背景:柴胡桂枝汤(CGD)是治疗流感和发烧的经典中药(TCM)处方,柴胡的成分,桂枝(桂枝),黄芩,党参(党参),甘草(知甘草)半夏(Fabanxia),生姜(盛江),白芍(白芍)与大枣(大枣)的比例为12:4.5:4.5:3:6:4.5:4.5:4.5:4。中医的疗效,如果有差异,取决于不同的提取方法和提取成分。
目的:本研究以不同提取方法评价CGD提取物的抗流感病毒作用。分析成分并探讨它们的相关性。
方法:分别用四种提取方法制备CGD,传统汤剂(TD),两步醇水提取(AWE),乙醇回流提取(AE)和水回流提取(WE)。基于流感小鼠模型,4种CGD提取物在体内抗流感病毒的功效用体重的治疗指数来评价,直肠温度,肺指数,小鼠胸腺指数和肺病毒载量。四种CGD提取物中的化学成分,通过UPLC-Q-Exactive/MS鉴定了在大鼠血液中吸收的具有原型或代谢物的化合物。采用偏最小二乘(PLS)方法探讨CGD提取物中成分变异与综合功效指数的相关性。通过分子对接进一步获得潜在的有效成分。
结果:与其他三种提取物相比,AWE具有最好的抗流感效果。它可以改善小鼠流感病毒感染引起的症状,增加体重和直肠温度,降低肺组织中的肺指数和病毒载量。从TD确定了129、144、140和129个组件,AWE,AE,分别是我们。鉴定的成分主要为黄酮类化合物,萜类化合物,有机酸,苯丙素类化合物,氨基酸,核苷,酚类物质,生物碱,等。口服给药后在大鼠血浆中检测到CGD的43个原型和49个代谢物。七个组成部分,肉桂醛,沃戈诺赛德,黄芩苷,黄芩素,没食子酸,oroxylinA-7-O-葡糖苷酸和香豆素,与抗流感作用显著相关,所有这些都与NA具有良好的结合活性,IL-6、STAT3、AKT1、EGFR和TNF。
结论:两步醇-水提取是CGD制剂的最佳选择。肉桂醛,沃戈诺赛德,oroxylinA-7-O-葡糖苷酸,香豆素,没食子酸,黄芩素和黄芩苷在抗流感作用中起着一定的作用,可作为CGD质量评价的潜在标记化合物。
BACKGROUND: Chaihu Guizhi decoction (CGD) is a classic Traditional Chinese Medicine (TCM) prescription for the treatment of influenza and fever, composes of Bupleuri Radix (Chaihu), Cinnamomi Ramulus (Guizhi), Scutellariae Radix (Huangqin), Codonopsis Radix (Dangshen), Glycyrrhizae Radix Et Rhizoma Praeparata Cum Melle (Zhigancao), Pinelliae Rhizoma Praeparatum (Fabanxia), Zingiberis Rhizoma Recens (Shengjiang), Paeoniae Radix Alba (Baishao) and Jujubae Fructus (Dazao) in the ratio of 12:4.5:4.5:4.5:3:6:4.5:4.5:4. The efficacy of TCM, if there are differences, depends on the different extraction methods and extracted components.
OBJECTIVE: This study was to evaluate the anti-influenza virus effect of CGD extracts with different extraction methods, analyze the components and explore their correlation.
METHODS: CGD were prepared with four extraction methods respectively, the traditional decoction (TD), two steps alcohol-water extraction (AWE), alcohol reflux extraction (AE) and water reflux extraction (WE). Based on the influenza mouse model, the efficacy of anti-influenza virus in vivo of the four CGD extracts were evaluated with the therapeutic index of body weight, rectal temperature, lung index, thymus index and lung viral load of mice. The chemical components in four CGD extracts, and compounds absorbed in rats blood with prototypes or metabolites were identified by UPLC-Q-Exactive/MS. The partial least squares (PLS) method was used to explore the correlation between the components variation in CGD extracts and the comprehensive efficacy index. The potential effective components were further accessed by molecular docking.
RESULTS: Comparing with the other three extracts, AWE has the best anti-influenza effect. It could ameliorate the symptoms caused by influenza virus infection in mice, increase body weight and rectal temperature, reduce the lung index and virus load in lung tissue. 129, 144, 140 and 129 components were identified from TD, AWE, AE, and WE respectively. The identified components were mainly including flavonoids, terpenoids, organic acids, phenylpropanoids, amino acids, nucleosides, phenols, alkaloids, etc. 43 prototypes and 49 metabolites of CGD were detected in rat plasma after oral administration. Seven components, cinnamaldehyde, wogonoside, baicalin, baicalein, gallic acid, oroxylinA-7-O-glucuronide and coumarin, showed significant correlation with anti-influenza effects, all of which had good binding activity with NA, IL-6, STAT3, AKT1, EGFR and TNF.
CONCLUSIONS: Two steps alcohol-water extraction was optimal for CGD preparation. Cinnamaldehyde, wogonoside, oroxylinA-7-O-glucuronide, coumarin, gallic acid, baicalein and baicalin play a certain essential role in anti-influenza effects and may be taken as a potential maker compounds for quality evaluation of CGD.