Breeding Of Inosine-Producing Strain And Studies On Its Fermentation Conditions

According to the theory of metabolic control fermentation, the dissertation focuses on the breeding and fermentation condition of inosine producing strain. The main research contents and results are as follows:(1) The method of ion exchange chromatography was defined to determinate inosine concentration quantitatively in the fermentation broth. The separation conditions of this method was optimized and compared with paper chromatography method. Both accuracy and precision are better than that of paper chromatography.(2) One inosine high-producing strain TX-3(Ade+Thi+8-AG +SG) was derived from Bacillus subtilis TY-21 with DES treatment, which could produce inosine 12.41 g/L under the fermentation condition without being optimized.(3) The batch fermentation condition of strain TX-3 in shake flask was studied in this dissertation. The proportions of seed medium and fermentation medium were optimized by pattern recognition and uniform design experiment. The seed culture conditions and fermentation conditions were also studied. Under the optimum condition, strain TX-3 could produce inosine 18.94g/L by shaking flask batch fermentation. Meanwhile the fed batch fermentation conditions were studied. The production of inosine under optimized fed batch fermentation was 24.79g/L.(4) The metabolic flux analysis was applied in inosine fermentation process based on the experimental data from batch fermentation in 5-liter fermentor. Based on the flux balance principle, the metabolic flux analysis method of inosine batch fermentation was defined .The whole metabolic process of inosine was analysed combined with fermentation process curve. Using metabolic flux balance model of inosine synthesis by Bacillus subtilis model, the metabolic flux distributions during the middle and last period were determination and the optimal flux distributions were calculated by linear program of MATLAB software .The flux analysis indicated that reducing the metabolic flux of EMP pathway were useful to Ile synthesis; Increasing activities of key enzymes in inosine biosynthesis pathway would increase inosine production. Reducing byproduct synthesis were important for inosine high producing, as well as the heightening soluble oxygen level.