瘤砂海葵衍生真菌株Arthrinium arundinis MA30之成分研究

Chemical Constituents from Palythoa sp. derived Arthrinium arundinis MA30

李元新(Yuen-Sing Lee)

指導教授 ： 李宗徽

國立臺灣大學/生命科學院/漁業科學研究所/碩士(2022年)

https://doi.org/10.6342/NTU202200789

摘要

本研究利用病原菌與病毒篩選活性平台 (Staphylococcus aureus, Candida albicans, Cryptococcus neoformans, Escherichia coli和Epstein-Barr virus) 來篩選具有抗菌和抗病毒潛力的海葵衍生真菌株。 實驗結果顯示，由瘤砂海葵 (Palythoa sp.) 單離出之衍生真菌株Arthrinium arundinis MA30之粗萃物具有抗 S. aureus、C. albicans 與EBV病毒的顯著效果，因而選定該菌株進行其天然物研究。接著，利用OSMAC (one strain many compound) 的方式進行了不同條件下的液態和糙米固態培養後，分離並純化此株真菌的次級代謝物，計獲得 32個化合物，再藉由各種光譜資料解析其結構，其中 6個為新化合物，分別為 arthrinoic acid (1)、hexylaconitic anhydride methyl ester (2)、iso-hydroxylated hexylitaconic acid (3) 、(3S,8R)-8-hydroxy-3-carboxy-2-methylenenonanoic acid (4)、arthripenoid G (5) 及arthripenoid H (6)，另外 26 個為已知化合物，分別為succinic acid (7)、sumiki's acid (8)、pyromucic acid (9)、4-(2-hydroxyethyl)benzoic acid (10)、4-hydroxyphenylacetic acid (11)、2-hydroxyphenylacetic acid (12)、methyl vanillate (13)、6,8-dihydroxy-3,4-dimethylisochromen-1-one (14)、3,4,8-THT (15)、2β-methyl-5-hydroxy-2H-1-benzopyran-4(3H)-one (16)、2β-methyl-4-methoxy-3,4-dihydro-2H-1-benzopyran-5-ol (17)、(+)-mellein (18)、(R)-2-hexyl-3-methylenesuccinic acid (19)、palmitic acid (20)、linoleic acid (21)、thalictric acid (22)、2-hexyl-3-methylmaleic anhydride (23)、TXIB (24)、anomalin A (25)、anomalin B (26)、1,3,5,6-tetrahydroxy-8-methylxanthone (27)、ergosterol (28)、cytochalasin E (29)、arthripenoid B (30)、arthripenoid C (31) 和arthripenoid F (32)。所有純物質在抗發炎活性平台的評估上，得知化合物5、6、27、30、31和32具有抑制小鼠神經膠細胞BV-2產生一氧化氮的 (nitric oxide, NO) 效果，於10 µM下抑制NO產生的比例介於80至100%之間，其中化合物27和31有不錯的活性且不具顯著的細胞毒性，與IC50分別為5.3 ± 0.6 和1.55 ± 0.4 μM。

關鍵字

瘤砂海葵衍生真菌 ； Arthrinium arundinis ； arthripenoid ； 天然物 ； 抗發炎 ； 細胞毒性

並列摘要

In an attempt to investigate bioactive fungal strains, preliminary field-sampling and bio-assays were conducted. The microorganisms used in antimicrobial and antiviral bio-assay platform included Staphylococcus aureus, Cryptococcus neoformans, Candida albicans, and Escherichia coli, as well as Epstein-Barr virus (EBV), respectively. From the macroalgae and sea anemone collected at northeastern coast of Taiwan (Badouzi), 220 fungal strains were isolated. Among these, Arthrinium arundinis MA30 from sea anemone was selected due to its significant inhibition against S. aureus, C. neoformans, and EBV. An OSMAC (one strain many compounds) strategy was applied, where Arthrinium arundinis MA30 was cultured in different environments, including liquid-state fermentation under shaking, liquid-state fermentation under aeration, and solid-state fermentation. As a result of isolation and purification on the fermented products, 32 compounds were identified and six of which were previously unreported compounds, including arthrinoic acid (1), hexylaconitic anhydride methyl ester (2), iso-hydroxylated hexylitaconic acid (3), (3S,8R)-8-hydroxy-3-carboxy-2-methylenenonanoic acid (4), arthripenoid G (5), and arthripenoid H (6), in addition of 26 known compounds, including succinic acid (7), sumiki's acid (8), pyromucic acid (9), 4-(2-hydroxyethyl)benzoic acid (10), 4-hydroxyphenylacetic acid (11), 2-hydroxyphenylacetic acid (12), methyl vanillate (13), 6,8-dihydroxy-3,4-dimethylisochromen-1-one (14), 3,4,8-THT (15), 2β-methyl-5-hydroxy-2H-1-benzopyran-4(3H)-one (16), 2β-methyl-4-methoxy-3,4-dihydro-2H-1-benzopyran-5-ol (17), (+)-mellein (18), (R)-2-hexyl-3-methylenesuccinic acid (19)，palmitic acid (20), linoleic acid (21), thalictric acid (22), 2-hexyl-3-methylmaleic anhydride (23), TXIB (24), anomalin A (25), anomalin B (26), 1,3,5,6-tetrahydroxy-8-methylxanthone (27), ergosterol (28), cytochalasin E (29), arthripenoid B (30), arthripenoid C (31), and arthripenoid F (32). The structures of these compounds were elucidated by comprehensive spectroscopic methods, and some of their absolute configurations were determined by Mosher’s reagent, circular dichroism, and single crystal X-ray diffraction. Of these compounds, 5, 6, 27, 30, 31, and 32 exhibited high inhibition rate towards nitric oxide (NO) production in mouse glial cells BV-2 cells, and the proportion of inhibiting NO production at 10 µM ranging from 80 to 100%. The IC50 values of the most active compounds 27 and 31 were 5.3 ± 0.6 and 1.55 ± 0.4 μM, respectively, without significant cytotoxicity.

並列關鍵字

Palythoa sp. derived fungi ； Arthrinium arundinis ； arthripenoid ； natural products ； anti-inflammation ； cytotoxicity

參考文獻

1. Ankisetty, S. and Slattery, M., Antibacterial secondary metabolites from the cave sponge Xestospongia sp.. Marine Drugs, 12. 10(5): p. 1037-1043.

2. Bao, J., He, F., Yu, J. H., Zhai, H. J., Cheng, Z. Q., Jiang, C. S., Zhang, Y. Y., Zhang, Y., Zhang, X. Y., Chen, G. Y., and Zhang, H., New chromones from a marine-derived fungus, Arthrinium sp., and their biological activity. Molecules, 2018. 23(8).

3. Carroll, A. R., Copp, B. R., Davis, R. A., Keyzers, R. A., and Prinsep, M. R., Marine natural products. Natural Product Reports, 2019. 36(1): p. 122-173.

4. Gao, S. S., Li, X. M., Du, F. Y., Li, C. S., Proksch, P., and Wang, B. G., Secondary metabolites from a marine-derived endophytic fungus Penicillium chrysogenum QEN-24S. Marine Drugs, 2010. 9(1): p. 59-70.

5. Heo, Y. M., Kim, K., Ryu, S. M., Kwon, S. L., Park, M. Y., Kang, J. E., Hong, J. H., Lim, Y. W., Kim, C., Kim, B. S., Lee, D., and Kim, J. J., Diversity and ecology of marine algicolous Arthrinium species as a source of bioactive natural products. Marine Drugs, 2018. 16(12): p. 508.