Understanding the mechanisms underlying the analgesic effect of new cyclooxygenase inhibitors is essential to identify dosing requirements in early stages of drug development. Accurate extrapolation to humans of in vitro and in vivo findings in preclinical species is needed to optimise dosing regimen in inflammatory conditions. The current investigation characterises the inhibition of prostaglandin E2 (PGE(2)) and thromboxane B2 (TXB(2)) by naproxen in vitro and in vivo in rat and human blood. The inhibition of PGE(2) in the absence or presence of increasing concentrations of naproxen (10(-8)-10(-1) M) was measured by ex vivo whole blood stimulation with LPS, whereas inhibition of TXB(2) was measured in serum following blood clotting. In further experiments, inhibition of PGE(2) and TXB(2) levels was also assessed ex vivo in animals treated with naproxen (2.5, 10, 25 mg kg(-1)). Subsequently, pharmacokinetic (PK)/pharmacodynamics (PD) modelling of in vitro and in vivo data was performed using nonlinear mixed effects in NONMEM (V). Inhibition of PGE(2) and TXB(2) was characterised by a sigmoid E(max) model. The exposure-response relationships in vitro and in vivo were of the same order of magnitude in both species. IC(80) estimates obtained in vitro were similar for PGE(2) inhibition (130.8 +/- 11 and 131.9 +/- 19 10(-6) M, mean +/- s.d. for humans and rats, respectively), but slightly different for TXB(2) inhibition (103.9+/-15 and 151.4 +/- 40 10(-6) M, mean +/- s.d. for humans and rats, respectively, P < 0.05). These differences, however, may not be biologically relevant. The results confirm the value of exposure-effect relationships determined in vitro as a means to predict the pharmacological activity in vivo. This analysis also highlights the need to parameterise concentration-effect relationships in early drug development, as indicated by the estimates of IC(80) for PGE(2) and TXB(2) inhibition.