The study of immobilization of glucose oxidase on hydrophilic-hydropholic silica gel

Objective: To evaluate the performances of glucose oxidase immobilized on hydrophilic-hydrophobic silica gel, which may be employed to prepare glucose sensor for the determination of glucose in body fluids. Methods: The silica gel was prepared from precursors γ-aminopropyltrimethoxysilane (APTMOS) and methyltrimethoxysilane (MTMOS) by sol-gel technique. Glucose oxidase (GOD) was covalently attached to the silica gel via carbodiimide coupling reaction between a carboxylic acid group on enzyme and an amine group of the silica gel under the participation of the linking reagents 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-hydroxysuccinimide. The performances of immobilized GOD were explored. Results: The optimum conditions were obtained as follows: volume fraction of APTMOS 70%, enzyme content given 16 800 U, the temperature of 35°C and buffer pH 5. 5. The decrement in the activity of immobilized GOD for the first 2 weeks was less than 10% of its original activity, and the activity of immobilized GOD retained more than 75% of its original activity after 1 month of testing. Six independently prepared immobilized GOD on the silica gel resulted in an average bioactivity of 1 290.9 μmol · min^-1 · g^-1 with an R. S. D. of 3.4%. The Michaelis constant (K_m) of immobilized GOD was 9.1 mmol · L^-1. Conclusion: Immobilizing GOD on the silica gel via the formation of peptide bonds is an outstanding enzyme immobilization method.