Spectrophotometric quantification of astemizole and its demethylated metabolite in urine after TLC separation

The coupling of TLC and UV measurement for determination of astemizole and its main metabolite, O-demethylated derivative, in urine has been investigated. The metabolite like the drug absorbs maximally at almost the same wavelengths, which makes their simultaneous UV determination in biological fluids quite inapplicable. TLC separation on silica gel F254 utilizing chloroform/methanol (85:15, v/v) achieved the best fractionation of the two compounds from the matrix-components of urine. Concentration levels of 0.5-140 micrograms/ml (ppm) in worked-up sample could be reached by adopting the spectrophotometric measurements at 286 nm for ethanolic extracts of the silica layers carrying each individual compound against a blank silica. Varying levels of the intact drug and its phenolic primary metabolite could be accurately traced in urine samples following a 10 mg single oral dose (approximately 12.5 micrograms/kg) after different time intervals up to 12 h. Synthetic preparation of the metabolite by demethylating astemizole is mentioned and its physicochemical characterization is briefly discussed.