酸枣仁汤组分配伍的抗焦虑作用及机理

【作者】 王守勇；

【导师】 谢鸣；

【作者基本信息】 北京中医药大学 ， 中医方剂学， 2003， 硕士

【摘要】 酸枣仁汤（SZRT）源于《金匮要略》，主治肝血不足、虚热内扰所致之虚烦证。现代临证除治疗失眠症外，还用于包括焦虑症在内的以情绪或神志障碍为主要表现的精神系统疾病。本文在前期工作证实SZRT水煎液具有确切的抗焦虑作用基础上，通过对水煎液中大体部位/组分的分离与重组，观察了不同组分配伍与抗焦虑作用的关系，并对酸枣仁汤组分有效配方的抗焦虑作用机理进行初步探讨。论文包括综述和实验研究两部分。综述包括了中药复方化学的研究进展和酸枣仁汤临床、药理及化学的研究现状二部分。实验研究包括组分分离、组分配伍的药效观察、有效组分的作用机理探讨三个部分。实验一：采用系统分离方法，从SZRT水煎液分得六个组分，并对各组分进行了定性检识和得率计算。各组分所含主要成分及得率如下：A（挥发油）、B（多糖，11.49%）、C（乙醚部分、含皂苷元及黄酮，0.15%）、D（皂苷及苷元，13.31%）、E（黄酮，0.22%）、F（不含上述组分的剩余部分，0.065%）。实验二：对分得的六个组分按L8（27）正交表配方，得成分群配方SZRT1-SZRT8。中药各组分别按20g/kg（昆明小鼠）和７.5g/kg（Wistar大鼠）灌胃，安定组按2mg/kg（昆明小鼠）和１mg/kg（Wistar大鼠）灌胃，给药10天后，进行高架十字迷宫（EPM）模型行为学测试。结果发现，小鼠EPM模型中，SZRT1（A＋B＋C＋D＋E＋F）、SZRT6（B+E）组小鼠进入开放臂次数比（OE%）和在开放臂停留时间比（OT%）均显著高于对照组（p<0.05），SZRT3、SZRT5组小鼠进入开放臂次数比显著高于对照组（p<0.05），但各组小鼠开放臂和封闭臂总的进入次数（Total arm entries）与对照组相比无显著差异（p>0.05）；大鼠EPM中，SZRT1、SZRT6组大鼠进入开放臂次数和在开放臂停留时间比均显著高于对照组（p<0.05），且大鼠在开放臂中Head-dipping次数显著高于对照组（p<0.05），但大鼠开放臂和封闭臂总的进入次数与对照组相比无显著差异（p>0.05）。在两个模型中，SZRT1、SZRT6的效用与安定一致；SZRT6与SZRT1组间各指标间均无显著性差异。结果表明：SZRT1和SZRT6有一定的抗焦虑作用，且与镇静催眠作用无关；在实验设定的SZRT不同成分群配方中，SZRT1和SZRT6与抗焦虑作用具有相关性；除了含原方全部组分的配方外，含B和E的SZRT6配方与抗焦虑关系最为密切。有关各成分群对OE%、OT%的影响的分析提示，组分C仅对OE%有显著影响（负影响），组分E仅对OT%有显著影响（正影响）;成分群B、E在抗焦虑效应方面有一定的协同作用。实验三：采用高效液相－电化学法和放免法检测上述实验中的动物中枢神经肽、单胺类递质以及血清细胞因子。神经肽的检测发现，较之对照组，SZRT1和SZRT6组小鼠全脑β－内啡肽含量显著升高（p<0.05）；SZRT6组大鼠下丘脑神经肽Y含量显著降低（p<0.05），SZRT1组亦降低；而杏仁核神经肽Y含量显著升高（p<0.05），与安<WP=6>定组一致。结果表明SZRT1和SZRT6可能有促进下丘脑分泌β－内啡肽，以及促进下丘脑神经肽Y转运至杏仁核的作用。单胺类递质检测发现， SZRT6组大鼠海马5－HT含量显著降低（p<0.05）, SZRT1组亦降低，其作用效果与安定一致，分析SZRT1和SZRT6可能有降低大鼠海马5－HT的释放作用。细胞因子检测发现，SZRT1和SZRT6组均使模型大鼠血清肿瘤坏死因子（TNFα）含量显著升高（p<0.05），SZRT6组白细胞介素2（IL-2）含量显著降低（p<0.05）。结果表明SZRT1和SZRT6对焦虑免疫功能具有一定调节作用。研究的初步结论:酸枣仁汤各组分与其抗焦虑效用具有一定的相关性，SZRT组分配方SZRT6的抗焦虑效应与含括SZRT水煎液全部成分的SZRT1相当，SZRT6主要为SZRT中的多糖和黄酮类成分，是该方抗焦虑作用的物质基础。酸枣仁汤全方和其组分有效配方抗焦虑效用可能与其对神经系统下丘脑β－内啡肽、中枢神经肽Y和以及海马5－HT的调节作用有关；还可能对免疫系统具有调节作用。提示SZRT及其组分配方通过多层次、多系统及多靶点产生效用，对于焦虑症以及伴随的免疫失调状态的防治具有优势。本研究为认识酸枣仁汤临床治疗焦虑症提供了现代科学的理解，为从化学组分配伍的角度认识中医方剂效用物质基础提供一个有用的研究模式，为抗焦虑中药复方新药的进一步研发奠定了良好的药学基础。更多还原

【Abstract】 Suanzaoren Tang(SZRT) is derived from "Synopsis of Prescriptions of the Golden Chamber", the indications is agitation of deficiency type result from insufficient liver blood, deficiency-fire stirring up inside. In modern clinic practice, it can treat many psychological disorders including anxiety as well as insomnia. On the basis of proving that SZRT decoction can treat anxiety, the relationship between different component compatibilities and their effects of treating anxiety was reviewed through isolating and recombining the main components in the decoction, meanwhile, the anxiolytic effect of effective components compatibilities was discussed elementarily .This paper includes summarization and experiment study two parts. The summarization includes study progress about compound prescriptions of traditional Chinese medicine and study status in quo of clinic, pharmacology and chemistry about SZRT. The experiment includes isolation of components, drug effect observation of component compatibilities, action mechanism of effective components.Experiment1: 6 component groups were isolated from SZRT decoction by means of system isolation, and the nature determination and obtain percent were made. The obtain percent is as follows:A(volatile oil),B(polysaccharide,11.49%), C(diethyl ether, sapogenin and flavone, 0.15%), D(saponin and sapogenin, 13.31%), E(flavone, 0.22%),F(the rest part, 0.065%).Experiment2:6 components were formulated according to orthogonal meter and got SZRT1-SZRT8. Kunming mice were administrated orally by herb medicine(20/kg/day) and Wister rats were administrated orally by herb medicine (7.5/kg/day), it’s 2mg/kg/day and 1mg/kg/day to Diazepam group, the elevated plus-muse(EPM) test was performed after 10 days of administration.The results were: percentage of open arm entries (OE%) and of times spent in the open arms (OT%) of SZRT1（A＋B＋C＋D＋E＋F）and SZRT6（B+E）mice were both higher than that of control group（p<0.05）. In the mouse EPM experiment, OE% of SZRT3 and SZRT5 groups were higher than that of control group（P<0.05）, but there was no significant difference between every group and control group about total number entries (P>0.05). In the<WP=8>rat EPM rats were higher than that of control group(P<0.05),but the total arm enteries had no significant difference compared with that of the control group(P>0.05). In two experiments, the effects of SZRT1 and SZRT6 were similar to Diazepam; there was no significant difference about the index between SZRT6 and SZRT1 group. The results showed: SZRT6 and SZRT1 had the effects of anti-anxiety, and it was not related to sedation and hypnosis effects; SZRT6(B+E) had close relationship with anti-anxiety effect besides the recipe including all components. The effects of all different components of SZRT on OE%、OT% showed components C only effected OE% significantly(minus),components E only effected OT% significantly(positive). Components B and E had cooperation effect in the respect of anti-anxiety.Experiment3: Central neuropeptide, monoamine transmitter and serum cytokine were measured by means of high performance liquid chromatography coupled with electrochemical detection (HPLC-ECD) and radioimmunoassay. The test of neuropeptide showed that the β-endorphin in SZRT1 and SZRT6 group mice increased significantly(P<0.05); Neuropeptide Y concentration in SZRT6 group rat hypothalamus decreased significantly(P<0.05), so did that in SZRT1. Neuropeptide Y in Nucleus amygdalae increased significantly(P<0.05), the same to diazepam group. The results showed SZRT1 and SZRT6 might accelerate the excretion of β-endorphin in hypothalamus and accelerate transport neuropeptide Y in hypothalamus to Nucleus amygdalae.The test of monoamine transmitter showed that 5-HT concentration in SZRT6 group rats’ hippocampus decreased significantly(P<0.05), SZRT1 group decreased without significant, the effects were similar to diazepam group, SZRT1 and SZRT6 might decrease the releasing of 5-HT in rat hippocampus. The test of cytokine showed that SZRT1 and SZRT6 cou更多还原