Sodium Tanshinone IIA Sulfonate (STS) is a water-soluble derivative of tanshinone IIA, an important lipophilic component isolated from the roots of Salvia miltiorrhiza. STS, as injection drug, is widely and successfully used in China for treating cardiovascular disease. Eight impurities were detected in STS bulk drug by LC-UV and LC-ESI-MS(n). Low pressure C(18) column chromatography and preparative HPLC were used for the isolation of three principal impurities. Based on LC/ESI-MS(n) and NMR analysis, related impurities were characterized as sodium (±)-hydroxyl tanshinone sulfonate (1), sodium tanshinone I sulfonate (2), sodium salvianolic acid sulfonate (3), sodium 1,2-dehydrotanshinone IIA sulfonate (4), sodium tanshinone IIB sulfonate (5), sodium 3-hydroxyl salvianolic acid sulfonate (6), sodium 1,16-dihydroxyl-15,16-dihydrotanshinone IIA sulfonate (7), and sodium methyl 1,2-dehydrotanshinonate sulfonate (8). The (1)H and (13)C NMR data of impurity 1 were reported in this paper for the first time. Also, to the best of our knowledge, impurity 4 had not been isolated as pure substance until now. The possible mechanism for the formation of impurities is also discussed.