Bacillus subtilis S21产抗菌脂肽分离鉴定及半固态发酵工艺初探

Isolation, identification of antimicrobial lipopeptide producing by Bacillus subtilis S21 and preliminary study on semisolid fermentation

采用酸沉、甲醇萃取和LH-20层析,从Bacillus subtilis S21发酵菌液纯化出两种抗菌脂肽,通过质谱鉴定两种抗菌脂肽分别为Surfactin和Fengycin,二级质谱表明Surfactin由β-OH脂肪酸链(C_(12-15))和Glu-Leu-Leu-Val-Asp-Leu-Leu(Ile)组成,Fengycin由β-OH脂肪酸链(C_(14-16))和Glu-Orn-Tyr-Thr-Glu-Val-Pro-Gln-Tyr-Ile组成。在前期探索试验基础上,以黄豆为主要培养基质,通过正交实验,确定了B. subtilis S21产Surfactin半固态发酵条件:无机盐体系添加20%,葡萄糖添加5%,发酵时间48 h,发酵温度37℃,验证实验表明此条件下Surfactin产量比landy培养基产量增加了180余倍。

Through acid precipitation,alcohol extraction and LH-20 column chromatography,two kinds of antibacterial lipopeptides were purified from fermented bacteria liquid of Bacillus subtilis S21,and mass spectrum revealed the lipopeptides were Surfactin and Fengycin.MS-MS revealed the sequence of Surfactin was β-OH-C12-15-Glu-Leu-Leu-Val-Asp-Leu-Leu(Ile)and Fengycin wasβ-OH-C14-16-Glu-Orn-Tyr-Thr-Glu-Val-Pro-Gln-Tyr-Ile.On the basis of preliminary exploration test,with soybean as the main culture medium,through orthogonal experiment,the semisolid fermentation conditions of Surfactin producing by B.subtilis S21 were determined,the inorganic ion system was added 20%,the glucose was added 5%,the fermentation time was 48 h and the fermentation temperature was 37℃,the test showed that the yield of Surfactin increased by 180 times more than that of the landy medium.