Stable multilayer electroactive films were grown on rough pyrolytic graphite (PG) electrodes by alternate adsorption of layers of polyanion poly(styrenesulfonate) (PSS) and positively charged myoglobin (Mb) from their aqueous solutions. Incorporation of large amounts of electroactive Mb was facilitated by high electrode surface areas and by adsorbing coiled PSS from 0.5 M NaCl solutions. Cyclic voltammetry of {PSS/Mb}n films showed a pair of well-defined, chemically reversible peaks at about -0.25 V vs SCE at pH 5.5, characteristic of the Mb heme Fe III /Fe II redox couple. Electroactivity was extended to 7 {PSS/Mb} bilayers on rough PG surfaces, in comparison to 2 electroactive layers on smooth gold coated with mercaptopropanesulfonic acid. Square wave voltammograms of {PSS/Mb}n films gave good fits by nonlinear regression analysis to a model featuring dispersion of formal potentials, providing average formal potentials and an apparent rate constant. Oxygen and trichloroacetic acid were catalytically reduced by Mb in {PSS/Mb}n films with significant decreases in the electrode potential required. Making such films on PG rather than on gold provides a larger number of electroactive layers with no need for chemical pretreatment of the electrode.

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