Florida manatees (Trichechus manatus latirostris) frequently present to rehabilitation care facilities for various conditions, including boat strike trauma, cold stress syndrome, and brevetoxicosis. Throughout the course of treatment, antimicrobial use to treat respiratory disease is frequently warranted. To date, clinicians have extrapolated dosages based on established information available in bovine and equine medicine. The routes of administration, efficacy, and treatment intervals are considerations in dealing with critical wild animals. The use of tulathromycin, a triamilide antibiotic, has been studied in multiple domestic species of economic importance, including cattle, small ruminants, and swine, and has revealed efficacy against respiratory diseases. Given this information, this antibiotic has also been used in manatees with positive clinical outcomes. This study employed sparse sampling and evaluated banked plasma samples at various time intervals post–tulathromycin administration obtained during the clinical treatment course of nine animals during their rehabilitation. Preliminary pharmacokinetic analysis following administration of a single dose estimated a half-life of 33.75 h and volume of distribution per fraction absorbed (Vz/F = 4.29 L/kg). The pharmacokinetic behavior of tulathromycin in Florida manatees can be used to optimize dosage regimens in this species.