Unusual response in mediated biosensors with an oxidase/peroxidase bienzyme system

Mediated biosensors consisting of an oxidase and peroxidase (POx) have attracted increasing attention because of their wide applicability. However, since most of oxidases utilize artificial electron acceptors in place of dioxygen, the competition between O2 and the electron acceptor in the mediated sensors is anticipated. This has been evidenced with a glucose oxidase (GOx)- and POx-coentrapped and ferrocene-embedded carbon paste electrode, which exhibits peak-shaped current-time curves at increased concentrations of glucose and also gives a peak-shaped calibration curve. Digital simulation has been applied to clarify the cause of such unusual responses, by taking into account the ping-pong enzyme kinetics on two- and three-substrate models for POx and GOx, respectively. The simulation has well reproduced such unusual responses and has clearly revealed that the depletion of O2 in the enzyme layer is the most important factor responsible for such unusual responses. To overcome such a drawback of oxidase/POx bienzyme sensors, it is expected to be essential to decrease the rate of the oxidase reaction. In contrast, increase in the POx activity is useful to improve the sensitivity. According to the simulation-based expectation, the GOx and POx concentrations in the bienzyme sensor are adjusted to exhibit normal behavior with high sensitivity.