蜜環菌栽培體之成份分析與抗發炎活性研究

Studies on chemical constituents and anti-inflammatory activity of cultivated Armillariella mellea

張景雯(Ching-Wen Chang)

指導教授 ： 盧虎生

共同指導教授 ： 盧美光(Mei-Kuang Lu)

國立臺灣大學/生物資源暨農學院/農藝學研究所/碩士(2011年)

https://doi.org/10.6342/NTU.2011.01012

摘要

蜜環菌 (Armillariella mellea) 是中國傳統的中草藥，在過去的研究中被指出其內含多醣體成份具抗發炎能力。因此，得到兼具產量與品質的蜜環菌，便是本研究的目標。本研究發現在培養四十九天時，蜜環菌的乾重產量可以達到15.73 ± 0.16 g/l，同時多醣體和酒精萃取物的產量也可以分別達在到0.63 ± 0.04 g/l和5.51 ± 0.38 g/l。蜜環菌多醣體中以半乳糖和葡萄糖為主要成份，含量分別為1071.30 ± 10.89 和525.45 ± 4.14 mmol/g polysaccharide。蜜環菌酒精萃取物中含有成份ADP、cytidine和adenosine，在每毫克酒精萃取物中含量分別為0.48、0.34和0.74 mg。在抗發炎能力測定中，將蜜環菌萃取出來的多醣體、含硫多醣體和酒精萃取物分別處理在小鼠巨噬細胞RAW264.7及人類臍靜脈細胞EAhy926上，並分別以脂多醣體 (lipoplysaccharide) 和腫瘤壞死因子 (tumor necrosis factor-a) 刺激細胞使其發炎後，偵測tumor necrosis factor-a (TNF-a)、interleukin-6 (IL-6) 和monocyte chemotactic protein-1 (MCP-1) 的含量。結果發現：在RAW264.7中，酒精萃取物的濃度和TNF-a及IL-6的產量有劑量正相關。此外，培養四十九天的蜜環菌多醣體500 mg/ml顯著抑制TNF-a 及IL-6分泌的能力達31.17%和62.74%。含硫多醣體對TNF-a及IL-6產量的抑制達100%及56.82%，其效果優於不含硫的多醣體。EAhy中MCP-1的含量可被培養四十九天蜜環菌的多醣體、酒精萃取物和含硫多醣體抑制到52.17％、37.49％和34.53％。以上結果皆證實蜜環菌萃取物有抗發炎的效果。綜上所述，在本研究的培養條件下，蜜環菌乾重和多醣體的產量在培養四十九天後都能有效的被提升，而半乳糖和葡萄糖則是蜜環菌多醣體中含量最多的單糖成份。除此之外，無論是蜜環菌的多醣體、含硫多醣體或酒精萃取物，都能有效抑制TNF-a、IL-6 和MCP-1等和發炎相關的細胞激素。

關鍵字

蜜環菌 ； 發炎 ； 細胞激素

並列摘要

Armillariella mellea was reported to exhibit anti-inflammatory activity, especially that of the polysaccharide (PS) . To mass produce high quality of A. mellea was the aim of the study. The maximum production of 15.73 ± 0.16 g/l was obtained at 49 days, with the yield of PS and ethanolic extract of 0.63 ± 0.04 g/l and 5.51 ± 0.38 g/l, respectively. Galactose and glucose was the major sugar of PS with value of 1071.30 ± 10.89 and 525.45 ± 4.14 mmol/g PS at 49-day-culture, respectively. The components of ethanolic extract included ADP, cytidine and adenosine with the value of 0.48, 0.34 and 0.74 mg/mg ethanolic extract, respectively. Effects of extracted PS, sulfated polysaccharide (SPS) and ethanolic extract from cultured A. mellea on lipopolysaccharide (LPS) and TNF-a induced inflammation-related cytokines in RAW264.7 macrophages and EAhy926 were evaluated. Secretions of tumor necrosis factor-a (TNF-a) and interleukin-6 (IL-6) were detected in RAW264.7. The results showed that inhibition of TNF-a and IL-6 production were in a dose-dependent manner with ethanolic extract of A. mellea. PS from 49-day-cultured mycelia at 500 mg/ml showed significant inhibition of TNF-a and IL-6 in the value of 31.17% and 62.74%, respectively. Furthermore, SPS exhibited more effective on inhibition of TNF-a and IL-6 secretion as compared that with non-SPS in the value of 100% and 56.82% suppression, respectively. On the other hand, the secretion of cytokine monocyte chemotactic protein-1 (MCP-1) was significantly reduced in 52.17%, 37.49% and 34.53% for 49-day-culture PS, ethanolic extract and sulfated-PS (SPS), respectively. All of the above results suggested that A. mellea exhibited anti-inflammatory activity. In conclusion, in our cultured condition, A. mellea's dry weight and PS production could be enhanced at 49-cultured days. Glactose and glucose was the major component of A. mellea PS. PS, ethanolic extract and SPS of A. mellea significantly inhibited the release of the inflammatory related cytokines of TNF-a, IL-6 and MCP-1.

並列關鍵字

Armillariella mellea ； anti-inflammatory ； TNF-a ； IL-6 ； MCP-1 ； IkB-a

參考文獻

Abbas, A. K., & Lichtmanl, A. H. (2004). Cellular and molecular immunology, 5th ed. Elsevier Pte. Ltd, Singapore. pp.247, 252-253, 262, 285-286, 490-491.

Alberto, A., Rosanna, C., & Gianluca, N. (1986). Structures of melleolides B-D, three antibacterial sesquiterpenoids from Armillaria mellea. Phytochemistry, 25, 471-474.

Anderson, J. B., Korhonen, K., & Ullrich, R. C. (1979). Biological species of Armillaria mellea in North America. Mycologia, 71, 402-414.

Balkwill, F. R., & Burke, F. (1989). The cytokine network. Immunology Today, 10, 299-304.

Baumgartner, K., Bhat, R., & Fujiyoshi, P. (2010). A rapid infection assay for Armillaria and real-time PCR quantitation of the fungal biomass in planta. Fungal Biology, 114, 107-119.