Determination of 8‐epi PGF2α concentrations as a biomarker of oxidative stress using triple‐stage liquid chromatography/tandem mass spectrometry |
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Authors: | Nasser E. Bastani Thomas E. Gundersen Rune Blomhoff |
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Affiliation: | 1. Institute of Basic Medical Sciences, University of Oslo, P.O. Box 1046 Blindern, N‐0316 Oslo, Norway;2. Vitas AS, Oslo Innovation Park, N‐0349 Oslo, Norway |
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Abstract: | F2‐isoprostanes are a family of prostaglandin F2‐like compounds that are formed by free‐radical‐catalyzed peroxidation of arachidonic acid. Several F2‐isoprostanes, but in particular 8‐epi PGF2α, are widely used as oxidative stress biomarkers. An analytical method based on liquid chromatography with negative electrospray ionization (ESI) coupled to tandem mass spectrometric detection (LC/MS/MS) was developed for the determination of 8‐epi PGF2α concentrations in human plasma, whole blood, erythrocytes and urine. 8‐epi PGF2α‐d4, a stable isotope derivative of 8‐epi PGF2α, was used as an internal standard (IS). A 50 µL sample was focused on‐column and separated on two 3 µm particle size SUPELCOSIL? ABZ+Plus HPLC columns (15 cm × 4.6 mm and 7.5 cm × 4.6 mm) connected in series. An Applied Biosystems 4000 Q TRAP LC/MS/MS system with ESI was operated in multiple reaction monitoring (MRM) mode with the precursor‐to‐product ion transitions m/z 353.4 → 193.1 (8‐epi PGF2α), 357.4 → 197.1 (8‐epi PGF2α‐d4), used for quantification. The assay was fully validated and found to have adequate accuracy, precision, linearity, sensitivity and selectivity. The mass limit of detection (mLOD) was 1 pg of analyte eluting from the column. The assay has been successfully applied to the analysis of human plasma, whole blood, erythrocytes and urine samples. Copyright © 2009 John Wiley & Sons, Ltd. |
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