Although biosynthesized nanoparticles are regarded as green products, research on their toxicity to aquatic food chains is scarce. Herein, biosynthesized silver nanoparticles (Alcea rosea-silver nanoparticles, AR-AgNPs) were produced by the reaction of Ag ions with leaf extract of herbal plant Alcea rosea. Then, the toxic effects of AR-AgNPs and their precursors such as Ag⁺ ions and coating agent (A. rosea leaf extract) on organisms of different trophic levels of a freshwater food chain were investigated. To the three studied aquatic organisms including phytoplankton (Chlorella vulgaris), zooplankton (Daphnia magna) and fish (Danio rerio), the coating agents of AR-AgNPs showed no toxic effects, and Ag⁺ ions were more toxic in comparison to AR-AgNPs. Further investigations revealed that the release of Ag⁺ ions from AR-AgNPs to the test media were not considerable due to the high stability of AR-AgNPs, thus the toxicity stemmed mainly from the particles of AR-AgNPs in all the three trophic levels. Based on values of 72-h EC₅₀ for C. vulgaris, 48-h LC₅₀ for D. magna and 96-h LC₅₀ for D. rerio, the most sensitive organism to AR-AgNPs exposure was D. magna (the second trophic level).