Abstract: The fingerprint of Gentiana scabra Bge. was established by electrospray ionization mass spectrometry (ESI-MS). Main secoiridoid glycosides of Gentiana scabra Bge. and their fragmentment pathway were analyzed by ESI-MS. Secoiridoid glycosides were extracted form Gentiana scabra Bge. by methanol supersonics，and extracts were analyzed directly in positive ion mode by ESI-MS. The results of ESI-MS fingerprint were processed by chemical pattern recognition method. The samples form autumn are in good quality. The result also can differentiate samples from different seasons. Secoiridoid glycosides are fragmented through losing H₂O, acetyl and glucosyl in ESI-MS². Electrospray ionization mass spectrometry method is good precision, stability and reproducibility. This method is simple, rapid， and can be used for identification and study of characteristic fingerprint.