Glucose oxidase, horseradish peroxidase and phenothiazine dyes-co-adsorbed carbon felt-based amperometric flow-biosensor for glucose.

To develop an amperometric flow-biosensor for glucose, the stabilizing effect of methylene blue (MB) toward adsorbed glucose oxidase (GOx) on carbon felt (CF) was successfully applied to prepare the GOx-modified CF-based enzyme reactor combined with a horseradish peroxidase (HRP)-modified CF-based H ₂ O ₂ detector. Upon mixing MB in the GOx-adsorption solution, the O ₂ -dependent GOx-activity was significantly increased with increasing concentration of MB in the GOx-adsorption solution. The GOx-immobilization protocol on CF is very straightforward [ i.e. , adsorption of the GOx/MB mixed aqueous solution for 5 min under ultrasound (US)-irradiation]. Under the optimized operational conditions ( i.e. , applied potential, 0 vs. Ag/AgCl; carrier pH, 5.0; carrier flow rate, 4.0 mL min ^-1 ), the resulting GOx/MB-CF-reactor and HRP/TN-CF-detector combined amperometric flow-biosensor exhibited sensitive, selective, reproducible and stable cathodic peak current responses to glucose with the following analytical performances: sensitivity, 6.22 μA mM ^-1 ; linear range, 0.01 to 1 mM; limit of detection, 9.6 μM (S/N = 3, noise level, 20 nA); sample throughput, 46-96 samples per h for 10-0.1 mM glucose. The developed amperometric flow-biosensor allowed the determination of glucose in beverages and liquors, and the analytical results by the sensor were in fairly good agreement with those by conventional spectrophotometry.