Investigation of metal activation of a partially purified polyphenol oxidase enzyme electrode

Biosensors can be developed using different biological materials and immobilization technologies. Enzymes are generally used in biosensor construction, and some enzymes need metal ions or small organic molecules as a cofactor for their activation. Polyphenol oxidases can be activated by several metal ions such as Cu2+, Mg2+, Zn2+, Mn2+, and Ni2+. In this study, a new measurement method has been developed that is based on the metal ion activation of the polyphenol oxidase enzyme used in the biosensor preparation, especially to determine the concentration of Mg2+ ions. Polyphenol oxidase (PPO) (EC 1.10.3.1) was partially purified from potato (Solanum tuberosum) by using (NH4)(2)SO4 precipitation, dialysis, and lyophylization processes. As a result of this processes, approximately 30-fold purification was achieved for PPO. For construction of the biosensor, the enzyme was immobilized on the dissolved oxygen probe membrane using gelatin and glutaraldehyde (2.5%). Using the biosensor, we obtained responses for catechol in the absence and presence of Mg2+ ions. Differences between the biosensor responses were related to the concentration of Mg2+ ions. The biosensor response depends linearly on concentration of Mg2+ ions between 0.05 and 7.5mM. In the optimization studies, phosphate buffer (pH 7.0, 50mM) and 35 degrees C were determined to be the optimum conditions. This project will be a novel biosensor study and it might bring a new term, 'activation based biosensor' into the biosensor area.