Eomecon chionantha Hance (ECH), a traditional Chinese herbal medicine, has been reported for the treatment of traumatic injuries and colitis. The current treatments for ulcerative colitis (TX) are unstable and have side effects, so ECH is potentially useful for treating this condition. However, the active ingredients and pharmacological mechanisms of ECH in treating UC remain unclear. In this study, 21 alkaloids were extracted and purified from the roots of ECH, among which 13 were extracted from this herb for the first time. Cur results showed that 12 ingredients may have effective pharmacological effects after absorption, distribution, metabolism, and excretion (ADME) screening. Network pharmacological analysis revealed that the active ingredients may have positive effects on 19 significant signaling pathways, such as on small cell lung cancer, serotonergic synapses, the IL-17 signaling pathway, Th17 cell differentiation, the estrogen signaling pathway, transcriptional misregulation in cancer, the PI3K-Akt signaling pathway, and others by targeting 23 proteins, including MAPK14, RXRA, GSK3B, CDK2, RXRB, HSP9OAA1, PTGS2, and ESR1. It is of great benefit to use separation, purification, and network pharmacology together to screen active natural products. This study indicated potential anti-UC mechanisms of the active ingredients of ECH and provides theoretical support for the treatment of UC using ECH.