枯草芽孢杆菌DC⁃1产维生素K2的发酵条件优化

为提高微生物发酵法生产维生素K2 的产量和多样性，对枯草芽孢杆菌DC?1 的发酵条件进行优化。采用单因素试验对其发酵条件包括碳源、氮源和无机盐进行初步优化，通过Plackett?Burman（PB）试验筛选出对DC?1产维生素K2具有显著性影响的因素，应用响应面Box?Behnken 设计优化获得DC?1 的最优发酵条件。结果表明，DC?1产维生素K2的最优发酵条件为甘油添加量36.5 g/L、糖蜜添加量31.8 g/L、黄豆粉添加量32.5 g/L、KH2PO4浓度0.3 g/100 mL、K2HPO4·3H2O 浓度0.8 g/100 mL、MgSO4·7H2O 浓度0.7 g/100 mL、NaCl 浓度0.5 g/100 mL。在该条件下，MK?7 产量达到94.66 mg/L，比初始发酵条件提高了48.04%；同时菌株DC?1产维生素K2 的多样性增加，经液相色谱?质谱联用（liquid chromatograph mass spectrometer，LC?MS）鉴定能同时产5 种MK 同系物，分别为MK?2、MK?3、MK?4 和MK?7，其中一个类似物未能在高分辨率质谱中找到，根据保留时间推测其为MK?5 或MK?6，可为维生素K2 的工业化生产和探索维生素K2 同系物的生物活性奠定基础。

Optimization of Fermentation Conditions for Vitamin K2 Production by Bacillus subtilis DC⁃1

In order to improve the yield and diversity of vitamin K2 produced by microbial fermentation，the fermentation conditions of Bacillus subtilis DC?1 were optimized.The fermentation conditions including carbon source，nitrogen source，and inorganic salt were optimized by single factor experiment.The factors that had a significant influence on vitamin K2 produced by DC?1 were screened by the Plackett?Burman（PB）experi?ment，and the optimal fermentation conditions of DC?1 were obtained by response surface Box?Behnken de?sign.The results showed that the optimal fermentation conditions for vitamin K2 produced by DC?1 were glyc?erol of 36.5 g/L，molasses of 31.8 g/L，soybean flour of 32.5 g/L，KH2PO4 of 0.3 g/100 mL，K2HPO4·3H2O of 0.8 g/100 mL，MgSO4·7H2O of 0.7 g/100 mL，and NaCl of 0.5 g/100 mL.Under these conditions，the yield of MK?7 reached 94.66 mg/L，which was 48.04% higher than the initial fermentation conditions.Meanwhile，the diversity of vitamin K2 produced by strain DC?1 increased，and a liquid chromatograph mass spectrometer（LC?MS）identified that it could produce five MK homologues simultaneously，namely MK?2，MK?3，MK?4，and MK?7，of which one analogue could not be found in the high?resolution mass spectrometry and was presumed to be MK?5 or MK?6 based on the retention time.This could lay a foundation for the industrial production of vi?tamin K2 and the exploration of the biological activity of vitamin K2 homologues.