Trollius ledebourii has been more involved in Mongolian medicine and is often used as a type of tea for heat-clearing and detoxifying in the populus. In this study, a rapid and sensitive method was established for the qualitative and quantitative analyses of the major constituents in T. ledebourii. Ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry was developed for the identification of the multi-constituents in T. ledebourii. A total of 37 chemical constituents in T. ledebourii extract were unambiguously or tentatively identified, including 17 flavonoid glycosides, 6 flavones, 3 flavonols, 1 dihydroflavone, 8 phenolic acids, 1 amide and 1 triterpene. Pectolinarin, naringenin, isorhamnetin, diosmetin, protocatechuic acid, paeonol, caffeic acid and ferulic acid were first detected in T. ledebourii and the buttercup family. High-performance liquid chromatography-quadrupole ion trap tandem mass spectrometry was applied for the simultaneous determination of 11 compounds, which were either with high contents or strong bioactivities. Satisfactory linearity was achieved with a wide linear range and fine determination coefficient (r > 0.9987). The overall recoveries ranged from 98.07 to 101.2%, and the precision in terms of RSD was <0.74%. The results might provide the basis for quality control analysis of T. ledebourii.