18] Convenient method for enzymic synthesis of [14C]nicotinamide riboside

Abstract

This chapter discusses the convenient method for enzymic synthesis of [ 14 C]nicotinamide riboside. Nicotinamide riboside can be prepared from nicotinamide adenine dinucleotide (NAD) by the procedures described by Kaplan and Stolzenbach. These procedures involve cleavage of the pyrophosphate bond of NAD with snake venom phosphodiesterase and separation of the nicotinamide mononucleotide (NMN) from 5'-AMP by Dowex 1-formate column chromatography. The NMN is then converted to nicotinamide riboside, by hydrolysis, with prostatic monoesterase. An alternative method for the preparation of nicotinamide riboside relies on the degradation of NAD, by a crude enzyme preparation, from Proteus vulgaris OX-19. This preparation normally degrades NAD to free nicotinic acid via NMN, nicotinamide riboside, and nicotinamide. Because of the heat stability of the enzymes catalyzing reactions, the enzymes that degrade the riboside and nicotinamide can be inactivated by heat treatment, providing a preparation that gives a high yield of nicotinamide riboside. The procedure described in the chapter provides a convenient method for the rapid preparation, in high yield, of [ 14 C]nicotinamide riboside, with essentially 100% radiochemical purity, starting with labeled NAD.