Anti-Tumor Chemical Constituents From Patrinia Villosa(Thunb.)Juss

Herba Patriniae is a perennial erect herb,named for the strong smell of decay of its underground roots.It is widely distributed in Eastern Asia and North America.There are more than 20 species in Patrinia genus,which were first recorded in one of the classic works of traditional Chinese medicine,<Shennong herbal classic>.Herba Patriniae is widely distributed in all parts of China,and its tender leaves can be used as food and it belongs to the food and medicine homologous plant.According to Chinese Pharmacopoeia(1977 Edition),Patrinia scabiosaefolia Fisch.ex Trev.and Patrinia villosa(Thunb.)Juss.are authentic.However,due to the variety of Patrinia,its easiness to be confused in practical application,Herba Patriniae is not included in the following editions of Chinese Pharmacopoeia.In the clinical practice,Herba Patriniae is mainly applied to clear heat and remove dampness,detoxify and fester,promote blood circulation and remove blood stasis and etc..Modern pharmacological researches show that it has clear pharmacological activities of anti-inflammatory,anti-tumor,anti-bacterial,and anti-virus.There are many bioactive components in P.scabiosaefolia and P.villosa,including phenylpropanoids,flavonoids,terpenoids,saponins,volatile oils,etc.At present,more than 200 components have been isolated and identified,more than one-third of which have been reported for the first time by our research group.On the basis of previous studies,we further studied the chemical constituents of P.villosa,screened the anti-tumor activity of the chemical constituents,and preliminarily analyzed the related mechanism of the active constituents of P.villosa.In this study,the dried leaves of Patrinia villosa(Thunb.)Juss were selected as the research object,and 70% ethanol was used to extract the concentrated extract.After the extract was dissolved in water,light petroleum ether,dichloromethane,and butanol were used to extract and the dichloromethane phase was systematically separated.The extracts were separated by normal silica gel column chromatography,ODS column chromatography,and Sephadex LH-20 column chromatography.After purification by semi-preparative HPLC,thirteen monomer compounds were finally obtained.The structures of the compounds were identified by NMR and MS,and seven of them have been identified at present.Two of them were new compounds including Impecylone A(1)and 5,7-dihydroxy-2-(3-hydroxy-4-methoxyphenyl)-3-pentadecyl-4H-chromen-4-one(2);and five of them were known compounds,including Methyl(E)-3-(3,4-dihydroxyphenyl)acrylate(3)5-hydroxy-2-(4-hydroxyphenyl)-6,7-dimethoxy-4H-chromen-4-one(4)Ethyl(E)-3-(3,4-dihydroxyphenyl)acrylate(5)Methyl 2-(4-hydroxyphenyl)acetate(6)and(E)-3-(4-hydroxy-3-methoxyphenyl)acrylic acid(7).We further evaluated the cytotoxicity of the isolated monomers in vitro,and three cell lines,Hep G2,A549,and A2780,were selected as evaluation models.The results showed that compound 1 could selectively inhibit the activity of tumor cells,and had a significant inhibitory effect on the proliferation of Hep G2 and A549 cells,with the IC50 values of 38.37 μM and 29.22 μM,respectively.The effects of compound 1 on apoptosis and cell cycle of Hep G2 and A549 cells were studied by flow cytometry.Apoptosis experiments showed that this compound could promote the apoptosis process of the two kinds of cells,and significantly increase the early apoptosis rate of A549 cells from 1.91% to 7.83%.The results of cell cycle experiments showed that compound 1 could increase the number of Hep G2 cells in G2/M phase to 26.6%,and decrease the number of cells in G0/G1 phase to 54.9%,while the number of cells in S phase was almost unchanged.To sum up,In this study,thirteen compounds were isolated from Patrinia villosa(Thunb.)Juss.and seven structures were identified,two of which were new compounds.The studies of the anti-tumor activity showed that compound 1 can selectively inhibit the activities of Hep G2 cells and A549 cells in vitro.And the inhibition may be caused by the irreversible damage of the DNA or chromosome of tumor cells,leading to the process of cell proliferation to be blocked in the G2/M phase,and further induce apoptosis of the tumor cells.The research of this subject can lay a foundation for further exploitation of Patrinia villosa(Thunb.)Juss.