臺灣博碩士論文加值系統

廉價的基質有助於提升暗醱酵產氫的競爭力，為發展非糧料源之模場產氫技術，本研究分別以60 L 及400 L 模場進行退漿廢水及濃縮糖蜜醱酵廢液之產氫試驗，期鹿簳?W模放大之相關參數並與商業
化程序接軌。此外，本研究亦以分子生物技術分析各系統之菌相結構，輔以觀察模場醱酵產氫系統內之微生物消長。
退漿廢水先以實驗室規模之CSTR (continuous flow stirred tank reactor)反應器進行各項環境因子及操作策略之試驗，結果顯示料源預熱處理(60℃、1小時)配合 HRT (hydraulic retention time) 12 h之操作，
可抑制非產氫菌的活性並改善系統操作之穩定性。最適之pH 值為5.5，然而卻無法穩態操作於HRT 8 h，而退漿廢水經糖化處理則可改善此問題，於HRT 8 h 其HPR (H2 production rate)、HY (H2 yield)及
HPE (overall H2 production efficiency)分別可達15.2 m3/m3/d、9.77 mmol-H2/g-hexose 及37.5%，若配合鹼處理(pH 10.0-10.5)則可進一步提升氫氣產率。
不幸地，經各項預處理之退漿廢水無法以60 L 模場順利啟動，然於濃縮飲料廢品中添加0-10% (總糖比例)之退漿廢水則可順利操作於HRT 6 h，HPR、HY 及HPE 分別可達20.2 m3/m3/d 、8.7
mmol-H2/g-hexose 及37.2%。菌群結構分析顯示添加0-10%退漿廢水醱酵產氫之模場操作之各試程其系統內均具有產氫菌Clostridium sp.，並以C. butyricum 為優勢產氫菌。然而，當退漿廢水添加量增至20%，系統產氫狀態隨即
降至零，菌群結構分析顯示系統中仍存有C. butyricum，據此推測無法順利產氫之主因應非生物性的問題，可能退漿廢水含有抑制產氫之化學物質。
以糖蜜醱酵液進行400 L 模場產氫試驗，系統可於HRT 6 h (Cs =40 g COD/L)成弗珧吽A但隨操作的持續進行，系統之產氫效能發生週期性升降的現象，透過基質預熱策略(60℃, 1 h)可明顯改善此問題，於HRT 4 h 及基質濃度70 g COD/L (總糖濃度17.7 g/L)之條件，HPR
及HY 分別可達17.8 m3/m3/d 及9.1 mol H2/g-hexose。菌群結構分析顯示，基質預熱可使原存於系統中之Corynebacterium glutamicum 趨於弱勢，使系統由不利於產氫之乳酸醱酵轉變成有利於產氫之丁酸醱
酵。另外，於預熱系統中發現存在可分解醣類及乳酸而產生氫氣之Megasphaera sp.，並由液相代謝物組成之乳酸含量降低推測其對於產氫具有相當程度的貢獻。
於弱勢，使系統由不利於產氫之乳酸醱酵轉變成有利於產氫之丁酸醱
酵。另外，於預熱系統中發現存在可分解醣類及乳酸而產生氫氣之
Megasphaera sp.，並由液相代謝物組成之乳酸含量降低推測其對於產
氫具有相當程度的貢獻。

The cheap substrate can improve competitiveness for dark
fermentative hydrogen production. In order to develop the pilot
technology of hydrogen production from non-grain feedstock, this study
explored the hydrogen production from desizing wastewater (DW) and
condensed molasses fermentation soluble (CMS) using 60 and 400 L
pilot-scale fermentors, respectively. It is expected that experiment results
obtain several related parameters for scale up and futher connect with
commercial processes. In addition, the microbial community structure in
systems was analysed by using molecular biological techniques.
DW was initially conducted the experiments of environmental
factors and operation strategies with a lab-scale continuous flow stirred
tank reactor (CSTR). The results showed that combination substrate
preheating (60℃, 1 h) and HRT (hydraulic retention time) 12 h could
inhibit the activity of non-hydrogen-producing bacteria and improve the
stability of system operation. The optimum pH value was 5.5, but the
CSTR reactor could not be operated steady at a HRT of 8 h. DW via
saccharification pretreatment could improve this problem, obtaining a H2
production rate (HPR), H2 yield (HY) and overall H2 production
efficiency (HPE) of 15.2 m3/m3/d, 9.77 mmol-H2/g-hexose and 37.5%,
respectively, at a HRT of 8 h. If the pretreatment processes combined
saccharification with alkali (pH 10.0-10.5) treatment, it could further
improve the hydrogen yield.
Unfortunately, the 60 L pilot-scalr reactor could not be started up by
feedimg variously pretreated DW. However, the waste concentrated
beverage added 0-10% (total sugar ratio) DW could be successfully
operated at HRT 6 h, getting a HPR, HY and HPE of 20.2 m3/m3/d, 8.7
mmol-H2/g-hexose and 37.2%, respectively. The analytical results of
microbial community structure showed that hydrogen-producing
Clostridium sp. existed in the reactor for three stages of addition of
0-10% DW, and C. butyricum was the predominant H2-producing
bactrrim in the system. However, hydrogen production decreased to near
zero when the addition of DW was increased to 20%. The analytical
results of microbial community structure showed that C. butyricum still
presented in the reactor. Accordingly, the main cause of stopping
hydrogen production should not result from propagation of non-H2-
producing bacteria in the fermentor. It is highly probable that chemical
substances of hydrogen inhibition existed in desizing water.
The 400 L pilot–scale bioreactor could successfully start up by
feeding CMS at HRT 6 h (Cs = 40 g COD/L). However, the performance
of hydrogen production became periodic fluctuation situation. This
problem could be improved by substrate preheating (60℃, 1 h), obtaining
a HPR and HY of 17.8 m3/m3/d and 9.1 mol H2/g-hexose, respectively, at .
HRT 4 h and substrate concentration 70 g COD/L (total sugar
concentration 17.7 g/L). The analytical results of microbial community
structure showed that Corynebacterium glutamicum presented in the
system became non-predominant bacterium by substrate preheating,
resulting in the shift from lactate fermentation into butyrate fermentation.
In addition, the Megasphaera sp., which can decompose carbohydrates
and lactate to produce hydrogen, was found that existed in the system of
substrate preheating. According to the desrese of lactate content in
soluble metabolites, there was speculatation that Megasphaera sp. gave
some contribution to hydrogen production.

誌謝 ..................................................................................................... I
摘要 ..................................................................................................... I
ABSTRACT ...................................................................................... III
目錄 .................................................................................................... V
圖目錄 ................................................................................................ X
表目錄 ............................................................................................. XII
符號說明 ......................................................................................... XIV
第一章 緒論 ...................................................................................1
1-1 前言 ...............................................................................................1
1-2 研究動機及目的 ..............................................................................2
1-3 研究架構 ........................................................................................3
第二章 文獻回顧 ............................................................................7
2-1 能源的重要性 .................................................................................7
2-2 氫能 ...............................................................................................8
2-2-1 氫能價值 ..............................................................................8
2-2-2 氫製備技術 ...........................................................................9
2-3 生物產氫 ...................................................................................... 10
2-3-1 光合作用 ............................................................................ 14
2-3-2 光醱酵 ................................................................................ 17
2-3-3 暗醱酵 ................................................................................ 19
2-4 厭氧醱酵產氫微生物之特性 ........................................................... 23
2-4-1 Enterobacter sp. .................................................................. 23
2-4-1 Bacillus sp. ......................................................................... 24
2-4-1 Clostridium sp. .................................................................... 27
醱酵產氫之模場操作
vii
逢甲大學e-Theses & Dissertations (99 學年度)
2-4-2 Klebsiella sp. ....................................................................... 32
2-4-3 Megasphaera sp. .................................................................. 36
2-5 環境因子對醱酵產氫的影響 ........................................................... 38
2-5-1 有機廢水 ........................................................................... 38
2-5-1-1 濃縮糖蜜醱酵液 ............................................................................ 38
2-5-1-2 紡織廠退漿廢水 ............................................................................ 43
2-5-1-3 濃縮飲料廢品 ................................................................................ 46
2-5-2 營養鹽及微量金屬 ............................................................... 46
2-5-3 水力滯留時間及有機負荷速率.............................................. 50
2-5-4 pH ...................................................................................... 52
2-5-5 ORP.................................................................................... 54
2-6 厭氧醱酵產氫系統 ........................................................................ 56
2-6-1 CSTR .................................................................................. 56
2-6-2 CIGSB ................................................................................ 57
2-7 顆粒污泥 ...................................................................................... 59
2-7-1 顆粒污泥形成機制 ............................................................... 59
2-7-2 顆粒污泥結構...................................................................... 62
2-8 分子生物技術於生物產氫之應用 .................................................... 64
2-8-1 分子生物技術...................................................................... 64
2-8-2 16S rDNA 及funct io nal gene ................................................ 66
2-8-3 聚合酶連鎖反應 ................................................................. 68
2-9 國內外模場產氫技術 ..................................................................... 73
第三章 研究材料及方法 ................................................................ 77
3-1 退漿廢水醱酵產氫 ...................................................................... 77
3-1-1 實驗室規模之產氫試驗 ........................................................ 77
3-1-1-1 產氫污泥及培養基配方 .............................................................. 77
3-1-1-2 退漿廢水前處理 ......................................................................... 77
醱酵產氫之模場操作
viii
逢甲大學e-Theses & Dissertations (99 學年度)
3-1-1-3 CSTR 反應器之產氫操作 ........................................................... 78
3-1-2 60 L 模場規模之產氫試驗 ................................................. 79
3-1-2-1 60 L 模場系統之設計 ................................................................. 79
3-1-2-2 60 L 模場系統之建造 ................................................................. 82
3-1-2-3 60 L 模場系統試車 ..................................................................... 83
3-1-2-4 60 L 模場系統改良 ..................................................................... 84
3-1-2-5 退漿廢水之模場產氫試驗 .......................................................... 87
3-1-2-6 退漿廢水與濃縮飲料廢品混合進料之模場產氫試驗 ................ 87
3-2 400 L 模場規模之濃縮糖蜜醱酵廢液產氫.............................. 87
3-2-1 產氫污泥及培養基配方 ................................................................. 87
3-2-2 濃縮糖蜜廢液之模場產氫試驗 ..................................................... 88
3-3 氣相組成分析............................................................................... 90
3-4 液相組成分析............................................................................... 91
3-5 水質分析 ...................................................................................... 92
3-5-1 一般水質分析 .................................................................... 92
3-5-2 總糖分析 ........................................................................... 92
3-6 菌相分析 ..................................................................................... 94
3-6-1 DNA 萃取 ........................................................................... 94
3-6-2 聚合酶鏈鎖反應 .................................................................. 96
3-6-3 變性梯度明膠電泳 ............................................................... 98
3-6-4 Acryla mide/Bis 膠體之DNA 純化 ......................................... 99
第四章 結果與討論 ..................................................................... 100
4-1 退漿廢水之產氫試驗 ................................................................... 100
4-1-1 退漿廢水性質 .................................................................. 100
4-1-2 實驗室規模之產氫試驗 ...................................................... 102
4-1-2-1 熱處理之效應 ........................................................................... 102
4-1-2-2 HRT 及基質濃度之效應 ........................................................... 103
4-1-2-3 pH 值之效應 ............................................................................. 106
4-1-2-4 糖化處理之效應 ....................................................................... 109
醱酵產氫之模場操作
ix
逢甲大學e-Theses & Dissertations (99 學年度)
4-1-2-5 鹼處理之效應 ........................................................................... 113
4-1-3 60 L 模場規模之產氫試驗 ......................................................... 117
4-1-3-1 以蔗糖進行模場試車 ..................................................... 117
4-1-3-2 糖化處理之退漿廢水模場產氫 ................................................ 123
4-1-3-3 糖化及鹼處理之退漿廢水模場產氫......................................... 128
4-1-3-4 退漿廢水與濃縮飲料廢品混合進料之模場產氫 ..................... 133
4-1-3-5 60 L 模場規模之菌群結構分析 ................................................ 140
4-3 400 L 模場規模之產氫試驗 ........................................................ 150
4-3-1 400 L 模場規模之糖蜜醱酵廢液產氫試驗 ......................... 150
4-3-1-1 基質效應 ................................................................................... 151
4-3-1-2 HRT 及OLR 效應 ..................................................................... 152
4-3-1-3 基質預處理效應 ....................................................................... 153
4-3-1-4 400 L 模場規模之菌群結構分析 .............................................. 157
第五章 結論與建議 ..................................................................... 159
5-1 結論 ........................................................................................... 159
5-2 建議 ........................................................................................... 161
參考文獻 .......................................................................................... 162
附錄 ................................................................................................. 178
附錄1 .............................................................................................. 178
作者簡歷 .......................................................................................... 179

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