香蕉屬於熱帶水果為全世界四大重要經濟作物之一，果皮占整根新鮮香蕉總重的 35-40%。香蕉皮含有豐富營養成分，相當適合作為益生菌生長的基質。本研究利用 MRS 培養基 (Man, Rogosa and Sharpe broth) /2.5% 乾重香蕉副產品，添加不同比例之酵母萃取物 (yeast extract) 和糖蜜 (molasses) 為發酵基質，觀察不同轉速下Lactobacillus plantarum (ATCC 8014) 產生胞外多醣 (exopolysaccharides, EPS)等特性。經實驗結果得知，胞外多醣的產量在第 48 小時最高，以酵母萃取物 4%、糖蜜 4%、轉速 100 rpm (Y4M4S100) 的組別含量最高。由 FTIR 測定能發現組別間的官能基相似，均屬酸性多醣。胞外多醣具有抗氧化 (ABTS+、DPPH與還原力)、吸附膽固醇、抑制酪胺酸酶之能力。透過體外細胞實驗得知，胞外多醣無細胞毒性且能增加小鼠脾臟細胞活性，促進免疫反應。藉由測定IFN-γ的含量發現，胞外多醣能減少經由脂多醣 (lipopolysaccharides, LPS) 引起的發炎反應，具調節免疫之潛力。最後使用反應曲面法 (response surface methodology, RSM) 分兩部分探討，以胞外多醣含量、吸附膽固醇、抑制酪胺酸酶和抗氧化能力之體外生理活性等指標進行最佳化，發現 Y4M1S61.576 為最適條件；第二部分為免疫調節之效果 (MTT、IFN-γ)，以 Y1.291M4S61.158 作為最佳條件。透過發酵可衍生生物活性物質並進階有效使用副產品，達到永續發展與循環經濟之目標。

Banana is the top four traded agricultural crop worldwide. However, the banana peel waste is account for 35-40% of the total weight. Although the peel is considered as by-products, it still has high nutrients, providing it an opportunity to be substrates for probitic fermentation. This study aimed at conducting Lactobacillus plantarum (ATCC 8014) fermentation with Man, Rogosa and Sharpe broth (MRS) plus 2.5% banana by-products, different orbital shaking rate, concentration of yeast extracts and molasses. The results showed that the highest exopolysaccharides (EPS) production in the fermentation with 4% of yeast extract, 4% molasses in 100 rpm (Y4M4S100) for 48 hours. The FTIR spectrum revealed that the functional groups among different groups corresponded to the acid polysaccharides. The EPS in this study were known for the several functions as antioxidant capacity (ABTS+, DPPH, reduction), cholesterol absorption in vitro, and tyrosinase inhibition in vitro. The EPS did not trigger cytotoxic effect and increase the mice spleen cell activity to promote immune response. The EPS inhibited the LPS/ConA-induced inflammatory cytokine (IFN-γ) and exhibited the immune modulatory potential. Optimization fermentation parameters were divided by two different orientations.The first orientation contained in vitro bioactivities of EPS content, cholesterol adsorption, tyrosinase inhibition and antioxidant capacity, and it showed that Y4M1S61.576 was the optimal condition. In immune modulation (MTT and IFN-γ), the optimal condition was Y1.315M4S60.922. This study derived high value bioactive compounds from banana by-products and achieved the goal of circular economy.

目錄
壹、前言 1
貳、研究動機 2
參、文獻回顧 3
一、副產品 3
1. 全球動態 3
2. 循環經濟 3
二、香蕉 3
1. 簡介 3
1.1 臺灣香蕉之產量與出口 4
1.2 全球香蕉產量 4
1.3 香蕉產生之副產品 4
2. 香蕉皮之價值 4
2.1 香蕉皮之營養成分 4
2.2 香蕉皮的應用發展 5
三、乳酸菌 5
1. 簡介 5
1.1 益生菌 5
1.2 乳酸菌發酵代謝途徑 6
1.3 乳酸菌發酵之機能特性 6
1.4 乳酸菌於發酵製品之應用 6
1.5 植物乳桿菌介紹 6
2. 胞外多醣 7
2.1 影響胞外多醣生成之因素 7
2.2 胞外多醣之生理功效 8
2.3 胞外多醣在食品上之運用 8
四、反應曲面法 8
肆、研究架構 9
伍、實驗設計 11
陸、材料與方法 12
一、實驗材料 12
1. 香蕉副產品 12
2. 實驗菌株 12
3. 實驗細胞 12
4. 微生物培養基 12
5. 藥品來源 12
二、儀器設備 15
三、實驗方法 16
1. 香蕉副產品之一般成分分析 16
1.1 水分含量 16
1.2 灰分含量 16
1.3 粗蛋白含量 16
1.4 粗脂肪含量 16
1.5 粗纖維含量 17
1.6 碳水化合物 17
2. 菌株之保存與活化 17
2.1 菌株之保存 17
2.2 菌株之活化 17
3. 生長曲線 17
4. 香蕉副產品發酵培養基 18
5. 乳酸菌發酵條件 18
5.1 乳酸菌含量測定 18
5.2 pH 值測定 18
5.3 可滴定酸度 18
5.4 總糖濃度測定 18
5.5 還原糖濃度測定 19
6. 胞外多醣測定 19
6.1 胞外多醣萃取 19
6.2 胞外多醣含量分析 19
7. 抗氧化能力 19
7.1 ABTS+ 自由基清除能力 19
7.2 DPPH 自由基清除能力 20
7.3 還原力測定 20
8. 抑制酪胺酸酶活性 20
9. 體外膽固醇吸附 20
10. 免疫反應實驗 21
10.1 脾臟細胞培養 21
10.2 細胞活性試驗 (MTT) 21
10.3 細胞激素 21
11. 傅立葉轉換紅外光譜 (FTIR) 21
12. 統計分析 22
柒、結果與討論 23
ㄧ、香蕉副產品之一般成分分析 23
二、添加不同比例香蕉副產品發酵之總糖、還原糖、pH、可滴定酸和乳酸菌數變化 23
1. 總糖、還原糖 23
2. pH、可滴定酸 24
3. 乳酸菌數 24
4. 胞外多醣含量 24
三、不同濃度糖蜜、酵母萃取物和轉速對於植物乳桿菌產胞外多醣的含量影響 25
1. 總糖、還原糖 25
2. pH與可滴定酸 25
3. 乳酸菌數 25
4. 胞外多醣含量變化 26
5. FTIR 圖譜 26
四、不同濃度糖蜜、氮源和轉速對於植物乳桿菌產胞外多醣之生理活性探討 27
1. 抗氧化能力 27
2. 抑制酪胺酸酶的能力 28
3. 體外吸附膽固醇能力 28
4. 胞外多醣對小鼠脾臟細胞活性之影響 28
5. 胞外多醣對脾臟細胞 IFN-γ和 IL-4 分泌之影響 29
五、以反應曲面法探討最適發酵條件 30
捌、結論 31
玖、貢獻與限制 32
拾、參考文獻 33
圖表 42
附錄 80

 

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