Peroxynitrite, hypochlorous acid

Fig. 7.3 Reactions showing the generation of ROS during lipid peroxidation and oxidative stress. Hydroxyl radical ( OH) lipid radical ( lipid), peroxyl radical (lipid-OO ) lipid peroxide (lipid-OOH) nitric oxide ( NO) nitrogen dioxide (N02) peroxynitrite anion (ONOO-) hypochlorous acid (HOC1), and hydrogen peroxide (H202)...

Pryor and coworkers have shown that peroxynitrite-mediated nitrosations and nitrations of phenols are modulated by CO2. The reaction was found to be first order with respect to peroxynitrite and zero order with respect to phenol, showing that an activated intermediate of peroxynitrite, perhaps the peroxynitrite anion-C02 adduct (0=N—OO—C02 ), is involved as the intermediate (equation 57) . At pH higher than 8.0, 4-nitrosophenol is the major product, whereas in acidic media significant amounts of the 2- and 4-nitrophenols were formed. Peroxynitrite also induces biological nitration of tyrosine residues of the proteins. The detection of 3-nitrotyrosine is routinely used as an in vivo marker for the production of the cytotoxic species peroxynitrite (ONOO ). It was shown that nitrite anion (N02 ) formed in situ by the reaction of nitric oxide and hypochlorous acid (HOCl) is similarly able to nitrate phenolic substrates such as tyrosine and 4-hydroxyphenylacetic acid . 

Free superoxide (O ) Perhydroxyl (HOi") Hydroxyl (-OH) Alkoxyl (LO-) Peroxyl (LOO-) Nitric oxide (-NO) Nitrogen dioxide (-NOi) Singlet oxygen (A O2) Hydrogen peroxide (H2O2) Hypochlorous acid (HOCl) Nitrous acid (HNO2) Peroxynitrite (ONOO ) Alkyl peroxynitrite (LOONO)... 

Peroxynitrite may be an important oxidant produced in acetaminophen-induced MPT. As discussed above, acetaminophen-induced MPT occurred with increased oxidation of the redox-sensitive dye DCFH2. This dye is readily oxidized by peroxynitrite but not by superoxide, hydrogen peroxide, or hypochlorous acid however, it may be oxidized by peroxide plus a peroxidase or a Fenton mechanism (ferrous ions plus peroxide) (Crow 1997 Myhre et al. 2003). Peroxynitrite is known to rapidly react with thiols such as N-acetylcyteine (Crow 2000), and N-acetylcysteine prevented acetaminophen-induced MPT and DCFH2 oxidation (Reid et al. 2005). The finding that nitration was predominantly in mitochondria of acetaminophen-treated mice supports the hypothesis that peroxynitrite formation occurred in that organelle (Cover et al. 2005). As pointed out above, the NOS isoform was probably not iNOS, which suggests involvement of another NOS species such as mitochondrial nitric oxide synthase (mtNOS) (Ghafourifar and Cadenas 2005). 

Fig. 3. Production of reactive species. (A) ROS can be produced from the weak radical oxygen in the mitochondria and endoplasmic reticulum, by various enzymatic reactions, and from oxyhemoglobin. Normally, nontoxic hydrogen peroxide can give rise to the powerful hydroxyl radical in the presence of transition metals (R5). Oxygen can also be induced to react with biomolecules by transition metals and enzymes. RNS can be produced by reaction of superoxide anion radical with the weak radical nitric oxide. These can react to form the powerful oxidant peroxynitrite/peroxynitrous acid, which can cause formation of other radicals, some with longer lives. See the text for details. SOD, superoxide dismutase. (B) Myeloperoxidase in leukocytes can produce the reactive species hypochlorous acid and tyrosyl radical. Unpaired electrons are indicated by the dense dots and paired electrons by the light ones.

Nitroxanthine is produced is produced as the major nitration product in reactions of 2 -deoxy-guanosine or ca//thymus DNA with nitryl chloride produced by mixing nitrite with hypochlorous acid, and 8-nitroguanidine was a minor product in these reactions (Chen et al. 2001). Formation of 8-nitroxanthine was also detected by xanthine reaction with various reactive nitrogen species, including nitryl chloride, peroxynitrite, nitronium tetra-fluoroborate, and heated nitric and nitrous acids. 

Substitution reactions may serve an antioxidant function. Halogenation and nitration of isoflavones is discussed later in this volume. These reactions imply that the proinflamitory oxidants (hypobromous acid, hypochlorous acid, and peroxynitrite) might be reduced or moderated by reaction with isoflavones in vivo. 

Oxygen is extremely important when discussing free radical reactions, and it is central to substances collectively referred to as reactive oxygen species (ROS) , which include both radical and non-radical substances. Radical ROS include hydroxyl, superoxide, peroxy, and alkoxy radicals. Non-radical ROS include hydrogen peroxide, hypochlorous acid, singlet oxygen, peroxynitrite, and ozone (7). 

Isoflavones have been shown to be beneficial in several chronic diseases in which oxidants are involved. Isoflavones are weak antioxidants when tested in vitro in die context of scavenging lipid peroxyl radicals. However, at sites of mflammation proinflammatory oxidants such as peroxynitrite (ONOO"), hypochlorous acid (HOCl), and hypobromous acid (HOBr) are formed that may also react widi isoflavones. We present evidence herein, using reverse-phase HPLC-mass spectrometry and proton NMR, that at concmitrations of diese oxidants formed at these local sites, nitration, chlorination, and bromination of isoflavones occurs. 

Free radicals produced in vivo include superoxide, the hydroxyl radical, nitric oxide, oxygen-centered organic radicals such as peroxyl and alkoxyl radicals, and sulfur-centered thiyl radicals. Other oxygen-containing reactive species that are not radicals are also formed. These include hydrogen peroxide, peroxynitrite, and hypochlorous acid. While these are not radical species, they are actually or potentially damaging oxidants. The collective term ROS is often used to describe both radical and nonradical species. 

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