利普司他汀产生菌的菌种选育及培养基优化

Mutation Breeding and Optimization of Fermentation Medium of Lipstatin Producing Strain

以产利普司他汀(lipstatin)的毒三素链霉菌(Streptomyces toxytricini)LIPS-352为出发菌(4 125 mg/L),利用紫外(UV)和亚硝基胍(NTG)诱变,获得一株高产突变株LIPS-N-27-46-42,发酵单位为8 122.9 mg/L。单因素试验结果表明,碳源和氮源对利普司他汀的合成有显著影响,其中葵花籽油较大豆油更适合作为利普司他汀合成的前体和碳源。进一步利用Box-Behnken中心组合设计优化发酵培养基组成,通过响应面分析得到较优培养基配方,使利普司他汀的产量达9 859.8 mg/L,与预测值(10 633.1 mg/L)的相对误差为7.27%。通过菌种选育和配方优化,利普司他汀的效价较出发菌株提高了139.0%。

The Streptomyces toxytricini strain LIPS-352, a producer of lipstatin of 4 125 mg/L, was mutated with UV and nitrosoguanidine. A mutant LIPS-N-27-46-42 was obtained, which yielded 8 122.9 mg/L lipstatin in shake flask cultures. The results of one-factor-at-a-time method showed that carbon sources （glycerol） and nitrogen sources （soybean meal and corn gluten） had a significant effect on the production of lipstatin. Meanwhile, it was found that sunflower oil was better than soybean oil in providing precursor and carbon source for lipstatin synthesis. Then the fermentation medium was further optimized by a Box-Behnken design with response surface methodology. The optimal medium was： glycerol 2.5%, soybean meal 3.35%, corn gluten 1.01%, and sunflower oil 7.56%. The yield of lipstatin was 9 859.8 mg/L, which was 21.4% higher than the original fermentation condition and had a relative error of 7.27% to the predicted value（10 633.1 mg/L）. Finally, a 139.0% increase was achieved compared to that of the original strain through mutation breeding and optimization of fermentation medium of lipstatin producing strain.