ADSORPTION AND DECHLORINATION OF CHLOROPHENOLS UNDER ACIDOGENIC CONDITIONS MUN CHEOK HONG (B. Eng (Hons), NUS) A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF PHILOSOPHY DIVISION OF ENVIRONMENTAL SCIENCE AND ENGINEERING NATIONAL UNIVERSITY OF SINGAPORE 2008 Acknowledgement I would like to extend my deepest appreciation to my thesis adviser, Professor Ng Wun Jern. Prof Ng’s “freehand” mentoring approach has given me the opportunity and freedom to explore the scientific world and grow at my own pace; yet he never fails to point the way when I reached the crossroads. Thank you, Prof Ng, for being a great mentor. My heartfelt gratitude is also extended to my other adviser, Dr He Jianzhong. Her openness in sharing her research and personal experience has been most helpful in my scientific work. I would also like to pay tribute to the late Professor Aziz, who was my adviser in the 1st and 2nd year of my PhD study before he passed away. Special thanks to Associate Professor Liu Wen-Tso and Associate Professor Jeffrey Obbard, for serving on my doctoral committee and giving valuable comments to my research work. I am also grateful for the valuable comments provided by Professor F. Michael Saunders during the thesis examination. I am also grateful to Assoc Prof Liu and his team for their help and advice rendered in the molecular biology work. ii To the wonderful staff at the Water Science and Technology Laboratory, many thanks, especially to Mr Michael Tan and Mr Chandra for all their assistance; Mdm Tan, Leng Leng, and Hwee Bee for their help in the administration and instrumentation. In addition, I would like to say a big thank you to all my fellow classmates in the laboratory. Your encouragements, criticisms, laughter and more importantly, your friendship has accompanied me through these four years. To my father: Thank you for being the unsung hero in my life. Without your support, all these would not have been possible. Lastly, this thesis is dedicated to my dear wife, Jingwen, with all my love. iii Table of Contents ACKNOWLEDGEMENTS ii TABLES OF CONTENTS iv SUMMARY x LIST OF SYMBOLS xiv LIST OF TABLES xvi LIST OF FIGURES xviii CHAPTER Introduction 1.1 Background 1.2 Problem Statement 1.3 Objectives 1.4 Organization of Thesis CHAPTER Literature Review 2.1 Chlorinated Organic Compounds – An Environmental Problem 2.1.1 9 Physical properties and toxicity of carbon tetrachloride and chlorophenols 10 2.2 Biological Treatment of Chlorinated Organics 12 2.3 Anaerobic Process 15 2.3.1 Anaerobic process instability 17 2.3.2 Inhibition of methanogenesis by the chlorinated compounds 21 iv 2.4 Proposed Solution 23 2.4.1 Phase separation of the anaerobic process 23 2.4.2 25 Use of the acidogenic biotreatment for treatment of potentially inhibitory compounds 2.4.3 Advantage of using acidogenic process for treatment of 26 chlorinated compounds 2.4.4 Knowledge gap in the understanding of dechlorination 28 under acidogenic condition 2.5 Summary 35 CHAPTER Evaluation of the Biodegradation Potential of Carbon Tetrachloride and Chlorophenols under Acidogenic Condition 36 3.1 Introduction 37 3.2 Materials and Methods 38 3.2.1 Acidogenic sequencing batch reactor 38 3.2.2 Biodegradation experiment and anaerobic toxicity assay 41 3.2.3 Sorption experiment 42 3.2.4 Analytical procedures 43 3.3 Results 3.3.1 3.3.2 45 Removal of carbon tetrachloride in the acidogenic environment 45 Carbon tetrachloride transformation pathway 46 v 3.3.3 Negligible removal of carbon tetrachloride by adsorption 48 3.3.4 Removal of chlorophenol in the acidogenic environment 50 3.3.5 CCl4 and chlorophenol inhibition on acidogens 53 3.4 Discussion 56 3.5 Conclusions 58 CHAPTER Biomass Sorption Chlorophenols under Acidogenic Condition 59 4.1 Introduction 60 4.2 Materials and Methods 62 4.2.1 Enriched acidogenic cultures 62 4.2.2 Biosorption experiment 63 4.2.3 Biomass surface properties 66 4.2.4 Determination of Freundlich’s isotherm constants 66 4.2.5 Analytical procedures 67 4.3 Results and Discussion 68 4.3.1 Increased in 2,4,6-TCP adsorption during fermentation 68 4.3.2 Effect of pH on chlorophenols adsorption 69 4.3.3 Effect of pH on the surface properties of acidogenic 4.3.4 biomass 72 Effect of the metabolic state of acidogens on adsorption 75 vi 4.3.5 Comparison of chlorophenols adsorption between acidogenic and anaerobic sludge 4.4 Conclusions 77 78 CHAPTER Acidogenic Sequencing Batch Reactor Start-up Procedures For Induction of 2,4,6-Trichlorophenol Dechlorination 79 5.1 Introduction 80 5.2 Materials and Methods 80 5.2.1 Setup of sequencing batch reactor 80 5.2.2 Start-up procedure 81 5.2.3 Experimental phase 82 5.2.4 Batch test 83 5.2.5 Theoretical calculation of changes in Gibbs free energy 5.2.6 5.3 of formation 84 Analytical procedures 85 Results 5.3.1 86 Changes in Gibbs free energy formation under acidic condition 5.3.2 Stepwise pH reduction to induce 2,4,6-Trichlorophenol dechlorination under acidic condition 5.3.3 5.3.4 86 87 Start-up procedure favourable for acidogenic 2,4,6-TCP dechlorination 92 Dechlorination activity inhibitors 97 vii 5.4 Discussion 98 5.5 Conclusions 102 CHAPTER Pentachlorophenol dechlorination by an acidogenic sludge 103 6.1 Introduction 104 6.2 Materials and Methods 104 6.2.1 Reactor setup 104 6.2.2 Effect of PCP loading on its adsorption and dechlorination 6.2.3 Effect of initial aqueous concentrations of PCP on its dechlorination kinetics 106 6.2.4 Competitive adsorption of PCP and 2,4,6-TCP 107 6.2.5 Calculations of changes in Gibbs free energy of 6.2.6 6.3 105 formation 108 Analytical procedures 108 Results 6.3.1 109 PCP dechlorination requires a 2,4,6-TCP acclimated acidogenic sludge 109 6.3.2 Ortho dechlorination of PCP to 3,4,5-TCP 111 6.3.3 PCP removal by adsorption and dechlorination 113 6.3.4 Effect of initial PCP aqueous concentrations on its 6.3.5 dechlorination kinetics 116 PCP inhibit 2,4,6-TCP dechlorination 120 viii 6.4 Discussion 121 6.5 Conclusions 127 Conclusions and Recommendations 128 Major Findings 128 7.1.1 Reductive dechlorination under acidogenic condition 128 7.1.2 Different dechlorination mechanism for CCl4 and Chapter 7.1 chlorophenols 7.1.3 Similar dechlorination mechanism for 2,4,6-TCP and PCP 7.1.4 130 131 Adsorption behaviour of chlorophenol under acidogenic condition 131 7.2 Conclusions and Implications 132 7.3 Recommendations 133 REFERENCES 135 APPENDIX 152 A1 Detail calculation of ∆Go at different pH 152 A2 Publications from this research work 158 ix Summary Carbon tetrachloride (CCl4) and chlorophenols – with their wide spread industrial use as solvent and biocidal agents - are frequently discharged with industrial effluents. Together with the primary organic wastes, these industrial wastewaters are often treated by the single phase anaerobic process. Although the process is very effective in degrading both the primary and chlorinated organics, there is a serious shortcoming during the operation. The process often suffers from instability due to the methanogens’ sensitivity to pH fluctuation, volatile fatty acids (VFA) accumulation and to the presence of CCl4 and chlorophenols even at low concentrations. During the simultaneous degradation of the primary and the chlorinated organics, the dechlorination process needs to proceed quickly in order to prevent accumulation of the chlorinated compounds. Failing this, the methanogens may be inhibited and will lead to VFA accumulation and decrease in pH. The methanogenic and dechlorination process will then fail and thereafter the reactor may take several months to recover. 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Chapman and Hall, New York. 151 Appendix A1 Detail calculation of ∆Go at different pH Table Values used in the calculation of ∆Go Free energy (kJ/mol) Compound pKa Molecular Anion PCP -112.3 -86.6 4.5 3,4,5-TCP -86.8 -42.1 7.84 2,4,6-TCP -104.6 -69.1 6.0 4-CP -53.1 0.5 9.41 H2 +# -39.83 H Cl -131.3 - Note: Concentration of PCP, 3,4,5-TCP, 2,4,6-TCP, 4-CP and Cl- are assumed to be M. All values were obtained from Dolfing (1992). # Gibbs free energy formation at pH 7.0 a) 2,4,6-TCP dechlorination to 4-CP Due to different pKa of 2,4,6-TCP and 4-CP, 2,4,6-TCP and 4-CP will exist in different form at different pH. There are total of possible cases for the calculation of ∆Go. They are 1) Case 1: pH < pKa of 2,4,6-TCP – 2.0 2) Case 2: pKa of 2,4,6-TCP – 2.0 < pH < pH where [2,4,6-TCP-] = [4-CP] 3) Case 3: pH where [2,4,6-TCP-] = [4-CP] < pH < pKa of 4-CP + 2.0 4) Case 4: pH > pKa of 4-CP + 2.0 Solving for pH where [2,4,6-TCP-] = [4-CP] 10 −6 [H + ] = [ H + ] + 10 −6 [ H + ] + 10 −9.41 Solving for [H+], pH = 7.7 152 1) Case 1: at pH < 4.0, Both 2,4,6-TCP and 4-CP exist in their molecular form, so there is only governing equation. 2,4,6-TCP + 2H2 = 4-CP + 2H+ + 2Cl- (1) ∆Go = -211.1 - 11.38pH 2) Case 2: 4. < pH < 7.7, It is assumed that the 2,4,6-TCP exist both in its molecular and anionic form; 4-CP only exist in its molecular form. Thus there are possible reactions 2,4,6-TCP + 2H2 = 4-CP + 2H+ + 2Cl- (1) ∆G1o = -211.1 - 11.38pH 2,4,6-TCP- + 2H2 = 4-CP + H+ + 2Cl- (2) ∆G2o = -246.6 - 5.69pH The relative distribution of [2,4,6-TCP] and [2,4,6-TCP-] at various pH are governed by: [2,4,6TCP ] : α = [H + ] [ H + ] + 10 −6 [2,4,6TCP − ] : α = 10 −6 [ H + ] + 10 −6 Hence, ΔG o = α ΔG1o + α ΔG 2o 153 3) Case 3: 7.7 < pH < 11.4, It is assumed that the 4-CP exist both in its molecular and anionic form; 2,4,6-TCP only exist in its anionic form. Thus there are possible reactions 2,4,6-TCP- + 2H2 = 4-CP + H+ + 2Cl- (2) ∆G2o = -246.6 - 5.69pH 2,4,6-TCP- + 2H2 = 4-CP- + 2H+ + 2Cl- (3) ∆G3o = -193 - 11.38pH The relative distribution of [4-CP] and [4-CP-] at various pH are governed by: [4CP ] : α = [H + ] [ H + ] + 10 −9.41 [4CP − ] : α = 10 −9.41 [ H + ] + 10 −9.41 Hence, ΔG o = α ΔG 2o + α ΔG3o 4) Case 4: at pH > 11.4, Both 2,4,6-TCP and 4-CP exist in their anionic form, so there is only governing equation. 2,4,6-TCP- + 2H2 = 4-CP- + 2H+ + 2Cl- (3) ∆Go = -193 - 11.38pH 154 b) PCP dechlorination to 3,4,5-TCP Due to different pKa of PCP and 3,4,5-TCP, PCP and 3,4,5-TCP will exist in different form at different pH. There are total of possible cases for the calculation of ∆Go. They are 1) Case 1: pH < pKa of PCP – 2.0 2) Case 2: pKa of PCP – 2.0 < pH < pH where [PCP-] = [3,4,5-TCP] 3) Case 3: pH where [PCP-] = [3,4,5-TCP] < pH < pKa of 3,4,5-TCP + 2.0 4) Case 4: pH > pKa of 3,4,5-TCP + 2.0 Solving for pH where [PCP-] = [3,4,5-TCP] 10 −4.5 [H + ] = [ H + ] + 10 − 4.5 [ H + ] + 10 −7.84 Solving for [H+], pH = 6.17 1) Case 1: at pH < 2.5, Both PCP and 3,4,5-TCP exist in their molecular form, so there is only governing equation. PCP + 2H2 = 3,4,5-TCP + 2H+ + 2Cl- (1) ∆Go = -237.1 - 11.38pH 155 2) Case 2: 2.5 < pH < 6.17, It is assumed that the PCP exist both in its molecular and anionic form; 3,4,5-TCP only exist in its molecular form. Thus there are possible reactions PCP + 2H2 = 3,4,5-TCP + 2H+ + 2Cl- (1) ∆G1o = -237.1 - 11.38pH PCP- + 2H2 = 3,4,5-TCP + H+ + 2Cl- (2) ∆G2o = -262.8 - 5.69pH The relative distribution of [PCP] and [PCP-] at various pH are governed by: [ PCP ] : α = [H + ] [ H + ] + 10 − 4.5 [ PCP − ] : α = 10 −4.5 [ H + ] + 10 − 4.5 Hence, ΔG o = α ΔG1o + α ΔG 2o 3) Case 3: 6.17 < pH < 9.94, It is assumed that the 4CP exist both in its molecular and anionic form; PCP only exist in its anionic form. Thus there are possible reactions PCP- + 2H2 = 3,4,5-TCP + H+ + 2Cl- (2) ∆G2o = -262.8 - 5.69pH PCP- + 2H2 = 3,4,5-TCP- + 2H+ + 2Cl- (3) ∆G3o = -217.9 - 11.38pH 156 The relative distribution of [3,4,5-TCP] and [3,4,5-TCP-] at various pH are governed by: [H + ] [3,4,5TCP ] : α = [ H + ] + 10 −7.84 [3,4,5TCP − ] : α = 10 −7.84 [ H + ] + 10 −7.84 Hence, ΔG o = α ΔG 2o + α ΔG3o 4) Case 4: at pH > 9.94, Both PCP and 3,4,5-TCP exist in their anionic form, so there is only governing equation. PCP- + 2H2 = 3,4,5-TCP- + 2H+ + 2Cl- (3) ∆Go = -217.9 - 11.38pH 157 A2 Publications from this research work 1. C.H. Mun, W.J. Ng, and J. He (2008). Evaluation of the biodegradation potential of carbon tetrachloride and chlorophenols under acidogenic condition. Journal of Environmental Engineering, 134(3), 177-183. 2. C.H. Mun, W.J. Ng, and J. He (2008). Acidogenic sequencing batch reactor start-up procedure for induction of 2,4,6-Trichlorophenol dechlorination. Water Research, 42(6-7), 1675-1683. 3. C.H. Mun, J. He, and W.J. Ng, (2008). Pentachlorophenol dechlorination by an acidogenic sludge. Water Research, 42(14), 3789-3798. 158 [...]... the dechlorination mechanisms The scopes of studies were as followed: 1) Determine the factors and conditions required to induce dechlorination of CCl4 and chlorophenols under acidogenic condition; 2) Elucidate the mechanism of dechlorination for CCl4 and chlorophenols under acidogenic condition; and 3) Determine the extent of chlorophenols adsorption on to acidogenic biomass 1.4 Organization of the... C) VFA profile change 6.3 112 Removal mechanism of PCP under acidogenic condition: A) Effect of PCP loading on its initial and the average specific dechlorination rate, B) Effect of PCP loading on PCP removal efficiency at the end of 4 days of incubation and C) Typical adsorption and dechlorination profile of PCP during each batch experiment 6.4 Typical adsorption and dechlorination profile of PCP during... B) and C) Relationships between chlorophenols adsorption and pH 4.4 70 Effect of pH on A) adsorption of 2-CP, B) relative hydrophobicity and C) zeta potential of the acidogenic biomass 74 4.5 Effect of metabolic state of acidogens on adsorption 75 4.6 Comparison of the adsorption of 2,4,6-TCP and PCP between conventional anaerobic and acidogenic biomass at pH 7.2 77 5.1 Relationship between pH and. .. metabolic profile of an acidogenic biomass when first exposed to 25 µM of 2,4,6-TCP and 9000 mg/L of sucrose: A and B) Biomass, VFA, sucrose and pH time profile of an acidogenic biomass in serum bottles and C) 2,4,6-TCP aqueous concentrations over the same incubation period 4.3 64 Enhanced adsorption of chlorophenols with decreasing pH from 7.2 to 4.5: A) Freundlich isotherms for adsorption of chlorophenols, ... this study 7.1 117 Effect of co-solvent and surfactants on 1) PCP adsorption capacity and its degradation kinetics and 2) Fermentation activity 6.5 110 123 Comparison of the factors affecting reductive dechlorination of CCl4, 2,4,6-TCP and PCP under acidogenic condition 129 xvii List of Figures Figure 2.1 Description Gibbs free energy of formation of chlorophenols under aerobic and anaerobic reaction... biomass adsorption of 2,4,6-TCP and PCP adsorption at various pHs 5.1 69 Experimental protocol for investigating factors affecting 2,4,6-TCP dechlorination 83 5.2 Amount of chlorophenols adsorbed onto biomass 60 days after startup 89 5.3 Factors affecting the 2,4,6-TCP dechlorination under acidogenic condition 5.4 94 Mass balance of 2,4,6-TCP and its dechlorination metabolites 96 xvi 5.5 Effect of inhibitors... potential of Carbon Tetrachloride and Chlorophenols under acidogenic condition Chapter 3 evaluates the biodegradation potential of 2 separate classes of chlorinated compounds, namely carbon tetrachloride (aliphatic) and chlorophenols (aromatic), under acidogenic conditions The compounds had different fates in the acidogenic process; 1) carbon tetrachloride was dechlorinated to dichloromethane where the dechlorination. .. only via adsorption Chapter 4 - Biomass sorption of Chlorophenols under acidogenic condition Chapter 4, the factors affecting the adsorption phenomenon of phenol and chlorophenol was further investigated It was found that the adsorption was governed primarily by the pH of the solution The pH affected the protonation/deprotonation of the chlorophenolic compound and the surface properties of the acidogenic. .. 0 and D) Primary to chlorinated organic ratio (26.3 mM of sucrose/ 100 μM 2,4,6-TCP) 5.5 93 Specific loading rate of 2,4,6-TCP on acidogenic bioreactor (Experiment 3, Reactor 1) 96 6.1 Performance of Reactor 4 over a period of 6 months 105 6.2 Initial adaptation of 2,4,6-TCP dechlorinating acidogenic sludge to PCP: A) Dechlorination of PCP to 3,4,5-TCP, B) Inhibition of 2,4,6TCP dechlorination, and. .. Simultaneous degradation of the primary and chlorinated organics 3.1 Page 17 Removal efficiencies and residual concentrations of CCl4 at various influent concentrations after 280 minutes of contact time 45 3.2 Effect of initial sucrose concentration on removal of 10 mg/L of CCl4 46 3.3 Degradation metabolites in acidogenic environment A) Degradation of CCl4 B) Formation and degradation of chloroform C) Final . the understanding of dechlorination under acidogenic condition 28 2.5 Summary 35 CHAPTER 3 Evaluation of the Biodegradation Potential of Carbon Tetrachloride and Chlorophenols under Acidogenic. Typical metabolic profile of an acidogenic biomass when first exposed to 25 µM of 2,4,6-TCP and 9000 mg/L of sucrose: A and B) Biomass, VFA, sucrose and pH time profile of an acidogenic biomass. isotherms for adsorption of chlorophenols, B) and C) Relationships between chlorophenols adsorption and pH. 70 4.4 Effect of pH on A) adsorption of 2-CP, B) relative hydrophobicity and C) zeta