Since surfactant or emulsifiers remained on the nanoparticle surface significantly affect the physicochemical properties, the biodegradation rate, the biodistribution, and the biocompatibility of nanoparticles, surfactant-free nanoparticles should be good candidate. surfactant-free PLGA nanoparticles were successfully prepared by both the dialysis method and the solvent diffusion method. The PLGA nanoparticles prepared using the solvent diffusion method has a smaller particle size than the dialysis method. The solvent diffusion method was better for a higher loading efficiency than the dialysis method but the nanoparticle yield was lower. Testosterone (TST) release from the PLGA nanoparticles was dependent on the particle size rather than the drug contents. Testosterone release from the PLGA nanoparticles prepared by the solvent diffusion method using acetone was faster than those prepared by the dialysis method. TST release from the PLGA nanoparticles prepared by the solvent diffusion method using acetone and the dialysis method using dimethylformamide (DMF) was completed for 4 days while the PLGA nanoparticles prepared by the dialysis method using acetone showed approximately 80% TST release after 4 days. Since the PLGA nanoparticle degradation ratio was below 20% within 5 days at all samples while TST release completed within 4 days, TST release was dependent on the diffusion mechanism rather than degradation.