Isolation and structural characterization of glucosylceramides from Ethiopian plants by LC/APCI-MS/MS

Chronic skin conditions such as atopic dermatitis, psoriasis and aged skin are characterized by defective skin barrier and dryness which are associated with reduced levels of skin ceramides (CERs). The beneficial effects of plant-derived CERs for skin hydration and skin barrier recovery have been shown in several studies. Although plenty of glucosylceramide (GlcCER)-based dietary supplements meant for skin barrier improvement have been marketed, there are limited commercial sources of plant GlcCERs. In an attempt to explore alternative GlcCER sources, a reversed phase LC-MS/MS method with atmospheric pressure chemical ionization (APCI) interface was developed for separation and structural identification of GlcCERs isolated from three plants. The GlcCERs were extracted from the seeds of grass pea (Lathyrus sativus L.), Ethiopian mustard (Brassica carinata) and haricot bean (Phaseolus vulgaris) and purified by column chromatography and preparative LC-MS. The individual GlcCER species were further separated and qualitatively analyzed by LC/APCI-MS/MS. The amount of GlcCERs in each plant was quantified by HPTLC. All GlcCER species detected in the three plants consisted of C18 di/trihydroxy sphingoid bases amide linked with hydroxy fatty acids (C14-C24). The trihydroxy SBs were acylated with very long chain FAs (C22-C24). The major GlcCERs derived from grass pea, Ethiopian mustard and haricot bean are composed of sphingenine (d18:1) linked to hydroxypalmitic acid (h16:0), 4-hydroxy-8-sphingenine (t18:1) coupled with hydroxynervonic acid (h24:1) and sphingadienine (d18:2) joined with h16:0, respectively. The GlcCERs contents in haricot bean (161.2mg/kg) and grass pea (130.0mg/kg) were found to be higher compared to Ethiopian mustard (71.8mg/kg). This qualitative and quantitative information suggests that the two plants of the Fabaceae family (haricot bean and grass pea) are potential alternative sources of GlcCERs for their use in products meant for the recovery of skin barrier function. The LC/APCI-MS/MS method described here has proven to be reliable for the screening of other potential plants containing GlcCERs.