Ethyl acetate fraction in ethanol extract from root of “Dai-Bai-Jie” (Marsdenia tenacissima): anti-tumor activity in A549 cancer cells

Abstract

OBJECTIVE

To evaluate the anti-tumor activity of ethyl acetate fraction (EFA), extracted with ethanol from the root of “Dai-Bai-Jie” in A549 cancer cells and its underlying mechanism.

METHODS

“Dai-Bai-Jie” was extracted with 95% ethanol-aqueous (DBJ-1), 50% ethanol-aqueous (DBJ-2), and water (DBJ-3) by reflux method. 95% ethanol-aqueous extract was separated byethyl acetate (EFA) and n-butyl alcohol (DBJ-5), consecutively. The SRB method was used to evaluate the cytotoxic activity. Annexin V-FITC staining was applied to observe the apoptosis and analyze the cell cycle activated by EFA in A549 tumor cell. Western blot was used to detect the apoptosis/related proteins expressions. A549 tumor cellsbearing nude mice model was employed to measure the tumor volume, mice weight, and tumor inhibition ratio in order to verify the antitumor activity in vivo.

RESULTS

DBJ-1 and EFA showed better cytotoxic activity on A549 tumor cells with IC₅₀ 25 and 3.5 µg/mL, respectively. EFA can exhibit the proliferation, arrest cell cycle at G0/G1 phase, and induce apoptosis in A549 tumor cells in vitro. The mechanisms of apoptosis induced by EFA may be associated with decreasing Bcl-2 protein expression and increasing p53, Bax, Caspase-3, and Caspase-8 proteins expression. EFA also possessed significant anti-tumor efficacy in nude mice, and little toxicity was observed in the host.

CONCLUSION

EAF could induce A549 tumor cells apoptosis and G0/G1 cell cycle arrest. A549 tumor cells apoptosis induced by EAF may be associated with the decrease in the ratio of Bcl-2 and Bax mRNA levels, and increase in the expression of p53, Caspase-3, and Caspase-8 proteins.