Studies On The Extraction Of Total Saponins From Clematis Florida Var.plena D.and Its Antitumor Activity

Clematis florida var.plena D.is a cultivated varieties from Ranunculaceae Clem atis,the root and rhizome is the medcinal parts which is widly uesd in east part of Fuji an,called twelve hour by the She nationality pepole.According to the clinical experien ce of local people,Clematis florida var.plena D.has certain curative effect on cancer,and some medical effect on diuresis,regulating the flow of vital energy and remove t he obstruction toit,laxative,detoxification,and so on.Our study focus on the pharmacologically active ingredient and the mechanism of Clematis florida var.plena D.in antitumor,aims to exploit the medicinal resources of national and guide the clinical medication.The optimization of the extraction process of total saponins from Clematis florida var.plena D.by ultrasonic method has been determined by response surface methodology after discussing the effects of time,solid-liquid ratio,temperature,ultrasonic power,times and ethanol concentration on the extraction yield of total saponins based on the single-factor experiments.The optimum extraction yield was 16.06 mg/g when the temperature,time and ethanol concentration were 53 ?,66 min,and 62%respectively,which was well matched with the predictive extraction yield 16.12mg/g.This study showed the process was feasible and reproducible.An anti-tumor system of the Clematis florida var.plena D.total saponins was established in vitro evaluation,aiming to observe antitumor spectrum and human esophageal carcinoma EC9706 cell line inhibition.The results show these saponins could inhibt the proliferation of human esophageal squamous cell carcinoma(EC 9706),human hepatoma cell line(HepG-2),human oral cancer cell line(KB)and human gastric cancer cell line(BGC-823).Among them,the max inhibition rate(81.5%)was detected in EC 9706 cells,moreover,the relationships of dose-effect and inhibition-time were found.After EC 9706 cells was exposed to the saponins,higher cell refraction,enhanced cell membrane permeability changes,more nuclear chromatin condensation,and more apoptotic body induced by DNA breakage were observed using cell morphology and nuclear fluorescence staining.The saponins could also induced EC9706 cells apoptosis after flow cytometry analysis.Our results first demonstrated that the Clematis florida var.plena D.total saponins could inhibit the proliferation of the EC9706 cells,moreover,these results provided reference information for the anti-tumor drug screening and clinical medication mechanism of the Clematis florida var.plena D.