The Sava River, the largest and most commercially valuable water body in the riparian countries, receives inputs of organic and inorganic compounds from a variety of domestic and industrial activities that may affect the health of human beings and wildlife. In this work, the chronic toxicity of sediment, sediment porewater and surface water from the Sava River and connecting tributaries to the freshwater algae Pseudokirchneriella subcapitata was assessed to characterise the potential impact on aquatic organisms. Samples of different environmental matrices were either tested directly (porewater) or subjected to pre-concentration (sediments and surface water) prior to testing in a 72 h growth inhibition assay using P. subcapitata. The results show that a combination of pre-concentration and bioassay testing was able to characterise the toxic potential and to identify several compartment-specific "hot spots" along the Sava River. Based on the resulting data, a crude risk assessment identified that some of the locations may represent a risk to algae. Additional testing using multiple species and exposure phases is required to conduct a thorough risk assessment, however.