Kinetic control by the substrate and the cosubstrate in electrochemically monitored redox enzymatic immobilized systems.: Catalytic responses in cyclic voltammetry and steady state techniques

The catalytic response of an immobilized redox enzyme connected to the electrode by a freely diffusing mediator (cosubstrate) may be kinetically controlled by the substrate and/or the cosubstrate. How the electrochemical responses are related to the rate constants, to the amount of enzyme on the electrode, to the substrate and cosubstrate concentration and to the mass transport parameter is systematically analyzed in the framework of cyclic voltammetry and steady state techniques (SST). Because of its frequent occurrence in practice, emphasis is put on the case of a fast enzymatic process, as compared to the diffusion of the cosubstrate, provision being made for Michaelis-Menten behavior for both substrate and cosubstrate. Within this framework, two situations of particular interest are discussed, namely the case of a negligible consumption of the substrate in the enzyme coating and the opposite case where the consumption of the substrate is so important that its diffusion toward the electrode controls the current. In the first case, plateau-shaped responses, independent of scan rate, are obtained in cyclic voltammetry, and likewise mass transport independent waves, in SST. In the second case, a curve exhibiting a sharp, discontinuous peak is obtained in cyclic voltammetry with a peak current proportional to the substrate concentration and square root of the scan rate. A discontinuity also appears in SST between the rising portion of the wave and the plateau current while the current is likewise proportional to substrate concentration. The combination of these two regimes accounts for a biphasic variation of the electrochemical signal with the substrate concentration. The relationships between the electrochemical responses and the kinetic characteristics of the enzymatic reaction form the bases of procedures for ascertaining the mechanism and measuring the key rate constants. In this connection, strategies for determining Michaelis-Menten characteristics of both the substrate and cosubstrate reactions are discussed. (C) 2002 Published by Elsevier Science B.V.