One-pot enzymatic production of deacetyl-7-aminocephalosporanic acid from cephalosporin C via immobilized cephalosporin C acylase and deacetylase

The three-enzyme three-pot using immobilized d-amino acid oxidase (DAAO), glutaryl-7-ACA acylase (GLA), and cephalosporin C deacetylase (CAH) was the main strategy for industrial conversion of cephalosporin C (CPC) to deacetyl-7-aminocephalosporanic acid (D-7-ACA), a key pharmaceutical intermediate that is used for the production of 3-vinyl substituted β-lactams antibiotics. In order to simplify this traditional process, we developed an one-pot bi-enzyme catalyzed procedure by coupling immobilized cephalosporin C acylase (CPCA) with immobilized cephalosporin C deacetylase (CAH), which contained two parallel cascade reactions: firstly, hydrolysis of CPC to 7-ACA by CPCA, then deacetylation of 7-ACA to D-7-ACA by CAH (route 1); or firstly, hydrolysis of CPC to deacetyl-CPC (D-CPC) by CAH, then de-α-aminoadipoylation of D-CPC to D-7-ACA by CPCA (route 2). A D-7-ACA yield of 78.39% was achieved in 30 min in a single reactor under the optimized enzyme amount. The specific productivity of D-7-ACA reached 10.85 g g−1 h−1 L−1, which increased approximately 4-fold compared to that obtained from the two-pot enzymatic process. The half-time of immobilized CPCA was approximately 18 h while immobilized CAH was approximately 534 h under the operational conditions. These results demonstrated the industrial feasibility of this one-pot process and its clear advantages towards the traditional three-enzyme three-pot process.