Immobilization of Glucose Oxidase on 4-(pyrrole-1-yl) Benzoic Acid Functionalized Carbon Nanotubes for Direct Electrochemistry and Glucose Sensing

In recent years, carbon nanotubes (CNT) have gained considerable attention, because of their remarkable electronic and mechanical properties, which have made them extremely attractive for a wide range of sensing applications from structural materials to nanoelectronic components. The ability of CNT-modified electrodes to promote electron transfer reactions has been documented in connection with important biomolecules [1]. Our goal is to explore new applications of CNT as an electrode material in facilitating the electron transfer between enzyme molecule and electrode in the bioelectrocatalytic system for glucose oxidation that is of potential utility for bioelectronic devices such as biosensors and biofuel cells.

Our research focuses on the direct electrochemical performance of glucose oxidase (GOx) immobilized on 4-(pyrrole-1-yl) benzoic acid (PyBA) modified multi-walled carbon nanotubes (CNTs). GOx is a homodimer containing two tightly bound flavin adenine dinucleotide (FAD) redox centers embedded deeply in the enzyme [2]. Modification of carbon nanostructures allow to obtain thin and organized films, causing a rapid and effective transfer of electrons between the active center of the biocatalyst (GOx) and the electrode surface. The presence of PyBA in our composite film significantly improves their stability and introduces new functional groups that have great importance in the enzyme immobilization process onto the CNTs-modified electrode [3].

In this work stable immobilization and direct electron transfer of glucose oxidase were achieved on the composite film modified glassy carbon electrode. The resulting electrode gave a well-defined redox peaks with a formal potential of about −440 mV (vs. Ag/AgCl) in 0.1 phosphate buffer solution pH=7.0. The electron transfer rate constant was estimated to be 3.15 s^-1, due to the combined contribution of CNTs/PyBA and GOx. Furthermore, the method for detecting of glucose was proposed based on the decrease of oxygen caused by the enzyme-catalyzed reaction between GOx and glucose. The low calculated apparent Michaelis–Menten constant (K_M^app) was 10.2 mM, implying the high enzymatic activity and affinity of immobilized enzyme for glucose. It can reasonably be expected that this observation might hold true for other noble carbon nanostructure-electroactive protein systems, providing a promising platform for the development of biosensors and biofuel cells.

References

[1] J.J.Gooding, R.Wibowo, J. Liu, W. Yang, D. Losic, S. Orbons, F.J. Mearns, J.G. Shapter, D.B. Ilibbert, J. Am. Chem. Soc. 125 (2003) 9006.

[2] H.J. Hecht, H.M. Kalisz, J. Hendle, R.D. Shmid, D.Schomburg, J. Mol. Biol. 229 (1993) 153.

[3] B. Kowalewska, M. Skunik, K. Karnicka, K. Miecznikowski, M. Chojak, G. Ginalska, A. Belcarz, P.J. Kulesza, Electrochim. Acta 53 (2008) 2408.