Study On Antimicrobial Componegnts Of Secondary Metabolites Produced By Bacillus TH07

Secondary metabolites from microorganisms are important source for natural antibacterial substances.With the development of scientific research,more and more metabolic compounds had been discovered,more and more duplication problems for analysis and identification of active substances,which is extremely detrimental to the drug research and development.In order to explore new natural antimicrobials,many laboratories have turned their attention to extreme environmental microbes that are more promising to discover new antimicrobial compounds.Thermophilic bacteria,a kind of extreme environmental microbe,may produce novel active secondary metabolites because of its particular growth environment.One strain of moderately thermophilic bacteria TH07(Bacillus)were studied here.The metabolites of TH07 exact by ethyl acetate had obvious inhibitory effect on TH08(Actinomycete).In order to trace the active ingredient that inhibits the growth of TH08,and to analyze the antibacterial mechanism,the following work had been done.Firstly,the fermentation conditions of TH07 were optimized.It does not grow at 28 ? and 37 ?,and grows slowly at 45 ?,but its growth is positively related to temperature between 50?65 °C.Based on results of response surface methodology,the optimum fermentation conditions for TH07 were obtained as following:initial medium pH of 9,medium volume of 381 mL with one liter conical flask,and inoculums size of 6.37%(V/V),fermentation temperature of 55? fermentation time 36 h.With these conditions,crude secreted metabolites of TH07 were extracted by ethyl acetate.Secondly,the extracted crude secreted metabolites were further separated,purified and primary identified.Three compounds were obtained from the crude extracts by thin layer chromatography and high performance liquid chromatography.Their structures were identified by mass spectrometry,nuclear magnetic resonance spectroscopy and infrared spectroscopy.They were Cyclo(tyrosine-proline),Cyclo(valine-proline)and Cyclo(phenylalanine-proline).The result revealed that three compounds had no antibacterial effects to the indicator strain of TH08.A mixture obtained by preparative liquid chromatography has a significant inhibitory effect on TH08.Its composition was roughly analyzed by gas chromatography mass spectrometry,and the results showed that the mixture contains long chain fatty acids,esters,amides and phenols,which suggests that these ingredients may be involved in the growth inhibition of TH08.