Study On Anti-tumor Active Substance Of Patrinia Heterophylla Bung

Objective:Patrinia heterophylla Bunge is a kind of perennial herb,its rhizome and root are mixed used with Patrinia scabra Bunge as traditional Chinese medicine named Mutouhui.Mutouhui was used to treat acute or chronic leukemia in the clinical practice,it also exists in many antitumor traditional Chinese medicine.Patrinia heterophylla Bunge is the main variety of Mutouhui while the study on its chemical components and pharmacologic action is not sufficient.This article aim to isolate and extract chemical extracts from the Patrinia heterophylla Bunge by using a chromatographic separation technique such as silica gel column,medium pressure reversed-phase column and Sephadex LH-20.And the structure of the compounds were identified on the basis of physicochemical properties and spectral data ¹H-NMR,¹³C-NMR and HR-ESIMS analyses.The toxic effects of each extract and compound on cell growth were determined by CCK-8kit.Methods:The dried whole plant of P.heterophylla Bunge was crushed and then extracted two times with 75%EtOH under reflux.Concentrated under reduced pressure,the residue was suspended in water and partitioned successively with petroleum ether,CHCl₃,EtOAc and n-BuOH three times.The organic phase was collected and evaporated to afford four parts.The chromatographic separation technique was used to isolating different compounds,such as silica gel column,medium pressure reversed-phase column and Sephadex LH-20column.?ESI-MS?and nuclear magnetic resonance spectroscopy?NMR?analysis to determine the structure of the compound.The human leukemia cell-HL-60,human chronic myeloid leukemia cell-K562 and human T cell leukemia cell-Jurkat were cultured adding five different parts of the plant and compounds,and DDP was used as positive drug.CCK-8 method was used to determine the inhibitory effect of different polarities and different monomers on cell growth.The preliminary anti-tumor activity was evaluated by IC₅₀.Results:Eleven compounds were isolated from chloroform and ethyl acetate extracts,and nine of them were identified by spectral data,namely,Narcotic Alcohol A???,1',3'-Dioxane[6,7]cyclopenta[c]pyran-3?1H?-one,hexahydro-2',2',4-trimethyl-,?4R,4aS,6R,7R,7aS????,?4S,4aS,6S,7S,7aS?-6-hydroxy-7-?hydroxymethyl?-4-methylhexahydrocyclopenta[c]pyran-3?1H?-one???,daucion 3-O-?-L-arabinose?1?4??-D-glucopyranosyl oleanolic acid28-O-?-D-glucopyranose?1?6?-?-D-glucopyranoside???,3-O-?-L-pyranose arabinose?1?4??-D-glucopyranosyl oleanolic acid 28-O-?-D-glucopyranose???,rhamnocitrin3-O-[?-L-r h a m n o p y r a n o s y l?1?4?-O-?-L-r h a m n o p y r a n o s y l?1?6?]-?-D-galactopyranoside???,Daucosterol???,ursolic acid???,?-sitosterol???.Among them,compound II andcompound III are the new iridoid compounds,the triterpenoid saponins compounds?and?,the flavonoid compound?and iridoid c o m p o u n d?w e r e i s o l a t e d f r o m t h i s g e n u s f o r t h e f i r s t t i m e.Compounds?and?showed significant inhibitory effects on HL-60,K562 and Jurkat cells,and other compounds had different inhibitory effects on the three leukemic cells.Compounds I and IV had no inhibitory effect on Jurkat cells.Compound?had little inhibitory effect on K562 and Jurkat cells,and compound?was not effective for the growth inhibition of three cells.At the same time,the inhibitory effects of the compounds on the cells showed a dose-dependent effect,with the increase of drug concentration,the the toxicity of cells be stronger.Compared the anti-tumor activity of compounds II,IIIwith?,it is presumed that cyclopentadieno[c]pyran and the 4-position methyl structure are effective groups for the inhibition of toxicity of the iridoid terpenoids,there is no correlation between the effect of pharmacological effects and 4-CH₃ cis or trans.Conclusion:At present,nine compounds were isolated from the P.heterophylla Bunge,including three iridoid terpenoids,three triterpenoid saponins and aglycone,one flavonoid and?-sitosterol and daucosterol.Compounds?and?showed better inhibitory effect on different leukemia cells,especially compound?inhibited the proliferation of chronic myeloid leukemia cell line K562 better than the positive drug?DDP?.In order to determine the material basis of anti-leukemia effect,we need to isolate and identify more iridoid and study the structure-activity relationship.Although the monomeric compounds showed in vitro toxicity to leukemic cell lines,the traditional Chinese medicine composition was complex and its pharmacological effects were characterized by multiple targets and multidirectional regulation.The exact mechanism of anti-tumor effect of chemical constituents need further study.