Mukohda Masashi, Ueno Shunji, Kamibayashi Masato, Okada Muneyoshi, Yamawaki Hideyuki, Hara Yukio
School of Veterinary Medicine, Kitasato University, Aomori, Japan.
J Vet Med Sci. 2010 Dec;72(12):1547-50. doi: 10.1292/jvms.10-0232. Epub 2010 Jul 22.
Estimation of radical scavenging capacity of lipophilic antioxidants by electron spin resonance (ESR) in vitro is a challenging issue due to their poor solubility in aqueous radical generating and measuring systems. Water-miscible organic solvents are used for this purpose. A novel radical trapping agent, 5-(2,2-dimethyl-1,3-propoxy cyclophosphoryl)-5-methyl-1-pyrroline N-oxide (CYPMPO), that has practical advantages over well-known trapping agents was synthesized. However, no available data for the influence of solvents in an ESR system that uses CYPMPO has been presented. The influences of six water-miscible organic solvents, acetonitrile (AcN), acetone, dimethyl sulfoxide (DMSO), ethanol, polyethylene glycol (PEG), and dimethoxyethane (DME), on ESR responses to Fenton Fe(2+)/H (2)O(2 )OH· and hypoxanthine/xanthine oxidase superoxide generation systems in vitro were studied. Reduction of the ESR signal to CYPMPO-OH· adducts by 55.86 ± 5.95 and 83.17 ± 2.50% compared with the control was observed in the presence of AcN and acetone, respectively, at a final concentration of 5% (v/v). AcN of less than 1% had minimal effects. DMSO, ethanol, PEG and DME at 5% (v/v) strongly inhibited the ESR signals and/or caused derangement in the signal patterns. The six water-miscible solvents at 5% (v/v) had no influence on the ESR spectra of CYPMPO-superoxide adducts. From these results, AcN, at less than 1% (v/v), is a useful water-miscible organic solvent for assessing radical scavenging capacities of lipophilic compounds in the CYPMPO-Fenton Fe(2+)/H(2)O(2) OH· reaction system in an ESR assay. Any of the solvents used in the present study can be used in a hypoxanthine/xanthine oxidase superoxide generation system.
由于亲脂性抗氧化剂在水性自由基产生和测量系统中的溶解度较差,因此通过电子自旋共振(ESR)体外评估其自由基清除能力是一个具有挑战性的问题。为此使用了与水混溶的有机溶剂。合成了一种新型自由基捕获剂5-(2,2-二甲基-1,3-丙氧基环磷酰基)-5-甲基-1-吡咯啉N-氧化物(CYPMPO),它比知名捕获剂具有实际优势。然而,尚未有关于在使用CYPMPO的ESR系统中溶剂影响的可用数据。研究了六种与水混溶的有机溶剂,乙腈(AcN)、丙酮、二甲基亚砜(DMSO)、乙醇、聚乙二醇(PEG)和二甲氧基乙烷(DME),对体外ESR对芬顿Fe(2+)/H(2)O(2) OH·和次黄嘌呤/黄嘌呤氧化酶超氧化物生成系统反应的影响。在终浓度为5%(v/v)的情况下,分别在AcN和丙酮存在下,观察到CYPMPO-OH·加合物的ESR信号与对照相比降低了55.86±5.95%和83.17±2.50%。浓度低于1%的AcN影响最小。5%(v/v)的DMSO、乙醇、PEG和DME强烈抑制ESR信号和/或导致信号模式紊乱。5%(v/v)的六种与水混溶的溶剂对CYPMPO-超氧化物加合物的ESR光谱没有影响。根据这些结果,浓度低于1%(v/v)的AcN是一种有用的与水混溶的有机溶剂,可用于在ESR测定中评估CYPMPO-芬顿Fe(2+)/H(2)O(2) OH·反应系统中亲脂性化合物的自由基清除能力。本研究中使用的任何一种溶剂都可用于次黄嘌呤/黄嘌呤氧化酶超氧化物生成系统。