| The biological properties of anthraquinones include antioxidation, antiinflammatation, antibacterial, antiproliferation of tumor cells and induction of apoptosis. Xanthophytum Attopvensis Pierre (Rubiaceae), which has been used in China for the treatment of icterus hepatitis and contains eight kinds of anthraquinones, is distributed in Guangdong, Hainan and Yunnan, etc.We isolated eight anthraquinones from Xanthophytum Attopvensis Pierre and the in vitro cytotoxicity of six anthraquinones against four human tumor cell lines HepG2, SGC-7901, Glc-82 and HL-60 by SRB assay was evaluated and their structure-activity relationships of them were analyzed. The growth curve assay, comet assay, FCM analysis and AO/EB double staining were used to investigate the inhibitory effect, DNA damage potential, cell cycle arrest and induction of apoptosis of damnacanthal on human leukemia HL-60 cells, respectively. Meanwhile, damnacanthal on human hepatoma HepG2 cells growth, cell migration, cytoskeleton and induction of apoptosis were investigated al well.The results of the experiments are as follows:(1) The results of SRB assay showed that 1,3-dihydroxy-2-aldehyde anthraquinone(1) and 1-mehtoxy-2-aldehyde-3-hydroxy anthraquinone (2) which do not bear the glycon group had higher cytotoxicity against four human tumor cell lines. Compared with the other four compounds which bear the glycon group, including 1-methoxy-2-hydroxymethyl-3-O-β-D-glucoside anthraquinone (3), 1-hydroxy-2-hydroxymethyl-3-O-β-D-xylosyl (1→6)-β-D-glucoside (4), 1-methoxy-2-hydroxy methylanthraquinone-3-O-β-D-xylosyl (1→6)-β-D-glucoside (5), 1, 3-dihydroxy-2- hydroxy methylanthraquinone-3-O-β-D-xylosyl (1→6)-β-D-glucoside (6), compounds 1 and 2 had about 4.7-12.7 times higher IC50 values.It is possibly because the sterically bulky glycon group makes the compound have higher stereospecific blockade so that the compound could not interact easily with its target molecules of cells.The cytotoxicity between compounds 1 and 2 was similar when C-1 site is replaced by–OH or–OCH3 group,indicating these two substituent groups contributed similarly to the cytotoxicity of the two compounds. Human tumor cell line HL-60 was more sensitive to treatment and the IC50 values were 1.2-2.5 times lower than the other three cell lines.(2) Damnacanthal had an inhibitory effect against the proliferation and DNA damage of HL-60 cells in a time- and dose- dependent manner. Compared with the control group of DNA damage, damnacanthal treated groups induced significant comet rates (P<0.01). G1 cell cycle arrest was observed when the cells were treated with 5, 10, 12.5 and 15μg mL-1 damnacanthal at 12-60 h. S and G2/M cell cycle arrest were when the cells were treated with 30 and 40μg mL-1 damnacanthal at 12-60 h, individually. Damnacanthal also exhibited a time- and dose- dependent manner of the induction of apoptosis. There was a significant difference (P<0.01) between the control and treated groups, except that of 2.5μg·mL-1 damnacanthal at 24 h (P<0.05) and 48 h (P<0.01).(3) Damnacanthal showed a significant inhibitory effect against the proliferation of HepG2 cells in a time- and dose- dependent manner. Damnacanthal also inhibited the HepG2 cell migration in a time- and dose- dependent manner.With the elongation of treated time and the increase of damnacanthal concentrations, the adhesion capacity of cells decreased and cells became round thus they underwent apoptosis. Meanwhile, compared with the control groups, damnacanthal treated group (4-20μg mL-1) induced significant apoptotic cells at 24h, 48h, 72h, respectively (P<0.01).The conclusions are as follows:(1) According to the guidelines of NCI, compound 1 and 2 worth further investigation. In addition, compounds 3, 4, 5 and 6 all bear the glycon group at C-3 sites so further studies need to be conducted to prove whether the glycon group could decrease the cytotoxicity.(2) Damnacanthal exhibited a significant cytotoxic effect and induced proliferation inhibition and apoptosis against HL-60 cells. Therefore, it could be regarded as a potential drug to treat human leukemia.(3) When exposed to damnacanthal, the cytoskeleton of HepG2 cells changed, the adhesion capacity of the cells decreased and cells became round and the subsequent cell migration was inhibited, thus cells underwent apoptosis. However, studies about the specific mechanisms of damnacanthal treated cells need further investigation thus it could offer credible proofs for the potential anti-tumor drug damnacanthal.
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