A previous study found that coke leachates (CL) collected from oil sands field sites were acutely toxic to Ceriodaphnia dubia; however, the cause of toxicity was not known. Therefore, the purpose of this study was to generate CL in the laboratory to evaluate the toxicity response of C. dubia and perform chronic toxicity identification evaluation (TIE) tests to identify the causes of CL toxicity. Coke was subjected to a 15-d batch leaching process at pH 5.5 and 9.5. Leachates were filtered on day 15 and used for chemical and toxicological characterization. The 7-d median lethal concentration (LC50) was 6.3 and 28.7% (v/v) for pH 5.5 and 9.5 CLs, respectively. Trace element characterization of the CLs showed Ni and V levels to be well above their respective 7-d LC50s for C. dubia. Addition of ethylenediaminetetraacetic acid significantly (p ≤ 0.05) improved survival and reproduction in pH 5.5 CL, but not in pH 9.5 CL. Cationic and anionic resins removed toxicity of pH 5.5 CL only. Conversely, the toxicity of pH 9.5 CL was completely removed with an anion resin alone, suggesting that the pH 9.5 CL contained metals that formed oxyanions. Toxicity reappeared when Ni and V were added back to anion resin-treated CLs. The TIE results combined with the trace element chemistry suggest that both Ni and V are the cause of toxicity in pH 5.5 CL, whereas V appears to be the primary cause of toxicity in pH 9.5 CL. Environmental monitoring and risk assessments should therefore focus on the fate and toxicity of metals, especially Ni and V, in coke-amended oil sands reclamation landscapes.