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1.山东中医药大学,济南 250355
2.山东中医药大学 第二附属医院,济南 250001
刘珊,博士,从事中药药性理论与临床应用研究、中药及复方药理研究,E-mail:liushan5463@163.com
王永春,硕士,主管技师,从事康复治疗技术理论与应用研究,E-mail:wyc19862012@163.com; *
徐凯勇,硕士,高级实验师,从事中药干预病证动物模型机制研究,E-mail:xkychina@163.com
收稿日期:2022-11-30,
网络出版日期:2023-02-24,
纸质出版日期:2023-05-05
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刘珊,王文越,王永春等.黄芪-当归药对益气活血作用的药效物质基础[J].中国实验方剂学杂志,2023,29(09):28-36.
LIU Shan,WANG Wenyue,WANG Yongchun,et al.Pharmacodynamic Material Basis of Astragali Radix and Angelicae Sinensis Radix Drug Pair in Supplementing Qi and Activating Blood Circulation[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(09):28-36.
刘珊,王文越,王永春等.黄芪-当归药对益气活血作用的药效物质基础[J].中国实验方剂学杂志,2023,29(09):28-36. DOI: 10.13422/j.cnki.syfjx.20230412.
LIU Shan,WANG Wenyue,WANG Yongchun,et al.Pharmacodynamic Material Basis of Astragali Radix and Angelicae Sinensis Radix Drug Pair in Supplementing Qi and Activating Blood Circulation[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(09):28-36. DOI: 10.13422/j.cnki.syfjx.20230412.
目的
2
探讨黄芪-当归药对(3∶1)对气虚血瘀模型大鼠的益气活血作用,从特征图谱、药效物质辨识、入血成分对比三方面入手,为临床合理用药及复方药效物质辨识与质量控制提供理论依据。
方法
2
使用高效液相色谱法(HPLC)建立黄芪-当归(3∶1)指纹图谱,采用超高效液相色谱-四级杆-静电场轨道阱联用技术(UPLC-Q-Exactive Orbitrap-MS)分析水煎液成分;选用SPF级成年雄性Wistar大鼠,随机分为空白组、模型组、黄芪-当归3∶1组及黄芪-当归5∶1组,每日通过控制食量及凉水游泳制备气虚血瘀模型,同时各组每天给药(水)1次,给药组剂量10.2 g∙kg
-1
,模型组和空白组给予等量的蒸馏水,连续15 d。观察大鼠一般状况;记录各组大鼠体质量,全血及血浆黏度,胸腺指数和脾脏指数,血清中三磷酸腺苷(ATP)、二磷酸腺苷(ADP)、大鼠血管性血友病因子(vWF)水平及ATP/ADP值;苏木素-伊红(HE)及扫描电镜观察血管内皮形态;UPLC-Q-Exactive Orbitrap-MS分析血清中的原型及代谢成分。
结果
2
建立了黄芪-当归(3∶1)指纹图谱;UPLC-Q-Exactive Orbitrap-MS共鉴定出体外化学成分49个,初步鉴定了11种原型成分吸收入血。与空白组比较,模型组大鼠体质量明显降低(
P
<
0.01);全血(高切、中切、低切)黏度及血浆黏度均明显升高(
P
<
0.05,
P
<
0.01);胸腺指数和脾脏指数均明显下降(
P
<
0.05,
P
<
0.01);血清ATP含量显著降低(
P
<
0.01),ADP含量显著升高(
P
<
0.01),ATP/ADP值显著降低(
P
<
0.01),vWF含量显著升高(
P
<
0.01);苏木素-伊红(HE)及扫描电镜结果显示,血管部分受损,表现为内膜结构紊乱,内皮凸起、缺损、粗糙,细胞黏附迁移明显。与模型组比较,黄芪-当归给药组体质量均明显增加(
P
<
0.01);全血及血浆黏度结果显示黄芪-当归3∶1组可明显降低全血低切黏度(
P
<
0.05);胸腺指数及脾脏指数结果显示黄芪-当归5∶1组能明显升高大鼠胸腺指数(
P
<
0.05);血清ATP、ADP水平结果显示黄芪-当归5∶1组对ATP、ADP水平作用更明显(
P
<
0.05),两组均能显著降低ATP/ADP值(
P
<
0.01);血清vWF水平结果显示黄芪-当归3∶1组vWF含量降低更明显(
P
<
0.05);HE及扫描电镜结果显示给药组对血管内皮损伤均有不同程度的改善,内膜结构较整齐。
结论
2
黄芪-当归3∶1及黄芪-当归5∶1组均对气虚血瘀模型大鼠宏观及微观指标有改善作用,但传统配比黄芪-当归5∶1益气效果更好,而黄芪-当归3∶1活血作用更好。
Objective
2
To evaluate the effect of Astragali Radix (AR)-Angelicae Sinensis Radix (ASR) drug pair on supplementing Qi and activating blood circulation in rats with Qi deficiency and blood stasis and provide a theoretical basis for clinical rational medication and identification and quality control of compound pharmacodynamic substances from the three aspects of characteristic map, identification of pharmacodynamic substances, and comparison of blood components.
Method
2
High-performance liquid chromatography (HPLC) was employed to establish the fingerprint of AR∶ASR (3∶1), and ultra-high performance liquid chromatography-Q-Exactive Orbitrap-mass spectrometry (UPLC-Q-Exactive Orbitrap-MS) was employed to analyze the ingredients of the decoction. Adult male Wistar rats with SPF grades were selected and randomly divided into a blank group, a model group, a 3∶1 group, and a 5∶1 group. The rat model of Qi deficiency and blood stasis syndrome was prepared by controlling food intake and swimming in cold water every day. In parallel, each group was given medicine (or water) once a day. The dose of drug groups was 10.2 g∙kg
-1
, and the model group and blank group were given the same amount of distilled water for 15 d. Animal behavior, body weight, whole blood and plasma viscosity, thymus index, spleen index, the levels of adenosine triphosphate (ATP), adenosine diphosphate(ADP), von willebrand factor (vWF), and ATP/ADP value in serum of rats were recorded. The morphology of vascular endothelium was observed by hematoxylin-eosin (HE) staining and scanning electron microscopy. UPLC-Q-Exactive Orbitrap-MS was used to analyze prototype and metabolic components in serum.
Result
2
The fingerprint of AR-ASR drug pair (AR-ASR 3∶1) was established. UPLC-Q-Exactive Orbitrap MS identified 49 chemical components
in vitro
and preliminarily identified 11 prototype components absorbed into blood
in vivo
. As compared with the blank group, the body mass decreased significantly (
P
<
0.01), the whole blood (high shear, middle shear, and low shear) viscosity and plasma viscosity were significantly increased (
P
<
0.05,
P
<
0.01), the thymus index and spleen index decreased significantly (
P
<
0.05,
P
<
0.01), serum ATP content decreased significantly (
P
<
0.01), ADP content increased significantly (
P
<
0.01), ATP/ADP value decreased significantly (
P
<
0.01), and vWF content increased significantly (
P
<
0.01). The results of HE staining and scanning electron microscopy showed that the vessels were partially damaged, showing the structural disorder of the intima, the bulge, defect, and roughness of the endothelium, and the obvious cell adhesion and migration in the model group. As compared with the model group, the body mass also increased significantly (
P
<
0.01). The results of whole blood and plasma viscosity showed that the whole blood low shear viscosity was significantly decreased in the 3∶1 group (
P
<
0.05). The results of thymus index and spleen index showed that 5∶1 group significantly increased the thymus index of rats (
P
<
0.05). The results of serum ATP and ADP levels showed that the 5∶1 group had more significant effects on ATP and ADP levels (
P
<
0.05), and both groups significantly reduced ATP/ADP values (
P
<
0.01). The results of serum vWF level showed that the vWF content in the 3∶1 group decreased significantly (
P
<
0.05). The results of HE staining and scanning electronic microscopy showed that the damage of vascular endothelium was improved in the treatment group and the structure of intima was neat.
Conclusion
2
AR-ASR drug pair can improve the macro and micro indexes of rats with qi deficiency and blood stasis in the 3∶1 and 5∶1 groups. Overall, the 5∶1 ratio has a better effect on supplementing Qi but 3∶1 ratio has a better effect on promoting blood circulation.
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