目的: 建立当归不同极性部位提取物中5个成分的含量测定方法;探讨中药当归不同极性部位提取物对血瘀证小鼠的干预作用。方法: 采用高效液相色谱(HPLC)法测定当归不同极性部位提取物中5个成分的含量;KM小鼠随机分为空白组、模型组、阳性对照组、当归石油醚组、正丁醇组、水部位组、乙酸乙酯组。实验第1天开始小鼠灌胃给药,空白组与模型组给予等量纯水。结果: 方法学考察各项指标符合规定,绿原酸、阿魏酸、洋川芎内酯Ⅰ、藁本内酯、欧当归内酯A的平均加样回收率分别为99.3%、101.0%、98.5%、97.2%、99.1%,RSD均小于3.0%;当归各实验组中血栓素(TXB2)含量下降,6-酮-前列腺素F1α(6-keto-PGF1α)含量明显升高(P＜0.05);模型组与空白组相比,血清组织型纤溶酶原激活剂(t-PA)含量下降,血清纤溶酶原激活剂抑制物(PAI-1)和环磷酸腺苷(cAMP)含量上升(P＜0.05);当归各组与模型组相比,血清t-PA含量升高,血清PAI-1和cAMP含量降低(P＜0.05);与模型组比较,当归提取物各给药组和阳性对照组,外周血中红细胞(RBC)、血红蛋白(HGB)、红细胞比容(HCT)均明显增加(P＞0.05);与模型组比较,当归各给药组及阳性对照组动物血清中白细胞介素-2(IL-2)、白细胞介素-3(IL-3)、白细胞介素-6(IL-6)和促红细胞生成素(EPO)含量明显增加(P＜0.05);与模型组比较,当归各给药组动物血清中粒细胞集落刺激因子(G-CSF)、巨噬细胞集落刺激因子(M-CSF)、血管细胞粘附因子-1(VCAM-1)含量均明显增加(P＜0.05);与空白组和阳性对照组比较,模型组动物脾脏指数明显增加(P＜0.05);与空白组和阳性对照组比较,模型组动物胸腺指数明显减小(P＜0.05)。结论: 本研究建立的含量测定方法专属性强,重复性好,准确度高,可用于绿原酸、阿魏酸、洋川芎内酯I、藁本内酯、欧当归内酯A的含量测定;当归可明显增加小鼠外周血RBC、PLT的数量,提高HGB含量,增加HCT比值;当归能够不同程度地提高与造血功能有关的细胞因子IL-3、EPO、G-CSF、M-CSF、VCMA-1的表达水平,对机体具有补血的作用;当归可以增加小鼠血清中6-keto-PGF1α的含量和降低血清中TXB2含量,还可以提高血清中t-PA含量、降低PAI-1含量;当归可以维持小鼠血液中TXB2/6-keto-PGF1α的动态平衡,调节血清中t-PA/PAI-1的平衡,纠正小鼠体内平衡失调的作用,当归具有补血作用。
Objective: To establish a method for determining the content of 5 components in extracts of different polar parts of Angelica sinensis (div.) Diels and to investigate the intervention effect of extract of Angelica sinensis (div.) Diels on mice with blood stasis syndrome. Methods: HPLC method was used to determine the contents of 5 components in the extracts of different polar parts of Angelica sinensis (div.) Diels. KM mice were randomly divided into blank group, model group, positive control group, Angelica sinensis (div.) Diels petroleum ether group, n-butanol group, water position group and ethyl acetate group. On the first day of the experiment, the mice in the blank group and the model group were given the same amount of pure water. Results: The methodological inspections met the requirements. The average sample recovery rates of chlorogenic acid, ferulic acid, ligustalide Ⅰ, ligustilide, and angelica lactone were 99.3%, 101.0%, 98.5%, 97.2% and 99.1%, respectively with RSD values less than 3.0%. The TXB2 content decreased in Angelica sinensis (div.) Diels experimental group, and 6-keto-PGF1α content increased significantly (P<0.05). Compared with the control group, the serum t-PA content decreased, and the serum PAI-1 and cAMP content increased (P<0.05). Compared with the model group, the serum t-PA content increased, and the serum PAI-1 and cAMP content decreased (P<0.05). Compared with the model group, RBC, HGB and HCT in peripheral blood of Angelica sinensis (div.) Diels extract were significantly increased in each group and positive control group (P>0.05). Compared with model group, IL-2, IL-3, IL-6 and IL-6 in serum of Angelica sinensis (div.) Diels and positive control group were significantly increased (P>0.05), and the content of EPO was significantly increased (P<0.05). Compared with the model group, the serum levels of G-CSF, M-CSF and VCAM-1 in Angelica sinensis (div.) Diels group were significantly increased (P<0.05). Compared with the blank group and the positive control group, the spleen index of the model group was significantly increased (P<0.05), and the thymus index of the model group was significantly decreased compared with the control group and the empty white group and the positive control group (P<0.05). Conclusion: The content determination method established in this study is specific, repeatable and accurat, and can be used for the content determination of chlorogenic acid, ferulic acid, ligustalide Ⅰ, ligustilide, and angelica lactone. Angelica sinensis (div.) Diels can significantly increase the number of RBC and PLT, HGB content and HCT ratio in peripheral blood of mice. Angelica sinensis (div.) Diels can improve the expression levels of IL-3, EPO, G-CSF, M-CSF and VCMA-1, which are related to hematopoietic function in varying degrees, and has the effect of enriching blood. Angelica sinensis (div.) Diels can increase the content of 6-keto-PGF1α in serum and reduce the content of TXB2 in serum of mice, and can also improve the blood circulation Angelica sinensis (div.) Diels can maintain the dynamic balance of TXB2/6-keto-PGF1α in the blood of mice, regulate the balance of t-PA/PAI-1 in the serum, and correct the imbalance in the body of mice. Angelica sinensis (div.) Diels has the effect of nourishing blood.
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