The Electrical Contacting of Enzymes in Mediator-containing Graphite Paste Composites

The Electrical Contacting of Enzymes in Mediator-containing Graphite Paste Composites 

Carbon paste electrodes have also been used as supports for multienzyme systems. For example, AChE and ChO have been coimmobilized in carbon pastes, either with monomeric TTF [157] or flexible ferrocene-containing polymers [158] as the electron mediator. The hydrolysis of acetylcholine is biocatalyzed by AChE, and then the choline produced is oxidized by the electrically contacted ChOx giving an analytical amperometric signal corresponding to the acetylchoKne concentration. 

It should be noted, however, that carbon pastes do not offer a general route to any desired assembly. Some enzymes, for example, fructose dehydrogenase [136, 159] and aldose dehydrogenase [160], have defied all efforts at incorporation into the matrix without loss of activity. It is possible to introduce these enzymes to the surface of preformed carbon paste electrodes while retaining their biocatalytic activity, but this procedure does not exploit the many advantages of the carbon paste technique [79, 80]. The construction and optimization of multicomponent matrices composed of graphite powder, organic oils, enzymes, redox mediators, cofactors, and other materials is still a subject of intensive study. 

We have already noted that the random functionahzation of the protein hackhone with electron relay groups leads to a mixture of products, the average of which is detected electrochemically. In order to accomplish the hest possible electron contacting, the mediator should be selectively placed in an optimum position between the redox-center and the enzyme periphery. In the case of surface-confined enzymes, the orientation of the enzyme-mediator assembly with respect to the electrode should also be optimized. These requirements have been addressed by the use of mediators that are covalently attached to the enzyme cofactor or to both the cofactor and to the electrode surface. This specific covalent attachment positions the mediator at exactly the point that is required, leading to much more homogeneous behavior of the many enzyme molecules in a sample. 

Electrical Contacting of Enzymes by Reconstitution of Apo-flavoenzymes with Relay-FAD Cofactor Units 

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