As an important grain production base and ecological barrier zone in China, Ulansuhai Basin provides a variety of important ecosystem services and ensures human well-being, and it is essential to maintain the sustainable development of the regional ecology–economy–society. Therefore, in order to explore the trade-offs and synergies between ecosystem services in Ulansuhai Basin, we first evaluated the spatio-temporal characteristics of five ecosystem services in 2000, 2005, 2010, 2015, and 2018 based on the InVEST model, including soil conservation, carbon storage, water production, water purification, and food supply. We then further analyzed the trade-offs and synergies of ecosystem services in Ulansuhai Basin and in different functional areas through using the Spearman correlation coefficient. The results show that different ecosystem services had obvious regional differences due to different land-use types in Ulansuhai Basin. Soil conservation, carbon storage, and water production were higher in the eastern region and lower in the central and western regions, while water purification and food supply were higher in the central region and lower in the eastern and western regions. Ecosystem services showed an overall increasing trend from 2000 to 2018. Moreover, trade-off was the dominant relationship between different ecosystem services, and trade-offs and synergies showed strengthening trends to a certain extent. The trade-offs and synergies of ecosystem services in different functional areas were obviously different. Our study aimed to clarify the trade-offs and synergies between ecosystem services and to propose ecological protection and management countermeasures and suggestions, which can provide decision-making reference for regional ecological protection and management.