Study On The Active Ingredients And Mechaniam Of Compound Sanhuang Decoction Against Candida Albicans

Chinese medicine believes that Candida albicans infection is due to dampness in the body Hot and damp stasis,coupled with Outside the toxin factor.Traditional Chinese medicine compound contains many kinds of active ingredients,has multi-faceted comprehensive utility,wide antibacterial spectrum,high efficiency and low toxicity.It is suitable for long-term prevention and application and has attracted more and more scholars' attention.Sanhuang Decoction from the "golden yi" volume fifteen,is composed of Coptis,Scutellaria and rhubarb Sanwei Chinese medicine,attending triple burner fever,irritability,language,constipation and other symptoms.Compound Sanhuang Decoction is a clinical side,laboratory pre-work according to the dry and dampness of the Sanhuang Decoction-based party,minus rhubarb plus cork,get better anti-Candida albicans compound Sanhuang Tang.This topic further study which of the effective components of the party have the role of inhibiting Candida albicans,and further study of these effective components inhibit Candida albicans mechanism of action.In this paper,the Sanhuang Decoction with the minimum inhibitory concentration as an index was separated by silica gel column chromatography,and the minimum inhibitory concentrations of the 10 components were determined.The best antibfungl components were screened out for further separation and purification.The antibfungl activity of Sanhuang Tang ethyl acetate layer petroleum ether:ethyl acetate=3:1(BWM-7)was the best,MIC is 0.03125mg/mL.The constituents were separated and purified.The constituents were identified by spectrum and physicochemical property.The compounds were identified as follows:Compound X,Compound Y,Compound Z.Secondly,the impact of BWM-7 on Candida albicans biofilm was investigated.The results showed that it could effectively inhibit the formation of C.albicans biofilm.Flocculation experiments and cell surface hydrophobicity experiments show that BWM-7 can reduce the flocculation ability of Candida albicans and cell surface hydrophobicity,inhibit the formation of Candida albicans biofilm reduce pathogenicity.BWM-7 treated Candida albicans after 1.5 h,Candida albicans survival rate has dropped significantly,and the role of fluconazole than Candida albicans survival rate is lower,indicating that BWM-7 against Candida albicans in vitro fungl onset time,and can inhibit the growth of Candida albicans within a certain period of time.The effect of BWM-7 on Candida albicans biofilm was examined by XTT,,crystal violet method,scanning electron microscopy,fluorescent dye and silica gel model.BWM-7 at 2 mg/mL significantly inhibited the formation of C.albicans biofilm.The effect of BWM-7 on the biofilm gene expression in candida albicans was detected by real-time quantitative PCR.The results showed that BWM-7 could significantly reduce the PLB2,LIP9,SAP5,Ndt80,EFG1 and BCR1 genes in C.albicans biofilms.Finally,the mechanism of inhibiting Candida albicans at the cellular level by BWM-7 was studied.The results showed that compared with BWM-7 untreated group,macrophages were pretreated with 6.25 mg/mL and 12.5 mg/mL BWM-7 for 24 h,and Candida albicans stimulated macrophages for 3 h.The results of Swiss-Giemsa staining,the number of macrophages phagocytizing yeast-type Candida albicans was 17%,indicating that BWM-7 can increase phagocytosis of Candida albicans by macrophages.Western blot detection of macrophages NF-?B,iNOS protein expression,the results show that Candida albicans stimulation of macrophages,the macrophages of the protein was significantly increased,the inflammatory response in cells increased,and HHH-7 pretreatment group,macrophages in each protein expression was significantly decreased compared with untreated.ELISA kit was used to detect the expression of cytokines IL-2,IL-12,IL-6 and TNF-? in cell culture supernatants.The results showed that after macrophages were stimulated by Candida albicans,the inflammatory response increased.after BWM-7 preconditioning macrophages,which was significantly lower than that in untreated cells,which was close to the normal expression of cytokines.The experimental results show that the antifungl effect of BWM-7 is the best by separating and purifying the antifungl active ingredients in Sanhuang Decoction.And studied its inhibitory effect on Candida albicans in vitro,as well as the cellular level.BWM-7 can stimulate macrophage phagocytosis after Candida albicans stimulated macrophages,and can reduce inflammation caused by Candida albicans infected macrophages reaction.BWM-7 not only antifungl effect but also improve the function of macrophages and reduce macrophage inflammatory response.