The voltammetry of cytochrome c oxidase immobilized in lipid bilayer membranes on gold electrodes and amperometric data of cytochrome c reacting at these electrodes under flow conditions are reported. A submonolayer of octadecyl mercaptan formed on electrodeposited silver anchors and becomes a part of the lipid bilayer membrane on the gold electrode. The supported lipid bilayer membrane containing cytochrome c oxidase is formed during a deoxycholate dialysis procedure. Slow scan rate cyclic voltammograms (20 mV/s) taken at the oxidase-modified electrodes show well-defined anodic waves. Fast scan rate cyclic voltammograms (200 mV/s) taken at the oxidase-modified electrodes show well-defined anodic and cathodic waves. Cyclic voltammograms taken at the oxidase-modified electrodes under 0.1 mM sodium cyanide show an increase (ca. 300%) in electrode capacitance and well-defined anodic and cathodic waves irrespective of scan rate. The voltammetric data are consistent with electron transfer of cytochrome c oxidase coupled with changes in nonfaradaic current and possibly diffusion of cytochrome c oxidase in a lipid multilayer structure. Quartz crystal microbalance data of cytochrome c binding to lipid bilayer membranes containing no cytochrome c oxidase under flow conditions are presented.

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A membraneassociated enzyme, cytochrome c oxidase, has been put in lipid bilayer membranes attached to gold electrode surfaces. Goldhhiol self-assembly chemistry and deoxycholate dialysis procedures have been used to insert cytochrome c oxidase into a lipid bilayer on gold surfaces with a controlled orientation. Voltammetric and spectroelectrochemical results indicate that direct electron-transfer communication between the gold electrode surface and cytochrome c oxidase has been achieved. Moreover, immobilized cytochrome c oxidase can both reduce and oxidize solution-resident cytochrome c by appropriately controlling the applied electrode potential.

In view of the observations, an adsorption mechanism is proposed as follows. Hydroxide ions are strongly adsorbed at the bronze surface in a deoxygenated system establishing a large negative potential (-1200 mV or less). When O2 molecules are introduced, they are also adsorbed at the surface, displacing the negatively charged hydroxide and resulting in a positive shift in electrode potential. A similar mechanism would also explain the O2 response in KC1.The success of EDTA titration in basic solutions could similarly be explained if the positive ions act in a manner similar to the oxygen molecules in displacing adsorbed OH-. The action of Li + in LiOH may have a similar explanation. CONCLUSIONThe tungsten bronzes have been shown to be highly useful as indicating electrodes in the potentiometric determination of dissolved oxygen. Obvious applications are foreseen in the environmental field, resulting from the high degree of sensitivity attainable and the magnitude of the potential change per unit change of oxygen concentration. The relative ease with which potentiometric measurements can be made and the simplicity of the equipment add to the utility of measuring systems using these electrodes. Work in progress includes the development of a portable device for monitoring dissolved oxygen in surface and waste waters. ACKNOWLEDGMENTThe authors wish to acknowledge the assistance of Howard R. Shanks of the Ames Laboratory for providing the tungsten bronze crystals which were used in this investigation and of Patrick R. Montoya for his help in conducting the experiments.

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