The use of enzymes requiring a cofactor as substrate in organic synthesis is still a problem since the cofactors are expensive. This study deals with a new approach consisting of using fragments of NAD+. Three fragments of NAD(H) are examined. The activities of NMN+ and NMNH are greatly improved by the addition of adenosine in ethanol oxidation and in cyclohexanone reduction, respectively. Nicotinamide monocucleoside is not active in the ethanol oxidation but the addition of AMP promotes this reaction.

Kinetic constants of horse-liver alcohol-dehydrogenase-mediated oxidation of alcohol by nicotinamide mononucleotide and nicotinamide ribose were determined and the role of different adenine moieties complementing the reaction mixtures was investigated. Five nicotinamide ribose analogs were synthesized and their activities as NAD' inhibitors and as cofactors in this dehydrogenase-mediated oxidation of alcohol were assayed. In the light of these results, structural requirements of the pyridine ribofuranosyl part of the NAD' are discussed.In a recent work, we have shown that the enzymatic activity of horse-liver alcohol dehydrogenase (HLADH), an NAD' -requiring enzyme, was partly retained when NMN+ or the nicotinamide ribose (NR') were used as cofactors. Moreover, the efficiencies of these NAD' fragments were strongly improved when the medium was supplemented with adenosine and AMP respectively [I].Many NAD + analogs [2] have been synthesized with, in particular, the nicotinamide-linked ribofuranosyl residue substituted by linear alkyl groups and assayed as cofactors. Only the analog with the butyl group was found to be slightly active, showing the importance of the nicotinamide-linked ri bofuranosyl.We describe in the present work the synthesis of four N R + analogs I -IV, where a cyclopentane diversely substituted is attached to the nicotinamide moiety, and one nicotinamide derivative V, where the nitrogen atom is substituted by a C 5 chain possessing four hydroxyl groups. We also describe the abilities of these derivatives to function as cofactors with HLADH and, in order tentatively to elucidate how adenosine and AMP activate the functional fragments, we present some kinetic studies of the enzymatic reactions. MATERIALS AND METHODSNMN' and NR' were prepared and purified according to [I] and [3] respectively. HLADH was from Sigma; NAD', ADP, AMP and adenosine were from Fluka.The enzyme kinetics were carried out in a 1-ml ultraviolet

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