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Rapid screening and identification of anticoagulation component from carthami flos by two‐dimensional thrombin affinity chromatography combined with HPLC‐MS/MS
Author(s) -
Qiao Yanru,
Shi Yingdi,
Wu Chen,
Hou Xiaofang,
Pan Xiaoyan,
Deng Zijie,
Wang Sicen
Publication year - 2021
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.202100092
Subject(s) - flos , chemistry , thrombin , chromatography , thrombin time , high performance liquid chromatography , coagulation , partial thromboplastin time , biochemistry , platelet , medicine , rutin , psychiatry , immunology , antioxidant
Carthami flos, commonly known as Honghua in China, is the dried floret of safflower and widely acknowledged as a blood stasis promoting herb. The study aimed at investigating the relationship between thrombin and carthami flos through a high‐performance thrombin affinity chromatography combined with a high‐performance liquid chromatography‐tandem mass spectrometry system. First, thrombin was immobilized on the glutaraldehyde‐modified amino silica gel to prepare the thrombin affinity stationary phase, which was packed into a small column (1.0 × 2.0 mm, id) for recognizing the anticoagulant active components of carthami flos. The target component was enriched and analyzed by the high‐performance liquid chromatography‐tandem mass spectrometry system. Finally, hydroxysafflor yellow A was screened out and identified as the active component. The anticoagulant effects of hydroxysafflor yellow A were analyzed by anticoagulant experiments in vitro, and the interaction of hydroxysafflor yellow A with thrombin was investigated by the molecular docking method. The results proved that hydroxysafflor yellow A (30 μg/mL, 0.05 mM) and carthami flos extract (30 μg/mL) could prolong activated partial thrombin time and thrombin time by 50 and 11%, respectively. Moreover, hydroxysafflor yellow A exhibits a good hydrogen bond field and stereo field matching with thrombin. Overall, it was concluded that hydroxysafflor yellow A might exert an anticoagulation effect by interacting with thrombin and thus could be potential anticoagulant drugs for the prevention and treatment of venous thrombosis.