高通量筛选法在测定腈水解酶活性中的应用

摘要

Nicotinic acid can be produced through direct hydrolysis of 3-cyanopyridine by Nitrilase.By high throughput screening,Nitrilase 131 was selected from 30 nitrilases as an applicable catalyst.It was measured that the conversion rate was the highest when its enzyme activity was 17.6U/mL and its specific activity was 35.2U/mg.At a temperature of 30℃ and a pH value of 8.0,when the initial substrate concentration was 8 ~ 10 g·L-1,3-cyanopyridine could be transformed into nicotinic acid completely with a pure product yield about 85%.The product structure was validated by MS and NMR.This study provides a new idea for the green technology of nicotinic acid's preparation.%腈水解酶可将3-氰基吡啶直接水解生成烟酸,应用高通量筛选法从30种腈水解酶中筛选出131粗酶作为催化剂,测得该酶的酶活17.6U/mL,比酶活35.2U/mg,转化率最高。在温度为30℃,pH为8.0,底物质量浓度为8～10g/L时,腈水解酶131能将3-氰基吡啶完全转化为烟酸,产物纯品收率为85%左右。并用质谱和核磁验证产物结构的正确性。为建立烟酸制备的绿色工艺提供了新的思路。