Comparison of two independent aromatic hydroxylation assays in combination with intracerebral microdialysis to determine hydroxyl free radicals

被引:33
|
作者
Themann, C [1 ]
Teismann, P [1 ]
Kuschinsky, K [1 ]
Ferger, B [1 ]
机构
[1] Univ Marburg, Fac Pharm, Inst Pharmacol & Toxicol, D-35032 Marburg, Germany
关键词
hydroxyl radicals; phenylalanine; salicylic acid; 6-hydroxydopamine; microdialysis; HPLC;
D O I
10.1016/S0165-0270(01)00370-3
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The phenylalanine- and salicylate assay were compared to investigate the production of hydroxyl free radicals. In vitro experiment: Phenylalanine (100 mu mol/1) or salicylic acid (100 mu mol/1) were incubated in a hydroxyl radical generating in vitro Fenton system with increasing concentrations (1.25-40 mu mol/1) of equimolar hydrogen peroxide and ferrous ions. Both, phenylalanine and salicylic acid were able to trap hydroxyl radicals in a reliable way indicated by the linear relationship between the concentration of the Fenton reagents and either the phenylalanine derived products (ortho-, meta-, para-tyrosine) or the salicylic acid-derived products (2,3- and 2,5-dihydroxybenzoic acid (DHBA)). In vivo experiment: Wistar rats were implanted with microdialysis probes and striatal perfusion with either 5 mmol/l phenylalanine or 5 mmol/l salicylic acid was performed. Addition of the dopaminergic neurotoxin 6-hydroxydopamine (100 mu mol/l, flow rate 2 mul/min, 60 min) to the perfusion fluid significantly increased the concentrations of ortho- and meta-tyrosine or 2,3-DHBA in comparison to control animals. All increases determined were rapidly reversible after changing back to pre-stimulation conditions. The results demonstrate that aromatic hydroxylation of phenylalanine or salicylic acid is a useful technique to investigate hydroxyl free radical formation in vitro and in vivo. Advantages and disadvantages of both methods are discussed. (C) 2001 Elsevier Science B.V. All rights reserved.
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页码:57 / 64
页数:8
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