Determination of 5-(3-Chlorophenyl)-4-hexyl-2,4-dihydro-3H-1,2,4-triazole-3-thione in Mouse Brain Tissue by Microwave-Assisted Extraction and High-Performance Liquid Chromatography with Fluorescence Detection

A sensitive and selective method was developed for the determination of 5-(3-chlorophenyl)-4-hexyl-2,4-dihydro-3H-1,2,4-triazole-3-thione (TP-315), an antiepileptic compound, in the tissue homogenate of mouse brains by microwave-assisted extraction and high performance liquid chromatography with fluorescence detection by use of an external standard. The analyses were performed on a C₁₈ column using a mobile phase consisting of 60% acetonitrile in water. Fluorescence detection was performed at an excitation wavelength of 263 nm and an emission wavelength of 411 nm. The method provided a high extraction yield with recoveries exceeding 90%. The precision values expressed by intra- and inter-day relative standard deviations were lower than 5%. The validated procedure was employed to develop a brain concentration profile after intraperitoneal administration of a single oral dose equal to 300 mg per kg of body weight. The time to the maximum anticonvulsant action of TP-315 in a mouse maximal electroshock-induced seizure test was 30 min after administration.