5,5-Dithiobis(2-nitrobenzoic acid) pyrene derivative-carbon nanotube electrodes for NADH electrooxidation and oriented immobilization of multicopper oxidases for the development of glucose/O2 biofuel cells

We report the functionalization of multi-walled carbon nanotubes (MWCNTs) electrodes by a bifunctional nitroaromatic molecule accomplished via pi-pi interactions of a pyrene derivative. DTNB (5,5'-dithiobis(2-nitrobenzoic acid)) has the particularity to possess both electroactivable nitro groups and negatively charged carboxylic groups. The integration of the DTNB-modified MWCNTs was evaluated for different bioelectrocatalytic systems. The immobilized DTNB-based electrodes showed electrocatalytic activity toward the oxidation of the reduced form of nicotinamide adenine dinucleotide (NADH) with low overpotential of -0.09 V vs Ag/AgCl at neutral pH. Glucose dehydrogenase was successfully immobilized at the surface of DTNB-based electrodes and, in the presence of NAD(+), the resulting bioelectrode achieved efficient glucose oxidation with high current densities of 2.03 mA cm(-2). On the other hand, the aromatic structure and the negatively charged nature of the DTNB provoked orientation of both laccase and bilirubin oxidase onto the electrode, which enhanced their ability to undergo a direct electron transfer for oxygen reduction. Due to the proper orientation, low overpotentials were obtained (ca. 0.6 V vs Ag/AgCl) and high electrocatalytic currents of about 3.5 mA cm(-2) were recorded at neutral pH in O-2 saturated conditions for bilirubin oxidase electrodes. The combination of these bioanodes and bilirubin oxidase biocathodes provided glucose/O-2 enzymatic biofuel cells (EBFC) exhibiting an open-circuit potential of 0.640 V, with an associated maximum current density of 2.10 mA cm(-2). Moreover, the fuel cell delivered a maximum power density of 0.50 mW cm(-2) at 0.36 V.