Gamma (gamma)-irradiation is finding increasing use in the sterilization of pharmaceutical products. However, irradiation also might affect the performance of drug delivery systems. In this study, the influence of gamma-irradiation on the characteristics of chitosan microparticles was investigated. The diclofenac sodium was incorporated into chitosan microparticles by spray-drying method. The chitosan microparticles (placebo and drug-loaded) were irradiated at doses of 5, 15, and 25 kGy using a (60)Co source. Later, the microparticles were characterized by Fourier transform infrared (FTIR) spectroscopy, electron paramagnetic resonance (EPR) spectroscopy, X-ray diffraction, differential scanning calorimetry, scanning electron microscopy, and atomic force microscopy. In addition, microparticles also were evaluated for their sizes, drug content, swelling, and drug release behavior. Encapsulation efficiency of irradiated and nonirradiated microparticles was essentially the same. Notably, surface roughness (rms) of placebo microparticles decreased significantly after gamma-irradiation when compared with nonirradiated placebo microparticles. FTIR spectroscopy revealed that gamma-irradiation of chitosan microparticles induced neither cross-linking nor formation of new group in the chitosan matrix. EPR spectroscopy results showed that the gamma rays induced only one kind of free radical in the chitosan matrix. Size, crystallinity, and thermal properties of the chitosan microparticles did not change much after gamma-irradiation. gamma-irradiated microparticles, exhibited a slightly higher drug release rate and low swelling capacity than the nonirradiated microparticles.