LC Determination of Malondialdehyde Concentrations in the Human Umbilical Vein Endothelial Cell Culture Medium: Application to the Antioxidant Effect of Vitexin-2″-O-rhamnoside
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Vitexin-2″-O-rhamnoside (VOR) presents the leaves of Crataegus pinnatifida Bge. var. major which plays a role in preventing human pathologies related to oxidative stress, but as the principal component in the leaves, little attention has been devoted to its study of antioxidation of VOR. A simple, rapid and sensitive high-performance liquid chromatography method was developed to determine malondialdehyde (MDA) in ECV304 cell culture medium induced by tert-butyl-hydroperoxide (TBHP). The preparation of analyzed samples involved a one-step derivatization with thiobarbituric acid (TBA). The separation was achieved using the Synergi Hydro-RP, a polar endcapped C18 column (250 × 4.6 mm, 4 μm pore size) with a linear gradient elution of acetonitrile and 10 mmol L−1 of ammonium acetate aqueous solution (pH 6.8). The calibration curve was linear over the ranges 0.0125–1.25 μmol L−1 MDA (r = 0.9962). The lower limit of quantification of MDA was 0.0125 μmol L−1. Relative standard deviations of intra-day and inter-day precision were less than 5.2 and 4.4%. The method with high recovery (95.4 ± 1.3%) was successfully applied to the investigation of antioxidation of VOR by determining MDA in ECV304 cell culture medium.
KeywordsColumn liquid chromatography Human umbilical vein endothelial cell Antioxidation Malondialdehyde Vitexin-2″-O-rhamnoside
This work was supported by the funds of Natural Science Foundation of Liaoning Province (20072073) and Scientific Research Planning of the Educational Department of Liaoning Province (2004D264), China.
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