Analysis Of Anti-Respiratory Syncytial Virus Effective Components Of Hibiscus Mutabilis Leaf And Its Anti-viral Mechanism

Objective:At the cellm Lar level,the anti-RSV active sites of Hibiscus mutabilis Leaves were screened through in vitro anti-viral tracing technology,and the resm Lts of anti-RSV were screened.The in vivo experiments were used as the main research content to confirm the anti-RSV effects of the anti-RSV active parts of Hibiscus mutabilis leaves,and further study its antiviral mechanism.Preliminary investigation of the chemical composition of the active site.Methods:Separate and purify the water extract of Hibiscus mutabilis Leaves by D101 macroporous resin,m Ltrafiltration and Extraction technology,get different separation and purification parts.The in vitro antiviral effect of different separation and purification sites was explored through in vitro antiviral experiments,and the separation and purification sites with better anti-RSV effects were selected.The effective parts against RSV were further separated and purified by the method of silica gel column chromatography,and combined with the in vitro antiviral experiment,the most effective active parts against RSV were selected.According to the screening resm Lts,determine the main research objects.Through the addition of drugs at different times,the antiviral mechanism of the active parts of Hibiscus mutabilis Leaf was initially explored.Establish pneumonia model of RSV-infected BALB / c mice,and explore the antiviral effect of the active part of Hibiscus mutabilis Leaf Establish pneumonia model of RSV-infected BALB / c mice,combined with ELISA kit method to detect the content of TNF-α,TNF-β,IFN-α and IL-4 in the serum of mice,and preliminary explore the immunity of the active parts of Hibiscus mutabilis Leaf Regm Lation effect.Establish a pneumonia model of RSV-infected BALB / c mice,detect the viral load,TLR3 and TLR4 expression levels in lung tissue by RT-PCR,and preliminary explore the effect of the active parts of Hibiscus mutabilis Leaf on the expression of RSV,TLR3 and TLR4 mRNA in mouse lung tissue.Combined with liquid-mass spectrometry technology,the chemical constituents of the active part of the leaves of Hibiscus mutabilis Leaf were preliminary explored.ResultFrom the point of view of antiviral index(TI value),the site with the best antiviral effect is the D101 macroporous resin 50% ethanol elution site(TI value is 43.17)and 25%elution site(TI value is 38.00)are better than positive the control drug ribavirin(TI value is 31.94).The effective part was further separated and purified by silica gel column chromatography.It was found that the silica gel column elution site M-18(ethyl acetate:methanol = 21: 4 eluted 50% ethanol site)had the best anti-RSV effect(TI value 66.17),so this part is the main research object.Through the experiment of adding drugs at different times,it was found that M-18 mainly played an antiviral role before the virus invaded the cell,that is,it played a preventive role against the virus.In vivo experiments showed that by comparing lung lesions,pathological slices,and virus re-separation among the groups,it was found that the active site of Hibiscus mutabilis Leaf M-18 can significantly reduce lung tissue lesions in mice,reduce lung index in mice,and reduce lung tissue in mice.Internal virus titer.Compared with the model group,the active site M-18 of Hibiscus can increase the concentration of IL-4 and IFN-α in the serum of mice,and reduce the concentration of TNF-α and TNF-β;Compared with the model group,the active site M-18 of Hibiscus can reduce the viral load in the lung tissue of mice,and down-regm Late the expression levels of TLR3 and TLR4.By referring to related literatures,rutin and kaempferol-3-O-[6"rhamnose(1-6)]-glucopyranoside were initially identified from the active part of M-18.ConclusionAfter being purified by D101 macroporous resin and silica gel column chromatography,the Hibiscus leaves have a good anti-RSV effect in vitro,which was verified by in vivo experiments and found to have a significant antiviral effect in vivo.In vitro experiments found that the active part of M-18 leaves M-18 mainly prevents the virus;In vivo experiments found that the active site of Hibiscus mutabilis Leaf M-18 may exert antiviral effects by reducing the content of inflammatory factors in mice serum,increasing the content of anti-inflammatory factors,and down-regm Lating the levels of TLR3 and TLR4 Subsequent experiments initially identified rutin and kaempferol-3-O-[6"rhamnose(1-6)]-glucopyranoside from the active parts of the leaves of Hibiscus.