Early warning signs: Applications of in vivo NMR spectroscopy to understand the impacts of industrial effluents.

Industrial effluents represent one of the most significant threats to water quality. To prevent any serious and sometimes irreversible damage to freshwater ecosystems, an improved understanding of how effluents impact aquatic environments is necessary. Environmental metabolomics is an ideal approach as it enables the identification of the impacts of a stressor on a model organism at the molecular level. In vivo 2D NMR is a powerful tool to understand responses to environmental stress directly in living organisms. Here, a framework to best obtain, process, and interpret in vivo NMR data is introduced within the context of water research. The impacts of unconcentrated wastewater effluents from five industrial sectors (wastewater treatment plant, cannabis production, containerboard, electronics, and electroplating) on Daphnia magna are explored. 2D NMR provides the spectral dispersion needed to discriminate a wide range of metabolites. Statistics are then used to filter the in vivo metabolome, leaving only the signals that are significantly changing, permitting targeted metabolite assignment. The subsequent pathway analysis enables the in-depth exploration of the metabolic impacts of each effluent. Remarkably similar metabolic responses were observed between some effluents, despite their significantly different compositions, while others showed unique metabolic profiles. These findings highlight the need for improved methods to relate the compositions of complex mixtures to their respective molecular level impacts. Overall, this work introduces in vivo NMR as a powerful approach for understanding the impacts of wastewater on aquatic environments. Results suggest that the approach is reliable and highly complementary to other, more routinely employed techniques.