In vivo distribution of free long-chain sphingoid bases in the human stratum corneum by high-performance liquid chromatographic analysis of strippings.
Conditions were established for the in vivo determination of free long-chain bases (phytosphingosine, sphingosine and sphinganine) in human stratum corneum layers by reversed-phase HPLC analysis of adhesive-tape strippings. Long-chain bases were first extracted from individual strippings with pure methanol in an ultrasound bath, then o-phthalaldehyde derivatives were separated on a C18 column with a gradient from methanol-water (80:20, v/v) to pure methanol. By performing a second series of strippings for protein determination it was possible to express the amounts of free long-chain bases per milligram of protein for each individual stratum corneum layer. Co-elution of endogenous phytosphingosine, sphingosine and spinganine was demonstrated in separate experiments by addition of standards to typical strips. From a study of three human volunteers, average concentrations of free long-chain bases in the stratum corneum were found to be 2.8, 1.2 and 0.5 nmol/mg of protein for phytosphingosine, sphingosine and sphinganine, respectively.
Laboratoires de Recherche Fondamentale, L'Oreal, Aulnay-sous-Bois, France.
N
N
Flamand
P
P
Justine
F
F
Bernaud
A
A
Rougier
Q
Q
Gaetani
D000328:Adult; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D007202:Indicators and Reagents; D008297:Male; D008875:Middle Aged; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011506:Proteins / Q000737:chemistry; D012867:Skin / Q000737:chemistry*; D013107:Sphingolipids / Q000032:analysis*; D013110:Sphingosine / Q000031:analogs & derivatives / Q000032:analysis; D009764:o-Phthalaldehyde / Q000737:chemistry
D007202:Indicators and Reagents; D011506:Proteins; D013107:Sphingolipids; D009764:o-Phthalaldehyde; D011493:Protein Kinase C; C012491:phytosphingosine; D013110:Sphingosine; C005682:safingol