Optimization of medium composition for the production of nattokinase by Bacillus natto NLSSE

Nattokinase, primarily isolated from a traditional fermented food in Japan---“ Natto” , is a novel fibrinolytic enzyme. It is considered to be a promising agent for thrombosis therapy. The composition of the medium for the production of nattokinase by Bacillus natto NLSSE was optimized with a stepwise strategy. Effects of various nitrogen and carbon sources were investigated. Among the tested nitrogen sources, soy peptone was found to be most promising. As to carbon sources, maltose was found to be most effective for nattokinase production. Fractional factorial design (FFD) was applied to elucidate the key ingredients in the media and the results indicated that the soy peptone and yeast extract gave significant negative influence on nattokinase production, while the calcium chloride gave strong positive effect. Central composite design (CCD) was employed to search for the optimal concentration of the three key components, and the experimental results were fitted with a second-order polynomial model at 98% level (p<0.02). Canonical analysis results showed that the response surface had a maximal point. According to the model, the predicted maximal nattokinase volumetric concentration was about 1270 u/ml and the corresponding concentration of soy peptone, calcium chloride and yeast extract were around 8.28 g/ml, 0.64 g/ml, 0.74 g/ml, respectively. The trial checking the developed medium showed a high nattokinase activity of 1300±30 u/ml (average of three repeats).