title: Quantitative determination of a novel insulin sensitizer and its para-hydroxylated metabolite in human plasma by LC-MS/MS.
abstract: I, 5-[3-[3-(4-phenoxy-2-propylphenoxy)-propoxy]-phenyl]-2,4-thiazolidinedione sodium salt, is a dual alpha/gamma peroxisome proliferator-activated receptor (PPAR) agonist for potential use in diabetic patients. The compound has a para-hydroxylated metabolite, II, which has also been shown to exhibit PPAR activity. An LC-MS/MS method for the simultaneous determination of I and its active metabolite (II) in human plasma has been successfully developed. The method consists of treating 0.5 ml plasma with ammonium acetate (pH 9.6; 50mM) and extracting I, II and internal standard (III, Fig. 2) with 5 ml ethyl acetate. The ethyl acetate is evaporated and the samples are reconstituted in 0.1 ml acetonitrile:0.1% formic acid (65:35, v/v). The entire extraction procedure, as well as sample collection, was performed in glass tubes and vials to overcome the analytes adherence to polypropylene. A linear HPLC gradient was used to separate the analyte, metabolite, internal standard, and other interfering, non-quantitated metabolites. Detection was by negative ionization MS/MS on a turbo ionspray probe. Precursor-->product ion combinations were monitored in multiple reaction monitoring (MRM) mode. The linear range is 0.05-20 ng/ml for I and 0.1-20 ng/ml for II. Recoveries were 59.4, 90.1 and 56.8% for I, II and III, respectively. Intraday variation using this method was <==7.0% for I and <==9.2% for II. The method exhibits good linearity and reproducibility for each analyte and good sensitivity, selectivity and robustness when used for the analysis of clinical samples.
authors.affiliation: Merck Research Laboratories, Department of Drug Metabolism, WP75A-303, West Point, PA 19486, USA. kerry_riffel@merck.com
authors.forename: Kerry A
authors.initials: KA
authors.lastname: Riffel
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Polinko
authors.forename: Hengchang
authors.initials: H
authors.lastname: Song
authors.forename: Ronda K
authors.initials: RK
authors.lastname: Rippley
authors.forename: Man-Wai
authors.initials: MW
authors.lastname: Lo
mesh_terms: D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D006801:Humans; D006900:Hydroxylation; D007328:Insulin / Q000378:metabolism; D013678:Technology, Pharmaceutical / Q000379:methods*; D045162:Thiazolidinediones / Q000097:blood* / Q000378:metabolism
chemical_list: D007328:Insulin; D045162:Thiazolidinediones; C089946:2,4-thiazolidinedione