Enzymes electrochemical processes

Electrochemical processes are particularly well suited for the manufacture of fine chemicals in view of their high sjjecificity (almost comparable to that offered by enzymes), the smaller number of steps required, adoption of milder conditions, lack of scale-up problems, avoidance of effluents, etc. The ease with which oxidation and/or reduction can be carried out with the practically mass-free clean electrons makes electrochemical processes well suited for such jobs, including paired synthesis in effect, we use electricity as a reagent . Consider a standard chemical oxidant like manganic or chromic sulphate. Here, a stoichiometric amount of the reduced salt will be formed the disposal of which can be a serious problem. If we adopt an electrochemical process, then the reduced salt is converted into the desired oxidized salt. 

The initial electrochemical and biological oxidation with xanthine oxidase are essentially identical. However, electrochemically 2,8-dioxyadenine the final product in the presence of xanthine oxidase is much more readily oxidizable than adenine 59) so that considerable further oxidation occurs. To the authors knowledge, 2,8-dioxyadenine is not a major metabolite of adenine in man or other higher organisms. Accordingly, it is likely that other enzymes accomplish further degradation of 2,8-dioxyadenine. The relationship between the products so formed and the mechanism of the reaction to the related electrochemical processes has yet to be studied. 

Ikariyama [2] described a unique method for the preparation of a glucose oxidase (GOD) electrode in their work. The method is based on two electrochemical processes, i.e. electrochemical adsorption of GOD molecules and electrochemical growth of porous electrode. GOD immobilized in the growing matrix of platinum black particles is employed for the microfabrication of the enzyme electrode. It demonstrated high performance with high sensitivity and fast responsiveness. 

An electron transfer type of enzyme sensor was thus fabricated by a electrochemical process. Although no appreciable leakage of ADH and MB from the membrane matrix was detected, NAD leaked slightly. To prevent this leakage, the ADH-MB-NAD/polypyrrole electrode was coated with Nation. A calibration curve is presented in Fig.25 for ethanol determination in an aquous solution with the enzyme sensor. Ethanol is selectively and sensitively determined in the concentration range from 0.1 nM to 10 mM. 

The pyridinium salt NAD 19a and its reduced form NADH 20a are important co-factors for many enzymes, fhe reduced form is involved in enzyme mediated reductions where it is converted to NAD. In natural systems, NAD is converted back to NADH by another enzyme-controlled process. Attempts to effect the direct electrochemical conversion of NAD to NADH are not very successful. Reduction on a mercury cathode at -1.1 V see on the first one-electron reduction wave leads to the radical-zwitterion, which reacts further to give dimers. Three stereoisomers of the 4,4 -dimer account for 90 % of the mixture and three 4,6 -dimers form the remainder [78]. Reduction at -1.8 V on the second reduction wave produces only 50 % of enzymatically active 1,4-NADH. The NAD analogue 19b shows related behaviour and one-electron reduction affords two diastereoisomers... 

Process control applications of enzyme electrochemical biosensors... 

The difficulty to transform CO2 into other organic compounds lies in its high thermodynamic stability. Typical activation energies for the dissociation and recombination ofC02 are of 535 and 13 kJ/mol, respectively [5], The activation can occur by photochemical or electrochemical processes, by catalytic fixation or by metal-ligand insertion mechanisms. As documented in different reviews, organometallic compounds, metallo-enzyme sites and well defined metallic surfaces are able to activate carbon dioxide [6-16],... 

Some methods for using enzymes in accelerating electrochemical processes have been developed and are shown in Fig. 17.9. In cases a and b there is an enzyme reaction whose intermediates, S, undergo electrochemical transformation at a much lower overpotential than the initial substrate, Sred... 

In c, d, and e we have the typical case of a bioelectrocatalyst where, through a mediator, there is electron transfer between the electrode and the enzyme active centre where the substrate is in its turn activated and reacts. In c the components are in solution in d and e the mediator or the enzyme are immobilized on the electrode surface, the electron transfer reaction occurring between mediator and electrode. In case/we have the ideal situation direct electron exchange between the electrode and active centre of the enzyme, the mediator being eliminated. It is, nevertheless, very difficult to reconcile the enzyme characteristics and the electrochemical process, and it continues to be important to find adequate mediators and enzyme immobilization procedures. 

Electrochemical Processes of Proteins and Enzymes at Command Interfaces... 

As pointed out before, to couple the redox enzyme to an electrode, a redox catalyst usually has to be applied that shuttles the electrons between the electrode and the protein (Fig. 3) (indirect electrochemical processes [19]). In certain cases, the transfer of the redox equivalents between the mediator and the cofactor also has to be catalyzed by an enzyme (see later discussions). 

According to the reaction scheme given in Fig. 26, the anaerobic oxidation of l-glycerol 3-phosphate was performed under the conditions of an indirect electrochemical process using a water-soluble ferrocene derivative as mediator in the presence of o-fruc-tose-1,6-diphosphate aldolase from rabbit muscle for the in situ generation of the carbohydrate product. With a mediator concentration fo 1.5 mM using about 80 U of immobilized enzyme, after 25 h a 75% turnover of the substrate L-glycerol phosphate... 

Pyruvate oxidase (Pyox) is a FAD- and thiamine diphosphate (ThDP)-dependent enzyme that catalyzes the reaction of pyruvate to give acetyl phosphate or vice versa (see Fig. 15). If used in the oxidative way, it can be activated and reactivated under nonaerobic conditions using ferrocene mediators. Kinetic parameters of the indirect electrochemical process using the enzyme incorporated into a biomimetic supported bilayer at a gold electrode have been reported [142]. Similarly, FAD-dependent amino oxidases may also be applied. 

Figure 4-12 Design of amperometric enzyme electrode based on anodic detection of hydrogen peroxide generated from oxidase enzymatic reaction (e.g., glucose oxidase) (A), and expanded view of the sensing surface showing the different membranes and electrochemical process that yield the anodic current proportional to the substrate concentration in the sample (B). (From Meyerhoff N, New in vitro analytical approaches for clinical chemistry measurements in critical care. Clin Chem I990 36 I570.)...

There are some exciting prospects—as yet unexplored—for the constructive interplay between electrochemical processes and membrane transport/ catalytic processes. This may lead to novel routes of organic chemical synthesis or removal of toxic substances from process streams. Many important enzymatic reactions involve oxidative or reductive transformations of the substrate. These usually require the participation of a cofactor which serves as the vehicle for transfer of electrons between the enzyme-bound substrate molecule and the participating oxidant or reductant in solution [71, 72], For such electron transfer to take place, both substrate molecule and cofactor must be localized in close proximity at the active site of the... 

About 300 pyridine nucleotide dependent enzymes are currently known. Many of them are in widespread use for analytical purposes. Therefore the determination of the coenzyme NAD(P)H is of great importance. In contrast to the enzyme-catalyzed oxidation of NAD(P)H, its anodic oxidation proceeds in two separate one-electron steps with radical intermediates (Elving et al., 1982). It requires an overvoltage of about 1 V. Furthermore, electrode fouling by the reaction products makes the electrochemical process poorly reproducible. Owing to the high electrode potential, other oxidizable substances interfere signifi-... 

Feedback. When an oxidoreductase enzyme is immobilized at the specimen surface, a redox mediator present in solution may be recycled by the diffusion-limited electrochemical process at the tip and electron exchange with the enzyme active site as described in Sec. I.C. The mass transport rate is defined by the tip radius and height of the tip above the specimen. The tip current depends on the mass transport rate and the enzyme kinetics. Kinetic information may therefore be obtained from the dependence of tip current on height, i.e., an approach curve. When the mediator is fed... 

Biomedical science and health care Electrochemical processes characteristic of living systems are reviewed, including such aspects as applications based on neuroscience, enzyme biocatalysis, adhesion and cell fusion, and electrophoresis. 

A single enzyme from the microbial cell is sufficient for the dehydrogenation of lactate to pyruvate, if the electron mediator is regenerated by a nonenzymic method, e. g., by an electrochemical process (62). 

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