三氯乙烯(trichloroethylene, TCE)為地底下之廣泛污染物，目前我國TCE污染場址已經有22處被列管為整治場址(土壤及地下水污染整治網, 2015)，TCE具有高的密度(1.46 g/mL)和低水溶性(1,100 mg/L)之特質，因此是屬於長期性的工作。現地生物復育(in situ bioremediation, ISB)是較為經濟可行的整治方式，惟生物降解需長期注入基質刺激微生物的生長及發展，促進還原脫氯反應(reductive dechlorination)，因此整治區經常選擇緩慢釋放有機基質，但基質之注入常造成阻塞及地下水酸化問題，使得基質之傳輸效果不佳，並影響地下水水質惡化。此外，有機基質在未飽和層中無法長時間停由於土壤層中，使得微生物生長有限，並無法與污染物接觸並降解，因此未飽和層無法達到整治成效。本研究將於土壤及地下水分別發展出適用於土壤層之阻斷式生物凝膠基質(blocked bio-gel substrate, BBS)及適用於地下水層緩衝膠體基質(buffered colloidal substrate, BCS)。本研究已開發出BBS，其界達電位為負值且表面具有網狀立體節構，能緩慢吸附污染物；由試驗結果得知BBS可讓營養素有效地附著於未飽和層中，並能長期提供碳源及營養物質供微生物利用，進而增加還原脫氯效果，使該微生物去除1,2-二氯乙烷，因此BBS適用作為未飽和土壤層之阻斷式生物凝膠基質，除可阻斷及侷限DNAPL之擴散和下滲外，亦可緩釋基質以加強未飽和層DNAPL之生物降解。另外本研究已研發出BCS，經批次試驗結果得知，其除了作為電子提供者促進生物降解外，希望可長期提供緩慢釋放鹼度提供飽和層地下水緩衝能力，維持最適生物反應之pH值(中性)，使地下水環境維持適合現地微生物生長之環境，並提升污染物降解效率。本研究於土壤及地下水分別發展出適用於土壤層之BBS及適用於地下水層之BCS，並且形成生物侷限及整治牆概念作污染源及邊界控制外，亦可達到風險管控之目的，使阻斷式生物凝膠基質及緩衝膠體基質成為一種更具經濟效益且突破傳統設計框架之綠色整治工法，符合現地、生物、被動之永續式綠色整治設計概念。

Soil and groundwater at many existing and former industrial areas and disposal sites are contaminated by halogenated organic compounds that were released into the environment. When they are released into the subsurface, they tend to adsorb onto the soils and cause the appearance of DNAPL (dense-non-aqueous phase liquid) pool. In this study, TCE will be used as the target compound for the feasibility and pilot-scale studies. One cost-effective approach for the remediation of the chlorinated-solvent contaminated aquifers is the installation of permeable reactive zones or barriers within aquifers. As contaminated groundwater moves through the emplaced reactive zones, the contaminants are removed, and uncontaminated groundwater emerges from the downgradient side of the reactive zones. Application of in situ anaerobic bioremediation is a feasible technology to remediate DNAPL site. However, enhanced in situ bioremediation requires the injection of primary substrates, which would cause the acidification and odor problems of the subsurface environment. This would deteriorate the groundwater quality and cause the increase in maintenance cost. The objective of this proposed study is to develop a blocked bio-gel substrate (BBS) and buffered colloid substrate (BCS) to bioremediate unsaturated and saturated zones contaminated with DNAPL, respectively. The BBS can contain and encapsulate the DNAPL in the unsaturated zone and prevent its further migration. The released substrate from the gel can enhance the reductive dechlorination of DNAPL in the unsaturated zone. The BCS can be applied in the saturated zone, which can release substrate from the colloid to enhance the reductive dechlorination of DNAPL in the saturated zone. Furthermore, the buffered colloid has the capability for pH control and prevents the decrease in pH value in groundwater. During the first-year research period, the unique BBS and BCS will be developed and the saponification will be studied. The BBS will contain slow carbon-releasing materials and biological gel to form a gel-like material. We will conduct the batch experiments to test the produced gel. The saponification test results would help on the design of a buffered colloid for pH control. The BCS has the capability for pH control and long-term substrate release. Several experimental conditions include the concentrations of contaminants and substrates, shacking speed, stability test, and percentage of each component will be tested. During the second calendar year, column experiments will be performed to evaluate the effectiveness of using the produced substrates as the primary substrates for DNAPL control and biodegradation. The possible TCE degradation byproducts will be also evaluated. During the third year of this proposed study, we will select a DNAPL-contaminated site to apply the designed substrates for field application. The results can be used to predict interactions and distribution of contaminant mass expected after substrate injection, and thus provides a more accurate estimate of the mass of DNAPL removed due to enhanced biodegradation. In this third calendar year, we will apply the Metagenomics technique and real-time polymerase chain reaction (PCR) to analyze the microbial diversity to obtain the metabolic routes of TCE biodegradation. Results of this study will aid in designing an in situ biobarrier system containing slowly released biocolloid for remedial application. The proposed treatment scheme would be expected to provide a more cost-effective alternative to remediate chlorinated-solvent contaminated aquifers. Knowledge obtained from this study will aid in designing a reactive barrier system containing inhibitive biological gel substrate and buffered colloid substrate for site remediation.

論文審定書 i
論文公開授權書 ii
謝 誌 iii
摘要 iv
Abstract v
目錄 vii
圖目錄 x
表目錄 xii
第一章 前言 1
1.1研究緣起 1
1.2研究目的 2
第二章 文獻回顧 3
2.1土壤及地下水受含氯脂肪族碳氫化物污染之近況 3
2.1.1 含氯脂肪族碳氫化物污染概述 3
2.1.2含氯脂肪族碳氫化物之性質及管制標準 6
2.1.3 含氯脂肪組碳氫化合物之傳輸 9
2.2常見的土壤與地下水污染整治技術 13
2.2.1 土壤與地下水生物整治技術 14
2.2.2 綠色整治技術 20
2.3 含氯碳氫化合物生物反應機制 22
2.3.1 含氯脂肪族碳氫化物好氧共代謝反應機制 22
2.3.2 含氯脂肪族碳氫化物厭氧還原脫氯反應機制 25
2.4 新穎土壤未飽和層與地下水飽和層生物整治技術 27
2.4.1 土壤阻斷式生物基質操縱技術 27
2.4.2 地下水緩衝膠體基質控制技術 31
2.5 分子生物技術運用於土壤與地下水整治 35
2.5.1 微生物多樣性之檢測方法 36
2.5.2 以即時定量PCR監測脫氯菌種與基因 37
第三章 實驗材料與方法 40
3.1 研究流程 40
3.2 實驗材料 42
3.2.1實驗藥品 42
3.2.2 實驗設備 42
3.3 阻斷式生物凝膠基質 43
3.3.1最佳合成配比與穩定性 43
3.3.2侷限試驗 43
3.3.3阻斷式生物凝膠基質分解實驗 43
3.4 緩衝膠體基質 43
3.4.1 緩衝膠體基質基本性質試驗 43
3.4.2緩衝膠體基質厭氧批次降解實驗 44
3.4.3現地模場試驗 46
3.5 實驗分析方法 47
3.5.1 水質分析 47
3.5.2 掃描式電子顯微鏡(SEM) 47
3.5.3 分子生物技術 48
第四章 結果與討論 54
4.1 阻斷式生物基質之基本性質 54
4.1.1阻斷式生物凝膠基質之合成 54
4.1.2 最佳合成配比及穩定性試驗 54
4.1.3侷限試驗 55
4.2 微生態系統阻斷式生物基質批次分解實驗 58
4.2.1 阻斷式生物凝膠基質/水對1,2-DCA之分配係數 58
4.2.2 BBS對於1,2-DCA之去除效率試驗 59
4.2.3 污染物吸附試驗 60
4.2.4 污染土阻絕試驗 63
4.2.5 環境掃描式電子顯微鏡觀察BBS 64
4.3 緩衝膠體基質之基本性質 66
4.3.1緩衝膠體基質之合成 66
4.3.2乳化試驗 66
4.3.3 穩定性試驗 68
4.4 微生態系統緩衝膠體基質批次生物降解實驗 70
4.4.1水質參數變化趨勢 70
4.4.2化學參數變化之趨勢 72
4.4.3污染物變化趨勢 75
4.4.4 土壤表面外觀 76
4.5 分子生物技術 77
4.5.1微生物菌相分析(DGGE) 77
4.5.2批次實驗之微生物菌種鑑定 80
4.5.3脫氯菌之菌量趨勢變化 89
4.5.4還原脫鹵酶基因數量變化趨勢 90
4.6模場試驗 92
4.6.1 DO、pH與ORP 93
4.6.2厭氧環境下的地下水化學參數變化 96
4.6.3三氯乙烯及副產物之降解趨勢 100
4.7模場分子生物技術分析 103
4.7.1 Dehalococcoides spp.菌屬數量變化趨勢 103
4.7.2還原脫鹵酶基因數量變化趨勢 104
4.7.3微生物菌相分析 107
4.7.4定序 110
4.8基質成本效益評估 112
第五章 結論與建議 114
5.1結論 114
5.2建議 115
參考文獻 116

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