Development and in vivo/in vitro evaluation of dexketoprofen trometamol (DT)-loaded nanosized, controlled release drug delivery system was aimed in this study.. DT-loaded Kollidon (R) SR (KSR) and Eudragit (R) RS 100 (ERS) polymeric nanoparticles were prepared using Nano Spray-Dryer (B-90, Buchi Labortechnik AG, Flawil, Switzerland). Both delivery systems prepared were characterized for selecting optimum formulations to investigate DT release profiles/kinetics, cytotoxicities and in vivo behaviors on animals. Structures of KSR and ERS were characterized by particle size and zeta potential measurements, shape and surface imaging, thermal analysis, X-ray diffraction, FTIR and H-1-NMR determinations. DT-loaded particles demonstrated nanostructured and spherical shape while in vitro relase studies showed extended release of DT incorporated. Korsmeyer-Peppas kinetic model was found to fit the best using DDSolver software program. Optimum formulations selected also showed prolonged analgesic activity in mice. Depending on in vitro and in vivo test results, formulations developed in this study seem to prolong release of DT from the nanoparticles prepared and are promising for extending analgesic activity.