title: A novel LC-MS/MS assay for the simultaneous determination of melatonin and its two major metabolites, 6-hydroxymelatonin and 6-sulfatoxymelatonin in dog plasma: Application to a pharmacokinetic study.
abstract: A convenient and specific liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous quantification of melatonin (MEL) and its major metabolites, 6-hydroxymelatonin (6-O-MEL) and 6-sulfatoxymelationin (S-O-MEL) in dog plasma. After plasma samples were deproteinized with acetonitrile, the post-treatment samples were analyzed on a Phenomenex Kinetex C18 column (50×2.1 mm, 1.7 μm) interfaced with a triple quadrupole tandem mass spectrometer. Electrospray ionization mode (ESI) and multiple reaction monitoring were used to assay MEL and its metabolites. Acetonitrile and 5 mM ammonium acetate were used as the mobile phase with a gradient elution at a flow rate of 0.2 mL/min. The analytical run time of 6.5 min was divided into two periods according to ionization mode. S-O-MEL was monitored in negative ionization mode (period 1), while MEL and 6-O-MEL were detected in positive ionization mode (period 2). All calibration curves showed good linearity (r>0.991) over the concentration range with a lower limit of quantification (LLOQ) of 0.02 ng/mL for MEL, 0.04 ng/mL for 6-O-MEL and 0.50 ng/mL for S-O-MEL. The intra- and inter-day precision was within 13.5% in terms of relative standard deviation (RSD%) and the accuracy within 13.0% in terms of relative error. This convenient and specific LC-MS/MS method was successfully applied to the pharmacokinetic study of MEL and its metabolites in Beagle dogs after an oral dose of 2.0mg MEL. After ingestion of MEL, S-O-MEL was the predominant component circulating in blood. 6-O-MEL showed similar pharmacokinetic profile to that of MEL.
authors.affiliation: Department of Pharmaceutical Analysis, Pharmacy School, Shenyang Pharmaceutical University, Shenyang 110016, China.
authors.forename: Huimin
authors.initials: H
authors.lastname: Zhao
authors.affiliation: Department of Pharmaceutical Analysis, Pharmacy School, Shenyang Pharmaceutical University, Shenyang 110016, China.
authors.forename: Yifei
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Pharmaceutical Analysis, Pharmacy School, Shenyang Pharmaceutical University, Shenyang 110016, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Yuan
authors.affiliation: Department of Pharmaceutical Analysis, Pharmacy School, Shenyang Pharmaceutical University, Shenyang 110016, China.
authors.forename: Shu
authors.initials: S
authors.lastname: Liu
authors.affiliation: Department of Pharmaceutical Analysis, Pharmacy School, Shenyang Pharmaceutical University, Shenyang 110016, China.
authors.forename: Shuang
authors.initials: S
authors.lastname: Man
authors.affiliation: Department of Pharmaceutical Analysis, Pharmacy School, Shenyang Pharmaceutical University, Shenyang 110016, China.
authors.forename: Haiyan
authors.initials: H
authors.lastname: Xu
authors.affiliation: Department of Pharmaceutical Analysis, Pharmacy School, Shenyang Pharmaceutical University, Shenyang 110016, China. Electronic address: lynn1975@163.com.
authors.forename: Xiumei
authors.initials: X
authors.lastname: Lu
mesh_terms: D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods; D004285:Dogs; D005260:Female; D008297:Male; D008550:Melatonin / Q000031:analogs & derivatives* / Q000097:blood* / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: C054513:6-sulfatoxymelatonin; D008550:Melatonin; C012365:6-hydroxymelatonin
keywords: 6-Hydroxymelatonin; 6-Sulfatoxymelatonin; LC–MS/MS; Melatonin; Pharmacokinetics