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Development of enzymatic resolution process of pantolactone by fungal lactonase

真菌内酯酶分解泛内酯工艺的研究进展

基本信息

批准号：
08556013
负责人：
SHIMIZU Sakayu
金额：
$ 6.4万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08556013/
关键词：
Lactonase Fusarium oxysporum Pantolactone Enzymatic resolution Immobilized enzyme

项目摘要

A novel enzymatic process for the optical resolution of racemic pantolactone through the stereospecific hydrolysis of D-pantolactone by lactonohydrolase of Fusarium oxysporum is developed. F.oxysporum showed high productivity of the enzyme and the cells containing the enzyme could be used repeatedly for this hydrolysis reaction. D-Pantolactone in a racemic mixture of pantolactone (700mg/ml) was almost stoichiometrically hydrolyzed to D-pantoic acid on incubation with F.oxysporum cells for 24h under conditions in which the pH was automatically maintained at 7.0 (optical purity for D-pantoic acid, 96% e.e.). In practice the hydrolysis of the D-pantolactone in a racemic mixture is achieved by immobilized cells of F.oxysporum as the catalyst. Stable catalysts with high hydrolytic activity can be prepared by entrapping the fungal cells into calcium alginate gels. When the immobilized cells were incubated in a reaction mixture containing 350g/l DL-pantolactone for 21h at 30ﾟC under the conditions of automatic pH control (pH 6.8-7.2), 90-95% of the D-pantolactone was hydrolyzed (optical purity, 90-97% e.e.). After repeated reactions for 180 times (i.e., 180 days), the immobilized mycelia retained more than 90% of their initial activity. The enzymatic resolution process of racemic pantolactone can skip several tedious steps which are necessary in the conventional chemical resolution process, and this is a considerable practical advantage.

开发了一种通过尖镰孢内酯水解酶立体定向水解 D-泛内酯来光学拆分外消旋泛内酯的新酶法。尖孢镰刀菌显示出该酶的高生产力，并且含有该酶的细胞可以重复用于该水解反应。泛解酸外消旋混合物 (700mg/ml) 中的 D-泛解酸在 pH 自动维持在 7.0 的条件下与尖孢镰刀菌细胞孵育 24 小时，几乎按化学计量水解为 D-泛解酸（D-泛解酸的光学纯度，96% e.e.）。在实践中，外消旋混合物中的D-泛内酯的水解是通过固定化尖镰孢细胞作为催化剂来实现的。通过将真菌细胞捕获到海藻酸钙凝胶中可以制备具有高水解活性的稳定催化剂。当固定化细胞在含有350g/l DL-泛内酯的反应混合物中于30℃、自动pH控制（pH 6.8-7.2）条件下孵育21h时，90-95%的D-泛内酯被水解（光学纯度为90-97% e.e.）。重复反应180次（即180天）后，固定化菌丝体保留了初始活性的90%以上。外消旋泛内酯的酶法拆分过程可以跳过传统化学拆分过程中必需的几个繁琐步骤，这是一个相当大的实用优势。