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Profile and quantification of human stratum corneum ceramides by normal-phase liquid chromatography coupled with dynamic multiple reaction monitoring of mass spectrometry: development of targeted lipidomic method and application to human stratum corneum of different age groups

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Abstract

Skin, the largest organ of the human body, serves as the primary barrier to the external environment. Ceramides are one of the main constituents of stratum corneum (SC), playing an important role in skin barrier function. Therefore, comprehensive profiling and quantification of SC ceramide is important. Herein, a new targeted lipidomic method for human SC ceramide profiling and quantification is presented and tested. Normal-phase high-performance liquid chromatography coupled with dynamic multiple reaction monitoring mass spectrometry (NP-HPLC-dMRM-MS) was used to separate ceramides into subclasses and then characterize different ceramides within each subclass on the basis of their characteristics. In total, 483 ceramides were quantified in a single run within 20 min, covering 12 subclasses as well as some glycosylated ceramides not previously reported. Each subclass had typical standard substances (if available) that served to establish representative standard curves and were used for related substances with no standards. Linearity range, limit of quantification (LOQ), limit of detection (LOD), precision, accuracy, stability, and matrix effects were validated. dMRM increased sensitivity and accuracy greatly compared with common MRM (cMRM). This method was successfully applied to the study of human SC from different age groups. A total of 193 potential biomarkers were found to indicate age differences between children and adults. This method is an innovative approach for high-throughput quantification of SC ceramide.

Method establishment (MRM spectra by the established method) and method application (score scatter plots of authentic samples)

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Abbreviations

APCI:

Atmospheric pressure chemical ionization

CE:

Collision energy

cMRM:

Common MRM

dMRM-MS:

Dynamic multiple reaction monitoring mass spectrometry

ESI:

Electrospray ionization

IS:

Internal standard

LOD:

Limit of detection

LOQ:

Limit of quantification

NP-HPLC:

Normal-phase high-performance liquid chromatography

QC:

Quality control

RSD:

Relative standard deviation

RT:

Retention time

SC:

Stratum corneum

SIM:

Selected ion monitoring

SPE:

Solid-phase extraction

TIC:

Total ion chromatography

TLC:

Thin-layer chromatography

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Acknowledgments

This work was supported by grants of the Natural Science Foundation of Beijing Municipality (7154197) and grants of the National Health and Family Planning Commission of the People’s Republic of China (201402001).

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Authors and Affiliations

  1. State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, No. 2 Nanwei Road, Xicheng District, Beijing, 100050, China

    Zhi-Xin Jia & Jin-Lan Zhang

  2. Beijing Children’s Hospital, Capital Medical University, 56 South Lishi Road, Xicheng District, Beijing, 100045, China

    Chun-Ping Shen & Lin Ma

Authors
  1. Zhi-Xin Jia
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  2. Jin-Lan Zhang
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Correspondence to Jin-Lan Zhang.

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All experimental procedures were reviewed and approved by the Ethics Committee of Beijing Children’s Hospital and the methods were carried out in accordance with the Declaration of Helsinki.

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The authors have no conflicts of interest to declare.

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Jia, ZX., Zhang, JL., Shen, CP. et al. Profile and quantification of human stratum corneum ceramides by normal-phase liquid chromatography coupled with dynamic multiple reaction monitoring of mass spectrometry: development of targeted lipidomic method and application to human stratum corneum of different age groups. Anal Bioanal Chem 408, 6623–6636 (2016). https://doi.org/10.1007/s00216-016-9775-6

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  • DOI: https://doi.org/10.1007/s00216-016-9775-6

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