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ABSTRACT
A novel amperometric biosensor for the determination of ethanol through one-step drop coating of a nanobiocomposite on a glassy carbon electrode is presented. The measurement of ethanol concentration is based on the production of reduced form of the β-nicotinamide adenine dinucleotide (NADH), which is the product of enzymatic reaction catalyzed by alcohol dehydrogenase. The enzyme was immobilized in a polystyrene sulfonate-multiwalled carbon nanotube composite that offers a stable environment for alcohol dehydrogenase. The performance of polystyrene sulfonate–multiwalled carbon nanotube–alcohol dehydrogenase nanobiocomposite–modified glassy carbon electrode was evaluated by cyclic voltammetry, electrochemical impedance spectroscopy, and amperometry. Enzymatic oxidation of ethanol was realized in the presence of NAD+ and was determined amperometrically. The concentrations of NAD+ and enzyme were optimized. Ethanol determination was performed using 7 IU alcohol dehydrogenase/electrode in the presence of 5 mM NAD+ in pH 8.8 pyrophosphate buffer. The polystyrene sulfonate–multiwalled carbon nanotube–alcohol dehydrogenase/glassy carbon electrode biosensor showed high sensitivity for ethanol of 15.6 µA·mM−1·cm−2 with a linear range from 70 to 420 µM. A detection limit of 19 µM was obtained and negligible interferences from glucose, ascorbic acid, uric acid, and acetaminophen were observed.