title: Quantitative analysis of ceramides using a novel lipidomics approach with three dimensional response modelling.
abstract: In the outermost layer of the skin, the stratum corneum (SC), ceramides form a diverse and essential pool of lipids. Due to their diversity and the limited availability of synthetic standards it is challenging to quantitatively analyse all SC ceramides independently. We aim to perform a detailed analysis of ceramides on SC harvested from in vivo and ex vivo skin, therefore, a LC/MS method was developed in which all steps from sample acquisition until data analysis were examined and optimized. Improving extraction efficiency of ceramides resulted in an increase in efficiency from 71.5% to 99.3%. It was shown that sample harvesting by tape-stripping in vivo was accurate and precise. A full scan MS method was developed, compatible with all sample types, enabling simultaneously qualitative and quantitative data analysis. A novel three dimensional response model was constructed to quantify all detected ceramides from full scan data using a limited amount of synthetic ceramides. The application is demonstrated on various SC sample types. When ex vivo SC was regenerated during human skin culture, increases are observed in the amount of the ceramide sphingosine subclasses, in mono unsaturated ceramides (which have an cis-double bond in the acyl chain), and ceramides with a short C34 carbon chain (ceramides with a total carbon chain of 34 carbon atoms), compared with native human skin. These changes in ceramide levels are also often encountered in diseased skin.
authors.affiliation: Leiden Academic Centre for Drug Research, Leiden University, Leiden, The Netherlands. Electronic address: boitenwa@lacdr.leidenuniv.nl.
authors.forename: Walter
authors.initials: W
authors.lastname: Boiten
authors.affiliation: Leiden Academic Centre for Drug Research, Leiden University, Leiden, The Netherlands. Electronic address: s.absalah@lacdr.leidenuniv.nl.
authors.forename: Samira
authors.initials: S
authors.lastname: Absalah
authors.affiliation: Leiden Academic Centre for Drug Research, Leiden University, Leiden, The Netherlands. Electronic address: rvreeken@ITS.JNJ.com.
authors.forename: Rob
authors.initials: R
authors.lastname: Vreeken
authors.affiliation: Leiden Academic Centre for Drug Research, Leiden University, Leiden, The Netherlands. Electronic address: bouwstra@lacdr.leidenuniv.nl.
authors.forename: Joke
authors.initials: J
authors.lastname: Bouwstra
authors.affiliation: Leiden Academic Centre for Drug Research, Leiden University, Leiden, The Netherlands. Electronic address: smedenjvan@lacdr.leidenuniv.nl.
authors.forename: Jeroen
authors.initials: J
authors.lastname: van Smeden
mesh_terms: D000328:Adult; D002138:Calibration; D002518:Ceramides / Q000032:analysis* / Q000737:chemistry; D005260:Female; D006801:Humans; D008297:Male; D055432:Metabolomics / Q000379:methods*; D012015:Reference Standards; D015203:Reproducibility of Results; D012867:Skin / Q000378:metabolism; D013237:Stereoisomerism
chemical_list: D002518:Ceramides
keywords: Extraction efficiency; Mass spectrometry; Skin; Stereoisomers; Stratum corneum; Tape-stripping