Orbifloxacin (ORBI) is a third-generation fluoroquinolone developed exclusively for use in veterinary medicine, mainly in companion animals. This antimicrobial agent has bactericidal activity against numerous gram-negative and gram-positive bacteria. A few chromatographic methods for its analysis have been described in the scientific literature. Here, coupling of sequential-injection analysis and solid-phase spectroscopy is described in order to develop, for the first time, a terbium-sensitized luminescent optosensor for analysis of ORBI. The cationic resin Sephadex-CM C-25 was used as solid support and measurements were made at 275/545 nm. The system had a linear dynamic range of 10-150 ng mL(-1), with a detection limit of 3.3 ng mL(-1) and an R.S.D. below 3% (n = 10). The analyte was satisfactorily determined in veterinary drugs and dog and horse urine.