Search

Bovine cytochrome c oxidase is immobilized in a gold substrate- supported lipid bilayer membrane to investigate reduction of oxygen using flow through detection technique. The results shown that system temperature, pH and component of solution affect the electroreductive rate of oxygen. In each case the reduction rate of oxygen is controlled by electron transfer. The oxidase modified electrode can be used for the direct electroreduction of oxygen to water in a biofuel cell.

Blood and tissue biochemical oxidation-reduction (redox) reactions are ubiquitous and are reflective of many important biological processes in the body, ranging from the state of cellular oxygenation to the overall antioxidant status. It is likely that, similar to acid-base balance, the body optimally operates within a narrow redox potential range made possible by various homeostatic mechanisms, and that deviation from this range will occur in tissue damage. A means to monitor the redox potential of blood or tissue would be valuable in both the diagnosis and treatment of diseases or conditions that adversely affect the body's redox potential. The biosensor described herein involves the immobilization of bovine cytochrome c oxidase (CCO) into electrode-supported lipid bilayer membranes. As a first proof of concept, the biosensor was used to potentiometrically monitor the concentration ratio of a redox pair (oxidized and reduced cytochrome c) in an in vitro system. The response of this modified electrode is reproducible and exhibits Nernstian behavior consistent with the four-electron reduction of the CCO. The oxidase-modified electrode can also operate as an amperometric biosensor for the detection of solution-resident ferrocytochrome c at concentrations as low as 0.1 /spl mu/M. Because this biosensor uses an electron sensor native to the body, it may be of future value to explore the biosensor as a point of care test to measure blood redox potential or perhaps as an implantable sensor to measure tissue redox potential in many settings.

Bovine cytochrome c oxidase has been successfully immobilized in electrode-supported lipid bilayer membranes to investigate the effect of cyanide binding on the oxidation of ferrocytochrome c and the electroreduction of dioxygen. Cyanide binding to oxidase was found to be reversible and exhibited 1:1 stoichiometry. Binding constants ( K i ) were also determined for binding of cyanide to the reduced (62 μM) and oxidized (195 μM) forms of the oxidase. The cytochrome c oxidase-modified electrodes described here could potentially be used as an amperometric biosensor for the detection of cyanide.

Stable lipid bilayer membranes containing cytochrome c oxidase from two sources (i.e. bovine and Bacillus stearothermophilus ) have been immobilized on gold quartz crystal microbalance (QCM) electrodes. The immobilized oxidase can be characterized using direct electrochemical methods. Amperometric flow injection analysis data of reduced cytochrome c reacting at oxidase-modified electrodes indicate that the kinetics are non-hyperbolic, which agrees with previous results found in the literature. The kinetics of the cytochrome c oxidase/cytochrome c redox couple are characterized by a high and low affinity regime each having a distinct K m value. The rate of mediated electron transfer from solution-resident reduced cytochrome c to the electrode is also affected by ionic strength. Electrochemical and QCM results suggest that the fastest rates of electron transfer are observed when the ionic strength is low enough to allow facile binding between the protein and the enzyme but high enough to allow the protein to dissociate from the oxidase.

In past work the direct electron transfer reactions of bovine cytochrome c oxidase in an electrode-supported lipid bilayer membrane have been studied. Its reaction with cytochrome c in solution was also studied and found to be consistent with previous solution studies. In this work it is shown that the electron transfer reactions of cytochrome c oxidase in this electrode-supported lipid bilayer membrane depend on the source of cytochrome c. This property has also been widely studied for solution samples. The differences in the electron transfer reaction rates correlate with the differences in the amino acid sequence for the cytochrome c molecules studied. Electrochemical results suggest that the dissociation of the cytochrome c/cytochrome c oxidase reaction complex is the rate-controlling step in this electron transfer mechanism for cytochrome c from some sources. Moreover, the electron transfer reaction mechanism exhibits biphasic reaction kinetics, which is consistent with earlier work on reactions between solubilized cytochrome c oxidase/cytochrome c samples. These results indicate that the cytochrome c oxidase modified electrodes described herein could be used to distinguish amino acid sequence variations in proteins such as cytochrome c, and this has potential relevance as a diagnostic for disease states.

Bovine cytochrome c oxidase has been successfully immobilized in an electrode-supported lipid bilayer membrane. The surface confined oxidase is capable of directly transferring electrons to the underlying electrode making characterization of the enzyme with direct electrochemical methods facile. Cyclic voltammetry indicates that the immobilized oxidase is capable of transferring electrons to the solid electrode over a wide range of pH and temperature values. Furthermore, the oxidase maintains its electron transfer properties at temperatures in excess of 80°C. Upon cooling the electrode back to room temperature, it was found that the enzyme still retained its electron transfer capabilities revealing the robust nature of the enzyme is this environment. In addition the oxidase continues to be capable of mediating electron transfer from solution-resident, reduce cytochrome c to the underlying metal electrode. The use of oxidase-modified electrodes enables the detection of solutions of reduced cytochrome c at concentrations as low as 0.25 μM.

The temperature dependence of the dissociation of cyanide (CN − ) from cyanomyoglobin (Mb(II)CN), the cyanide complex of myoglobin, has been studied by cyclic voltammetry. Direct electrochemical reduction of cyanometmyoglobin initiates the dissociation of the cyanide ligand. The temperature dependence of the heterogeneous electron transfer kinetics, the kinetics of cyanide dissociation, and the formal potential for Mb(III)CN have been determined over the temperature range of 10–50°C. The latter information has been used to determine the reaction center entropy change (Δ S rc ) of this reaction to be −90±5 J mol −1 K −1 . The water complex of myoglobin, aquometmyoglobin (Mb(III)H 2 O), was also studied and the reaction center entropy for this system was determined to be −40±4 J mol −1 K −1 . Differential scanning calorimetry was also used to determine the transition temperature and enthalpy changes for each component of these reactions. The results given here provide an indirect model for the in vivo reaction of the oxymyoglobin complex (Mb(II)O 2 ), that is, the dissociation of dioxygen from the reduced form of myoglobin.

Scanning force microscopic images of cytochrome c oxidase immobilized within an electrode-supported lipid bilayer membrane are reported. These images represent the first direct evidence that the microscopic architecture of this important model system effectively mimics the microstructure proposed for the inner mitochondrial membrane. The images reveal (1) that the oxidase is present as monomers and small aggregates within the supported membrane and (2) that the oxidase constitutes {approximately}20% of the lipid bilayer membrane. The latter finding allows an estimation of the lower limit for the minimum turnover rate ({approximately}0.5 electrons/s) required for the transition of the oxidase from its resting to pulsed kinetic state.

The effect of electrostatic substitutions in the heme pocket of myoglobin on the ligand binding and electrochemical properties of three variants of horse heart myoglobin (S92D, V67R, and V68H) have...

Cytochrome c oxidase immobilized in lipid bilayer membranes on gold electrodes mediates electron transfer between reduced cytochrome c in solution and the electrode. Under flow conditions, fixed potential amperometry of anaerobic solutions of reduced cytochrome c shows changes in the rate of enzyme mediated electron transfer from cytochrome c to oxidase modified electrodes. These data are consistent with literature that describes both activation of the resting oxidase state to the pulsed oxidase state and subsequent decay of the pulsed oxidase state to the resting oxidase state.

Electrochemistry and spectroelectrochemistry were used to study solutions of horse myoglobin, both by itself and complexed with cyanide or imidazole, at indium-tin oxide electrodes. In the absence of ligands other than water, myoglobin exhibited slow quasi-reversible heterogeneous electron transfer kinetics, with a formal heterogeneous electron transfer rate constant ( k °′) value of 1.6(±0.4)×10 −5 cm s −1 . Differences from simple simulations in the reverse sweep (anodic) cyclic voltammograms are proposed to be due to residual amounts of dioxygen in solution and/or in the protein molecule. Those differences were not seen in the anodic waveforms of simultaneously acquired derivative cyclic voltabsorptometry (DCVA) experiments. Cyanomyoglobin exhibited faster heterogeneous electron transfer kinetics, with a k °′ value of 6.0(±1.0)×10 −4 cm s −1 . Values for the homogeneous rate constants for dissociation ( k f ) and association ( k b ) of cyanide from and to the electrochemically reduced protein were determined to be 0.08(±0.03) s −1 and 0.06(±0.03) M −1 s −1 , respectively. Imidazolemyoglobin was found to transfer electrons faster than cyanomyoglobin, with a k °′ value of 2.0(±0.5)×10 −3 cm s −1 . The values for the homogeneous rate constants k f and k b were 0.40(±0.1 5) s −1 and 1.1(±0.1) M −1 s −1 , respectively.

The objective of this work is to accurately and reproducibly control the amount of alkanethiol self-assembled onto the surface of a gold quartz crystal microbalance (QCM) electrode. This reaction is the first step in preparing bilayer membranes on gold electrodes that contain cytochrome c oxidase. In this work procedures for cleaning the surface of gold QCM electrodes, including use of an oxygen plasma, a UV surface cleaner, and aggressive chemical treatment of the surface, still led to rates of octadecyl mercaptan (OM) self-assembly that were highly irreproducible, varying over several orders of magnitude in time. Electrodeposition of a silver monolayer onto the surface of gold QCM electrodes affords a surface at which the OM self-assembly reaction can be easily controlled. Using the QCM to monitor the self-assembly reaction of OM on the silver-coated gold QCM electrode surface, a selected sub-monolayer coverage can be reliably prepared in about 10 min.

Examples of facile electron transfer between electrode surfaces and electron transfer proteins were first reported nearly twenty years ago. Substantial progress has been achieved in understanding the fundamental requirements that must be met in order to observe such reactions. Despite the large body of work that has now been published on a host of examples of such reactions, there continues to be a lack of understanding over just what conditions must be met in order to use direct electrochemical methods to characterize the electron transfer thermodynamics and kinetics of electron transfer proteins. Part of the problem lies in the inherent difficulties associated with using solid electrodes. Equally as important is the wide range of attention that has been paid to the purity of the electron transfer protein solutions being studied. The goal here is to bring together those aspects of this problem that enjoy some degree of agreement among scientists in this field and to reflect upon those issues tha...

The direct heterogeneous electron transfer reactions of cytochrome c from different vertebrate species are compared in this report. A kinetic study revealed a biphasic temperature dependence at neutral pH for the formal heterogeneous electron transfer rate constants of these cytochromes. The properties of the electrode-solution interface are believed to cause this biphasic kinetic effect. This effect does not correlate with the conformational transitions evident in both formal potential and circular dichroism (CD) measurements as a function of temperature. The conformational transitions seen in the formal potential and CD measurements arise from the alkaline isomerization and heme crevice changes. These are subtle effects compared with the structural changes associated with unfolding of the protein that are evident in differential scanning calorimetric experiments. The conformational stabilities of cytochrome c between species relate to the differences in their amino acid sequences. The amino acid interactions which stabilize the structure of these different cytochrome c molecules have been evaluated and will be discussed.

The electron-transfer reactions of high-spin metmyoglobin and low-spin cyanometmyoglobin have been studied electrochemically. The striking differences seen for the reactions of these complexes are apparent when the solutions studied contain both metmyoglobin and cyanometmyoglobin. The redox potential for cyanometmyoglobin has shifted negatively over 400 mV, from +0.046 to -0.385 (±0.015) V vs NHE, and the rate of electron transfer has increased by more than 1 order of magnitude compared with that for metmyoglobin, from 7×10 -6 to 5,4 (±0.9)×10 -4 cm s -1

Cytochrome c oxidase is the terminal enzyme in mammalian respiration, and one of its main functions is to catalyze the reduction of oxygen under physiological conditions. Direct reduction of oxygen at electrodes requires application of substantial overpotentials. In this work, bovine cytochrome c oxidase has been immobilized in electrode-supported lipid bilayer membranes to investigate the electroreduction of oxygen under flow conditions. The effect that temperature, solution pH, and solution composition have on the reduction of oxygen by this novel enzyme-modified electrode is reported. Results indicate that the electroreduction of oxygen is most pronounced at low pH (6.4) and elevated temperature (38 degrees). At an applied potential of -350 mV vs. Ag/AgCl (1M KCl), a current density of ca. 7 microA/cm2 was obtained. The current responses obtained at these electrodes are stable over a period of ca. 10-14 days (10-15% decrease in response). The cytochrome c oxidase-modified electrodes described here could potentially be used for the direct electroreduction of oxygen to water in a biofuel cell.

The characterization of the dynamics of conformational changes that accompany the ligand binding and dissociation reactions of myoglobin may provide insights into the events that control the physiological function of this oxygen storage protein. The cyanometmyoglobin system was chosen for this study because cyanide binds to the metmyoglobin state and dissociates upon reduction. The potential step spectroelectrochemical method was used here because the rate of cyanometmyoglobin reduction at an electrode can be controlled so that the rate of dissociation of the reduced cyanomyoglobin can then be followed.

Cyclic voltammetry has been used to study the heterogeneous electron transfer kinetics of horse heart cytochrome c in pH 7 tris/cacodylate media at several electrode surfaces. Reversible voltammetric responses (formal heterogeneous electron transfer rate constant>10−2 cm/s) were observed at bare gold electrodes and at tin-doped indium oxide semiconductor electrodes for certain experimental conditions. Quasireversible voltammetric responses were more typically observed at fluorine-doped tin oxide semiconductor electrodes, bare platinum electrodes, and at the indium oxide electrodes. Reaction rates at bare metal electrodes were strongly dependent on pretreatment procedures and experimental protocol. Reaction rates at metal oxide electrodes were strongly dependent on solution conditions, pretreatment procedures, and on the hydration state of the electrode surface. A general mechanistic scheme involving both interfacial electrostatic and chemical interactions is proposed for cytochrome c electrode reactions. The asymmetric distribution of surface charges on cytochrome c appears to play a dominant role in controlling electron transfer rates by its interaction with the electric field at the electrode surface. Electron transfer distances are also considered, and it is concluded that electron transfer between an electrode surface and the exposed heme edge of properly oriented cytochrome c molecules involves maximum distances of ca. 0.6–0.9 nm.

An electrochemical thin-layer cell, 0.5 ml in volume and 0.25 mm in pathlength, employing an optically transparent gold minigrid electrode has been constructed and used in redox studies of photosynthetic electron-transport components. Light-induced absorption changes accompanying P700 photo-oxidation in photosystem-I subchloroplasts were measured at a series of electrochemically poised potentials at 88°K. The fluorescence-yield changes in photosystem-II subchloroplasts were measured as the poised potential was varied electrochemically. The utility of the cell was further demonstrated by a redox titration of soluble (spinach) ferredoxin using circular dichroism to monitor changes in the redox state. One important advantage of electrochemical vs chemical titration of photosynthetic electron-transport components is that it is possible to attain much more negative potentials than is thermodynamically possible in chemical titrations. This permits the exhaustive titration of certain components at physiologically reasonable pH values not possible chemically.

The biphasic behavior of the formal potential (Eo') of cytochrome c with variations of pH (2.5–9) and temperature (4–52° C) is discussed. From an analysis of the dependence of Eo' on pH and temperature it was found that the biphasic behavior of Eo' is due to ferricytochrome c and not ferrocytochrome c. Four conformational states in ferricytochrome c have been identified from the electrochemical measurements. The linear change in Eo' of the individual states with pH and temperature was explained by a simple equation of the electrode reaction with an apparent protonation number. A pH versus temperature diagram which shows the stable regions of the four states has been obtained. These four states correspond to states that have been suggested by other methods such as spectrophotometry and resonance Raman spectroscopy. Since the biphasic changes of Eo' with pH and temperature were very sharp, a conformational transition instead of a change by an equilibrium reaction was introduced to explaine this feature. The high temperature dependence of Eo' (2 mV K−1) of some states has been explained partly by the concentration dependent effect of protons on the entropy change of the electrode reaction.

The indirect coulometric titration of cytochrome c oxidase and dioxygen using cytochrome c as a mediator is described. Results of both the indirect coulometric titrations and the cyclic voltammetric experiments reported herein verify that the reaction mechanism involves the catalytic regeneration of the electroactive species, the cytochrome c mediator, with the selective reduction of cytochrome c oxidase alone. During the indirect coulometric titrations dioxygen is reduced to water only by cytochrome c oxidase and not by either direct reduction at the electrode surface or reaction with cytochrome c. This system utilizes the electron transfer selectivity of cytochrome c for cytochrome c oxidase over dioxygen and offers a means by which the reaction of cytochrome c oxidase and dioxygen can be examined.

The first reduction of 1,1′-dimethyl-r,r′-bipyridinium dichloride (methyl viologen, MV +- results in the formation of both a monomeric, MV ± , and a dimeric (MV +- 2 product. The molar absorptivities and the wavelengths of maximum absorption for both reduction products can be determined by computer decomposition of the u.v.-visible absorption spectra resulting from the chemical and in situ electrochemical reduction of MV 2+ . The FORTRAN computer program, SPECSOLV, is used to establish the spectral parameters of MV± and (MV±) 2 .

Spectroelectrochemical and electrochemical methods were used to investigate the characteristics of heterogeneous electron transfer between cytochrome c and indium oxide electrodes. A linear temperature dependence of the formal potential of cytochrome c was observed from 5 to 75 °C in acidic media. This behavior is attributed to a linear variation in the conformation of ferricytochrome c that results in an increase in solvent exposure of the solvent-exposed heme edge. A break in the linear temperature dependence of the formal potential occurred at 40 °C in alkaline media. This reflects a distinct conformational change that accompanies the onset of thermal denaturation of ferricytochrome c. The small change in reaction center entropy, ΔS°rc, of ca. −54 J K−1 mol−1 in neutral and acidic media (5 to ⩾ 55 °C) and in alkaline media (below 40 °C) is consistent with a small shift to a more stable conformation of cytochrome c that occurs upon reduction. Adsorption of reactant and product was detected. The strength and type of adsorption were found to be temperature- and pH-dependent. The characteristics of electron transfer between cytochrome c and an electrode depend on bulk solvent properties and electrode surface characteristics.

On decrit les effets de la temperature sur a la fois le potentiel standard et la vitesse de transfert d'electron heterogene pour le cytochrome c a des electrodes d'oxyde d'indium dopees a l'etain

The kinetics of viologen cation radicals reacting at hydrogen-evolving gold and nickel electrodes in pH 6–8 electrolytes have been investigated. Visible absorption spectroscopy was used to follow the course of the reaction in an optically transparent thin-layer electrochemical cell under quasi-steady-state conditions. The spectroelectrochemical data were analyzed using classical kinetics and yielded zero-order behavior with respect to the viologen cation radical. For methyl viologen cation radical at gold, a formal zero-order rate constant evaluated at zero hydrogen overpotential was found to be 1.0 × 10 −13 mol s −1 cm −2 . At nickel the comparable rate constant was nearly two orders of magnitude larger than at gold. Increasing pH from 6 to 8 at gold electrodes shifted both the hydrogen evolution and the methyl viologen cation radical reaction 60–70 mV/pH unit in a negative direction. The diquat cation radical behaved in a similar manner. The proposed mechanism involves a fast, non-rate-limiting, chemical reaction between the viologen cation radical and adsorbed hydrogen atom(s). Results are interpreted in terms of previous proposed hydrogen evolution reaction mechanisms.

Channel-flow hydrodynamic voltammetry was used to study the direct electron-transfer reactions of two metalloproteins, myoglobin and cytochrome c, under steady-state conditions at methyl viologen modified (MVM) gold foil electrodes. Utilization of a dual working electrode cell with this technique permitted determination of the heterogeneous electron-transfer kinetics for both the reduction and oxidation of myoglobin. The formal heterogeneous electron-transfer rate constants in pH 7.00 phosphate buffered solutions were found to be 8.9 (±1.5)×10−5 cm s−1 for the reduction, and 7.7 (±1.2)×10−5 cm s−1 for the reoxidation of myoglobin. The transfer coefficient values obtained were 0.21 (±0.01) for the reductive (α) and 0.82 (±0.01) for the oxidative (1−α) electrode reactions. Ionic strength and pH dependences were observed in these direct electron-transfer reactions. Collective current efficiency measurements in the myoglobin experiments indicated that an overall simple charge-transfer process occurred at the respective electrode interfaces. A formal rate constant of 3.4 (±0.2)×10−5 cm s−1 with a transfer coefficient of 0.25 (±0.01) for the reduction (α) of cytochrome c was obtained by this hydrodynamic technique. The use of channel-flow hydrodynamic voltammetry in characterizing an electrode reaction as well as an interpretation of the data presented are discussed.

The heterogeneous electron transfer kinetics of soluble spinach ferredoxin reacting at modified gold electrodes have been characterized by both steady state and transient electrochemical methods. The steady state analysis was conducted at a methyl viologen modified gold disk electrode using rotating disk hydrodynamic voltammetry. The results presented are the first reported steady state kinetic parameters observed for the unmediated reduction of ferredoxin at this surface. Single potential step chronoabsorptometry was utilized to observe the transient kinetic behavior of ferredoxin at modified gold minigrid electrodes. The kinetic parameters reported herein were evaluated using a recetly advanced, generalized analysis that is applicable to heterogeneous electron transfer systems that exhibit any degree of reversibility. Both analyses yield a formal heterogeneous electron transfer rate constant, ks,h0', and electrochemical transfer coefficient, α, in the following ranges: 1×10−4 cm/s

The electrochemical properties of three dipyridinium salts were studied in order to assess their behavior for use as mediators in titrations of biological molecules. The compounds studied were 1,1′-trimethylene-2,2′-dipyridinium dibromide, 5,5′-dimethyl-1,1′-trimethylene-2,2′-dipyridinium dibromide and 4,4′-dimethyl-1,1′-trimethylene-2,2′-dipyridinium mide. Differential pulse polarographic analysis showed that each of these compounds undergoes two successive one-electron reductions, and that the formal potentials of these reduction steps are independent of pH. Cyclic voltammetry at the hanging mercury drop electrode (HMDE) indicated that in all cases the first reduction step is reversible. However, the sample potentials measured during controlled potential electrolysis of these compounds were not as negative as theoretically expected, indicating decomposition of the electrolysis products. This study concludes, however, that these mediators can be used in the indirect coulometric titration of large biological redox molecules if there is sufficient separation between the formal potential, E 1 0′ , of the mediator and the E 0′ of the biological molecules. This condition prevents the accumulation of the product of the electrode reaction, the cation radical and therefore the decomposition of the mediator.

Description de cette methode permettant de determiner les parametres de la cinetique de transfert d'electrons heterogene dans le cas de systemes redox reversibles. Exemple dans le cas de la reduction de l'hexacyanoferrate, dans des solutions de pH7, en presence d'electrodes optiquement transparentes d'oxyde d'etain, dopees avec des fluorures

The heterogeneous electron transfer reactions of cytochromec occurring at platinum, gold and mercury electrodes are shown to be quasi-reversible. In each case the electrodes have not been modified and the cytochromec samples are native. This work extends previous work and demonstrates that biological molecule electron transfer reactions can be studied at clean metal surfaces to gain fundamental knowledge of the mechanisms of these reactions.

Differential pulse polarography at the dropping mercury electrode and differential pulse voltammetry at the carbon paste electrode are used for direct determinations of pentachlorophenol at concentrations down to 0.27 ppm. PCP is electrochemically reduced in phosphate buffers of pH 8 to produce a concentration-dependent current peak at —0.8 V vs. Ag/AgCl. The procedure requires only 15 min. Cyclic voltammetry at the hanging mercury drop electrode is used to evaluate the electrochemical reaction and to establish the reversibility of the PCP electrode reaction.

Cyclic voltammetry (CV) and differential scanning calorimetry (DSC) have been used to study the thermal denaturation of the coupled cytochrome c/cytochrome c oxidase/dioxygen system in solution. CV has also been used to determine the rates of homogeneous electron transfer between cytochrome c and cytochrome c oxidase over a temperature range from 10 to 45°C. These electron transfer reactions have been initiated by direct reduction of cytochrome c at indium oxide electrodes. Denaturation mechanisms have been characterized by both CV and DSC demonstrating the complementary nature of the information provided from these different methods. Cytochrome c oxidase denaturation resulted in a large decrease in the thermal stability of cytochrome c while the presence of cytochrome c had no measurable effect on the thermal stability of cytochrome c oxidase. The kinetics of denaturation have been followed by CV following the rapid increase in sample temperature and mechanisms for denaturation have been suggested.

Very little information has been reported describing the heterogeneous electron transfer kinetics of biological molecules at electrodes. We report here the first application of a recently developed spectroelectrochemical technique to the measurement of the heterogeneous electron transfer kinetics of a biological molecule, sperm whale myoglobin, at a methyl viologen modified gold minigrid electrode. The overpotential dependence of the heterogeneous electron transfer rate constant for the reduction of myoglobin at this surface gives rise to values of the formal heterogeneous electron transfer rate constant [ko′f,h = 3.88 (± 0.07) × 10−11 cm/s] and the transfer coefficient [α = 0.88 (± 0.01)] for this electrocatalyzed process. The importance of studying the heterogeneous electron transfer kinetics of biological molecules lies in the fact that many physiological electron transfer reactions occur heterogeneously. Though myoglobin does not function in this manner physiologically, our initial study has been directed at this molecule owing to its stability and ready availability. It is expected that this technique will be applied to other optically transparent electrodes and other biological molecules thereby providing new insights into understanding biological redox reactions.

Very little information has been reported describing the heterogeneous electron transfer kinetics of biological molecules at electrodes. We report here the first application of a recently developed spectroelectrochemical technique to the measurement of the heterogeneous electron transfer kinetics of a biological molecule, sperm whale myoglobin, at a methyl viologen modified gold minigrid electrode. The overpotential dependence of the heterogeneous electron transfer rate constant for the reduction of myoglobin at this surface gives rise to values of the formal heterogeneous electron transfer rate constant [ko′f,h = 3.88 (± 0.07) × 10−11 cm/s] and the transfer coefficient [α = 0.88 (± 0.01)] for this electrocatalyzed process. The importance of studying the heterogeneous electron transfer kinetics of biological molecules lies in the fact that many physiological electron transfer reactions occur heterogeneously. Though myoglobin does not function in this manner physiologically, our initial study has been directed at this molecule owing to its stability and ready availability. It is expected that this technique will be applied to other optically transparent electrodes and other biological molecules thereby providing new insights into understanding biological redox reactions.