Accurate and sensitive liquid chromatography/tandem mass spectrometry simultaneous assay of seven steroids in monkey brain

Abstract

BackgroundFollowing its secretion mainly by the adrenal glands, dehydroepiandrosterone (DHEA) acts primarily in the cells/tissues which express the enzymes catalyzing its intracellular conversion into sex steroids by the mechanisms of intracrinology. Although reliable assays of endogenous serum steroids are now available using mass spectrometry (MS)-based technology, sample preparation from tissue matrices remains a challenge. This is especially the case with high lipid-containing tissues such as the brain. With the combination of a UPLC system with a sensitive tandem MS, it is now possible to measure endogenous unconjugated steroids in monkey brain tissue. MethodsA Shimadzu UPLC LC-30AD system coupled to a tandem MS AB Sciex Qtrap 6500 system was used. ResultsThe lower limits of quantifications are achieved at 250pg/mL for DHEA, 200pg/mL for 5-androstenediol (5-diol), 12pg/mL for androstenedione (4-dione), 50pg/mL for testosterone (Testo), 10pg/mL for dihydrotestosterone (DHT), 4pg/mL for estrone (E1) and 1pg/mL for estradiol (E2). The linearity and accuracy of quality controls (QCs) and endogenous quality controls (EndoQCs) are according to the guidelines of the regulatory agencies for all seven compounds. ConclusionWe describe a highly sensitive, specific and robust LC–MS/MS method for the simultaneous measurement of seven unconjugated steroids in monkey brain tissue. The single and small amount of sample required using a relatively simple preparation method should be useful for steroid assays in various peripheral tissues and thus help analysis of the role of locally-made sex steroids in the regulation of specific physiological functions.