纳豆菌液态发酵荞麦产纳豆激酶及其代谢特性分析

对纳豆枯草芽孢杆菌（Bacillus subtilis natto）液态发酵荞麦产纳豆激酶揺瓶、2.5 L发酵罐条件进行优化。对比自制大豆分离蛋白酶解物与商业大豆蛋白胨作为补充氮源时菌体生长规律以及代谢物质（酶、可溶性蛋白、还原糖、多酚及抗氧化物质）变化规律。纳豆菌液态发酵荞麦产纳豆激酶2.5?L发酵罐最优条件为荞麦浸泡6?h后，按料液比1∶10（g/mL）加水打浆，加入0.4%?α-淀粉酶，90?℃加热40?min，补充NS37071酶解12?h时所得酶解物，调节发酵培养基pH?7.0，接种量3%，通气量3.5?L/min，转速300?r/min，装液量1.2?L，发酵36?h。与商业大豆蛋白胨相比，补充大豆分离蛋白酶解物时，纳豆菌生长对数期较长，可溶性蛋白与还原糖的消耗量较大，在36?h趋于平稳，纳豆激酶活力持续上升至36?h达到最大值，酚类物质比溶出速率在12?h达到最大值，抗氧化物质比生成速率在6?h达到最大值。以荞麦为原料，补充自制大豆分离蛋白酶解物，通过优化纳豆菌液态发酵条件，可制备具有高纳豆激酶活力（152.5?FU/mL）、富含谷物多酚（0.109?mg/mL）且具有强抗氧化活性（27.43?μmol/mL）的发酵产物。

Nattokinase Production and Metabolic Characteristics during Submerged Fermentation of Buckwheat Using Bacillus subtilis natto

The submerged fermentation conditions of buckwheat for nattokinase production in shake flasks and a 2.5 L fermentor using Bacillus subtilis natto were optimized. Changes in bacterial growth and the metabolism of nutrients (nattokinase, soluble protein, reducing sugar, phenolics and antioxidants) during fermentation using soybean protein isolate hydrolysate and commercial soybean peptone as a supplementary nitrogen source were comparatively studied. The optimized fermentation conditions in a 2.5 L fermentor were obtained as follows: 6 h of buckwheat soaking, pulping at a solid-to-water ratio of 1:10 (g/mL), addition of 0.4% α-amylase, heating for 40 min at 90 ℃, supplementation of 12 h hydrolyzed soybean protein isolate (SPI) with a commercial protease preparation (NS37071), pH adjustment to 7.0, inoculum size 3%, ventilation rate 3.5 L/min, stirring speed 300 r/min, medium volume 1.2 L, and 36 h fermentation. Supplementation of SPI hydrolysate prolonged the logarithmic growth phase and led to increased consumption of reducing sugar and soluble protein as compared with soybean peptone. Residual reducing sugar and soluble protein tended to level off at 36 h of fermentation with supplementation of SPI hydrolysate, and simultaneously the maximum nattokinase activity was obtained. In addition, the specific dissolution rate of phenolics and the specific formation rate of antioxidants reached a maximum at 12 and 6 h, respectively. The nattokinase activity of the fermentation products obtained under the optimized conditions was 152.5 FU/mL, which were rich in phenolics (0.109 mg/mL) and antioxidants (27.43 μmol/mL).