近平滑假丝酵母全细胞催化不对称还原反应：底(产)物抑制及其解除

Candida parapsilosis Whole Cell Catalyzed Asymmetric Reduction:Inhibition and Removal of Substrate(Product)

以不对称还原4-氯乙酰乙酸乙酯(COBE)制备(S)-4-氯-3-羟基丁酸乙酯(S-CHBE)为模式反应,从实验室保藏菌种中筛选获得高效生物催化剂近平滑假丝酵母(Candida parapsilosis)CCTCC M 203011,底物浓度为20g/L时产物产率达96%,光学纯度(e.e值)大于99%.底(产)物分别具有对细胞(酶)的毒害作用和对酶促反应的抑制作用,是影响不对称还原反应催化效率的主要因素.采用水-有机双相原位萃取和分批补料联用策略,原位萃取产物并控制水相底物浓度不高于4g/L,可将底物浓度提高到60g/L,产率达95%;同时采用高生物量策略,将产物总体积浓度提高到230g/L,e.e值保持99%以上.

Asymmertric reduction synthesis of（S） -4-chloro-3-hydroxybutyrate ethyl（S-CHBE） from 4-chloro-3-oxo-butyrate ethyl（COBE） was introduced as model reaction. Candida parapsilosis CCTCC M 203011,an efficient biocatalyst,was obtained from laboratory preserved strains,which could reduce COBE to S-CHBE with high efficiency and enantioselectivity. 96% in yield and over 99% in e.e were achieved respectively when COBE was 20 g/L. However,the product inhibition to enzymatic reaction and substrate toxicity to enzyme/microbial cell were the main reasons which restricted the reaction efficiency. Coupled with aqueous-organic biphasic system and substrate feeding strategy,the product was removed in situ and the substrate concentration in aqueous phase controlled below 4 g/L,the accumulated concentration of COBE was increased to 60 g/L with 95% in yield. And a higher product titer（approximately 230 g/L） and high optical purity（99%） were achieved through increasing the usage of cell.