Lipidomic Analysis of Prostanoids by Liquid Chromatography–Electrospray Tandem Mass Spectrometry
Lipidomics aim to generate qualitative and quantitative information on different classes of lipids and their species, and when applied in conjunction with proteomic and genomic assays, facilitate the comprehensive study of lipid metabolism in cellular, organ, or body systems. Advances in mass spectrometry have underpinned the expansion of lipidomic methodologies. Prostanoids are potent autacoids present in a plethora of cellular systems, known best for their intimate role in inflammation. Electrospray ionisation (ESI) allows the efficient ionisation of prostanoids in aqueous systems. ESI can be readily coupled to liquid chromatography (LC) followed by tandem mass spectrometry (MS/MS)-based detection, thus allowing the development of a potent and selective LC/ESI–MS/MS quantitative assays. The protocol we describe in this chapter outlines the steps we follow to (a) extract prostanoids from solid or liquid samples, (b) semi-purify the metabolites using solid phase extraction (c) set-up the HPLC separation using reverse phase chromatography and (d) set-up the MS/MS assay using a triple quadrupole mass spectrometer. The experimental details and notes presented here are based on the detailed protocols followed in our group.
Key wordsProstaglandins Thromboxanes Dihydro-prostaglandins Electrospray ionisation Tandem mass spectrometry HPLC Lipidomics
This project has been supported in part by the Wellcome Trust Grant WT077714. We acknowledge the excellent technical support provided by Andrew Healey, Analytical Centre, University of Bradford.
- 1.Nicolaou, A. (2004) Prostanoids, Bioactive Lipids. Nicolaou, A., Kokotos, G. editors, 197–222.Google Scholar
- 12.Kempen, E. C., Yang, P., Felix, E., Madden, T. & Newman, R. A. (2001) Simultaneous quantification of arachidonic acid metabolites in cultured tumor cells using high-performance liquid chromatography/electrospray ionization tandem mass spectrometry, Anal Biochem. 297, 183–90.PubMedCrossRefGoogle Scholar
- 14.Yue, H., Strauss, K. I., Borenstein, M. R., Barbe, M. F., Rossi, L. J. & Jansen, S. A. (2004) Determination of bioactive eicosanoids in brain tissue by a sensitive reversed-phase liquid chromatographic method with fluorescence detection, J Chromatogr B Analyt Technol Biomed Life Sci. 803, 267–77.PubMedCrossRefGoogle Scholar