Evaluation Of The Main Material Basis And Bioactivity Of Bacillus Megaterium L2

In this paper we improved the fermentation medium and the optimum fermentation conditions of the high yield antibacterial active components of Bacillus megateriu L2,and the strain by separation and purification of the material composition,and the identification of the chemical structure.Later a series of determination on bacteriostatic activity and mechanism of bacteriostasis were carried out on the obtained LE4-5 components.Finally only two monomeric compound were isolated and identified and further experiment about their Bacteriostatic activity was performed.The resu Lts were showed as follows:1?The single factor test Plackett-Burmania(PB)design and response surface analysis(RSM)were used to analysis and optimize the fermentation cu Lture conditions of Bacillus megaterium L2.Resu Lts showed that the best fermentation conditions were as follows:glucose content 1.5%,peptone content 25%,beef paste 4.58g/L,sodium chloride 3.23g/L,liquid volume 50 m L/250m L,inocu Lation volume 6%,initial p H 7,temperature 30?.After48 hour fermentation,the diameter of inhibition zone in the optimized fermentation broth was34.99(±0.14)mm,which was close to the predicted value of the equation 34.88mm.By contrast,the diameter was 14.55(±0.43)mm under the original fermentation condition,which indicated that the bacteriostatic activity of the optimized fermentation broth increased by140.48%than original one.2?Using GC-MS analysis predicted that the main substance component of LE4-3 may be ethyl phenylacetate,ethyl linoleate,methyl phenylacetate,ethyl ethyl stearate,etc.While in LE4-5,substance components may be Linoleic acid,methyl linoleate,pentadecanoic acididine,and Oleic acid.3?The separation and purification of the secondary metabolites from the strain were carried out by atmospheric pressure silica gel column chromatography,Sephadex LH-20 gel column chromatography,preparation thin layer chromatography and other techniques.The molecu Lar weight and chemical structure of the two pure products were determined by u Ltraviolet spectroscopy,ESI-MS and nuclear magnetic resonance spectroscopy.The resu Lts showed that BM-3 had a molecu Lar weight of 337 and BM-4 was palmic acid.The bacteriostatic activity of BM-3 and BM-4 was determined by using Ralstonia solanacearum Q11-2 and Agrobacterium tumefaciens T-37 as test strains.The resu Lts showed that BM-3have antibiotic effect on both tested strains,but BM-4 did not work.4?Using Ralstonia solanacearumas as tested strain,the bacteriostatic activity of LE4-5and the mechanism of its inhibition were studied.The changes of electrical conductivity and the leakage of total nucleic acid protein showed that the LE4-5 component had a significant effect on the permeability of the cell membrane,and the LE4-5 component cou Ld produce the effect more quickly on a long enough time scale.The LE4-5 component caused more damage to cell membrane permeability,resu Lting in more effusion of cell contents.According to resu Lts from OD_260nmand OD_280nm,LE4-5 components had a influence on cells,causing outflows of nucleic acid and protein at the beginning.It is suggested that the LE4-5component plays an important role in cell membrane permeability and whether it causes nuclear dissolve needs further study.