Nicotinamide-Adenine Dinucleotides (NAD, NADP, NADH, NADPH)

Publisher Summary The nicotinamide dinucleotides, NAD and NADH, and nicotinamide dinucleotide phosphates, NADP and NADPH, occur in all living cells. Within cells, the nicotinamide dinucleotides are not distributed evenly over the various subcellular regions, for example, mitochondria and cytosol. In whole blood, the nicotinamide dinucleotides occur only within the cell elements. This chapter describes the methods that allow the determination of the state of reduction of nicotinamide dinucleotides by the measurement of the content of NADH and NAD or NADPH and NADP. The catalytic method also offers a simple determination of the total nicotinamide dinucleotide content. Nicotinamide dinucleotides can be determined by three different enzymatic methods. The spectrophotometric absorption allows particularly accurate measurement when the content of nicotinamide dinucleotides in the extracts is sufficiently high. Measurements at lower concentrations are possible only with special equipment (high-sensitivity difference spectrophotometer). The lower contents of nicotinamide dinucleotides can be measured by fluorimetric methods. Both methods are stoichiometric substrate assays. Even greater sensitivity is possible in the measurement of nicotinamide dinucleotides by the catalytic assay.