Quantitative determination of a novel dual PPAR α/γ agonist using on-line turbulent flow extraction with liquid chromatography–tandem mass spectrometry

Turbulent flow chromatography (TFC) is a technique for the direct and efficient analysis of drugs and metabolites in biological matrices. We report here TFC on-line with an HPLC–MS/MS assay for the determination of 5-[2,4-dioxothiazolidin-5-yl)methyl]-2-methoxy-N-[[(4-trifluoromethyl)phenyl]methyl]benzamide (I, MK-0767, KRP297, Fig. 1) in plasma. Samples were transferred using an automated system followed by the addition of internal standard (II), prepared in 0.1 M ammonium acetate (pH 4.0). The plasma samples were directly injected onto a C18 turbulent flow column on-line with an HPLC–MS/MS system, and the analytical column used was a ThermoHypersil Keystone C18. Detection was achieved by MS/MS, using positive ionization on a TurboIonSpray® probe, operated in multiple reaction monitoring (MRM) mode. The linear range was 4–2000 ng/mL for I when using 50 μL of plasma. The method exhibited good linearity and reproducibility. The method also showed good selectivity and ruggedness when applied to clinical samples, and was successfully cross-validated with a conventional off-line SPE, LC–MS/MS method.