Covalent Immobilization of Glucose Oxidase on Magnetite Particles via Graft Polymerization of Acrylic Acid

A new technique for immobilizing enzyme molecules on magnetite particles via the graft polymerization of acrylic acid is presented. The polymerization of acrylic acid was carried out in a redox system consisting of ceric ion and mercapto groups introduced onto magnetite particles. In the course of the polymerization, poly(acrylic acid) was attached to the magnetite particles. Glucose oxidase was covalently immobilized on the magnetite particles by the condensation reaction with the carboxyl groups of the poly(acrylic acid). It was shown that 2.8 mg of glucose oxidase was immobilized on 1 g of the magnetite attached with poly(acrylic acid), and the immobilized glucose oxidase had a specific activity of 81 units/mg, which was 50% of that of the native enzyme. Due to the immobilization, the optimum pH for glucose oxidase was shifted to a higher value and the temperature dependency of activity was diminished. A kinetic study of the glucose oxidation reaction with immobilized enzyme showed that the immobilization limited accessibility of glucose molecules to the active sites of the enzyme and caused a decrease of the maximum reaction rate. Glucose oxidas immobilized on magnetite particles kept 95% of its original activity in water over a period of 9 months.