Peroxynitrite (ONOO⁻) is a potent oxidant that mediates the oxidation of both nonprotein and protein sulfhydryls. It is formed in vivo from the reaction of superoxide (O₂⁻) and nitric oxide (NO), which are produced by endothelial cells, macrophages, and neutrophils. The reaction of peroxynitrite with free cysteine and the single thiol of albumin has apparent second-order rate constants of 5,900 M⁻¹·s⁻¹ and 2,600–2,800 M⁻¹·s⁻¹, respectively, at pH 7.4 and 37°C, which are 3 orders of magnitude higher than those for hydrogen peroxide (H₂O₂) with sulfhydryls. Unlike H₂O₂, which oxidizes thiolate anion, peroxynitrite reacts preferentially with the undissociated form of the thiol group. Peroxynitrite oxidizes cysteine to cystine and the thiol group of bovine serum albumin (BSA) to an arsenite nonreducible product, suggesting oxidation beyond sulfenic acid. Peroxynitrous acid is a less effective thiol-oxidizing agent than its anion, with oxidation presumably mediated by decomposition products such as hydroxyl radical and nitrogen dioxide. The reactive peroxynitrite anion may exert cytotoxic effects by oxidizing tissue sulfhydryls. Endothelial cells produce and release NO and superoxide, which can react to form peroxynitrite. Peroxynitrite has a pKa of 6.8 at 37°C and decomposes rapidly, forming hydroxyl radical-like species. Peroxynitrite is highly reactive and can oxidize deoxyribose and dimethyl sulfoxide at acidic pH. It also produces nitrogen dioxide, a strong oxidant with significant cytotoxic potential. The study investigated the reactions of peroxynitrite with protein and nonprotein sulfhydryl groups and compared them with H₂O₂-mediated thiol oxidation. Sulfhydryls are common targets for free radicals, and their oxidation is a key mechanism of free radical-mediated toxicity. Thiols are critical for enzyme activity and protein conformation. Peroxynitrite oxidizes sulfhydryls about 10³ times faster than H₂O₂ at 37°C and pH 7.4, suggesting it is an important mechanism of oxygen radical-mediated toxicity. The study found that peroxynitrite reacts with sulfhydryls more rapidly than H₂O₂, with rate constants of 2,600–2,800 M⁻¹·s⁻¹ for BSA and 5,900 M⁻¹·s⁻¹ for cysteinePeroxynitrite (ONOO⁻) is a potent oxidant that mediates the oxidation of both nonprotein and protein sulfhydryls. It is formed in vivo from the reaction of superoxide (O₂⁻) and nitric oxide (NO), which are produced by endothelial cells, macrophages, and neutrophils. The reaction of peroxynitrite with free cysteine and the single thiol of albumin has apparent second-order rate constants of 5,900 M⁻¹·s⁻¹ and 2,600–2,800 M⁻¹·s⁻¹, respectively, at pH 7.4 and 37°C, which are 3 orders of magnitude higher than those for hydrogen peroxide (H₂O₂) with sulfhydryls. Unlike H₂O₂, which oxidizes thiolate anion, peroxynitrite reacts preferentially with the undissociated form of the thiol group. Peroxynitrite oxidizes cysteine to cystine and the thiol group of bovine serum albumin (BSA) to an arsenite nonreducible product, suggesting oxidation beyond sulfenic acid. Peroxynitrous acid is a less effective thiol-oxidizing agent than its anion, with oxidation presumably mediated by decomposition products such as hydroxyl radical and nitrogen dioxide. The reactive peroxynitrite anion may exert cytotoxic effects by oxidizing tissue sulfhydryls. Endothelial cells produce and release NO and superoxide, which can react to form peroxynitrite. Peroxynitrite has a pKa of 6.8 at 37°C and decomposes rapidly, forming hydroxyl radical-like species. Peroxynitrite is highly reactive and can oxidize deoxyribose and dimethyl sulfoxide at acidic pH. It also produces nitrogen dioxide, a strong oxidant with significant cytotoxic potential. The study investigated the reactions of peroxynitrite with protein and nonprotein sulfhydryl groups and compared them with H₂O₂-mediated thiol oxidation. Sulfhydryls are common targets for free radicals, and their oxidation is a key mechanism of free radical-mediated toxicity. Thiols are critical for enzyme activity and protein conformation. Peroxynitrite oxidizes sulfhydryls about 10³ times faster than H₂O₂ at 37°C and pH 7.4, suggesting it is an important mechanism of oxygen radical-mediated toxicity. The study found that peroxynitrite reacts with sulfhydryls more rapidly than H₂O₂, with rate constants of 2,600–2,800 M⁻¹·s⁻¹ for BSA and 5,900 M⁻¹·s⁻¹ for cysteine