Biamperometric determination of ethanol, lactate and glycerol using immobilised enzymes in flow streams

Abstract

Alcohol dehydrogenase was successfully immobilised on a nylon tube and on silanised glass beads packed in a short column, by means of covalent bonding. Lactic dehydrogenase and glycerol dehydrogenase were immobilised on titanium dioxide beads by adsorption, and on silanised glass beads by covalent bonding. All immobilisations by covalent bonding were achieved by means of glutaraldehyde cross-linking. These enzyme reactors were used in a flow system to measure ethanol, lactate and glycerol by catalysing their reaction with NAD. The NADH produced was coupled with hexacyanoferrate(III) ion, using diaphorase as a catalyst. The hexacyanoferrate(II) ion produced was measured biamperometrically using two open-tubular carbon electrodes. Measurement of standards in serum control samples was demonstrated.