title: The development of an ultra performance liquid chromatography-coupled atmospheric pressure chemical ionization mass spectrometry assay for seven adrenal steroids.
abstract: An ultra performance liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (UPLC-APCI-MS) method was developed for the separation and quantification of adrenal steroid metabolites from heterologous expression media. Steroids were extracted by liquid-liquid extraction, separated on a Waters UPLC BEH C18 column, ionized by APCI, and detected using a triple quadrupole mass spectrometer in APCI positive mode with single ion monitoring. The incorporation of UPLC enabled the detection of seven structurally closely related steroids at between 5 and 40 ng/ml using run times of 11 min. The adrenal steroidogenic enzyme cytochrome P450 17-hydroxylase/17,20-lyase (CYP17) was expressed in the yeast Pichia pastoris and in nonsteroidogenic COS-1 cells, and used as a model system to evaluate the detection and quantification of adrenal steroid metabolites by UPLC-APCI-MS.
authors.affiliation: Department of Biochemistry, University of Stellenbosch, Stellenbosch 7602, South Africa.
authors.forename: Karl-Heinz
authors.initials: KH
authors.lastname: Storbeck
authors.forename: Norbert W
authors.initials: NW
authors.lastname: Kolar
authors.forename: Marietjie
authors.initials: M
authors.lastname: Stander
authors.forename: Amanda C
authors.initials: AC
authors.lastname: Swart
authors.forename: Désirée
authors.initials: D
authors.lastname: Prevoo
authors.forename: Pieter
authors.initials: P
authors.lastname: Swart
mesh_terms: D000311:Adrenal Glands / Q000737:chemistry*; D000818:Animals; D001274:Atmospheric Pressure; D019556:COS Cells; D002522:Chlorocebus aethiops; D002853:Chromatography, Liquid / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D013256:Steroids / Q000032:analysis*
chemical_list: D013256:Steroids