The radionuclide contamination of the environment is an abiotic stress factor that influences biological systems. Plants growing in contaminated areas for many generations provide a unique opportunity to study adaptive strategies aimed at maintaining homeostasis under elevated radiation levels. Using non-targeted metabolomics approaches, we investigated the metabolomic profiles of Achillea millefolium L. plants from the Chernobyl exclusion zone. Amino acid biosynthesis pathways (arginine, glycine, serine, threonine, and proline) and metabolites associated with nitrogen mobilization, cell wall response to injury, photosynthetic efficiency, and defence responses were highly affected in plants from contaminated plots. Our results suggest that these changes may be involved in the adaptive strategies of A. millefolium plant to chronic radiation exposure.