Serum metabolomic method was used to investigated the anti-diabetic effects and mechanism of Ginseng berry (GB) on high-fat diet combined streptozotocin induced type II diabetes mellitus (T2DM) rats based on ultra high performance liquid chromatography coupled with quadrupole Exactive orbitrap mass spectrometry (UHPLC-Q-Exactive Orbitrap/MS). Serum samples from control group, T2DM group, metformin treatment group, and GB ginsenoside treatment group rats were collected after intervention. The biochemical parameters of serum were firstly analyzed. Then metabolomic studies based on UHPLC-Q-Exactive Orbitrap/MS and multivariate statistical analysis were performed for the pattern recognition and characteristic metabolites identification. The differential metabolites were analyzed by KEGG metabolic pathway to study the potential mechanism. The treatment of GB ginsenoside significantly reduced the blood glucose level, increased the content of serum SOD, and reduced the content of malondialdehyde. Respectively 16, 9, and 24 differential metabolites were found and identified in T2DM compared to control group, metformin compared to T2DM group and GB compared to T2DM group. Metabolic pathways analysis indicated that GB ginsenoside regulated bile acid metabolism, arachidonic acid metabolism, glucuronization to play a role in the treatment of T2DM. This study verified the anti-diabetic and anti-oxidation effects of ginseng berry, elaborated that GB regulated the secretion of bile acids, activated GLP-1 pathway, increased the secretion of insulin, promoted the hydrolysis of fat and triglyceride, inhibited the activity of 5α - reductase, reduced weight and insulin resistance, so as to improved and treated T2DM, and laid the foundation for the further development and utilization.