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研究生:田育民
研究生(外文):Yu-MinTien
論文名稱:厭氧流體化床優植菌種代謝廚餘以促進纖維水解及產氫功能
論文名稱(外文):Bioaugmented Anaerobes Digest Kitchen Waste to Promote Cellulose Hydrolysis and Hydrogen Generation with Anaerobic Fluidized Bed Process
指導教授:鄭幸雄鄭幸雄引用關係
指導教授(外文):Sheng-Shung Cheng
學位類別:碩士
校院名稱:國立成功大學
系所名稱:環境工程學系碩博士班
學門:工程學門
學類:環境工程學類
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:172
中文關鍵詞:纖維素Clostridium sp. TCW1厭氧流體化床氫氣CMCase
外文關鍵詞:CelluloseClostridium sp. TCW1AnFBhydrogenCMCase
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本研究先以都市污水廠的厭氧消化污泥作為多樣性菌種來源,以批次方式測試消化污泥對純纖維素基質的水解,酸化及產氣的潛能。批次操作的前33~40小時內有明顯的產氫表現,氫氣可占所產氣體的40%以上;而後甲烷的產氣表現迅速提升,但產氣組成中的甲烷比例普遍不到40%,剩下的氣體組成皆為二氧化碳。實驗進行了99小時,甲烷產氣速率趨緩,此時批次瓶內的pH降到5.4,推測是因低pH影響甲烷菌的活性。99小時後,纖維素約去除34%,CMCase活性可累積到0.08 U mL-1。
而後以纖維素水解菌Clostridium sp. TCW1作為菌源,測試其對濾紙,蔗渣,狼尾草以及稻桿等纖維性基質的產氫潛能。其中以狼尾草試驗組有最高的產氫速率15 mL-H2 L-1 hr-1,產氫yield約10.4 mmole-H2 g-NG-1。以濾紙作為基質實驗組中,可觀測到TCW1分解纖維素的液相代謝產物主要有乳酸,乙酸,丁酸以及乙醇;另有少量甲酸及丁醇的生成。其中乳酸及乙酸在批次進行第218小時以後有明顯消耗情形,同時丁酸大量生成,因此推測TCW1可能也有同時代謝乳酸及乙酸生成丁酸的能力。
接著運轉100 L厭氧流體化床(AnFB)反應槽,植種TCW1優勢纖維素降解菌,饋料以CMC來增殖優勢菌於反應槽,為第1試程。而後再饋料以木質纖維(tissue),並以低進流負荷VLR 0.94 g-COD L-1 d-1,15天的水力停留時間(HRT)啟動連續流試程,為第2試程。試程中,pH控制在6.2以上,氣相的氫氣比例最高可達30%,穩定時的氫氣比例為13%;產氫速率為15 mL-H2 L-1 d-1,產氫yield約0.64 mmole-H2 g-COD-1。整個第2試程的纖維素降解率可達98%,槽內的總非纖維素懸浮固體量可累積到549 g SS。
第3試程將VLR提升至1.13 g-COD L-1 d-1,HRT降至12.5天,pH維持在6.2左右。試程後期槽內發生甲烷化,甲烷比例累積到30%,氫氣比例降到3%以下,其餘皆為CO2,甲烷產氣速率為103 mL-CH4 L-1 d-1,產氫速率約3 mL-H2 L-1 d-1。推測因低體積負荷及較長的水力停留時間導致甲烷菌有機會在反應槽中生長出來。整個第3試程的固態碳水化合物(纖維素)降解率仍有99%,但槽內的非碳水化合物懸浮固體含量降至206 g SS。第4試程為了抑制槽內的甲烷化,以澱粉類廚餘(SKW)將體積負荷提升至約5 g-COD L-1 d-1,HRT降至10天,但產氣還是以甲烷及二氧化碳為主;將SKW負荷提升至9 g-COD L-1 d-1以後甲烷的產氣表現才被抑制住,顯示甲烷菌較不適合在高負荷環境下生長。此時氫氣比例可提升至40%,產氫速率約835 mL-H2 L-1 d-1。而後隨即將澱粉類廚餘換成蔬菜類廚餘(VKW),產氫速率馬上下降,約20 mL-H2 L-1 d-1,氫氣比例亦降至12%左右,槽內亦出現肉眼可辨識的纖維性固體物累積。木質纖維及VKW的饋料8天後換回1 g L-1 d-1木質纖維與9 g L-1 d-1 SKW的進料,產氫速率及氫氣比例逐漸上升,但槽內的纖維性固體物累積情況未消失。
量測AnFB操作期間各試程槽內的纖維素水解酵素活性來進一步了解纖維素水解情況。第3試程操作第232天槽內CMCase活性約0.15 U mL-1,接近本研究測到的最高值0.17 U mL-1;而第4試程進料澱粉類廚餘後(第356天)槽內的CMCase活性只有0.02 U mL-1,接近偵測極限。且由掃描式電子顯微鏡(SEM)的觀察,饋料廚餘後的菌相以懸浮在液相中的為主,少有附著在纖維性固體物上的菌體。推測進料澱粉廚餘後帶進利用澱粉的微生物,與槽內的纖維素水解菌產生生長空間的競爭,使得纖維素水解菌及酵素逐漸流失,而造成纖維素降解率下降。此假設仍有待驗證。
The objective of this study is to construct a system that hydrolyzes cellulose and produces hydrogen simultaneously using anaerobic fluidized bed (AnFB) reactor. A thermophilic cellulolytic microbe, Clostridium sp. TCW1 was chosen as an inoculation for AnFB. There were three Runs in AnFB process operation. 1st Run was to start up this system and enrich biomass of cellulose hydrolyzing microbes, 2nd Run was to increase cellulose volumetric loading rate (VLR) to 1.16 g-COD/L/d, and 3rd Run was adding starch kitchen waste (SKW) to increase hydrogen production. In 1st Run, the hydrogen production rate was 0.02±0.01 L-H2/L/d, hydrogen yield was 0.77±0.57 mmole-H2/g-COD, and the total solid amount at Day 111 was 2,046 g SS. In 2nd Run, the hydrogen production rate was 0.004 L-H2/L/d, hydrogen yield was 0.10 mmole-H2/g-COD, and the total solid amount at Day 232 and Day 248 were 538 g SS and 206 g SS respectively, there were biomass washed out and degraded during 2nd Run. In 3rd Run, the hydrogen production rate is up to 1.0 L-H2/L/d. The enzyme activity of Endo-β-1,4-glucanase (CMCase) of AnFB operation process at Day 232 in 2nd Run was 0.15 U/mL, it near the maximum activity 0.17 U/mL of batch culture though biomass content in AnFB was not enough (538 g); at Day 280 in 3rd Run, CMCase activity down to 0.02 U/mL when vegetable kitchen waste (VKW) was fed to AnFB, it seems VKW was not favored for cellulase production in this process operation.
摘要 I
Abstract III
致謝 IV
目錄 V
表目錄 VIII
圖目錄 X
第一章、 前言 1
第二章、 文獻回顧 4
2-1. 全球能源發展 4
2-2. 植物細胞及纖維素結構 8
2-2-1. 植物細胞分類 9
2-2-2. 植物細胞壁的構成 17
2-2-3. 纖維素結構 19
2-3. 厭氧微生物分解有機物之代謝機制 20
2-3-1. 厭氧有機物代謝概論 20
2-3-2. 纖維素水解機制 22
2-3-3. 厭氧碳水化合物代謝及有機酸醇生成機制 27
2-3-4. 厭氧生物產氫機制 30
2-3-5. 厭氧生物產甲烷機制 36
2-4. 厭氧生物反應槽批次及連續流之單元操作功能 41
2-5. 纖維水解以及厭氧產氫產甲烷醱酵之微生物菌群 44
2-5-1. 纖維素水解菌群 44
2-5-2. 厭氧產氫醱酵菌群 49
2-5-3. 厭氧甲烷醱酵菌群 53
第三章、 材料與方法 59
3-1. 基質成分特性分析(項目) 59
3-1-1. 總有機碳 60
3-1-2. 揮發酸分析IC 60
3-1-3. 氣體組成分析 - GC-TCD 61
3-2. 批次基質微生物分解實驗方法(BHP, BMP) 62
3-2-1. Owen營養鹽組成 62
3-2-2. Angelidaki營養鹽組成 62
3-2-3. 營養液調配 63
3-2-4. 批次產氣實驗植種來源 64
3-2-5. 批次產氣實驗數據模式迴歸法 64
3-3. 厭氧流體化床硬體改良 66
3-3-1. 反應槽組成結構 66
3-3-2. 反應槽功能設備改良 70
3-4. 操作策略與試程 76
3-5. 碳水化合物分析方法 80
3-5-1. 碳水化合物分析- Phenol-Sulfuric Acid method 80
3-5-2. 纖維素分析 81
3-6. 纖維素水解酵素分析方法 82
3-7. 厭氧微生物特性分析方法 86
3-7-1. 掃描式電子顯微鏡 Scanning Electron Microscope (SEM) 86
第四章、 結果與討論 87
4-1. 微生物利用纖維性基質之批次實驗生物分解性探討 87
4-1-1. 都市污水廠厭氧消化污泥分解纖維素產氣潛能探討 87
4-1-2. TCW1優勢菌分解濾紙,蔗渣,狼尾草及稻桿纖維物料之產氫潛能探討 91
4-2. 厭氧流體化床連續流操作分解功能探討 95
4-2-1. 第一試程,CMC批次進料之功能評估 95
4-2-2. 第二試程,木質纖維(tissue)進料再啟動功能評估 100
4-2-3. 第三試程,提升VLR至1.13 g-COD L-1 d-1之功能評估 107
4-2-4. 第四試程,混合木質纖維及廚餘進料之功能評估 116
4-3. 試程質量平衡探討 118
4-3-1. 質量平衡計算 118
4-3-2. 各試程氮平衡探討 119
4-3-3. 各試程電子平衡探討 122
4-3-4. 代謝產物與產氫關係探討 125
4-4. 鹼液回饋與產氣關係 129
4-4-1. 醱酵系統之酸鹼平衡 129
4-4-2. 碳酸鹽鹼液調配 131
4-4-3. 反應槽之酸化程度探討 133
4-5. 固態有機氮v.s.非碳水化合物VSS,及TCOD/TOC探討 136
4-6. 纖維水解酵素活性探討 138
4-6-1. TCW1的CMCase基本特性測試 138
4-6-2. 廚餘添加對纖維水解酵素活性影響 143
4-7. 菌相觀察 147
4-7-1. 掃描式電子顯微鏡(SEM)菌相觀察 147
第五章、 結論與建議 157
5-1. 結論 157
5-2. 疑問與建議 159
第六章、 參考文獻 160
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