de Avellar Isa G J, Magalhães Marta M M, Silva André B, Souza Luciana L, Leitão Alvaro C, Hermes-Lima Marcelo
Oxyradical Research Group, Departamento de Biologia Celular, Universidade de Brasília, Brasília, DF 70910-900, Brazil.
Biochim Biophys Acta. 2004 Nov 18;1675(1-3):46-53. doi: 10.1016/j.bbagen.2004.08.006.
It is widely believed that the iron chelator 1,10-phenanthroline (phen) is able to fully block the Fenton reaction by forming a complex (Fe(phen)3(2+), also known as ferroin) that cannot react with H2O2. We observed that phen cannot fully prevent 2-deoxyribose (5 mM) degradation induced by Fenton reagents (30 microM Fe(II) plus 100-500 microM H2O2); protection varied from 55% to 66% when the phen/Fe(II) ratio was 3:1 to 20:1. Inhibition of 2-deoxyribose damage was nearly unchanged if phen was pre-incubated with Fe(II). Moreover, preformed Fe(phen)3(2+) complex added to the solution containing H2O2 was able to induce 2-deoxyribose degradation and methane sulfinic acid formation from the oxidation of 5% DMSO. The partially protective effect of phen was unchanged with the use of either phosphate or HEPES as buffers (5 mM, pH 7.2), or in unbuffered media (pH 5.1). Both DMSO oxidation and 2-deoxyribose degradation correlated with the increase in Fe(phen)3(2+) concentration. Strand breaks in plasmid pTARGETtrade mark DNA induced by Fenton reagents (1 microM Fe(II) plus 25 microM H2O2) in HEPES buffer could only be partially prevented by phen, even when the chelator was 16 times more concentrated than Fe(II). In these experiments, Fe(phen)3(2+) and DNA were pre-incubated from 1 to 10 min before addition of H2O2. Moreover, a high level of DNA strand breakage was observed when iron and phen are added to the reaction immediately before H2O2. On the other hand, phen fully prevented 2-deoxyribose degradation induced by the autoxidation of 30 microM Fe(II) in phosphate-buffered (3 to 30 mM) media. Our data provide evidence that the Fe(phen)3(2+) complex induces in vitro oxidative damage in the presence of H2O2 (possibly by means of Fe(phen)3(2+) dissociation into Fe(phen)2(2+)), but they show that the complex cannot undergo autoxidation.
人们普遍认为，铁螯合剂1,10 - 菲咯啉（phen）能够通过形成一种不能与过氧化氢反应的络合物（Fe(phen)3(2+)，也称为亚铁菲咯啉）来完全阻断芬顿反应。我们观察到，phen不能完全阻止芬顿试剂（30 microM Fe(II)加100 - 500 microM过氧化氢）诱导的2 - 脱氧核糖（5 mM）降解；当phen/Fe(II)比例为3:1至20:1时，保护率在55%至66%之间变化。如果phen与Fe(II)预孵育，对2 - 脱氧核糖损伤的抑制作用几乎不变。此外，将预先形成的Fe(phen)3(2+)络合物添加到含有过氧化氢的溶液中，能够诱导2 - 脱氧核糖降解以及由5%二甲基亚砜氧化形成甲磺酸。使用磷酸盐或HEPES作为缓冲液（5 mM，pH 7.2），或在无缓冲介质（pH 5.1）中，phen的部分保护作用不变。二甲基亚砜氧化和2 - 脱氧核糖降解都与Fe(phen)3(2+)浓度的增加相关。在HEPES缓冲液中，芬顿试剂（1 microM Fe(II)加25 microM过氧化氢）诱导的质粒pTARGETtrade mark DNA链断裂只能被phen部分阻止，即使螯合剂的浓度比Fe(II)高16倍。在这些实验中，在添加过氧化氢之前，Fe(phen)3(2+)和DNA预先孵育1至10分钟。此外，当在过氧化氢加入前立即将铁和phen加入反应中时，观察到高水平的DNA链断裂。另一方面，phen完全阻止了在磷酸盐缓冲（3至30 mM）介质中30 microM Fe(II)自氧化诱导的2 - 脱氧核糖降解。我们的数据提供了证据表明，Fe(phen)3(2+)络合物在过氧化氢存在下诱导体外氧化损伤（可能是通过Fe(phen)3(2+)解离为Fe(phen)2(2+)），但表明该络合物不会发生自氧化。
