Study On The Chemical Constituents And Anti H5N1 Activity Of Campylotropis Hirtella(franch.)schindl

Since the first report of human infection with the H5N1 bird flu virus in 1997, the number of human infection with H5N1 avian influenza have been more than 700 people according to WHO statistics as of July 2014, and the mortality rate as high as 60%. Because of its variability and resistance, there are not yet appropriate and efficient drugs for clinical treatment.Due to herbs contains novel structure, looking for anti viral activity of compounds from herbs is becoming more and more popular. Our previous study has filter out some herbs which have activity against H5N1 virus from more than 80 kinds of herbs.We selected Campylotropis hirtella(Franch.)Schindl as the researching object, and study on the chemical constituents of ethyl acetate fraction of 60% ethanol extract.Campylotropis hirtella(Franch.)Schindl,commonly known as Dahehong, Xiudingzi, Yougen, Bianzaojiao, is a plant belongs to Campylotropis hirtella pulse family, which medicinal part is the roots. It widely grows in the province of Yunnan, Guizhou and Sichuan in China, and has the pharmacological activity of hemostasis, odynolysis and antiinflammatory. It contains lignans, coumarins, flavonoids, tannins and so on.To study the chemical constituents of Campylotropis hirtella(Franch.)Schindl, recrystallization, column chromatography on silica gel, ODS; Sephadex LH 20 chromatography techniques were used. 29 compounds were isolated, and the structures of the compounds were identified by physicochemical properties and spectral data. In which 25 compounds were identified as fllows: Hedysarimcoumestan I(1), 4′,6′,7 trihydroxy 2′ methoxy 3 arylcoumarin(2), Hedysarimcoumestan B(3), isotrifoliol(4), quercetin(5), kaempferol(6), dihydrokaempferol(7),(+) catechin(8), umbelliferone(9), Aesculin(10), diosgenin(11), rutin(12), sophororicoside(13), daidzein(14), orobol(15), ferulic acid(16), maltol(17), salicylic acid(18), 1,3 dihydroxy anthraquinone(19),(2R,3R) 3,4′ dihydroxy 7 methoxy 8 methyl flavan(20), daucosterol(21), β sitosterol(22),(2R) 4′,7 dihydroxy flavan(23), 3,4 dihydroxybenzoic acid(24), p hydroxybenzoic acid(25). Compound 1 and 2 were new compounds; compounds 3、4、10、13、15、19、20、23 were isolated from this genus for the first time.The total extract, n Bu OH phase, ethyl acetate phase and CHCl3 phase, and the compounds of Campylotropis hirtella(Franch.)Schindl were tested for anti HPAI in vitro, using MDCK cell lines infected by H5N1 and employing cytopathic effect(CPE) method. The total extracts of Campylotropis hirtella(Franch.)Schindl possess potent cytotoxicity at the effective dose of 150 g/m L. Furthermore, n Bu OH phase and ethyl acetate phase showed much better cytotoxicity against H5N1 virus than that of the total extract of Campylotropis hirtella(Franch.)Schindl. In screening for anti H5N1 virus cytotoxic agents from all compounds of Campylotropis hirtella(Franch.)Schindl, compounds 15 and 14 exhibited potent anti H5N1 virus activity in MDCK cell cultures which did not show obvious cytopathic effect on cell viability, proliferation or metabolism. At the effective dose of 50.0 and 200.0 g/m L, respectively.