BuyangHuanwu decoction (BHD) is widely used as a traditional herbal medicine because of its antithrombotic effect, which is attributed to the inhibition of platelet aggregation; however, its active compounds remain unknown. In this study, we developed a method involving platelet binding, solid-phase extraction, and HPLC-MS/MS for screening BHD compounds with potential anti-platelet aggregation properties. Five compounds showing platelet binding affinity were identified as 6-hydroxykaempferol-di-O-glucoside, paeoniflorin, calycosin-7-O-β-d-glucoside, galloylpaeoniflorin, and formononetin-7-O-β-d-glucoside. The results of anti-platelet aggregation experiments in vitro confirmed that these compounds inhibited adenosine diphosphate-induced platelet aggregation. Our results suggest that a platelet binding assay combined with solid-phase extraction and HPLC-MS/MS is an effective method for screening anti-platelet aggregation agents in traditional Chinese medicines.

补阳还五汤具有抗血栓形成作用，可抑制血小板凝集，因此被广泛用作传统草药。然而，其活性化合物仍是未知的。在这项研究中，我们开发了一种涉及血小板结合，固相萃取和HPLC-MS / MS的方法，用于筛选具有潜在抗血小板凝集特性的BHD化合物。五种化合物表示血小板结合亲和性被鉴定为6- hydroxykaempferol二ø葡糖苷，芍药苷，毛蕊-7- ö -β- d葡糖苷，galloylpaeoniflorin，和芒柄花黄素-7- ö -β- d葡糖苷。体外抗血小板凝集实验结果证实这些化合物抑制了二磷酸腺苷诱导的血小板凝集。我们的结果表明，结合固相萃取和HPLC-MS / MS的血小板结合测定是筛选中药中抗血小板凝集剂的有效方法。