Covalent Immobilization of Uricase on Amino-Functionalized Magnetic Bentonite: Enhanced Stability and Reusability for Enzyme Applications

Enzyme immobilization on efficient and reusable supports is a promising strategy to improve enzyme stability and expand their applications in biocatalysis and biosensing. In this study, uricase was covalently immobilized onto amino-functionalized magnetic bentonite (Fe۳O۴@BNT-NH۲-GA) using glutaraldehyde as a crosslinking agent. The successful fabrication of the magnetic support and enzyme immobilization were confirmed by FTIR, XRD, and SEM analyses. The Bradford assay revealed an immobilization efficiency of ۷۴.۹%, indicating effective attachment of uricase onto the functionalized support. The catalytic properties of free and immobilized uricase were evaluated under different temperature and pH conditions. Both forms exhibited maximum activity at ۴۰ °C and pH ۸.۵; however, the immobilized enzyme demonstrated enhanced tolerance toward thermal and pH variations compared with the free enzyme. The improved stability was attributed to the covalent attachment of uricase onto the magnetic bentonite surface, which restricted conformational flexibility and provided a protective microenvironment for the enzyme. Moreover, the immobilized uricase exhibited good operational stability, retaining ۵۶.۲% of its initial activity after seven consecutive reaction cycles. These findings demonstrate that Fe۳O۴@BNT-NH۲-GA is an effective, magnetic, and reusable support for uricase immobilization, offering potential applications in biocatalytic processes and biosensor development.

کلیدواژه ها:

Urate oxidase ، Magnetic bentonite ، Biocatalyst ، Enzyme immobilization ، Covalent immobilization