title: Development and validation of an UPLC-MS/MS assay for quantitative analysis of the ghrelin receptor inverse agonist PF-5190457 in human or rat plasma and rat brain.
abstract: PF-5190457 is a ghrelin receptor inverse agonist that is currently undergoing clinical development for the treatment of alcoholism. Our aim was to develop and validate a simple and sensitive assay for quantitative analysis of PF-5190457 in human or rat plasma and rat brain using liquid chromatography-tandem mass spectrometry. The analyte and stable isotope internal standard were extracted from 50 μL plasma or rat brain homogenate by protein precipitation using 0.1% formic acid in acetonitrile. Chromatography was carried out on an Acquity UPLC BEH C18 (2.1 mm × 50 mm) column with 1.7 μm particle size and 130 Å pore size. The flow rate was 0.5 mL/min and total chromatographic run time was 2.2 min. The mobile phase consisted of a gradient mixture of water: acetonitrile 95:5% (v/v) containing 0.1% formic acid (solvent A) and 100% acetonitrile containing 0.1% formic acid (solvent B). Multiple reaction monitoring was carried out in positive electro-spray ionization mode using m/z 513.35 → 209.30 for PF-5190457 and m/z 518.47 → 214.43 for the internal standard. The recovery ranged from 102 to 118% with coefficient of variation (CV) less than 6% for all matrices. The calibration curves for all matrices were linear over the studied concentration range (R(2) ≥ 0.998, n = 3). The lower limit of quantification was 1 ng/mL in rat or human plasma and 0.75 ng/g in rat brain. Intra- and inter-run mean percent accuracies were between 85 and 115% and percent imprecision was ≤15%. The assays were successfully utilized to measure the concentration of PF-5190457 in pre-clinical and clinical pharmacology studies of the compound.
authors.affiliation: Clinical Pharmacokinetics Research Laboratory, Department of Biomedical and Pharmaceutical Sciences, University of Rhode Island, 495A College of Pharmacy, 7 Greenhouse Road, Kingston, RI, 02881, USA.
authors.forename: Mwlod
authors.initials: M
authors.lastname: Ghareeb
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Leggio
authors.forename: Ayman
authors.initials: A
authors.lastname: El-Kattan
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Akhlaghi
mesh_terms: D000818:Animals; D001384:Azetidines / Q000097:blood / Q000378:metabolism; D001921:Brain / Q000378:metabolism*; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D057230:Limit of Detection; D051381:Rats; D054440:Receptors, Ghrelin / Q000037:antagonists & inhibitors*; D013141:Spiro Compounds / Q000097:blood / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D001384:Azetidines; C000606714:PF-5190457; D054440:Receptors, Ghrelin; D013141:Spiro Compounds