Automated assay for nicotinamide adenine dinucleotide (NAD+)

Sulfur mustard (HD), a vesicating chemical warfare compound, has been shown to deplete the nicotinamide adenine dinucleotide (NAD+) content in several cell systems and tissues. This NAD+ depletion has been proposed as an indicator of HD exposure and can be used to evaluate potential antivesicant compounds. To examine NAD+ levels, an automated method based on the alcohol dehydrogenase cycling assay of Jacobson and Jacobson and utilizing a Cobas FARA clinical analyzer has been developed. Automation of this assay led to smaller sample volumes and more efficient use of personnel and resources. The usefulness of this automated method was tested by evaluating the protection, if any, by the topical application of vitamin D or betamethasone against HD-induced NAD+ depletion in skin punches from the hairless guinea pig. The results showed that the samples exposed to HD exhibited significant decreases in NAD+ levels when compared with controls. However, neither vitamin D nor betamethasone demonstrated protection against HD-induced NAD+ depletion. In fact, betamethasone exacerbated the NAD+ depletion when compared with the HD exposed group. This assay appears to be useful for testing potential antivesicant compounds using both in vivo and in vitro exposure systems.