Methodological approach to the intracrine study and estimation of DHEA and DHEA-S using liquid chromatography–tandem mass spectrometry (LC–MS/MS)

• Steroid analysis using LC–MS/MS is reliable and sensitive.
• Sample processing is very important for quantification of steroids.
• Derivatization technique increases the sensitivity and specificity.
• 13C-labeled substrates combined with LC–MS/MS are useful for intracrine research.

A reliable and sensitive method for analyzing steroids using liquid chromatography tandem mass spectrometry (LC–MS/MS) is required for research concerning dehydroepiandrosterone (DHEA), which plays a central role in steroid hormone biosynthesis and metabolism. Furthermore, after the first proposal of the concept of intracrine DHEA, stable isotope tracer analysis, which is useful for structural recognition as well as determination of steroids, has been required to evaluate physiological action and hormone biosynthesis/metabolism in target organs. We describe sample processing and analysis methods for simultaneous quantification of multiple hormones, including DHEA, in serum, saliva and tissue using LC–MS/MS. A derivatization technique compatible with each functional group for measuring 3β-hydroxy-5-enes, such as DHEA and 5α/5β-steroids, with high sensitivity and specificity is also described. Finally, we describe a newly developed method for intracrine research using stable isotope-labeled 13C-steroid substrates with tracer analysis of their metabolites by LC–MS/MS.This article is part of a Special Issue entitled’Essential role of DHEA’.