The acute ecotoxicity of different diameters of silica and polyethyleneimine polystyrene (PS-PEI)nanoparticles (NPs)was assessed on a test battery of aquatic organisms representing different trophic levels. Daphnia magna, Thamnocephalus platyurus, Pseudokirchneriella subcapitata and Vibrio fischeri, were employed in a series of standard acute ecotoxicity tests and work was complemented with two cytotoxicological end points on a rainbow trout gonadal cell line (RTG-2). Physico-chemical characterization of the NPs was performed in the different test media employed, using dynamic light scattering (DLS) and zeta potentiometry. In contrast to silica NPs exposure, for which no effect was observed for concentrations up to 1000 µg ml-1 for all in vivo aquatic organisms tested, significant toxicity was detected after exposure to PS-PEI NPs at concentrations from 0.40 µg ml-1 to 416.5 µg ml-1. Differing sensitivities for each NP diameter for the different organisms was observed as: P. subcapitata > D. magna > T. platyurus > V. fischeri. The effects observed were dependent in some cases on the NP size, a higher effect being observed for the larger NPs. Finally, cytotoxicity studies showed an effect at the highest concentrations for both sets of NPs which was greater in the case of the PS-PEI NPs. However, as agglomeration and sedimentation of the nanoparticles was observed at these concentrations, the cytotoxicity studies were found not to be a reliable ecotoxicity test model.