舞菇Grifola frondosa菌絲體液態發酵多醣體及其免疫活性之探討

Production and immunomodulating activities of extracellular polysaccharides from submerged cultivation of Grifola frondosa

阮胡尼(Huu Nghi Nguyen)

指導教授 ： 郭嘉信

國立屏東科技大學/農學院/熱帶農業暨國際合作系所/碩士(2010年)

https://doi.org/10.6346/NPUST.2010.00188

摘要

舞菇 Grifola frondosa 為可食用性的擔子類 Basidiomycete 真菌，其在分類學上為Aphyllopherales目、Polyporaceaehas科。舞菇的子實體屬於菇菌類，中文別名為“灰樹花”日本人稱為“Maitake”。舞菇可促進健康對身體有益因此也被當作一種藥用菇菌類。有許多文獻證實由菇菌類中所萃取出來的多醣體具有抗癌及刺激免疫力提升等活性。本實驗利用三角瓶搖瓶培養舞菇菌絲體，並探討其搖瓶轉速、接種量及最初pH值對菌絲體型態的影響及在培養期間所產生的菌絲體量與胞外多醣體量。而且，我們利用此三種因子發展出最適化的數學模式來探討菌絲體量及胞外多醣體的產生。另外，本實驗也探討其胞外多醣體特性及其免疫活性。
實驗結果顯示培養期間搖瓶轉速、接種量及最初pH值顯著的影響其菌絲體型態進而影響其菌絲體量與胞外多醣體產生。低搖瓶轉速(100 rpm)產生小且髮狀的菌絲體型態但其菌絲體及胞外多醣體產量高(分別為5.30 mg/ml及2.14 mg/ml) ，而高搖瓶轉速(150 rpm)產生較大菌絲體團但較低的菌絲體含量及胞外多醣體。另一方面，較低的接種量(2%)產生較小的菌絲體團且較少的菌絲體量，但產生較多的胞外多醣體(2.46 mg/ml)，而4%的接種量產生較大的菌絲體團且較多的菌絲體量但較少量的胞外多醣體(1.68 mg/ml)。最初培養基pH值在5.5及6.0時皆可促
進其菌絲體量與胞外多醣體的產生。菌絲體產量最適的條件為搖瓶轉速105 rpm、接種量3.1%及最初pH值5.5，在此條件下，其最高的菌絲體含量可達5.28mg/ml。而關於胞外多醣體產生的最適化條件分別為搖瓶轉速111 rpm、接種量3.3%及最初pH5.6，在此條件下，其最高的菌絲體含量可達2.35mg/ml。HPLC-RI試驗結果顯示舞菇菌絲體胞外多醣體有兩種其分子量分別為21.16及1.69 kDa。其胞外多醣體顯示可使其PBMC細胞增殖提升其免疫活性，在波長450 nm時，舞菇胞外多醣體組的吸光值為1.12 高於控制組及LPS組的吸光值0.74及0.79。

關鍵字

舞菇 ； 菌絲體量 ； 胞外多醣體 ； 免疫活性 ； 最適化

並列摘要

The edible mushroom Grifola frondosa is a Basidiomycete fungus belonging to the order Aphyllopherales, and family Polyporaceaehas. Polysaccharides extracted from the mushroom have been reported to have antitumor and immunomodulating activities. Other medical uses of the mushroom include antioxidant, antidiabetic, and cardiovascular activities. In the present research, G. frondosa was cultivated in shaking flasks and investigated the effects of agitation speeds, inoculum ratios, and initial pH on the pellet morphology, biomass, and extracellular polysaccharide production. Furthermore, mathematical models for optimization of the three factors for biomass and extracellular polysaccharide production were developed. In addition, immunomodulating activities of extracellular polysaccharides produced by submerged cultivation under optimized conditions was also evaluated.
The results showed that the culture parameters including agitation speed, inoculum ratio, and initial pH influenced pellet morphology,
biomass, and extracellular polysaccharide production. Low agitation speed at 100 rpm resulted in small and hairy pellets but greater amount of biomass and extracellular polysaccharides which were 5.30 mg/ml and 2.14 mg/ml, respectively, while higher agitation speed at 150 rpm led to larger pellet size but less biomass and extracellular polysaccharides which were 5.03 mg/ml and 1.62 mg/ml, respectively. On the other hand, less inoculum ratio 2% led to smaller pellet size and less amount of biomass 4.51 mg/ml while maximum extracellular polysaccharide production 2.46 mg/ml, whereas 4% of inoculum resulted in larger pellet size and greater amount of biomass production 5.21 mg/ml but less amount of extracellular polysaccharides 1.68 mg/ml. In addition, initial pH 5.5 and 6.0 were favorable for both biomass and extracellular polysaccharide production. The optimal conditions for biomass production were agitation speed at 105 rpm, inoculum ratio 3.1%, and initial pH 5.5. Under this condition, the predicted maximum biomass was 5.28 mg/ml. Regarding to optimization of extracellular polysaccharide production, the optimal conditions were agitation speed at 111 rpm, inoculum ratio 3.3%, and initial pH 5.6. Under this situation, the predicted maximum extracellular polysaccharides was 2.35 mg/ml. HPLC-RI results indicated that, two extracellular polysaccharides with molecular weight of 21.16 and 1.69 kDa were identified. The polysaccharides showed immunomodulating activity by increasing the proliferation of peripheral blood mononuclear cells. Optical density at 450nm of the treatment with polysaccharide from G. frondosa was 1.12 higher than that of control and LPS treatments which were 0.74 and 0.79, respectively. The findings may be useful in future for development of proper fermentation techniques for production of G. frondosa products.

並列關鍵字

Grifola frondosa ； biomass ； extracellular polysaccharides ； immunomodulating activities ； optimization

參考文獻

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