Rapid sample preparation and simultaneous quantitation of prostaglandins and lipoxygenase derived fatty acid metabolites by liquid chromatography-mass spectrometry from small sample volumes

Background: Fatty acid metabolites play a key role in numerous physiological and pathological processes. A rapid liquid chromatography-mass spectrometry assay for the simultaneous determination of prostanoids, isoprostane and lipoxygenase (LOX) derived fatty acid metabolites in a small biological sample of only 20 mu L was developed. Methods: Human plasma samples were applied to a filter spot, extracted without prior derivatization and analyzed within 13 min. Detection of metabolites was performed on a triple quadrupole mass spectrometer in negative multiple-reaction monitoring detection mode. Application of this assay to various biological matrices was performed. Results: The validated assay was linear over the concentration range of 5-500 nmol/ L for prostanoids and isoprostane, 50-5000 nmol/ L for LOX-derived metabolites and 400-40,000 nmol/ L for fatty acids. Limits of quantitation were 0.4-233 nmol/ L, depending on the metabolite. Plasma samples from diabetic patients and controls showed significant increases in (+/-)9-HODE and 15(S)-HETE with p-values of 0.019 and 0.024, respectively. Conclusions: The small amount of 20 mL sample volume used in this assay and the demonstrated application to various sample types makes it an ideal routine analysis method for fatty acid metabolites. The resulting values for LOX-derived metabolites in diabetes mellitus type 2 samples support earlier findings about the role of lipid oxidation products in diabetes.