通过电子顺磁共振（EPR）和自旋捕集技术研究活性氧和氮物种的反应。

Reactions of active oxygen and nitrogen species studied by EPR and spin trapping.

作者信息

Carmichael A J, Steel-Goodwin L, Gray B, Arroyo C M

机构信息

Radiation Biophysics Department, Armed Forces Radiobiology Research Institute, Bethesda, Maryland 20889-5603.

出版信息

Free Radic Res Commun. 1993;19 Suppl 1:S1-16. doi: 10.3109/10715769309056s1.

DOI:10.3109/10715769309056s1

PMID:8282210

Abstract

The reactions of hydrogen peroxide (H2O2) with nitrite (NO2-) and of superoxide (O2-.) with nitric oxide (NO.) were studied using EPR and spin trapping techniques. These reactions reportedly have a common peroxynitrite (OONO-) intermediate. It has been suggested that this intermediate when protonated rapidly decomposes producing hydroxyl radicals (.OH) and the nitrogen dioxide radical (NO2.). The production of .OH in the reaction between H2O2 and NO2- was confirmed in spin trapping experiments using the spin trap 5,5-dimethyl-1-pyrroline-1-oxide (DMPO). H2O2 and NO2- were mixed at neutral pH and then the pH was decreased to pH 3-3.5 in the presence of DMPO or DMPO and ethanol. In these experiments, the EPR spectrum of the DMPO-OH adduct was obtained in addition to a weak EPR spectrum consisting of a triplet of triplets (a N = 1.415 mT and a N beta = 0.35 mT) indicating the addition of a nitrogen centered radical to DMPO. The formation of .OH was confirmed using ethanol as an .OH scavenger. The DMPO-hydroxyethyl adduct was produced from the reaction of .OH with ethanol. However, in experiments using an excess of ethanol, the formation of DMPO-OH was not prevented. This suggests that the DMPO-OH formed in the decomposition of HOONO does not entirely originate from a direct addition of .OH to DMPO. The reaction of O2-. with NO. was carried out in deaerated and air-saturated solutions at pH 12.3 where the dismutation of O2-. is minimal. The pH was then decreased to pH 3-3.5 in the presence of DMPO or DMPO and ethanol. In these experiments, the most prominent EPR spectrum obtained was a triplet of triplets (aN = 1.415 mT and a beta N = 0.35 mT) suggesting the addition of a nitrogen centered radical to DMPO. The formation of DMPO-OH was minimal and there was no formation of DMPO-hydroxyethyl adducts in the presence of ethanol. The results suggest that NO. in solution yields additional reactive species which act as nitrating agents in the presence of DMPO.

摘要

利用电子顺磁共振（EPR）和自旋捕集技术研究了过氧化氢（H₂O₂）与亚硝酸盐（NO₂⁻）以及超氧阴离子（O₂⁻·）与一氧化氮（NO·）的反应。据报道，这些反应有一个共同的过氧亚硝酸根（OONO⁻）中间体。有人提出，该中间体质子化后会迅速分解，产生羟基自由基（·OH）和二氧化氮自由基（NO₂·）。在使用自旋捕集剂5,5 - 二甲基 - 1 - 吡咯啉 - 1 - 氧化物（DMPO）的自旋捕集实验中，证实了H₂O₂与NO₂⁻反应中·OH的产生。在中性pH下将H₂O₂和NO₂⁻混合，然后在DMPO或DMPO与乙醇存在的情况下将pH降至3 - 3.5。在这些实验中，除了由三重态的三重态组成的弱EPR谱（aN = 1.415 mT和aNβ = 0.35 mT）表明有一个以氮为中心的自由基加成到DMPO上之外，还获得了DMPO - OH加合物的EPR谱。使用乙醇作为·OH清除剂证实了·OH的形成。·OH与乙醇反应生成了DMPO - 羟乙基加合物。然而，在使用过量乙醇的实验中，并未阻止DMPO - OH的形成。这表明HOONO分解过程中形成的DMPO - OH并非完全源于·OH直接加成到DMPO上。O₂⁻·与NO·的反应在pH 12.3的除氧和空气饱和溶液中进行，此时O₂⁻·的歧化作用最小。然后在DMPO或DMPO与乙醇存在的情况下将pH降至3 - 3.5。在这些实验中，获得的最显著的EPR谱是三重态的三重态（aN = 1.415 mT和aβN = 0.35 mT），表明有一个以氮为中心的自由基加成到DMPO上。DMPO - OH的形成极少，并且在乙醇存在的情况下没有形成DMPO - 羟乙基加合物。结果表明，溶液中的NO·会产生额外的活性物种，这些活性物种在DMPO存在时充当硝化剂。