The pharmacokinetics of the novel cephalosporin cefovecin were investigated in a series of in vivo, ex vivo and in vitro studies following administration to adult cats at 8 mg/kg bodyweight. Bioavailability and pharmacokinetic parameters were determined in a cross-over study after intravenous (i.v.) and subcutaneous (s.c.) injections. [14C]cefovecin was used to evaluate excretion for 21 days after s.c. administration. Protein binding was determined in vitro in feline plasma and ex vivo in transudate from cats surgically implanted with tissue chambers. After s.c. administration, cefovecin was characterized by rapid absorption with mean peak plasma concentrations of 141+/-12 microg/mL being achieved within 2 h of s.c. injection with full bioavailability (99%). The mean elimination half-life was 166+/-18 h. After i.v. administration, volume of distribution was 0.09+/-0.01 L/kg and mean plasma clearance was 0.35+/-0.04 mL/h/kg. Approximately 50% of the administered radiolabelled dose was eliminated over the 21-day postdose period via urinary excretion and up to approximately 25% in faeces. In vitro and ex vivo plasma protein binding ranged from 99.8% to 99.5% over the plasma concentration range 10-100 microg/mL. Ex vivo protein binding in transudate was as low as 90.7%. From 8 h postdose, concentrations of unbound (free) cefovecin in transudate were consistently higher than in plasma, with mean unbound cefovecin concentrations being maintained above 0.06 microg/mL (MIC90 of Pasteurella multocida) in transudate for at least 14 days postdose. The slow elimination and long-lasting free concentrations in extracellular fluid are desirable pharmacokinetic attributes for an antimicrobial with a 14-day dosing interval.