Studies On The Constituents From The Leaves And Stem Bark Of Garcinia Bracteata And Their Antitumor Activities

In searching for the bioactive constituents from the genus Garcinia, 13 species of the genus Garcinia were screened via the growth inhibitory activities against HL-60 cell line, and TLC, HPLC-DAD, NMR and G-quadruplex binding experiments. Garcinia bracteata C. Y.Wu showed stronger growth inhibitory activity and G-quadruplex binding effect.G. bracteata, a plant belonging to the family of Guttiferae, is distributed throughout Thailand, Cambodia, and the southern part of China. Based on the review on the chemical constituents and pharmacological research of Garcinia, we launched a systematic study to investigate the chemical constituents in the ethanol extract of G. bracteata.By means of many different chromatographic methods such as silica gel, Sephadex LH-20,ODS column chromatography, preparative HPLC and preparative TLC, 35 compounds were isolated from the leave of G. bracteata. By physico-chemical data and spectral methods, thirty of them were identified as garcibracteatones A-C (1-3), isobractatin B(4), 13-hydroisobractatin A (5), 13-hydroisobractatin B (6), 8a-methoxy-8, 8aα-dihydrobractatin (7),8a-ethoxy-8,8α-dihydrobractatin (8),garcibracteatone D (9),8p-methoxy-8,8aβ-dihydroneobractatin (10), bractatin (11), 3-O-methylbractatin (12),isobractatin (13), neobractatin (14), 3-O-methyoneobractatin (15), neoisobractatin A (16),neoisobractatin B (17), bracteaxanthone Ⅶ (18), bracteaxanthone Ⅷ (19), bracteaxanthone Ⅸ(20), allanxanthone A (21), 1,6-dihydroxy-2,4-diprenylxanthone (22), 5-O-methylxanthone V1(23), 10- O-methylmacluraxanthone (24), pinetoxanthone (25), subelliptenone H (26),toxyloxanthone A (27), 1,2,8-trihydroxyxanthone (28), 3β-hydroxy-glutin-5-ene (29),taraxerol (30). Among them, compounds 1 ～10］18～20 are determined as new compounds,and compounds 1-3 were novel highly rearranged prenylxanthones. The absolute configurations of all the cage xanthones were studied with the aid of NOESY and CD spectra, and determined by X-ray for the first time.45 compounds were isolated from the stem barks of G. bracteata and characterized as 1,4,5,6-tetrahydroxyxanthone (31), bracteaxanthone Ⅲ-Ⅳ (32～35), 1,4,5,6-tetrahydroxy-7-(3-methylbut-2-enyl)xanthone (36), 1,4,6-trihydroxy-5-methoxy-7-(3-methylbut-2-enyl)xanthone (37), garcinenone A (38), 1,4,5,6-tetrahydroxy-7,8-di(3-methylbut-2-enyl)xanthone(39), garciniaxanthone E (40), 6-desoxyjacreubin (41), 6-desoxyjacareubin (42),symphoxanthone (43), 1 -O-methylsymphoxanthone (44), garciniaxanthone H (45),12b-hydroxy-des-D-garcigerrin A (46), globuxanthone (47), morusignin 1 (48), 1,7-dihydroxyxanthone (49), 1, 5-dihydroxy-3-methoxyxanthone (50), 1,5-dihydroxy-3,8-dimethoxyxanthone (51), 1, 4-dihydroxy-5, 6-dimethoxyxanthone (52), subelliptenone G(53), 1,2,5-trihydroxyxanthone (54), 2,5-dihydroxy-l -methoxyxanthone (55), 2,6-dihydroxy-1, 5-dimethoxyxanthone (56), 1,3,5,6-tetrahydroxyxanthone (57), 1,3,6,7-tetrahydroxyxanthone (58),1, 2, 5-trihydroxy-6-methoxyxanthone (59),1,5-dimethoxy-3－hydroxyxanthone (60), 1,3,7-trihydroxyxanthone (61), 1,3,5-trihydroxyxanthon (62),garciduol A (63), apigenin (64), luteolin (65), kaempferol (66), quercetin (67),6-(3-methylbut-2-enyl)-apigenin (68), rutin (69), montroumarin (70), volkensiflavone (71),morelloflavone (72), gustastatin (73), garcinielliptone (74), morolic acid acetate (75).Among them, compounds 31-35 are determined as new compounds. Compounds 21, 30,70,73 are isolated from genus Garcinia for the first time.The antitumor activities of 14 caged polyprenylated xanthones isolated from the leave of G. bracteata were tested. All of them showed potent cell growth inhibitory activities against HL-60 and K562 cells, in which compounds 1 and 12 were the most potent, respectively.The structure-activity relationship of caged polyprenylated xanthones was discussed. All of the isolated 26 simple xanthones were evaluated for their cell growth inhibitory effects against HL-60 cell line. Almost all of the tested compounds exhibited moderate inhibitory activities against the HL-60 cell line, with GI50 values ranging from 2.8 to 47.9 μM. A preliminary structure-activity relationship was discussed. Of these compounds, the prenylated xanthones exhibited more potent effects, of which 39, 40, and 47 are the most effective compounds with GI50 values of 2.8, 3.1, and 3.4 μM, respectively.The apoptotic effects of compounds 4, 12, 15 in HL-60 cells were also examined by morphological observation after AO and EB staining and FACS analysis. Compounds 4, 12,15 induced HL-60 cells to undergo apoptosis in a dose- and time-dependent manner.Through 1H-NMR experiment, compound 37 was found to bind with G-quadruplex DNA,of which binding pattern was studied to explore the possible mechanism of activity via molecular docking models by computer calculation.