| With the rapid development of traditional Chinese medicine(TCM)industry,the demand for TCM is also increasing day by day.However,most traditional Chinese medicine preparations only extract a few types of chemical components,resulting in a large number of traditional Chinese medicine residues,which cause great pressure on the environment;at the same time,some traditional Chinese medicine grows in the western region with fragile ecology,and the long-term unplanned mining causes great damage to the local ecological environment.It is one of the important ways to realize the sustainable development of traditional Chinese medicine resources to fully tap the residual resource value of traditional Chinese medicine residues and promote the recycling of traditional Chinese medicine resources.Sophora flavescens is mainly distributed in the northwest of China.Its medicinal materials are mainly used in the preparation of alkaloid components of Sophora flavescens in the traditional Chinese medicine industry,at the same time,a lot of acid water is produced to extract drug residues.However,there are a lot of isopentenyl flavones in the residue of Sophora flavescens(acid)water extraction.The content of these flavones is considerable,the preparation process is simple,and they have a wide range of biological activities.The value of isopentenyl flavonoids in the residue of water extraction of Sophora flavescens can be found as the basis for the resource recycling and sustainable development of Sophora flavescens.In this study,the active parts and monomers of isopentenyl flavones were prepared from the dregs of Sophora flavescens,and the antibacterial structure-activity relationship and mechanism were studied.On the basis of the above-mentioned in vitro study,the whole animal model of mastitis in mice was further used to explore the active site of isopentenyl flavone of Sophora flavescens and its antibacterial and anti-inflammatory activities and mechanism in vivo.In order to expand the application of isopentenyl flavones in medicine,the disease targets corresponding to isopentenyl flavones of Sophora flavescens were mined by network pharmacology technology.The results showed that these components had a high matching degree to type 2 diabetes mellitus.Furthermore,molecular docking technology was used to evaluate the binding ability of isopentenyl flavones of Sophora flavescens to key targets of diabetes mellitus The content of isopentenyl flavonoids in the dregs of Sophora flavescens water extraction can provide basis for the research and development of new type 2 diabetes drugs.Through the research of this paper,we hope to provide support for the research of isopentenyl flavonoids in the residue of water extraction of Sophora flavescens in animal husbandry instead of antibiotics and the creation of new type 2 diabetes drugs,and provide scientific basis for the discovery of resource value of water extraction residue of Sophora flavescens and the recycling of resources of Sophora flavescens.This paper is divided into four chapters,which are summarized as follows:Chapter 1:Literature researchIn this part,the structural characterization and pharmacological activities of Sophora flavescens isopentenyl flavonoids were summarized,and the industrialization status of these components was analyzed,so as to provide a theoretical basis for the development and utilization of Sophora flavonoids.Chapter 2:Study on antibacterial activity,structure-activity relationship and antibacterial mechanism of Sophora flavescens isopentenyl flavonoids1.Taking the content of total isopentenyl flavonoids in Sophora flavescens as an index,the extraction technology of total isopentenyl flavonoids from Sophora flavescens was studied by method of orthogonal processing investigation.The experimental results showed that the best extraction technology of isopentenyl flavonoids from Sophora flavescens was as follows:ethyl acetate percolation method with material-liquid ratio of 1:4,soaking time of 24 h,flow rate of3ml·min~-11 and percolation of 6 times.According to this method,the total flavonoid yield of Sophora flavescens was 2.53%,and the content of total flavonoids in the extract was 50.12%by UV spectrophotometry.At the same time,the flavonoids in Sophora flavonoids were qualitatively analyzed by LC-MS.A total of 20 flavonoids were identified,of which 18 were isopentenyl flavonoids.2.Total isopentenyl flavonoids of Sophora flavescens were systematically isolated and 6monomer compounds(Isoxanthohumol,Kushenol,Kurarinone,Sophoraflavanone G,Kushenol A and Leachianone A)were obtained.The inhibitory effect of Sophora flavescens isopentenyl flavonoids on Escherichia coli,Shigella dysenteriae,Clostridium perfringens and Staphylococcus aureus was evaluated by broth dilution method with the minimum inhibitory concentration(MIC)as index.At the same time,the antibacterial activity of these flavonoids was compared with that of non-isopentenyl flavonoids,and the antibacterial structure-activity relationship of these flavonoids was discussed.The results showed that these compounds had no significant antibacterial activity against Gram-negative Escherichia coli and Shigella dysenteriae,but had significant antibacterial activity against Gram-positive Staphylococcus aureus and Clostridium perfringens.The structure-activity relationship showed that the antibacterial activity of isopentenyl flavonoids was significantly stronger than that of non-isopentenyl flavonoids,and their 8-position isopentenyl substitution and 3,5-position hydroxylation enhanced their antibacterial activity,while 5-position methylation weakened their antibacterial activity.3.Taking kurarinone as an example,the bacteriostatic mechanism of these compounds on the growth of Staphylococcus aureus was discussed by nuclear magnetic metabonomics.The results showed that the antibacterial activity may be related to the interference of kurarinone on the tRNA biosynthesis of aminoacyl groups of Staphylococcus aureus,glutathione metabolism;glycine,serine and threonine metabolism,as well as alanine,aspartic acid and glutamic acid metabolism.Chapter 3:Evaluation of the effect and mechanism of Sophora flavescens isopentenyl flavonoids and their compatibility on mastitis in mice induced by Staphylococcus aureus1.Adopt the micro chessboard dilution method,with FIC Index is the evaluation standard to evaluate the inhibitory effect of isopentenyl flavanone and its combination group on Staphylococcus aureus.At the same time,the mouse mastitis model caused by Staphylococcus aureus was established to evaluate the antibacterial and anti-inflammatory activities in vivo.The results of antibacterial experiment in vivo showed that isopentenyl flavanone and its combination group had certain improvement on the indicators of mastitis in mice,isopentene flavanone and its combination group had certain improvement on the indicators of mastitis in mice The effect of the combination of alkenyl flavone and Sophora flavescens Glycyrrhiza group on the organ index and the amount of bacteria in the tissues of the mastitis mice was better than that of the single group,and the effect of Isoamyl flavone of Sophora flavescens was better than that of flavone of Glycyrrhiza flavescens and Sophora flavescens alkaloids;but the effect of flavone of Glycyrrhiza on the histopathology of the mammary gland of the model mice and the decrease of the inflammatory factors IL-1β,IL-2 and TNF-αwere better than that of Isoamyl Sophora flavescens Flavonoids and alkaloids of Sophora flavescens,and isopentenyl flavonoids of Sophora flavescens and flavonoids of Glycyrrhiza have synergistic effect.The combination group of isopentenyl flavanone and alkaloid of Sophora flavescens showed more synergistic effect in bacteriostasis,while the combination group of isopentenyl flavanone and glycyrrhizin showed more synergistic effect in anti-inflammatory.2.Based on UPLC-QTOF/MS technique,metabonomics was used to study the intervention mechanism of Sophora flavescens isopentenyl flavonoids on mastitis mice induced by Staphylococcus aureus.The results showed that 45 differential metabolites were found in urine,blood and tissue samples of mice.The study showed that isopentenyl flavone of Sophora flavescens could inhibit the mastitis activity of mice,which might affect the propionate generation Metabolism processes of metabolism,such as metabolism of metabolism,ether lipid metabolism,histidine metabolism and terpene skeleton biosynthesis,are related.Chapter 4:Network pharmacology study on Sophora flavescens isopentenyl flavonoidsTo predict the pharmacological activity,target and signal pathway of isopentenyl flavonoids of Sophora flavescens,and to provide theoretical guidance for the discovery of its resource value.Through tcmsp,CNKI and PubMed database,isoamyl flavones of Sophora flavescens were collected;by Swiss target prediction platform,isoamyl flavones of Sophora flavescens were predicted;by David platform,disease correlation analysis was carried out;by Cytoscape 3.7.2 The network of traditional Chinese medicine compound target was constructed;PPI network was constructed by using string database;KEGG pathway and go function enrichment analysis were carried out for the genes enriched by disease,and the results were visualized by using version 3.6.3 software;molecular docking research was carried out by using autodock via 1.1.2.A total of 52 isopentenyl flavonoids of Sophora flavescens were found corresponding to 170 target genes.The results of disease correlation analysis showed that the target genes of isopentenyl flavonoids of Sophora flavescens were closely related to type 2diabetes.PPI network analysis showed that the treatment of type 2 diabetes by isopentenyl flavonoids of Sophora flavescens might be related to PTPN1、AKT1 and HSD17B1 and other gene proteins;go and KEGG The results showed that there were 147 biological processes(P<0.05)and 31 signal pathways(P<0.05)related to the effect of isopentenyl flavonoids in Sophora flavescens on the treatment of type 2 diabetes mellitus.The results of molecular docking showed that alpinumisolavone and PTPN1 had strong binding capacity,and the binding capacity was stronger than the positive drugs glibenclamide,flavescenone C and kushenol The binding ability of V and HSD17B1 is better than that of glibenclamide,and kushenol f has the strongest binding ability with AKT1,and the binding ability is slightly weaker than that of glibenclamide.The results showed that isopentenyl flavonoids of Sophora flavescens is a potential active component in the treatment of type 2 diabetes.It can act on PTPN1,AKT1,HSD17B1 and other targets,regulate multiple signal pathways,so as to play a therapeutic role in type 2 diabetes. |
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