Combining EPR and ESI-MS/MS to study the reactivity of alkylthiols and cysteine towards 2-methyl-2-nitrosopropane (MNP)

Abstract

An approach combining EPR spectroscopy and electrospray ionisation mass spectrometry was employed to study the reactivity of two alkylthiols and of cysteine towards 2-methyl-2-nitrosopropane (MNP) in the presence of H₂O₂. Depending on the experimental conditions, various nitroxides could be EPR-detected, while their diamagnetic derivatives were structurally characterised after tandem mass spectrometry experiments. Sample dilution in methanol before electrospray ionisation was also found to generate methyl-hydroxylamine derivatives and could thus constitute a simple derivatisation process to stabilise hardly MS-detectable radical species. Upon white light irradiation, the three thiols tested led to alkoxynitroxides according to an inverted spin trapping process. On the other hand, only the reduced form of cysteine was found to react with MNP in the dark, yielding an alkylthionitroxide adduct via a Forrester–Hepburn mechanism. Besides a better understanding of the reactivity of thiols towards nitroso compounds, the results obtained underlined the particular behaviour of cysteine, suggesting that the use of a molecule as simple as MNP could allow a nitroxide moiety to be linked to a cysteinyl residue.