Development of a Complex Material Composed of Enzymes-Inorganic Nanosheets-Magnetic Beads for Coupled Enzyme Reaction and Its Application to Glucose Detection

Abstract

A complex material, composed of glucose oxidase (GOx), horseradish peroxidase (HRP), titanate nanosheets (TiOx), and magnetic beads (MB), was developed, and its performance for glucose detection was investigated. The GOx/HRP/TiOx/MB complex was successfully prepared by utilizing electrostatic interactions among these components in acetate buffer at pH 4. The resultant GOx/HRP/TiOx/MB complex showed an excellent magnetic response and high retention ability for enzymes immobilized in the complex. Upon the addition of glucose to an acetic buffer solution containing the GOx/HRP/TiOx/MB complex and o-phenylenediamine (OPD), an increase in absorption derived from oxidation products of OPD was observed. The absorbance at 450 nm (A₄₅₀) increased with an increase in the concentration of glucose, while the addition of fructose, galactose, mannose, sucrose, and maltose, did not cause a significant change in A₄₅₀ value. The results indicate that the detection method based on the GOx/HRP/TiOx/MB complex has the detection ability and selectivity for glucose. Since the immobilization method for enzymes onto magnetic beads used in this study is simple and quick to operate, the method is expected to be a useful technique for preparing various types of enzyme-immobilized magnetic beads that can be used as attractive analytical tools.