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Development and validation of LC/APCI-MS method for the quantification of oat ceramides in skin permeation studies

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Abstract

Ceramides (CERs) are the backbone of the intercellular lipid lamellae of the stratum corneum (SC), the outer layer of the skin. Skin diseases such as atopic dermatitis, psoriasis, and aged skin are characterized by dysfunctional skin barrier and dryness which are associated with reduced levels of CERs. Replenishing the depleted epidermal CERs with exogenous CERs has been shown to have beneficial effects in improving the skin barrier and hydration. The exogenous CERs such as phyto-derived CERs (PhytoCERs) can be delivered deep into the SC using novel topical formulations. This, however, requires investigating the rate and extent of skin permeation of CERs. In this study, an LC/APCI-MS method to detect and quantify PhytoCERs in different layers of the skin has been developed and validated. The method was used to investigate the skin permeation of PhytoCERs using Franz diffusion cells after applying an amphiphilic cream containing PhytoCERs to the surface of ex vivo human skin. As plant-specific CERs are not commercially available, well-characterized CERs isolated from oat (Avena abyssinica) were used as reference standards for the development and validation of the method. The method was linear over the range of 30–1050 ng/mL and sensitive with limit of detection and quantification of 10 and 30 ng/mL, respectively. The method was also selective, accurate, and precise with minimal matrix effect (with mean matrix factor around 100%). Even if more than 85% of oat CERs in the cream remained in the cream after the incubation periods of 30, 100, and 300 min, it was possible to quantify the small quantities of oat CERs distributed across the SC, epidermis, and dermis of the skin indicating the method’s sensitivity. Therefore, the method can be used to investigate the skin permeation of oat CERs from the various pharmaceutical and cosmeceutical products without any interference from the skin constituents such as the epidermal lipids.

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Abbreviations

CER:

Ceramide

DR:

Dermis

EP:

Epidermis

ELSD:

Evaporative light scattering detector

FA:

Fatty acid

GlcCER:

Glucosylceramide

LC:

Liquid chromatography

LOD:

Limit of detection

LOQ:

Limit of quantification

MS/MS:

Tandem mass spectrometry

MF:

Matrix factor

SIM:

Selected ion monitoring

S/N:

Signal to noise ratio

RSD:

Relative standard deviation

SB:

Sphingoid base

SC:

Stratum corneum

SD:

Standard deviation

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Acknowledgements

The authors would like to thank Manuela Woigk, Claudia Bruhne, Andrea Stennett, and Anja Ehrlich for their excellent technical assistance. Efrem N. Tessema greatly acknowledges the financial support provided by the German Academic Exchange Service (DAAD).

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Authors and Affiliations

  1. Department of Pharmaceutical Technology and Biopharmaceutics, Institute of Pharmacy, Martin Luther University Halle-Wittenberg, Wolfgang-Langenbeck-Str. 4, 06120, Halle (Saale), Germany

    Efrem N. Tessema & Reinhard H. H. Neubert

  2. Department of Pharmaceutics and Social Pharmacy, School of Pharmacy, College of Health Sciences, Addis Ababa University, Addis Ababa, Ethiopia

    Tsige Gebre-Mariam

  3. Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, 06120, Halle (Saale), Germany

    Andrej Frolov

  4. Department of Dermatology and Venereology, Medical Faculty, Martin Luther University Halle-Wittenberg, Ernst-Grube-Str. 40, 06120, Halle (Saale), Germany

    Johannes Wohlrab

  5. Institute of Applied Dermatopharmacy, Martin Luther University Halle-Wittenberg, Weinbergweg 23, 06120, Halle (Saale), Germany

    Johannes Wohlrab & Reinhard H. H. Neubert

Authors
  1. Efrem N. Tessema
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  2. Tsige Gebre-Mariam
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  3. Andrej Frolov
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  4. Johannes Wohlrab
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  5. Reinhard H. H. Neubert
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Corresponding author

Correspondence to Reinhard H. H. Neubert.

Ethics declarations

An ethical clearance was obtained from the Ethics Committee of the Faculty of Medicine, Martin Luther University Halle-Wittenberg for the SC lipid extraction and ex vivo permeability studies. An informed consent was also obtained from the volunteer subjects.

Conflict of interest

The authors declare that they have no conflict of interest.

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Tessema, E.N., Gebre-Mariam, T., Frolov, A. et al. Development and validation of LC/APCI-MS method for the quantification of oat ceramides in skin permeation studies. Anal Bioanal Chem 410, 4775–4785 (2018). https://doi.org/10.1007/s00216-018-1162-z

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  • DOI: https://doi.org/10.1007/s00216-018-1162-z

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