Study On The Attenuate And Synergistic Mechanism Designed For Mixed-decoction Of Radix Aconiti Lateralis Praeparata And Radix Glycyrrhizae In Sini Decoction Based On Biopharmaceutics

Sini Decoction (SND), a classical Traditional Chinese Medicine formula recorded in Treatise on Febrile Diseases(Shanghan Lim), which is composed of Radix Aconiti Lateralis Preparata. Radix Glycyrrhizae preparata and Rhizoma Zingiberis, is a representative formula, with the function of restoring yang from collapse. Modern clinical and recent researches have shown that SND can protect the myocardium effectively during myocardial ischemia reperfusion injury. Mixed-decoction of Radix Aconiti Lateralis Praeparata and Radix Glycyrrhizae belongs to one of the basic requirements for Chinese decoction, and the purpose of attenuate and synergistic can be reached according to mixed-decoction of Chinese medicine compound.ObjectiveThis thesis is guided by integrally concept and systematic ideas, which are suitable for the whole concept and dialectical view of Traditional Chinese Medicine. The idea that attenuate and synergistic effects of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae in SND are related to a variety of chemical components from effects on organisms and the organisms reply, will be proposed. Based on the theory of biopharmaceutics, the analytical tools of LC-MS, GC-MS and metabonomics, and the research pattern of’toxic substance group-weakened-synergistic materials group-synergy’ corresponding metabolic characteristics both in vivo and in vitro will be used together, which are usually preferred in studying the attenuate and synergistic mechanism of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae in SND. We aim to explore the material base and mechanisms of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae in SND, in order to provide a new bunch of ideas for studying preparation theory of mixed-decoction of Chinese medicine compound integrity and systemically.Method and Result1.Study preparation technology and quality standard of SND:The contents of Glycyrrhizic Acid, DDA(Mesaconitine, Aconitine and Hypaconitine),6-gingerol and the value of dry extract were set as indexes for comparative analysising SND prepared by different processes between the traditional method and Chinese Pharmacopoeia (2010,1) method. Then the traditional process of SND were chosen. The content of Monoester Diterpenoid Alkaloids (Benzoylmesaconine, Benzoylaconine, and Benzoylneoline), Glycyrrhizic Acid and the ratio of extractum were used as chemical properties. The function of anti-hypoxia of mice is set as the pharmacodynamic index. Firstly, the optimal technical conditions were selected by orthogonal design L9(34). Then ANN model was established to obtain the optimum extraction parameters, and proved by the experiments. The optimal technical parameters were as follows:extracting3times in all, once1.5h, adding8times amount of water. TLC was used to identify Radix Glycyrrhizae and Rhizoma Zingiberis. and the contents of MDA and Glycyrrhizic Acid in SND were determined by HPLC in the research of the quality standard. It was confirmed that the content of MDA should be more than25.00μg·mL-1in SND (0.8g·mL-1), and the content of Glycyrrhizic Acid should be more than8.00mg·mL-12. Compare content change of DDA and pharmacokinetics of hypaconitine between the mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae or not in SND:The simultaneous determination method of DDA in supernatant of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae or not in SND was estimated. The pharmacokinetics of hypaconitine mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae or not in SND on mice were contracted.The results showed that the content of DDA in SND was lower than the one of SND without Radix Glycyrrhizae, and there were significant differences between them (p<0.01).At the same time, though the absorption rate of hypaconitine was slowed, the total absorption area of hypaconitine was added after mixed-decoction of Radix Glycyrrhizae in SND.3. Metabonomics research on the effects of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae or not in SND on MI-RI mouse cardiomyocytes based on GC-MS:A myocardial ischemia reperfusion injury model of H9c2cardiomyocytes in vitro was established.The effect of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae or not in SND on myocardial ischemia reperfusion injury in H9c2 cardiomyocytes was studied.and the metabonomic method using GC-MS was developed to research the metabolic profile of extracellular fluid. Compared with the SND without Radix Glycyrrhizae, the protective effect of myocardial ischemia reperfusion injury model of H9c2cardiomyocyte from SND was stronger. The attenuation and synergistic mechanism of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae in SND were related with the glycolysis, lipid metabolism, citrate cycle and nitrogen metabolism of amino acids metabolism.4.Metabolomic fingerprinting analyze and metabonomics research of mice plasma on mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae or not in SND based on GC-MS:Metabolomic fingerprint and metabonomics of mouse plasma based on GC-MS were adopted. From the integrally modes of metabolic regulation,it is proved that the attenuation and synergistic mechanism of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae in SND were related with the glycolysis,lipid metabolism, citrate cycle and some amino acids metabolism.ConclusionIn this thesis,based on biopharmaceutics theory, we have confirmed that mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae in SND, which is the main toxic component of Radix Aconiti Lateralis Preparata united in vitro firstly, as well as liberated and absorbed slowly while in vivo, and realised the’attenuation and synergistic". It doses not only reveal that the attenuate and synergistic mechanisms of mixed-decoction of Radix Aconiti Lateralis Preparata and Radix Glycyrrhizae in SND are related with the glycolysis, lipid metabolism, citrate cycle and some amino acids metabolism, but also provide a new method and idea for studying preparation theory of mixed-decoction of Chinese medicine compound integrally and systematically.