Study On The Purification Of Total Saponin From Clematis Chinensis Osbeck And The Effects On Acute Promyelocytic Leukemia NB4 Cell Line In Vitro

Objective: Study on the purification of total Saponin from Clematis chinensis Osbeck and the effects on Acute Promyelocytic Leukemia NB4 cell line in vitro to provide theory evidence for clinical application. Methods: (1)To establish a method for the separation and purification of total saponin from Radix Clematidis by macroporous absorptive resin and detect the content of the extraction. (2)The NB4 cells were independently treated with ATRA and total saponin combined with differernt concentrations in five days.The proliferative inhibition rates of NB4 cell induced by total saponin and ATRA were calculated in every day. Cell differentiations were demonstrated by Giemsa stain,transmission electron microscope(TEM) and NBT reduction assay at different time. The expressions of CD11b and CD33 and apoptosis rates induced by these drugs were measured by flow cytometry(FCM) at different time. The cell apoptosis rate induued by these drugs were measured by flow cytometry (FCM).Results: (1) The RSDs of precision testing, and reproducibility testing in purifications of total saponin from Clematis using macroporous resin, were 0.85% and 1.35% respectively. Total saponins of Radix Clematidis were enriched in the 70 % ethanol eluted solution fraction, and obtained with a yield rate of 23.14±0.12%, and the recovery rate reached 4.55% with the RSD of 0.89%. (2) Clematis Total Saponins on NB4 cells significantly inhibited the growth with a dose-effect dependence;But during our experimental condition, NB4 cells treated had revealed no significant role in the differentiation rate induced, but the mature granulocyte cell ratios were increased along with the increased concentrations; NB4 cells were significantly induced to apoptosis, and the induced-apoptosis rates were time dependent and dose dependent in the range of 25μg/ml to 100μg/ml. Conclusion: (1)The effective composition total Saponin in the Radix Clematidis was efficaciously extracted by macroporous absorptive resin.(2) The total Saponin can obviously inhibit the proliferation of NB4 cells and induced to apoptosis, While no statistical significance in the differentiation phenomenon be observed...