利用变温提高汉逊酵母发酵生产谷胱甘肽的研究

Study on temperature-shift strategy on the overproduction of glutathione by Hansenula polymorpha

依据温度对细胞代谢过程的影响,建立了一种提高谷胱甘肽(Glutathione,GSH)产量的方法。通过单因素实验确定了多形汉逊酵母DL-1的最适生长温度、GSH合成的最佳温度和培养基初始pH分别为30、37℃和pH6.5;接着,通过变温调控分批发酵,确定最佳变温条件为:45℃培养16h后将温度降低为37℃继续培养32h,采用该方法摇瓶发酵后GSH总量及GSH含量分别为420.76mg/L和105.19mg/g,比恒温培养(37℃)分别提高了2.34倍和1.27倍,即胞内GSH积累幅度显著升高;最后,通过5L发酵罐扩大培养,生物量和GSH总量分别达42.35g/L和927.68mg/L。实验结果表明该方法可用于提高利用多形汉逊酵母的GSH产量。

An optimal two-stage temperature control strategy was established to improve the yield and the productivity of glutathione in the batch fermentation, according to the effects of temperature on cell metabolism. Through single-factor experiments,the optimum growth temperature of Hansenula polymorpha DL-1, the optimum biosynthesis temperature of GSH and initial pH value of medium were obtained. The result showed that the effect of temperature on glutathione production was different from that on cell growth and maximum glutathione was obtained at 37~C,but the optimum growth temperature was at 30~C. When the temperature shifted from 45~Cto 37℃for further cultivation of 16h,a glutathione yield of 420.16mg/L and an intracellular glutathione content of 105.19mg/g were achieved,which was 2.34 and 1.27 times than that of the constant temperature fermentation,respectively. Subsequently,scale-up studies were conducted in a 5L fermenter,a cell dry weight of 42.35g/L and a glutathione content of 927.68mg/L were achieved. Therefore,the temperature- shift strategy would be a valuable alternative to improve glutathione production.