Soil solutions were collected from soils contaminated differently by the emissions from a copper smelter and incubated with beech litter. Five times in 30 days of incubation, soil solutions were acquired with MacroRhizon samplers and examined using the chemometric approach and two ecotoxicological assays: Microtox with Vibrio fischeri bacteria and Phytotoxkit with Sinapis alba seeds. Copper speciation in soil solutions was modeled in the MinteQ program. Application of beech litter to soils resulted in a considerable increase of copper solubility. The toxicity of soil solutions was associated with total copper concentrations in solutions, copper speciation, and pH values. The toxicity measured by the Phytotoxkit test in soil solutions collected from non-amended soils was higher than that measured in Microtox, and was attributed to high concentrations of copper organic complexes in solutions. The application of beech litter, rich in dissolved organic carbon DOC, resulted in a radical increase of soil solution toxicity to both indicating organisms. This effect, particularly well expressed in the case of V. fischeri, was apparently caused by a decrease in pH and associated increase of total Cu concentrations in solutions.