Isolation of ceramide fractions from human stratum corneum lipid extracts by high-performance liquid chromatography

This report describes a procedure to isolate ceramides from human stratum corneum lipid extracts using preparative liquid chromatography (LC) and semi-preparative high-performance liquid chromatography (HPLC) methods, which is economical in construction and operation. The composition of isolated individual 'peaks' of separated ceramide fractions is reported. These peaks were quantified by scanning of the chromatographic plates stained after high performance thin-layer chromatography (HPTLC). The HPTLC was done by automated multiple development (AMD). The enrichment of the ceramides by liquid chromatography turned out to be necessary for their separation from more polar lipids of the human stratum corneum lipid extract. Only the evaporative light scattering detector gave a stable baseline, reproducible results and eliminated the 'solvent front' in which peaks of interest could coelute. Fluorescence measurements of extracted human stratum lipids indicated lipofuscin-like substances.