Generation of peroxynitrite from reaction of N-acetyl-N-nitrosotryptophan with hydrogen peroxide over a wide range of pH values

Abstract

The novel reaction of N-acetyl-N-nitrosotryptophan (NANT) with hydrogen peroxide to yield peroxynitrite is demonstrated. Quantum chemical calculations performed at CBS-QB3 level of theory predicted that the reaction of N-nitrosoindole with both H₂O₂ and its corresponding anion is thermodynamically feasible. At pH 13, the formation of peroxynitrite from the bimolecular reaction of NANT with H₂O₂ is unequivocally demonstrated by ¹⁵N NMR spectrometry. In order to prove the intermediacy of peroxynitrite from the NANT–H₂O₂ system at neutral (7.4) and acidic pH (4.5), the characteristic pattern of CIDNP (chemically induced dynamic nuclear polarization) signals were recorded, i.e. enhanced absorption in the ¹⁵N NMR signal of nitrate and emission in the ¹⁵N NMR signal of nitrite. Most interestingly, the NANT–H₂O₂ system nitrated N-acetyltyrosine at pH 4 via recombination of freely diffusing nitrogen dioxide and tyrosyl radicals, but nitration was negligible at pH 7.4. Since the combination between NANT and H₂O₂ is slow, endogenous N-nitrosotryptophan residues cannot act as a “carrier” for peroxynitrite.