(1)H NMR-based metabolomics investigation of Daphnia magna responses to sub-lethal exposure to arsenic, copper and lithium.

Metal and metalloid contamination constitutes a major concern in aquatic ecosystems. Thus it is important to find rapid and reliable indicators of metal stress to aquatic organisms. In this study, we tested the use of (1)H nuclear magnetic resonance (NMR) - based metabolomics to examine the response of Daphnia magna neonates after a 48h exposure to sub-lethal concentrations of arsenic (49μgL(-1)), copper (12.4μgL(-1)) or lithium (1150μgL(-1)). Metabolomic responses for all conditions were compared to a control using principal component analysis (PCA) and metabolites that contributed to the variation between the exposures and the control condition were identified and quantified. The PCA showed that copper and lithium exposures result in statistically significant metabolite variations from the control. Contributing to this variation was a number of amino acids such as: phenylalanine, leucine, lysine, glutamine, glycine, alanine, methionine and glutamine as well as the nucleobase uracil and osmolyte glycerophosphocholine. The similarities in metabolome changes suggest that lithium has an analogous mode of toxicity to that of copper, and may be impairing energy production and ionoregulation. The PCA also showed that arsenic exposure resulted in a metabolic shift in comparison to the control population but this change was not statistically significant. However, significant changes in specific metabolites such as alanine and lysine were observed, suggesting that energy metabolism is indeed disrupted. This research demonstrates that (1)H NMR-based metabolomics is a viable platform for discerning metabolomic changes and mode of toxicity of D. magna in response to metal stressors in the environment.