以豆粕粉为氮源的枯草芽孢杆菌液态发酵生产纳豆激酶

Process optimization and validation of Bacillus subtilis liquid fermentation for nattokinase production using soybean cake powder as the nitrogen source

枯草芽孢杆菌液态发酵通常采用蛋白胨作为氮源生产纳豆激酶,导致生产成本过高,发酵液中酶活性较低。采用价格低廉的豆粕粉作为氮源不仅大幅度降低了原料成本,而且显著提高了发酵液中的酶活性。在摇瓶实验中优化确定豆粕粉最佳添加量,并在此基础上优化确定了最佳接种量、发酵液初始p H、发酵时间。最终摇瓶发酵液中酶活达到5 471 IU/m L,是等量胰蛋白胨作为氮源发酵的3.6倍。上述摇瓶发酵参数进一步在7 L发酵罐和200 L中试水平发酵罐进行验证,并优化确定发酵过程中控制溶氧水平为30%,纳豆激酶活性分别达到6 717 IU/m L和4 236 IU/m L。

Tryptone was selected as the nitrogen source in traditional Bacillus subtilis liquid fermentation, which led to high production costs and low nattokinase activities. In this study, tryptone was replaced with low-cost soybean cake powder as the nitrogen source in Bacillus subtilis liquid fermentation to produce nattokinase. The amountof soybean cake powder, the optimal inoculum size, media initial pH and fermentation time were confirmed after optimization at shake flask level. The nattokinase activities in shake flask level arrived at 5 471 IU/mL, which was 3.6 times of that from fermentation with same amount of tryptone as nitrogen source. The above parameters at shake flask fermentation level were further verified in 7 L and 200 L fermentation tank, and the optimal dissolved oxygen level was confirmed to be 30%. The nattokinase activities in 7 L and 200 L fermentation tank respectively reached 6 717 IU/ mL and 4 236 IU/mL.