Fluorimetric determination of norfloxacin in plasma and urine samples after thin-layer chromatographic separation

A thin-layer chromatographic (TLC) assay for the determination of norfloxacin (1-ethyl-6-fluoro-1,4-dihydro-4-oxo-7- (piperazinyl)-3-quinolinecarboxylic acid), a new antibacterial agent, in human plasma and urine is described. Norfloxacin was extracted from plasma samples using commercial cartridges, urine samples were simply diluted with methanol. Plasma extracts or urine samples were applied to TLC plates (silica gel). Chromatography was performed with a mobile phase consisting of methanol, ethanol and water, the atmosphere in the tank being saturated with ammonia. The fluorescence intensity of norfloxacin was enhanced by dipping the plate into a paraffin/citric acid mixture. Peaks were quantified by fluorimetric measurement (excitation 313 nm/emission 390 nm, cut-off filter). The method showed acceptable precision and linearity in the range of 0.01 to 4 micrograms/ml for plasma and 1 to 100 micrograms/ml for urine. The assay was shown to be applicable to human plasma and urine samples.