Carmustine-loaded poly(lactic acid) (PLA) nanoparticles were prepared with a spontaneous emulsification-solvent diffusion method by using Pluroinc F68 as emulsifying agent, and the influence of micellar behavior of surfactant in aqueous solution on the sizes of the nanoparticles was studied. The results revealed that F68 dissolved as unimers in water or sodium acetate-acetic acid buffer with pH value ranged from 5.0 to 5.5, resulting in larger PLA particles (226 nm) or clumps. In acidic media with pH of 4.0 and 4.5, however, F68 aggregated into micelles over the concentration of 3.7 and 4.8 mg/mL, as a result, ultrasmall nanoparticles with a size 50 and 80 nm could be obtained, respectively. The change in size had less effect on the drug content and entrapment efficiency, and the nanoparticles strongly enhanced the cytotoxic effect of the loaded drug in vitro, and this effect being more relevant for prolonged incubation times. (C) 2008 Wiley Periodicals, Inc.