曇花及台灣穗花杉之活性成分研究

本研究乃將曇花(Epiphyllum oxypetalum Haw)與台灣穗花杉(Amentotaxus formosana Li)分別進行成分分離及生物活性探討。將曇花(E. oxypetalum)花朵利用乙酸乙酯萃取並進行成分分離，得到十個化合物，其中包含九個已知化合物，分別為：acetoxymethylfurfural (1)，1,1'-di-a-furaldehydic dimethyl ester (2)，b-sitosterol (3)， 3b-hydroxy-24-methylene-cholest-5-ene (4)，5-hydroxy- methylfurfural (6)，kaempferol (7)，kaempferol-3-O-b-D-gluco- pyranoside (8) ，stigmasterol-3-O-b-D-glucopyranoside (9)，kaempferol- 3-O-L-rhamnopyranosyl-(1à6)-b-D-glucopyranoside (10) 及一個新天然物3b-acetyl-24-methylenecholest-5-ene (5)。將以上所得到的部份化合物分別進行抗發炎活性篩選，結果顯示，2，4，8 及10並不呈現抗發炎 (anti-inflammatory effect)作用；7對肥胖細胞 (mast cell)之脫顆粒 (degranulation)反應並無抑制作用，但對嗜中性白血球 (neutrophil)以fMLP/CB (N-formylmethionyl- leucyl-phenylalanine/ cytochalasin B)所誘發之b-glucuronidase及lysozyme釋放具有很強的抑制作用，其IC50值分別為18.5 ± 1.0及21.2 ± 2.1 mM，比所使用的positive control，trifluoperazine作用還強；對小神經膠質細胞 N9 (murine microglial cell line N9)以LPS/IFN-g (lipopolysacharide/ interferon-g)誘發之TNF-α (tumor necrosis factor α)的形成活性亦具顯著之抑制活性，其在30 mM濃度下具有43.3 ± 10.1%之抑制活性; 5對嗜中性白血球 (neutrophil) 以fMLP/CB所引發之b-glucuronidase釋放具有顯著的抑制作用，其IC50值為23.0 + 0.7。台灣穗花杉(A. formosana)的葉及心材利用氯仿萃取並進行成分分離，於葉部分離得到十一個化合物，其中包含五個已知化合物：b-sitosterol (3)，(+)-ferruginol (17)，(+)-sugiol (18)，7a-hydroxy-8,12-abietadien-11,14-dione (19)，blumenol-A (20)，五個新化合物 (3b,23S)-3,23-dimethoxy-24-methylene-9,19-cyclolanostane (12)，(3b,23S)-3,23-dimethoxy-24-methylenelanost-8-ene (13)，(3b,23R)- 3-methoxy-24-methylenelanost-8-en-3-ol (14)，(23S)-23-methoxy-24- methylene-lanost-8-en-3-one (15)，(3b,24x)-3-methoxy-24-methyl- 9,19-cyclolanostan-25-ol (16)，及一個新天然物3b-methoxy- 24-methylene-9,19-cyclolanostane (11)。於心材分離得到十個化合物，其中包含七個已知化合物：7-hydroxycadalin (21)，coniferaldehyde (22)，amentotaxin BA (25)，2-benzo[1,3]dioxol-5yl-3-methoxy- acrylic acid methyl ester (26)，piperonylic acid (27)，ent-8(14),15- sandaracopimara-dien-2a,18-diol (29)，sandaracopimaradien-2a,18,19- triol (30)，二個新骨架化合物amentotaxin WA (23) 及amentotaxin WB (24)，及一個新化合物2-benzo[1,3]dioxol-5yl-3-methoxy-acrylic acid (28) 。將以上所得部分化合物: 17a，18，23及皮部所得之6a-hydroxy-ent-kaur-16-en-15-one及ent-kaur-16-en-15-one，分別進行細胞毒殺活性(cytotoxic activity)篩選，結果顯示，6a-hydroxy-ent-kaur-16-en-15-one及ent-kaur-16-en-15-one 對人類肝癌細胞(Hep G2、Hep 3B)、人類直腸腺癌細胞( HT-29、HCT 116)具有很顯著的細胞毒活性； 17a對人類肝癌細胞(Hep G2、Hep 3B) 及23 對人類肝癌細胞(Hep G2) 及人類直腸腺癌細胞(HT-29)皆具邊際效應(marginal effects)的活性。

關鍵字

台灣穗花杉；曇花；抗發炎；細胞毒殺活性

並列摘要

In a research of bioactive constituents of Epiphyllum oxypetalum Haw, including nine known compounds, acetoxymethylfurfural (1), 1, 1'-di-??-furaldehydic dimethyl ester (2), ??-sitosterol (3), 3??-hydroxy- 24-methylenecholest-5-ene (4), 5-hydroxymethylfurfural (6), kaempferol (7), kaempferol-3-O-??-D-glucopyranoside (8), stigmasterol-3-O-??-D- glucopyranoside (9), kaempferol-3-O-??-L-rhamnopyranosyl-(1??6)- ??-D-glucopyranoside (10), and a new natural product, 3??-acetyl-24- methylenecholest-5-ene (5), were isolated and their structures were determined by NMR and MS studies. Compound 5 showed significant inhibitory effects on the release of ??-glucuronidase from rat neutrophils with IC50 values of 23.0 ± 0.7 ?嵱 after stimulated with fMLP/CB (N-formylmethionyl-leucyl-phenylalanine / cytochalasin B) with IC50 values of 23.0 ± 0.7 ?嵱. Compound 7 indicated potent inhibitory effects on the release of ??-glucuronidase and lysozme from rat neutrophils stimulated with fMLP/CB with IC50 values of 18.5 ± 1.0 ?嵱 and 21.2 ± 2.1 ?嵱, respectively. Compound 4 also showed significant inhibitory effects on TNF-α(tumor necrosis factor α) formation from N9 cells stimulated LPS/IFN-?? (lipopolysacharide/ interferon-??). These results indicated that 5 and 7 were active constituent of E. oxypetalum. As part of a continued search for bioactive constituents from formosan plant, five known compound, ??-sitosterol (3) ，(+)-ferruginol (17)，(+)-sugiol (18)，7??-hydroxy-8,12-abietadien-11,14-dione (19)，blumenol-A (20), five new compounds, (3??,23S)-3,23-dimethoxy-24- methylene-9,19-cyclolanostane (12)，(3??,23S)-3,23-dimethoxy-24-methyl- enelanost-8-ene (13)，(3??,23R)-3-methoxy-24-methylenelanost-8-en- 3-ol (14)，(23S)-23-methoxy-24-methylene-lanost-8-en-3-one (15)， (3??,24??)- 3-methoxy-24- methyl-9,19-cyclolanostan-25-ol (16) and a new natural product, 3??-methoxy-24-methylene-9,19-cyclolanostane (11) were isolated from the leaves of Amentotaxus formosana Li. Seven known compounds, 7-hydroxycadalin (21)，coniferaldehyde (22)，amentotaxin BA (25)，2-benzo[1,3]dioxol-5yl-3-methoxy- acrylic acid methyl ester (26)，piperonylic acid (27)，ent-8(14),15-sandaracopimara-dien-2??,18- diol (29)，sandaracopimaradien-2??,18,19-triol (30), two novel terpenoids with a novel skeleton, amentotaxin WA (23) and amentotaxin WB (24), and a new compound, 2-benzo[1,3]dioxol-5yl-3-methoxy- acrylic acid (28) were isolated from the heart woods of A. formosana. Their structures were elucidated from spectroscopic data, and the structure of 13 was further confirmed by X-ray crystallographic analysis. The cytotoxicity of 17a and 18, 23, and 6??-hydroxy- ent-kaur-16-en-15-one and ent-kaur-16-en-15-one, isolated from the leaves, heart woods, and barks of A. formosana, respectively, were studied against a number of cancer cell lines. Compounds 6??-hydroxy- ent-kaur-16-en-15-one and ent-kaur-16-en-15-one showed significant cytotoxic activities against the Hep G2, Hep 3B, HT-29, and HCT 116. Compound 17a and 23 showed marginal activity against Hep G2 and Hep 3B, and Hep 3B and HT-29, respectively, while compound 18 did not show significant cytotoxic activity against the cell lines.