Reversed-phase liquid chromatographic method for the determination of 7-nitrobenz-2-oxa-1,3-diazol-4-yl-labelled lipid analogues

• Published in: Journal of chromatography. B, Biomedical sciences and applications Biomedical sciences and applications, 1998-06, Vol.710 (1), p.9-16
• Main Authors: Frijters, Charles M.G, Tuijn, Coosje J, Hoek, Frans J, Groen, Albert K, Oude Elferink, Ronald P.J, N. Zegers, Bart
• Format: Article
• Language: English
• Subjects: Analytical, structural and metabolic biochemistry; Animals; Bile - metabolism; Biological and medical sciences; Chromatography, Liquid - methods; Fluorescent Dyes; Fundamental and applied biological sciences. Psychology; Glucosylceramides - isolation & purification; Glucosylceramides - metabolism; Lipids; Mice; Other biological molecules; Oxadiazoles - isolation & purification; Oxadiazoles - metabolism; Sphingolipids; Sphingomyelins - isolation & purification; Sphingomyelins - metabolism
• ISSN: 0378-4347; 1387-2273
• DOI: 10.1016/S0378-4347(98)00132-7

This paper reports the development of a dual column system for the simultaneous separation of fluorescent short-chain ceramide, 6-[(7-nitrobenz-2-oxa-1,3,-diazol-4-yl[NBD])amino]hexanoyl-sphingosine and its metabolites, C6-NBD-sphingomyelin and C6-NBD-glucosylceramide, as well as the fluorescent derivatives of choline and serine phosphatides. The method enables the separation of these lipids in a single run on the basis of the polarity of their headgroups and hydrophobicity of their acyl backbone. The fluorescent properties of the NBD-label make it possible to quantitate small amounts of NBD-lipid analogues. The sensitivity of the presented method thus permits the use of small sample volumes and the determination of NBD-lipid analogues secreted into mouse bile directly, without prior extraction or concentration steps.