Preliminary Studies On Chemical Constituents And Anti-inflammatory And Anti-hepatocarcinoma Activities In Vitro Of Ethly Acetate Extract From Viburnum Taitoense Hayata

Objective:To investigate the chemical compositions and isolate the chemical constituents of the ethyl acetate extract from Viburnum taitoenes Hayata,andfurthermoretestingtheiranti-inflammatoryand anti-hepatocarcinoma activities in vitro,which would further enriched both the research data of genus Viburnum and the theory of drug use in Viburnum taitoenes Hayata.Methods:The ethyl acetate extract of Viburnum taitoenes Hayata?JME?was prepared by immersing with ethyl acetate at room temperature.The contents of total triterpenoids,total phenolic components and total flavonoids of JME were determined by colorimetry,respectively.The cell model which used to evaluate the in vitro anti-inflammatory activity of JME was established by inducing RAW264.7 mouse macrophages with lipopolysaccharide?LPS?release nitric oxide?NO?,tumor necrosis factor?TNF-??,interleukin-1?and interleukin-6?IL-6?;the effect of JME on cell viability was examined by MTT method.The chemical constituents of JME were studied with comprehensive application of repeated silica gel column chromatography?CC?,Sephadex LH-20 CC and so on.The chemical structures of the isolated compounds were identified on the basis of physicochemical properties and modern spectroscopic methods.Also,the in vitro anti-inflammatory activity of the chemical constituents from JME were evaluated for their inhibitory effects on the NO production in LPS-induced RAW264.7 mouse macrophages,and their effects on cell viability were examined by MTT method.Besides,for the triterpenoids from JME,their cytotoxicity on human hepatoma cell line Bel 7402 were tested by MTT assay,which would help in evaluating their anti-hepatocarcinoma activities.Results:The chemical composition investigation of JME showed that the content of total triterpenoids,total phenolic components and total flavonoids in JME were 14.96?0.17%,3.46?0.04%,2.38?0.04%,respectively.The in vitro anti-inflammatory activity experiments showed that JME could inhibit the release of NO in a concentration-dependent manner with an IC₅₀ value of 60.08?1.10?g/mL?P<0.01?,and also inhibit the release of inflammatory factors such as TNF-?,IL-1?and IL-6,which exhibited certain anti-inflammatory activity in vitro.15 compounds were isolated from JME,namely:3?,6?-dihydroxyoleana-11,13?18?-dien-28-acid?1?,3?-hydroxy oleana-11,13?18?-diene-28-acid?2?,12-ene-oleana-28-acid-3?-palmitate?3?,3?-acetyl cocodiol?4?,corosolic acid?5?,uvaol?6?,ursolic acid?7?,quercetin?8?,quercetin-3-O-?-L-rhamnoside?9?,kaempferol?10?,7-hydroxy-6-methoxycoumarin?11?,trans-coumaric acid?12?,daucosterol?13?,?-sitosterol?14?,palmitic acid?15?.Among them,compound 1 was a new compound,compounds 2-5,9,11 were isolated from genus Viburnum for the first time,and all the compounds were obtained from the species V.taitoenes Hayata for the first time.Compounds 1,5,8,10 showed inhibitory effects on NO production in LPS-stimulated RAW264.7 macrophages with IC₅₀0 values of44.16±0.91?M,25.52±0.56?M,25.46±0.62?M,54.11±0.40?M,respectively;Compounds 3 and 7 showed strong cytotoxicity on RAW264.7macrophages while compounds 2,4,6,9,11-12 were inactive with inhibition rate<50%at the highest concentration tested?60?M?.Compounds 3-5,7showed different levels of cytotoxicity on human hepatoma cell line Bel 7402with IC₅₀ values of 52.43±0.61?M,58.41±0.55?M,24.29±0.68?M,39.58±3.47?M,respectively;while compounds 1-2,6 were inactive with inhibition rate<50%at the highest concentration tested?60?M?.Conclusion:In this paper,the chemical constituents and biological activities of V.taitoenes Hayata were studied and 15 compounds were isolated,one of which was a new compound and six of which were isolated from genus Viburnum for the first time.Pharmacological experiments showed that JME can inhibit the production of NO on LPS-induced RAW264.7 macrophages in a concentration-dependent manner,and inhibit the release of inflammatory factors such as TNF-?,IL-1?and IL-6,which exhibited certain anti-inflammatory activity in vitro.Compounds 1,5,8,10 showed inhibitory effects on NO production in LPS-stimulated RAW264.7 macrophages and compounds 3-5,7showed different levels of cytotoxicity on human hepatoma cell line Bel 7402.