Ecotoxicological effect factors are part of the analysis of relative impacts by chemical contaminants on ecosystems. Uncertainty distributions, represented by the 90% confidence interval, belonging to ecotoxicological effect factors for freshwater ecosystems were determined. This study includes 869 high production volume chemicals, related to 7 nonspecific toxic modes of action (TMoAs). The ecotoxicological effect factors are divided into a TMoA-specific part and a chemical-specific part. The 90% confidence interval of the TMoA-specific part of the effect factor ranges from 23 orders of magnitude for acrylate toxicity to 2 orders of magnitude for nonpolar narcosis. The range in the TMoA-specific part of the effect factor is mainly caused by uncertainty in the spread in toxic sensitivity between species (σ_j). Average uncertainty in the chemical-specific part of the effect factors depends on the number of species tested and ranges on average from a factor of 5 for more than 3 species tested to a factor of about 1,000 for 2 species tested. Average uncertainty in the ecotoxicological effect factors ranges from a factor of 100 for more than 3 species tested to a factor of nearly 10,000 for 2 species tested. It is recommended that the ecotoxicological effect factor of a chemical is based on toxicity data of at least 4 species.