Drug and metabolite analysis. Serum samples and liver homogenates were mixed with acetonitrile containing ethopropazine as internal standard (IS). After vortexing and centrifugation, protein was removed, and the supernatant was diluted and transferred to autosampler vials for liquid chromatographytandem mass spectrometry (LC/MS/MS) analysis. LC/MS/MS analysis was performed by use of a Shimadzu LC-20 high performance liquid chromatography (HPLC) system coupled to a QTRAP 3200 tandem quadrupole mass spectrometer (AB SCIEX, Foster City, CA) operated under control of Analyst v. 1.4.2 software. The Ascentis Express C18 HPLC Column (5 cm 3 2.1 mm, 2.7 lm) was maintained at 50°C. A volume of 2 ll was injected into the HPLC system and eluted with a gradient based on 10mM ammonium acetate in H 2 O (solvent A) and methanol (solvent B): 0-0.5 min, 10% solvent B; 0.5 -1 min, 10-98% solvent B; 1-4 min, 98% solvent B; 4-6 min, 10% solvent B; flow rate, 0.3 ml/min. Positive mode electrospray ionization was used for all analyses. Mass spectrometry parameters, including declustering potential and collision energy, were optimized independently for each analyte and IS. Multiple reactions monitoring the m/z transitions were used for the quantitative analysis of AMD (m/z 646.1/201.1), DEA (m/z 618.1/547.0), and ethopropazine (m/z 313.1/114.1). The LC/MS/MS method achieved lower limits of quantification: 50 ng/ml for AMD and 5 ng/ml for DEA. Analytical reproducibility was judged to be ± 10.2% in the middle of the calibrated range of concentrations. The Pierce BCA protein assay kit (Thermo Scientific, Rockford, IL) was used to determine protein concentration in the liver homogenates.