Abstract
Phosphoinositides derive from the phospholipid, phosphatidylinositol (PtdIns), by phosphorylation of the inositol ring in the lipid head group. The determination of phosphoinositide species is a particular challenge, because the structurally similar inositolphosphate-head groups must be analyzed as well as the lipid-associated fatty acids. The method presented in this chapter consists of two steps: First phosphoinositides are separated by thin-layer chromatography (TLC) according to their characteristic head groups and the individual lipids are isolated. Second, the fatty acids associated with each isolated lipid are analyzed using a gas-chromatograph (GC). The combination of these two classical methods for lipid analysis, TLC and GC, provides a cost-efficient and reliable alternative to lipidomics approaches requiring more extensive instrumentation.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Palmer S, Hawkins PT, Michell RH, Kirk CJ (1986) The labelling of polyphosphoinositides with [32P]Pi and the accumulation of inositol phosphates in vasopressin-stimulated hepatocytes. Biochem J 238:491–499
Heilmann I (2009) Using genetic tools to understand plant phosphoinositide signalling. Trends Plant Sci 14:171–179
König S, Ischebeck T, Lerche J, Stenzel I, Heilmann I (2008) Salt stress-induced association of phosphatidylinositol-4,5-bisphosphate with clathrin-coated vesicles in plants. Biochem J 415:387–399
König S, Mosblech A, Heilmann I (2007) Stress-inducible and constitutive phosphoinositide pools have distinct fatty acid patterns in Arabidopsis thaliana. FASEB J 21:1958–1967
Saavedra L, Balbi V, Lerche J, Mikami K, Heilmann I, Sommarin M (2011) PIPKs are essential for rhizoid elongation and caulonemal cell development in the moss Physcomitrella patens. Plant J 67:635–647
König S, Hoffmann M, Mosblech A, Heilmann I (2008) Determination of content and fatty acid composition of unlabeled phosphoinositide species by thin layer chromatography and gas chromatography. Anal Biochem 278:197–201
Goebbels S, Oltrogge JH, Kemper R, Heilmann I, Bormuth I, Wolfer S, Wichert SP, Mobius W, Liu X, Lappe-Siefke C, Rossner MJ, Groszer M, Suter U, Frahm J, Boretius S, Nave KA (2010) Elevated phosphatidylinositol 3,4,5-trisphosphate in glia triggers cell-autonomous membrane wrapping and myelination. J Neurosci 30:8953–8964
Walsh JP, Caldwell KK, Majerus PW (1991) Formation of phosphatidylinositol 3-phosphate by isomerization from phosphatidylinositol 4-phosphate. Proc Natl Acad Sci USA 88:9184–9187
Perera IY, Davis AJ, Galanopoulou D, Im YJ, Boss WF (2005) Characterization and comparative analysis of Arabidopsis phosphatidylinositol phosphate 5-kinase 10 reveals differences in Arabidopsis and human phosphatidylinositol phosphate kinases. FEBS Lett 579:3427–3432
Cho MH, Chen Q, Okpodu CM, Boss WF (1992) Separation and quantification of [3H]inositol phospholipids using thin-layer-chromatography and a computerized 3H imaging scanner. LC-GC 10:464–468
Hornung E, Korfei M, Pernstich C, Struss A, Kindl H, Fulda M, Feussner I (2005) Specific formation of arachidonic acid and eicosapentaenoic acid by a front-end Delta5-desaturase from Phytophthora megasperma. Biochim Biophys Acta 1686:181–189
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2013 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Heilmann, M., Heilmann, I. (2013). Mass Measurement of Polyphosphoinositides by Thin-Layer and Gas Chromatography. In: Munnik, T., Heilmann, I. (eds) Plant Lipid Signaling Protocols. Methods in Molecular Biology, vol 1009. Humana, Totowa, NJ. https://doi.org/10.1007/978-1-62703-401-2_3
Download citation
DOI: https://doi.org/10.1007/978-1-62703-401-2_3
Published:
Publisher Name: Humana, Totowa, NJ
Print ISBN: 978-1-62703-400-5
Online ISBN: 978-1-62703-401-2
eBook Packages: Springer Protocols