Direct electrochemistry of membrane-entrapped horseradish peroxidase. Part II: Amperometric detection of hydrogen peroxide

Direct (unmediated) electrochemistry of horseradish peroxidase (HRP) immobilized within a polymeric film is investigated at a pyrolytic graphite electrode by de cyclic voltammetry, in the absence and in the presence of hydrogen peroxide. Under the latter condition. a reduction wave centered at approx. -280 mV (vs. SCE) is observed, the intensity of which is strictly dependent on the hydrogen peroxide concentration. This permits a voltammetric investigation of the electrocatalytic reduction of hydrogen peroxide. Flow and flow-injection measurements carried out at constant potential under the same conditions, support voltammetric data. The suitability of the immobilized HRP-based electrodic system to monitor the presence of important analytes, such as glucose or choline, in solution, is also discussed. To this issue, suitable amounts of HRP and glucose oxidase (or, in turn, choline oxidase) were simultaneously entrapped in the polymer. The results obtained are of potential value for basic and applied biochemistry and represent a first step for construction of a mediator-free (third-generation) biosensor which may find application in the biosensoristic area. (C) 1998 Elsevier Science S.A. All rights reserved