Flavonoids are the main bioactive components of Radix Scutellariae, a well-known traditional Chinese medicine. After oral administration of Radix Scutellariae extract （RSE）, the bile, plasma and urine of rats were analyzed to identify the absorbed parent components and metabolites by ultra-fast liquid chromatography coupled with ion trap time-of-flight mass spectrometry （UFLC-IT-TOF/MS）. A total of 36 different flavonoids, including 13 new metabolites, were identified or tentatively characterized in bile, plasma and urine of rats based on their fragmentation patterns. Among them, 16 flavonoids were identified in RSE in vitro, 25 flavonoids in rat bile, 15 flavonoids in rat plasma, and 14 flavonoids in rat urine. The results indicated that glucuronidation, sulfation and methylation were the major metabolic pathways of flavonoids. It is the first time that the metabolism of RSE was studied comprehensively.
