Antibiotics and pesticides, as well as various emerging contaminants that are present in surface waters, raise significant environmental concerns. Advanced oxidation processes, which are employed to eliminate these substances, have demonstrated remarkable effectiveness. However, during the degradation process, by-products that are not completely mineralized are generated, posing a substantial risk to aquatic ecosystem organisms; therefore, it is crucial to assess effluent ecotoxicity following treatment. This study aimed to assess the toxicity of effluents produced during the removal of amoxicillin and glyphosate with a Fenton-type process using a laminar structure catalyzed with iron (Fe) and copper (Cu). The evaluation included the use of , , and , and mutagenicity testing was performed using strains TA98 and TA100 of . Both treated and untreated effluents exhibited inhibitory effects on root growth in , even at low concentrations ranging from 1% to 10% /. Similarly, negative impacts on the growth of algal cells of were observed at concentrations as low as 0.025% /, particularly in cases involving amoxicillin-copper (Cu) and glyphosate with copper (Cu) and iron (Fe). Notably, in the case of , mortality was noticeable even at concentrations of 10% /. Additionally, the treatment of amoxicillin with double-layer hydroxides of Fe and Cu resulted in mutagenicity (IM ≥ 2.0), highlighting the necessity to treat the effluent further from the advanced oxidation process to reduce ecological risks.