ABSTRACT Four controlled microcosms were prepared, one of which served as control, whereas the other three received treatments of 10, 50, and 150 mg/L of florfenicol, an antibacterial drug developed initially for aquaculture applications. Water and sediment samples were collected according to a pre‐set time schedule to determine its fate in the aquatic environment. The results demonstrated that florfenicol amine was not detectable in any treatment groups. The half‐life of florfenicol in pond water was 30.76, 52.92, and 51.48 hours for the 10, 50, and 150 mg/L treatment groups, respectively; the corresponding half‐lives in sediment were 25.42, 38.40, and 53.59 days. Further, the 50 and 150 mg/L florfenicol treatment groups inhibited alkaline protease activity; however, all treatment groups inhibited sediment microflora structural diversity.