A New Approach to the Co-immobilization of Alcohol Dehydrogenase and NAD on Glutaraldehyde-activated Sepharose and Its Application to the Enzymatic Analysis of Ethanol

Abstract

A new method is described for the co-immobilization of enzyme alcohol dehydrogenase (ADH) and coenzyme NAD onto Sepharose gel. In order to increase the degree of freedom of NAD, spacer molecules were introduced to the support by allowing it to repeatedly react with hexamethylenediamine and glutaraldehyde, and ADH and NAD were co-immobilized to the spacer-modified Sepharose. The results suggest that the re-usability of Sepharose-bound ADH and NAD (AN-Seph) depended both on the concentration and the reaction time of glutaraldehyde to a great extent. Furthermore, it is suggested that AN-Seph prepared under the optimum conditions might be applicable to the determination of ethanol.