P6: Optimization and Characterization of Glucose Oxidase Immobilization on Carbon Nanotube

The immobilized glucose oxidase is necessary for the development of enzymatic biofuel cells which can be applied to power sources of implantable devices as well as biosensors. We aimed to maximize the loading of glucose oxidase (GO_x) on multi-wall carbon nanotubes (MWCNTs) by covalent binding. One mg of MWCNT was carboxylated and reacted with N-Hydroxysuccinimide (NHS) in the presence of 1-ethyl-3(3-dimethylaminopropyl)carbodiimide (EDC), resulting in a NHS ester, which was then reacted with primary amine (NH₂) of glucose oxidase to form amide crosslinks. The concentrations of EDC, NHS, and glucose oxidase greatly influenced on the immobilization efficiency and thus these factors were optimized using Box-Behnken design. Physical adsorption was minimized when GOx-bound MWCNT was washed with Tween-20 solution. The amount of immobilized GOx was maximum, 3.3 mg-GOx/mg-CNT, when concentration of EDC, NHS and protein was 52 mM, 430 mM and 8.7 mg/mL, respectively and washed with 5 mg/mL of Tween-20 solution. The formation of chemical bonding between GOx and MWCNT, and GOx on the surface of MWCNT was observed by FT-IR and SEM. Consumption of glucose and production of gluconic acid were observed in enzyme assay using immobilized GOx.