Isolation, chemical characterization, and subcellular distribution of 1-O-alkyl-2-acetyl-sn-glycerol in Tetrahymena pyriformis cells.
1-O-Hexadecyl-2-acetyl-sn-glycerol, the immediate precursor of platelet- activating factor (PAF) in its de novo formation, was detected in the protozoon Tetrahymena pyriformis. It was purified from the total lipid extract by TLC, after successive developments in two different solvent systems. Characterization was assessed by (a) gas-liquid chromatography with electron capture detection, and (b) gas chromatography combined with mass spectrometry in selected ion monitoring mode, after derivatization with heptafluorobutyric acid anhydride and tert-butyldimethylchlorosilane/imidazole, respectively. Its quantity was found to be 0.1 nmol/10(7) cells from the GC-MS, using authentic alkylacetylglycerol as external standard. Cell fractionation revealed that alkylacetylglycerol is located exclusively in the microsomal fraction of the protozoon. Previously, we have reported the occurrence of PAF in the microsomal fraction, as well as a dithiothreitol-insensitive CDP-choline: cholinephosphotransferase activity that utilizes exogenous alkylacetylglycerol as substrate in the mitochondrial and microsomal fractions. The above findings indicate that PAF can be formed in the cell by the de novo pathway.
Tellis
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Department of Chemistry, University of Ioannina, Greece.
Tsoukatos
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DC
Lekka
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ME
D000818:Animals; D002849:Chromatography, Gas / Q000379:methods; D005466:Fluorocarbons; D005995:Glyceryl Ethers / Q000032:analysis* / Q000302:isolation & purification; D007093:Imidazoles; D007202:Indicators and Reagents; D008861:Microsomes / Q000737:chemistry*; D010972:Platelet Activating Factor; D012821:Silanes; D013769:Tetrahymena pyriformis / Q000737:chemistry*
D005466:Fluorocarbons; D005995:Glyceryl Ethers; D007093:Imidazoles; D007202:Indicators and Reagents; D010972:Platelet Activating Factor; D012821:Silanes; C024004:t-butyldimethylchlorosilane; C007924:heptafluorobutyric anhydride; C029899:imidazole