A rapid and reproducible method with high selectivity was developed for simultaneous determination of a promising anti-brain tumor agent CAT3 and its two metabolites PF403 and GLU-PF403 in mouse plasma and brain. An economic deproteinization with septuple acetonitrile (v/v) was applied to pretreat the samples in this study. All analytes were well retained and separated on a CAPCELL CORE PC (2.7 μm, 2.1 mm I.D. × 150 mm, SHISEIDO Technologies) column with an eluting solvent of acetonitrile /water containing 0.1 % formic acid (v/v) at the flow rate of 0.2 mL per minute. The detection was carried out on a Q Exactive high resolution mass spectrometer equipped with a HESI ion source in parallel reaction monitoring (PRM) mode. The corresponding transitions for quantitation were 434.23→ 70.07 for CAT3, 350.17→70.07 for PF403, 526.21→70.07 for GLU-PF403, 364.19→70.07 for IS-1 and 625.18→317.07 for IS-2, respectively. A well-linear fit curve was achieved among the range of 0.1∼50 ng/mL for CAT3, 0.2∼100 ng/mL for PF403 and 2.5∼600 ng/mL for GLU-PF403 both in mouse plasma and brain homogenate. The intra-/inter-day accuracies of three analytes were within ±14.5 % and precisions were below to 13.44 %. The mean values of recovery of three compounds in mouse plasma and brain homogenate were among 98.06 ∼ 118.63 % and 81.04∼108.69 %. The analytes in NaF-treated ice cold blood of mouse was stable within tested 30 min. Plasma and brain homogenate samples had no obvious changes during all storage, sample treatment and analytic process of mouse plasma sample. The reproducible and reliable method was well employed to the research of CAT3 pharmacokinetic characteristics in mouse plasma and brain after a single intragastric administration at dose of 10 mg/kg.
Keywords: Anti-brain tumor agent; CAT3; LC–MS/MS; Pharmacokinetic.
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