The preparation and purification of no gas and the use of no releasers: The application of no donors and other agents of nitrosative stress in biological systems

Cylinders and lecture bottles are often the source of nitric oxide (NO) in studies of the biological chemistry of this remarkable molecule. The NO from both sources will probably contain NO2 (and N2O) formed by disproportionation of NO. The NO2 must be removed by passing the NO through a thoroughly deoxygenated sequence of traps containing sodium hydroxide solution and then water. The presence of NO2 in aqueous solutions of NO may be determined readily using 2,2' azino(3-ethylbenzothiazoline-6-sulfonic acid). NO2 oxidizes this compound to a long-lived radical anion, the concentration of which may be determined spectrophotometrically. The formation of NO by the decomposition of nitrous acid (via its disproportionation to nitrate and NO) and by the use of commercially available NO-releasing compounds with defined half-lives are also discussed. Other reactions that lead indirectly to the formation of NO are noted. In all cases, care must be taken to exclude oxygen to minimize as much as possible the formation of NO2 (and, consequently, the nitrosating agent N2O3)The uses of these methods for generating NO and the reactivity of related compounds are illustrated with examples of studies of nitrosative stress.