Mechanistic studies of nitric oxide formation from s-nitrosothiols

Abstract

A study of the reactions of S-nitrosothiols in solution was undertaken. S-Nitrosothiols were known to produce NO, a physiologically important regulatory chemical, in solution but the mechanism of the reaction was unknown. Kinetic measurement of the S-nitrosothiol decomposition spectroscopically was erratic and irreproducible. The reaction was found to be mediated by adventitious Cu(^2+) in the water supplies used. The reaction of Cu(^2+) with S-nitrosothiols was studied. The rate equation was established and the reaction was found to be first order in S-nitrosothiol and the added Cu(^2+). Second order rate constants for the reaction were determined so that the effect of structure could be studied. S-Nitirosothiols containing no other functional groups did not react with Cu(^2+). S-Nitrosothiols containing a P-amino group were found to be the most reactive. The S-niti-osothiols were thought to be bidentately bound about the Cu(^2+). which leads to rate limiting S-N bond breaking producing the thiyl radical and nitiic oxide. A similar reaction was found to occur between S-nitrosothiols and Cu+ and Fe2+ ions, but no other transition metals appeared to catalyse nitiic oxide formation from S-nitrosothiols. The reaction of-S-nitrosothiols with other thiols was also Studied. The ti-ansnitrosation reaction was rapid and the variation of the observed rate constant with pH over the range 6-13 produced a sigmoidal curve which was indicative of a reaction involving attack by tiuolate anion. It is believed that this reaction could be important in vivo as a mechanism for storing NO in the form of a stable S-niti-osothiol and then transferring the NO group to a more reactive S-niti-osothiol for rapid NO release. The effect of the thiol group alone was also investigated and depending on the concentration of thiol used the reaction was either catalysed or inhibited. Other conditions such as changing pH, adding metal chelators and the oxygen dependency of the reaction were studied. Methods of measuring NO levels in solution were tested, the best being the use of a NO specific electi-ode. The reactions of NO with thiols were also studied to see if a mechanism for in vivo formation of S-nitrosothiols could be determined.