Study On Screening Of Dandelion Active Ingredients For Inhibiting Vaginal Diseases And Its Mechanism

Dandelion is also known as Pugongcao,Popoding and so on,which was included in the 2015 edition of the Chinese Pharmacopoeia as the dandelion of Asteraceae.Dandelion is widely used in clinical practice,mainly including antibacterial,anti-inflammatory and analgesic,anti-virus,anti-tumor,anti-cancer and other aspects,especially in the more application of antibacterial and anti-inflammatory and good results.In this study,7 kinds of dandelion samples(Ⅰ,Ⅱ,Ⅲ,Ⅳ,Ⅴ,Ⅵ and Ⅶ)were isolated and purified by using a series of methods including hot water extraction,80% alcohol extraction,and ethyl ether Soxhlet extraction.And antimicrobial active ingredients were screened against Candida albicans(C.albicans),Gardnerella vaginalis(G.vaginalis),and Staphylococcus aureus(S.aureus).The components of dandelion extract were analyzed and detected by high-performance liquid chromatography(HPLC),ultra high-performance liquid chromatography tandem mass spectrometry(UPLC-MS/MS)and gas chromatography mass spectrometry(GC-MS),respectively.The antibacterial mechanism of dandelion extract on C.albicans and G.vaginalis was studied by analyzing extract on the minimum inhibitory concentrations,cell ultrastructure,antibacterial kinetics,cell membrane permeability and the effects of cell wall glucan synthesis of C.albicans and G.vaginalis.The main findings were as follows:(1)The yield of sample Ⅰ,sample Ⅱ,sample Ⅲ,sample Ⅳ,sample Ⅴ,Sample Ⅵ,and sample Ⅶ were 0.4679%,0.0198%,0.0195%,0.0191%,0.0246%,0.0221%,and2.8697%,respectively.Among 7 kinds of dandelion extracts,dandelion sample Ⅰ had strong antibacterial activity against C.albicans,G.vaginalis and S.aureus,and sample Ⅶ had strong antibacterial activity against S.aureus.(2)HPLC combined with UPLC-MS/MS analysis showed that the 8 phenolic compounds were separated from dandelion sample Ⅰ by HPLC.Peak 1,peak 2,peak 3,peak 4,peak 5and peak 6 were 4-coumaric acid,ferulic acid,quercetin pentoside,3,5-di-O-caffeoylquinic acid,4,5-di-O-caffeoylquinic acid,luteolin,respectively.peak 7 and peak 8 were two unknown compounds.And the relative contents were 11.45%,3.96%,10.48%,34.24%,3.91%,11.80%,3.65% and 4.21%,respectively.Among them,3,5-di-O-caffeoylquinic acid was the highest content compound in sample Ⅰ.The GC-MS results showed that there were36 compounds in the dandelion sample Ⅶ.The higher content components successively were9,12,15-octadecanoic acid(Z,Z,Z)(27.69%),9,12-octadecadienoic acid(Z,Z)(19.99%),hexadecanoic acid(15.54%),3,7,11,15-tetramethyl-2-hexadecene-1-alcohol(9.72%),campesterol(6.20%)and octadecanoic acid(3.99%).The sum of the relative contents of the 6substances accounted for 73.42% of the sample Ⅶ.(3)The minimum inhibitory concentration(MIC)of dandelion sample Ⅰ on C.albicans and G.vaginalis were 32.0 mg/m L.The results of scanning electron microscopy revealed that sample Ⅰ could destroy the integrity of C.albicans and G.vaginalis cell membrane,causing leakage of intracellular substances and disorder of cell metabolism.The antibacterial kinetics analysis showed that the inhibitory effect of sample Ⅰ on C.albicans and G.vaginalis were mainly acted on its growth during the exponential phase,which led to a significant reduction in its growth rate.Membrane permeability study discovered that sample I increased the permeability of C.albicans and G.vaginalis membranes,resulting in large amounts of nucleic acids inside C.albicans and G.vaginalis,which affected C.albicans and G.vaginalis metabolism and growth.Found by infrared spectroscopy,sample Ⅰ maybe destroyed the glycosidic bond of C.albicans cell wall β-(1-3)-D glucan and changed the cell wall structure,thereby inhibiting cell growth.Therefore,dandelion sample I can destroy cell membrane of C.albicans and G.vaginalis,and may also affect the synthesis of C.albicans cell wall glucan.Then,cell growth is inhibited and eventually leads to its death.