Reaction in dimethyl sulfoxide of nicotinamide 8-bromoadenine dinucleotide with the disodium salt of 3-mercaptopropionic acid afforded nicotinamide-8-(2-carboxyethylthio)adenine dinucleotide, a new NAD' analogue functionalized at the adenine C-8 position by an w-carboxylic side chain. Carbodiimide coupling of the latter derivative to high-molecular-weight water-soluble (polyethyleneimine, polylysine) and insoluble (aminohexyl-Sepharose) polymers gave the corresponding macromolecular NAD+ analogues. These derivatives have been shown to be enzymically reducible. The polyethyleneimine analogue showed a substantial degree of efficiency relative to free NAD ' with yeast alcohol dehydrogenase (47 %) but a considerably lower one with rabbit muscle lactate dehydrogenase (3 %); the polylysine analogue showed a low degree of efficiency with both enzymes (5 -6 %).The use in the industrial, analytical or biomedical field of enzymes immobilized by physical entrapment in porous structures permeable to low-molecularweight substrates and products is becoming increasingly interesting. However, this immobilization technique, to be economically applicable also to enzymes utilizing readily dissociable coenzymes (like the NAD' and NADP'-dependent dehydrogenases), requires coupling of the cofactor, through a stable linkage, to hydrosoluble polymers, in such a way as to assure retention of coenzymic activity. In fact, by entrapment in the porous structure of such a macromolecular soluble cofactor derivative together with the enzyme (or multienzyme system) its loss by diffusion is prevented, and the interaction between coenzyme and enzyme can be satisfactorily performed with both in solution.Recently the synthesis and coenzymic activity of macromolecular soluble NAD' [l -51 and NADP'[6] derivatives from these and other laboratories has been reported. In all these derivatives the site of attachment, through a suitable linkage, of the cofactor molecule to the polymer was the exocyclic adenine amino group (P). In pursuing our studies[4] on macromolecular soluble NAD + derivatives we were interested in synthesising new NAD' derivatives of this type attached at a different site of the adenine nucleus to the same polymers, and to compare their coenzymic efficiencies to the ones of the corresponding N6 derivatives.

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