title: LC-MS/MS method for the determination of two metabolites of tribendimidine, deacylated amidantel and its acetylated metabolite in plasma, blood and dried blood spots.
abstract: Tribendimidine has emerged as potential alternative to praziquantel in the treatment of Opisthorchis viverrini infections. To support its clinical development program, a quantitative high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) assay was developed for tribendimidine's degradation product deacylated amidantel (dADT) and its acetylated metabolite adADT. Analytical sample preparation included protein precipitation for blood and plasma, and direct processing of dried blood spots (DBS). The analytes were detected by multiple reaction monitoring with electrospray ionization in the positive mode (dADT: 178.3→133.1 m/z, adADT: 220.4→175.1 m/z, tribendimidine 294.3→249.0 m/z). A pentafluorophenyl (PFP) phase Kinetex analytical column (2.6 μm, 100 Å, 50 mm × 4.6 mm) with a 6 min lasting mobile phase gradient program of ammonium acetate and acetonitrile was applied. The method was validated with respect to precision, accuracy, linearity, sensitivity, and selectivity. The analytical range in plasma and blood was 1-1000 ng/ml and in DBS 10-2000 ng/ml (R(2)>0.99). Recoveries determined using four different human blood batches were in the range of 70-90%. Inter- and intra-assay accuracy and precision deviations were at least ≤12.2%. dADT and adADT were stable within the autosampler for 72 h (10°C), for 4 h at room temperature, for 3 month at -80°C, and after three freeze and thaw cycles. DBS samples should be stored at -20°C. The validation results demonstrated that the LC-MS/MS method is precise, accurate and selective and can be applied for pharmacokinetic studies with tribendimidine.
authors.affiliation: Department of Medical Parasitology and Infection Biology, Swiss Tropical and Public Health Institute, Basel, Switzerland; University of Basel, P.O. Box, CH-4003 Basel, Switzerland.
authors.forename: Urs
authors.initials: U
authors.lastname: Duthaler
authors.affiliation: Department of Medical Parasitology and Infection Biology, Swiss Tropical and Public Health Institute, Basel, Switzerland; University of Basel, P.O. Box, CH-4003 Basel, Switzerland. Electronic address: jennifer.keiser@unibas.ch.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Keiser
authors.affiliation: Department of Pharmaceutical Sciences, Division of Pharmaceutical Technology, University of Basel, Klingelbergstrasse 50, CH-4056 Basel, Switzerland.
authors.forename: Jörg
authors.initials: J
authors.lastname: Huwyler
mesh_terms: D000107:Acetylation; D000969:Antinematodal Agents / Q000097:blood* / Q000378:metabolism; D001801:Blood Stains; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D057230:Limit of Detection; D015394:Molecular Structure; D010655:Phenylenediamines / Q000097:blood* / Q000378:metabolism; D012015:Reference Standards; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000969:Antinematodal Agents; D010655:Phenylenediamines; C056017:tribendimidine; C019144:amidantel
keywords: Anthelmintic drug; Deacylated amidantel; Opisthorchis viverrini; Pharmacokinetics; Tribendimidine