A simple, rapid, highly sensitive and reproducible quantification method for plasma malondialdehyde by high‐performance liquid chromatography
1998; Wiley; Volume: 12; Issue: 5 Linguagem: Inglês
10.1002/(sici)1099-0801(199809/10)12
ISSN1099-0801
AutoresKenji Fukunaga, Munehiro Yoshida, Naoki Nakazono,
Tópico(s)Metabolomics and Mass Spectrometry Studies
ResumoA simple, rapid, highly sensitive and reproducible quantification method for plasma malondialdehyde (MDA) was developed using high-performance liquid chromatography (HPLC). The present method is based on the thiobarbituric acid (TBA) reaction and reversed-phase HPLC separation with fluorescence detection. For sample preparation, an aliquot of 5 μL plasma was mixed with 0.2% (w/v) TBA in 0.1 M sodium acetate buffer, pH 3.5. After heating at 95°C for 60 min, the reaction solution was centrifuged and an aliquot of 10 μL supernatant was injected into the HPLC system without neutralization or extraction procedures. The TBA–MDA adduct was separated on a reversed-phase column and quantified by a fluorescence detection (λ ex = 515 nm; λ em = 553 nm). The mobile phase was a mixture of acetonitrile:water (7:3, v/v) under isocratic conditions at ambient temperature. These procedures gave good results with regard to sensitivity (detection limit S/N = 3, 0.5 fmol per injection), linearity (0.5–500 fmol per injection), precision (between-assay C.V = 3.1%, within-assay C.V = 1.2%) and recovery (98.6%). The simple sample procedure, the highly sensitive detection limit and the stability of the TBA–MDA adduct make the present method applicable to numerous clinical samples. As many as 250–300 samples per day can be assayed using an autosampler. © 1998 John Wiley & Sons, Ltd.
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