Some observations on the preparation and properties of dihydronicotinamide-adenine dinucleotide

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Abstract
NADH2 prepared by enzymic reduction of pure NAD by the method of Rafter & Colowick (1957), and isolated as the sodium salt, gives higher maximum rates of reduction of acetaldehyde with liver alcohol dehydrogenase at pH6 than a number of commercial preparations of high purity. Maximum values of 2.4 for the extinction ratio E260/E340 and 2% for the proportion of inactive material absorbing at 340 m[mu] are taken as criteria of a satisfactory preparation, and are lower than those for most commercial preparations and for products obtained by chemical reduction of NAD. Methods involving enzyme inactivation by heat or isolation by drying aqueous solutions are not satisfactory for the preparation of pure NADH2. The compound is not stable at room temperature, nor in concentrated solutions. It appears that an inhibitor of liver alcohol dehydrogenase which competes with the coenzyme is formed, and is present in some commercial preparations. Kinetic studies of the inhibition of liver alcohol dehydrogenase by adenosine diphosphate ribose are briefly reported, and the results are discussed with reference to the possible magnitude.