Environmental context. Polycyclic aromatic hydrocarbons (PAHs) are common contaminants, but there has been limited research investigating the responses of earthworm exposure to sub-lethal PAH concentrations. In this study, ¹H nuclear magnetic resonance (NMR) metabolomics was used to characterise the metabolic responses of Eisenia fetida earthworm exposure in contact tests to 10, 50 and 100 μg cm^–2 naphthalene, phenanthrene and pyrene. The findings of this study highlight the potential of metabolomics as a tool for monitoring earthworm responses to sub-lethal concentrations of problematic environmental contaminants.

Abstract. Metabolic responses of earthworm exposure to the polycyclic aromatic hydrocarbons (PAHs) naphthalene, phenanthrene and pyrene in contact tests were measured using ¹H nuclear magnetic resonance (NMR). Novel metabolites were not detected but principal component analysis (PCA) showed that earthworms exposed to 10, 50 and 100 μg cm^–2 naphthalene, phenanthrene and pyrene differed from unexposed (control) earthworms. Partial least-squares-discriminant analysis (PLS-DA) showed that earthworms had statistically significant responses to PAH exposure, except for 10 μg cm^–2 naphthalene and 50 μg cm^–2 pyrene. Leucine, valine, alanine, lysine and maltose were identified as potential response indicators of PAH exposure, but whether the concentration of these metabolites increased or decreased was PAH- and concentration-dependent. These initial findings reveal the potential of metabolomics for monitoring earthworm responses to sub-lethal PAH exposure and highlight the role of metabolomics as a future tool in ecotoxicology.