Studies On Warm And Cold Nature Of JiangHuang And YuJin Based On Metabonomics

The properties of tradional Chinese medicinal herbs, which are the essential basis of the analyses and clinical usage of Chinese medicinal herbs, are summarized in medical practice and on the basis of the theories of Yin-Yang, Hot-Cold,Toxicity and other theories of therapeutic principles of traditional Chinese medicine(TCM).The reason that Chinese medicinal herbs can prevent and treat diseases is due to their two internal natures:chemical components and the biological activities. Because the source of tradional Chinese medicinal herbs is widely, the chemical components were various and the biological activities were related with too many active aspects. So it was lacking the proper methods and objects to study and research the property theory of traditional Chinese medicinal herbs by modern scientific language.Jianghuang and Yujin which come from the rhizome and tuberous root of Curcuma longa L. are used as two different Chinese medicinal herbs. In the property theory of traditional Chinese medicinal herbs, it was believed that Yujin has cold nature and Jianghuang has hot nature. Accordingly Jianghuang was used to promote circulation of blood and arrest pain while Yujin was used to circulate the blood and normalize the gallbladder and liver to cure jaundice in clinic. As Jianghuang and Yujin come from the same plant of Curcuma longa L, they have similar chemical components and effective activities. So Jianghuang and Yujin were selected as objects, which makes it easier to study and compare the chemical components and the biological activities so as to explain why they have hot and cold nature.The metabonomics/metabolomics gives a good and new idea to study the plant secondary metabolites and the therapeutic evaluation of TCM. As a part of system biology, metabonomies apply modern chemical analysis technology such as HPLC-MS、GC-MS、NMR to determine active alterations of endogenous metabolites of biological organism and subsequently reveal changes in the whole process, including occurrence, development,and consequences in response to stimuli or drug treatment. Metabonomies, defined as "the quantitative measurement of the multiParametrie time-related metabolic responses of a complex system to a pathophysiological intervention or genetic modification" has been widely used in the researches of pharmacology and biological seiences.In this thesis firstly we established HPLC-DAD-MS and GC-MS methods to analysis and compare the the plant secondary metabolites of the rhizome(Jianghuang) and tuberous root(Yujin) of Curcuma longa L. Principle Component Analysis(PCA) was used to discovery the biomarkers between Jianghuang and Yujin from chromatograms. Secondly we establish a GC-MS method to analyse plasmar metabolites and a HPLC-MS method to determine brain catecholamine metabolic profiling.With the methods we evaluated the Heat-Syndrome model(damp-heat jaundice model) and Cold-Syndrome model(cold-damp jaundice model). Pattern recognition was used to distinguish Heat-Syndrome model and Cold-Syndrome model. Based on this, we evaluated drug effect of Jianghang,Yujin,and curcumin on Heat-Syndrome model and Cold-Syndrome model. According to different effects on Heat-Syndrome model and Cold-Syndrome model, we can explain and illustrate hot and cold nature of Jianghuang and Yujin.Main contents are summarized below:1. The plant secondary metabolites analysis of Jianghang and Yujin by HPLC-DAD-MS and GC-MS.In order to analyse and compare the the plant secondary metabolites including volatile terpenes and nonvolatile curcuminoids of Jianghuang and Yujin, the metabolite profilings was established by HPLC-DAD-MS and GC-MS.18 samples of Curcuma longa L inluding the rhizome and tuberous roots were collected from the regions in Sichuan province of China. The samples analyzed by HPLC-DAD-MS were processed by ultrasonic extraction in methanol.While the samples analysed by GC-MS were processed by Soxhlet extraction with light petroleum. After peaks of every chromatograms synchronized, multivariate analysis was performed using the SIMCA-P 12 version (Umetrics AB, Ume□, Sweden) based on the all peak areas(peak area/sample weight). Principal component analysis (PCA) was used and reveal the differences between the the rhizome and tuberous roots.Form Scores Polts of PCA based on HPLC or GC, there are both obvious difference between Jianghuang and Yujin. Based on chromatograms of HPLC-DAD-MS, there are 10 biomarkers found between Jianghuang and Yujin. Seven of them were identified as Bisdemethoxycurcumin、Demethoxycurcumin、Curcumin Dihydrocurcumin Ar-turmerone、a,β-turmerone (bisacumol)、Zingiberene (B-bisabolene, B-curcumene),which are all higher in Jianghuang than Yujin.Based on chromatograms of GC-MS,14 biomarkers were found and identified whose contents are obvious different between Jianghuang and Yujin.2. Evaluation of Hot and cold nature of Jianghuang and Yujin by metabonomics method of GC-MS technologyThe GC/MS method was established to analyse the blood plasma samples and the Heat-Syndrome model(damp-heat jaundice model) and Cold-Syndrome model(cold-damp jaundice model) rats blood plasma was introdued to GC/MS. Then applying the GC/MS method, the blood plasma samples from the Heat-Syndrome model and Cold-Syndrome model rats treated by Jianghuang、Yujin and curcumin were detected by the machine.With the PCA and PLS-DA, the status of drug effect was evaluated.With the multianalysis to the metabonomic datas of GC/MS, the Heat-Syndrome group、Cold-Syndrome group and the normal group can be divided into three groups accordingly. And compared with normal group, the biomarkers of the Heat-Syndrome and Cold-Syndrome group were found and identified such as aminoacids, organic acids. Then we compared the efficiency of jianghuang,Yujin and curcumin on the Cold-Syndrome model and the Heat-Syndrome model. The result suggest that Jianghuang and curcumin have more regulation effect on Cold-Syndrome model than Yujin based on the metabonomic datas. While Yujin have more regulation effect on Heat-Syndrome model than Jianghuang and curcumin.3. Evaluation of Hot and cold nature of Jianghuang and Yujin Based on brain catecholamine Metabolic ProfilingHPLC-MS method for the quantitative determination of 7 catecholamines in brain samples was developed. And Principal Component Analysis(PCA) was processed to analyze the catecholamine metabolic profiling. Heat-Syndrome model and Cold-Syndrome model rats was firstly introdued to HPLC-MS. Then applying the the method, the samples of the Heat-Syndrome model and Cold-Syndrome model rats treated by Jianghuang、Yujin and curcumin were detected.The results suggest Heat-Syndrome model and Cold-Syndrome model can be distingushed by catecholamine metabolic profiling. Jianghuang、Yujin and curcumin has different effect for the Heat and Cold Syndrome model based on Brain catecholamines Metabolic Profiling.