耦合透性化辅酶循环制备手性(R)-苯基乙醇

Coupling Permeabilized Cofactor Recycling for Chiral (R)-Phenethanol Preparation

摘要:

耦合透性化博伊丁假丝酵母（Candida boidinii, C. boidinii）加强近平滑假丝酵母（Candida parapsilosis, C. parapsilosis）内羰基还原酶（Carbonyl reductase, CR）催化反应中的辅酶循环与反复利用，构建有效的手性(R)-苯基乙醇透性化细胞耦合制备体系。实验结果如下：透性化的C. parapsilosis细胞底产物渗透效果增强，底物转化率由33.5%提升到43.8%；进一步外添加辅酶后，底物转化率由43.8%提高到了52.1%；耦合透性化C. boidinii细胞并同时外添加辅酶可大大增强辅酶循环效率，底物转化率可提高至71.8%。优化透性化细胞耦合体系反应条件，当以0.2 g近平滑假丝酵母（C. parapsilosis）为基准时，在C. parapsilosis与C. boidinii细胞质量比为1.5:1，底物浓度为10 mmol/L，辅酶浓度为底物浓度的1%（即0.1 mmol/L）时，底物转化率可达78.2%。离心法收集细胞进行多批次反应，当反应12批次时，耦合体系仍然保持有55%底物转化率，此时总的底物转化率达70.3%，产物经测定为(R)-苯基乙醇，对映体过量值(e.e.值)达98.2%。

Abstract:

The cofactor recycling and reutilization of carbonyl reductase catalysis in Candida parapsilosis (C. parapsilosis) was reinforced by coupling with permeabilized cell of Candida boidinii (C. boidinii). Thus an effective permeabilized cell coupling system was constructed for preparation chiral (R)-phenethanol. The experiment results were as follows: substrate and production infiltration ability of permeabilized C. parapsilosis were enhanced, its substrate conversion rate was improved from 33.5% to 43.8%; the substrate conversion rate was further improved from 43.8 to 52.1% by adding cofactor in addition; the cofactor recycling efficiency could be greatly enhanced by coupling with permeabilized C. boidinii with adding cofactor in addition, the substrate conversion rate could reach as 71.8%. The permeabilized cell coupling system reaction conditions were optimized as: when the C. parapsilosis cell mass was 0.2 g, the cell mass ratio of C. parapsilosis and C. boidinii was 1.5:1, substrate concentration was 10 mmol/L, cofactor concentration was 1% of substrate concentration (as 0.1 mmol/L), its substrate conversion rate could reach as 78.2%. The cells were re-collected by centrifugation for multibatches reaction. The substrate conversion rate could still keep at 55% when the coupling system was reacted for 12 batches, the total substrate conversion rate was 70.3%, the production was determined as (R)-phenethanol, its enantiomeric excess(e.e.) was 98.2%.