Abstract: The purpose of this study is proposing a methodology to determine the ecological base flow of a river. In the past in Taiwan, the ecological base flow was calculated according to the basin area, and the influence of the water supply was not considered. This study proposes a methodology combining the water distribution optimization model with the Riverine Habitat Simulation System model to demonstrate the impact on the aquatic habitat for indicator species with different ecological base flow settings. The most suitable value of the ecological base flow between the indicators of the Shortage Index and the average value of Percentage Usable Area is determined using trade-off analysis and the incremental method. The result shows for the indicator species in the study area, Tou-Qian River Basin in Northern Taiwan, the most appropriate ecological base flow is 3.75 m3/s and it’s the optimal use of the ecological base flow between the water supply and the ecology.