Synthesis of R and S tritiated reduced beta-nicotinamide adenine dinucleotide 2' phosphate.
Nicotinamides are ubiquitous cofactors used by many biological systems as redox agents. Stereospecifically labeled cofactors are useful in many studies of nicotinamide-dependent enzymes. Enzyme-directed synthesis of these cofactors is rather common but their stability imposes significant challenges on yield, purity, and preservation. This paper presents the stereospecific synthesis of reduced R- and S-[4-3H] beta-nicotinamide adenine dinucleotide 2' phosphate (NADPH). The method of Valera et al. [Biochem. Biophys. Res. Commun. 148 (1987) 515] was modified to a synthetic procedure that produces both isotopic diastereomers within 2h with an improved yield of 75-90% after purification and lyophilization. In the synthesis, [4-3H]NADP+ was generated as an intermediate (which can be isolated if desired). The specific radioactivities reported here are 2.7 and 1.1 Ci/mmol for the S and R diastereomers, respectively. Specific radioactivities ranging from carrier-free to trace labeling can be achieved with a minor change to the procedure.
McCracken
Jocelyn A
JA
Department of Chemistry, University of Iowa, Iowa City, IA 52242, USA.
Wang
Lin
L
Kohen
Amnon
A
D000426:Alcohol Dehydrogenase; D002851:Chromatography, High Pressure Liquid; D007553:Isotope Labeling; D009249:NADP / Q000138:chemical synthesis* / Q000302:isolation & purification; D010084:Oxidation-Reduction; D013056:Spectrophotometry, Ultraviolet; D013237:Stereoisomerism; D013997:Time Factors; D014316:Tritium
D014316:Tritium; D009249:NADP; D000426:Alcohol Dehydrogenase