Determination of 5-HT receptor antagonists, MEFWAY and MPPF using liquid chromatography electrospray ionization tandem mass spectrometry in rat plasma and brain tissue

A simple, selective, and sensitive liquid chromatography electrospray ionization tandem mass spectrometry (LC–ESI-MS/MS) method was validated for the determination of 4-fluoromethyl-N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridyl)cyclohexane-1-carboxamide (MEFWAY) and 4-fluoro-N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridyl)benzamide (MPPF) in rat plasma and brain samples, respectively. Plasma and brain samples were extracted with a mixture of acetonitrile and methanol (1:1, v/v) and then separated on a C18 column (Gemini 3 μm 110 Å, 50 × 2.00 mm ID, Phenomenex, USA). Quantitation was performed using LC–ESI-MS/MS in multiple-reaction monitoring (MRM) mode with positive ion electrospray ionization (ESI). The limit of quantification (LOQ) of 5 ng/mL and 1 ng/mL were obtained in 50 μL brain homogenate and plasma, respectively. The analytical linear ranges of this method were 1–4000 ng/mL in plasma and 5–4000 ng/mL in brain homogenate with a correlation coefficients (R2) greater than 0.9993. The intra- and inter-day precision and accuracy values were within the assay validation guideline (lower than 13.0%). The analytes in plasma and brain samples were stable after three freeze–thaw cycles, long-term storage (one month at −80 °C), and short-term (4 h) storage at room temperature. The present method was successfully applied to plasma-brain pharmacokinetic studies to investigate brain penetration of a single dose of MEFWAY and MPPF in rats.

► A simple, rapid, and sensitive LC–ESI-MS/MS method of MEFWAY and MPPF.
► Validation for the determination of MEFWAY and MPPF in rat plasma and brain.
► The sample preparation followed by one-step protein precipitation using acetonitrile and methanol.
► The assay variability limits set forth in the FDA guidelines.
► The present method can be applied to plasma-brain pharmacokinetic studies to investigate brain penetration in rats.