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        HPLC Determination of Malondialdehyde in ECV304 Cell Culture Medium for Measuring the Antioxidant Effect of Vitexin-4"-O-glucoside

        Ying, Xi-Xiang,Li, Hai-Bo,Chu, Zheng-Yun,Zhai, Yan-Jun,Leng, Ai-Jing,Liu, Xun,Xin, Chun,Zhang, Wen-Jie,Kang, Ting-Guo 대한약학회 2008 Archives of Pharmacal Research Vol.31 No.7

        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 $Synergi^{TM}$ Hydro-RP, a polar end-capped $C_{18}$ column ($250{\times}4.6\;mm$, $4\;{\mu}m$), 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 $0.0125-1.25\;{\mu}M$ 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 $96.9\;{\pm}\;1.6%$. The lower limit of quantification (LLOQ) of MDA was $0.0125\;{\mu}M$. 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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        HPLC Determination of Malondialdehyde in ECV304 Cell Culture Medium for Measuring the Antioxidant Effect of Vitexin-4"-O-glucoside

        Xi-xiang Ying,Hai-bo Li,Zheng-yun Chu,Yan-jun Zhai,Ai-jing Leng,Xun Liu,Wen-jie Zhang,Ting-guo Kang,Chun Xin 대한약학회 2008 Archives of Pharmacal Research Vol.31 No.7

        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 SynergiTM Hydro-RP, a polar end-capped C18 column (250×4.6 mm, 4 μm), 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 0.0125-1.25 μM 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 96.9 ± 1.6%. The lower limit of quantification (LLOQ) of MDA was 0.0125 μM. 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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