Enzyme glucose oxidase electrodes

Maines A, Ashworth D, Vadgama P. Diffusion restricting outer membranes for greatly extended hnearity measurements with glucose oxidase enzyme electrodes. Analytica Chimica Acta 1996, 333, 223-231. 

Figure 19. (A) The reconstitution of apo-glucose oxidase on a PQQ-FAD monolayer assembled on an Au electrode. (B) Cyclic voltammograms of the PQQ-FAD-reconstituted glucose oxidase on an Au electrode (a) in the absence of glucose (b) with glucose, 80 mM. Recorded in 0.1 M phosphate buffer, pH 7.0, under Ar, at 35°C potential scan rate, 5 mV s. Inset calibration curve corresponding to the amperometric responses (measured by chronoamperometry, = 0.2 V vs. SCE) of the PQQ-FAD-reconstituted glucose oxidase enzyme electrode at different concentrations of glucose.

In actual studies, the problem is more complicated, since one must determine which case applies from the experimental results, that is, // as a function of o>, Fp, C, and . Procedures and diagnostic criteria are given in reference 80. Also discussed there are several experimental studies featuring analysis of results using this approach. In addition, this reference covers situations where the catalyst and the electron carrier are different species, such as in the glucose oxidase enzyme electrode. 

Mell L.D. and Maloy J.T. (1976) Amperometric response enhancement of the immobilized glucose oxidase enzyme electrode. Ami/. Chem., AS, 1597-1601. 

The enzyme is used in an enzyme electrode in which a tube is sealed at its lower end with a cellulose acetate membrane. An outer membrane of collagen is also attached to the end of the electrode tube and glucose oxidase enzyme is contained in the space between the two diaphragms. 

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. 

Scheme 8 Assembly of a photoisomerizable glucose oxidase monolayer electrode and the reversible photoswitchable activa-tion/deactivation of the bioelectrocatalytic functions of the enzyme electrode.

The influence of different spacer and coupling molecules on the relative performances of seventeen glucose oxidase-nylon electrodes (NGO) fabricated from the same batch of fresh enzyme have been conveniently established in the FIA mode with standard glucose( 1 mM) ... 

Polypyrrole thin film doped with glucose oxidase (PPy-GOD) has been prepared on a glassy carbon electrode by the electrochemical polymerization of the pyrrole monomer in the solution of glucose oxidase enzyme in the absence of other supporting electrolytes. The cyclic voltammetry of the PPy-GOD film electrode shows electrochemical activity which is mainly due to the redox reaction of the PPy in the film. Both in situ Raman and in situ UV-visible spectroscopic results also show the formation of the PPy film, which can be oxidized and reduced by the application of the redox potential. A good catalytic response to the glucose and an electrochemical selectivity to some hydrophilic pharmaceutical drugs are seen at the PPy-GOD film electrode. 

Polypyrrole Film Formation in Glucose Oxidase Enzyme Solution. Cyclic voltammograms recorded in the GOD and pyrrole solution showed an anodic peak current (E = 1.08 V), which suggested the polymerization of pyrrole in the above solution. However, the polymerization potential moved toward the more positive direction compared to the polymerization potential of PPy doped with Cl ( pa < 1.0 V). This is due to the fact that the polymerization is more difficult to take place in enzyme solution than in Cl solution because the enzyme solution is a much weaker electrolyte than NaCl it may also be due to the less conductive nature of the PPy-GOD film as compared to that of the PPy-Cl film. The polymerization current level was much lower in the enzyme solution than in the Cl solution because of the poor charge-transport property of the enzyme protein molecules. It was found that the constant current method was more suitable than the controlled potential method for making the PPy-GOD film on the GC electrode. 

Figure 17.3 The pH dependence of a glucose oxidase membrane electrode at different enzyme loadings and different glucose concentrations electrode surface, 0.22 mm electrode potential, +600 mV against Ag/AgCl phosphate buffer, 0.66 molT (reproduced with the permission of Elsevier Science Publishers BV). 

Enzymes are often employed in the chemical layer to impart the selectivity needed. We saw an example of this in Chapter 13 when discussing potentiometric enzyme electrodes. An example of an amperometric enzyme electrode is the glucose electrode, illustrated in Figure 15.4. The enzyme glucose oxidase is immobilized in a gel (e.g., acrylamide) and coated on the surface of a platinum wire cathode. The gel also contains a chloride salt and makes contact with silver-silver chloride ring to complete the electrochemical cell. Glucose oxidase enzyme catalyzes the aerobic oxidation of glucose as follows ... 

The response of a surface antigen-antibody reaction can also be mediated by an electron transfer reagent. This has been demonstrated by Robinson et al. in their studies on electrochemical immunoassays for hCG and thyroxine. In the analysis of hCG, a two-site amperometric immunoassay was developed in which monoclonal capture antibodies were immobilised on the surface of a glassy carbon electrode. A second antibody against hCG was labelled with glucose oxidase. The electrode was used both to separate free from bound enzyme-antibody conjugate and to assay the enzyme activity electrochemically by use of dimethylaminomethyl ferrocene as an electron transfer mediator. This method was found to correlate well with an immunoradiometric assay. In the analysis of thyroxine, another ferrocene derivative, namely ferrocenemonocarboxylic acid, was used as the electron transfer mediator. ... 

The electropolymerlzatloa was done with the enzyme already Immobilized on the electrode surface, resulting in a complete glucose biosensor. The limiting factor is now the electropolymerized film and the length of time that common interferents can be screened from the electrode surface. The cross-linked glucose oxidase enzyme is stable for months cross-linked on RVC, with or without the polymer film. Although the polymer film does provide additional enzyme stability. 

Schuhmann, W., Ohara, T., Heller, A., and Schmidt, H.-L., Electron transfer between glucose oxidase and electrodes via redox mediators bound with flexible chains to the enzyme surface, J. Am. Chem. Soc., 113, 1394-1397 (1991),... 

Biosensors are often based on the use of an enzyme with specific response. Polymerization of pyrrole can be carried out in aqueous solution at moderate pH values. In those conditions enzymes can be entrapped in the growing film of the polymer formed on the electrode. This has been applied in particular to the glucose-oxidase enzyme, in order to build a glucose sensor [412 18]. 

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