Amperometric on-line sensor for continuous measurement of hypoxanthine based on osmium-polyvinylpyridine gel polymer and xanthine oxidase bienzyme modified glassy carbon electrode

Osmium-polyvinylpyridine gel polymer (Os-gel-horseradish peroxide (HRP)) an d xanthine oxidase (XOD) bienzyme-based amperometric biosensor was develope d for use in a thin-layer radial flow cell for on-line measurement of hypox anthine. Glassy carbon electrodes (GCEs) were initially cast-coated with Os -gel-HRP with a surface coverage of 7.1 mu l/cm(2) to sense hydrogen peroxi de produced from XOD catalytic enzyme reaction based on HRP catalytic reduc tion mediated by osmium, Then, bovine serum albumin (BSA)/XOD was cross-lin ked to the Os-gel-HRP-coated glassy carbon electrodes (GCEs) with glutarald ehyde vapor for the measurement of hypoxanthine at 0.0 mV (versus Ag/AgCl). The resultant HRP and XOD bienzyme-based sensors (Os-gel-HRP/XOD/GCEs) wer e tested with techniques of cyclic voltammetry (CV), on-line amperometry in a flow system and flow injection analysis (FLA). In continuous-flow ampero metric experiments, the Os-gel-HRP/XOD/GCEs were quick to stabilize backgro und current and show linear and sensitive response towards hypoxanthine in a concentration range from 0.5 to 80 mu M with a detection limit of 0.2 mu M (S/N=3). The specificity against ascorbic acid was enhanced by negatively shifting the operating potential to -200 mV versus Ag/AgCl and was demonst rated to be acceptable for on-line performance. The interference from highe r than physiological levels of dopamine (DA), norepinephrine (NE), serotoni n, their principal metabolites and uric acid was minimal. The dependency of sensor response on pH value of the buffer and operating potential was also tested and optimized. (C) 2000 Elsevier Science B.V. All rights reserved.