High-performance ion-exchange separation of oxidized and reduced nicotinamide adenine dinucleotides

High-performance anion-exchange chromatography of oxidized and reduced forms of nicotinamide adenine dinucleotide (NAD) and nicotinamide adenine dinucleotide phosphate (NADP) on a Pharmacia Mono Q anion-exchange column is reported. Microgram quantities of all four nucleotides can be separated at pH 7.7 in approximately 20 min. For preparative purposes, greater than 7 mg of NADH can be purified in a single injection, and the peak fractions have an A260 of greater than 80 OD units with an A260/A340 ratio of 2.25.