Mixture Toxicity of Three Unconventional Gas Fracking Chemicals, Barium, O‐Cresol, and Sodium Chloride, to the Freshwater Shrimp Paratya australiensis

The 96‐h acute toxicity of barium (Ba(2+)), o‐cresol, and sodium chloride (NaCl) to Paratya australiensis was assessed in single, binary, and ternary combinations in addition to three biochemical assays: glutathione S‐transferase, acetylcholinesterase, and sodium–potassium adenosine triphosphatase. The 96‐h lethal concentrations that expressed 50% mortality (LC50) in the single‐toxicant exposures were Ba(2+) = 23.4 mg/L, o‐cresol = 12.2 mg/L, and NaCl = 4198 mg/L. Mortality from o‐cresol exposure occurred between 11 and 22 mg/L, whereas Ba(2+) was more gradual across 10–105 mg/L, and most of the NaCl mortality occurred between 2050 and 4100 mg/L. Toxic units were used to assess the binary and ternary interactions of the toxicants. A more than additive effect was observed for most combinations in the binary chemical exposures, with the ternary combinations yielding highly synergistic interactions. Greater synergism was observed with the 96‐h LC50 of o‐cresol in combination with the three concentrations of NaCl (1025, 2050, and 3075 mg/L) compared with Ba(2+), with toxic units of 0.38, 0.48, and 0.10 (o‐cresol) and 0.71, 0.67, and 0.50 (Ba(2+)). No notable enzyme activity trends were observed in the enzyme biomarker responses from both individual and mixture exposures. Although acute single‐species toxicity tests tend to underestimate the effects of Ba(2+), o‐cresol, and NaCl on populations, communities, and ecosystems in seminatural (e.g., mesocosms) and natural systems, there are currently no published acute toxicity data available for P. australiensis and the three toxicants used in the present study. The present study shows that chemicals with different toxicity mechanisms can potentially lead to more synergistic responses. Environ Toxicol Chem 2023;42:481–494. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.