Direct measurement of oxy-radicals using HPLC-ECD
Liu, Y.; Zhang, J.T.
Yao Xue Xue Bao 28(6): 416-421
1993
ISSN/ISBN: 0513-4870 PMID: 8249598 Document Number: 418567
A direct, sensitive, simple and specific high-pressure liquid chromatographic (HPLC) method was used for the quantitation of hydroxyl radicals by means of oxy-radical trapping of DMPO to form DMPO-OH adducts. The DMPO-OH adduct peak was separated successfully and identified by HPLC-ECD with a Waters ODS reversed phase 10 mu-m column. The mobile phase composed of citric acid/sodium acetate (citric acid-30 mmol/L-sodium acetate 50 mmol/L-3% acetonitrile, pH 5.1), at a flow rate of 1.2 ml/min and detection potential of 0.6 V with Ag/AgCl as reference electrode. Both EDTA-Fe-2+-H-2O-2(FeSO-4300 mu-mol/L, EDTA 300 mu-mol/L, H-2O-2180 mu-mol/L and DMPO 2 mmol/L) and H-2O-2 photolysis (H-2O-218 mmol/L and DMPO 2 mmol/L photolysis for 6 min) systems were taken to produce hydroxyl free radicals for screening new drugs and studying the mechanism of action. The relative standard deviations were 6.1 and 8.0% respectively. The sensitivity of the method was shown to be similar to that of ESR. The method for detection of superoxide anions with HPLC-ECD was also described.