Design of a Reagentless Enzymatic Amperometric Alcohol Biosensor: Yeast Alcohol Dehydrogenase and Nicotinamide Adenine Dinucleotide on Vertically Aligned Carbon Nanofibers

A reagentless amperometric enzymatic biosensor was constructed on a carbon substrate for the detection of ethanol, methanol, and isopropanol. Yeast alcohol dehydrogenase (YADH), an oxidoreductase enzyme, and its cofactor nicotinamide adenine dinuleotide (NAD⁺) were immobilized by adsorption and covalent attachment to the carbon substrate. Carbon nanofibers grown by plasma enhanced chemical vapor deposition (PECVD) were chosen as the electrode material due to their excellent structural and electrical properties.

Electrochemical techniques were employed to test the function and performance of the constructed biosensor. Characterization of the electrode was performed using NADH. This allowed the function of the electrode to be examined as well as to determine the oxidation peak potential of NADH. Subsequently, amperometric measurements were conducted for the detection of ethanol, methanol, and isopropanol to determine the response in electrical current as a result of an increase in analyte concentration. The storage stability, reusability, and response time of the biosensor was also examined.

Carbon nanofibers were found to be an effective strategy for building a biosensor. At a working potential of 0.9 V, electrodes with covalently attached and adsorbed enzyme were found to provide a strong current response. The electrical current responses to methanol and isopropanol were found to be on average one order of magnitude lower than that of the ethanol experiments. The response was stable, storage stability of the sensors was excellent, and the response time to analyte additions was quick.