Next Level of p-Phenylene Diisothiocyanate-based Covalent Immobilization of β-d-Galactosidase: technical optimization an application

Abstract In this study, a continuous lactose hydrolysis process in a fixed-bed reactor was developed using β-galactosidase covalently immobilized on resin beads via 1,4-phenylenediisothiocyanate (PDC) as linker. Process conditions, including temperature, enzyme loading, perfusion speed, and repeated perfusion of the same substrate solution were systematically varied. The highest glucose yields were obtained at 55°C, with increased yields observed at low perfusion speeds, high enzyme loadings, and upon repeated perfusions. Under optimized cycle perfusion over 72 h, final lactose conversion reached approximately 90% at 37°C and 80% at 22°C. A hydrolysis process in a fixed-bed reactor was successfully established, although further optimization is required.