Peroxynitrite does not decompose to singlet oxygen ( ¹ Δ _g O ₂ ) and nitroxyl (NO ⁻ )

Abstract

], peroxynitrite (ONOO ؊ ) decomposes after protonation to singlet oxygen ( 1 ⌬gO2) and singlet oxonitrate (nitroxyl, 1 NO ؊ ) in high yield. They claimed to have observed nitrosyl hemoglobin from the reaction of NO ؊ with methemoglobin; however, contamination with hydrogen peroxide gave rise to ferryl hemoglobin, the spectrum of which was mistakenly assigned to nitrosyl hemoglobin. We have carried out UV-visible and EPR experiments with methemoglobin and hydrogen peroxide-free peroxynitrite and find that no NO ؊ is formed. With this peroxynitrite preparation, no light emission from singlet oxygen at 1270 nm is observed, nor is singlet oxygen chemically trapped; however, singlet oxygen was trapped when hydrogen peroxide was also present, as previously described [Di Mascio, P., Bechara, E. )] is an inorganic oxidant of biological importance that is produced from superoxide and nitrogen monoxide (1). The mechanisms by which peroxynitrite or peroxynitrous acid [pK a ϭ 6.7 at 37°C (2)] oxidizes biomolecules are being investigated by various groups. There are two types or reaction: reactions between biomolecules and peroxynitrite (or peroxynitrous acid) that are first-order in peroxynitrite and in the biomolecule, and reactions that are first-order in peroxynitrite and zero-order in the biomolecule. In the latter case, either peroxynitrous acid undergoes homolysis to nitrogen dioxide and the hydroxyl radical (3-7), or it forms an activated intermediate (2,8). In the absence of oxidizable molecules the radicals recombine to nitrate, or the activated intermediate undergoes an internal isomerization. The rate constant for this process, which is first-order in peroxynitrite, is 3.9 s Ϫ1 at 37°C (2). Whether radicals are intermediates is not relevant for the present discussion. All involved agree that at acid pH and at low temperatures peroxynitrous acid isomerizes to nitrate quantitatively, whereas at higher pH nitrite and dioxygen in a 2 to 1 ratio also are found.Recently, a mechanism was proposed, in which peroxynitrite forms, after protonation, a cyclic intermediate, which dissociates to form oxonitrate(1Ϫ)-the commonly used name ''nitroxyl'' is outdated-and singlet oxygen (9):These products have been postulated before by Khan (10). As described in the more recent article (9), Khan et al. used methemoglobin (MetHb) to trap oxonitrate(1Ϫ), and 9,10-diphenylanthracene and 2,3-dimethyl-2-butene to trap singlet oxygen. Their mechanism does not explain the quantitative formation of nitrate, and we therefore investigated how these results (9) could have been obtained. We show here that singlet oxygen is not formed when peroxynitrite free of hydrogen peroxide is used, that the evidence for oxonitrate(1Ϫ) is based on a misinterpretation of the hemoglobin UV-visible spectra, and that the formation of these products and the postulated cyclic precursor, 4-hydrido-4-azy-1,2,3-trioxy[04]cycle, or trioxazetidine, is thermodynamically unlikely. The experimental evidence presented by Khan et al. (9) is best explained by the presence...

Arsenic-induced oxidative stress in the common bean legume, Phaseolus vulgaris L. seedlings and its amelioration by exogenous nitric oxide

“…Vertical bar represents means (n04) ± SE. Means followed by the same capital letter were not significantly different at P<0.05 for leaves; means followed by the same lower case letter were not significantly different at P<0.05 for roots enzymatic defense, NO itself has the capacity to detoxify ROS directly (Martinez et al 2000;Wendehenne et al 2001;Jin et al 2010). Therefore, it has the ability to prevent SODgenerated excess H 2 O 2 production in the SNP + As-treated P. vulgaris seedlings.…”

Cytochrome c-promoted cardiolipin oxidation generates singlet molecular oxygen

“…Tetramyristoyl (TMCL) and tetraoleoyl cardiolipin (TOCL) were purchased from Avanti Polar Lipids Inc. (Alabaster, AL). The disodium salt of anthracene-9,10-diyldiethane-2,1-diyl disulfate (EAS) and 3,3′-(1,4-epidioxynaphthalene)-1,4-diylbis-[N-(2,3-dihydroxypropyl)propanamide] (DHPNO 2 ) were synthesized as described by Di Mascio and Sies 31 and Martinez et al 32 respectively. Standard EAS endoperoxide (EASO 2 ) was prepared by photooxidation in D 2 O and purified by HPLC using the conditions described for EASO 2 analysis and quantified spectrophotometrically (ε 212 = 40 738 M −1 cm −1 ).…”

“…Standard EAS endoperoxide (EASO 2 ) was prepared by photooxidation in D 2 O and purified by HPLC using the conditions described for EASO 2 analysis and quantified spectrophotometrically (ε 212 = 40 738 M −1 cm −1 ). 32 Deuterated phosphate buffer at specified pD values ( pDvpH + 0.4) was prepared by mixing stock solutions of K 2 HPO 4 and KH 2 PO 4 prepared in D 2 O. All solvents were of HPLC grade and were acquired from Mallinckrodt Baker (Phillipsburg, NJ).…”

Differential responses of hydrogen peroxide, lipid peroxidation and antioxidant enzymes in Azolla microphylla exposed to paraquat and nitric oxide

“…These peroxynitrite are less toxic than peroxides, thus limit cellular damage (Hasanuzzaman et al 2010). These peroxynitrite might have reacted with H 2 O 2 to yield a nitrite anion and oxygen (Martinez et al 2000). This could be the mechanism of cytoprotective action of NO in Azolla fronds.…”