Preparative separation of sphingolipids and of individual molecular species by high-performance liquid chromatography and their identification by gas chromatography-mass spectrometry.
Six fractions containing tri- to pentaglycosylceramides were isolated from the green, fresh water alga Chlorella kessleri, grown heterotrophically, by using preparative high-performance liquid chromatography (HPLC). Up to twelve fractions were obtained by further reversed-phase HPLC of each glycosylceramide. The use of a polar capillary column with Supelcowax 10 as the stationary phase allowed an excellent separation of the individual molecular species of ceramides, even though the separation did not occur when the ceramides differed only in the position of the amide bond. The individual molecular species (even if present in mixtures) were identified by gas chromatography-chemical ionization mass spectrometry. The evidence for a complete structure was obtained by enzyme splitting with alpha- and beta-galactosidases (the sequence of monosaccharides) and by negative ionization fast atom bombardment mass spectrometry. More than 400 molecular species of glycosylceramides were identified.
Institute of Microbiology, Czechoslovak Academy of Sciences, Prague.
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D002518:Ceramides / Q000032:analysis; D055598:Chemical Phenomena; D002621:Chemistry; D002708:Chlorella; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D009844:Oligosaccharides / Q000032:analysis; D013107:Sphingolipids / Q000032:analysis* / Q000302:isolation & purification
D002518:Ceramides; D009844:Oligosaccharides; D013107:Sphingolipids