Journal
MOLECULAR AND CELLULAR BIOCHEMISTRY
Volume 234, Issue 1, Pages 379-385Publisher
SPRINGER
DOI: 10.1023/A:1015998732066
Keywords
hydroxyl radical; 3,4-DHBA; 4-HBA; microdialysis; HPLC; cerebral ischemia and reperfusion
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Funding
- NCRR NIH HHS [P20 RR15636] Funding Source: Medline
- NHLBI NIH HHS [R21 HL60326] Funding Source: Medline
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Aromatic hydroxylation from the reaction between hydroxyl radical and salicylate or its related compounds has been often utilized as a marker for the generation of hydroxyl radicals. We have investigated several technical aspects of applying this method to study hydroxyl radical production during cerebral ischemia and reperfusion using the hydroxylation of 4-hydroxybenzoic acid (4-HBA) to form 3,4-dihydroxybenzoic acid (3,4-DHBA). 4-HBA was administered to rats either through intravenous infusion, or by way of an in vivo microdialysis probe implanted in the brain. Dialysate containing 3,4-DHBA was collected and analyzed by HPLC with electrochemical detection. An endogenous compound was found to co-elute with 3,4 -DHBA but could be separated by varying the chromatographic conditions. Because of interrupted blood flow during cerebral ischemia and reperfusion, delivery of 4-HBA through the microdialysis probe is a preferred method to systemic administration such as intravenous infusion. It is concluded that the oxidation of 4-HBA to 3,4-DHBA can be a reliable and accurate indicator for the formation of hydroxyl radical in vivo if the experiments are well designed to avoid potential pitfalls associated with technical difficulties of the method.
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