An Os(byp) 2 ClPyCH 2 NHPoly(allylamine) hydrogel mediator for enzyme wiring at electrodes C. Danilowicz, a * E. CortoÂn, b F. Battaglini b and E. J. Calvo b a Facultad de Farmacia y Bioquimica, Universidad de Buenos Aires, Junin 956, AR-1113, Buenos Aires, Argentina b INQUIMAE, Departamento de QuõÂmica InorgaÂnica, AnalõÂtica y QuõÂmica FõÂsica Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellon 2, Ciudad Universitaria, AR-1428 Buenos Aires, Argentina (Received 27 October 1997; in revised form 29 December 1997) AbstractÐAn Os hydrogel based on the covalent attachment of Os(byp) 2 ClPyCHO to PAA-NH 2 was cross-linked with glucose oxidase (GOx), lactate oxidase (LOx) and horseradish peroxidase (HRP) on GC and Au electrodes by PEG-400 bifunctional reagent. Single layer monoenzyme (GOx or LOx) and bien- zyme (HRP-GOx) single layer modi®ed electrodes were prepared with the Os moieties acting as ``electron wires or electron shuttles''. Cyclic voltammetry showed diusional charge propagation in the gel which resulted more stable than similar ferrocene based gels reported before. In solutions containing the sub- strates, catalytic currents were obtained due to enzyme catalysis for the oxidation of glucose and lactate by the respective enzymes mediated by the Os polymer either by detecting directly the anodic current in a single enzyme electrode or indirectly by further reducing the peroxide formed in the aerobic enzymatic cycle at the Os-wired HRP. A rotating disc electrode (RDE) and a wall jet electrode (WJE) were employed as hydrodynamic electrodes in order to correct the amperometric response for substrate concentration polarization in the external electrolyte. # 1998 Elsevier Science Ltd. All rights reserved Key words: enzyme electrodes, GOx, LOx, HRP, FIA, biosensor, Os polymer. INTRODUCTION Biosensors based on amperometric devices with im- mobilized enzymes on the surface of an electrode have been thoroughly described to recognize the analyte at the molecular level and to generate an analytical signal by an electrochemical reaction which acts as a transducer [1]. Electrically wired enzymes by redox hydrogels were invented by A. Heller [2] and provide three dimension electronic conductivity between enzymes and electrodes. Hydrogels provide enhanced enzyme stability in an environment with high water content where soluble biochemicals such as glucose or lac- tate can diuse towards the speci®c enzyme. The biosensor response results by shuttling electrons between the enzyme redox prosthetic group (FAD, NAD(P), Heme, etc.) and the underlying electrode. Examples of this analytical strategy are biosensors based on glucose oxidase, lactate oxidase and horse- radish peroxidase [3±12] with applications ranging from implantable clinical sensing to food analysis. Other dierent redox polymers such as polysilox- anes were also used for biosensor design and proved to be eective to transfer charge to redox enzymes [13±15]. In our hands, hydrogels formed by cross-linking ferrocene poly(allylamine), (Fc-PAA), and glucose oxidase (GOx) were investigated by cyclic voltam- metry (CV) and electrochemical impedance spec- troscopy (EIS). The eect of enzyme and substrate concentration, electrode potential, nature of sub- strate and redox mediator on the biosensor signal, electrostatic eects on the catalytic current and the Electrochimica Acta, Vol. 43, No. 23, pp. 3525±3531, 1998 # 1998 Published by Elsevier Science Ltd. All rights reserved Printed in Great Britain 0013±4686/98 $19.00 + 0.00 PII: S0013-4686(98)00100-5 *Author to whom correspondence should be addressed. 3525