Simultaneous determination of 3,3′,4′,5,7-pentamethylquercetin and its possible metabolite 3,3′,4′,7-tetramethylquercetin in dog plasma by liquid chromatography–tandem mass spectrometry and its application to preclinical pharmacokinetic study

A sensitive, simple and rapid ultra fast liquid chromatography (UFLC)–ESI-MS/MS method was established for the simultaneous determination of 3,3′,4′,5,7-pentamethylquercetin (PMQ) and its possible metabolite 3,3′,4′,7-tetramethylquercetin (TMQ) in dog plasma using 4′,5,7-trimethylapigenin (TMA) as the internal standard. The plasma sample was pretreated with acetonitrile for protein precipitation and the analytes were separated on an Ultimate XB-CN column (5 μm, 2.1 mm × 150 mm) with the mobile phase consisting of acetonitrile and water (2:1, v/v). Detection was performed on a triple-quadrupole tandem mass spectrometer under a positive multiple reaction-monitoring mode (MRM). The mass transition ion-pair was followed as m/z 373.1–312.1 for PMQ, 359.1–344.0 for TMQ and 313.1–298.1 for TMA. The validated concentration ranged from 1.272 to 3060 ng/mL for PMQ and from 10.35 to 1725 ng/mL for TMQ. The lower limit of quantifications for PMQ and TMQ were 1.272 ng/mL and 10.35 ng/mL, respectively. The developed-method was successfully applied for the pharmacokinetic study of PMQ and its metabolite TMQ in dogs following a single oral dose.