Development of enzymatic resolution process of pantolactone by fungal lactonase

提出了一种利用尖孢镰刀菌内酯水解酶立体特异性水解D-pantolactone的光学分解外消旋体的新酶解工艺。尖孢镰刀菌对该酶的产率较高，含有该酶的细胞可重复用于该水解反应。在pH自动维持在7.0的条件下（D-pantoic acid的光学纯度为96% e.e.e），与oxysporum细胞孵育24h后，pantolactone外消旋混合物（700mg/ml）中的D-Pantolactone几乎被化学定量水解为D-pantoic acid。在实践中，D-pantolactone在消旋混合物中的水解是通过固定的尖孢霉细胞作为催化剂来实现的。将真菌细胞包埋在海藻酸钙凝胶中，可制备具有高水解活性的稳定催化剂。将固定的细胞在含有350g/l DL-pantolactone的反应混合物中，在30‰的温度下（pH值为6.8 ~ 7.2），自动控制pH值，培养21h， 90-95%的D-pantolactone被水解（光学纯度为90-97% e.e）。经过180次（即180天）的重复反应，固定化菌丝保留了90%以上的初始活性。外消旋环内酯的酶解过程可以跳过常规化学分解过程中几个繁琐的步骤，这是一个相当大的实用优势。

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.