The effects of ionizing radiation to human health are of great concern in the field of space exploration and for patients considering radiotherapy. However, to date, the effect of high-dose radiation on metabolism in the liver has not been clearly defined. In this study, ¹H nuclear magnetic resonance (NMR)-based metabolomics combined with multivariate data analysis was applied to study the changes of metabolism in the liver of C57BL/6 mouse after whole-body gamma (3.0 and 7.8 Gy) or proton (3.0 Gy) irradiation. Principal component analysis (PCA) and orthogonal projection to latent structures analysis (OPLS) were used for classification and identification of potential biomarkers associated with exposure to gamma and proton radiation. The results show that the radiation exposed groups can be well separated from the control group. Where the same dose was received, the proton exposed group was nevertheless well separated from the gamma-exposed group, indicating that different radiation sources induce different alterations in the metabolic profile. Common among all high-dose gamma and proton exposed groups were the statistically decreased concentrations of choline, O-phosphocholine and trimethylamine N-oxide, while the concentrations of glutamine, glutathione, malate, creatinine, phosphate, betaine and 4-hydroxyphenylacetate were statistically and significantly elevated. Since these altered metabolites are associated with multiple biological pathways, the results suggest that radiation induces abnormality in multiple biological pathways. In particular, metabolites such as 4-hydroxyphenylacetate, betaine, glutamine, choline and trimethylamine N-oxide may be prediagnostic biomarkers candidates for ionizing exposure of the liver.