Phase transfer bioconversion of penicillin G into 6-APA by immobilized penicillin acylase in recycling aqueous two-phase systems with a light-sensitive copolymer

Abstract

Immobilized penicillin acylase was used for bioconversion of penicillin G into 6-APA in aqueous two-phase systems consisted of a light-sensitive polymer (PNBC 300000) and Dextran 20000. Partition coefficients of 6-APA was found to be about 8.4 in the presence of 50 mmol/L KCl. Results of enzyme kinetic studies showed that the reaction reached equilibrium at 6 h or so. The conversion ratio of PG(Na) were 82.6% (pH 7.8, 2.2% substrate concentration and 20°C), which was about 20% higher than that of the control in the reaction of single aqueous phase buffer. Three repeated batches of reactions were carried out. In aqueous two-phase systems, substrate penicillin G, products 6-APA and phenylacetate acid are biased in top phase, while immobilized penicillin acylase is completely partitioned in bottom. Substrate penicillin G entering into bottom phase, is catalyzed to 6-APA and phenylacetate acid, then the products diffused into top phase. Finally, inhibition of substrate and products was removed which led to the in improvement of products yield. Moreover, immobilized enzyme has higher efficiency than immobilized cells and occupy smaller volume. Comparing with free enzyme, immobilized enzyme has higher stability, longer working life, completely partitioned in bottom phase and the capacity of recycling. Bioconversion in two-phase systems using immobilized penicillin acylase showed outstanding advantage. The light-sensitive copolymer forming aqueous two-phase systems could be reused by laser radiation at 488 nm, with the recovery of 95%-98%.