The dried fruit of Evodia rutaecarpa (Juss.) Benth. (ER) is a well-known traditional Chinese medicine and has been used in treatment of headache, abdominal pain, migraine, chill limbs, diarrhea, nausea, hyperbaropathy, dysmenorrheal and postpartum hemorrhage. In this work, a comprehensive characterization of chemical constituents in ER extract is carried out by high performance liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry. The increased power of higher mass resolution and exact mass measurements for precursor and fragment ions lead to detection of 3 acylgluconic acids, 3 flavonol glycosides, 6 limonoids and 24 alkaloids in ER, of which 6 quinolone alkaloids has not been reported previously to our best knowledge. This research enriches our knowledge of chemical constituents of ER and provides a reliable guide for further pharmacology and active mechanism study on this herb.