ELECTROCATALYTIC OXIDATION OF REDUCED NICOTINAMIDE COENZYMES AT METHYLENE GREEN-MODIFIED ELECTRODES AND FABRICATION OF AMPEROMETRIC ALCOHOL BIOSENSORS

Chemically modified electrodes with Methylene Green adsorbed on the graphite surface and incorporated into carbon paste exhibit excellent electrocatalytic ability for oxidation of NADH. Alcohol dehydrogenase, nicotinamide adenine dinucleotide (NAD+) and mediator were incorporated into a carbon paste matrix to yield an alcohol sensor. The enzyme and cofactor retain their bioactivity within the matrix. The surface of the sensor is protected by coverage with a poly(ester sulphonic acid) cation exchanger to form a membrane, which effectively prevents the aqueous soluble species from dissolving out from the enzyme electrode and reduces or eliminates the interference from electroactive anions. The oxidation current of the NADH formed by enzymatic reaction serves as the response for target analytes. The reagentless sensor shows steady-state signals within 50 s, owing to the intimate contact between the biocatalytic and sensing sites. The influence of various experimental conditions was explored for optimum analytical performance. The sensor remained relatively stable for about 15 days.