Retention behaviour of ceramides in sub-critical fluid chromatography in comparison with non-aqueous reversed-phase liquid chromatography

• Published in: Journal of Chromatography A Vol. 883; no. 1; pp. 211 - 222
• Main Authors: Gaudin, Karen, Lesellier, Eric, Chaminade, Pierre, Ferrier, Danielle, Baillet, Arlette, Tchapla, Alain
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 23.06.2000; Elsevier
• Subjects: Analytical chemistry; Ceramides; Ceramides - analysis; Chemistry; Chromatographic methods and physical methods associated with chromatography; Chromatography, Liquid - methods; Exact sciences and technology; Other chromatographic methods; Ceramides; Retention chart; Retention behaviour; Subcritical fluid chromatography; Phase composition; Chromatographic retention; HPLC chromatography; Temperature effect; Retention factor; Experimental study; Mobile phase; Modeling; Chromatography; Operating conditions; Reversed phase chromatography; Pressure effect; Scattered light; Ceramide; Property structure relationship; Detection; Non aqueous solvent; Subcriticality; Comparative study; Stationary phase
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(00)00445-3

This study was devoted to the development of an analytical method for ceramide analysis in packed subcritical fluid chromatography (pSubFC). Monofunctional grafted silica support was found to be more suitable for ceramide analysis. Five Kromasil columns were coupled and the parameters, temperature, pressure and percentage of organic modifier in CO 2 were optimised, considering selectivity and analysis time. The final conditions were 31°C, 6% of methanol (MeOH) and 13 MPa. In these conditions the selectivity for structural differences (methylene group, unsaturation or two different bases) were studied. As classically observed, the methylene selectivity decreased with the increase of the eluotropic strength. Moreover, unlike in non-aqueous reversed-phase liquid chromatography (NARP–LC), adding a further unsaturation and two further methylene groups on ceramide results to an increase of retention in pSubFC. Moreover, this last technique allowed to separate ceramides with the same total number of carbons containing unsaturated fatty acids, when the distribution of carbon number of the two chain is very different. These results had enabled to plot retention chart in order to predict ceramide structure in view to identify additional ceramide. This retention chart was finally compared with the one already obtained in NARP–LC.