Metabolomics, the exponentially developing technique, could provide a systemic mapping in toxicology by directly measuring small molecular metabolites. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) was found to be neurotoxic in mammalian animals. In this study, we employed liquid chromatography/quadrupole time-of-flight mass spectrometry for non-targeted analysis of metabolic profiling in hippocampal sample sets of the rats exposed to TCDD. Hippocampal metabolome from different ages of the healthy rats (4-week, 12-week and 20-week) was also deciphered. The relationship between the two tested cases was unlocked to delineate TCDD toxicity associated with ageing. Tandem mass spectrometry fragmentation in conjunction with metabolic database searching and compared to authentic standards was utilized for metabolite identification. As a consequence, the reduced levels of phenylalanine and leucine/isoleucine as well as the up-regulation of inosine and hypoxanthine were highlighted for understanding of TCDD toxicity related to age in rats and the trajectory was depicted by principal components analysis.