臺灣博碩士論文加值系統

多環芳香族碳氫化合物 (polycyclic aromatic hydrocarbons, PAHs) 在環境中是一種具有毒性難以分解的污染物，所以從環境中去除 PAHs 是一個相當重要的研究，而在許多的生物研究中 PAHs 已被證實可以在各種不同的厭氧條件下被微生物降解。本研究是以硫酸還原菌 (sulfate reducing bacteria, SRB) 為優勢菌的微生物族群來降解Fluorene，以中央合成設計法 (central composite design, CCD) 進行批次式實驗操作條件的規劃，同時找出硫酸鹽以及微生物濃度的最佳操作條件。再以最佳操作條件，改變 Fluorene 的濃度 (0-100 mg/L) 研究 Fluorene 對 SRB 的抑制。
實驗結果顯示以 SRB 為優勢菌的微生物族群可以利用 Fluorene 作為基質而生長，而 Fluorene 的零階、一階降解速率常數分別為0.21 mg/L-d 和 0.1/d。藉由反應曲面法 (response surface methodology, RSM) 找出硫酸鹽和微生物濃度的最佳操作範圍以及發展出一個 Fluorene 去除百分比的二階模式，找到最佳的操作點硫酸鹽濃度為14.4 mM，微生物濃度為37.8 mg/L，在此最佳條件下 Fluorene 的降解百分比為90%。
抑制實驗的數據以 Haldane’s equation 模式套配可得到動力參數 �慆ax、Ks以及 Ki，其數值分別為0.35/d、0.07 mg/L 以及600 mg/L。由於本研究所使用的營養鹽中含有酵母萃取物以及溶劑，微生物可以利用作為碳源使得無法在抑制實驗中觀察到 Fluorene 對 SRB 的毒性影響，但是可以推測 Fluorene 的濃度高於600 mg/L 時會對 SRB 造成抑制。另外在螢光原位雜交法 (fluorescent in-situ hybridization, FISH) 的分析，可以證明 SRB 在實驗所使用的微生物族群中為主要的優勢菌種，所佔的比例為85%。

Polycyclic aromatic hydrocarbons (PAHs) are one of the most important contaminants released in the environment. Anaerobic degradation of PAHs has been demonstrated in several microcosm studies. In this study, anaerobic fluorene degradation by sulfate reducing bacterial enrichment culture was investigated. Subsequently, batch biodegradation experiments were designed using the central composite design (CCD) and carried out to estimate the optimum operating conditions of sulfate reducing bacteria (SRB) and sulfate concentrations. Biodegradation rates of fluorene were 0.21 mg/L-d in zero-order and 0.1/d in first-order kinetics.

Response surface methodology (RSM) was used to optimize the sulfate and biomass concentrations, and was used to develop a model for the responses. Applying the desirability function, the optimum conditions of sulfate and biomass concentrations were found as 14.4 mM and 37.8 mg/L, respectively. The highest fluorene removal percentage of 90% was observed at optimum conditions.

In addition, the inhibition kinetics of fluorene biodegradation by SRB were investigated over a range of initial fluorene concentrations (from 0 to 100 mg/L) using the Haldane’s model. The kinetics parameters obtained were �慆ax of 0.35/d, Ks of 0.07 mg/L and Ki of 600 mg/L. Due to the addition of solvents (dichloromethane, N,N-dimethylformamide) and yeast extract in the PAHs biodegradation experiments, microorganisms grew using DMF and/or yeast extract which result no toxicity in the inhibition study. However, the fluorene biodegradation by SRB can be inhibited when fluorene concentration is higher than 600 mg/L. The FISH analysis showed that SRB was the major species in the enriched bacterial culture.

Contents
中文摘要 I
Abstract III
Contents V
List of Tables VII
List of Figures VIII
Chapter 1 Introduction 1
Chapter 2 Literature Review 3
2.1 Introduction to PAHs and Fluorene 3
2.2 Environmental Fate of PAHs and their Problems in the Environment 5
2.3 Anaerobic Biodegradation of PAHs 5
2.3.1 PAHs Biodegradation under Sulfate Reducing Conditions 8
2.4 Factors Influencing Biodegradation of PAHs 11
2.4.1 Concentration of Electron Acceptor 11
2.4.2 Biomass Concentration 12
2.4.3 pH and Temperature 12
2.5 Anaerobic PAHs Biodegradation Kinetics 16
Chapter 3 Materials and Methods 18
3.1 Introduction 18
3.2 Materials 18
3.3 Incubation 22
3.4 Experimental Methods and Design 23
3.4.1 Treatability Study 23
3.4.2 Optimization Study 25
3.4.3 Inhibition Study 26
3.5 Analytic Methods 27
3.5.1 PAH Extraction 27
3.5.2 PAH Quantification 27
3.5.3 Bacterial Cell Density Measurement 28
3.5.4 FISH Analysis 30
Chapter 4 Result and Discussion 33
4.1 Enrichment of SRB 33
4.2 Treatability of SRB Enrichment Culture 35
4.3 Optimization Study 39
4.3.1 Result of CCD 39
4.3.2 Response Surface and Contour Plot 44
4.3.3 Optimization of Fluorene Biodegradation 46
4.4 Confirmation Study 46
4.4.1 Evaluation of Predicted Model 46
4.4.2 Observations of FISH Analysis 51
4.4.3 Kinetics of Fluorene Biodegradation 53
4.5 Inhibition Study 58
4.5.1 Biodegradation of Fluorene 58
4.5.2 Inhibition Kinetics 60
Chapter 5 Summary and Conclutions 62
References 63

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