| Objective: Objective to investigate the mechanism of rheumatoid arthritis(RA)attack,and investigate the active components,targets and related pathways of Fufang sifangteng,investigate the anti RA active components,targets and related pathways of Fufang sifangteng,so as to provide basis for elucidating the mechanism of Fufang sifangteng in the treatment of RA.The preparation of compound tetragonium granules,which has the effect of treating rheumatism and arthralgia,was established,and the preparation technology,the draft quality standard and the stability were investigated.Methods:(1)Combined with the literature reports,the main chemical components in the prescription of Fufang sifangteng were analyzed,and the potential Q-marker component library was predicted,TCMSP database was used to predict the potential Q-marker components,TTD database was used to search for RA disease targets,and the target proteins were mapped to String and KEGG databases to predict their pathways and their relationship,signal pathways and key target proteins related to biological effects were screened,and the protein-protein interaction was analyzed.The gene expression microarray of RA disease was retrieved from the GEO microarray database in NCBI,and the RA gene expression data was downloaded from the GEO database.After processing the data with R language,the bioinformatics analysis was carried out to mine the differentially expressed genes and abnormal signal pathways in the process of RA disease,Q-biomarkers were predicted by combining differential genes with chemical targets.HPLC and UPLC-QTOF/ MS were used to analyze the main potential Q-marker chemical components in the samples of Fufang sifangteng,The visualization network of "potential quality markers,action targets and Q-biomarkers" was constructed by Cytoscape.The Q-marker was determined by the strategy of "Q-biomarkers → action targets → Q-marker".According to the analysis of the effectiveness and testability of potential q-marker components,the q-marker component library was established.(2)The extract yield and bergenin content were used as indexes.L9(34)orthogonal test was used to optimize the extraction,concentration and drying process.The purification,concentration and drying process were determined by comparative experiments.According to the angle of repose,bulk density,hygroscopicity and molding condition of particle excipients,the excipients and adhesive were determined.(3)After the preparation process was established and verified to be feasible,the systematic investigation method was adopted.The related tests were carried out for the description,identification,inspection of related items,the content of brassinogon as the index,and the related tests,and the quality standard research method of compound tetragonium tetragondii granules was established and its stability was investigated.Results:(1)32 potential q-marker components were screened and 147 target points were obtained.Through TTD database search and screening,88 RA targets were obtained.The data of GEO chip were compared and analyzed to obtain 884 different genes.According to the screening criteria: logfc > 2 or logfc <-2,P < 0.05,144 differential genes were screened by geo chip analysis.The differential genes of GEO chip,potential targets of q-marker components and RA targets were analyzed together.The top ten targets with the highest correlation were IL10、MCL1、PTGS1、PTGS2、MMP9、AKT1、CXCL8、STAT3、IL6、TNF、JUN.The targets were inversely correlated with the drug targets potential Q-marker ranking with high relevance.Nine components without relevant targets and standard reference substance,such asβ-terpineol,decarine,bergamot lactone,homosacroidin,daucosterol,trigonelline,noraconitine,isomonoemodin and isozipraeruptorin,were removed.According to the validity and testability analysis of potential q-marker components,Q-marker components were finally determined as six components by UPLC-QTOF/ MS,they are: bergenin,resveratrol,osthole,hesperidin,gallic acid,dihydrocaryol angelic acid ester.According to the results of HPLC,bergenin content was determined as the evaluation index of subsequent preparation process and quality standard.(2)The preparation process was determined as follows: the prescription amount of medicinal materials was soaked in 8 times of water for 3 h,heated and refluxed for 30 min,three times,combined with filter solution,the filtrate was centrifuged at 4000 rpm for 10 min,and then concentrated into a clear paste with relative density of 1.20-1.35(60℃).Add appropriate amount of auxiliary materials to the paste,mix well,and dry in a vacuum microwave dryer;beat the dried paste into fine powder,add auxiliary materials to 300 g,mix well,spray 90% ethanol to make granules,dry in a blast drying oven at60℃,and then complete the granules.(3)Objective to establish a TLC method for the identification of tetragonum,Zanthoxylum nitidum and Angelica pubescens in Fufang sifangteng granules.The samples showed the same color spots on the corresponding position of the control chromatography,but the negative samples do not show the same color spots on the same position.The separation is good and the spots were clear.A method for the determination of Bergenin in Fufang Sifang granules was established.The content limit ofBergenin in Fufang Sifang granules was determined to be no less than 10.03mg/g.Conclusion:(1)Based on the concept of Q-marker,combined with network pharmacology analysis and GEO chip verification,this study established the quality control traceability system of Fufang sifangteng,and analyzed the pharmacodynamic mechanism of Fufang sifangteng in the treatment of RA,so as to provide the basis for improving the quality and quality control level of traditional Chinese medicine.It provides a new way to study the pathogenesis of RA and the prevention and control effect of compound tetragonum on RA according to bioinformatics knowledge.(2)In this study,the preparation technology of Fufang sifangteng was established,the quality standard research method of Fufang sifangteng granules was established,and the stability of Fufang sifangteng granules was investigated.The development of the preparation was completed,which provided the basis and reference for the follow-up production and clinical research of the preparation. |
PDF Full Text Request