Covalent immobilization of GL-7-ACA acylase on silica gel through silanization

GL-7-ACA acylase was covalently immobilized on silica gels modified with 3-aminopropyltriethoxysilane followed sequentially by treatment with glutaraldehyde. The overall immobilization procedure has several steps such as pretreatment of silica gel, modification of carrier surface by silanization, crosslinking by glutaraldehyde and coupling of GL-7-ACA acylase on the silica gel. In this study, each step in the immobilization procedure was optimized individually to improve the efficiency of the overall immobilization procedure of GL-7-ACA acylase. The optimal conditions of each step were determined as follows; pretreatment by 35% hydrogen peroxide, silanization by 10% 3-APTES at pH 9.0 for 120 min, crosslinking by using glutaraldehyde modified under alkaline (pH 8.5) solution at 60 °C for 20 min and coupling in 1.0 M phosphate buffer (pH 8.0). Especially, the adverse effect of glutaraldehyde, which was observed in this study, was reduced by modification of glutaraldehyde. Through the modification of glutaraldehyde, activity of immobilized GL-7-ACA acylase was increased up to about 60% compared to that obtained by using untreated glutaraldehyde. The immobilized GL-7-ACA acylase was tested for long-term stability and it was found that about 70% activity was retained after using it 20 times.