Nie W, Wei G, Du G, Li Y, Chen J
Key Laboratory of Industrial Biotechnology, Ministry of Education, Southern Yangtze University, Wuxi, China.
Lett Appl Microbiol. 2005;40(5):378-84. doi: 10.1111/j.1472-765X.2005.01687.x.
To study the effect of low pH stress on glutathione (GSH) synthesis and excretion capability of GSH fermentation production in Candida utilis.
When C. utilis WSH 02-08 was cultivated in a glucose-ammonium sulfate medium without pH control, GSH leakage occurred when the pH of the medium decreased to 1.5. However, analysis of the cell viability indicated that the cells were not lysed. To further study the effect of low pH stress on GSH production, pH-controlled batch cultures were conducted, where the pH was switched from 5.5 to 1.2 at 24 h and maintained at 1.2 for 6 h. Nearly all intracellular GSH was leaked into the medium and the cell viability decreased dramatically, conceiving a long-term exposure of strain WSH 02-08 at low pH environment led to a complete cell lysis. A critical point (treated at pH 1.2 for 3 h) was experimentally determined, where most cells were alive but suffering a low pH stress. Low pH-stressed C. utilis cells displayed an increased intracellular GSH synthesis and export capability, which protected the cells against short-term low pH treatment.
Using this knowledge, a low pH-stress strategy was developed and applied in fed-batch production of GSH and 197.3 mg l-1 of GSH was secreted into the medium. The GSH-specific production yield could be increased from 2.11 to 2.67% (w/w), and the total GSH concentration could reach 737.1 mg l-1 and increased by 24.9%.
This is the first report of GSH secretion of C. utilis at low pH. This study demonstrated the importance of the physiology-based fermentation strategy in the production of useful metabolites.
研究低pH胁迫对产朊假丝酵母谷胱甘肽（GSH）合成及GSH发酵生产排泄能力的影响。
当产朊假丝酵母WSH 02 - 08在无pH控制的葡萄糖 - 硫酸铵培养基中培养时，培养基pH降至1.5时会发生GSH泄漏。然而，细胞活力分析表明细胞未裂解。为进一步研究低pH胁迫对GSH产生的影响，进行了pH控制的分批培养，在24 h时将pH从5.5切换至1.2并维持6 h。几乎所有细胞内的GSH都泄漏到培养基中，细胞活力急剧下降，推测菌株WSH 02 - 08在低pH环境下长期暴露会导致细胞完全裂解。通过实验确定了一个临界点（在pH 1.2处理3 h），此时大多数细胞存活但遭受低pH胁迫。低pH胁迫的产朊假丝酵母细胞表现出细胞内GSH合成和输出能力增强，这保护细胞免受短期低pH处理。
基于此知识，开发了一种低pH胁迫策略并应用于GSH的补料分批生产，197.3 mg l-1的GSH分泌到培养基中。GSH的比生产产量可从2.11%提高到2.67%（w/w），GSH总浓度可达737.1 mg l-1，提高了24.9%。
这是关于产朊假丝酵母在低pH下分泌GSH的首次报道。本研究证明了基于生理学的发酵策略在有用代谢产物生产中的重要性。
