We investigated the allelopathic activity of two submersed macrophytes with different growth forms and nutrient uptake modes, Ceratophyllum demersum and Najas marina ssp. intermedia. A bioassay-directed method development revealed optimal extraction solvents for allelochemicals from both macrophytes. For Najas, 50% methanol and for Ceratophyllum 50% acetone yielded the strongest inhibition in the agar-diffusion assay with various filamentous or chroococcal cyanobacteria as target species. Further fractionation by liquid-liquid extraction (LLE) and solid phase extraction (SPE) procedures showed that both aquatic plants appear to have more than one active fraction, one being hydrophilic and one moderately lipophilic. The water-soluble allelochemicals may inhibit phytoplankton whereas the lipophilic allelochemicals may act through direct cell-cell contact, e.g., against epiphytes. Both macrophytes also exuded allelopathically active compounds into the surrounding medium as shown by SPE of their incubation water.