The water conservation zone of the Yellow River (including the source region of the Yellow River, the Weihe River Watershed controlled by the Huaxian Station, and the Yiluo River Watershed) is the main runoff producing area. Identifying the comprehensive change characteristics and spatial differences of flow regime can provide an important scientific basis for analyzing the water cycle evolutions and their causes in the Yellow River Basin under the changing environment. Current research focused on the influence of environmental change on runoff magnitude in the Yellow River Basin, but cannot comprehensively explain the change characteristics and spatial differences of flow regimes. In this study, 31 source small watersheds in the water conservation zone of the Yellow River were taken as the research areas. The characteristics of flow regimes, including the magnitude, rate of change, frequency, duration, and timing metrics, were comprehensively considered. The Theil-Sen trend-free pre-whitening with Mann-Kendall (TFPW-MK) trend analysis and Sen's slope estimator methods were used to detect the variation of flow regime metrics in all the watersheds from 2000 to 2020. The hierarchical clustering method was used to identify the main patterns of flow regime changes and spatial distribution characteristics. The results show that: 1) At most watersheds, significant increases in all magnitude metrics, rate of change in high flow, and low flow frequency were detected, while rate of change in low flow, high flow frequency, and all duration and timing metrics showed a significant decrease. 2) The typical flow regimes found in the Huangshui River, the Tao River, downstream of the Weihe River, and the Luohe River were more sensitive to climate change, mainly showed significant increase in high flow magnitude but decrease in timing, and significant increase in low flow frequency but decrease in duration and timing metrics. 3) The variation of flow regime characteristics was divided into three patterns. The first pattern accounted for 42%, located in the Taohe River, the Huangshui River, the trunk and tributaries of the Weihe River, and the main stream of the Yi River, with significant increases in magnitude, frequency, and rate of change, and significant decreases in duration and timing. The second pattern accounted for 35%, located in the upstream and downstream of the three major watershed divisions, with metrics of low flow increased significantly except for a decrease in rate of change; while the five type metrics in high flow all decreased significantly. The third pattern accounted for 23%, located in the middle and lower reaches of the three watershed divisions, with significant increases in magnitude and frequency, and significant decreases in rate of change, duration, and timing.