S-nitroglutathione, a product of the reaction between peroxynitrite and glutathione that generates nitric oxide

Peroxynitrite (ONOO-) has been shown in studies on vascular relaxation and guanylate cyclase activation to react with glutathione (GSH), generating an intermediate product that promotes a time-dependent production of nitric oxide (NO), In this study, reactions of ONOO- with GSH produced a new substance, which was characterized by liquid chromatography, ultraviolet spectroscopy, and electrospray tandem mass spectrometry, The mass spectrometric data provided evidence that the product of this reaction was S-nitroglutathione (GSNO(2)) and that S-nitrosoglutathione (GSNO) was not a detectable product of this reaction. Further evidence was obtained by comparison of the spectral and chromatographic properties with synthetic standards prepared by reaction of GSH with nitrosonium or nitronium borofluorates, Both the synthetic and ONOO-/GSH-derived GSNO(2) generated a protonated ion, GSNO(2)H(+) at mit 353, which was unusually resistant to decomposition under collision activation, and no fragmentation was observed at collision energy of 25 eV, In contrast, an ion at mit 337 (GSNOH(+)), generated from the synthetic GSNO, readily fragmented with the abundant loss of NO at 9 eV, Reactions of ONOO- with GSH resulted in the generation of NO, which was detected by the head space/NO-chemiluminescence analyzer method, The generation of NO was inhibited by the presence of glucose and/or CO2 in the buffers employed. Synthetic GSNO(2) spontaneously generated NO in a manner that was not significantly altered by glucose or CO2. Thus, ONOO- reacts with GSH to form GSNO(2), and GSNO(2) decomposes in a manner that generates NO.