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New Approaches to Lipid Analyses of Lipoproteins and Cell Membranes

  • Chapter
Membrane Fluidity

Part of the book series: Experimental Biology and Medicine ((EBAM,volume 1))

Abstract

Conventional and capillary gas chromatography-mass spectrometry (GC/MS) was used to determine the molecular association and positional distribution of fatty acids in diacylglycerols and of molecular association of nitrogenous bases and fatty acids in ceramides following their conversion to the tert-butyl dimethyl silyl (t-BDMS) ethers. The diacylglycerols and ceramides were generated from the corresponding glycerophospholipids and sphingomyelins by hydrolysis with phospholipase C of the original lipoproteins and cell membranes or appropriate total lipid extracts thereof. The abundant M-57 ion provided the molecular weights for both diacylglycerols and ceramides. The abundance ratio of the ions due to losses of the acyloxy radical (M-RCOO) from position 1 and from position 2 indicated the proportion of the reverse isomers of the saturated diacylglycerols. The ratio of the reverse isomers in unsaturated diacylglycerols could be similarly determined after reducing the sample with hydrogen (unsaturated species having higher carbon numbers) or with deuterium in the presence of Wilkinson’s catalyst. The alkylacyl and alkenylacyl glycerols were similarly identified by abundant ions at m/e values corresponding to [M-57], [acyl + 74], [RO + 114] and [R-CH=CHO + 114]. The long chain bases in the ceramides were identified via the intense ions CH3(CH2)n-(CH=CH)m-CH2O-t-BDMS and the fatty acids via the intense ions CH3(CH2)n-(CH=CH)m-CH2O-t-BDMS and the fatty acids via the intense ions CH3(CH2)n-(Ch=CH)m-CONHCH-Ch2O-t-BDMS. These methods are suitable for work with stable isotope-labelled phospholipids provided the isotopes are present in sufficient excess at least in one of the neutral lipid moieties of the molecular species. The assays usually do not require silver nitrate prefractionation of the sample and can be executed in the microgram range of material.

The above methods have been applied in analyses of molecular species of phosphatidylcholines and sphingomyelins of high density lipoproteins (HDL) of human plasma, and of the free diacylglycerol fraction obtained from an isolated rat liver during perfusion with deuterium oxide.

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Authors and Affiliations

  1. Banting and Best Dept. of Medical Research, University of Toronto, Toronto, Ontario, M5G 1L6, Canada

    A. Kuksis & J. J. Myher

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  1. A. Kuksis
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Editors and Affiliations

  1. University of Ottawa, Canada

    Morris Kates

  2. University of Toronto, Canada

    Arnis Kuksis

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Kuksis, A., Myher, J.J. (1980). New Approaches to Lipid Analyses of Lipoproteins and Cell Membranes. In: Kates, M., Kuksis, A. (eds) Membrane Fluidity. Experimental Biology and Medicine, vol 1. Humana Press. https://doi.org/10.1007/978-1-4612-6120-9_1

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  • DOI: https://doi.org/10.1007/978-1-4612-6120-9_1

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