Abstract

A direct spectrofluorimetric method was developed and validated for the simultaneous determination of levofloxacin (LFX) and ciprofloxacin (CFX) in human urine without chromatographic separation. The two antibiotics were discriminated by their distinct excitation/emission maxima (286/465 nm for LFX at pH 7 and 270/429 nm for CFX at pH 9) and by their pH-dependent fluorescence. Fluorescence lifetimes were also measured (6.79 ns for LFX and 1.65 ns for CFX) and are discussed as an additional selectivity criterion for time-resolved detection. Calibration was linear from 0.04 to 0.75 µg mL⁻¹, as confirmed by analysis of variance and lack-of-fit testing. The limits of detection were 0.58 ng mL⁻¹ for LFX and 1.55 ng mL⁻¹ for CFX. In spiked urine from healthy volunteers, analysed by standard addition after deproteinization, mean recoveries were 103.0% for LFX and 99.0% for CFX, with intra-day relative standard deviations below 2%. The slopes of the standard-addition and calibration curves did not differ significantly, indicating no significant matrix effect. Common inorganic ions did not interfere at the tested levels, whereas ofloxacin and norfloxacin interfered above 0.12–0.25 µg mL⁻¹ because of spectral overlap; strategies to mitigate this interference are discussed. The method is rapid and inexpensive, is suitable for laboratories without chromatographic equipment, and can support pharmacokinetic studies and the monitoring of fluoroquinolones in urine.

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