Plant Activity and Organic Contaminant Processing by Aquatic Plants A Dissertation Presented to The Academic Faculty By Jacqueline M. Tront In Partial Fulfillment Of the Requirements for the Degree Doctor of Philosophy in The School of Civil and Environmental Engineering Georgia Institute of Technology April, 2004 Copyright © Jacqueline M. Tront, 2004 Plant Activity and Organic Contaminant Processing by Aquatic Plants Approved by: Dr. F. Michael Saunders, Advisor Dr. Marc E. Frischer Dr. Ching-Hua Huang Dr. Frank Löffler Dr. Sotira Yiacoumi Date Approved: April 12, 2004 This work is deadicated to my family I have always relied on their friendship, love, and support as the stable platform from which I can reach for the stars ACKNOWLEDGEMENT As I come to the end of a long career as a student at Georgia Tech, I would like to thank my family, friends, and advisors for their advice, encouragement and support. Their continuing guidance had been critical in my pursuit of this endeavor. I am very grateful to the American Association of University Women, Department of Energy Education Research and Development Agency (ERDA), the School of Civil Engineering at Georgia Tech and the Georgia Tech Regional Engineering Program (GTREP) who have generously provided the financial support for this research. I would like to thank my committee members, Dr. Marc Frischer, Dr. Ching-Hua Huang, Dr. Frank Löffler, and Dr. Sotira Yiacoumi. Their counsel on both my research and professional development are greatly appreciated. I would also like to thank the rest of the faculty of the environmental engineering program at Georgia Tech for providing a positive environment that facilitates learning and collaboration. I would especially like to thank Dr. Saunders for being a wonderful advisor, teacher and mentor. Dr. Saunders has endured endless research meetings, countless manuscript drafts and plenty of high emotion. He has also helped me to celebrate my small triumphs and has pushed me to improve my understanding of the environmental engineering field through critical evaluation of my work and that of others. He has helped me to grow intellectually, professionally and personally through his counsel and example. I greatly appreciate the support of the student community in the environmental engineering program at Georgia Tech. There are too many names to list, but I know that many of the friendships I have made while I have been a graduate student at Georgia Tech will last a lifetime. I would also like to thank my friends that belong to the world outside of graduate school. Although infrequently acknowledged, their support and understanding has always been valued. I would like to thank Andrew for his love and friendship. I would not have made it without his encouragement, I have relied on his strength and support while we were both in Georgia and since he moved to the cold, cold north. Finally, I would like to recognize the support of my family as the most critical factor in my success in academics and enjoyment in all of life’s endeavors. I have always relied on their friendship, love, and support as the stable platform from which I can reach for the stars. iii TABLE OF CONTENTS Acknowledgement iii Table of Contents iv List of Tables ix List of Figures xi List of Acronyms and Symbols xx Summary xxv Chapter 1 Introduction 1 Problem Statement 2 Objectives 3 Chapter 2 Literature Review 5 Description of Aquatic Plants 6 Uptake, Sorption, Enzymatic Processing of Organic Contaminants in 7 Aquatic Plant Systems Uptake of Organic Contaminants by Aquatic Plants 8 Uptake of Pesticides by Aquatic Plant Systems 9 Uptake of Nitroaromatics by Aquatic Plant Systems 12 Model of Uptake Kinetics of Technetium-99 by Lemna. minor 13 Uptake of Chlorinated Aliphatics by Aquatic Plants 14 Uptake of Chlorinated Phenols by Plants 14 Predictive Relationships to Describe Uptake in Terrestrial 16 Plant Systems Enzymatic Processing of Organic Contaminants by Aquatic Plants 17 Transformation Processes Involved in Enzymatic Processing 17 by Plants Transformation of Pesticide Compounds by Aquatic Plants 23 Transformation of TNT by Aquatic Plants 26 Transformation of Chlorinated Aliphatics by Aquatic Plant 27 Transformation of Chlorinated Phenols by Plants 27 Metabolism of Chlorinated Phenols by Microorganisms 31 Effects of Inhibition on Aquatic Plant Uptake and Enzymatic 32 Processing of Organic Contaminants iv Plant Toxicity Responses 36 Toxicity Assessment Background 36 Plant Exposure Protocols 47 Toxic Effect Endpoints 48 pH Effects 49 Biological Indicators 52 Organism Level Indicators 53 Cellular Level Indicators 57 Characterization of Chlorinated Phenols 61 Industrial Use of Chlorophenols 62 Relevant Chemical and Physical Properties of Chlorinated Phenols 63 Vapor Pressure 63 Acidity 65 Solubility 68 Volatilization 70 Hydrophobicity 73 Air-Water Partitioning 76 Organic Matter Partitioning 76 Chlorophenol Toxicity 78 Environmental Persistence of Chlorophenols 81 Photolytic, Photooxidative and Hydrolytic Degradation 81 Microbial Degradation 82 Fluorinated Analogs of Chlorophenols 83 Physical and Chemical property variations 84 Biological Activity of Fluorinated organics 86 Fluoride in Biological Systems 87 Nuclear Magnetic Resonance 88 Analytical Overview 88 Identification and Detection of Metabolites 91 Chapter 3 Materials and Methods 94 Experimental Reactor Protocol 94 v Oxygen Production Rate Measurements 98 14C Studies 100 Analytical Methods 104 HPLC 104 GC-TCD 105 Nuclear Magnetic Resonance 105 Sample Preparation 105 Instrumentation Used 106 Plant Sources and Stock Cultures 107 Chapter 4 Uptake and Sequestration of Organic Contaminants by Aquatic 108 Plants Photolysis Controls 108 Inactivated Plant Controls 111 Active Plant Uptake of Organic Contaminants 117 Contaminant Uptake by L. minor 120 Solvent Enhanced Contaminant Uptake. 125 Photoperiod Effects on Contaminant Uptake 125 Contaminant Uptake by M. aquaticum 128 Effect of Chemical Structure and Physicochemical Parameters on 132 Rate of Halogenated Phenols Uptake by Aquatic Plants Halogenated Phenols Examined 134 Calculation of Molecular Physicochemical Properties 135 Inactivated Controls 136 Plant Uptake of Halogenated Phenols 136 Comparison of Halogenated-Phenol Uptake with Literature Predictive 147 Relationships Effect of Speciation Internal to Plant on Contaminant Uptake Rate 156 Discussion 158 Chapter 5 Transformation and Sequestration of Chlorophenols by Aquatic 162 Plants Sequestration of Contaminants by Aquatic Plants 162 2,4,5-Trichlorophenol Sequestration by L. minor and M. aquaticum 162 2,4-Dichlorophenol Sequestration by L. minor 172 vi Measurement of Parent Compounds and Metabolic Products Sequestered in 178 Plants Using 19F NMR Identification of Chlorophenols in Liquid Extracts via 19F NMR 180 Quantification of Chlorophenols in Liquid Extracts via 19F NMR 191 Discussion 200 Chapter 6 Role of Plant Activity and Contaminant Speciation in Aquatic Plant 203 Assimilation of Organic Contaminants Assessment of Plant Activity (α) 203 Oxygen Production Rate (α) as a Measure of Plant Activity 204 Integration of Plant Activity, Contaminant Speciation and Contaminant 210 Uptake Rate Plant Activity Effects on Contaminant Uptake and Assimilation 210 pH Dependant Contaminant Uptake 219 Discussion 227 Chapter 7 A Conceptual Model for Contaminant Uptake by Aquatic Plants 230 Incorporating of Inhibition, Plant Activity, and Contaminant Speciation Effects of Inhibition of on Uptake of 2,4,5-TCP by L. minor 233 Plant Activity and Inhibition (α, β) 233 Relative Plant Activity and Uptake of 2,4,5-TCP. 234 Conceptual Model to Describe Effects of Inhibition on Contaminant 242 Uptake Contaminant Uptake, pH, and Plant Activity 245 Contaminant Partitioning Into Plants 246 Enzymatic Contaminant Processing by Plants 248 Prediction of Contaminant Uptake 249 Prediction of Contaminant Uptake Given Relative Inhibition (β) and 249 Plant Activity (α). Toxicity Relationship for Relative Plant Activity 254 Comparison of Contaminant Uptake and Enzymatic Processing 256 Predictions with Independent Data Discussion 272 vii Chapter 8 Anaerobic Microbial Degradation of Plant Sequestered 280 Contaminant Reduction of Halogenated Phenols by Desulfitobacterium sp. strain 280 Viet1 Reduction of Plant-Sequestered 2,4-DCP by strain Viet1 287 Discussion 294 Chapter 9 Summary of Results 303 Uptake of Halogenated Phenols by Aquatic Plants 303 Sequestration and Metabolism of Halogenated Aromatics by Aquatic 313 Plants Role of Aquatic Plants in Natural and Engineered Systems 315 Future Research Needs 318 Literature Cited 321 Vita 341 viii LIST OF TABLES Table 2.1. Identified metabolic products of chlorinated phenols. 35 Table 2.2. Reported toxicity information for Lemna species. 40 Table 2.3. Comparison of estimated vapor pressure of 2,4,5-TCP with literature 64 data. Table 2.4. Comparison of estimated values of the acidity constant of 2,4,5-TCP 66 with literature data. Table 2.5. Comparison of estimated and literature values of the aqueous solubility 69 of 2,4,5-TCP. Table 2.6. Comparison of estimated and literature values for Henry’s Law 72 constant of 2,4,5-TCP. Table 2.7. Comparison of estimated and literature values of the octanol-water 75 partition coefficient of 2,4,5-TCP. Table 2.8. 2,4,5-TCP toxicity data (from Mackay et al., 1985). 79 Table 2.9. 2,4-DCP toxicity data (from Mackay et al., 1985). 80 Table 2.10. Comparison of physical and chemical properties for fluorinated 85 analogs. Table 3.1. Experimental series detailed herein with corresponding contaminants 97 examined, plants used and experimental parameters used. The chapter of thesis where experimental results are described is listed. Table 4.1. Values of C0, fitted closely correspond to the concentration measured at 126 t=5 min and do not agree with initial concentrations. Table 4.2. Halogenated phenol experimental data. 141 Table 5.1. Material balance for 14C-labeled 2,4,5-trichlorophenol in L. minor 167 system. Table 5.2. Material balance for 14C-2,4,5-trichlorophenol and M. aquaticum 171 system. Table 5.3. Material balance for 14C-2,4-dichlorophenol and L. minor system. 177 ix Table 6.1. Example data for examination of error associated with normalization of 221 k to α.