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›› 2011, Vol. 11 ›› Issue (6): 1038-1043.

• 生化工程专栏 • 上一篇    下一篇

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

乐庸堂 徐岩 穆晓清   

  1. 江南大学生物工程学院 教育部工业生物技术重点实验室,江南大学生物工程学院 教育部工业生物技术重点实验室,江南大学生物工程学院
  • 收稿日期:2011-09-23 修回日期:2011-11-17 出版日期:2011-12-20 发布日期:2011-12-20
  • 通讯作者: 穆晓清

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

YUE Yong-tang XU Yan MU Xiao-qing   

  1. School of Biotechnology,Jiangnan University, Key Laboratory of Industrial Biotechnology of Ministry of Education and School of Biotechnology, SouthenYangtze UniVersity Key Laboratory of Industrial Biotechnology of Ministry of Education and School of Biotechnology, SouthenYangtze UniVersity
  • Received:2011-09-23 Revised:2011-11-17 Online:2011-12-20 Published:2011-12-20
  • Contact: MU Xiao-qing

摘要: 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20 g/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4 g/L,可将底物浓度提高到60 g/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230 g/L,e.e值保持99%以上.

关键词: 近平滑假丝酵母, 不对称还原, (S)-4-氯-3-羟基丁酸乙酯, 底物分批补给, 产物原位萃取

Abstract: 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.

Key words: Candida parapsilosis, asymmetric reduction, (S)-4-chloro-3-hydroxybutyrate ethyl, substrate feeding, in situ product removal

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