Properties of immobilised penicillin G Acylase in beta-lactam antibiotic synthesis

The beta-lactam antibiotics are the most important class of antibiotics used today. In the last decade the production routes of these antibiotics have shifted from chemical routes to more environmentally benign routes using the enzyme penicillin G acylase. For both practical and economical reasons the enzyme is immobilised. As the kinetically controlled synthesis reaction is performed in aqueous medium with a hydrolytic enzyme, hydrolysis of the activated acyl donor, e.g. D-phenyl glycine amide and the antibiotic product, e.g. cephalexin, ampicillin, is unavoidable. The immobilisation of the enzyme further promotes the unwanted hydrolysis reactions due to diffusion limitation. This was shown in chapters 3 and 4 for penicillin G acylase immobilised on Eupergit C, Eupergit C 250 L, and Sepabeads. The average catalytic activities per immobilised enzyme molecule were investigated using two active-site titration methods, one using a fluoride ion-selective electrode for the first time (chapter 5). Thus, the work described in this thesis has resulted in a better understanding of the effects of diffusion limitation on the catalytic performance of immobilised penicillin acylase in beta-lactam antibiotic synthesis