Formation of Iodinated Disinfection By-Products From

Home , Acetrizoic acid, Diodone, Iobenzamic acid, Iobitridol, Iocarmic acid, Iocetamic acid

FORMATION OF IODINATED DISINFECTION BY-PRODUCTS FROM

IODINATED X-RAY CONTRAST MEDIA, IOPAMIDOL, IN THE PRESENCE

OF NOM AND CHLORINATED OXIDANTS.

A Thesis

Presented to

The Graduate Faculty of The University of Akron

In Partial Fulfilment

of the Requirements for the Degree

Master of Science

Elizabeth Ann Crafton

December, 2014 FORMATION OF IODINATED DISINFECTION BY-PRODUCTS FROM

IODINATED X-RAY CONTRAST MEDIA, IOPAMIDOL, IN THE PRESENCE

OF NOM AND CHLORINATED OXIDANTS.

Elizabeth Ann Crafton

Thesis

Approved: Accepted:

______Advisor Department Chair Dr. Stephen E. Duirk Dr. Wieslaw Binienda

______Committee member Dean of the College Dr. Teresa J. Cutright Dr. George K. Haritos

______Committee member Interim Dean of Graduate School Dr. Lan Zhang Dr. Rex Ramsier

______Date ii ABSTRACT

The objective of this study was to investigate the formation of iodinated disinfection

by-products (iodo-DBPs) where iodinated x-ray contrast media (ICM), Iopamidol, acted as source of iodine, with respect to pH (6.5, 7.5, 8.5 and 9.0) in the presence of natural organic matter (NOM) and chlorinated oxidant. This was achieved by varying the NOM concentration as well as iopamidol concentrations.

Iodinated trihalomethane (iodo-THM) formation was highest at pH 9.0 for chlorine and at pH 6.5 for monochloramine. A considerable increase in formation was observed from pH 6.5 to 7.5 and from 8.5 to 9.0 with respect to chlorine as oxidant.

Monochloramine expressed a decreasing trend from pH 6.5 to 9.0. The most predominately formed iodo-THM was dichloroiodomethane, 57.5 nM for chlorine at pH 9.0 (DOC = 2.51 mg/L-L) and 35.2 nM for monochloramine at pH 6.5 (DOC =

2.51 mg/L-L). Chloroform formation was also impacted by the introduction of iopamidol to reactor. Monochloramine produced a wider variety of iodo-DBPs at lower concentrations in comparison to chlorine.

Variation of NOM proved to impact the formation of iodo-DBPs. When NOM levels were reduced to a quarter of the original capacity (2.51 – 0.63 mg/L-L) dichloroiodomethane formation doubled at pH 9.0 with respect to chlorine (110.5 nM). Monochloramine expressed a decreasing trend with respect to decreasing levels of NOM with the source water. For both oxidants, half and quarter capacity of NOM,

1.26 and 0.63 mg/L-L respectively, expressed considerably similar formation.

iii Additionally, two-way analysis of variance (two way-ANOVA) tables were generated for two response variables, chloroform and dichloroiodomethane. With respect to each oxidant, iopamidol and pH were evaluated while NOM level remained constant and NOM and pH were evaluated while iopamidol remained constant. In regard to chloroform as the response variable with varied DOC, significance was yielded for both DOC (p-value of 0.0004) and pH (p-value of 0.0325) with respect to monochloramine: whereas chlorine only exhibited significance in regard to pH (p- value of 0.0052). Varied iopamidol expressed significance for both iopamidol (p- value of < 0.0001) and pH (p-value of 0.0014) with respect to monochloramine. In regard to chlorine, significance was only determined for pH (p-value of < 0.0001).

With respect to dichloroiodomethane acting as the response variable amongst varied

DOC, significance was determined in regard to pH for both chlorine (p-value of

0.0009) and monochloramine (p-value of 0.0012). Varied iopamidol concentration, significance was found for iopamidol for chlorine (p-value of 0.0064) and pH for monochloramine (p-value of 0.0029). While the findings of said statistical analysis were in most cases significant, considering the interaction plots and the lack of parallelism expressed, one can assume interaction between independent variables (i.e. factors). While it cannot be determined if the lack of parallelism is due to randomness or interaction, past work supports the assumption of interaction over randomness.

iv ACKNOWLEDGEMENTS

First and foremost, I like to thank my advisor, Dr. Stephen Duirk for numerous years of support and guidance. I would also like to thank Dr. Teresa Cutright for her continued inspiration and mentorship. I would like to thank my committee for their support throughout this work and their input, Dr. Stephen Duirk, Dr, Teresa Cutright

and Dr. Lan Zhang. I would like to thank my friends/peers: Edward Machek, Nana

Ackerson, Mallory Crow and Alexis Killinger. I would like to express my deepest

gratitude for my parents, Jay and Diane Crafton. I would also like to thank both of my

sisters, Dr. Sarah Crafton and Rachel Crafton-Stiver, for being impeccable role

models whom I can always look to for support and guidance. This work is dedicated

to my uncle and fellow research scientist, Dr. John Shainoff.

TABLE OF CONTENTS Page LISTS OF TABLES ...... viii

LISTS OF FIGURES ...... xiv

CHAPTER

I. INTRODUCTION ...... 1

1.1 Background ...... 1

1.2 Problem Statement ...... 6

1.3 Specific Objectives ...... 7

II. LITERATURE REVIEW ...... 9

2.1 Iodinated X-Ray Contrast Media ...... 9

2.2 Occurrence of ICM in Water and Wastewater ...... 12

2.3 Chemical Oxidation and Disinfection By-Products (DBPs) ...... 14

2.3.1 Chemical Oxidation ...... 14

2.3.2 Disinfection By-Product Formation ...... 15

2.3.3 Aqueous Chlorine Reactivity and DBP Formation...... 17

2.3.4 Chloraime ...... 19

2.4. ICM Transformation and Iodo-DBPs ...... 20

2.4.1 ICM Transformation ...... 21

2.4.2 Microbial Transformation ...... 21

2.5 Toxicity ...... 22

vi III. MATERIALS AND METHODS ...... 27

3.1 Chemicals and Reagents ...... 27

3.2 Source Water Characterization ...... 28

3.3 Experimental Methods ...... 33

3.3.1 DBP Formation Experiments with Cleveland Source Water ...... 34

3.4 Analytical Procedures ...... 35

3.4.1 Disinfection By-product ...... 35

3.6 Analyses of DBPs ...... 38

IV. RESULTS AND DISCUSSION ...... 66

4.1 Introduction ...... 66

4.2 Predominant Observed Trends ...... 67

4.3 Statistical Analysis ...... 118

V. CONCLUSIONS AND RECOMMENDATIONS ...... 125

5.1 Introduction ...... 125

5.2 Conclusions ...... 126

5.3 Recommendations ...... 129

REFERENCES ...... 131

APPENDIX ...... 146

vi i LIST OF TABLES

Table Page

3.1: Source water characteristics from Cleveland water ...... 29

3.2: Florescence EEM regions proposed by Chen et al. (2003) ...... 31

3.3: Florescence regions for Cleveland source waters, 1 mg/L C ...... 31

3.4: Oven temperature programming for THMs and HANs analysis on GC/μECD ... 38

3.5: Oven temperature programming for HAAs analysis on GC/μECD ...... 39

3.6: Limit of quantification (LOQ) for the detection of DBPs ...... 65

4.1: p-values for two-way ANOVA (two factors) where α = 0.05 (95% significance) with respect each oxidant where the response variable (Y) was chloroform. ... 121

4.2: p-values for two-way ANOVA (two factors) where α = 0.05 (95% significance) with respect each oxidant where the response variable (Y) was dichloroiodomethane...... 122

A1: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 146

A2: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 147

A3: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L...... 148

viii A4: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L...... 148

A5: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L ...... 149

A6: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L ...... 149

A7: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L ...... 150

A8: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L...... 150

A9: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L...... 151

A10: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L ...... 151

A11: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L...... 152

A12: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L...... 152

ix A13: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 153

A14: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 153

A15: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L...... 154

A16: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L...... 155

A17: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 1.0 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 156

A18: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 1.0 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 156

A19: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 1.0 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L...... 157

A20: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 1.0 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L...... 157

A21: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 158

x A22: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L ...... 158

A23: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L- L ...... 159

A24: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L- L ...... 159

A25: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L ...... 160

A26: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L ...... 160

A27: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L- L ...... 161

A28: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L- L ...... 161

A29: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L ...... 162

A30: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L ...... 162

A31: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L- L ...... 163

xi A32: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L- L ...... 163

A33: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L) ...... 164

A34: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L) ...... 165

A35: Two-way ANOVA where the response variable was dichloroiodomethane (CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L) ...... 166

A36: Two-way ANOVA where the response variable was dichloroiodomethane (CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L) ...... 167

A37: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM) ...... 168

A38: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM) ...... 169

A39: Two-way ANOVA where the response variable was dichloroiodomethane (CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM) ...... 170 xii A40: Two-way ANOVA where the response variable was dichloroiodomethane (CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM) ...... 171

xiii LIST OF FIGURES

Figure Page

2.1: The chemical structures of ICM of common use within the medical field (Duirk et al. 2011) ...... 11

2.2: Iodo-DBP formation pathway due to iopamidol reacting with chlorinated oxidants in the presence of NOM (Duirk et al., 2011) ...... 24

3.1: Fluorescence excitation-emission spectrum (EEM) of Cleveland source water. [DOC] = 2.51 mg/L, SUVA254 = 1.17 L/mg.m ...... 32

3.2: Calibration curve for CHCl3using chloroform. [CHCl3] = 0 – 1000 nM ...... 40

3.3: Calibration curve for CHBr2Cl using dibromochloromethane. [CHBr2Cl] = 0 – 300 nM ...... 41

3.4: Calibration curve for CHBrI2 using bromodiiodomethane. [CHBrI2] = 0 – 125 nM ...... 42

3.5: Calibration curve for CHClI2 using chlorodiiodomethane. [CHClI2] = 0 – 250 nM ...... 43

3.6: Calibration curve for CHBr2I using dibromoiodomethane. [CHBr2I] = 0 – 250 nM ...... 44

3.7: Calibration curve for CHBrClI using bromochloroiodomethane. [CHBrClI] = 0 – 250 nM ...... 45

3.8: Calibration curve for CHBr3 using bromoform. [CHBr3] = 0 – 500 nM ...... 46

3.9: Calibration curve for CHCl2I using dichloroiodomethane. [CHCl2I] = 0 – 500 nM ...... 47

3.10: Calibration curve for CHCl2Br using bromodichloromethane. [CHCl2Br] = 0 – 400 nM ...... 48

3.11: Calibration curve for CHI3 using iodoform. [CHI3] = 0 – 50 nM ...... 49 xiv 3.12: Calibration curve for CAN using chloroacetonitrile. [CAN] = 0 – 500 nM ...... 50

3.13: Calibration curve for DCAN using dichloroacetonitrile. [DCAN] = 0 – 500 nM ...... 51

3.14: Calibration curve for TCAN using trichloroacetonitrile. [TCAN] = 0 – 125 nM ...... 52

3.15: Calibration curve for BAN using bromoacetonitrile. [BAN] = 0 – 125 nM ...... 53

3.16: Calibration curve for DBAN using dibromoacetonitrile. [DBAN] = 0 – 250 nM ...... 54

3.17: Calibration curve for BCAN using bromochloroacetonitrile. [BCAN]=0–250 nM ...... 55

3.18: Calibration curve for IAN using iodoacetonitrile. [IAN] = 0 – 31 nM ...... 56

3.19: Calibration curve for CAA using chloroacetic acid. [CAA] = 0 – 250 nM ...... 57

3.20: Calibration curve for DCAA using dichloroacetic acid. [DCAA] = 0 – 500 nM ...... 58

3.21: Calibration curve for TCAA using trichloroacetic acid. [TCAA] = 0 – 250 nM59

3.22: Calibration curve for BCAA using bromochloroacetic acid. [BCAA]=0–250 nM ...... 60

3.23: Calibration curve for BDCAA using bromodichloroacetic acid. [BDCAA] = 0 – 250 nM ...... 61

3.24: Calibration curve for BAA using bromoacetic acid. [BAA] = 0 – 1000 nM ..... 62

3.25: Calibration curve for DBAA using dibromoacetic acid. [DBAA] = 0 – 500 nM ...... 63

3.26: Calibration curve for IAA using iodoacetic acid. [IAA] = 0 – 125 nM ...... 64

4.1: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 68

xv 4.2: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 1.26 mg/L-L...... 69

4.3: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 0.63 mg/L-L...... 70

4.4: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 71

4.5: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 72

4.6: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 1.26 mg/L-L...... 73

4.7: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 0.63 mg/L-L...... 74

4.8: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 6.5; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 75

4.9: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 7.5; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 77

4.10: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 8.5; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 79

4.11: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH)

xvi

at pH 9.0; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 81

4.12: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 6.5; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 82

4.13: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 7.5; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 84

4.14: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 8.5; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 86

4.15: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 9.0; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 88

4.16: Observed CHCl2I formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 90

4.17: Observed trichloroacetic acid formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 91

4.18: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 92

4.19: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 1.26 mg/L-L...... 93

4.20: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

xvii

(Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 0.63 mg/L-L...... 94

4.21: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 95

4.22: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 1.0 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 96

4.23: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L- L...... 97

4.24: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 1.26 mg/L- L...... 98

4.25: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 0.63 mg/L- L...... 99

4.26: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 6.5; [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 100

4.27: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 7.5; [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 101

4.28: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 8.5; [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 102

4.29: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water

xviii (Cleveland, OH) at pH 9.0; [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C...... 103

4.30: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 6.5; [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 104

4.31: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 7.5; [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 105

4.32: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 8.5; [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 106

4.33: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 9.0; [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L...... 107

4.34: Speciation of hypochlorous acid (HOCl/OCl-) and iopamidol (depicted as IP/IP- ), alphas (αo and α1) derived from pH and respective pKa...... 113

4.35: Speciation of hypochlorous acid (HOCl/OCl-) and iopamidol (depicted as IP/IP- - ), alphas (αo and α1) derived from pH and respective pKa, [Cl2 ]T = 100 μM, [iopamidol] = 5 μM...... 114

A1: Interaction plot where the response variable was chloroform (CHCl3) formation at 72 hours, factors (x-axis) were pH and DOC (2.51, 1.26 and 0.63 mg/L-L) where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 172

A2: Interaction plot where response variable (y-axis) was chloroform (CHCl3) formation at 72 hours, factors (x-axis) was pH and DOC (2.51, 1.26 and 0.63 mg/L-L) where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 173

A3: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factor (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and DOC (2.51, 1.26 and 0.63 mg/L-L) where the reactor vessel contained

xix

iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 174

A4: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and DOC (2.51, 1.26 and 0.63 mg/L-L) where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 175

A5: Interaction plot where response variable (y-axis) was chloroform (CHCl3) formation at 72 hours, factor (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 176

A6: Interaction plot where response variable (y-axis) was chloroform (CHCl3) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 177

A7: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 178

A8: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C ...... 179

xx CHAPTER I

INTRODUCTION

1.1 Background

Chlorination of water was originally employed to inactivate pathogenic microorganism as means of public health protection, which resulted in massive reduction of common waterborne gastrointestinal diseases (Akin et al., 1982). Additionally, disinfection via chemical oxidation is employed oxidize organic micropollutants (anthropogenic and natural) (Nriagu and Simmons, 1994). Ideally, chemical oxidation of micropollutants results in their respective terminal end products (CO2 and H2O) or an intermediate product that possesses a lesser toxicological effect and is more readily biodegradable

(Nriagu and Simmons, 1994). Chemical oxidation can also be utilized to induce the precipitation of insoluble inorganic metals from the water column (Crittenden et al.,

2012). Nevertheless, chemical oxidants retain the ability to facilitate the formation of potentially harmful by-products (i.e., disinfection by-products), some of which may be considerably more toxic than respective parent compound (Krasner et al., 2006; Simmons et al., 2002; Bichsel and von Gunten, 2000). Initially, the formation of disinfection by- products (DBPs) went undetected until the early 1970s when chloroform was first reported in chlorinated drinking water (Bryant et al., 1992; Bellar et al., 1974; Rook

1 1974). Since 1974, more than 600 DBPs have been identified within drinking water

(Richardson et al., 2007). The majority being halogens (i.e., chlorine, bromine, and iodine) incorporated into organic carbon resulting in the formation of unidentified DBPs

(Richardson et al., 2002; Weinberg, 1999). Previously, DBPs were defined as the reaction of natural organic matter (NOM) with oxidants, which are employed during the disinfection processes. Since, DBPs have been redefined as the reactions of said oxidant with anthropogenic contaminants such as pharmaceuticals, personal care products, and pesticides (Duirk et al. 2011). The formation of DBPs is greatly dependant upon aqueous conditions such as; concentration and type of disinfectant, pH, temperature, source water characteristics, the presence of inorganic precursor, as well as the treatment process

(Krasner, 2009; Richardson et al., 2007; Ueno et al., 1996). Source water characteristics heavily influence DBP formation and speciation. The United States Environmental

Protection Agency (US EPA) currently regulates two groups of organic DBPs, trihalomethanes (THMs) and haloacetic acids (HAAs), totalling 9 compounds (US EPA,

2006). Of the 9 regulated DBPs, 4 are THMs (chloroform (CHCl3), bromodichloromethane (CHBrCl2), dibromochloromethane (CHBr2Cl) and bromoform

(CHBr3)) and 5 are HAAs (monochloroacetic acid (CAA), dichloroacetic acid (DCAA), trichloroacetic acid (TCAA), bromoacetic acid (BAA) and dibromoacetic acid (DBAA))

(US EPA, 2006). In addition to the aforementioned regulated compounds, the US EPA also regulates bromate and chlorite (US EPA, 2006). Regulated groups of DBPs have been assigned a maximum contamination level (MCL) (THMs 80 μg/L, HAAs 60 μg/L, bromate 10 μg/L and chlorite 1.0 mg/L) (Weinberg et al., 2002).

As previously stated, in addition to NOM, anthropogenic micropollutants (i.e. ICM) react

with residual chemical oxidants thus resulting in stable transformation products, which

may potentially be more toxic than the parent compound. Transformation of said

anthropogenic micropollutants may contribute to an observed elevation in regulated

DBPs (THMs and/or HAAs) or potentially form unknown, and therefore unregulated,

DBPs that possess unknown toxicity. Each oxidant and respective precursor possesses

vastly different formation, which produce an array of DBPs. Previously, iodinated-DBPs

(iodo-DBPs) were thought to be the result of oxidation of iodide present in source water

to hypoiodous acid (HOI) by hypochlorous acid (HOCl), which rapidly incorporates into

organic matter present (Bischell and von Gunten 1999, Duirk et al., 2011; Hansen et al.,

2011; Gilfedder et al., 2009).

In the presence of a chemical oxidant (i.e. chlorine, chloramine or ozone), iodide is

promptly oxidized to hypoiodous acid (HOI) (Garland et al., 1980; Nagy et al., 1988;

- Kumar et al., 1986). Hypoiodous acid (HOI) is further oxidized to iodate (IO3 ), additionally, the hypoiodous acid (HOI) can react with present NOM consequently inducing the formation of iodo-DBPs (Bichsel and von Gunten, 1999b; Richardson et al.,

2008; Krasner et al., 2006). Conversely, source waters were documented to possess

considerably low concentrations of natural iodide (I-), or concentrations below detection limits (Richardson et al., 2008). Corresponding analysis determined iodide concentrations to be insufficient in correlation to observed iodo-DBP formation thus suggesting an alternative source of iodide and contradicting the previous perception of

natural iodide as the primary source of iodo-DBP formation (Bichsel and von Gunten,

3 2000 & 1999a; Richardson et al., 2008). Alternative sources of iodide were investigated, which revealed iodinated x-ray contrast media (ICM) as a source of iodide (Duirk et al.,

2011).

Iodinated x-ray contrast media (ICM) are derivatives of 2,4,6-triiodobenzoic acid. ICMs are an intravenous, and at times intra-arterial administrated radio-contrast that which are most commonly used within the radiology sector of modern western medicine. ICMs are used to enhance visibility of vascular structures and organs during radiographic examinations to achieve diagnostic x-ray imaging. Water-soluble ICMs are most abundantly used in computed topography (CT) scans, utilization also occurs for gastrointestinal exam when the traditionally used barium sulfate can potentially inhibit exam or compromise the results of said exam. ICMs are unmetabolized and renally excreted for the body within 24 hours of intravenous or intraarterial administration due to their inert properties (Perez et al., 2006; Steger-Hartmann et al., 2000). ICM consumption is estimated to be 3.5 x 106 kg/year, of which solely Germany contributes 5 x 105 kg/year (Steger-Hartmann et al., 1999). During 1999, United States consumption was estimated to be 1.33 x 106 kg (Steger-Hartmann et al., 1999). Since ICMs are resistant to conventional water and wastewater treatment, their presence within the water infrastructure (e.g., source water, drinking water plant/distribution, wastewater collection/plant, etc) and abundance of which are of imperative concern. Documented low removal efficiency during conventional wastewater treatment processes (10%) has lead to ICM detection, at relatively high concentrations (>1 μg/L), within natural water sources (Drews et al., 2001; Ternes and Hirsch, 2000; Hirsch et al., 2000; Putschew et

4 at., 2000). ICMs have also been identified as the main contributor to total adsorbable organic halogens, more specifically adsorbable organo-iodine (AOI) observed within clinical wastewater (Gartiser et al. 1996; Seitz et al. 2006). It is speculated that there are approximately 69 pharmaceutical agents present within wastewater produced by medical facilities, of which 52 active pharmaceutical agents were detected (McArdell et al. 2010).

Of the active agents detected, iopamidol was determined to be present at the highest concentrations (mg/L range) thus confirming the frequent usage and high dosage administered at medical imaging facilities (McArdell et al. 2010).

Chlorine is a very commonly used oxidant that can be employed in multiple forms; solid

(Ca(OCl)2), liquid (NaOCl) and gaseous (Cl2). Aqueous chlorine efficiently and inexpensively inactivates pathogenic microorganisms found in drinking water.

Simultaneously, aqueous chlorine can react with several anthropogenic organic micropollutants thus resulting in toxic, yet stable, transformation products (Duirk and

Collette, 2006; Gallard and Von Gunten, 2002; Arnold, et al, 2008). Four common pathways can describe chlorine’s reaction with water constituents: addition, substitution, oxidation and light decomposition (Gang et al., 2003; Johnson and Jensen, 1986).

Speciation of chlorine is dependant upon the pH of the aqueous solution. Given that drinking water criteria recommend a pH between 6.5-8.5 and the pKa of chlorine; the predominant chlorine species observed are hypochlorous acid (HOCl) and hypochlorite ion (OCl-) (equilibrium dictated below) (Deborde and Von Gunten, 2008).

An alternative oxidant in lieu of chlorine used within water treatment for disinfection is chloramine, which includes mono-, di- and tri-chloramine. Chloramine is produced via a 5

substitution reaction consisting of free chlorine and ammonia. It has been determine that

monochloramine (NH2Cl) is the predominate species present within drinking water

conditions, that being a pH range of 6.5–8.5 (Vikesland et al., 1998). While monochloramine possesses equivalent oxidizing aptitude as chlorine, it is ultimately a weaker disinfectant (Wolfe et al., 1984). Utilization of monochloramine is not without difficulty considering the lack of stability at neutral pH despite the absence of organic/inorganic compounds; in addition to the largely pH dependant, auto- decomposition that is observed (Jafvert and Valentine, 1992).

1.2 Problem Statement

In the presence of a chemical oxidant (chlorine or chloramine), iodinated-DBPs were thought to be the result of the oxidation of iodide present in source water to HOI by

HOCl, rapidly followed by supplemental incorporation with organic matter present

(Bischell and von Gunten 1999, Duirk et al., 2011; Gilfedder et al., 2009; Hansen et al.,

2011; Krasner et al., 2006; Richardson et al., 2008). Formed HOI may also be further

- oxidized to IO3 in the presence of chlorine in lieu of reincorporation (Bichsel and von

Gunten, 1999b). Natural iodide concentrations proved to be insufficient in correlation to

iodo-DBP formation suggesting an alternative source and contradicting the former

perception of natural iodide as the principal source of iodide in the formation of iodo-

DBPs (Bichsel and von Gunten, 2000 & 1999a; Richardson et al., 2008). Iodinated x-ray

contrast media (ICM) proved to be a valid source of iodide (Duirk et al., 2011).

Furthermore, documented low removal efficiency during conventional wastewater 6

treatment processes (≈10%) has lead to ICM detection, at relatively high concentrations

(>1 μg/L), within natural water sources (Drews et al., 2001; Ternes and Hirsch, 2000;

Hirsch et al., 2000; Putschew et at., 2000). When investigated, 52 active pharmaceutical

agents were detected, iopamidol was determined to be present at the highest

concentrations (mg/L range) (McArdell et al. 2010). Duirk et al. (2011) demonstrated

the release of organically bound iodide from the aromatic ring of iopamidol in the

presence of chlorinated chemical oxidants. Release of said organically bound iodide

gave way to the incorporation with NOM resulting in the formation of iodo-DBPs; in the

- absence of NOM, IO3 and only trace levels of iodo-DBPs were formed (Duirk et al.,

2011). Additionally, hypochlorite ion (OCl-) is believed to be the primary reactant in the

degradation of iopamidol with both chlorine and monochloramine (Duirk et al., 2011).

1.3 Specific Objectives

The following objective were considered throughout the completion of this work:

1. Investigate the formation of iodinated disinfection by-products (iodo-DBPs) where

ICM, Iopamidol, acted as the source of iodine, with respect to pH (6.5, 7.5, 8.5 and

9.0) in the presence of NOM and chlorinated oxidant. Additionally, the effect of

varied NOM as well as varied iopamidol concentrations were explored. Experiments

were performed in source water obtained from the Cleveland water treatment plant

(Cleveland, OH). The data was obtained from experiments in which iopamidol was

introduced into a batch reactor containing source water (Cleveland, OH) at varying

concentrations (5.0, 2.5, 1.0 and 0.0 μM) over the pH range of 6.5 – 9.0 (6.5, 7.5, 8.5

and 9.0). Additionally, iopamidol concentration was held constant (5.0 μM) while

TOC was varied via dilution with dionized water (2.51, 1.26 and 0.63 mg/L-L) over

the pH range of 6.5 – 9.0 (6.5, 7.5, 8.5 and 9.0). Source water experiments were

performed in the absence of iopamidol, with each chlorinated oxidant to act as a

control over the pH range of 6.5 – 9.0 (6.5, 7.5, 8.5 and 9.0). The chlorinated

oxidants that were evaluated were aqueous chlorine and monochloramine.

2. A statistical analysis was performed on derived data set (two-way ANOVA and

interaction plots) with the objective to obtain significance of respective factors

(iopamidol, pH and DOC) were the response variables were chloroform (CHCl3) and

dichloroiodomethane (CHCl2I). Two-way analysis of variance (two way-ANOVA)

tables were generated for two response variables, chloroform and

dichloroiodomethane. With respect to each oxidant, iopamidol and pH were

evaluated while NOM level remained constant and NOM and pH were evaluated

while iopamidol remained constant. Corresponding interaction plots were generated

for each respective two-way ANOVA table.

CHAPTER II

LITERATURE REVIEW

2.1 Iodinated X-Ray Contrast Media

Derivatives of 2,4,6-triiodobenzoic acid, ICMs are an intravenous, and at times intraarterial administrated radio-contrast that are most commonly used within the radiology sector of modern western medicine. ICMs are used to enhance visibility of vascular structures and organs during radiographic examinations to achieve diagnostic x-ray

imaging. ICM are also used within oncology to achieve improved visibility of certain

neoplastic growths. The medical field utilizes both oil-based ICM and water soluble

ICM. Oil-based ICM are used during sialography and uterosalpingography exam. A

sialographiy is a radiographic examine where the salivary glands are evaluated for normal

biological fucntion. Uterosalpingography is also a radiological procedure in which the

uterine cavity as well as the shape and patency of fallopian tubes are evaluated for

abnormalities. Water-soluble ICMs are most abundantly used in CT scans, utilization

also occurs for gastrointestinal examines when the traditionally used barium sulfate can

potentially inhibit exam or compromise the results of said exam. Post examination, the

unmetabolized ICM is renally excreted from the body within 24 hours of intravenous or

9 intraarterial administration due to their chemical and biological stability and inert properties (Perez et al., 2006; Steger-Hartmann et al., 2000). It was reported by Speck

and Hübner-Steiner (1999) that approximately 100 grams of ICM are administered for each radiographic examination. Perez and Barcelo (2007) reported a maximum of 200 grams ICM administered per diagnosis. Seitz et al. (2005) reported a required dosage of

200 grams, approximately half of which is iodine. When ICMs are administered to a patient on a nephrotoxic antibiotic (Gentamicin), extensive renal damaged is observed unlike like that observed in a patient solely on nephrotoxic antibiotic (Jensen et al.,

2013).

Globally, ICM consumption is estimated to be 3.5 x 106 kg/year, of which solely

Germany contributes 5 x 105 (Steger-Hartmann et al., 1999). During 1999, United States consumption was estimated to be 1.33 x 106 kg (Steger-Hartmann et al., 1999). These large molecules (≈ 600 – 700 Da) have similar tri-iodinated benzene moiety with side chains comprised of hydroxyl, carboxyl and amide moieties (Figure 2.1) that increase aqueous solubility (Krause and Schneider, 2002).

Figure 2.1: The chemical structures of ICM of common use within the medical field (Duirk et al. 2011)

According to the World Health Organisation (WHO) Collaborating Centre for Drug

Statistics Methodology, there are over 35 ICMs and these can be categorized as water soluble, nephrotropic, high osmolar ICM; water soluble, nephrotropic, low osmolar ICM; water soluble, hepatotropic ICM; and non-water soluble ICM. Examples of water soluble, nephrotropic, high osmolar ICM include diatrizoic acid, metrizoic acid, iodamide, iotalamic acid, ioglicic acid, acetrizoic acid, iocarmic acid, methiodal, and diodone. Metrizamide, iohexol, ioxaglic acid, iopamidol, iopromide, iotrolan, ioversol, iopentol, iodixanol, iomeprol, iobitridol and ioxilan are examples of water soluble, nephrotropic, low osmolar ICM. Iodoxamic acid, iotroxic acid, ioglycamic acid, adiopiodone, iobenzamic acid, iopanoic acid, iocetamic acid, sodium iopodate, tyropnoic acid and calcium iopodate are in the water soluble, hepatotropic ICM category. Majority of the non-water soluble ICM are ethyl ester of iodised fatty acid, iopydol, propyliodone, iofendylate (http://www.whocc.no).

2.2 Occurrence of ICM in Water and Wastewater

Since ICMs are resistant to conventional water and wastewater treatment, their presence within the water infrastructure (e.g., source water, drinking water plant/distribution, wastewater collection/plant, etc) and abundance of which are of imperative concern.

Documented low removal efficiency during conventional wastewater treatment processes

(10%) has lead to ICM detection, at relativlely high concentrations (>1 μg/L), within natural water sources (Drews et al., 2001; Ternes and Hirsch, 2000; Hirsch et al., 2000;

Putschew et at., 2000). Documented aqueous environments of impact include surface

12 water, river, creaks, groundwater and wastewater effluents (Drews et al., 2001; Ternes and Hirsch, 2000; Hirsch et al., 2000; Putschew et at., 2000). Additionally, ICMs have been detected within wastewater produced from medical imaging facilities (Ziegler et al.,

1997; Gartiser et al., 1996). ICMs have also been identified as the main contributor to total adsorbable organic halogens, more specifically adsorbable organo-iodine (AOI) observed within clinical wastewater (Gartiser et al. 1996; Seitz et al. 2005). It is speculated that approximately 69 pharmaceutical agents may be present within wastewater and are produced by medical facilities, 52 active pharmaceutical agents were detected, iopamidol was determined to be present at the highest concentrations (mg/L range) thus confirming the frequent usage and high dosage administered at medical imaging facilities (McArdell et al. 2010). Treatment processes were evaluated for the waste stream produced by medical imaging facilities. Use of a biologically activated membrane showed less than 20% removal of pharmaceuticals, which includes ICMs

(McArdell et al. 2010). Post treatment, use of powder activated carbon (PAC) proved a removal efficiency of approximately 70% of ICMs, iopamidol resulting in a preeminent residual concentration of 900 μg/L (McArdell et al. 2010). Low adsorption efficiency can be attributed to the high polarity exhibited by ICMs (Steger-Hartmann et al., 1999).

Iopamidol has been detected in aquatic environments that receive effluent flow from municipal and sewage treatment plants at a median concentration of 0.49 μg/L (Ternes and Hirsch, 2000; Putschew et al., 2001). Furthermore, iopamidol has been detected within the influent of sewage treatment plants at equivalent concentrations to the effluent

(Ternes and Hirsch, 2000). The highest concentrations of ICMs within domestic effluent

13 are observed during the weekdays (Monday through Friday), which corresponds to medical application (Drewes et al., 2001; Seitz et al., 2005). Finally, Iopamidol was detected up and downstream of a sewage treatment plant in southern Germany; observed concentrations were 470 and 510 ng/L, respectively (Seitz et al., 2005).

2.3 Chemical Oxidation and Disinfection By-Products (DBPs)

Chemical oxidation is a commonly employed method of disinfection within water treatment. Disinfection by-products are a direct result of said chemical oxidation thus awareness of contingency is imperative.

2.3.1 Chemical Oxidation

Within water treatment, disinfection via the use of chemical oxidants is employed to inactivate pathogenic microorganisms as well as oxidize organic micropollutants

(anthropogenic and natural) (Nriagu and Simmons, 1994). Chemical oxidation of micropollutants ideally results in their respective terminal end products (CO2 and H2O) or an intermediate product that possesses a lesser toxicological effect and is more readily biodegradable (Nriagu and Simmons, 1994). Chemical oxidation can also be utilized to induce the precipitation of insoluble inorganic metals from the water column (Crittenden et al., 2012). Commonly used oxidants consist of, but are not limited to aqueous chlorine, chlorine dioxide, ozone, hydrogen peroxide, potassium permanganate and chloramines (mono-, di- and tri- chloramine) (Crittenden et al., 2012; Nriagu and

Simmons, 1994). However, chemical oxidants are capable of forming potentially 14

harmful by-products (i.e., disinfection by-products), some of which can be considerably

more toxic than the parent compound (Krasner et al., 2006; Simmons et al., 2002;

Bichsel and von Gunten, 2000).

2.3.2 Disinfection By-Product Formation

Chlorination of water was originally employed to inactivate pathogenic microorganism as

means of public health protection, which resulted in massive reduction of common water-

borne gastrointestinal diseases (Akin et al., 1982). Initially, the formation of DBPs went

undetected until the early 1970s when chloroform was first reported chlorinated drinking

water (Bryant et al., 1992; Bellar et al., 1974; Rook 1974). Since, more than 600 DBPs

have been identified within drinking water (Richardson et al., 2007). However, the

majority of halogens (i.e., chlorine, bromine, and iodine) incorporated into organic

carbon thus resulting in the formation of DBPs have yet to be identified (Richardson et

al., 2002; Weinberg, 1999).

Previously, DBPs were defined as the reaction of NOM with oxidants employed during

drinking water disinfection processes. However, DBPs have been redefined as due to the

reactions with anthropogenic contaminants such as pharmaceuticals, personal care

products, and pesticides (Duirk et al. 2011). The modified definition encompasses any

transformation products formed from the reaction of organic material, not specifically

NOM, and residual disinfectant. The formation of said DBPs is greatly dependant upon aqueous conditions such as; concentration and type of disinfectant pH, temperature,

15 source water characteristics, the presence of inorganic precursor, as well as the treatment process (Krasner, 2009; Richardson et al., 2007; Ueno et al., 1996). Source water characteristics heavily influence DBP formation and speciation (i.e., SUVA254 (derived in

Chapter 3), total TOC concentration, soluble microbial products, regional assessment of fluorescence excitation - emission matrix (EEM) spectra and the concentrations of bromide or iodide). Using source water characteristics and oxidant type, DBP research has provided countless models that have the ability to not only describe degradation and formation pathways but also predict DBP formation in finished water (Amy et al., 1987;

Clark et al., 2001, Cowman et al., 1996; Duirk et al., 2005; Duirk et al., 2006; Duirk et al., 2007; von Guten et al., 2002). The US EPA currently regulates two groups of organic DBPs, trihalomethanes (THMs) and haloacetic acids (HAAs), totalling 9 compounds (US EPA, 2006). Of the 9 regulated compounds, 4 are THMs (chloroform

(CHCl3), bromodichloromethane (CHBrCl2), dibromochloromethane (CHBr2Cl) and bromoform (CHBr3)) and 5 are HAAs (monochloroacetic acid (CAA), dichloroacetic acid (DCAA), trichloroacetic acid (TCAA), bromoacetic acid (BAA) and dibromoacetic acid (DBAA)) (US EPA, 2006). In addition to the aforementioned regulated compounds, the US EPA also regulates bromate and chlorite (US EPA, 2006). Regulated groups of

DBPs have been assigned a maximum contamination level (MCL) (THMs 80 μg/L,

HAAs 60 μg/L, bromate 10 μg/L and chlorite 1.0 mg/L) (Weinberg et al., 2002).

However, the most genotoxic and cytotoxic DBPs have yet to be regulated (Richardson et al., 2008 and Plewa et al., 2002).

2.3.3 Aqueous Chlorine Reactivity and DBP Formation

As previously mentioned, chlorine is a very commonly used oxidant that can be

employed in multiple forms; solid (Ca(OCl)2), liquid (NaOCl) and gaseous (Cl2).

Aqueous chlorine efficiently and inexpensively inactivates pathogenic microorganisms

found in drinking water. Simultaneously, aqueous chlorine can react with several

anthropogenic organic micropollutants thus resulting in toxic, yet stable, transformation products (Duirk and Collette, 2006; Gallard and Von Gunten, 2002; Arnold, et al, 2008).

As mentioned earlier, four common pathways can describe chlorine’s reaction with water

constituents: addition, substitution, oxidation and light decomposition (Gang et al., 2003;

Johnson and Jensen, 1986). Addition and substitution pathways consist of the addition or

substitution of chlorine onto the molecular structure of NOM thus producing chlorinated,

organic intermediates that result in DBP formation upon further decomposition (van

Hoof, 1992).

Speciation of chlorine is dependant upon the pH of the solution. Given that drinking

water criteria recommend a pH between 6.5-8.5 and the pKa of chlorine; the predominant

chlorine species observed are HOCl and OCl- (equation 2.1) (Deborde and Von Gunten,

2008). Quantification of the total concentration of said predominant chlorine species represents what is known as free chlorine. Within drinking water conditions, HOCl is dominant reactive species due to the relative abundance with respect to additional chlorine species that may be present (Morris 1978). In the presence of other halides,

HOCl remains dominant reactive species.

17 + = 7.54 ( 2.1) − + 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 ↔ 𝑂𝑂𝑂𝑂𝑂𝑂 𝐻𝐻 𝑝𝑝𝐾𝐾𝑎𝑎 The most frequent oxidation reaction that occurs with HOCl under given drinking water

conditions is dictated below, where “X” represents an organic or inorganic compound in

equation 2.2 (Deborde and von Gunten, 2008). The reaction was assumed to be an

elemental stoichiometric oxidation reaction and assumed to be an overall second – order

reaction, first – order with respect to chlorine ([HOCl]T) and organic or inorganic

compound ([X]T) (Deborde and von Gunten, 2008). The representative differential

equation describing said reaction is dictated below, equation 2.3; variables within

equation 2.3 are also defined below. Furthermore, the apparent second order rate constant

was determined to be dependant upon pH (Deborde and von Gunten, 2008).

+ (2.2)

𝐻𝐻𝑂𝑂𝑂𝑂𝑂𝑂 𝑋𝑋 → 𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃𝑃 [ ] = [ ] [ ] (2.3) 𝑑𝑑 𝑋𝑋 𝑇𝑇 𝑘𝑘𝑎𝑎𝑎𝑎𝑎𝑎 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 𝑇𝑇 𝑋𝑋 𝑇𝑇 𝑑𝑑𝑑𝑑 =

𝑘𝑘𝑎𝑎𝑎𝑎𝑎𝑎 𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎𝑎 𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠𝑠 𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜𝑜 𝑟𝑟𝑟𝑟𝑟𝑟𝑟𝑟 𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐𝑐 [ ] = [ ] + [ ] − 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 𝑇𝑇 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 𝑂𝑂𝑂𝑂𝑙𝑙 [ ] = [ ] + [ ] − 𝑋𝑋 𝑇𝑇 𝐻𝐻𝐻𝐻 𝑋𝑋 Copious research has verified suggestive variability in reactivity with respect to HOCl

and OCl- for a given compound (Armesto et al., 1994a; Rebenne et al., 1996; Abia et al.,

1998; Gallard and von Gunten, 2002; Gallard et al., 2004; Deborde et al., 2004; Dodd et al., 2005). As previously mentioned, “X” within equation 2.2 can represent either an

organic or inorganic compound. In regard to inorganic compounds (i.e. cyanide and

nitrite), HOCl oxidizes through an electrophilic attack. Deborde and von Gunten (2008)

determined foreseen susceptibility of chlorine reactivity can be observed with respect to

nucleophilic properties. HOCl is generally the dominant species reacting with organic

compounds by oxidation, addition and electrophilic substitution reaction pathways. Said reaction pathways result in the formation of more oxidized or chlorinated compound due to HOCl’s ability to provoke changes within the molecular structure of the parent compound (Dore, 1989). Aromatic compounds and bound moieties undergo an electrophilic substitution reaction in the presence of chlorine where the initial attack predominantly transpires in ortho- or para- position to substituent (Deborde and von

Gunten, 2008; Roberts and Caserio, 1968). The substituent is of imperative influence on the respective aromatic ring throughout the electrophilic substitution reaction. Deborde and von Gunten, (2008) described that the properties of the electron donor within said substituent increase of the charge density of aromatic ring thus giving way to faster substitution reaction.

2.3.4 Chloraime

An alternative oxidant in lieu of chlorine used within water treatment for disinfection is chloramine, which includes mono-, di- and tri-chloramine. Chloramine is produced via a substitution reaction consisting of free chlorine and ammonia. It has been determine that monochloramine (NH2Cl) is the predominate species present at drinking water

conditions, that being a pH range of 6.5 – 8.5 (Vikesland et al., 1998). While 19

monochloramine possesses equivalent oxidizing aptitude as chlorine, it is ultimately a

weaker disinfectant (Wolfe et al., 1984). Utilization of monochloramine is not without

difficulty considering the lack of stability at neutral pH in the absence of

organic/inorganic compounds due to chloramine disproportionation (Jafvert and

Valentine, 1992). That being the oxidation of ammonia coupled with the reduction of active chlorine, auto-decomposition rates increase as the ratio of chlorine to ammonia is

increased (Jafvert and Valentine, 1992; Vikesland et al., 2000). Vikesland and Valentine

(2000) determined that within solution, monochloramine reacted with aqueous ferrous

iron via an autocatalytic reaction. This is supported by iron oxide product of said aqueous

phase reaction increased the overall reaction kinetics giving way to a highly reactive

ferrous iron surface complex (Vikesland and Valentine 2000). Monochloramine has also

been determined to react with organic substance found within water known as

dimethylamine (NMA) thus producing N-nitrosodimethylamine (NDMA) (Mitch and

Sedlak, 2002; Choi and Valentine, 2001). This is accomplished through the oxidation

NMA to unsymmetrical intermediate, dimethylhydrazine resulting in further oxidation to final product of NDMA (Mitch and Sedlak, 2002; Choi and Valentine, 2001).

2.4. ICM Transformation and Iodo-DBPs

Transformation of ICMs occurs in a variety of ways, each possessing unique and

respective pathways given present conditions, which are subjective. Iodo-DBP formation, consists of but is not limited to the chemical oxidation of ICMs.

2.4.1 ICM Transformation

In regard to the treatment of pharmaceuticals, partitioning (adsorption via activated

carbon or membrane separation) and transformation (oxidation via aerobic/anaerobic

microbial degradation or chemical oxidation) are utilized (Adams, 2009; Grassi et al.,

2012). While microbial transformation will be discussed briefly, this review will focus

on chemical oxidation of ICMs via chlorinated oxidants.

2.4.2 Microbial Transformation

ICM are known to be highly water-soluble and occur in drinking water sources; however, some investigations have shown commonly used ICM have been transformed during biological wastewater treatment. Schulz et al. (2008) investigated microbial transformation of iopromide and were able to identify 12 transformation products (within water/soil matrixes) via high performance liquid chromatography coupled with ultraviolet

(HPLC-UV) and liquid chromatography coupled with tandem mass spectrometry (LC-

MS). German based research team, Kormos et al (2010), also evaluated aerobic

transformation of ICMs, pathways and products, within batch reactors consisting of

soil/water and river sediment/water, which the water was groundwater obtained from a

deep well. The two soil types were; loamy sand soil and upper ploughed agricultural soil

with 2.3% and 0.9% organic matter respectively, and both soils have been subjected to

irrigation with secondary treated wastewater effluent and sludge for approximately 50

years. Similar to Schulz et al. 2008, HPLC-UV and LC-MS were utilized to identify

potential microbial transformation products (TPs). Kormos et al (2010) determined that 21 diatrizoate was not susceptible to microbial transform, whereas the nonionic ICMs,

iohexol, iomeprol and iopamidol, did undergo microbial transformation producing 11, 15

and 8 TPs respectively at neutral pH (≈ 7). Seven of the TPs observed by Kormos et al.

(2010) were not previously identified within pear-reviewed literature thus requiring mass

fragmentation experiments in order to elucidate the chemical structure of the unknown

TPs. Chemical oxidation of anthropogenic micropollutants (i.e. pharmicuticals - ICM),

in addition to NOM, via residual chemical oxidants results in stable transformation

products that can potentially be more toxic than the parent compound. These

transformation products may contribute to elevated levels of regulated DBPs (THMs

and/or HAAs) or potentially form unknown, and therefore unregulated, DBPs that

possess unknown toxicity. Each oxidant and precursor yields different DBPs, of various

magnitudes. As previously discussed, the modifications to the formal definition of DBPs

were made to account to emerging environmental pollutants; such as ICM and their role

in the formation of iodinated DBPs (iodo-DBPs). Formerly, iodo-DBPs were thought to

be a result of iodide present in the source water being oxidized to HOI by HOCl, which

rapidly incorporates into organic matter present (Bischell and von Gunten 1999, Duirk et

al., 2011; Hansen et al., 2011; Gilfedder et al., 2009). Of the aforementioned species,

iodide (I-) naturally exists in seawater (global concentration approximately 30 μg/L) and can potentially exist in rainwater and freshwater sources (Gilfedder et al., 2009, 2008;

Schwehr and Santschi, 2003; Yokota et al., 2004). Moran et al. (2002) detected iodine (I2)

within major United States, Canadian and European rivers ranging from 0.5 to 212 μg/L.

In the presence of a chemical oxidant, chlorine, chloramine or ozone, iodide is promptly

oxidized to HOI (Garland et al., 1980; Nagy et al., 1988; Kumar et al., 1986). HOI is

- further oxidized IO3 in the presence of chlorine or ozone (Bichsel and von Gunten,

1999b). Regardless, the HOI formed from I- can react with NOM thus inducing the formation of iodo-DBPs (Richardson et al., 2008; Krasner et al., 2006). Conversely,

source waters were documented to possess considerably low concentrations of natural I-,

or concentrations below detection limits (Richardson et al., 2008). Corresponding

analysis determined said iodide concentrations to be insufficient in correlation to formed

iodo-DBPs thus suggesting an alternative source of iodide (Richardson et al., 2008). Thus

contradicting the previous perception that natural iodide, within source water, being the

primary source of iodo-DBP formation (Bichsel and von Gunten, 2000; 1999a).

Alternative sources of iodide and their respective contribution to iodo-DBP formation

were investigated, which revealed ICMs as an organic source of iodide (Duirk et al.,

2011). Duirk et al. (2011) demonstrated the release of organically bound iodide from the

ring of an aromatic compound in the presence of chlorinated chemical oxidants and

subsequently incorporating into the NOM structure resulting in the formation of iodo-

- DBPs. In the absence of NOM, IO3 and only trace levels of iodo-DBPs were formed

(Duirk et al., 2011). It is hypothesized that the chlorinated chemical oxidants cleaves

moieties of ICM thus changing the polarity and stimulating the release of free iodide,

which can be utilized in the formation of iodo-DBPs (Duirk et al., 2011). Additionally,

hypochlorite ion is believed to be the primary reactant in the degradation of iopamidol

with both chlorine and monochloramine (Duirk et al., 2011). This is supported by the observed pH dependency and limited availability hypochlorite ion during

23 monochloramine auto-decomposition of resulting in lower iodo-DBP concentrations compared to aqueous chlorine (Duirk et al., 2011). In correlation with the previously discussed occurrence study by Richardson et al., (2008); Duirk et al., (2011) determined iopamidol to be a precursor to iodo-DBP formation in the presence of both chlorine and monochloramine. In the presence of chlorine, iopamidol produced up to 212 nM of dichloroiodomethane and 3.0 nM of iodoacetic acid (Duirk et al., 2011). The proposed reaction pathway from Duirk et al. (2011) is depicted in Figure 2.2.

Figure 2.2: Iodo-DBP formation pathway due to iopamidol reacting with chlorinated oxidants in the presence of NOM (Duirk et al., 2011)

2.5 Toxicity

As previously mentioned, the most common and regulated groups of DBPs are THMs and HAAs. It has been shown that regulated DBPs are not only carcinogenic but genotoxic and cytotoxic as well (Richardson et al., 2007). Epidemiologic studies have connected a preeminent elevation in the risk of cancer to the ingestion of chlorinated water; forms of cancer consist of the pancreas, stomach, kidney, rectum, Hodgkin’s and

non Hodgkin’s lymphoma (Bull et al., 1995; Koivusalo et al., 1994; Morris et al., 1992).

In addition to cancer, spontaneous abortions and birth defects have also been linked to

DBP exposure (Nieuwenhuijsen et al. 2000; Waller et al. 2001). Further investigation of

concentrated extracts from drinking water samples have shown to be toxic in several in

vivo and in vitro bioassays (Wilcox and Williamson, 1986). Within strains of Salmonella,

among the glutathione S-transferase theta (GSTT1-1) enzyme, brominated species of the

regulated THMs (bromodichloromethane, dibromochloromethane and bromoform)

stimulated genotoxicity (Kogevinas, et al., 2010; Kargalioglu, et al., 2002). Richardson et al. (2007) went on to determine the dissipation of genotoxicity of said brominated species in the absence of the GSTT1 -1 enzyme. Plewa et al. (2002) conducted a study on mammalian cell cytotoxicity and genotoxicity of brominated and chlorinated HAAs utilizing Chinese Hamster Ovarian (CHO) cells; bromoacetic acid was determined to be the most genotoxic and cytotoxic. Furthermore, it was determined that brominated HAAs were found to be more cytotoxic and genotoxic than their chlorinated counterparts (Plew et al. 2002); while, dichloroacetic acid and trichloroacetic acid were determined to be mutagenic in mouse lymphoma cells (Harrington-Brook et al., 1998). Across the United

States of America and Canada, 23 cities were chosen to partake in a study of chlorinated and chloraminated drinking waters to measure five iodo-acids and two THMs, resulting

in the determination that iodinated DBPs are highly genotoxic and cytotoxic (Richardson

et al., 2008). Of the investigated iodinated DBPs, iodoacetic acid was determined to be

the most genotoxic within mammalian cells (Richardson et al., 2008). In addition, it was

determined that iodo-THMs to be less cytotoxic than iodo-acids with the exception of

25 iodoform. Plewa et al. (2004) determined iodoacetic acid to be highly cytotoxic and more genotoxic in mammalian cells than bromoacetic acid. Duirk et al. (2011) confirmed previously determined cytotoxicity and genotoxicity of iodo-DBPs in mammalian cells upon dosing chlorinated/chloraminated source water (Athen-Clark County) with iopamidol at a pH of 7.5. Duirk et al. (2011) also provided cytotoxicity ranking, in descending order, for chlorinated, iopamidol spiked source waters; iodoacetic acid (IAA)

> chlorodiiodomethane >dichloroiodomethane > iodoform > bromochloroiodomethane.

The same ranking was provided for chloraminated waters; IAA > chlorodiiodomethane > dichloroiodomethane. Duirk et al. (2011) additionally noted that iodo-DBPs induce the highest genotoxicity.

CHAPTER III

MATERIALS AND METHODS

3.1 Chemicals and Reagents

Iopamidol was purchased from U.S. Pharmacopeia (Rockville, MD, USA). Commercial

(10-15%) sodium hypochlorite (NaOCl) that contained equimolar amounts of OCl- and

Cl- was purchased from Sigma Aldrich (St. Louis, MO, USA). Standards for the DBP analysis were from the following sources: iodoacetic acid and iodoform from Sigma

Aldrich (St. Louis, MO, USA), haloacetic acid mix from Restek (Bellefonte, PA,

USA)(in which methyl tert-butyl ether (MtBE) acted as the solvent for discrete concentrations of monochloro-, monobromo-, dichloro-, trichloro-, bromochloro-, dibromo-, bromodichloro-, chlorodibromo-, and tribromoacetic acid), trihalomethane mix was purchased through Chem Service (West Chester, PA, USA) (comprised of discrete concentrations of chloroform, bromoform, bromodichloromethane, and dibromochloromethane), iodo-trihalomethanes were individually purchased from CanSyn

Chem Corporation (Toronto, ON, Canada) (dichloroiodo-, dibromoiodo-, bromochloroiodo-, chlorodiiodo-, and bromodiiodomethane), Chloro-, dichloro-, and trichloroacetonitrile were purchased from ChemService (West Chester, PA, USA), bromoacetonitrile and dibromoacetonitrile were purchased from Arcos Organics (Geel,

27 Belguim) and bromochloroacetonitrile and iodoacetonitrile were purchased from

Crescent Chemical and Alfa Aesar (Ward Hill, MA, USA) respectively. All DBP

standards were purchased at the highest purities available. Additional organic and inorganic chemicals used were certified American Chemical Society (ACS) reagent grade and used without supplementary purification.

For experiments, deionized water (18.2 MΩ.cm-1) was obtained via a Barnstead ROPure

Infinity/NANOPure system (Barnstead-Thermolyne Corp. Dubuque, IA, USA).

Experimental pH was monitored with Orion 5 star pH meter equipped with Ross ultra combination electrode (Thermo Fisher Scientific, Pittsburgh, PA, USA). The pH of each experiment was adjusted via drop-wise addition of 0.1 N H2SO4 and 0.1 N NaOH. All glassware and polytetrafluoroethylene (PTFE) items were soaked in an aqueous chlorine or base bath for 24 hours. Upon removal, all items were rinsed with a considerable amount of deionized water and dried prior to experimental usage.

3.2 Source Water Characterization

Source water was obtained from the intake of Cleveland Garret Morgan Drinking Water

Treatment Plant in Cleveland, Ohio (USA). Source water characteristics from the

Cleveland water treatment plant are depicted below within Table 3.1. Total organic carbon (TOC) concentrations were measured using Shimadzu TOC analyzer (Shimadzu

Scientific, Columbia, MD, USA) which was calibrated according to Standard Method

505A (APHA et al, 1992). The ultraviolet absorbance at a wavelength of 254 nm

(UV254) and spectral characteristics of the NOM were measured with Shimadzu UV 1601

UV visible spectrophotometer in accordance with Standard Method 5910B (APHA et al,

1998). The specific ultraviolet absorbance at 254 nm (SUVA254) was calculated from the relation: SUVA = 100 x UV DOC . This analysis was performed due to DBP

254 254 formation has been linked to ⁄water characteristics such as SUVA254, bromide concentration and DOC concentration (Njam et al., 1994).

Table 3.1: Source water characteristics from Cleveland water Cleveland

source water

DOC (mg/L C) 2.51

Bromide (µM) < 0.5

Iodide (µM) < 0.5

-1 UV254 (cm ) 0.029

SUVA254 (L/mg-m) 1.17

Supplemental characterization of source water was achieved via florescence spectroscopy, which yielded the EEM spectra. Sample preparation for florescence spectra detection was achieved by the execution of the method developed by Chen et al.

(2003) with minor modifications. Sulfuric acid was utilized to acidify water samples in order to obtain a pH of 2.75 – 3.25, thus resulting in the removal of inorganic carbon.

Furthermore, samples were diluted to obtain a final DOC concentration of 1 mg/L with

0.01 M KCl to allow direct comparison of fluorescence intensities (Nguyen et al., 2005).

The EEM florescence spectra were obtained with an F-7000 FL fluorescence spectrophotometer (Hitachi Hi-Tech, Tokyo, Japan). This spectrophotometer utilizes a 29 xenon lamp as light source. The excitation and emission slit were both set to a band-pass of 10 nm. The spectra of the source water samples were measured at successive emission spectra, 2 nm intervals from 290 to 550 nm, while acquiring excitation wavelengths spaced at 5 nm from 204 to 404 nm. The resulting spectra were then merged into an

EEM, which was then constructed using SigmaPlot 12.0 (SPSS Inc.) to generate a contour map of fluorescence intensity with the regional integration (Figures 3.1). Chen et al. (2003) proposed a quantitative technique, entitled Florescence Regional Integration

(FRI), in order to quantify multiple EEM peaks of various shapes. The FRI technique is utilized to integrate the volume under EEM region (Table 3.2). This quantitative technique has also been used to analyse all wavelength-dependent florescence intensity data from EEM spectra (Marhuenda-Egea et al., 2007). Depicted within Table 3.2 are the five disparate regions proposed by Chen et al. (2003). The five distinctive regions depicted in the Cleveland source water NOM EEM are displayed in Table 3.3.

Table 3.2: Florescence EEM regions proposed by Chen et al. (2003) Excitation Emission Range

Regions Representation Range (nm) (nm)

I Aromatic 200 – 250 280 – 330

II Aromatic protein-like 200 – 250 330 – 380

III Fulvic acids 200 – 250 380 – 550

IV Soluble microbial by-products 250 – 400 280 – 380

V Humic acids 250 – 400 380 – 550

Table 3.3: Florescence regions for Cleveland source waters, 1 mg/L C Cleveland

Fluorescence Regions %

Aromatics (I) 2.4 22.7

Aromatic Protein-Like (II) 3.3 31.1

Fulvics (III) 3.0 28.7

Microbial (IV) 1.2 11.2

Humics (V) 0.7 6.3

Total 10.5 100

400 0 380 5 10 360 15 20 25 340

320 V 300 IV Excitation (nm) Excitation 280

260

240

I II III 220

300 320 340 360 380 400 420 440 460 480 500 520 540 Emission (nm)

Figure 3.1: Fluorescence excitation-emission spectrum (EEM) of Cleveland source water. [DOC] = 2.51 mg/L, SUVA254 = 1.17 L/mg.m

The EEM validates that the source water possesses a considerable percent volume within

region II, which corresponds to aromatic protein-like moieties. With respect to fulvic and humic regions, Cleveland source water exhibits a lower percent volume when compared to additional local source waters (Akron and Barberton) (Ackerson 2014). Humic acid is

a dominant component of humic substances, which are a result of plant material

degradation within terrestrial and aquatic environments by biological and natural

chemical processes (Hudson et al., 2007). Humic and fulvic acids have been known to

vary with respect to; vegetation in proximity to the watershed, algal content and

concentration, and possibly the season of year (Singer, 1994; Kavanaugh et al., 1980).

Leaf litter is an imperative source of fulvic acid (Schlesinger, 1997). The plant materials

in the watershed may be a result of the land coverage such as farms, forests or other

vegetative coverage. Aromatic protein-like moieties present in source water may

originate from bacterial communities (Elliot et al., 2006), potentially from the enzyme required for degradation of leaf matter/litter (Allan and Castillo, 2007; Benfield, 2006;

Suberkropp and Klug, 1976). Although, the relatively high percent volume of aromatic proteins observed within the Cleveland source water may be attributed to past and current algae blooms. The Cleveland source water does not contain appreciable quantities of

soluble microbial by-products.

3.3 Experimental Methods

The experimental procedures consisted of source waters experiments, which contain

NOM. Below is the procedure for making chloramine stock solutions. Pre-formed

chloramie solution was prepared to avoid the artifacts caused by the reactions of excess

free chlorine that may briefly exist when forming monochloramine in-situ (Duirk et al,

2005). Pre-formed monochloramine solution was prepared by mixing 5.64 mM

ammonium chloride with 3.7 mM hypochlorous acid to achieve a Cl/N molar ratio of 0.7

in a 10 mM carbonate buffer solution. The solution, under rapid mix condition on a

magnetic stir plate using a PTFE stir bar at a pH 8.5, was allowed to react and reach

equilibrium for 30 min. A higher pH (8.5) was used to minimise monochloramine

decomposition and to ensure monochloramine remains the active species (Symons et al,

1998) in the aqueous solution. The concentration of the preformed monochloramine was

checked with UV visible spectrophotometer and FAS/DPD titration (APHA et al., 2005).

3.3.1 DBP Formation Experiments with Cleveland Source Water

The Cleveland source water was filtered through 0.45 μm Whatman nylon membrane

filters (Whatman, West Chester, PA, USA) and stored at 4°C prior to use. Chlorination

and chloramination kinetic experiments were conducted under a pseudo first order

conditions using [Cl2]T:[iopamidol] = 20:1 over the pH range of 6.5 – 9. In order to

determine the effect of NOM on DBP formation, the NOM concentration in the

Cleveland source water was decreased by ½ and ¼ via dilution with deionized water.

Aqueous solutions were prepared in batch reactors containing NOM, iopamidol and

buffer in 500 mL Erlenmeyer flasks for each pH respectively. Buffer was used to

maintain the pH of the solution, 4 mM of phosphate buffer (for pH 6.5 and 7.5) and

borate buffer (for pH 8.5 and 9.0) were used to maintain the pH. Under rapid mix

condition, using a magnetic stir plate and a PTFE-coated stir bar, relatively high

concentration of aqueous chlorine was added to the aqueous solution at the requisite

[Cl2]T:[iopamidol] ratio. The relatively high concentration of aqueous chlorine was used to ensure excess oxidant was present within the aqueous mixture throughout the duration 34 of the experiment, thus not limiting the reaction. Prior to the addition, the chlorine

concentration was checked using ferrous ammonium sulphate (FAS)/N, N′-diphenyl-p-

phenylenediamine (DPD) titration (APHA et al., 2005). In order to ensure a uniform

mixture, stirring was maintained for about 3 min. Triplicate analysis was desired

therefore the aliquots of the aqueous solution were transferred into three 128 mL amber

vials with PTFE septa and stored headspace free at 25±1°C in an incubator for a reaction

time of 72 hours. A similar experimental protocol as depicted above was utilized with

pre-formed monochloramine (that is zero minute of free aqueous chlorine contact time).

3.4 Analytical Procedures

Analytical procedures will be discussed with respect analyte of interest. Furthermore,

additional steps may be required in order to perform the desired analysis given the

analytical equipment employed.

3.4.1 Disinfection By-product

THMs, HANs and HAAs were extracted using a micro liquid-liquid extraction with

MtBE at acidic pH. The THMs analysed included bromodichloromethane (CHBrCl2), dibromochloromethane (CHBr2Cl), chloroform (CHCl3), dichloroiodomethane (CHCl2I),

bromochloroiodomethane (CHBrClI), bromoform (CHBr3), dibromoiodomethane

(CHBr2I), chlorodiiodomethane (CHClI2), bromodiiodomethane (CHBrI2), and iodoform

(CHI3). Also chloroactonitrile (CAN), trichloroacetontrile (TCAN), dichloroacetonitrile

(DCAN), bromochloroacetonitrile (BCAN), dibromoacetonitrile (DBAN)

bromoacetonitrile (BAN), and iodoacetontrile (IAN) were the HAN compounds analysed.

THMs and HANs were extracted using the US EPA method 551.1 (Munch and Hautman,

1998) with minor modifications. The sample was acidified with 5 mL of concentrated sulphuric acid 30 g of anhydrous sodium sulphate salt (dried at 100°C) was added. Then,

3 mL of MtBE and 10 μL of 123.9 mM of 1,2-dibromopropane internal standard were added into the acidified sample to achieve approximately 12.4 μM internal standard in the sample. MtBE was used to extract non-dissociated acidic compounds (APHA AWWA and WEF, 1995). The salt was added to decrease the activity of inorganic compounds and increase the activity of the organic compounds, increasing partitioning of the DBPs from the aqueous phase to the organic phase (MtBE) (US EPA, 2013), thus increasing the extraction efficiency. The 100 mL sample, in the 128 mL amber bottles were capped with polyseal cone-lined cap, hand-shaken for a minute and then shaken on the wrist action shaker (Burrell Scientific, Pittsburgh, PA, USA) for 30 minutes. After the mechanical shake, the sample poured into a 100-mL volumetric flask and left to settle for

3 minutes. A disposable Pasteur pipette was used to transfer the organic layer (i.e.,

MtBE) into a 2 mL GC autosampler vial after being filtered through another Pasteur

pipette filled with glass wool and dried anhydrous sodium sulphate salt in order to deplete

the water from the organic extract. The extracted sample was then split – 0.5 mL used for derivatization with diazomethane for HAAs analysis and the remaining used for

THMs and HANs analyses. The extracted samples were stored in the freezer.

HAAs were measured using a modified US EPA method 552.1 (Hodgeson and Becker,

1992), which uses liquid-liquid extraction with MTBE, derivatization with diazomethane

and analysis with GC/MS. The HAA compounds analysed were chloroacetic acid

(CAA), bromoacetic acid (BAA), dichloroacetic acid (DCAA), trichloroacetic acid

(TCAA), iodoacetic acid (IAA), bromochloroacetic acid (BCAA), bromodichloroacetic

acid (BDCAA) and dibromoacetic acid (DBAA). A 0.5 mL aliquot of the extracted

sample was methylated with diazomethane for the production of methyl ester or other

derivatives for gas chromatographic separation (APHA, AWWA and WEF, 1995).

Diazomethane was generated by adding 0.367 g diazald and 1 mL carbitol (2-[2-

ethoxyethoxy] ethanol) to the inner tube of the diazomethane generator. Then 3 mL of

MtBE was added to the outer tube of the diazomethane generator. The two parts of the

generator were assembled and the lower part of the outer tube was immersed in ice bath

to ensure an isothermal condition of 0°C was maintained. After equilibrating to 0°C, 1.5

mL of KOH (37%) was slowly injected (drop-wise) into the generator through the septum

to initiate the reaction. The apparatus was shaken gently by hand to ensure uniform

mixture of reactants in the inner tube while avoiding spill into the outer tube. When the solution in the outer tube becomes yellow it is an indication of excess diazomethane. The apparatus with the solution was left to stand for 50 minutes, after which the tube was opened to destroy unreacted diazomethane with activated silica. After preparing the diazomethane, about 0.5 mL of the extracted sample was transferred into another GC autosampler vial and 250 μL of the diazomethane added to it. The sample was allowed to

derivatize for 30 minutes to allow adequate methylation of the HAAs, and then 1 – 3

grains of activated silica were added to the sample to destroy any excess diazomethane.

3.6 Analyses of DBPs

The extracted and derivatized samples were analyzed with 7890A GC system equipped

with 63Ni microelectron capture detector (μECD) (Agilent Technologies, Santa Clara,

CA, USA). A 30-m Restek Rxi–5Sil MS with Integra- Guard GC column (Restek

Corporation, Bellefonte, PA, USA) internal diameter 0.25 mm, film thickness 0.5 μm was

usedachieved separation of analytes. Samples were delivered by 7693 autosampler

(Agilent Technologies, Santa Clara, CA, USA). Splitless injections were achieved by

injecting 1 μL of the sample on the column. The flowrate through the column as 1 mL/min, the temperature of the μECD was 250°C, and the make-up gas was ultrahigh purity nitrogen gas with flow rate of 19 mL/min. The carrier gas employed was helium gas (ultrahigh purity). There were two oven temperature programming used – one for analysis of THMs and HANs (Table 3.4) and the other for HAAs analysis (Table 3.5).

Table 3.4: Oven temperature programming for THMs and HANs analysis on GC/μECD Rate (°C/min) Temperature (°C) Hold time (min) Run time (min)

Initial 50 10 10

Ramp 1 2.5 65 0 16

Ramp 2 5 85 0 20

Ramp 3 7.5 205 0 36

Ramp 4 10 280 0 43.6

Table 3.5: Oven temperature programming for HAAs analysis on GC/μECD Rate (°C/min) Temperature (°C) Hold time (min) Run time (min)

Initial 50 10 10

Ramp 1 0.25 50.5 5 17

Ramp 2 0.25 52 5 28

Ramp 3 0.25 62.5 0 70

Ramp 4 35 280 0 76.214

THMs, HANs and HAAs calibration standards were prepared by spiking each standard

into deionized water. Calibration curves were created by serial dilution of the initial

spiked standard. The known concentrations of the THMs and HANs standards were

extracted utilizing the aforementioned extraction procedure described above using 10 μL

of 123.9 mM 1,2-dibromopropane internal standard to achieve about 12.4 μM internal

standard within each respective sample. The HAAs standards, of known concentrations

were derivatized with diazomethane after extraction using the same volume and concentration of 1,2-dibromopropane as internal standard. A calibration curve of concentration of the standard versus the relative response of the standard solution to the internal standard was developed to calculate the concentrations of the DBPs in the samples. The relative response of standard to the internal standard is referred to in the calibration curve as response ratio (shown on respective axis). The calibration curves for

all the standard solutions are shown in figures 3.2 to 3.36. The concentration of the

specific DBP was calculated from the equation of the line of best fit of the corresponding

standard curve. The limits of quantification (LOQ) for the DBPs are shown in table 3.6. 39

1200

1000

800 ](nM)

3 600 y = 1471.4x R² = 0.9785 [CHCl 400

200

0 0.0 0.2 0.4 0.6 0.8 1.0

Response ratio

Figure 3.2: Calibration curve for CHCl3using chloroform. [CHCl3] = 0 – 1000 nM

350

300

250

200 Cl](nM) 2 150 y = 109.38x

[CHBr R² = 0.9974 100

0 0.0 0.5 1.0 1.5 2.0 2.5 3.0

Response ratio

Figure 3.3: Calibration curve for CHBr2Cl using dibromochloromethane. [CHBr2Cl] = 0 – 300 nM

160

140

120

100 ](nM) 2 80

[CHBrI 60 y = 135.95x R² = 0.9925 40

0 0.0 0.2 0.4 0.6 0.8 1.0

Response ratio

Figure 3.4: Calibration curve for CHBrI2 using bromodiiodomethane. [CHBrI2] = 0 – 125 nM

300

250

200 ](nM) 2 150 y = 266.06x

[CHClI R² = 0.9993 100

0 0.0 0.2 0.4 0.6 0.8 1.0 Response ratio

Figure 3.5: Calibration curve for CHClI2 using chlorodiiodomethane. [CHClI2] = 0 – 250 nM

43 300

250

200 I] (nM) I]

2 150 y = 1152.7x

[CHBr R² = 0.9941 100

0 0.00 0.05 0.10 0.15 0.20 0.25 Response ratio

Figure 3.6: Calibration curve for CHBr2I using dibromoiodomethane. [CHBr2I] = 0 – 250 nM

44 300

250

200

150 y = 1595.3x R² = 0.998 [CHBrClI](nM) 100

0 0.00 0.02 0.04 0.06 0.08 0.10 0.12 0.14 0.16 0.18 Response ratio

Figure 3.7: Calibration curve for CHBrClI using bromochloroiodomethane. [CHBrClI] = 0 – 250 nM

45 600

500

400 ](nM)

3 300 y = 204.28x R² = 0.9968 [CHBr 200

100

0 0.0 0.5 1.0 1.5 2.0 2.5 3.0

Response ratio

Figure 3.8: Calibration curve for CHBr3 using bromoform. [CHBr3] = 0 – 500 nM

600

500

400

y = 2013.1x I] (nM) I]

2 300 R² = 0.9985 [CHCl 200

100

0 0.00 0.05 0.10 0.15 0.20 0.25 0.30

Response ratio

Figure 3.9: Calibration curve for CHCl2I using dichloroiodomethane. [CHCl2I] = 0 – 500 nM

400

300

Br](nM) 200 2 y = 158.05x R² = 0.9965 [CHCl

100

0 0.0 0.5 1.0 1.5 2.0 2.5 3.0

Response Ratio

Figure 3.10: Calibration curve for CHCl2Br using bromodichloromethane. [CHCl2Br] = 0 – 400 nM

y = 68.338x ](nM) 3 30 R² = 0.9933 [CHI 20

0 0.0 0.2 0.4 0.6 0.8 1.0

Response ratio

Figure 3.11: Calibration curve for CHI3 using iodoform. [CHI3] = 0 – 50 nM

600

500

400

300

[CAN](nM) y = 478.7x 200 R² = 0.9988

100

0 0.0 0.2 0.4 0.6 0.8 1.0

Response ratio

Figure 3.12: Calibration curve for CAN using chloroacetonitrile. [CAN] = 0 – 500 nM

600

500

400

300 y = 2852.9x [DCAN](nM) 200 R² = 0.9991

100

0 0.00 0.02 0.04 0.06 0.08 0.10 0.12 0.14 0.16 0.18 0.20

Response ratio

Figure 3.13: Calibration curve for DCAN using dichloroacetonitrile. [DCAN] = 0 – 500 nM

140

120

100

[TCAN](nM) y = 160.4x R² = 0.9631 40

0 0.0 0.2 0.4 0.6 0.8 1.0

Response ratio

Figure 3.14: Calibration curve for TCAN using trichloroacetonitrile. [TCAN] = 0 – 125 nM

140

120

100

y = 129.6x 60 R² = 0.9987 [BAN](nM)

0 0.0 0.2 0.4 0.6 0.8 1.0 1.2

Response ratio

Figure 3.15: Calibration curve for BAN using bromoacetonitrile. [BAN] = 0 – 125 nM

300

250

200

150 y = 346.63x R² = 0.9977 [DBAN](nM) 100

0 0.0 0.2 0.4 0.6 0.8

Response ratio

Figure 3.16: Calibration curve for DBAN using dibromoacetonitrile. [DBAN] = 0 – 250 nM

300

250

200

150 y = 372.69x R² = 0.9931 [BCAN](nM) 100

0 0.0 0.2 0.4 0.6 0.8

Response ratio

Figure 3.17: Calibration curve for BCAN using bromochloroacetonitrile. [BCAN]=0–250 nM

y = 33.062x 15

[IAN](nM) R² = 0.9946

0 0.0 0.2 0.4 0.6 0.8 1.0 1.2

Response ratio

Figure 3.18: Calibration curve for IAN using iodoacetonitrile. [IAN] = 0 – 31 nM

300

250

200

150 y = 773.91x

[CAA](nM) R² = 0.9997 100

0 0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.35

Response ratio

Figure 3.19: Calibration curve for CAA using chloroacetic acid. [CAA] = 0 – 250 nM

600

500

400

300

y = 376.03x [DCAA](nM) 200 R² = 0.9693

100

0 0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6

Response ratio

Figure 3.20: Calibration curve for DCAA using dichloroacetic acid. [DCAA] = 0 – 500 nM

300

250

200

150

[TCAA](nM) y = 100.41x 100 R² = 0.9986

0 0.0 0.5 1.0 1.5 2.0 2.5 3.0

Response ratio

Figure 3.21: Calibration curve for TCAA using trichloroacetic acid. [TCAA] = 0 – 250 nM

300

250

200

150 y = 105.74x

[BCAA](nM) R² = 0.9969 100

0 0.0 0.5 1.0 1.5 2.0 2.5 3.0

Response ratio

Figure 3.22: Calibration curve for BCAA using bromochloroacetic acid. [BCAA]=0–250 nM

300

250

200

150 y = 85.951x R² = 0.9982 [BDCAA](nM) 100

0 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 Response ratio

Figure 3.23: Calibration curve for BDCAA using bromodichloroacetic acid. [BDCAA] = 0 –250 nM

1200

1000

800

600 y = 1378.1x [BAA](nM) R² = 0.991 400

200

0 0.0 0.2 0.4 0.6 0.8 Response ratio

Figure 3.24: Calibration curve for BAA using bromoacetic acid. [BAA] = 0 – 1000 nM

62 600

500

400

300

[DBAA](nM) y = 139.48x 200 R² = 0.9949

100

0 0 1 2 3 4 Response ratio Figure 3.25: Calibration curve for DBAA using dibromoacetic acid. [DBAA] = 0 – 500 nM

63 140

120

100

y = 83.588x 60

[IAA](nM) R² = 0.9896

0 0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6 1.8 Response ratio

Figure 3.26: Calibration curve for IAA using iodoacetic acid. [IAA] = 0 – 125 nM

64 Table 3.6: Limit of quantification (LOQ) for the detection of DBPs THMs LOQ HANs LOQ HAAs LOQ (nM) (nM) (nM)

CHCl 1.0 CAN 1.0 CAA 1.0 3

CHBrCl2 1.0 TCAN 1.0 BAA 1.0

CHBr2Cl 1.0 DCAN 1.0 DCAA 1.0

CHClBrI 1.0 BAN 1.0 TCAA 1.0

CHCl2I 0.2 BCAN 1.0 IAA 0.2

CHClI2 0.2 DBAN 1.0 BCAA 1.0

CHBr3 1.0 IAN 0.2 BDCAA 1.0

CHBr2I 0.2 DBAA 1.0

CHBrI2 0.2

CHI3 0.2

CHAPTER IV

RESULTS AND DISCUSSION

4.1 Introduction

This chapter evaluates the findings obtained from the investigation of iopamidol transformation in the presence of a chlorinated oxidants and respective contribution to the formation of DBPs with respect to pH and concentrations of iopamidol and total organic carbon (TOC). This work compliments that of Pushpita Kumkum in 2013 and Nana

Ackerson in 2014. To reiterate, the data to be discussed was obtained from experiments in which iopamidol concentration was varied in batch reactors containing source water

(Cleveland, OH), amongst a buffer and internal standard, over the pH range of 6.5 – 9.0.

Additionally, iopamidol concentration was held constant while TOC was varied via dilution with dionized water over the pH range of 6.5 – 9.0. Source water experiments were performed in the absence of iopamidol, where the respective chlorinated oxidant, buffer and internal standard were present to act as a control over the pH range of 6.5 –

9.0. This chapter possesses two sections; the first section will evaluate the predominant trends that were observed within the data and provide a possible explanation of said trends. While the second section will statistically evaluate the data in order to support or dispute the proposed explanation.

66 4.2 Predominant Observed Trends

First and foremost, it should be noted that the observed trends will be discussed

separately with respect to each of the two chlorinated oxidants that were investigated.

The observed major trends within the chlorine data set were with respect to pH (6.5, 7.5,

8.5 and 9.0) and TOC levels (2.51, 1.26 and 0.63 mg/L-L). Figure 4.1 (displayed on consecutive page) exhibits the iodo-DBP formation at 72 hours in the presence of iopamidol (5 μM) and aqueous chlorine ([Cl2]T = 100 μM) at full DOC capacity (2.51

mg/L-L) with respect to pH. Within said figure, the predominantly observed trend is

contingent upon pH. At pH 6.5 the lowest formation is observed (6.4 nM), a considerable

increase is observed formation from 6.5 to 7.5 (≈ 18.6 nM), a slight increase in formation

is observed from 7.5 to 8.5 (≈ 11.2 nM) and a considerable increase is observed from 8.5

to 9.0 (≈21.3), which is the highest observed formation.

60 CHCl2I CHBrI2 50

30 Concentration,nM 20

0 6.5 7.5 8.5 9.0 pH Figure 4.1: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L.

68 Figure 4.2 (shown below) expresses the iodo-DBP formation at 72 hours in the presence of iopamidol (5 μM) and aqueous chlorine ([Cl2]T = 100 μM) at half DOC capacity (1.26 mg/L-L) with respect to pH. The aforementioned predominant observed trend with respect to pH in Figure 4.1 is also observed within Figure 4.2, if not strengthened despite the reduction in DOC. Case in point, increase from pH 6.5 to 7.5 was approximately 32.9 nM, slight increase from 7.5 to 8.5 being 15.8 nM and considerable increase from 8.5 to

9.0 being 48 nM thus strengthened the aforementioned trend with respect to pH.

120

100 CHCl2I

Concentration,nM 40

0 6.5 7.5 8.5 9.0 pH Figure 4.2: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH);

[Iopamidol] = 5 μM, [Cl 2]T = 100 μM, [Buffer] = 4m M, temperature = 25 °C, [TOC] = 1.26 mg/L-L.

Furthermore, the data continues to exhibit said predominant observed trend when the

DOC capacity is further reduced to quarter capacity (0.63 mg/L-L), Figure 4.3 (shown below). Case in point, an increase from pH 6.5 to 7.5 was approximately 29.4 nM, slight increase from 7.5 to 8.5 was 18 nM and a considerable increase from 8.5 to 9.0 was 56.3 nM.

140

120 CHCl2I

100

60 Concentration,nM 40

0 6.5 7.5 8.5 9.0 pH Figure 4.3: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamid ol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl ] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2 T 0.63 mg/L-L.

Figure 4.4 (shown below) exhibits iodo-DBP formation at 72 hours in the presence of iopamidol (2.5 μM) and aqueous chlorine ([Cl2]T = 100 μM) at full DOC capacity (2.51 mg/L-L) with respect to pH. Overall, formation considerably decreased when iopamidol concentration was reduced (5.0 μM to 2.5 μM), the same trend was observed with respect to pH as previously discussed, maximum formation at pH 9.0 which was approximately

10.4 nM. It should be noted, upon further reduction of iopamidol (2.5 μM to 1.0 μM) iodo-DBP formation was observed although it was below quantification limit.

12 CHCl2I CHBr2I 10

6 Concentration,nM 4

0 6.5 7.5 8.5 9.0 pH Figure 4.4: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L.

71 Figure 4.5 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of aqueous chlorine ([Cl2]T = 100 μM) at full DOC capacity (2.51 mg/L-L) with respect to

pH. Within the source water (Figure 4.5), iodo-DBP formation was only observed at pH

6.5 and 9.0, pH 6.5 exhibited minimal formation while 9.0 exhibited a relatively larger

magnitude of formation (respective to pH 6.5).

CHCl2I 25 CHClI2 CHBrClI

Concentration,nM 10

0 6.5 7.5 8.5 9.0 pH Figure 4.5: Observed Iodo-DBP formation at 72 hours as a function of pH where the

reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L -L.

Figure 4.6 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of

aqueous chlorine ([Cl2]T = 100 μM) at half DOC capacity (1.26 mg/L-L) with respect to

pH, minimal formation is observed (< 2.5 nM) across all pHs with no predominant trend.

3.0

CHCl2I 2.5

2.0

1.5

Concentration,nM 1.0

0.5

0.0 6.5 7.5 8.5 9.0 pH Figure 4.6: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 1.26 mg/L -L.

Figure 4.7 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of

aqueous chlorine ([Cl2]T = 100 μM) at quarter DOC capacity (0.63 mg/L-L) with respect to pH, minimal formation is observed (< 1.2 nM) across all pH. Despite the lack of iodo-

DBP formation at pH 6.5, the aforementioned predominant observed trend with respect to pH is observed, that being minimal formation at pH 6.5, a considerable increase is observed formation from 6.5 to 7.5, a minimal increase in formation is observed from 7.5 to 8.5 and a considerable increase is observed from 8.5 to 9.0, which is the highest observed formation.

1.6

1.4 CHCl2I 1.2

1.0

0.8

0.6 Concentration,nM

0.4

0.2

0.0 6.5 7.5 8.5 9.0 pH

Figure 4.7: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 0.63 mg/L-L.

Figure 4.8 (shown below) displays select DBP formation (chloroform,

dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5 μM) and aqueous chlorine ([Cl2]T = 100 μM) at pH 6.5. Chloroform decreased from full (2.51

mg/L-L) to half (1.26 mg/L-L) DOC capacity by approximately half and then remained

approximately the same at quarter (0.63 mg/L-L), trichloroacetic acid formation showed

a considerable increase from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity and

remained approximately the same at quarter (0.63 mg/L-L), dichloroiodomethane

formation remained approximately the same despite DOC variation.

1200

CHCl3 1000 TCAA CHCl2I

800

600

Concentration,nM 400

200

0 0.63 1.26 2.51 [TOC], mg/L-L Figure 4.8: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH

6.5; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

Figure 4.9 (shown on consecutive page) displays select DBP formation (chloroform,

dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5 μM) and aqueous chlorine ([Cl2]T = 100 μM) at pH 7.5. Chloroform exhibited a slight decrease

(approximately 51.4 nM) from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity

and remained approximately the same at quarter (0.63 mg/L-L) (approximately 7.3 nM

increase). Trichloroacetic acid considerably increased from full (2.51 mg/L-L) to half

(1.26 mg/L-L) DOC capacity (approximately 305.1 nM), formation slightly increased at

quarter DOC (0.63 mg/L-L) (approximately 35.1 nM). Dichloroiodomethane formation

slightly increased from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity

(approximately 14.71 nM) and remained approximately the same at quarter DOC (0.63

mg/L-L)(3.51 nM decrease).

1200

CHCl3 1000 TCAA

CHCl2I

800

600

Concentration,nM 400

200

0 0.63 1.26 2.51 [TOC], mg/L-L Figure 4.9: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 7.5; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

77 Figure 4.10 (shown on consecutive page) displays select DBP formation (chloroform,

dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5.0 μM) and aqueous chlorine ([Cl2]T = 100 μM) at pH 8.5. Chloroform formation decreased from full

(2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity (147.4 nM decrease) and continued to

slightly decrease from half (1.26 mg/L-L) to quarter (0.63 mg/L-L) DOC (approximately

77.1 nM). Trichloroacetic acid formation increased from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity (approximately 351.3 nM) and decreased from half (1.26 mg/L-

L) to quarter (0.63 mg/L-L) DOC (approximately 143.8). Dichloroiodomethane

formation increased from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity

(approximately 19.32 nM) and remained approximately the same at quarter (0.63 mg/L-

L) DOC (approximately 1.3 nM decrease).

1200

CHCl3 1000 TCAA CHCl2I

800

600

Concentration,nM 400

200

0 0.63 1.26 2.51 [TOC], mg/L-L Figure 4.10: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 8.5; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

79 Figure 4.11 (shown on consecutive page) displays select DBP formation (chloroform,

dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5.0 μM) and

aqueous chlorine ([Cl2]T = 100 μM) at pH 9.0. Chloroform formation increased from full

(2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity (approximately 159.9 nM) and

continued to increase from half (1.26 mg/L-L) to quarter (0.63 mg/L-L) DOC capacity

(approximately 106.6 nM). It should be noted that this trend differs from the

aforementioned trends at previously discussed pHs (6.5, 7.5 and 8.5). Trichloroacetic

acid increases from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity

(approximately 133.2 nM) and decreased from half (1.26 mg/L-L) to quarter (0.63 mg/L-

L) DOC capacity (approximately 181.6 nM). Dichloroiodomethane formation increased

from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity (approximately 45.96 nM)

and remained approximately the at quarter (0.63 mg/L-L) DOC capacity (approximately

6.95 nM).

1200

CHCl3 1000 TCAA CHCl2I

800

600

Concentration,nM 400

200

0 0.63 1.26 2.51 [TOC], mg/L-L Figure 4.11: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous chlorine an d source water (Cleveland, OH) at pH

9.0; [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

Figure 4.12 (shown below) displays select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous chlorine ([Cl2]T = 100 μM) at full DOC (2.51 mg/L-L) capacity and pH 6.5. Chloroform formation decreased from 5.0

μM to 2.5 μM, formation remained approximately the same for 2.5 μM, 1.0 μM and 0.0

μM (approximately 16.1 nM variance amongst varied concentration). Trichloroacetic acid formation increased from 5.0 μM to 2.5 μM, decreased from 2.5 μM to 1.0 μM and decreased from 1.0 μM to 0 μM. Dichloroiodomethane concentration decreased with respect to decreasing iopamidol concentration.

1000

CHCl3 TCAA 800 Cl2I

600

400 Concentration,nM

200

0 0.0 1.0 2.5 5.0 [Iopamidol], µM Figure 4.12: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH)

at pH 6.5; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L- L. Figure 4.13 (shown on consecutive) displays select DBP formation (chloroform,

dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous chlorine ([Cl2]T = 100 μM) at

full DOC (2.51 mg/L-L) capacity and pH 7.5. Chloroform formation increased from 5.0

μM to 2.5 μM (approximately 35.6 nM) and then decreased with respect to iopamidol

concentration. Trichloroacetic acid formation increased from 5.0 μM to 2.5 μM

(approximately 240.6 nM) and then continued to decrease with respect to decreasing iopamidol concentration. Dichloroiodomethane formation decreased with respect to decreasing iopamidol concentration.

1000

CHCl3 TCAA 800 CHCl2I

600

400 Concentration,nM

200

0 0.0 1.0 2.5 5.0 [Iopamidol], µM

Figure 4.13: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 7.5; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L- L.

84 Figure 4.14 (displayed on consecutive page) displays select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous chlorine ([Cl2]T = 100 μM) at full DOC (2.51 mg/L-L) capacity and pH 8.5. Chloroform formation decreased from 5.0

μM to 2.5 μM, an increase was observed at 1.0 μM and decreased formation was observed at 0.0 μM. Trichloroacetic acid formation increased from 5.0 μM to 2.5 μM

(approximately 116.3 nM), then decreased with respect to decreasing iopamidol concentration. Dichloroiodomethane formation decreased with respect to decreasing iopamidol concentration.

1000

CHCl3 TCAA 800 CHCl2I

600

400 Concentration,nM

200

0 0.0 1.0 2.5 5.0 [Iopamidol], µM

Figure 4.14: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 8.5; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L- L.

86 Figure 4.15 (shown on consecutive page) displays select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous chlorine ([Cl2]T = 100 μM) at full DOC (2.51 mg/L-L) capacity and pH 9.0. Chloroform formation decreased from 5.0

μM to 2.5 μM, increased from 2.5 μM to 1.0 μM and then decreased from 1.0 μM to 0.0

μM. Trichloroacetic acid formation increased from 5.0 μM to 2.5 μM (approximately

54.5 nM) and then decreased with decreasing iopamidol concentration.

Dichloroiodomethane formation decreased with decreasing iopamidol concentration.

1000 CHCl3 TCAA CHCl2I 800

600

400 Concentration,nM

200

0 0.0 1.0 2.5 5.0 [Iopamidol], µM

Figure 4.15: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH) at pH 9.0; [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L- L.

Figure 4.16 (shown on consecutive page) depicts dichloroiodomethane formation with respect to DOC level (2.51, 1.26 and 0.63 mg/L-L), in the presence of aqueous chlorine

([Cl2]T = 100 μM) as a function of pH. Formation at pH 6.5 remained the same (0.4 nM variance), pH 7.5 displays a 14.7 nM increase from full (2.51 mg/L-L) to half (1.26 mg/L-L) capacity and a 3.5 nM decrease from half (1.26 mg/L-L) to quarter (0.63 mg/L-

L) capacity, pH 8.5 exhibits an 19.3 nM increase from full (2.51 mg/L-L) to half (1.26 mg/L-L) capacity and a 1.3 nM decrease from half (1.26 mg/L-L) to quarter (0.63 mg/L-

L) capcity, pH 9.0 exhibits a 46 nM increase from full (2.51 mg/L-L) to half (1.26 mg/L-

L) capacity and a 7 nM increase from half (1.26 mg/L-L)to quarter (0.63 mg/L-L) capacity.

89 140

120 2.51 mg/L-L 1.26 mg/L-L 0.63 mg/L-L 100

60 Concentration(nM) 40

0 6.5 7.5 8.5 9.0 pH

Figure 4.16: Observed CHCl2I formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

Figure 4.17 (shown on next page) displays trichloroacetic acid formation with respect to

DOC level (2.51, 1.26 and 0.63 mg/L-L), in the presence of aqueous chlorine ([Cl2]T =

100 μM) as a function of pH. Formation exhibited a 192.6 nM increase from full (2.51 mg/L-L) to half (1.26 mg/L-L) capacity and a 43.7 nM decrease from half (1.26 mg/L-L) to quarter (0.63 mg/L-L) at pH 6.5, a 305.1 nM increase was observed at from full (2.51 mg/L-L) to half (1.26 mg/L-L) capacity and a 35.1 nM increase from half (1.26 mg/L-L) to quarter (0.63 mg/L-L) capacity at pH 7.5, a 351.3 nM increase was observed from full

(2.51 mg/L-L) to half (1.26 mg/L-L) capacity and a 143.8 nM decrease from half (1.26

mg/L-L) to quarter (0.63 mg/L-L) capacity at pH 8.5, a 133.2 nM increase was observed

from full (2.51 mg/L-L) to half (1.26 mg/L-L) and a 181.6 nM decrease was observed from half (1.26 mg/L-L) to quarter (0.63 mg/L-L) capacity at pH 9.0.

500

2.51 mg/L-L 1.26 mg/L-L 400 0.63 mg/L-L

300

200 Concentration(nM)

100

0 6.5 7.5 8.5 9.0 pH Figure 4.17: Observed trichloroacetic acid formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

In regard to the monochloramine data set, the observed major trends within the were with

respect to pH (6.5, 7.5, 8.5 and 9.0) and TOC levels (2.51, 1.26 and 0.63 mg/L-L).

91 Figure 4.18 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of iopamidol (5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at full DOC capacity (2.51 mg/L-L) with respect to pH. Iodo-DBP formation decreased with respect to increasing pH, highest formation was observed at pH 6.5 and lowest formation was observed at pH 9.0.

CHCl2I CHClI 40 2 CHBrClI CHBr2I CHBrI 30 2 CHI3

20 Concentration,nM

0 6.5 7.5 8.5 9.0 pH Figure 4.18: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L- L.

Figure 4.19 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of iopamidol (5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at half DOC

capacity (1.26 mg/L-L) with respect to pH. Iodo-DBP formation decreased with respect

to increasing pH, highest formation was observed at pH 6.5 and lowest formation was

observed at pH 9.0.

CHCl2I CHClI2 20 CHBrClI CHBrI2 CHI 15 3

10 Concentration,nM

0 6.5 7.5 8.5 9.0 pH Figure 4.19: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 1.26 mg/L-L.

93 Figure 4.20 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of iopamidol (5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at quarter DOC

capacity (0.63 mg/L-L) with respect to pH where iodo-DBP formation exhibit the same

trend as full (2.51 mg/L-L) and half (1.26 mg/L-L) DOC capacity.

CHCl2I CHClI2 20 CHBrClI CHBrI2 CHI3 15

10 Concentration,nM

0 6.5 7.5 8.5 9.0 pH Figure 4.20: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH 2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 0.63 mg/L -L.

Figure 4.21 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of iopamidol (2.5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at full DOC

capacity (2.51 mg/L-L) with respect to pH. Aforementioned decreasing trend with

respect to increasing pH was observed in regard to iodo-DBP formation.

CHCl2I 20 CHClI2 CHBrClI CHBr I 15 2 CHBrI2 CHI3 10 Concentration,nM

0 6.5 7.5 8.5 9.0 pH Figure 4.21: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 2.5 μM, [NH Cl] = 100 μM, [Buffer] = 4mM, 2 temperature = 25 °C, [TOC] = 2.51 mg/L-L.

Figure 4.22 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of iopamidol (1.0 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at full DOC

capacity (2.51 mg/L-L) with respect to pH. Iodo-DBP formation exhibits the same

aforementioned trend, decreasing formation with respect to increasing pH.

CHCl3

CHCl2I

15 CHBr2I IAN

10 Concentration,nM

0 6.5 7.5 8.5 9.0 pH

Figure 4.22: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 1.0 μM, [NH 2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L -L.

Figure 4.23 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence

of aqueous monochloramine ([NH2Cl] = 100 μM), in the absence of iopamidol, at full

DOC capacity (2.51 mg/L-L) with respect to pH. Observed iodo-DBP formation trend

coincides with previous discussed trend, decreases as pH increases.

16 CHCl I 14 2 CHClI2 12 CHBrClI CHBr2I 10 CHBrI2 CHI3 8

6 Concentration,nM

0 6.5 7.5 8.5 9.0 pH Figure 4.23: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH);

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L.

Figure 4.24 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of aqueous monochloramine ([NH2Cl] = 100 μM), in the absence of iopamidol, at half

DOC capacity (1.26 mg/L-L) with respect to pH. Iodo-DBP formation was minimal (<

1.2 nM) and possessed no predominant trend.

1.4

CHCl2I 1.2

1.0

0.8

0.6 Concentration,nM 0.4

0.2

0.0 6.5 7.5 8.5 9.0 pH Figure 4.24: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 1.26 mg/L-L. 2

Figure 4.25 (shown below) exhibits the iodo-DBP formation at 72 hours in the presence of aqueous monochloramine ([NH2Cl] = 100 μM), in the absence of iopamidol, at quarter

DOC capacity (0.63 mg/L-L) with respect to pH. Iodo-DBP formation was minimal (<

1.0 nM), possessing no predominant formation trend.

1.2

1.0 CHCl2I

0.8

0.6

Concentration,nM 0.4

0.2

0.0 6.5 7.5 8.5 9.0 pH Figure 4.25: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH);

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 0.63 mg/L-L.

Figure 4.26 (shown below) exhibits select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at pH 6.5. DBP formation decreases as

DOC capacity decreases, dichloroiodomethane formation remained the same at half (1.26

mg/L-L) and quarter (0.63 mg/L-L) DOC capacity (0.52 nM variance between half and

quarter).

CHCl3 TCAA CHCl I 30 2

20 Concentration,nM

0 0.63 1.26 2.51 [TOC], mg/L-L

Figure 4.26: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 6.5; [Iopami dol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

100

Figure 4.27 (shown below) exhibits select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at pH 7.5. Overall, DBP formation decreased as DOC capacity decreased, dichloroiodomethane formation remained the same at half (1.26 mg/L-L) and quarter (0.63 mg/L-L) DOC capacity (0.96 variance).

CHCl3 30 TCAA CHCl2I

20 Concentration,nM

0 0.63 1.26 2.51 [TOC], mg/L-L Figure 4.27: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 7.5; [Iopamidol] = 5 μM, [NH 2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

101

Figure 4.28 (shown below) exhibits select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at pH 8.5. DBP formation decreased as

DOC capacity decreased, dichloroiodomethane remained the approximately the same for

half (1.26 mg/L-L) and quarter (0.63 mg/L-L) DOC capacity (0.53 variance between half

and quarter).

CHCl3 TCAA CHCl I 30 2

20 Concentration,nM

0 0.63 1.26 2.51 [TOC], mg/L-L Figure 4.28: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 8.5; [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

102 Figure 4.29 (shown below) exhibits select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to TOC, full (2.51 mg/L-L),

half (1.26 mg/L-L) and quarter (0.63 mg/L-L), in the presence of iopamidol (5 μM) and aqueous monochloramine ([NH2Cl] = 100 μM) at pH 9.0. As previously described,

formation decreased as DOC capacity decreased. Dichloroiodomethane remained the

approximately the same for half (1.26 mg/L-L) and quarter (0.63 mg/L-L) DOC capacity

(0.47 variance between half and quarter).

CHCl3 TCAA CHCl I 30 2

20 Concentration,nM

0 0.63 1.26 2.51 [TOC], mg/L-L Figure 4.29: Observed DBP formation at 72 hours as a function of TOC where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 9.0; [Iopami dol] = 5 μM, [NH 2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C.

103

Figure 4.30 (shown below) displays select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous monochloramine ([NH2Cl] =

100 μM) at full DOC (2.51 mg/L-L) capacity and pH 6.5. Chloroform formation increased with decreasing iopamidol concentration expect for 1.0 μM (lowest), dichloroiodomethane formation decreased with decreasing iopamidol concentrations.

CHCl3 50 TCAA CHCl2I

Concentration,nM 20

0 0.0 1.0 2.5 5.0 [Iopamidol], µM Figure 4.30: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 6.5; [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L.

104 Figure 4.31 (shown below) displays select DBP formation (chloroform,

dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous monochloramine ([NH2Cl] =

100 μM) at full DOC (2.51 mg/L-L) capacity and pH 7.5. Formation with respect to each

DBP maintained the same trend as previously mentioned (pH 6.5).

CHCl3 50 TCAA CHCl2I 40

Concentration,nM 20

0 0.0 1.0 2.5 5.0 [Iopamidol], µM Figure 4.31: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 7.5; [NH2 Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L -L.

105

Figure 4.32 (shown below) displays select DBP formation (chloroform, dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous monochloramine ([NH2Cl] =

100 μM) at full DOC (2.51 mg/L-L) capacity and pH 8.5. The aforementioned trend was once again observed with respect to each DBP.

CHCl3 50 TCAA

CHCl2I

Concentration,nM 20

0 0.0 1.0 2.5 5.0 [Iopamidol], µM Figure 4.32: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 8.5; [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L.

106

Figure 4.33 (shown below) displays select DBP formation (chloroform,

dichloroiodomethane and trichloroacetic acid) with respect to iopamidol concentration

(5.0 μM, 2.5 μM and 1.0 μM), in the presence of aqueous monochloramine ([NH2Cl] =

100 μM) at full DOC (2.51 mg/L-L) capacity and pH 9.0. Again, the same formation

trend was observed with respect to each DBP.

CHCl3 50 TCAA CHCl2I 40

Concentration,nM 20

0 0.0 1.0 2.5 5.0 [Iopamidol], µM Figure 4.33: Observed DBP formation at 72 hours as a function of iopamidol where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH) at pH 9.0; [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25 °C, [TOC] = 2.51 mg/L-L.

107

With respect to chlorine, iodo-DBP formation, specifically iodo-THM formation,

increased as pH increased (6.5, 7.5, 8.5 and 9.0). While the data may have exhibited an

increasing tread with respect pH, it was by no means proportional. From pH 6.5 to 7.5 a

drastic increase in formation (THMs and iodo-THMs) occurred, this is attributed to the

necessary and sufficient concentration of the OCl-, a strong nucleophile, which may be

responsible for the induction of iopamidol transformation (Duirk et al. 2011). Duirk et

al. (2011) theorized the strong nucleophile, OCl-, attacked the partially positive resonance structure of iopamidol. Hence why Duirk et al. (2011) originally proposed OCl- as being

responsible for the initial attack on one of the amide side chains of the iopamidol

molecule thus resulting in a primary amine transformation product. Removal of said

amide side chain, exposes electronegative iodine on the aromatic ring leaving said iodine

susceptible to hypochlorous acid; an electrophile, that will rapidly oxidize iodine from

the benzene ring to form HOI (Duirk et al. 2011).

Additionally, it should be noted that while this work supports the proposed theory as well

as the complimentary work of Kumkum (2013) and Ackerson (2014) it contradicts the

conventional iodide oxidation pathway (Bichsel and von Gunten, 2000; Bichsel and von

Gunten, 1999b). Nonetheless, the contradiction of the conventional pathway does not

reduce the validity of the proposed, arguable the opposite considering the increasing

support from recent work. Case in point, the work of Kumkum (2013) and Ackerson

(2014) determined the greatest reduction in iopamidol surrogate, total organic iodine

(TOI), at pH 7.5 and least at 9.5, which coincides with the speciation between

hypochlorous acid and hypochlorite ion given the associated pKa of 7.5 (speciation

108

depicted in Figure 4.34). This trend may be viewed for full TOC capacity (2.51 mg/L-L)

within Figure 4.1 (shown earlier) with respect to iodo-THM formation. Furthermore, this trend was maintained, arguably strengthened, with a reduction in TOC level (2.51, 1.26 and 0.63 mg/L-L), which may be viewed in Figures 4.1, 4.2 and 4.3 (shown earlier)

respective to decreasing TOC level. Additionally, Figure 4.16 (shown earlier) further supports aforementioned, strengthened trend in regard to dichloroiodomethane formation.

In previous work, the rapid oxidation of iodide to HOI in the presence of aqueous

chlorine has been investigated (Bichsel and von Gunten, 1999a; Nagy et al., 1988). In the presence of aqueous chlorine, HOI can disproportionate to iodate or iodide or to

iodine monoxide. However, disproportionation to iodine monoxide was determined to

relevant pathway in the fate of HOI given the excessively slow rate at which it occurs (1

– 10 μg/L HOI, pH 6 – 8, [CO3]T = 0 – 5 mM) (Hua et al., 2006; Bichsel and von Gunten,

1999a). Corresponding work performed by Nana Ackerson (2014) determined a

decreasing trend in iodate formation with respect to pH (lowest observed at pH 9.5) thus

suggesting the active oxidant in the formation of HOI is hypochlorous acid.

Furthermore, HOI can then be subsequently oxidized to iodate or be re-incorporated with

NOM to form iodo-DBPs (Bichsel and von Gunten, 1999a; Duirk et al. 2011; Hua et al.,

2006). Additionally, if hypochlorite was responsible for oxidation resulting in HOI then

the only a slight increase in iodate and/or iodo-DBP formation from pH 8.5 to 9.0 would

be exhibited due to the minor increase in hypochlorite ion abundance given the pKa (7.5).

Ackerson (2014) determined that iodate formation remained approximately the same for

pH 8.5 and 9.0, greatest at 7.5; this trend may be interrupted as hypochlorite being the 109

active oxidant. Nevertheless, if iodate remained approximately the same (at pH 8.5 and

9.0) and if hypochlorite was the active oxidant in HOI formation then one would expect

to observed approximately the same magnitude of iodo-DBP formation at pH 8.5 and 9.0.

In comparison to the work of Duirk et al. (2011), the pH range was extended in order to

include pH 9.0. Due to the addition and evaluation of pH 9.0, a second drastic increase

was observed with respect to THM and iodo-THM formation from pH 8.5 to 9.0, with a

focal point on CHCl3 and CHCl2I. The data expressed a considerable increase from pH

8.5 to 9.0 thus contradicting the expected behaviour if hypochlorite was in fact the active

oxidant. Additionally, to satisfy mass balance, the observed reduction in iodate

formation should be reflected in iodo-DBP formation. Ackerson (2014) determined the

reduction in iodate formation was in fact reflected in TOI; only a fraction of TOI is

representative of known iodo-DBPs while the majority remains unknown transformation

products. The considerable difference between units should be noted, iodate (mM) and

iodo-DBPs (nM). At pH 9.0, while the population of unknown transformation products is lessened, it still remains considerably high. Regardless, the magnitude of formation at pH 9.0 may be attributed to the interaction of hypochlorous acid, an electrophile, and the negatively charged, conjugate base of iopamidol. The proposed can be supported by the evaluation of speciation via the alphas, which are derived from the pKa and pH.

First and foremost, it should be noted, in regard to the speciation of iopamidol, there are

22 hydrogen atoms that can protonate thus 23 alphas exist. Within this work, only the first protonation will be considered given the increasing difficulty per protonation and

110

respective pH at which they would occur. The alphas (αo and α1) for both chlorine (pKa

7.5) and iopamidol (pKa 10.7) were derived as follows,

+ − + 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 ↔ 𝑂𝑂𝑂𝑂𝑂𝑂 𝐻𝐻 [ ] = [ ] + [ ] − − 𝐶𝐶𝐶𝐶2 𝑇𝑇 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 𝑂𝑂𝑂𝑂𝑂𝑂 [ ][ ] = [ − ]+ 𝑂𝑂𝑂𝑂𝑂𝑂 𝐻𝐻 𝐾𝐾𝑎𝑎 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 [ ] [ ] = = [ ] ++ [ ] 𝐻𝐻 𝐻𝐻𝐻𝐻𝐻𝐻𝐻𝐻 𝛼𝛼𝑜𝑜 + − 𝐻𝐻 𝐾𝐾𝑎𝑎 𝐶𝐶𝐶𝐶2 𝑇𝑇 [ ] = = [ ] + [ ]− 𝐾𝐾𝑎𝑎 𝑂𝑂𝑂𝑂𝑂𝑂 𝛼𝛼1 + − 𝐻𝐻 𝐾𝐾𝑎𝑎 𝐶𝐶𝐶𝐶2 𝑇𝑇 + − + 𝐶𝐶17𝐻𝐻22𝐼𝐼3𝑁𝑁3𝑂𝑂8 ↔ 𝐶𝐶17𝐻𝐻21𝐼𝐼3𝑁𝑁3𝑂𝑂8 𝐻𝐻 [ ] = [ ] + [ ] − 𝐶𝐶17𝐻𝐻22𝐼𝐼3𝑁𝑁3𝑂𝑂8 𝑇𝑇 𝐶𝐶17𝐻𝐻22𝐼𝐼3𝑁𝑁3𝑂𝑂8 𝐶𝐶17𝐻𝐻21𝐼𝐼3𝑁𝑁3𝑂𝑂8 [ ] [ ] = = [ ] ++ [ ] 𝐻𝐻 𝐶𝐶17𝐻𝐻22𝐼𝐼3𝑁𝑁3𝑂𝑂8 𝛼𝛼𝑜𝑜 + 𝐻𝐻 𝐾𝐾𝑎𝑎 𝐶𝐶17𝐻𝐻22𝐼𝐼3𝑁𝑁3𝑂𝑂8 𝑇𝑇 [ ] = = [ ] + [ −] 𝐾𝐾𝑎𝑎 𝐶𝐶17𝐻𝐻21𝐼𝐼3𝑁𝑁3𝑂𝑂8 𝛼𝛼1 + 𝐻𝐻 𝐾𝐾𝑎𝑎 𝐶𝐶17𝐻𝐻22𝐼𝐼3𝑁𝑁3𝑂𝑂8 𝑇𝑇 The calculated alphas were than graphed against pH to produce Figure 4.34 (shown on

consecutive page) where IP represents iopamidol and IP- represents the conjugate base of

iopamidol. The intersection of hypochlorous acid and the conjugate base of iopamidol

fell at pH 9.0, which may be correlated to larger than anticipated magnitude of iodo-DBP formation at said pH. To account for the considerable difference of concentration 111 between hypochlorous acid and iopamidol, given that hypochlorous acid is to be in great excess for DBP research, an additional speciation diagram was generated to properly weight speciation (Figure 4.35 – shown on consecutive page). The intersection of hypochlorous acid and the conjugate base of iopamidol then fell between pH 9.5 – 10.0

([iopamidol] = 5μM). In order to prove the proposed hypothesis, future work would need to expand the pH range to include 10.0 and 11.0. Given the weighted speciation diagram, the optimal pH for hypochlorous acid and the conjugate base of iopamidol should approximately be 10.0. Meaning if future work determined that formation decreased from 9.0 to 10.0 then the hypothesis would be rejected. Conversely, if future work determined an increase in formation from pH 9.0 to 10.0 and a reduction from 10.0 to

11.0 then the proposed hypothesis would be supported. Additionally, the utilization of chlorine within the reactor should be closely monitored. Excessive utilization of chlorine would further support proposed hypothesis. Within this work, a 20:1 ratio of aqueous chlorine to iopamidol was employed, which is a common ratio amongst DBP research.

This allows the oxidant to be in sufficient excess where it may be considered constant and the reaction can then be viewed as a pseudo first order reaction in lieu of a second order reaction. Aforementioned excessive use of chlorine, despite the initial excess

(20:1), would rule out the employment of pseudo first order reaction kinetics. It should be noted that drinking water criteria requires a pH within the range of 6.5 – 8.5 which is why Duirk et al. (2011) originally utilized said range. The pH range for this work was expanded past aforementioned range with a desire to increase the knowledge of

112 iopamidol’s role in iodo-DBP formation. Further expansion of the pH range for future work can be supported by the demand to obtain knowledge of said iodo-DBP formation.

Figure 4.34: Speciation of hypochlorous acid (HOCl/OCl-) and iopamidol (depicted as - IP/IP ), alphas (αo and α1) derived from pH and respective pKa.

113 Speciation - Concentration 0.00012

0.0001

)

μM 0.00008 αo - HOCl α1 - OCl- 0.00006 αo - IP α1 - IP- 0.00004 Concentration (

0.00002

0 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 pH

Figure 4.35: Speciation of hypochlorous acid (HOCl/OCl-) and iopamidol (depicted as - - IP/IP ), alphas (αo and α1) derived from pH and respective pKa, [Cl2 ]T = 100 μM, [iopamidol] = 5 μM.

In regard to varied DOC levels of source water via dilution, when DOC was reduced

from full capacity (2.51 mg/L-L) to half (1.26 mg/L-L) dichloroiodomethane (CHCl2I)

formation (Figure 4.16 – shown earlier); remained equivalent at pH 6.5, exhibited a slight

increase at pH 7.5 and 8.5 (8.5 > 7.5) and doubled at pH 9.0 (approximately 45.96 nM

increase). When DOC capacity was further reduced (0.63 mg/L-L), CHCl2I exhibited almost identical formation to that of half DOC capacity (1.26 mg/L-L) (approximately

6.95 nM increase). Trichloroacetic acid increased from full (2.51 mg/L-L) to half (1.26 mg/L-L) DOC capacity for all pHs (9.0< 6.5< 7.5< 8.5) and decreased from half (1.26

114

mg/L-L) to quarter (0.63 mg/L-L) DOC capacity for pH 6.5, 8.5 and 9.0(6.5< 8.5< 9.0)

and increased at pH 7.5. In regard to chloroform (CHCl3) formation upon reduction of

DOC (2.51 to 1.26 mg/L-L), formation was reduced by approximately half for pH6.5,

slight decrease in pH 7.5 and 8.5 (8.5 > 7.5) and increased considerably at pH 9.0

(approximately 159.9 nM increase). When DOC was further reduced from half (1.26 mg/L-L) to quarter (0.63 mg/L-L), CHCl3 exhibited a similar trend in which reduction at

pH 6.5 was observed, slight increase at pH 7.5, slight reduction at pH 8.5 and considerable increase at pH 9.0 (approximately 106.6 nM increase). It should be noted that the trend observed at pH 9.0 drastically differs from the trend observed at other pHs

(6.5, 7.5 and 8.5) with respect to chloroform formation. Given the initial competition between iopamidol and DOC, a reduction in DOC should give away to proportionally increased formation of iodo-DBPs, which proved accurate for full to half capacity DOC reduction (2.51 to 1.26 mg/L-L). Conversely, further reduction of DOC (1.26 to 0.63 mg/L-L) failed to produce additional iodo-DBP formation. One could argue that necessary concentration of DOC required to induce the observed formation would be that of quarter capacity (0.63 mg/L-L). Thus suggesting an alternative limiting reactant within half capacity (1.26 mg/L-L), potentially iopamidol, meaning an excess of DOC. While

DOC capacity was reduced, iopamidol and chlorine concentrations remained constant

([iopamidol]=5μM, [Cl2-] = 100μM). Reduction of DOC, decreased competition

(Iopamidol and DOC) enough to allow the reaction between iopamidol and aqueous chlorine to proceed to full capacity thus explaining the virtually identical formation at half and quarter DOC capacity for chlorine ([CHCl2I] = 103.5 nM at half DOC pH 9.0

115

and [CHCl2I] = 110.5 nM at quarter DOC pH 9.0). This trend was also apparent within

the monochloramine data set ([CHCl2I] = 20.0 nM at half DOC pH 9.0 and [CHCl2I] =

19.5 nM at quarter DOC pH 9.0). Upon reduction of DOC, formation at pH 9.0 doubled

while the remaining pHs exhibited a slight increase, additionally supporting the

interaction between hypochlorous acid and the conjugate base of iopamidol. This can be

further investigated by reduction of DOC to that of half or quarter coupled with varied

iopamidol concentrations (2.5 and 1.0 μM). Iodo-THM, specifically dichloroiodomethane (CHCl2I) formation should decrease at half and quarter DOC when iopamidol concentration is decreased but remain equivalent respective to half and quarter.

Meaning the optimal DOC conditions for iodo-DBP, respective to Cleveland source water, formation lie between half (1.26 mg/L-L) and quarter (0.63 mg/L-L) capacity of

DOC. It should be noted that complementary work performed by Ackerson (2014) in addition to Duirk et al. (2011) reported trace levels of iodo-DBP formation in the absence of DOC.

Within this work, the maximum dichloroiodomethane formation observed was 110.5 nM

(DOC = 0.63 mg/L-L, pH 9.0), while Duirk et al. (2011) observed 212 nM (DOC = 2.1 mg/L-L, pH 8.5). The difference in formation at drastically different DOC levels may be attributed to the percent volume of fulvic vs humic acid. Duirk et al. (2011) determined that iodo-DBP formation is larger in the presence of humic acid in lieu of fulvic acid.

Given the difference in iodo-DBP formation, one could argue that the source water used within this work contained less humic acid and more fulvic acid in comparison. Fulvic acid may be more competitive with iopamidol hence the lower formation when present

116

and increased formation when humic is present. Additionally, SUVA254 for the source

water used within this work was 1.17 L/m-mg and 4.88 L/m-mg for the source water used

by Duirk et al. (2011). DBP formation has been linked to SUVA254 as well as DOC

concentration (Njam et al., 1994). It should also be noted that the analytical equipment

and methods differed between this work and that of Duirk et al. (2011). This work utilized gas chromatography coupled with 63Ni microelectron capture detector (μECD)

(as described in chapter 3) while Duirk et al. (2011) utilized gas chromatography/electron

ionization coupled with mass spectrometry (MS) in selected ion monitoring (SIM) mode.

With respect to the monochloramine data set, while an increased variety of iodo-DBPs formed, the magnitude of said formation was considerably less than what was observed within the chlorine data set. The highest formation was observed at pH 6.5 and the lowest at pH 9.0 (Figure 4.18, 4.19 and 4.20 – shown earlier). Formation also decreased as DOC level (2.51, 1.26 and 0.63 mg/L-L) decreased (Figure 4.26 – shown earlier). Both aforementioned trends differed from that exhibited by chlorine. Complementary work performed by Nana Ackerson (2014) determined no significant degradation of iopamidol in the presence of monochloramine over 168-hour time frame. As previously stated, degradation of iopamidol was tracked via surrogate (TOI). Monochloramine has been documented to react with iopamidol to form iodo-DBPs in the presence of NOM (Duirk et al., 2011). Meaning the iodide on the aromatic ring of iopamidol may be oxidized to

HOI (Bichsel and von Gunten, 1999b). HOI has shown to be stable in the presence of monochloramine hence the lack of formation of iodate found by Ackerson (2014)(Bichsel and von Gunten, 1999a). The trends observed within this work reinforced the findings of

117

Duirk et al. (2011) considering the virtually identical observed trends. It should be noted, the magnitude of observed iodo-DBP formation was higher within this work.

4.3 Statistical Analysis

The objective of this statistical analysis was to provide insight for future work and aid in future determination of a statistical model and experimental design to strengthen the validity of the proposed. Within in this work, triplet analysis was utilized which is representative of sub-sampling in regard to statistical analysis of data. Given the sample size, the sub-sampling values (S = 3) were averaged and presented with corresponding

95% confidence intervals, this value will be taken to represent the value of the respective response variable (Y) in a randomized complete block design (RCBD). This approach assumes no interaction between blocking factors, which can be verified by interaction plots. The reduced power of the statistical test is accepted due the economical efficiency of this approach considering experimental cost. If statistical significance can be determined given aforementioned conditions, future work would be greatly supported thus likely to be funded.

As briefly mentioned, a randomized complete block design (RCBD) was utilized where the null hypothesis taken was that of iopamidol exhibiting no effect (Ho = 0), which is represent by the following statistical model,

= + + +

𝑌𝑌𝑖𝑖𝑖𝑖 𝜇𝜇 𝜏𝜏𝑖𝑖 𝛽𝛽𝑗𝑗 𝜀𝜀𝑖𝑖𝑖𝑖

118 Where Yij represents the observed values for treatment i and block j, μ represents the unknown overall mean, τi represents the treatment effect, βj represents the block effect

and εij represents the error term for treatment i, block j. The statistical objective is to

estimate the unknown overall mean by the derived data set,

𝜇𝜇 𝑌𝑌

119

𝐸𝐸�𝑌𝑌𝑖𝑖𝑖𝑖� 𝑌𝑌�𝚤𝚤𝚤𝚤 =

𝑌𝑌�𝚤𝚤𝚤𝚤 𝑌𝑌

120 significance in regard to pH (p-value of 0.0052) and not DOC. Continuing the utilization

of chloroform as the response variable (Table 4.1), varied iopamidol expressed

significance for both iopamidol (p-value of < 0.0001) and pH (p-value of 0.0014) with respect to monochloramine. In regard to chlorine (Table 4.1), significance was only determined for pH (p-value of < 0.0001) and not for iopamidol. The lack of significance may be attributed to interaction, thorough discussion to follow.

Table 4.1: p-values for two-way ANOVA (two factors) where α = 0.05 (95% significance) with respect each oxidant where the response variable (Y) was chloroform.

Factors Chlorine Monochloramine

IP 0.1334 < 0.0001

pH < 0.0001 0.0014

DOC 0.7078 0.0004

pH 0.0052 0.0325

With respect to dichloroiodomethane acting as the response variable (Table 4.2 – shown

on consecutive page) amongst varied DOC, significance was determined in regard to pH

for both chlorine (p-value of 0.0009) and monochloramine (p-value of 0.0012), while

significance was not determined for DOC for both oxidants. When dichloroiodomethane

was utilized as the response variable (Table 4.2), coupled with varied iopamidol

concentration, significance was found for iopamidol for chlorine (p-value of 0.0064) and

121 pH for monochloramine (p-value of 0.0029), while significance was not found for pH

respective to chlorine and iopamidol respective for monochloramine. Again, the lack of

significance may be attributed to interaction given the experimental design.

Table 4.2: p-values for two-way ANOVA (two factors) where α = 0.05 (95% significance) with respect each oxidant where the response variable (Y) was dichloroiodomethane. Factors Chlorine Monochloramine

IP 0.0064 0.096

pH 0.0712 0.0029

DOC 0.0903 0.1009

pH 0.0009 0.0012

While the findings of said statistical analysis were in most cases significant, considering

the interaction plots (appendix: Figure A1 – A8) and the lack of parallelism expressed,

one can assume interaction between independent variables (i.e. factors). While it cannot

be determined if the lack of parallelism is due to randomness or interaction, past work

supports the assumption of interaction over randomness (Duirk et al., 2011).

Additionally, the magnitude of interaction displayed in said interaction plots given the

aforementioned assumption cannot be quantified, however, it appears to be present in

varying degree (i.e. some plots possess what appears to be close to parallelism while

other do not). Regardless, it is required to place an interaction term into the statistical

122 model to account for said interaction between factors. Thus altering the experimental design to that of a generalized randomized complete block design (GRCBD) with sub- sampling, which will require an increase in sample size. A recommendation for future work would be to increase the sample size by 1 while mimicking the current sub- sampling (S = 3). This would vastly increase the power of the statistical test and strengthen the validity of conclusions drawn from ANOVA tables and interaction plots while minimizing the cost to investigator. Furthermore, upon completion or in lieu of the aforementioned recommendation given finical accessibility, a factorial design may be necessary in order to properly cross multiple independent variables to profoundly evaluate interaction. This would require observations at every combination of independent variables (factors) and levels of said variables; derived data would then be coupled with response surface methodology for analysis. Although it should be noted the cost to perform this work would greatly increase in comparison the cost of this work or that of the work to increase the sample size by 1.

A full factorial design would allow the investigation of each independent variable (factor) on the response variable (i.e. CHCl3 or CHCl2I formation), it will also allow the investigation of the effects on the response variable due to interaction between independent variables (factors). Given the four independent variables (factors) within this work and the levels associated to each said independent variable (factor), a full factorial design is extremely financially unfeasible. A fractional factorial design has the potential to make the needed assessment of interaction between factors with a more feasible experimental cost. A fractional factorial design would reduce the number of

123

observation by meticulous selection to omit the unnecessary. This design exploits the

sparsity-of-effect principle, which states most systems are dominated by main effects and

low-order interaction while high-order interaction can be neglected; main effect being

that of the effect of independent variables (factors) on the dependant variable (response

variable)(D. Montgomery, 2013). An additional recommendation to increase the power

of the statistical test and properly represent interaction would be the completion of

factional factorial design upon determination of the proper sub-set of observations and proper omissions and coupled with response surface methodology for analysis. However, this approach will still require an increase in the sample size and potentially additional experiments to expand on current observations thus increasing the fiscal and time commitment.

Additionally, a more readily available, timely and fiscally sound recommendation for future work would be the execution of an unreplicated factorial design. This would require the assumption that high-order interactions are negligible thus allowing combination their respective mean squares to estimate error. This approach would also utilize the sparsity-of-effect principle. This recommendation would require a limited number of experiments to expand on the current set of observations thus accounting for all combinations of factors and respective levels of said factors, followed by proper statistical analysis.

124

CHAPTER V

CONCLUSIONS AND RECOMMENDATIONS

5.1 Introduction

Iopamidol was investigated in source water obtained from the Cleveland water treatment

plant (Cleveland, OH). Transformation of iopamidol in the presence of a chlorinated oxidant and respective contribution to the formation of iodinated disinfection by- products, with respect to pH, total organic carbon (TOC) and iopamidol concentration was evaluated. The data was obtained from experiments in which iopamidol was introduced into a batch reactor containing source water (Cleveland, OH), amongst a buffer and internal standard, at varying concentrations (5.0, 2.5, 1.0 and 0.0 μM) over the pH range of 6.5 – 9.0 (6.5, 7.5, 8.5 and 9.0). Additionally, iopamidol concentration was held constant (5.0 μM) while TOC was varied via dilution with dionized water (2.51,

1.26 and 0.63 mg/L-L) over the pH range of 6.5 – 9.0 (6.5, 7.5, 8.5 and 9.0). Source water experiments were performed in the absence of iopamidol, where the respective chlorinated oxidant, buffer and internal standard were present to act as a control over the pH range of 6.5 – 9.0 (6.5, 7.5, 8.5 and 9.0). The chlorinated oxidants that were evaluated were aqueous chlorine and monochloramine. A statistical analysis was performed on derived data set (two-way ANOVA and interaction plots) with the

125 objective to obtain significance of respective factors (iopamidol, pH and DOC) were the

response variables were chloroform (CHCl3) and dichloroiodomethane (CHCl2I). It should be noted that the aforementioned research objective have been met and recommendations were given for future work.

5.2 Conclusions

1. Iodinated trihalomethane (iodo-THM) formation was highest at pH 9.0 for

chlorine and at pH 6.5 for monochloramine. A considerable increase in formation

was observed from pH 6.5 to 7.5 and from 8.5 to 9.0 with respect to chlorine as

oxidant. Monochloramine expressed a decreasing trend from pH 6.5 to 9.0. The

most predominately formed iodo-THM was dichloroiodomethane, 57.5 nM for

chlorine at pH 9.0 (DOC = 2.51 mg/L-L) and 35.2 nM for monochloramine at pH

6.5 (DOC = 2.51 mg/L-L). Chloroform formation was also impacted by the

introduction of iopamidol to reactor. Monochloramine produced a wider variety

of iodo-DBPs at lower concentrations in comparison to chlorine.

2. From pH 6.5 to 7.5 a drastic increase in formation (THMs and iodo-THMs)

occurred, this is attributed to the necessary and sufficient concentration of the

hypochlorite ion (OCl-), a strong nucleophile, which induces the transformation of

iopamidol. Supporting hypochlorite (OCl-) as being responsible for the initial

attack on one of the amide side chains of the iopamidol molecule thus resulting in

a primary amine transformation product. In theory, removal of said amide side

126

chain, exposes electronegative iodine on the aromatic ring leaving said iodine

susceptible to hypochlorous acid; an electrophile, that will rapidly oxidize iodine

from the benzene ring to form hypoiodous acid (HOI).

3. The magnitude of formation at pH 9.0 may be attributed to the interaction of

hypochlorous acid, an electrophile, and the conjugate base of iopamidol. The

calculated alphas were than graphed against pH, the intersection of hypochlorous

acid and the conjugate base of iopamidol fell at pH 9.0 – 9.5, which may be

correlated to larger than anticipated magnitude of formation at said pH. Speciation

diagram placed the intersection of hypochlorous acid and the conjugate base of

- iopamidol between pH 9.5 – 10.0 ([iopamidol] = 5 μM and [Cl2 ]T = 100 μM).

Upon reduction of DOC, formation at pH 9.0 doubled while the remaining pHs

exhibited a slight increase, additionally supporting the interaction between

hypochlorous acid and the conjugate base of iopamidol.

4. When NOM levels were reduced to a half of the original capacity (2.51 – 1.26

mg/L-L), increased formation was observed (approximately doubled at pH 9.0).

Further reduction of DOC (1.26 mg/L-L to 0.63 mg/L-L) failed to produce

additional iodo-DBPs. Suggesting that the necessary concentration of DOC

required to induce the observed formation would be that of quarter capacity (0.63

mg/L-L). While DOC capacity was reduced, iopamidol and chlorine

concentrations remained constant ([iopamidol]=5μM, [Cl2-] = 100μM).

Reduction of DOC, decreased competition (Iopamidol and DOC) enough to allow

the reaction between iopamidol and aqueous chlorine to proceed until the

127

exhaustion of iopamidol thus explaining the virtually identical formation at half

and quarter DOC capacity for chlorine ([CHCl2I] = 103.5 at half DOC pH 9.0 and

[CHCl2I] = 110.5 at quarter DOC pH 9.0). Meaning the optimal DOC conditions

for iodo-DBP formation, respective to Cleveland source water, lies at quarter

(0.63 mg/L-L) capacity of DOC.

5. Two-way analysis of variance (two way-ANOVA) tables were generated for two

response variables, chloroform and dichloroiodomethane. With respect to each

oxidant, iopamidol and pH were evaluated while NOM level remained constant

and NOM and pH were evaluated while iopamidol remained constant. In regard

to chloroform as the response variable and varied DOC, significance was yielded

for both DOC (p-value of 0.0004) and pH (p-value of 0.0325) with respect to

monochloramine: whereas chlorine only exhibited significance in regard to pH (p-

value of 0.0052). Varied iopamidol expressed significance for both iopamidol (p-

value of < 0.0001) and pH (p-value of 0.0014) with respect to monochloramine.

In regard to chlorine, significance was only determined for pH (p-value of <

0.0001). With respect to dichloroiodomethane acting as the response variable

amongst varied DOC, significance was determined in regard to pH for both

chlorine (p-value of 0.0009) and monochloramine (p-value of 0.0012). Varied

iopamidol concentration, significance was found for iopamidol for chlorine (p-

value of 0.0064) and pH for monochloramine (p-value of 0.0029). While the

findings of said statistical analysis were in most cases significant, considering the

interaction plots and the lack of parallelism expressed, one can assume interaction

128

between independent variables (i.e. factors). While it cannot be determined if the

lack of parallelism is due to randomness or interaction, past work supports the

assumption of interaction over randomness.

5.3 Recommendations

1. The role of DOC and proposed hypothesis can be further investigated by

reduction of DOC to that of half and quarter capacity coupled with varied

iopamidol concentrations (2.5 and 1.0 μM). This would also complete factor

combinations to strengthen statistical analysis, allowing analysis to be performed

as an unreplicated full factorial. Iodo-THM, specifically dichloroiodomethane

(CHCl2I) formation should decrease at half and quarter DOC when iopamidol

concentration is decreased. Furthermore, formation should remain approximately

equivalent respective to half and quarter with respect to iopamidol concentration

reduction. Optimal DOC conditions for iodo-THM formation, respective to

Cleveland source water, should lie between half (1.26 mg/L-L) and quarter (0.63

mg/L-L) capacity of DOC, favouring quarter.

2. In order to prove the proposed hypothesis of the reaction between hypochlorous

acid and the conjugate base of iopamidol, future work would need to expand the

pH range to include 10.0 and 11.0. Given the weighted speciation diagram, the

optimal pH for hypochlorous acid and the conjugate base of iopamidol should

approximately be 10.0. Meaning if future work determined that formation

decreased from 9.0 to 10.0 then the hypothesis would be rejected. Conversely, if

129

future work determined an increase in formation from pH 9.0 to 10.0 and a

reduction from 10.0 to 11.0 then the proposed hypothesis would be supported.

Additionally, the utilization of chlorine within the reactor should be closely

monitored. Excessive utilization of chlorine would further support proposed

hypothesis, ruling out the ability to assess said reaction as pseudo first order.

3. In regard to statistical analysis and experimental design, a factorial design may be

necessary in order to properly cross multiple independent variables to profoundly

evaluate interaction. An additional recommendation to increase the power of the

statistical test and properly represent interaction would be the completion of

factional factorial design upon determination of the proper sub-set of observations

and proper omissions and coupled with response surface methodology for

analysis.

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APPENDIX

Table A1: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH);

[Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 478.3 ± 53.9 530.1 ± 204.2 594.7 ± 28.2 769.3 ± 42.4

CHCl2Br 59.7 ± 17.7 70.2 ± 12.1 74.1 ± 4.3 68.8 ± 4.7

CHClBr2 5.5± 1.1 6.1 ± 0.2 8.7 ± 0.2 9.2 ± 1.2

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 7.4 ± 6.0 4.7 ± 2.0 3.0 ± 0.6 3.0 ± 0.7 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs CAA 3056.8 ± 884.1 3451.6 ± 223.7 4302.6 ± 684.4 4384.1 ± 694.7 DCAA 428.0 ± 36.9 1094.1 ± 186.1 694.9 ± 76.5 966.4 ± 152.1 TCAA 55.5 ± 16.4 122.5 ± 18.0 45.1 ± 0.8 82.2 ± 12.0 BAA 739.2 ± 204.1 566.4 ± 152.1 152 ± 39.2 38.1 ± 3.4 BCAA 23.2 ± 18.0 36.9 ± 21.8 28.9 ± 27.7 45.2 ± 29.9

146 Table A2: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland,

OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 6.4 ± 5.7 25.0 ± 18.0 36.2 ± 19.2 57.5 ± 8.2

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5

CHBrI2 < 0.5 < 0.5 < 0.5 2.8 ± 0.8

CHI3 < 0.5 < 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 11.5 ± 0.9

147 Table A3: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland,

OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L. pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 31.3 ± 3.5 27.3± 3.3 16.5 ± 2.5 19.1 ± 6.0

CHCl2Br 12.8 ± 1.7 4.8 ± 0.3 0.8 ± 9.0E-02 0.9 ± 0.1

CHClBr2 1.5 ± 0.2 < 1 < 1 < 1

CHBr3 1.2 ± 0.1 < 1 < 1 < 1 HANs CAN 8.9 ± 0.2 7.4 ± 1.8 9.2 ± 6.0E-02 6.6 ± 1.3 TCAN < 1 < 1 < 1 < 1 BAN 1.9 ± 0.2 1.5 ± 0.5 2.0 ± 0.3 1.2 ± 0.4 HAAs CAA 29.8 ± 2.8 5.5 ± 0.7 2.3 ± 0.2 1.7 ± 0.2 DCAA 7.2 ± 10.7 9.4 ± 9.8 7.3 ± 1.0 11.8 ± 0.7 TCAA < 1 < 1 < 1 < 1 BCAA < 1 < 1 < 1 < 1

Table A4: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L. pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 35.2 ± 4.2 22.7 ± 1.1 2.0 ± 0.3 1.7 ± 0.1

CHClI2 17.4 ± 2.5 9.0 ± 1.1 < 0.5 < 0.5 CHBrClI 6.2 ± 0.8 1.7 ± 0.2 < 0.5 < 0.5

CHBr2I 0.9 ± 0.1 < 0.5 < 0.5 < 0.5

CHBrI2 4.4 ± 0.7 1.2 ± 7.0E-02 < 0.5 < 0.5

CHI3 11.3 ± 2.2 3.1 ± 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

148

Table A5: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH);

[Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 260.0 ± 7.1 478.7 ± 34.5 447.3 ± 26.9 929.2 ± 44.2

CHCl2Br 22.2 ± 3.8 134.4 ± 17.21 269.9 ± 78.3 302.3 ± 61.6

CHClBr2 6.5 ± 7.4 3.3 ± 0.3 4.9 ± 0.3 5.4 ± 8.0E-02 HANs CAN 2.6 ± 4.1 2.1 ± 0.8 < 1 3.5 ± 0.4 HAAs CAA 1339.0 ± 120.9 1910.3 ± 59.5 3099.5 ± 915.0 2832.7 ± 454.1 DCAA 667.9 ± 68.7 1645.7 ± 34.3 1109.2 ± 142.9 742.2 ± 184.7 TCAA 248.1 ± 15.9 427.6 ± 21.5 396.4 ± 35.9 215.4 ± 28.2 BCAA 26.6 ± 4.9 33.2 ± 6.6 39.2 ± 4.0 35.5 ± 4.8

Table A6: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland,

OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 6.8 ± 2.8 39.7 ± 8.4 55.5 ± 3.1 103.5 ± 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

149

Table A7: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland,

OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 13.6 ± 0.5 10.3 ± 0.8 7.0 ± 0.8 6.6 ± 0.6

CHCl2Br 3.6 ± 0.8 1.1 ± 0.1 < 1 < 1

CHClBr2 < 1 < 1 < 1 < 1

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 6.6 ± 0.6 6.7 ± 1.3 6.0 ± 0.2 6.6 ± 0.2 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs CAA 10.3 ± 1.6 3.8 ± 0.1 < 1 < 1 DCAA 9.6 ± 1.4 8.0 ± 0.3 6.2 ± 1.9 5.1 ± 1.0 TCAA < 1 < 1 < 1 < 1 BCAA < 1 < 1 < 1 < 1

Table A8: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 20.0 ± 0.4 11.9 ± 0.3 1.0 ± 0.4 0.8 ± 0.6

CHClI2 10.9 ± 0.4 6.6 ± 0.7 < 0.5 < 0.5 CHBrClI 2.2 ± 0.1 0.6 ± 6.0E-02 < 0.5 < 0.5

CHBr2I < 0.5 < 0.5 < 0.5 < 0.5

CHBrI2 1.9 ± 0.1 < 0.5 < 0.5 < 0.5

CHI3 12.3 ± 0.2 3.7 ± 1.0 < 5 < 5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA 0.6 ± 0.3 < 0.5 < 0.5 < 0.5

150

Table A9: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH);

[Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L.

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 222.0 ± 15.1 486.0 ± 59.3 370.2 ± 23.5 1035.8 ± 27.1

CHCl2Br 15.7 ± 1.5 111.4 ± 8.7 265.2 ± 12.0 291.9 ± 77.4 HANs CAN 5.1 ± 1.4 3.3 ± 1.8 2.0 ± 1.6 < 1 HAAs CAA 690.8 ± 39.8 1087.5 ± 136.0 1969.8 ± 37.9 1715.5 ± 377.4 DCAA 627.2 ± 23.8 2027.2 ± 140.5 1157.9 ± 100.1 889.3 ± 12.4 TCAA 204.4 ± 35.5 462.7 ± 12.0 252.6 ± 63.6 33.8 ± 2.0 BCAA 13.5 ± 4.0 3.6 ± 0.9 < 1 < 1

Table A10: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland,

OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 6.8 ± 0.9 36.2 ± 9.2 54.2 ± 3.6 110.5 ± 11.8 HAAs IAA 15.2 ± 5.3 < 0.5 < 0.5 < 0.5

151

Table A11: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 10.5 ± 0.9 9.0 ± 0.6 7.4 ± 1.7 6.9 ± 0.7

CHCl2Br 2.0 ± 1.1 < 1 < 1 < 1

CHClBr2 < 1 < 1 < 1 < 1

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 6.8 ± 0.4 7.1 ± 0.4 7.9 ± 1.1 7.2 ± 0.3 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs CAA 4.7 ± 2.2 2.4 ± 0.3 < 1 < 1 DCAA 7.0 ± 1.6 7.5 ± 1.4 17.5 ± 24.5 5.9 ± 2.2 TCAA < 1 < 1 1.79 ± 2.6 1.0 ± 0.7 BCAA < 1 < 1 < 1 < 1

Table A12: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 19.5 ± 1.5 12.8 ± 0.4 1.5 ± 0.2 1.2 ± 0.72

CHClI2 10.3 ± 1.0 9.1 ± 0.5 < 0.5 < 0.5 CHBrClI 1.3 ± 0.2 < 0.5 < 0.5 < 0.5

CHBr2I < 0.5 < 0.5 < 0.5 < 0.5

CHBrI2 1.3 ± 0.2 < 0.5 < 0.5 < 0.5

CHI3 19.5 ± 2.8 10.0 ± 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

152

Table A13: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland,

OH); [Iopamidol] = 2.5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 343.7 ± 14.2 565.7 ± 56.9 573.5 ± 6.0 652.1 ± 38.6

CHCl2Br 64.4 ± 3.4 82.5 ± 10.9 84.4 ± 4.8 83.7 ± 10.5

CHClBr2 7.0 ± 0.6 9.2 ± 1.2 10.7 ± 0.4 11.9 ± 1.7

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 6.5 ± 5.1 5.9 ± 3.1 4.1 ± 0.3 3.2 ± 0.2 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs CAA 3114.4 ± 804.2 3800.5 ± 391.1 4413.2 ± 432.8 3947.2 ± 72.9 DCAA 745.9 ± 107.1 2097.1 ± 123.2 900.7 ± 20.4 723.9 ± 33.9 TCAA 322.8 ± 1.1 363.1 ± 78.9 161.4 ± 20.6 136.7 ± 20.8 BCAA 51.5 ± 1.8 113.4 ± 15.6 61.9 ± 7.1 70.4 ± 28.1

Table A14: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland,

OH); [Iopamidol] = 2.5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 1.7 ± 0.4 4.4 ± 0.5 7.3 ± 0.3 10.4 ± 1.2

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5

CHBr2I 0.6 ± 4.0E-02 < 0.5 < 0.5 < 0.5

CHI3 < 0.5 < 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

153

Table A15: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 2.5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 41.5 ± 2.2 29.9 ± 5.2 22.9 ± 1.3 21.8 ± 3.2

CHCl2Br 19.6 ± 0.7 5.7 ± 2.9 1.5 ± 1.2 1.8 ± 1.1

CHClBr2 3.6 ± 6.0E-02 < 1 < 1 < 1

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 2.2 ± 0.2 1.2 ± 0.1 1.2 ± 1.0E-02 1.3 ± 0.2 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 DBAN < 1 < 1 < 1 < 1 HAAs CAA 40.2 ± 2.1 13.4 ± 0.4 3.1 ± 1.5 0.7 ± 1.3 DCAA 15.9 ± 2.0 14.2 ± 2.6 3.6 ± 1.3 1.5 ± 2.9 TCAA < 1 < 1 < 1 < 1 BCAA 2.0 ± 0.7 < 1 < 1 < 1

154

Table A16: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 2.5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 23.0 ± 0.8 10.1 ± 0.5 < 0.5 < 0.5

CHClI2 5.2 ± 0.3 1.5 ± 0.1 < 0.5 < 0.5 CHBrClI 2.1 ± 1.3 < 0.5 < 0.5 < 0.5

CHBr2I 1.2 ± 2.0E-02 < 0.5 < 0.5 < 0.5

CHBrI2 1.7 ± 0.1 < 0.5 < 0.5 < 0.5

CHI3 1.2 ± 0.1 < 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

155

Table A17: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland,

OH); [Iopamidol] = 1.0 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 367.7 ± 4.5 513.6 ± 105.8 629.6 ± 63.8 746.5 ± 150.3

CHCl2Br 91.0 ± 1.9 104.1 ± 20.5 114.5 ± 13.1 129.1 ± 30.2

CHClBr2 10.6 ± 0.2 12.3 ± 2.4 16.8 ± 1.9 21.4 ± 5.7

CHBr3 < 1 < 1 < 1 1.2 ± 0.3 HANs CAN 2.5 ± 0.2 5.2 ± 4.0 2.3 ± 0.9 1.8 ± 0.5 TCAN 2.2 ± 0.3 1.2 ± 0.1 < 1 BAN < 1 < 1 < 1 < 1 HAAs CAA 3802.7 ± 1573.0 3724.0 ± 581.1 3624.4 ± 304.6 3439.0 ± 446.0 DCAA 442.3 ± 26.8 860.3 ± 60.2 670.6 ± 27.6 386.7 ± 2.0 TCAA 132.5 ± 20.1 148.5 ± 30.2 71.6 ± 3.8 58.2 ± 20.6 BAA 75.4 ± 147.9 < 1 < 1 < 1 BCAA 49.4 ± 4.8 81.3 ± 14.0 76.7 ± 30.6 48.8 ± 3.5

Table A18: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland,

OH); [Iopamidol] = 1.0 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I < 0.5 < 0.5 < 0.5 2.9 ± 1.7

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5

CHBrI2 < 0.5 < 0.5 < 0.5 < 0.5

CHI3 < 0.5 < 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA 5.9 ± 0.8 4.9 ± 0.6 3.6 ± 0.7 < 0.5

156

Table A19: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 1.0 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 16.0 ± 2.4 11.1 ± 0.6 8.2 ± 1.2 8.0 ± 0.7

CHCl2Br 8.2 ± 0.9 1.98 ± 0.2 < 1 < 1

CHClBr2 1.7 ± 0.5 < 1 < 1 < 1

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 3.2 ± 2.1 3.0 ± 0.2 2.8 ± 0.1 2.8 ± 0.1 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 BCAN 1.4 ± 1.8 < 1 < 1 < 1 HAAs CAA 39.1 ± 3.0 10.6 ± 0.6 1.9 ± 0.1 1.6 ± 0.3 DCAA 14.1 ± 7.4 12.6 ± 4.1 7.5 ± 2.8 10.0 ± 3.1 TCAA < 1 < 1 < 1 < 1 BCAA 1.7 ± 1.4 < 1 < 1 < 1

Table A20: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water

(Cleveland, OH); [Iopamidol] = 1.0 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 11.4 ± 3.1 3.0 ± 0.2 0.8 ± 0.1 0.7 ± 0.1

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5 CHBrClI < 0.5 < 0.5 < 0.5 < 0.5

CHBr2I 1.0 ± 1.4 < 0.5 < 0.5 < 0.5

CHBrI2 < 0.5 < 0.5 < 0.5 < 0.5

CHI3 < 0.5 < 0.5 < 0.5 < 0.5 HANs IAN 5.2 ± 8.3 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

157 Table A21: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 374.2 ± 107.8 443.4 ± 50.2 521.9 ± 5.7 716.3 ± 87.5

CHCl2Br 80.0 ± 6.1 82.4 ± 9.0 90.3 ± 0.7 117.1 ± 14.5

CHClBr2 7.0 ± 0.8 9.2 ± 0.9 11.2 ± 0.3 18.4 ± 2.6

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 7.4 ± 0.4 4.3 ± 1.0 3.7 ± 0.2 4.3 ± 1.3 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 BCAN HAAs CAA 1984.3 ± 65.0 2086.8 ± 384.2 2579.9 ± 75.5 3003.8 ± 120.0 DCAA 383.0 ± 135.3 541.3 ± 45.5 341.6 ± 96.6 274.9 ± 35.6 TCAA 24.1 ± 5.4 120.7 ± 5.6 18.3 ± 3.8 15.3 ± 9.1 BAA 46.4 ± 1.6 19.8 ± 7.6 9.6 ± 0.7 8.3 ± 1.8 BCAA 15.2 ± 4.4 39.1 ± 5.0 27.4 ± 8.7 23.5 ± 3.7

Table A22: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs

CHCl2I 2.8 ± 0.4 < 0.5 < 0.5 22.1 ± 2.7

CHClI2 < 0.5 < 0.5 < 0.5 0.9 ± 0.1 CHBrClI < 0.5 < 0.5 < 0.5 1.8 ± 0.2

CHI3 < 0.5 < 0.5 < 0.5 < 0.5 HANs

IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs

IAA < 0.5 < 0.5 < 0.5 2.6 ± 0.2

158

Table A23: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs

CHCl3 50.0 ± 4.1 53.8 ± 0.6 43.5 ± 10.5 38.0 ± 7.3 CHCl2Br 18.2 ± 6.6 9.0 ± 1.6 4.0 ± 2.2 2.5 ± 0.7

CHClBr2 2.0 ± 0.2 1.3 ± 0.4 1.1 ± 0.3 < 1 CHBr3 < 1 < 1 < 1 < 1 HANs CAN 5.5 ± 2.1 6.7 ± 1.8 6.0 ± 1.9 5.8 ± 1.6 TCAN < 1 1.1 ± 0.7 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs

DCAA 55.2 ± 26.1 65.5 ± 74.3 85.7 ± 32.0 < 1 TCAA 3.26 ± 0.1 3.23 ± 0.3 4.22 ± 1.1 4.1 ± 0.2 BCAA 13.8 ± 5.1 10.6 ± 7.4 16.6 ± 4.2 < 1

Table A24: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH);

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 2.51 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs

CHCl2I 12.9 ± 1.5 8.2 ± 1.0 2.7 ± 0.4 3.1 ± 0.6

CHClI2 4.2 ± 0.4 2.1 ± 0.3 0.7 ± 0.1 0.7 ± 0.5 CHBrClI < 0.5 1.1 ± 1.0 0.8 ± 0.8 < 0.5

CHBr2I 1.2 ± 0.2 < 0.5 < 0.5 < 0.5

CHBrI2 1.5 ± 0.1 < 0.5 0.6 ± 0.9 < 0.5

CHI3 1.2 ± 0.2 < 0.5 < 0.5 < 0.5 HANs

IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs

IAA 7.4 ± 0.2 2.8 ± 2.7 5.0 ± 0.8 < 0.5

159

Table A25: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 231.9 ± 25.2 333.0 ± 23.6 390.3 ± 54.2 494.0 ± 35.4

CHCl2Br 68.0 ± 7.3 85.8 ± 3.1 93.9 ± 10.8 98.1 ± 8.4

CHClBr2 7.1 ± 1.0 10.9 ± 0.8 15.9 ± 1.7 17.5 ± 0.7

CHBr3 < 1 < 1 < 1 < 1 HANs CAN 3.0 ± 2.1 1.0 ± 0.1 1.2 ± 0.9 < 1 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs DCAA 36.3 ± 14.5 43.9 ± 10.9 36.3 ± 10.7 27.4 ± 25.7 TCAA 44.7 ± 8.1 35.6 ± 10.3 20.0 ± 1.9 14.1 ± 5.5 BCAA 6.9 ± 2.7 7.7 ± 1.7 6.2 ± 2.3 4.6 ± 4.0

Table A26: Observed Iodo-DBP formation at 72 hours as a function of pH where the

reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs

CHCl2I 2.3 ± 0.3 < 0.5 1.2 ± 0.1 1.8 ± 0.3

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

160 Table A27: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH);

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 16.4 ± 2.3 17.2 ± 4.9 16.9 ± 4.7 15.1 ± 3.7

CHCl2Br 4.2 ± 0.5 1.7 ± 0.6 < 1 < 1

CHClBr2 < 1 < 1 < 1 < 1

CHBr3 < 1 < 1 < 1 < 1 HANs CAN < 1 < 1 < 1 < 1 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs DCAA 145.2 ± 7.5 < 1 27.7 ± 0.1 < 1 TCAA 8.3 ±1.0 < 1 2.4 ± 0.2 < 1 BCAA 14.9 ± 3.0E-02 < 1 1.4 ± 0.1 < 1

Table A28: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH);

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 1.26 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs

CHCl2I 0.63 ± 0.1 1.1 ± 0.1 < 0.5 < 0.5

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5

CHBr2I < 0.5 < 0.5 < 0.5 < 0.5

CHBrI2 < 0.5 < 0.5 < 0.5 < 0.5

CHI3 < 0.5 < 0.5 < 0.5 < 0.5 HANs

IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs

IAA < 0.5 < 0.5 < 0.5 < 0.5

161

Table A29: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 175.4 ± 16.1 240.4 ± 13.2 288.0 ± 25.1 275.4 ± 12.3

CHCl2Br 38.0 ± 3.9 53.3 ± 1.0 65.0 ± 3.5 55.3 ± 2.4

CHClBr2 2.8 ± 0.4 5.0 ± 1.0E-02 9.5 ± 1.0 9.0 ± 0.6

CHBr3 < 1 < 1 < 1 < 1 HANs CAN < 1 < 1 < 1 < 1 TCAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs CAA 511.2 ± 48.1 688.2 ± 28.9 847.7 ± 133.1 698.6 ± 43.6 DCAA 70.0 ± 32.7 63.1 ± 9.8 55.8 ± 17.0 27.6 ± 2.4 TCAA 3.7 ± 0.8 6.4 ± 0.5 3.1 ± 0.8 3.0 ± 0.8 BCAA < 1 11.8 ± 1.5 5.5 ± 1.8 3.4 ± 0.7

Table A30: Observed Iodo-DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L pH DBPs 6.5 7.5 8.5 9 THMs

CHCl2I < 0.5 < 0.5 0.7 ± 0.1 1.1 ± 0.3

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

162 Table A31: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH);

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl3 10.8 ± 1.6 8.7 ± 3.1 4.7 ± 0.6 5.6 ± 1.2

CHCl2Br 1.4 ± 0.3 < 1 < 1 < 1

CHClBr2 < 1 < 1 < 1 < 1 HANs CAN < 1 < 1 < 1 < 1 BAN < 1 < 1 < 1 < 1 HAAs CAA 5.2 ± 0.6 2.6 ± 0.4 < 1 < 1 DCAA 4.1 ± 0.7 3.2 ± 0.1 < 1 < 1 TCAA < 1 < 1 < 1 < 1 BCAA < 1 < 1 < 1 < 1

Table A32: Observed DBP formation at 72 hours as a function of pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH);

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, [TOC] = 0.63 mg/L-L

pH DBPs 6.5 7.5 8.5 9 THMs CHCl2I 0.6 ± 0.1 0.9 ± 0.02 < 0.5 1.0 ± 0.1

CHClI2 < 0.5 < 0.5 < 0.5 < 0.5 CHBrClI < 0.5 < 0.5 < 0.5 < 0.5

CHBrI2 < 0.5 < 0.5 < 0.5 < 0.5 HANs IAN < 0.5 < 0.5 < 0.5 < 0.5 HAAs IAA < 0.5 < 0.5 < 0.5 < 0.5

163

Table A33: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM,

[Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L)

164

Table A34: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol]

= 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L)

165

Table A35: Two-way ANOVA where the response variable was dichloroiodomethane

(CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH);

[Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L)

166

Table A36: Two-way ANOVA where the response variable was dichloroiodomethane

(CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH);

[Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, DOC was varied (2.51, 1.26 and 0.63 mg/L-L)

167

Table A37: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM)

168

Table A38: Two-way ANOVA where the response variable was chloroform (CHCl3) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L,

[NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM)

169

Table A39: Two-way ANOVA where the response variable was dichloroiodomethane

(CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-

L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM)

170

Table A40: Two-way ANOVA where the response variable was dichloroiodomethane (CHCl2I) formation at 72 hours, blocking factor was pH where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C, Iopamidol concentration was varied (5.0, 2.5, 1.0 and 0.0 μM)

171

Figure A1: Interaction plot where the response variable was chloroform (CHCl3) formation at 72 hours, factors (x-axis) were pH and DOC (2.51, 1.26 and 0.63 mg/L- L) where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C

172

Figure A2: Interaction plot where response variable (y-axis) was chloroform (CHCl3) formation at 72 hours, factors (x-axis) was pH and DOC (2.51, 1.26 and 0.63 mg/L-L) where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C

173 Figure A3: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factor (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and DOC (2.51, 1.26 and 0.63 mg/L-L) where the reactor vessel contained iopamidol, aqueous chlorine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C

174

Figure A4: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and DOC (2.51, 1.26 and 0.63 mg/L-L) where the reactor vessel contained iopamidol, aqueous monochloramine and source water (Cleveland, OH); [Iopamidol] = 5 μM, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C

175

Figure A5: Interaction plot where response variable (y-axis) was chloroform (CHCl3) formation at 72 hours, factor (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C

176

Figure A6: Interaction plot where response variable (y-axis) was chloroform (CHCl3) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C

177

Figure A7: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous chlorine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [Cl2]T = 100 μM, [Buffer] = 4mM, temperature = 25° C

178 Figure A8: Interaction plot where response variable (y-axis) was dichloroiodomethane (CHCl2I) formation at 72 hours, factors (x-axis) were pH (6.5, 7.5, 8.5 and 9.0) and iopamidol concentration (5.0, 2.5, 1.0 and 0.0 μM) where the reactor vessel contained aqueous monochloramine and source water (Cleveland, OH); [DOC] = 2.51 mg/L-L, [NH2Cl] = 100 μM, [Buffer] = 4mM, temperature = 25° C

179