Electrochemistry of reconstituted glucose oxidase on carbon paste electrodes

Abstract

Flavine Adenine Dinucleotide (FAD) was covalently immobilised onto glassy carbon matrix using a 13-carbon atom long spacer arm. FAD modified electrodes offer a convenient handle for immobilising glucose oxidase (GOD) enzyme for direct electron transfer. The electrochemical characteristics of immobilised FAD were compared with free FAD in solution at blank paste electrode surface. The prominent peaks at −475 mV and −601 mV indicate the positive coupling of FAD onto the carbon matrix. In our case we observed that reduction peaks are well separated from the oxidation and the immobilised FAD shows more reversible behavior compared to the free FAD in solution. GOD apoenzyme has been prepared by acidification of GOD solution (5 mg/ml) in buffer (100 mM sodium acetate) using ammonium sulfate solution at pH 1.4. The apoenzyme was coupled to the FAD modified matrix by incubating the matrix with the solution of apoenzyme for 4–12 h. The paste electrode with reconstituted GOD was investigated for its electrochemical characteristics and for its response with substrate (glucose solution). Our major finding is that the reconstituted enzyme shows better electron transfer rates compared to normal enzyme. The reason for this can be attributed to the long spacer arm holding the electroactive FAD which facilitates better electron transfer between enzyme redox center and electrode surface. The above technique appears to be a promising approach to be used in biosensor application.