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Microanalytical systems for separations of stratum corneum ceramides
Author(s) -
Quinton Loïc,
Gaudin Karen,
Baillet Arlette,
Chaminade Pierre
Publication year - 2006
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.200500362
Subject(s) - chemistry , chromatography , stratum corneum , ceramide , elution , formic acid , chromatography detector , polyvinyl alcohol , pullulan , acetone , analytical chemistry (journal) , high performance liquid chromatography , organic chemistry , biochemistry , polysaccharide , medicine , apoptosis , pathology
The small amount of lipids from human skin obtained with noninvasive sampling method led us to investigate microanalytical separation techniques. The lipid class analysis was performed with a micro polyvinyl alcohol‐silica (PVA‐Sil®) column. The gradient elution was from heptane to acetone/butanol 90:10 v/v in 4%/min at 78 μL/min. In addition an evaporative light scattering detector (ELSD) was modified for micro‐LC. All solvents contained 0.1% of triethylamine and formic acid in stoichiometric amount, which increased the ELSD response. In these conditions, the cholesterol eluted before free fatty acid, and squalene and triglycerides close to the dead volume. The various ceramide classes eluted following the order of the increased number of hydroxyl groups. The LOD for ceramides was 2.2 ng. The advantages of this method are the use of a normal stationary phase more reliable due to its chemical stability, its surface homogeneity and its development in microchromatography without chlorinated solvents which offers small LOD and the whole profile of lipids present in stratum corneum (SC). A method using a narrow‐bore PVA‐Sil column was used to collect ceramide fraction. Then the molecular species were analysed with a porous graphitic carbon column in capillary LC using a gradient from CH 3 OH/CHCl 3 70:30 v/v to CHCl 3 at 2%/min with a flow rate at 5 μL/min. The LOD obtained for ceramide was 1 ng. Both methods were assessed with SC samples obtained by rinsing a 5.7 cm 2 area of the forearm with 25 mL of ethanol.