Abstract: AimTo develop HPLC-UV/ELSD and HPLC-UV/MS³ fingerprints of Japan Jiuxin pill using multi-dimensional hyphenated methods and study on its chemical base. MethodsSeparation was performed on Altima C₁₈ analytical column (250 mm×4.6 mm ID, 5 μm). The mobile phases consisted of water containing 0.2% formic acid and acetonitrile was used as gradient elute. The flow rate was 1.0 mL·min^-1. An electrospray ion trap mass spectrometer was utilized for qualitative analysis and both positive and negative secondary ion scan mode were applied. ResultsHPLC-UV/ELSD and HPLC-UV/MS³ fingerprints of Japan Jiuxin pill with well separation and reproducibility were obtained. Totally 21 components in the complex formula were identified and the chemical structural information of glycocholic acid and its isomer were suggested. ConclusionThe research is propitious to offer an effective and reliable pattern for analyzing Japanese Kampo Medicines as well as its quality control.