Naphthenic acids are key contributors to toxicity of heavy oil refining effluents.

Oil refining produces vast quantities of wastewater with harmful contaminants that can be released back into the environment with a possible risk of toxicity to aquatic wildlife and human populations. Hence the importance of adequate wastewater treatment to achieve safe effluents that protect both ecological and human health. However, some refining effluents are linked to serious pollution problems even after treatment, partly because little progress has been made in determining the causative agents of the observed biological effects, resulting in non-targeted treatment. Here, we followed an effect-directed analysis (EDA) approach using Aliivibrio fischeri as biosensor to show that naphthenic acids (NAs) are important components of refining wastewater resulting from the processing of heavy crude oil. Furthermore, we demonstrate that besides mixture effects, NAs have a significant contribution to the toxicity exerted by these effluents. Profiling of the NA mixture was conducted using high resolution liquid chromatography-Orbitrap, which evidenced that O2 NAs corresponded to 90% of the NAs detected. Our findings contrast with previous reports where classic NAs have been found between 15% and 72% and could explain the significant biological effects observed in A. fischeri. This study broadens the body of evidence pointing at mixture effects and low-concentration pollutants as the cause of toxicity from RWW, in addition to NAs resulting from the processing of heavy crude oil. Our results can serve as a starting point for setting better effluent discharge standards relevant to oil refining wastewater resulting from heavy crude oil and help improve wastewater treatment plants to reduce effluent toxicity.