Anodes enzymes

Do both mechanisms play a part At the present time several distinguishing experiments could be devised. One has been done by Bockris and Tunuli, who showed that when photo system I is absorbed upon platinum, and irradiated with light, the ensuing photoelectrochemical reaction is cathodic, but when photo system II replaced photo system I and I was irradiated, the corresponding reaction was anodic. This seems to fit well the theory of photosynthesis, in which photo system I and photo system II are analogous to cathodic and anodic enzyme electrodes, respectively. 

The degree of enzyme purity will ultimately affect fuel cell performance, particularly when enzyme preparations are used to form immobilized films on electrode surfaces in DET reactions. Contaminating proteins that do not provide electron transfer effectively foul the electrode. When enzyme immobilization techniques are specific to the enzyme, then enzyme purity may not be as much as an issue, but rarely the immobilization technique is absolutely specific to the cathodic or anodic enzyme. For example, an attractive immobilization strategy is to link a particular enzyme to an electrode via its cofactor (e.g., flavin adenine dinucleotide (FAD), nicotinamide adenine dinucleotide (NAD), etc.) [59]. The cofactor is linked to the electrode material first and then the apoenzyme is allowed to naturally bind to the cofactor all other proteins in the enzyme preparation that cannot bind the cofactor remain unbound and can be removed. Enzymes used in fuel cells are not so unique, and proteins in the immobilizing preparation may use the same cofactor but not the same fuel during fuel cell analysis or operation. 

TABLE 14.1 Common EEC Anode Enzymes Studied Using Enzyme-Mediated FB Imaging Mode in SECM... 

Table 14.1 lists the anode enzymes studied using SECM in FB mode and reported in the literature. Although the enzyme-mediated FB has primarily been used for imaging enzymes that catalyze oxidation reactions (anode enzymes for EFCs), there are examples of using this mode for studying reduction reactions using enzymes such as nitrate reductases [32,33] and horseradish peroxidase (HRP) [34]. 

The most usual choice for anodic enzyme has been glucose oxidase (GOx) [53], which, when under mediated ET conditions, can effectively electro-oxidize glucose. The enzyme carries a flavin (flavin adenine dinucleotide (FAD)) buried deep within the enzyme, which makes DET difficult. Although DET with GOx has been reported in many different studies [54-57], there is an ongoing debate as to whether true DET is achieved, or whether the observed bioelectrocatalytic currents are due to naturally mediated glucose oxidation by free FAD, which has diffused out firom the active centers of some partly denatured enzyme molecules. 

The Katz research group published two consecutive reports in 2012 of glucose/02 EFCs operating in mollusks [129,130]. The electrodes were fabricated using a DET-based design based on CNT paper (buckypaper), with laccase and PQQ-dependent GDH as cathodic and anodic enzymes, respectively. The enzymes were immobilized on nanostmetured electrodes by cross-linking. Such a DET-based approach simplifies... 

Functionalized conducting monomers can be deposited on electrode surfaces aiming for covalent attachment or entrapment of sensor components. Electrically conductive polymers (qv), eg, polypyrrole, polyaniline [25233-30-17, and polythiophene/23 2JJ-J4-j5y, can be formed at the anode by electrochemical polymerization. For integration of bioselective compounds or redox polymers into conductive polymers, functionalization of conductive polymer films, whether before or after polymerization, is essential. In Figure 7, a schematic representation of an amperomethc biosensor where the enzyme is covalendy bound to a functionalized conductive polymer, eg, P-amino (polypyrrole) or poly[A/-(4-aminophenyl)-2,2 -dithienyl]pyrrole, is shown. Entrapment of ferrocene-modified GOD within polypyrrole is shown in Figure 7. 

In presence of the enzyme glucose oxidase, an aqueous solution of glucose undergoes oxidation to gluconic acid with formation of hydrogen peroxide which can be determined by anodic oxidation at a fixed potential. 

When the electrode is placed in an aqueous solution of glucose which has been suitably diluted with a phosphate buffer solution (pH 7.3), solution passes through the outer membrane into the enzyme where hydroxen peroxide is produced. Hydrogen peroxide can diffuse through the inner membrane which, however, is impermeable to other components of the solution. The electrode vessel contains phosphate buffer, a platinum wire and a silver wire which act as electrodes. A potential of 0.7 volts is applied to the electrodes (the apparatus shown in Fig. 16.17 is suitable) with the platinum wire as anode. At this electrode the reaction H202->02 + 2H+ +2e takes place, and the oxygen produced is reduced at the silver cathode ... 

Figure 17.17 Schematic representation of a single-compartment glucose/02 enzyme fuel cell built from carbon fiber electrodes modified with Os -containing polymers that incorporate glucose oxidase at the anode and bilirubin oxidase at the cathode. The inset shows power density versus cell potential curves for this fuel cell operating in a quiescent solution in air at pH 7.2, 0.14 M NaCl, 20 mM phosphate, and 15 mM glucose. Parts of this figure are reprinted with permission from Mano et al. [2003]. Copyright (2003) American Chemical Society.

Minteer and co-workers have also exploited the broad substrate specificity of PQQ-dependent alcohol dehydrogenase and aldehyde dehydrogenase from Gluconobacter species trapped within Nahon to oxidize either ethanol or glycerol at a fuel cell anode [Arechederra et al., 2007]. Although the alcohol dehydrogenase incorporates a series of heme electron transfer centers, it is unlikely that many enzyme molecules trapped within the mediator-free Nahon polymer are electronically engaged at the electrode. 

Of direct interest for biofuel cell applications are the reported reduction of O2 by multi-copper oxidases on carbon nanotube electrodes [Yan et al., 2006 Zheng et al., 2006] and the oxidation of H2 by hydrogenase covalently bound to carbon nanotubes [Alonso-Lomillo et al., 2007]. The hydrogenase/nanotube anode is extremely stable (>1 month), and shows 33-fold enhanced enzyme coverage compared with similarly treated graphite of the corresponding geometric surface area. A. vinosum... 

Enzymes are efficient catalysts for cathodic and anodic reactions relevant to fuel cell electrocatalysis in terms of overpotential, active site activity, and substrate/reaction specificity. This means that design constraints (e.g., fuel containment and anode-cathode separation) are relaxed, and very simple devices that may take up ambient fuel or oxidant from their environment are possible. While operation is generally confined to conditions close to ambient temperature, pressure, and pH, and power densities over about 10 mW cm are rarely achieved, enzyme fuel cells may be particularly useM in niche environments, for example scavenging trace H2 released into air, or sugar and O2 from blood. Thus, trace or unusual fuels become viable for energy production. 

Although the inhibition-based biosensors are sensitive, they are poor in selectivity and are rather slow and tedious since the analysis involves multiple steps of reaction such as measuring initial enzyme activity, incubation with inhibitor, measurement of residual activity, and regeneration and washing. Biosensors based on direct pesticide hydrolysis are more straightforward. The OPH hydrolyzes ester in a number of organophospho-rus pesticides (OPPs) and insecticides (e.g. paraoxon, parathion, coumaphos, diazinon) and chemical warfare agents (e.g. sarin) [53], For example, OP parathion hydrolyzes by the OPH to form p-nitrophenol, which can be measured by anodic oxidation. Rainina... 

Figure 6.7 illustrates the voltammetric response of the third-generation SOD-based 02 biosensors with Cu, Zn-SOD confined onto cystein-modified Au electrode as an example. The presence of 02" in solution essentially increases both the cathodic and anodic peak currents of the SOD compared with its absence [150], Such a redox response was not observed at the bare Au or cysteine-modified Au electrodes in the presence of 02". The observed increase in the anodic and cathodic current response of the Cu, Zn-SOD/cysteine-modified Au electrode in the presence of 02 can be considered to result from the oxidation and reduction of 02, respectively, which are effectively mediated by the SOD confined on the electrode as shown in Scheme 3. Such a bi-directional electromediation (electrocatalysis) by the SOD/cysteine-modified Au electrode is essentially based on the inherent specificity of SOD for the dismutation of 02", i.e. SOD catalyzes both the reduction of 02 to H202 and the oxidation to 02 via a redox cycle of its Cu (II/I) complex moiety as well as the direct electron transfer of SOD realized at the cysteine-modified Au electrode. Thus, this coupling between the electrode and enzyme reactions of SOD could facilitate the development of the third-generation biosensor for 02". ...

Contrary to traditional fuel cells, biocatalytic fuel cells are in principle very simple in design [1], Fuel cells are usually made of two half-cell electrodes, the anode and cathode, separated by an electrolyte and a membrane that should avoid mixing of the fuel and oxidant at both electrodes, while allowing the diffusion of ions to/from the electrodes. The electrodes and membrane assembly needs to be sealed and mounted in a case from which plumbing allows the fuel and oxidant delivery to the anode and cathode, respectively, and exhaustion of the reaction products. In contrast, the simplicity of the biocatalytic fuel cell design rests on the specificity of the catalyst brought upon by the use of enzymes. 

Provided that the required enzymes can be immobilized at, and electrically communicated with, the surface of an electrode, with retention of their high catalytic properties and there is no electrolysis of fuel at the cathode or oxidant at the anode, or a solution redox reaction between fuel and oxidant, the biocatalytic fuel cell then simply... 

FIGURE 12.2 Schematic depicting a direct electron transfer process between an enzyme and the electrode, acting as an anode in this case. 

In redox mediation, to have an effective electron exchange, the thermodynamic redox potentials of the enzyme and the mediator have to be accurately matched. For biocatalytic electrodes, efficient mediators must have redox potentials downhill from the redox potential of the enzyme a 50 mV difference is proposed to be optimal [1, 18]. The tuning of these potentials is a compromise between the need to have a high cell voltage and a high catalytic current. Furthermore, an obvious requirement is that the mediator must be stable in the reduced and oxidized states. Finally, for operation of a membraneless miniaturized biocatalytic fuel cell, the mediators for both the anode and the cathode must be immobilized to prevent power dissipation by solution redox reactions between them. 

In this section, the enzymes, and associated substrates, used as biocatalysts in anodes are presented. For the development of biocatalytic anodes, there is a wide range of fuels available for use as substrates, such as alcohols, lactate, hydrogen, fructose, sucrose, all of which can be oxidized by biocatalysts. The fuel that is the most widely considered, however, in the context of an implantable biocatalytic fuel cell is glucose. We shall focus our attention on this fuel, but will mention briefly research on the use of some other fuels in biocatalytic anodes. 

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