HPLC/MS(N) analysis of retinoids

James E Evans, Peter McCaffery

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

This protocol describes a highly sensitive and selective method to quantify retinoids using normal-phase HPLC with online APCI MS(N). The retinoids are key regulators of gene expression, retinol being oxidized via a retinaldehyde intermediate to retinoic acid (RA) which activates specific nuclear receptors, the signalling of which is turned off by oxidative inactivation of the ligand to 4-oxo-RA and other metabolites. Many of these retinoids are present only transiently at low concentrations in tissues and during analysis are labile to heat, light, and oxygen. HPLC with online APCI MS(N) provides a rapid technique to quantify these retinoids simultaneously. Techniques to extract the retinoids and prevent their degradation are described, with an emphasis on transcriptionally active RA. RA controls patterning of gene expression in the embryo, organizing embryonic morphology in the central nervous system. Similarly, a patterned distribution of RA controls function of the adult CNS, a tissue particularly difficult to analyse for RA because of its high lipid content. To understand how these patterns are organized in the brain and change over time, it is essential to determine the concentration of RA in small areas of tissues, and techniques of exquisite sensitivity are indispensable.
Original languageEnglish
Pages (from-to)149-162
Number of pages14
JournalMethods in Molecular Biology
Volume652
DOIs
Publication statusPublished - May 2010

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Retinoids
Tretinoin
High Pressure Liquid Chromatography
Retinaldehyde
Keto Acids
Gene Expression
Regulator Genes
Cytoplasmic and Nuclear Receptors
Vitamin A
Embryonic Structures
Central Nervous System
Hot Temperature
Oxygen
Ligands
Lipids
Light
Brain

Keywords

  • animals
  • chromatography, high pressure liquid
  • isomerism
  • mass spectrometry
  • rats
  • retinoids
  • silicon dioxide

Cite this

HPLC/MS(N) analysis of retinoids. / Evans, James E; McCaffery, Peter.

In: Methods in Molecular Biology, Vol. 652, 05.2010, p. 149-162.

Research output: Contribution to journalArticle

Evans, James E ; McCaffery, Peter. / HPLC/MS(N) analysis of retinoids. In: Methods in Molecular Biology. 2010 ; Vol. 652. pp. 149-162.
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