Study On The Screening Of High-yielding Strain And Optimization Of Submerged Fermentation Of Nattokinase

Nattokinase is one kind of serine protein metabolized by Bacillus subtils natto. In 1987, Japan scholar Xu Jianyangxing discovered it in natto food and named it nattokinase. Nattokinase has good effect on dissolving thrombus, compared with traditional thrombolytic drug like streptokinase and urokinase, nattokinase has many advantages such as pharmacological action direction and diversification, longer half-life in vivo, smaller relative molecular weight to be absorbed by gastrointestinal system directly, higher security, cheaper price and so on. The study of how to promote enzyme activity of nattokinase is becoming a focus in the whole professional fields.This experiment got one kind of nattokinase high-producing strains by isolating strains from natto food which imported from Japan. We processed these strains by UV mutagenesis and selected strains by Bacillus subtils natto death time curve. After many repeated experiments, obtained the strains which with higher enzyme producing ability. (We temporarily designated YNS-3).The strains which were selected after mutagenesis with higher enzyme producing ability were considered as parent strains, optimized the nattokinase components and the conditions of fermentation. The results are as follows: the best nitrogen source of the fermentation is bean sprout powder, adding 8g/100mL; the best carbon source of the fermentation is glucose, adding 2%; the optimum concentration of MgSO4 is 0.05%; the optimum fermentation temperature is 38℃, pH=7; adding 2% strains which are fermented 24 hours; when we use shaking flask to ferment, the best rocking bed velocity is 150r/min, the volume of Erlenmeyer-flask is 200mL/500mL. After three-day fermenting, nattokinase reaches the highest enzyme activity.At last, the comparative experiments validate that the strains which are fermented in optimization culture medium have higher enzyme activity. Enzyme activity can reach 846IU/mL, 53.23% more than original strains that are fermented in common medium. It can prove that using the selected and mutagenesis strains and the optimization culture medium, the purpose of getting the strains with higher enzyme producing ability and better conditions of the fermentation is achieved.