| Background and SignificanceThis study is part of basic research on the composition of the law of Ephedra sinica herbal pairs, funded by the National Natural Science Foundation.Couple medicine also called drug pairs is a relatively fixed concered application form in clinic. Research single herb is difficult to reflect its role in prescription, but prescription composed by lots of drugs,It is difficult to draw accurate conclusions. As drug compatibility drugs on the prototype, the research can make up just less than the above studies. Therefore, it is necessary for drugs on the historical process of development and research of modern systematic induction and awareness in order to carry out their law more in-depth exploration, scientific guidance to clinical application.Herba Ephedrae-Radix Aconiti Later alls couplet medicines used for clinical in the book of Zhang Zhong jing there are Mahuang Fuzi Xixin Tang、Mahuang Fuzi Gancao Tang、Wutou Tang、Mahuang Fuzi Tang. Herba phedrae and Radix Aconiti Lateralis couplet medicines are different in different soup prescription dose and efficacy,the couplet medicines come from Mahuang Fuzi Xixin Tang. Mahuang Fuzi Xixin Tang indicate in cold table real and yang in the fever. Herba Ephedrae-Radix Aconiti Lateralis a extenrior and a interior,a false and a real complish complementary.The secret of prescription is dose, this article set Herba Ephedrae and Radix Aconiti Lateralis different compatibility proportion by its material basis in order to find the best ratio, compatibility law of the drug on the ratio of research from the chemical point of view. Through its metabolomics research to compare the differences of the endogenous substances and determine the compatibility of the corresponding biological targets, further investigate the drug from the biological point of view on the compatibility of connotation.Objection(1) By measuring Herba Ephedrae-Radix Aconiti Lateralis different proportions main active ingredient content, quantitative research Ephedra-aconite compatibility variation of main chemical components.(2) By comparing Herba Ephedrae-Radix Aconiti Lateralis different compatibility fingerprint composition changes, further research on the compatibility of the laws affecting other components.(3) The use of metabolomics methods by comparing Herba Ephedrae-Radix Aconiti Lateralis combination on subacute granulomatous inflammation model in rats urine biomarker changes, explore the compatibility of the metabolic pathway.Methods(1) The couplet medicines Herba Ephedrae and Radix Aconiti Lateralis in Mahuang Fuzi Xixin decoction were taken as study object of Herba Ephedrae-Radix Aconiti Lateralis (1:1), the compatibility ratio of Herba Ephedrae-Radix Aconiti Lateralis was multiplied floating up and down basing on the ratio of the couplet medicines reported in the literature. Herba Ephedrae water extract, Radix Aconiti Lateralis water extract, Herba Ephedrae-Radix Aconiti Lateralis (1:0.5) water extract, Herba Ephedrae-Radix Aconiti Later alls (1-1) water extract, Herba Ephedrae-Radix Aconiti Lateralis (1:2) water extract and mahuang decoction were individually prepared. The content of Norpseudoephedrine (NMP), Norephedrine (NME), Ephedrine (E), Pseudoephedrine (PE) and Methylephedrine (ME), which are the major active compounds of Herba Ephedrae, were analyzed by high performance liquid chromatography. The content of Benzoylmesaconine and BenzoylHypacoitine, which are the major active compounds of Radix Aconiti Lateralis in the extracts were analyzed by high performance liquid chromatography.(2) By using gradient elution HPLC chromatogpram obtained Herba Ephedrae, Radix Aconiti Lateralis, Herba Ephedrae-Radix Aconiti Lateralis(1:0.5,1:1,1:2), Similarity software used to analyze the chromatogram obtained ephedra ephedra common pattern and determine their common peaks. Determined by UV absorption chromatogram peaks of its ingredients.Compare Herba Ephedrae-Radix Aconiti Lateralis different proportion fingerprint ingredient Herba Ephedrae decotion rate of change of total peak area.(3) According to Herba Ephedrae-Radix Aconiti Lateralis has anti-inflammatory effects selection subacute granulomatous inflammation model in rats modeling,24rats were randomly divided into five groups namely normal group, model group, Herba Ephedrae group, Radix Aconiti Lateralis group, Herba Ephedrae-Radix Aconiti Lateralis group each group of six rats. Modeling were collected before and six days after modeling the urine, the urine was derivatized after centrifugation. Using GC/MS in rat urine metabolites full scan analysis, the resulting GC/MS data import XCMS for peak identification, peak alignment, less solvent peak, impurity peak processing. Exporting data normalized peak area, including sample generated name, the normalized peak area two-dimensional matrix. SIMCA-P software using rat urinary metabolite profiling of the principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA), PCA difference visually observe the different groups; PLS-DA between further study group the degree of difference, and in accordance with the importance of metabolite projection (VIP) Evaluation of different substances associated with the classification level to VIP>1metabolites as potential biomarkers. Using SPSS13.0software for potential biomarkers for statistical analysis, the difference was statistically significant selection of the material as the ultimate biomarkers. Through three days with the normal group, Herba Ephedrae group, Radix Aconiti Lateralis group, Herba Ephedrae-Radix Aconiti Lateralis group and model group were conducted PLS-DA analysis, analysis of significant differences in biomarkers that were identified through the database endogenous marker, combined with now some knowledge clarify Herba Ephedrae before and after compatibility Radix Aconiti Lateralis metabolic pathways are affected.Results(1) Compared with Herba Ephedrae single decoction, Herba Ephedrae-Radix Aconiti Lateralis compatibility of different degrees of five effects of ephedra alkaloids, its dissolution rate and aconite alkaloids in drugs on the proportion related to the greater proportion, the dissolution rate is smaller. Including Herba Ephedrae-Radix Aconiti Lateralis1:1,1:2group five kinds of aconite alkaloids were significantly lower (P<0.05). Decoction demethylation pseudoephedrine, ephedrine, ephedrine, pseudoephedrine, ephedrine methyl descending order of Herba Ephedrae> Herba Ephedrae-Radix Aconiti Lateralis1:0.5>1:1>1:2. NMP five kinds of rate of change of alkaloids is8%-40%, NME is10%-44%, E is10%-44%, PE is7%-43%, ME is10%-44%. Radix Aconiti Later alls decoction with a single flavor than Herba Ephedrae-Radix Aconiti Lateralis compatibility of Benzoyl-mesaconine and BenzoylHypacoit-ine gradually decreased and compatibility with Herba Ephedrae in the proportion related to Herba Ephedrae the greater the ratio, the greater the reduction Radix Aconiti Lateralis main active ingredient. Including Herba Ephedrae-Radix Aconiti Lateralis1:1,1:2group were significantly lower (P<0.05). Rate of change of two alkaloids Benzoylmesaconine rate is10%-27%, BenzoylHypacoitine change rate is9%-24%.(2) Ephedra single decoction were obtained13common peaks. Herba Ephedrae-Radix Aconiti Lateralis different compatibility proportion varying degrees, affect ingredient ephedra fingerprint patterns, compatibility of total peak area was significantly reduced as ephedrine, ephedrine, pseudoephedrine, meth ephedrine,11unknown peak,12unknown peak. Compatibility of total peak area was significantly higher for7unknown peaks,8unknown peak,9unknown peak,10unknown peak. Compatibility of total peak area has a slight influence of demethylation pseudoephedrine,6unknown peak,13unknown peak.(3) The metabonomic technique PCA and PLS-DA analysis methods and score plots, urinary metabolites changed significantly after modelling and divided into two categories, which displaying the similarities and differences between Results showed that the model group if ok. Model Id group, moving away from the blank group, the maximum in the model3d group, model4d group, after modeling, endogenous productthe type, concentration and relative proportions of the location of the gradual pullback to the blank group, but ultimately failed and the blank group overlap, indicating that the material metabolism of the model group showed a certain degree of self-regulation trend observed within the time limit, but could not be restored to their normal state. For the three days of the blank group, Herba Ephedrae group, Radix Aconiti Lateralis, Herba Ephedrae-Radix Aconiti Lateralis group together with the model group for PCA analysis, model group in the second quadrant, blank group in the first quadrant, indicating successful modeling; give Herba Ephedrae, Radix Aconiti Lateralis, Herba Ephedrae-Radix Aconiti Lateralis compared with model group, the distinction clear, Herba Ephedrae-Radix Aconiti Lateralis group in the first quadrant, Herba Ephedrae group in the third quadrant, Radix Aconiti Lateralis group in the fourth quadrant, deviate significantly from the model group, indicating that the three dose groups were played on the model intervention;3d group gradually to close to the direction control group, suggesting that the treatment group over time according to the model group trajectory regression blank group. Intuitive PCA above analysis shows that Herba Ephedrae-Radix Aconiti Lateralis group show metabolic state than Herba Ephedra group, Radix Aconiti Lateralis group closer to the control group.For the three days of the blank group, Herba Ephedrae group, Radix Aconiti Lateralis group, Herba Ephedrae-Radix Aconiti Lateralis group and model group were analyzed for PLS-DA obtained characterize each treatment group and the model group differences in biomarkers, including organic acids, fatty acids, amino acids and amines. Normal group with the model group PLS-DA analysis, obtained characterize differences in model group and normal group biomarkers, including: higher than the control group of n-propylamine, pyruvic acid, benzamide, phenylacetic acid, leucine, lauric acid, phenylalanine, isobutyric acid, tyrosine, tryptophan, isonicotinamide; levels lower than the control group of citric acid, hippuric acid, isoleucine. Herba Ephedrae group biomarkers include:higher than model group hippuric acid; level lower than the model group, valine, benzamide, phenylacetic acid, isobutyric acid, tyrosine, tryptophan, isonicotinamide. Biomarkers Radix Aconiti Lateralis group comprising:level higher than the model group, hippuric acid, isoleucine; level lower than the model group, glutaric acid, valine, benzamide, phenylacetic acid, lauric acid, phenylalanine, isobutyric acid, tyrosine, tryptophan. Herba Ephedrae-Radix Aconiti Lateralis group of biomarkers include: higher than the model group, citric acid, hippuric acid; level lower than the model group, n-propylamine, pyruvic acid, benzamide, acid, leucine, lauric acid, suberic acid, phenylalanine, isobutyric acid, tyrosine, tryptophan, isonicotinamide.Conclusion(1) After Herba Ephedrae compatibility with Radix Aconiti Lateralis, Herba Ephedrae in the dissolution rate of the active ingredient significantly reduced, the dissolution rate of drugs on Radix Aconiti Lateralis in the proportion related to the greater proportion of the dissolution rate is smaller; Herba Ephedrae and compatibility of Radix Aconiti Lateralis, with the single flavor compared monkshood, Radix Aconiti Lateralis in the dissolution rate of the active ingredient was significantly reduced, with increasing doses of Radix Aconiti Lateralis, monkshood Herba Ephedrae on the amount of fried greater impact; Herba Ephedrae-Radix Aconiti Lateralis compatibility with each other after the dissolution of the active ingredient has a certain inhibition.(2) Herba Ephedra e-Radix Aconiti Lateralis decoction of different compatibility fingerprint ingredient ephedra total peak area affected in varying degrees, the peak area change a few numbers to be specific to the analysis of peak(3) Herba Ephedrae, Radix Aconiti Lateralis, Herba Ephedrae-Radix Aconiti Lateralis inflammation model group levels of biomarkers in urine showed different degrees of correction, three-administered group showed some improvement in the metabolic state of inflammation model, through the study of metabolic pathways of Herba Ephedrae-Radix Aconiti Lateralis can regulate the body’s overall metabolism, energy metabolism, including Herba Ephedrae-Radix Aconiti Lateralis group compatibility of biomarkers callback greatest extent that Herba Ephedrae-Radix Aconiti Lateralis compatibility phenological models for intervention in chronic inflammatory efficacy of best... |
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