L-乳酸高产菌株的选育及发酵条件的研究

Study on the Breeding and Cultivation of L-Lactic Acid Producing Strain

采用亚硝基胍诱变筛选高产L-乳酸米根霉菌株,对其高产机理进行分析,并研究了不同条件下突变株发酵产乳酸的情况。研究结果表明:亚硝基胍诱变米根霉As3.3462,获得高产L-乳酸的正向突变株。通过对该高产突变株和原始株在不同时段的产酸量和产乙醇量进行比较,发现突变株的碳代谢系统中丙酮酸分支点流入乙醇途径和乳酸途径的流量改变,46h时乙醇产量从11.476g/L减少到7.038g/L,乳酸产量从64.8g/L增加到73.8g/L。运用正交试验设计理论,对高产L-乳酸高产菌株的发酵培养基进行了优化,确定了最佳发酵培养基为(g/L):葡萄糖160,尿素2,KH2PO40.3,MgSO4·7H2O0.25,ZnSO4·7H2O0.08。

Rhizopus oryzae is an important fungi which can metabolize to produce high optical purity L- lactic acid. In this study, N-methyl-N＇-nitro-N-nitrosoguanidine （NTG） and diethyl sulphate （DES） were used to mutagenize Rhizopus oryzae As3. 3462 and a mutant with high-producing L-lactic acid was screened out. The fermentation processes of the two Rhizopus oryzae strains were researched and the data indicated that the flux of lactic acid and ethanol was varied at the fermentation stage of 46hr, namely, the concentration of ethanol decreased from 11. 476 g/L to 7.038 g/L, but the concentration of lactic acid increased from 64.8g/L to 73.8 g/L,these changes indicated that the flux of pyruvate was shifted. From the shake-flask experiments, the optimal fermentation medium consisted of the following：glucose 160g,/L, urea 2 g/L, and rotation 220 r/min. The optimal fermentation medium of mutant strain with a higher and stable yield of L-lactic acid were studied by orthogonal design theory. From the shake-flask experiments, the optimal fermentation medium consisted of the following （g/L） ：glucose 160, Urea 2, KHEPO4 0. 3, MgSO4 · 7H2O 0. 25 ,ZnSO4· 7H2O 0. 08.