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KMID : 0043320080310070878
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Archives of Pharmacal Research
2008 Volume.31 No. 7 p.878 ~ p.885
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HPLC Determination of Malondialdehyde in ECV304 Cell Culture Medium for Measuring the Antioxidant Effect of Vitexin-4"-O-glucoside
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Ying Xi-xiang
Li Hai-Bo
Chu Zheng-Yun
Zhai Yan-Jun
Liu Xun
Xin Chun
Zhang Wen-Jie
Kang Ting-Guo
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Abstract
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To investigate the antioxidant effect of vitexin-4"-O-glucoside, a flavone glycoside, isolated from the leaves of Crataegus pinnatifida Bge. var. major, we developed a simple and sensitive high-performance liquid chromatography (HPLC) method to determine levels of malondialdehyde (MDA) in ECV304 cell culture medium after induction by tert-butyl-hydroperoxide (TBHP). The preparation of analyzed samples involved a one-step derivatization with thiobarbituric acid (TBA). HPLC analysis was performed on a Hydro-RP, a polar end-capped column (, ), using an acetonitrile-ammonium acetate aqueous solution (10 mM, pH 6.8) as the mobile phase under linear gradient conditions with UV detection at 532 nm. The calibration curve was linear over MDA (r=0.9951). Relative standard deviations (RSDs) of intra-day and inter-day precision were less than 6.1% and 5.0%, respectively. The mean recovery was . The lower limit of quantification (LLOQ) of MDA was . This chromatographic method was successfully applied to investigating the in vitro antioxidant effect of vitexin-4"-O-glucoside. Vitexin-4"-O-glucoside (120 M) protected ECV304 cells from peroxidation induced by TBHP.
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KEYWORD
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High-performance liquid chromatography, Vitexin-4"-O-glucoside, ECV304 cells, Antioxidant effect, Malondialdehyde
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