安徽工程大学学报2024,Vol.39Issue(1) :8-14.

三相法提取猪肝酯酶及其在制备D-7-ACA中的应用

Extraction of Esterase from Pig Liver by Three-phase Partitioning and Its Application in Preparation of D-7-ACA

张呈彦 程瑶纳 王雨轩 王堃宁 魏胜华
安徽工程大学学报2024,Vol.39Issue(1) :8-14.

三相法提取猪肝酯酶及其在制备D-7-ACA中的应用

Extraction of Esterase from Pig Liver by Three-phase Partitioning and Its Application in Preparation of D-7-ACA

张呈彦 1程瑶纳 1王雨轩 1王堃宁 1魏胜华2
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作者信息

  • 1. 安徽工程大学 生物与食品工程学院,安徽 芜湖 241000
  • 2. 安徽工程大学 生物与食品工程学院,安徽 芜湖 241000;微生物发酵安徽省工程研究中心,安徽 芜湖 241000;安徽省工业微生物分子育种工程实验室,安徽 芜湖 241000
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摘要

酯酶广泛存在于动物、植物和微生物中,具有作用范围广、底物特异性高等特点.该酶能够催化许多含有酯、酰胺、硫酯或乙酰基的化合物的水解与合成,在生物催化领域具有重要的用途.对来源于猪肝中的酯酶采用三相分离的方法进行了提取,并利用猪肝酯酶的水解特性,研究了其催化7-氨基头孢烷酸(7-ACA)脱乙酰基制备去乙酰基-7-氨基头孢烷酸(D-7-ACA)的过程.主要研究结果:确立了三相法分离提取猪肝酯酶的方法,考察了有机溶剂种类、有机溶剂用量、硫酸铵浓度以及体系的pH和提取温度等条件对三相法分离提取猪肝酯酶的影响.实验结果表明在硫酸铵质量分数为45%、异丁醇与水相之比为1∶1、pH7.0、温度为30℃的条件下,酶活回收率为54.84%,纯化倍数为6.32倍.以猪肝酯酶为催化剂,研究了猪肝酯酶催化7-ACA脱乙酰基制备D-7-ACA,在pH7.5、温度35℃、底物浓度2%、酶添加量30 U/mL的反应条件下,7-ACA转化率达到94.33%.

Abstract

Esterase is widely distributed in animals,plants,and microorganisms.It has wide substrate spe-cificity and can catalyze hydrolysis and synthesis of a variety of compounds containing esters,amides,sul-fones,or acetyl groups.Esterase has important applications in the field of biocatalysis.This paper studies the extraction of the enzymology of esterase from pig liver,and its ability to catalyze the deacetylation of 7-aminocephalosporanic acid to produce deacetyl-7-aminocephalosporanic acid.The main results are as follows:Extraction of esterase from pig liver by three-phase partitioning has been established.The effects of different organic solvents,organic solvents dosage,ammonium sulfate concentration,and the pH of the system and extraction temperature on the three-phase extraction of pig liver esterase have been investigated.The experimental results showed that under the conditions of ammonium sulfate mass frac-tion 45%,isobutanol to water ratio of 1∶1,pH 7.0,and temperature 30 ℃,the enzyme activity recovery rate was 65.79%,and the purification factor was 2.31.With pig liver esterase as a catalyst,the prepara-tion of D-7-ACA by deacetylation of 7-ACA was studied.The conversion rate of 7-ACA reached 94.33%when the preparation conditions were pH 7.5,temperature 35 ℃,substrate concentration 2% and en-zyme addition was 30 U/mL.

关键词

猪肝酯酶/三相法/7-氨基头孢烷酸/去乙酰基-7-氨基头孢烷酸

Key words

pig liver esterase/three-phase partitioning/7-ACA/D-7-ACA

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基金项目

安徽省高等学校自然科学研究项目(KJ2021A0512)

出版年

2024
安徽工程大学学报
安徽工程大学

安徽工程大学学报

影响因子:0.289
ISSN:2095-0977
参考文献量20
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