title: A LC-MS/MS method to evaluate the hepatic uptake of the liver-specific magnetic resonance imaging contrast agent gadoxetate (Gd-EOB-DTPA) in vitro and in humans.
abstract: Gadoxetate (Gd-EOB-DTPA, Primovist®) is a frequently used liver-specific magnetic resonance imaging (MRI) contrast agent which disposition is so far not fully understood in humans. Here, we describe the development and validation of a selective and sensitive quantification method to measure cellular in vitro concentrations as well as human serum concentrations of gadoxetate. The drug was measured after protein precipitation with acetonitrile and ethyl acetate-mediated sample concentration using amoxicillin as internal standard and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) for detection. Hydrophilic interaction chromatography (HILIC) was performed by using the column Atlantis® HILIC Silica (2.1 mm × 100 mm), a step-elution gradient with acetonitrile and ammonium acetate (5mM, pH 3.8) as mobile phases and a flow rate of 200 μl/min. The MS/MS detection was done in the negative multiple reaction monitoring (MRM) mode by monitoring the m/z transitions 681.3/635.2 for gadotrexate and 363.8/222.7 for the internal standard. The method was validated between 5 and 4000 ng/ml in serum and between 1.25 and 500 ng/ml in cell lysates. The method was shown to possess sufficient specificity, accuracy, precision and stability without any matrix effects, thereby fulfilling current bioanalytical guidelines. The developed assay was successfully applied to quantify gadoxetate in cellular uptake studies in OATP1B1-transfected cell lines and to monitor serum concentrations-time profiles from a clinical pilot study performed in healthy volunteers carrying the wild-type or the functionally relevant variants T521C (*5) and A388G (*1b) of the hepatic uptake transporter OATP1B1.
authors.affiliation: Department of Clinical Pharmacology, University of Greifswald, Friedrich-Loeffler-Str. 23d, D-17487 Greifswald, Germany.
authors.forename: Jia
authors.initials: J
authors.lastname: Jia
authors.forename: Markus
authors.initials: M
authors.lastname: Keiser
authors.forename: Ali
authors.initials: A
authors.lastname: Nassif
authors.forename: Werner
authors.initials: W
authors.lastname: Siegmund
authors.forename: Stefan
authors.initials: S
authors.lastname: Oswald
mesh_terms: D002460:Cell Line; D002853:Chromatography, Liquid / Q000379:methods*; D003287:Contrast Media / Q000493:pharmacokinetics*; D019786:Gadolinium DTPA / Q000493:pharmacokinetics*; D006801:Humans; D066298:In Vitro Techniques; D008099:Liver / Q000378:metabolism*; D008279:Magnetic Resonance Imaging / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D003287:Contrast Media; C073590:gadolinium ethoxybenzyl DTPA; D019786:Gadolinium DTPA