FLOW-INJECTION ANALYSIS OF ETHANOL BASED ON OXYGEN-CONSUMPTION USING ALCOHOL-DEHYDROGENASE AND NADH OXIDASE FROM BACILLUS-LICHENIFORMIS

A flow-injection analysis system for the determination of ethanol was developed incorporating an oxygen electrode and an enzyme reactor pack ed with Sepharose 4B on which alcohol dehydrogenase (ADH) and NADH oxi dase (NOD) from Bacillus licheniformis were co-immobilized. Although t he response of the system depended on the concentration of FAD added i nto a carrier solution, it was linearly related to the ethanol concent ration over the range 0.25 - 1.0 mM, even in the absence of FAD, by co ntrolling the immobilized amount of ADH and NOD. The linear relationsh ip between the ethanol concentration and the response in the presence of 60 muM FAD was observed over the range 0.1 - 1.5 mM. When the syste m was applied to the determination of ethanol in various kinds of alco holic beverages, the results obtained under both conditions were compa rable to those obtained by the F-kit method.