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研究生:林姍樺
研究生(外文):Shan-hua Lin
論文名稱:以固態栽培豬肚菇及特有化合物之產製
論文名稱(外文):The solid-state cultivation of Clitocybe maxima and the production of special compounds
指導教授:王進琦
指導教授(外文):Jinn-Chyi Wang
學位類別:碩士
校院名稱:大仁科技大學
系所名稱:食品科技研究所
學門:農業科學學門
學類:食品科學類
論文種類:學術論文
論文出版年:2012
畢業學年度:100
語文別:中文
論文頁數:128
中文關鍵詞:γ-胺基丁酸麥角固醇海藻糖反應曲面法豬肚菇
外文關鍵詞:Response Surface MethodologyClitocybe maxima
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中文摘要
豬肚菇又名酒杯菇,是一種質脆味佳及具有一些生理效用之新興的食藥用菇類,因此頗有開發及推廣之前景。
目前食藥用菇類多使用木屑基質進行人工栽培,須耗用大量木屑,且栽培後之廢棄基質對林業及環境均會造成某種程度之負擔及傷害。本研究在探討豬肚菇以液態培養時菌絲產量及使用穀類基質進行固態栽培時子實體產量及生物效率 (biological efficiency) 之適化條件;另外,針對幾種具生理功能之特用成分,包括海藻糖 (trehalose)、麥角固醇 (ergosterol) 及γ-胺基丁酸 (γ-aminobutyric acid, GABA) 分析其於培養期間之產製情形;並且亦探討利用穀物基質產製γ-胺基丁酸之適化基質組成。
研究結果發現,液態培養菌絲產量最佳之適化條件為 C/N 60、溫度 23 ℃ 及 pH 6.5,而在生長曲線測定時,培養至第 20 天菌絲體可達最佳產量 5.35 mg (乾重)。使用穀類基質進行固態栽培時,子實體產量、子實體長出數量及子實體高度最佳之適化基質組成為糙米 30 g、2% peptone 及 1% yeast extract,栽培期間子實體中海藻糖含量在第 30 天可達最高量之 310.90 mg/g,而麥角固醇之含量亦在第 30 天達最高量 (4.34 mg/g),其生物效率為 59.57%,子實體產量為 17.90 g 。至於 GABA 產量最高之基質為含 5% Glucose 及0.5% Glutamate 者,其在培養至第 27 天時,GABA 有最高之產率 (14.2 mg/g),因此利用穀物基質栽培豬肚菇除可收穫子實體外,亦可利用栽培過程生產具生理效用之化合物,並且收成後之基質亦具食用價值,俾使菇類栽培能達到之完全利用之效益。
The Clitocybe maxima is also named as “wine glasses mushroom”. It’s a crisp and well-tasted new edible and medicinal mushroom, and which have a few physiological effects. They shall be explored and developed for their economical values. Much sawdust could be used for the cultivation of mushrooms artificially at present, and the spent substrate should make overload and harm to the wood industry and environment in some levels. This study explored the optimization for the production of biomass in the liquid culture and the yield of fruiting bodies and biological efficiency in the solid-state culture. Some compounds of physiological activities were determined, including trehalose, ergosterol, and γ-aminobutyric acid (GABA) during cultivation. The optimal cereal-based substrates for the production of GABA in the solid-state culture were also studied by using Response Surface Methodology.
The results showed that the optimal coditions for the biomass production in the liquid culture were C/N 60, 23 ℃, and pH 6.5. The biomass reached to the highest production (5.35 mg, dry weight) for culturing 20 days in the growth curve. The highest yield should be obtained in the substrate containing brown rice 30g, 1% peptone, and 1% yeast extract. The trehalose of fruiting body reached to the highest amount of 310.90 mg/g for culturing 30 days, while the ergosterol was 4.34 mg/g. The yield and biological efficiency of fruiting body was 17.9 g and 59.57%, respectively. The production of GABA was 14.2 mg/g in the optimal cereal-based substrate, containing 5% glucose and 0.5% glutamate for culturing 27 days. In addition to the harvest of edible fruiting bodies, some compounds of physiological activities from the spent substrate and fruiting bodies of Clitocybe maxima might be also obtained. Therefore, the solid-state culture of Clitocybe maxima with the cereal-based substrate might be able to utilize totally.
中文摘要……………………………………………………………….. I
英文摘要………………………………………………………………..II
誌謝……………………………………..................................................III
目錄………………………………………………………………… …IV
圖表目錄……………………………………………………………… IX
附錄……………………………………………………………………XII
壹、前言…………………………………………………………………1
貳、文獻回顧……………………………………………………………4
一、真菌的介紹……………………………………………………4
二、食藥用菇類的介紹……………………………………………4
(一) 菇類之一般特性…………………………………………4
(二) 台灣重要菇類之產值、出口量及生產量……………….5
(三) 菇類之利用………………………………………………6
(四) 菇類應用於健康食品……………………………………8
(五) 菇類之特殊成分及生理效用……………………………9
(六) 菇類的栽培……………………………………………..11
1. 菇類栽培之內在影響因子………………………….11
(1) 碳源……………………………………………11
(2) 氮源……………………………………………11
(3) 無機鹽…………………………………………12
(4) 生長因子………………………………………13
2. 菇類栽培之外在影響因子………………………….13
(1) 溫度……………………………………………13
(2) 水分……………………………………………14
(3) 濕度……………………………………………14
(4) 氧與二氧化碳…………………………………14
(5) 酸鹼度…………………………………………15
(6) 光照……………………………………………15
(七) 菇類研究及開發之展望………………………………..16
三、菇類之特有化合物…………………………………………..16
(一) 抗生素 (Antibiotics)……………………………….…..16
(二) 凝集素 (Agglutinin)……………………………………16
(三) 萜類化合物 (Terpenoids)……………………………...17
(四) 多醣類 (Polysaccharides)……………………………...17
(五) 海藻糖 (Trehalose)…………………………………….17
(六) 麥角固醇 (Ergosterol)…………………………………19
(七) γ-胺基丁酸 (γ-Aminobutyric acid, GABA)……… ..….19
四、豬肚菇之介紹…………………………………………………20
(一) 豬肚菇之類緣關係……………………………………..20
(二) 豬肚菇之生長特性……………………………………..20
(三) 豬肚菇之利用價值……………………………………..20
(四) 豬肚菇之相關研究……………………………………..21
五、反應曲面法 ( Response Surface Methodology, RSM)……….21
(一) 反應曲面法之介紹……………………………………..21
(二) 反應曲面法之優點……………………………………..21
(三) 反應曲面法試驗設計步驟……………………………..22
(四) 反應曲面法探討最適化步驟…………………………..23
(五) 二水準因子設計………………………………………..24
(六) 陡升路徑法 (Method of steepest ascent)……………...24
(七) 中央合成設計 (Central composite design)……………25
叁、材料與方法………………………………………………………..27
一、實驗架構……………………………………………………..27
二、材料……………………………………………………………27
(一) 菌種來源及保存………………………………………..27
(二) 種菌之製備……………………………………………..27
(三) 試藥……………………………………………………..27
(四) 儀器……………………………………………………..28
三、方法……………………………………………………………28
(一) 液態培養豬肚菇之適化條件…………………………..28
(二) 液態培養豬肚菇之生長曲線…………………………..28
(三) 液態培養豬肚菇產製特有化合物 ………………..…..29
(四) 固態栽培豬肚菇穀物基質之培養基組成 ………..…..29
(五) 固態栽培豬肚菇產製特有化合物……………………..29
(六) 固態栽培豬肚菇產製γ-胺基丁酸之適化基質組成…..29
(七) 分析項目………………………………………………..30
1. 還原糖……………………………………………….30
2. 氮含量……………………………………………….30
3. 水分含量…………………………………………….31
4. 水活性……………………………………………….31
5. pH ………………………………………………….32
6. 二氧化碳濃度……………………………………….32
7. 生物效率………………………………..…………...32
8. 海藻糖……………………………………………….32
9. 麥角固醇…………………………………………….33
10. γ-胺基丁酸………………………………………….33
(八) 統計分析………………………………………………..34
肆、結果………………………………………………………………..35
伍、討論………………………………………………………………..40
陸、結論…………………………………………………………………45
柒、參考文獻…………………………………………………………..46

圖表目錄
圖1 實驗架構圖………………………………………………………57
圖2 還原糖之標準曲線………………………………………………58
圖3 氮含量之標準曲線………………………………………………59
圖4 二氧化碳濃度檢測之裝置………………………………………60
圖5 海藻糖之標準曲線………………………………………………61
圖6 麥角固醇之標準曲線……………………………………………62
圖7 γ-胺基丁酸之標準曲線………………………………………….63
圖8 以不同溫度及pH 對豬肚菇菌絲生長之等高線圖…................64
圖9 液態培養豬肚菇在不同時間,菌絲體之變化…………………65
圖10 液態培養豬肚菇在不同時間,pH之變化…………………….66
圖11 液態培養豬肚菇在不同時間,發酵液中還原糖之變化……..67
圖12 液態培養豬肚菇在不同時間,發酵液中氮含量之變化…..…..68
圖13 液態培養豬肚菇在不同時間,菌絲體中海藻糖之變化………69
圖14 液態培養豬肚菇在不同時間,菌絲體中麥角固醇之變化……70
圖15 液態培養豬肚菇在不同時間,菌絲體中γ-胺基丁酸的變
化………………………………………………………………..71
圖16 固態栽培豬肚菇在不同穀類基質,培養20天之產率………...72
圖17 固態栽培豬肚菇在不同穀類基質,培養20天之生物效率…...73
圖18 固態栽培豬肚菇在不同時間,基質水含量之變化……………74
圖19 固態栽培豬肚菇第15天時,菌絲糾結及原基體形成……….75
圖20 固態栽培豬肚菇第20天時,菌絲糾結及原基體形成……….76
圖21 固態栽培豬肚菇第25天時子實體形成 (A &;B)……………...77
圖22 固態栽培豬肚菇第30天之成熟子實體……………………….78
圖23 固態栽培豬肚菇第35天之成熟子實體……………………….79
圖24 固態栽培豬肚菇在不同時間,基質水活性之變化…………..80
圖25 固態栽培豬肚菇在不同時間,基質pH之變化……………….81
圖26 固態栽培豬肚菇在不同時間,基質上部空間二氧化碳濃度之
變化……………………………………………………………..82
圖27 固態栽培豬肚菇在不同時間,基質及子實體中海藻糖之變化………………………………………………………………..83
圖28 固態栽培豬肚菇在不同時間,基質及子實體中麥角固醇之變
化………………………………………………………………..84
圖29 固態栽培豬肚菇在不同時間,基質及子實體中γ-胺基丁酸之
變化……………………………………………………………..85
圖30 不同濃度之Glucose及Glutamate 對豬肚菇產製γ-胺基丁酸之
等高線圖………………………………………………………..86
圖31 固態栽培豬肚菇在不同時間,基質、菌絲體及子實體中γ-胺
基丁酸之變化…………………………………………..………87

表1 液態培養豬肚菇之培養基組成…………………………………88
表2 豬肚菇菌絲生長獨立變因所代表之實際值……………………89
表3 固態栽培豬肚菇之培養基組成…………………………………90
表4 豬肚菇產製γ-胺基丁酸獨立變因所代表之實際值……………91
表5 以中央合成設計反應曲面法之組合條件及豬肚菇菌絲之產量……………………………………………………………..…..92
表6 C/N、溫度及pH 對豬肚菇生長菌絲體之影響……………….93
表7 在不同穀物基質,豬肚菇子實體高度、數量及重量………….94
表8 以中央合成設計反應曲面法之組合條件及豬肚菇產製γ-胺基丁
酸…………………………………………………………………95
表9 葡萄糖及麩胺酸對豬肚菇生長過程產製γ-胺基丁酸之影響………………………………………………………………....96
附錄
附圖1 真菌之分類……………………………………………………97
附圖2 海藻糖之結構…………………………………………………98
附圖3 海藻糖在酵母菌中之生合成途徑……………………………99
附圖4 海藻糖在酵母菌中之三種主要生合成途徑……………..…100
附圖5 海藻糖在植物中之代謝途徑………………………………..101
附圖6 麥角固醇之生合成途徑……………………………………..102
附圖7 γ-胺基丁酸之結構…………………………………………..103
附圖8 γ-胺基丁酸之代謝途徑……………………………………...104
附圖9 豬肚菇之類緣關係…………………………………………..105
附圖10 二變數因子系統之反應曲面圖……………………………106
附圖11 中央合成設計法之星點與中心點實驗圖…………………107
附表1 台灣在1991至2010年菇類之產值…………………………108
附表2 台灣在2001至2010年菇類之出口量……………………..109
附表3 台灣在2001至2010年菇類之產量………………………..110
附表4 衛生署驗證通過的健康食品之菇類及其功用……………..111
附表5 豬肚菇及其他同屬不同菇種菌之相關研究………………..112
附表6 23因子設計…………………………………..………………114
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