As basis for evidence-based analgesia refinement, species-specific pharmacokinetic and tolerability profiles of carprofen were determined in rats for least aversive administration routes and prolonged treatment. Further, potential influence on behavioral pain indicators was evaluated. LC-MS/MS determined plasma concentrations in Sprague-Dawley rats (n = 21/sex) after subcutaneous (s.c.) injection (5 mg/kg) and during a 5-day treatment via the drinking water (d.w., 10 mg/kg/24 h). Irwin test parameters, clinical scoring, body weight, body temperature, fluid and food intake, grimace scale, burrowing, nesting, hematology, and histopathology were investigated. Plasma concentrations early after injection were higher in females, reached a maximum (C