Investigating the Electrocatalytic Oxidation of Dihydronicotinamide Adenine Dinucleotide at Nitrogen-Doped Carbon Nanotube Electrodes: Implications to Electrochemically Measuring Dehydrogenase Enzyme Kinetics

摘要

Nitrogen-doped carbon nanotubes (N-CNTs) have been shown to be electrocatalytic toward the oxidation of dihydronicotinamide adenine dinucleotide (NADH), the reduced form of the coenzyme necessary for enzymatic turnover in NAD+-dependent dehydrogenases. The observed oxidation potential of the electrocatalyst, however, still shows a significant overpotential, suggesting that even for effective electrocatalysts, electrooxidation may be kinetically controlled. We demonstrate using the Koutecky-Levich rotating disk electrode technique that the observed electron transfer rate constant (kobs) is a function of potential over a wide potential window; however, kobs could only be accurately measured for a portion of that window for the electrocatalytic N-CNTs. More importantly, electrochemically measured enzyme kinetics, acquired after adsorption of glucose dehydrogenase onto the N-CNTs, are never independent of potential, even when the electron transfer rate constant is too fast to measure by the rotating disk technique. Thus, electrochemically obtained kinetics (e.g., KM app and Vmax) are actually measuring the electrochemical kinetics of NADH oxidation at the electrode surface, rather than the spontaneous and potential-independent enzymatic turnover.