Most plants exhibit strong tolerance to excess molybdenum (Mo). However, the metabolic profile and tolerance mechanisms of plants in response to excess Mo remain unknown. We comprehensively analyzed changes in the metabolic profiles of leaves and roots in soybean (Glycine max L.) seedlings cultured under normal-Mo and excess-Mo conditions by using ultra performance liquid chromatography (UPLC) combined with MS/MS (mass spectrometry). There were 42 differential metabolites in the roots and 19 differential metabolites in the leaves in response to excess Mo stress. In roots, the organic acids, levels of gluconic acid, D-glucarate and citric acid increased by 107.63-, 4.42- and 2.87-folds after excess Mo exposure. Several hormones (salicylic acid, jasmonic acid) and lipids (PG, MG, DG etc) also increased significantly under excess Mo condition. Metabolites related to ascorbate-glutathione metabolism and flavonoid and isoflavone biosynthesis notably accumulated in roots. Only lipid metabolism and salicylic acid accumulation were induced in leaves under excess Mo stress. It is speculated that organic compounds such as 2-oxoarginine, L-nicotine, gluconic acid, D-glucurate, and citric acid played important roles to chelate Mo and reduce its toxicity. Signaling molecules (JA, SA, and some lipids) and non-enzyme antioxidants such as flavonoids/isoflavones act synergistically to detoxify ROS and contribute to Mo tolerance in soybean seedlings. More metabolic pathways were induced by Mo excess in roots than in leaves, suggesting that roots play more implant role in Mo tolerance.