An on-line high performance liquid chromatography (HPLC)-diode array detector (DAD)-mass spectrometry (MS) analytical method has been developed to simultaneously separate and identify seventeen main constituents of Chuanxiong (Ligusticum chuanxiong Hort.), a traditional Chinese medicinal herb. In three Chuanxiong samples, nine compounds were unequivocally determined as vanillin (1), ferulic acid (2), senkyunolide I (4), senkyunolide H (5), senkyunolide A (6), coniferyl ferulate (7), Z-ligustilide (8), neocnidilide (9) and 3-butylidenephthalide (10) by comparing their t, UV, and MS data with those obtained for the authentic compounds. Furthermore, additional eight compounds were tentatively identified as senkyunolide J (11), senkyunolide F (12), 3-butylphthalide (13), cnidilide (14), riligustilide (15) or Z,Z′-6,8′,7,3′-diligustilide (16), tokinolide B (17), levistolide A (18) and senkyunolide P (19), based on their MS data and the comparison of their UV spectra with the published references. This is the first report to describe the development of an on-line HPLC-DAD-MS method to simultaneously analyse seventeen main constituents present in Chuanxiong and to construct the chemical profiles of various Chuanxiong samples.