Mechanisms of Cigarette Smoke Toxicity: the Inactivation of Human Alpha-1-proteinase Inhibitor by Nitric Oxide/isoprene Mixtures in Air

A mixture of nitric oxide (NO) and isoprene in air has been studied as a model for gas-phase cigarette smoke. We have shown that this model system duplicates many of the properties of cigarette smoke including the inactivation of human alpha-1-proteinase inhibitor (a1PI). In this study, buffered solutions of a1PI were exposed to puffs of air containing 300 ppm NO and 400 ppm isoprene. Bubbling of the NO/air/isoprene gas stream directly through buffered protein solutions causes a1PI to undergo a fast loss of inhibitory capacity. This fast inactivation is not observed when a1PI is exposed to aqueous extracts of the NO/air/isoprene mixture. Both direct exposure and exposure to aqueous extracts, however, cause a1PI to undergo a slow loss of activity that continues for several days as the protein is incubated in the buffer solutions. Gas-phase cigarette smoke has already been shown to cause this same two-phase inactivation of a1PI. The inactivation of a1PI by the model system is dependent on the presence of oxygen in the gas stream, suggesting that the oxidation of nitric oxide to nitrogen dioxide in air is involved in the formation of the inactivating species. The nature of these species remains to be determined; however, small alkoxyl or peroxyl radicals (such as are spin-trappable from gas-phase smoke as well as from the NO/air/isoprene system) do not appear to inactivate a1PI. One possibility is that the inactivating species are metastable compounds formed by radical processes in the gas phase of both cigarette smoke and our model system. Our data suggest that one possible class of species is peroxynitrates.