title: Measurement of steroid synthesis in zona glomerulosa cells by liquid chromatography-electrospray ionization-mass spectrometry: inhibition by nitric oxide.
abstract: A liquid chromatography-electrospray ionization-mass spectrometry method was developed to simultaneously determine the concentrations of aldosterone, corticosterone, cortisol, deoxycorticosterone, pregnenolone, and progesterone in bovine adrenal zona glomerulosa (ZG) cells. Steroids were extracted by liquid-liquid extraction, separated on a reverse-phase C18 column, ionized by electrospray, and detected by single-quadrupole mass spectrometry in a positive ion mode. All steroids formed sodium adducts at high abundance. Factors affecting the formation and signal of sodium adducts were investigated. The limits of detection (S/N=3) using selected ion monitoring are 2 pg for these steroids and 10 pg for pregnenolone. DETA NONOate, a nitric oxide donor, inhibited the basal, angiotensin-II-stimulated, and 25-hydroxycholesterol-stimulated syntheses of these steroids in ZG cells in a concentration-dependent manner. The technique demonstrates the ability to determine the individual steroid in each enzymatic step of aldosterone synthesis and the activity of steroidogenic enzymes in adrenal ZG cells.
authors.affiliation: Department of Pharmacology and Toxicology, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA.
authors.forename: Kasem
authors.initials: K
authors.lastname: Nithipatikom
authors.forename: Blythe B
authors.initials: BB
authors.lastname: Holmes
authors.forename: Marilyn A
authors.initials: MA
authors.lastname: Isbell
authors.forename: Craig J
authors.initials: CJ
authors.lastname: Hanke
authors.forename: Celso E
authors.initials: CE
authors.lastname: Gomez-Sanchez
authors.forename: William B
authors.initials: WB
authors.lastname: Campbell
mesh_terms: D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid; D003903:Deuterium; D009569:Nitric Oxide / Q000378:metabolism*; D012015:Reference Standards; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization; D013256:Steroids / Q000096:biosynthesis*; D015384:Zona Glomerulosa / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
chemical_list: D013256:Steroids; D000804:Angiotensin II; D009569:Nitric Oxide; D003903:Deuterium