耐盐芽孢杆菌B11抗菌活性物质的分离纯化及其特征分析

对耐盐芽孢杆菌B11产生的抗菌活性物质进行了分离纯化及特性研究。从盐碱土壤中分离得到一株对金黄色葡萄球菌具有强拮抗活性的耐盐芽孢杆菌B11,其发酵液中抗菌活性物质具有良好的热稳定性、p H稳定性和紫外光照稳定性,100℃加热1 h,抑菌活性仅下降11.0%,且在p H为3和11环境下,抑菌活性也仅下降11.4%和10.9%,对酸碱环境具有耐受性。发酵液经预处理、离心、阳离子阴离子交换层析,经正丁醇抽提和半制备高效液相色谱得到活性物质A1和A2,其中A1分子量为1423.8 u且带有多电荷,氨基酸分析仪鉴定A1为非核糖体多肽(NRPS),对金黄色葡萄球菌的最小抑菌浓度为8.0μg/m L;A2活性很弱且含量较低,初步鉴定为小分子抗菌物质,分子量为373 u。耐盐芽孢杆菌B11的抗菌代谢产物应用于食品防腐具有很大的潜力。 

Isolation and purification of antimicrobial substance from Bacillus sp.B11 and analysis of its properties

Study the purification and characterization of antimicrobial substance from Bacillus sp.B11.A halotolerant bacterium B11 isolated from saline- alkali soil showed strongly antagonistic against Staphylococcus aureus,and its antimicrobial substance showed excellent stability towards p H,temperature and ultraviolet.The antimicrobial activity was not influenced disposing for 1 h under 100 ℃,and only 11.0% loss of activity was suffered.After treatment at p H3 and p H11,antimicrobial activities decreased 11.4% and 10.9%,respectively,which indicated the antimicrobial substance of Bacillus sp. B11 was stable to acid and alkali. Fermentation broth was used to isolate antimicrobial substances with a procedure involving pretreatment of fermentation broth,cation- exchange chromatography,anion- exchange chromatography,rotary evaporation after butanol extraction,and preparative HPLC on C18 column. Two elution peaks,named A1 and A2,which possessed the antimicrobial activity were eluted with preparative HPLC,and the molecular weight of multi- charge A1 was 1423.8 u.The purified antimicrobial peptide A1 was determined as non- ribosomal synthetic peptide( NRPS) by amino acid analyzer,and its minimum inhibitory concentration against Staphylococcus aureus was 8.0 μg / m L.At the same time,the antimicrobial substance A2 was determined as low- molecular- weight antibiotic substance combined its antimicrobial activity and molecular mass,and its molecular mass was 373 u. Therefore,the antibacterial metabolites from halotolerant bacterium B11 may have potential application in food industry.