| Objective: In this study,the chemical constituents,the establishment of fingerprints,the experimental study of in vitro and in vivo pharmacodynamics,the analysis of data spectral effects and the combination of multi-omics were studied.The purpose of this study is to clarify the pharmacodynamic substance basis of Shuanghuanglian preparation,further reveal the compatibility law of Shuanghuanglian compound preparation,and explore the target and mechanism of action of the drug.We hope to form a system model with the analysis of the compatibility of the refined side with the spectrum-effect related technology as the core.At the same time,through the system to optimize addition and subtraction of the prescriptions to provide technical support for the development of new Chinese medicines.Methods: 1.According to the 2015 Chinese Pharmacopoeia method to make Shuanghuanglian preparation(compound,disassembled and single-flavored medicine preparations).Then we established the method of qualitative and quantitative by LC-QTOF/MS technology.2.We conducted the experiments of antibacterial and anti-RSV of Shuanghuanglian preparation.Among them,we use TI as the evaluation index for anti-RSV and the inhibition rate as an indicator of inhibition in vitro.We evaluated the anti-RSV effect of Shuanghuanglian preparation by measuring the body weight,organ index,pathological section and inflammatory factor content of mice to judge the in vitro and in vivo efficacy of Shuanghuanglian preparation.3.We used GRA combined with PLS analysis methods to find key medicinal ingredients in Shuanghuanglian preparation to infer the compatibility of the drug.We used the LS-SVM method to establish a spectral-effect correlation mathematical model and then established a spectral-effect correlation quality evaluation system for Shuanghuanglian preparation.4.Finally,we performed proteomics and metabolomics analysis on mouse lung tissue using LC-MS/MS technology.Data were processed by software such as R language,XCMS,SPSS,and SIMCA-P.By analyzing the differential changes of proteins and endogenous metabolites in mouse lung tissue,we screened differential proteins and potential biomarkers to explore the target and mechanism of Shuanghuanglian preparation.Result: 1.Using HPLC-TOF/MS/MS to analysis 93 compounds and inferring the possible structures of them,among them 45 compounds come from Honeysuckle,26 from Astragalus and 22 from Forsythia.Then,48 peaks with stable peaks and good linearity in the Shuanghuanglian preparation were selected for relative quantification,and their relative peak areas were determined.2.The results of PLS analysis showed that the body weight,organ index,pathological section of lung tissue and inflammatory factors of the model mice were severely pathologically marked,indicating that the model was successfully replicated.After different drug interventions,the model mice showed different degrees of recovery.Among them,Shuanghuanglian oral preparation has the best overall effect,and Forsythia has a relatively good anti-RSV effect in vivo.The results of in vitro experiments showed that the 50 batches of Shuanghuanglian oral liquid purchased had a great difference in efficacy.TI was 4-40 and the inhibition rate was between 65%-95%.The anti-RSV effect of the prepared Shuanghuanglian preparation was significant,and the antibacterial results showed that the Shuanghuanglian preparation compound and the Forsythia effect were significantly more than 85%.3.The results of GRA combined with PLS analysis showed that the main components affecting the antibacterial results in vitro include quercetin,hyperoside,forsythin and other substances from forsythia.It is indicated that the main component of Shuanghuanglian preparation in inhibiting the growth of Staphylococcus aureus in vitro is from Forsythia.The main component affecting the anti-RSV results in vitro is from honeysuckle,including caffeic acid,3,5-O-dicaffeoylquinic acid and oxydecaine.The mathematical model established by LS-SVM found that the mathematical model constructed by the model had a deviation of 0.35% for each batch of samples,and the relative deviation of the samples was within 5%.4.Proteomics studies identified 38 related differential proteins,including Cd81,Oas3,and Rock1.These differential proteins are mainly involved in signaling pathways including immune system processes,defense response processes,arachidonic acid binding processes,and Toll-like receptor binding processes.Metabolomics studies have shown that a variety of endogenous metabolite metabolic abnormalities in the model mice include: taurine,glucose,amino acids,glycerophospholipids.Shuanghuanglian oral solution can be adjusted specifically.It mainly involves metabolic pathways such as TCA cycle,amino acid metabolism,glycerophospholipid metabolism,pantothenic acid and Co A biosynthesis.Conclusion: 1.Shuanghuanglian compound preparation with multiple medicinal ingredients has a good therapeutic effect on inhibiting Staphylococcus aureus and anti-RSV in vitro.2.The correlation analysis between fingerprint and drug efficacy further explained the compatibility of Shuanghuanglian compound preparation.The main component of Shuanghuanglian preparation in vitro inhibition is mainly from Forsythia.The main component of anti-RSV in vitro is mainly from Honeysuckle.The results quantitatively analyzed the main pharmacodynamic components and their affiliation in Shuanghuanglian group,and scientifically explained the compatibility of Shuanghuanglian formula,which provided new ideas and methods for the study of compound Chinese medicine compatibility and component new drugs.3.The mathematical model constructed by Shuanghuanglian preparation can not only evaluate the quality of the drug based on the fingerprint information,but also predict the efficacy of the drug according to the fingerprint.It can also provide the establishment of the comprehensive quality evaluation system for Shuanghuanglian oral liquid spectrum effect.Based on the basis,further realize the comprehensiveness,rationality and accuracy of drug evaluation.4.Multi-omics combination analysis found that Shuanghuanglian compound preparation is closely related to the inflammatory process and immune process in the treatment of RSV-infected mice.Metabolic disorders of various endogenous substances in the model group,including taurine,phenylalanine,glucose,and glycerophospholipids.Shuanghuanglian compound preparation can play a good role in regulating a variety of metabolic pathways such as amino acid metabolism,glycerophospholipid metabolism and TCA cycle.Shuanghuanglian compound preparation can play a good role in regulating a variety of metabolic pathways such as amino acid metabolism,glycerophospholipid metabolism and TCA cycle.This regulation gradually restores the metabolic level of the body to normal.This method is in line with the holistic principle of the role of traditional Chinese medicine,and has obvious advantages in the research of the mechanism of action of Chinese medicine compound. |
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