Li Linxiang, Abe Yoshihiro, Kanagawa Kiyotada, Shoji Tomoko, Mashino Tadahiko, Mochizuki Masataka, Tanaka Miho, Miyata Naoki
Kyoritsu University of Pharmacy, 1-5-30 Shibakoen, Minato, Tokyo 105-8512, Japan.
Anal Chim Acta. 2007 Sep 19;599(2):315-9. doi: 10.1016/j.aca.2007.08.008. Epub 2007 Aug 6.
Hydroxyl radical formation by Fenton reaction in the presence of an iron-chelating agent such as EDTA was traced by two different assay methods; an electron spin resonance (ESR) spin-trapping method with 5,5-dimethyl-1-pyrroline N-oxide (DMPO), and high Performance liquid chromatography (HPLC)-fluorescence detection with terephthalic acid (TPA), a fluorescent probe for hydroxyl radicals. From the ESR spin-trapping measurement, it was observed that EDTA seemed to suppress hydroxyl radical formation with the increase of its concentration. On the other hand, hydroxyl radical formation by Fenton reaction was not affected by EDTA monitored by HPLC assay. Similar inconsistent effects of other iron-chelating agents such as nitrylotriacetic acid (NTA), diethylenetriamine penta acetic acid (DTPA), oxalate and citrate were also observed. On the addition of EDTA solution to the reaction mixture 10 min after the Fenton reaction started, when hydroxyl radical formation should have almost ceased but the ESR signal of DMPO-OH radicals could be detected, it was observed that the DMPO-OH* signal disappeared rapidly. With the simultaneous addition of Fe(II) solution and EDTA after the Fenton reaction ceased, the DMPO-OH* signal disappeared more rapidly. The results indicated that these chelating agents should enhance the quenching of [DMPO-OH]* radicals by Fe(II), but they did not suppress Fenton reaction by forming chelates with iron ions.
在存在诸如乙二胺四乙酸（EDTA）等铁螯合剂的情况下，通过芬顿反应产生羟基自由基的过程，采用两种不同的测定方法进行追踪；一种是使用5,5 - 二甲基 - 1 - 吡咯啉N - 氧化物（DMPO）的电子自旋共振（ESR）自旋捕获法，另一种是使用对苯二甲酸（TPA，一种羟基自由基荧光探针）的高效液相色谱（HPLC） - 荧光检测法。从ESR自旋捕获测量结果来看，观察到随着EDTA浓度的增加，它似乎抑制了羟基自由基的形成。另一方面，通过HPLC测定监测发现，芬顿反应产生的羟基自由基不受EDTA的影响。对于其他铁螯合剂，如次氮基三乙酸（NTA）、二乙烯三胺五乙酸（DTPA）、草酸盐和柠檬酸盐，也观察到了类似的不一致效应。在芬顿反应开始10分钟后向反应混合物中加入EDTA溶液时，此时羟基自由基的形成应该几乎停止，但仍能检测到DMPO - OH自由基的ESR信号，结果观察到DMPO - OH信号迅速消失。在芬顿反应停止后同时加入Fe(II)溶液和EDTA时，DMPO - OH信号消失得更快。结果表明，这些螯合剂应该增强了Fe(II)对[DMPO - OH]*自由基的猝灭作用，但它们并没有通过与铁离子形成螯合物来抑制芬顿反应。
