Title | Benefits of prolonged gradient separation for high-performance liquid chromatography-tandem mass spectrometry quantitation of plasma total 15-series F-isoprostanes. |
Publication Type | Journal Article |
Year of Publication | 2006 |
Authors | Taylor AW, Bruno RS, Frei B, Traber MG |
Journal | Anal Biochem |
Volume | 350 |
Issue | 1 |
Pagination | 41-51 |
Date Published | 2006 Mar 01 |
ISSN | 0003-2697 |
Keywords | Adolescent, Adult, Arachidonic Acid, Ascorbic Acid, Chromatography, High Pressure Liquid, Dinoprost, Enzyme-Linked Immunosorbent Assay, F2-Isoprostanes, Female, Gas Chromatography-Mass Spectrometry, Humans, Mass Spectrometry, Reproducibility of Results, Tobacco Smoke Pollution |
Abstract | The F(2)-isoprostanes are products of free-radical-induced oxidation of arachidonic acid (AA) that are stereoisomers of prostaglandin F(2alpha) (PGF(2alpha)). We describe a method for quantitation of several 15-series PGF isomers (15-PGFs) and AA by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS-MS). Plasma samples were subjected to alkaline hydrolysis and acidified, and total (free + esterified) 15-PGFs and AA were extracted with organic solvents. The analytes were separated by gradient reverse-phase HPLC and detected by multiple reaction monitoring on a triple-quadrupole mass spectrometer, using deuterated internal standards for quantitation. The assay had a linear range of 1-40 pg of 8-iso-PGF(2alpha) on column and can quantify as little as 40 pg/mL (0.11 nM) in plasma. Outcomes significantly correlated (p < 0.0001) with data obtained by gas chromatography-mass spectrometry GC-MS or enzyme-linked immunosorbent assay. All plasma 15-PGF isomers increased over time with in vitro cigarette smoke exposure and correlated (p < 0.0001) with each other. The same strong inter-15-PGF correlations were observed in plasma from healthy young adult subjects. The coefficients of variation of HPLC-MS-MS measurements (24-32%) were smaller than those obtained by GC-MS (53%). Thus, HPLC-MS-MS potentially offers greater precision and allows quantitation of more compounds with simpler sample preparation than existing methods. Ours is the first validated quantitative assay using HPLC-tandem MS applied to plasma total 15-PGFs. |
DOI | 10.1016/j.ab.2005.12.003 |
Alternate Journal | Anal. Biochem. |
PubMed ID | 16448621 |
Grant List | AG05144 / AG / NIA NIH HHS / United States AG16835 / AG / NIA NIH HHS / United States DK59576 / DK / NIDDK NIH HHS / United States |