title: Chemical derivatization of neurosteroids for their trace determination in sea lamprey by UPLC-MS/MS.
abstract: This article describes the development and validation of a sensitive and robust method for the determination of neurosteroids in sea lamprey, an ancestral animal in vertebrate evolution. Chemical derivatization was used to enhance the detection of neurosteroids containing ketone function by ultra-high performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Iminooxy derivatives of 12 oxosteroids and three internal standards were monitored by positive electrospray tandem mass spectrometry using the neutral loss of sulfate. Limit of quantification, extraction recovery and matrix effect were first evaluated and SPE using C18 sorbent was selected after comparison with liquid/liquid extraction and protein precipitation. Matrix effect ranged from 89.6% to 113.1% in plasma and from 79.8% to 100.0% in the brain. Recovery values ranged from 80.0% to 103.8% in plasma and from 86.3% to 107.9% in the brain. Chromatographic separation was achieved by reverse phase chromatography (2.1mm×100mm, 1.7µm particle size, C18) with a binary gradient between methanol and 0.1% formic acid in water. Limit of quantification ranged from 5 to 10pg/mL and was up to 80 times lower than those for non-derivatized steroids. Accuracy and precision parameters were determined and inter- and intra-day at three concentrations: 50, 500 and 5000pg/mL. This method was applied to analyze real samples. progesterone (P), pregnenolone (P5), 7-hydroxy-pregnenolpne (7P5), 17-hydroxy-pregnenolpne (17P5)dehydroepiandrosterone (DHEA), androstenedienone (A4), testosterone (T), dihydrotestosterone (DHT), allopregnanolone (THP), 11-hydroxy-androstenedienone (11A4) and 11-Deoxycortisol (S) were measured in sea lamprey brain and plasma matrixes.
authors.affiliation: Department of Fisheries and Wildlife, Michigan State University, Room 13 Natural Resources Building, 480 Wilson Road, East Lansing, MI 48824 USA.
authors.forename: Ugo
authors.initials: U
authors.lastname: Bussy
authors.affiliation: Department of Fisheries and Wildlife, Michigan State University, Room 13 Natural Resources Building, 480 Wilson Road, East Lansing, MI 48824 USA.
authors.forename: Mar
authors.initials: M
authors.lastname: Huertas
authors.affiliation: Department of Fisheries and Wildlife, Michigan State University, Room 13 Natural Resources Building, 480 Wilson Road, East Lansing, MI 48824 USA.
authors.forename: Yu-Wen
authors.initials: YW
authors.lastname: Chung-Davidson
authors.affiliation: Department of Fisheries and Wildlife, Michigan State University, Room 13 Natural Resources Building, 480 Wilson Road, East Lansing, MI 48824 USA.
authors.forename: Ke
authors.initials: K
authors.lastname: Li
authors.affiliation: Department of Fisheries and Wildlife, Michigan State University, Room 13 Natural Resources Building, 480 Wilson Road, East Lansing, MI 48824 USA. Electronic address: liweim@msu.edu.
authors.forename: Weiming
authors.initials: W
authors.lastname: Li
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D002851:Chromatography, High Pressure Liquid; D059625:Liquid-Liquid Extraction; D008297:Male; D018377:Neurotransmitter Agents / Q000032:analysis* / Q000097:blood*; D048428:Petromyzon; D052616:Solid Phase Extraction; D053719:Tandem Mass Spectrometry
chemical_list: D018377:Neurotransmitter Agents
keywords: Chemical derivatization; ESI; LC-MS/MS; Neurosteroids; Petromyzon marinus