赤芍抗凝血活性成分的筛选  

Screening of anticoagulant active components of Paeonia lactiflora

作　　者：黄晨存 严晟 赵元 黄诗怡 袁志鹰[1] 许光明[1] HUANG Chen-cun;YAN Sheng;ZHAO Yuan;HUANG Shi-yi;YUAN Zhi-ying;XU Guang-ming(College of Pharmacy,Hunan University of Chinese Medicine,Changsha 410208,China)

机构地区：[1]湖南中医药大学药学院,湖南长沙410208

出　　处：《中国药物化学杂志》2023年第3期181-190,共10页Chinese Journal of Medicinal Chemistry

基　　金：湖南省自然科学基金项目(2021JJ30511);湖南省中医药科研项目(2021062)。

摘　　要：目的建立超滤质谱快速筛选赤芍活性成分的方法,从中得到可能的潜在抗凝血活性成分。方法对赤芍不同极性部位成分进行鉴定,以凝血酶(thrombin,THR)为靶标,亲和超滤法逆靶向筛选赤芍中的凝血酶抑制剂,并对得到的活性成分进行分析、鉴定;采用生色底物法及体外抗凝法对其进行活性验证,通过分子对接对其结合能力进行初步分析。结果经对赤芍不同极性部位成分进行定性分析,检测出17个成分,并通过亲和超滤质谱法筛选得到一个可能的抗凝血活性成分——鞣花酸。鞣花酸浓度为1 g·L^(-1)时对凝血酶百分抑制率可达98.34%;体外凝血实验发现鞣花酸凝血酶时间(thrombin time,TT)与空白组相比有显著性差别(P<0.01),表明其为潜在的凝血酶直接抑制剂;分子对接结果显示鞣花酸与凝血酶活性结合位点多、结合能低,具有良好的结合效果。结论建立了一种快速筛选赤芍中抗凝血活性成分的方法,筛选结果表明鞣花酸可能为潜在的凝血酶抑制剂,为中药的靶向治疗、机制探索和质量控制提供了有价值的参考。To establish an affinity ultrafiltration mass spectrometry method for rapid screening of active components of Paeonia lactiflora,and to obtain it's anticoagulant active components,the components of different polar parts of P.lactiflora were identified,and thrombin inhibitors in P.lactiflora were inversetargeted screened by affinity ultrafiltration with thrombin(THR)as the target,and the obtained active components were analyzed and identified.Its activity was tested by chromogenic substrate method and in vitro anticoagulant method,and its binding ability was preliminarily analyzed by molecular docking.In this experiment,the active components in different polar parts of P.lactiflora were qualitatively analyzed,and 17 components were detected.An anticoagulant active component,ellagic acid,was screened by affinity ultrafiltration mass spectrometry.When the concentration of ellagic acid was 1 g·L^(-1),the inhibition rate of thrombin was 98.34%.In vitro coagulation experiments showed that the thrombin time(TT)of ellagic acid was significantly different from the blank group(P<0.O1),indicating that it was a potential direct thrombin inhibitor.Molecular docking results showed that ellagic acid had many binding sites with thrombin activity,low binding energy,and good binding effect.The screening results showed that ellagic acid might be a potential thrombin inhibitor.At the same time,a rapid screening method for anticoagulant active ingredients in P.lactiflora was established,which provided valuable reference for targeted therapy,mechanism exploration and quality control of traditional Chinese medicine.

关 键 词：亲和超滤质谱 赤芍 凝血酶 活性筛选 分子对接 鞣花酸 

分 类 号：R284[医药卫生—中药学]