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A simple and precise method for the routine determination of platelet-activating factor in blood and urine

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Lipids

Abstract

A simple and precise method is described for the measurement of platelet-activating factor (PAF) in blood and urine. The method involves the isolation of PAF from blood samples by two successive steps. In the first step, blood proteins are precipitated with ethanol and the “free” PAF, i.e. the PAF which is extractable with ethanol, is recovered. In the second step, “bound” PAF, i.e., PAF not extractable with ethanol, is extracted from the protein precipitate with chloroform/methanol/water. The extraction of PAF from urine samples requires only the ethanol extraction step. “Free” and “bound” PAF are then each fractionated by silicic acid column chromatography, and the methanol/water eluent containing PAF is then further fractionated by high-performance liquid chromatography using an isocratic solvent system of acetonitrile/methanol/water. PAF is then quantitated by measuring its ability to induce platelet aggregation in an aggregometer. Application of the method to blood and urine samples from twenty-three healthy volunteers revealed PAF levels in blood of 140–480 pg/mL (630–254.4 pg “free” PAF/mL and 64–225.6 pg “bound” PAF/mL), and of 1.2–4.0 pg PAF/mL in urine. The method overcomes various technical problems and was shown to be very precise. It should prove useful for monitoring PAF levels in various disease conditions.

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Abbreviations

BSA:

bovine serum albumin

HPLC:

high-performance liquid chromatography

LPC:

lysophosphatidylcholine

LPE:

lysophosphatidylethanolamine

PAF:

platelet-activating factor, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine

PC:

phosphatidylcholine

PE:

phosphatidylethanolamine

SM:

sphingomyelin

TLC:

thin-layer chromatography

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Authors and Affiliations

  1. Department of Chemistry, Panepistimioupolis, University of Athens, GR-157 71, Athens, Greece

    Constantinos A. Demopoulos & Smaragdi Antonopoulou

  2. Department of Dietetics, Harokopio University of Home Economics, GR-176 71, Athens, Greece

    Nikolaos K. Andrikopoulos

Authors
  1. Constantinos A. Demopoulos
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  2. Nikolaos K. Andrikopoulos
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  3. Smaragdi Antonopoulou
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Demopoulos, C.A., Andrikopoulos, N.K. & Antonopoulou, S. A simple and precise method for the routine determination of platelet-activating factor in blood and urine. Lipids 29, 305–309 (1994). https://doi.org/10.1007/BF02536336

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  • DOI: https://doi.org/10.1007/BF02536336

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