Objective To establish a high choline diet mouse model and investigate the effect of high choline diet on serum amino acid metabolism in mice.Methods Twenty SPF-grade 4-week-old mice were randomly divided into two groups， the control group was given ordinary mouse food and laboratory drinking water， and the high-choline diet group were given 3% choline aqueous solution instead of daily drinking water. Serum was collected after 8 weeks of feeding， and serum amino acids levels were quantified by ultra-performance liquid chromatography tandem triple quadrupole mass spectrometry （UPLC-MS/MS）. Principal component analysis and orthogonal partial least squares-discriminant analysis were used to determine the similarity of amino acid composition between the high choline diet group and the control group.Results Compared with the control group， the levels of cystine， serine， alanine， glutamic acid， phenylalanine， asparagine， threonine， proline， trans-4-hydroxyproline and branched-chain amino acids were significantly higher and the levels of lysine were significantly lower in the high-choline diet group， with isoleucine levels of 34.20±2.71 and 72.59±10.42 μmol/L； the levels of leucine were 68.59±4.06 and 113.04±14.64 μmol/L； the levels of valine were 129.86±10.54 and 161.95±16.54 μmol/L， respectively， and the differences between groups were statistically significant （P<0.05）.Conclusion The high choline diet cause significant changes in serum amino acid metabolism in mice.