Fluorometric determination of clobazam, a 1,5-benzodiazepine, in human plasma

A fluorometric procedure for clobazam, a 1,5-benzodiazepine, based on a fluorophore formed upon irradiation of the drug using short wavelength UV light (254 nm) for 35 min is presented. Fluorescence is linear over a 100-6400-ng/ml range using excitation and emission wavelengths of 350 and 400 nm, respectively. Application of the method to the determination of clobazam in spiked human plasma samples revealed that the drug can be determined at nanogram per milliliter levels with an accuracy of 1-5%. The procedure is specific for clobazam in samples containing its major plasma metabolite, N-desmethylclobazam, and also in samples containing 1,4-benzodiazepines and other selected drugs. A plasma level-time profile after oral administration of a single 40-mg dose of clobazam to a healthy adult male is also illustrated.