Measurement of hydroxyl radical in rat blood vessel by microbore liquid chromatography and electrochemical detection: an on-line microdialysis study.
Salicylic acid (0.5 mM) is used as a trapping reagent of hydroxyl radical, and the formed 2,3- and 2,5-dihydroxybenzoic acids were collected via an on-line microdialysis device from the blood vessels. This study revealed the use of a sensitive liquid chromatographic system with electrochemical detection for the determination of 2,3- and 2,5-dihydroxybenzoic acids. Mobile phase consisted of 0.1 M monochloroacetic acid, 10 mM EDTA, 0.5 mM sodium octylsulfate, 20% acetonitrile and 5% tetrahydrofuran in 1 l (pH 3.0 adjusted with 1 M NaOH), and the flow-rate of 0.05 ml/min were found to be optimum. Isocratic separation of these adducts on a microbore column (reversed-phase C18, 150x1 mm I.D., 5 microm) was achieved within 10 min. The optimal applied potential of dihydroxybenzoic acids was set at 750 mV based on a hydrodynamic study. This method has the detection limits of 1.3 pmol/ml (or 0.2 ng/ml) for 2,3- and 2,5-dihydroxybenzoic acids in Ringer solution (at signal-to-noise ratio=3).
Tsai
T H
TH
Department of Pharmacology, National Research Institute of Chinese Medicine, and National Yang-Ming University, Taipei, Taiwan. thtsai@cma23.nricm.edu.tw
Cheng
F C
FC
Hung
L C
LC
Chen
C F
CF
D000818:Animals; D001808:Blood Vessels / Q000737:chemistry*; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D004566:Electrodes; D005841:Gentisates / Q000032:analysis; D062385:Hydroxybenzoates / Q000032:analysis; D017665:Hydroxyl Radical / Q000032:analysis*; D007202:Indicators and Reagents; D007601:Jugular Veins / Q000737:chemistry; D008297:Male; D017551:Microdialysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D020156:Salicylic Acid; D012680:Sensitivity and Specificity
D005841:Gentisates; D062385:Hydroxybenzoates; D007202:Indicators and Reagents; D017665:Hydroxyl Radical; C009135:2,3-dihydroxybenzoic acid; D020156:Salicylic Acid