探討稻殼酸水解液中有害物質的移除對Pichia stipitis BCRC 21775生產乙醇之影響

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研究生：	劉家宏 Chia-Hung Liou
論文名稱：	探討稻殼酸水解液中有害物質的移除對Pichia stipitis BCRC 21775生產乙醇之影響 Effect of detoxification methods of dilute-acid hydrolyzates for fermentation by Pichia stipitis BCRC 21775 on ethanol production
指導教授：	徐敬衡 Chin-Hang Shu
口試委員:
學位類別：	碩士 Master
系所名稱：	工學院 - 化學工程與材料工程學系 Department of Chemical & Materials Engineering
畢業學年度：	96
語文別：	中文
論文頁數：	107
中文關鍵詞：	酵素、生質酒精、稀酸水解
外文關鍵詞：	laccase, Agaricus blazei Murrill, Pichia stipitis
相關次數：	點閱：14 下載：0
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最近幾年，由於現代工業的發展，使得化石燃料快速的消耗，造成能源價格節節高漲。因此，目前全世界各國都在積極進行新能源之研究開發，其中，生質能( biomass energy )因其原料取自大自然，且能生生不息地被循環使用，故成為最受矚目的替代能源之一。
纖維素是一種生質能的原料，將纖維素水解後，生成容易發酵的單醣，再經微生物發酵後，所產生的酒精即為生質酒精。一般纖維素水解的化學方法包含了稀酸水解和濃酸水解。本實驗中採用稀酸水解法，稀酸水解雖然反應速度快，但因為不具選擇性，會有大量副產物的生成，如:乙酸(Acetic acid)、Furfural、HMF、LDP…等等，使得酒精產率過低而不具經濟價值。在本研究中，選擇酸水解條件為1% H2SO4 、酸水解時間為60分鐘、固液比為10%、121∘C、1.2atm，所產生的酸水解液，則利用Agaricus blazei Murrill 所產的laccase酵素及百貴LC-A1活性碳移除其中的有害物質。利用添加laccase酵素可移除酸水解液中53%的LDP，而添加百貴LC-A1活性碳則可100%移除酸水解液中的乙酸。
本研究中，以攪拌式反應器來進行液態發酵培養酵母菌來產生酒精，所使用的菌株為 Pichia stipitis BCRC 21775。在實驗中，我們探討添加不同活性單位的laccase 酵素及活性碳，對Pichia stipitis BCRC 21775產生酒精的影響。
實驗結果發現如下:
(1) 在本研究稀酸水解條件下，LDP ( Lignin degradation products ) 為抑制酒精產率的主要物質。
(2) 在以不同來源的laccase降解LDP實驗中，可以發現由A. blazei 所
產的laccase其降解LDP能力高於傳統T. versicolor所產laccase，
且經A. blazei laccase處理的酸水解液的酒精產量也優於傳統
T. versicolor laccase 處理的酸水解液。
(3) 經處理的酸水解液其酒精產量、菌體生長都比未處理的高，且發酵時間也比未處理的短，其中以添加10000U/L laccase酵素其酒精產量(5.62g/L→10.71g/L)、比生長速率μ (0.006→0.013)為未處理的兩倍，菌體生長為未處理的1.85倍。

During hydrolysis of lignocellulosic materials a wide range of compounds which are inhibitory to microorganisms are formed or released. Based on their origin the inhibitors are usually divided in three major groups: weak acids, furan derivatives, and phenolic compounds. These compounds limit effcient utilisation of the hydrolysates for ethanol production by fermentation.
This research discusses the generation of detoxification methods , and the effect of these on fermentation yield and productivity. Choosing acid-hydrolysates conditions , reaction time was sixty - minuite and solid /liquid ratio was ten percent , and the reducing sugar conversion was 83% . Then use enzyme laccase and activated charcoal removal of toxic compounds in the acid-hydrolysates , the ethanol yield and specific growth rate ehanced two times , the biomass yield ehanced 1.85 times .

摘要	I
目錄	III
圖索引	VII
表索引	X
第一章	緒論..............................................1
1-1	研究動機..........................................1
1-2	研究目的..........................................2
第二章 文獻回顧..........................................3
2-1  生質能源的定義及分類..............................3
2-1-1  生質能源的定義................................3
2-1-2  生質能源的分類................................5
2-2  木質纖維素........................................6
2-3  前處理............................................8
2-3-1  機械破碎法...................................10
2-3-2  氨纖維爆破法.................................10
2-3-3  蒸氣爆破法...................................11
2-3-4  二氧化碳爆破法...............................12
2-3-5  超音波微波處理法.............................12
2-4  生質原料轉換成生質酒精程序.......................13
2-4-1  稀酸水解.....................................13
2-4-2  濃酸水解.....................................17
2-4-3  酵素水解.....................................20
2-5  酸水解液中副產物去除方法.........................22
2-5-1  汽提法( Air Stripping ) .........................22
2-5-2  閃蒸法( Flash evaporation ) .....................23
2-5-3  過量鹼處理法( over-liming ) ....................23
2-5-4  活性碳吸附法.................................24
2-5-5  酵素處理法...................................25
2-6  漆氧化酶.........................................27
2-6-1  漆氧化酶之來源...............................29
2-6-2  漆氧化酶之結構...............................30
2-6-3  漆氧化酶之反應機制...........................32
2-6-4  漆氧化酶之應用...............................35
2-7 稻殼的應用........................................38
第三章、實驗材料與方法..................................40
3-1 實驗材料..........................................40
3-1-1 實驗菌株......................................40
3-1-2 實驗藥品......................................40
3-1-3 實驗儀器及其他設備............................41
3-1-4 實驗裝置......................................43
3-2 實驗方法..........................................44
3-2-1 Pichia stipitis 菌種保存及培養方法.................44
3-2-2 Agaricus blazei Murrill 菌種保存及培養方法.........46
3-2-3 操作條件......................................48
3-2-4 前處理........................................49
3-2-5 分析方法......................................51
3-2-5-1 水解液處理流程............................51
3-2-5-2 菌重分析..................................53
3-2-5-3 葡萄糖殘量分析............................54
3-2-5-4 Laccase 酵素活性分析.......................55
3-2-5-5 有害物質濃度分析..........................58
3-2-5-6 酒精濃度分析..............................63
3-2-5-7 蛋白質含量測定............................64
第四章、結果與討論......................................65
4-1 酵母菌平面培養情形................................65
4-2 稀酸水解條件分析及抑制作用之影響..................66
4-2-1 稀酸處理時間之影響............................66
4-2-2 稀酸處理固液比之影響..........................69
4-2-3 酸水解液有害物質分析..........................72
4-2-4 酸水解液分析..................................74
4-3 巴西洋菇平面培養情形..............................75
4-4 巴西洋菇產Laccase酵素活性分析.....................76
4-5 Laccase酵素降解LDP之影響.........................77
4-6 活性碳吸附酸之影響.................................80
4-7 搖瓶實驗...........................................81
4-8不同處理方式攪拌式發酵槽培養實驗....................86
4-8-1發酵動力曲線比較................................86
4-8-2 不同處理方式對菌體生長及酒精產量之影響.........88
第五章、結論與建議.......................................92
5-1 結論..............................................92
5-2 建議..............................................93
參考文獻...............................................94

                                

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