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Plasma disposition of florfenicol in channel catfish was investigated after an oral multidose (10 mg/kg for 10 days) administration in freshwater at water temperatures ranging from 24.7 to 25.9 °C. Florfenicol concentrations in plasma were analyzed by means of liquid chromatography with MS/MS detection. After the administration of florfenicol, the mean terminal half-life (t1/2), maximum concentration at steady-state (Css(max)), time of Css(max) (Tmax), minimal concentration at steady-state (Css(min)), and Vc/F were 9.0 h, 9.72 μg/mL, 8 h, 2.53 μg/mL, and 0.653 L/kg, respectively. These results suggest that florfenicol administered orally at 10 mg/kg body weight for 10 days could be expected to control catfish bacterial pathogens inhibited in vitro by a minimal inhibitory concentration value of <2.5 μg/mL.