BACKGROUND TongMai Keli (TM) is a widely used traditional Chinese medicine preparation for the treatment of cardiovascular and cerebrovascular diseases. It is composed of Puerariae Lobatae Radix (roots of Pueraria lobata (Willd.) Ohwi), Salviae Miltiorrhizae Radix (roots of Salvia miltiorrhiza Bge.), and Chuanxiong Rhizoma (rhizomes of Ligusticum chuanxiong Hort.). The aim of this study is to identify the in vivo metabolites of TM, and to elucidate the pharmacokinetics of TM constituents and their metabolites. METHODS For metabolites identification, TM was orally administered to rats (n=3), and the metabolites in plasma were identified by UHPLC/DAD/qTOF-MS analysis and β-glucuronidase hydrolysis. For pharmacokinetic study, rats (n=10) were treated with TM at a clinical dose, and the plasma was analyzed by LC/MS/MS. RESULTS A total of 25 metabolites from TM were identified in rats plasma. Glucuronide and sulfate conjugations were the major metabolic reactions, and produced 14 metabolites. The analytical method for pharmacokinetic study was fully validated with good linearity (r>0.99), wide dynamic ranges (6-6000 ng/mL), and low variations (<14.3%). The plasma concentration-time curves of puerarin and nine metabolites were profiled. CONCLUSIONS Isoflavones from Puerariae Lobatae Radix were the major metabolites in rat plasma after oral administration of TM. Puerarin and other isoflavone glycosides could reach their first C(max) within 30 min, and were then rapidly eliminated, followed by their phase II metabolites.