使用高效液相色谱-电子顺磁共振联用仪（HPLC-EPR）和高效液相色谱-电子顺磁共振-质谱联用仪（HPLC-EPR-MS）鉴定大鼠脑匀浆与亚铁离子/抗坏血酸体系反应混合物中形成的自由基。

Identification of a radical formed in the reaction mixture of rat brain homogenate with a ferrous ion/ascorbic acid system using HPLC-EPR and HPLC-EPR-MS.

作者信息

Kumamoto Kazumasa, Hirai Tomihiro, Kishioka Shiroh, Iwahashi Hideo

机构信息

Department of Pharmacology, Wakayama Medical University, Wakayama, Japan.

出版信息

Free Radic Res. 2007 Jun;41(6):650-4. doi: 10.1080/10715760701236758.

DOI:10.1080/10715760701236758

PMID:17516237

Abstract

Identification of a free radical is performed for the reaction mixture of rat brain homogenate with a ferrous ion/ascorbic acid system using EPR, high performance liquid chromatography-electron paramagnetic resonance spectrometry (HPLC-EPR) and high performance liquid chromatography-electron paramagnetic resonance-mass spectrometry (HPLC-EPR-MS). EPR measurements of the reaction mixtures showed prominent signals with hyperfine coupling constants (alpha(N) = 1.58 mT and alpha(H)beta = 0.26 mT). No EPR spectrum was detectable without rat brain homogenate, suggesting that the radical is derived from rat brain homogenate. An HPLC-EPR analysis of the reaction mixture showed a peak with retention time of 33.7 min. An HPLC-EPR-MS analysis of the peak gave two ions at m/z 224 and 137, suggesting that alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone (4-POBN)/ethyl radical adduct forms in the reaction mixture.

摘要

利用电子顺磁共振（EPR）、高效液相色谱 - 电子顺磁共振光谱法（HPLC - EPR）和高效液相色谱 - 电子顺磁共振 - 质谱法（HPLC - EPR - MS），对大鼠脑匀浆与亚铁离子/抗坏血酸体系的反应混合物进行自由基鉴定。反应混合物的EPR测量显示出具有超精细偶合常数（α(N) = 1.58 mT和α(H)β = 0.26 mT）的显著信号。没有大鼠脑匀浆时未检测到EPR光谱，这表明该自由基源自大鼠脑匀浆。反应混合物的HPLC - EPR分析显示出一个保留时间为33.7分钟的峰。对该峰的HPLC - EPR - MS分析给出了m/z 224和137处的两个离子，表明反应混合物中形成了α-(4 - 吡啶基 - 1 - 氧化物)-N - 叔丁基硝酮（4 - POBN）/乙基自由基加合物。