibition to Gram positive bacteria (Bacillus and Staphylococcus aureus) and Candida albicans, especially to Vancomycin-resistant Enterococcus (VRE) and Vancomycin-resistant Staphylococcus aureus (VRSA). Y-1 has 83% antibacterial activity after water bath treatment at 100 ℃ for 60 minutes which means it has great heat-resistant capability, and Y-1 also has ability to clean away biofilms produced by Staphylococcus aureus. After LC-Mass analysis, it showed that Y-1 was a compound with small molecular weight.
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