United States Office of Science Environmental Protection and Technology July 2003 Agency Washington, D. C.

Office of Water Drinking Water Criteria Document Brominated Acetic Acids DRAFT Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD i Draft, do not cite or quote Acknowledgements Chemical Manager/ Lead Scientist:

Diana Wong, Ph. D., DABT ( Office of Water)

Contractor Authors:

Lynne Haber, Ph. D. ( TERA) Bonnie Stern, M. P. H., Ph. D. ( GRAM, Inc.) Claudine Kasunic ( GRAM, Inc.)

EPA Internal Reviewer:

John Lipscomb, Ph. D., DABT ( NCEA/ ORD) Linda Teuschler, Ph. D. ( NCEA/ ORD)

OST Mail Code 4304T EPA 822 R 03 015 Title: Drinking Water Criteria Document for Brominated Haloacetic Acids: External Review Draft Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD ii Draft, do not cite or quote Table of Contents List of Figures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . iv List of Tables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v I. Executive Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I 1 II. Physical and Chemical Properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . II 1 III. Toxicokinetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 1 A. Absorption . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 1 B. Distribution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 6 C. Metabolism. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 12 D. Excretion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 18 E. Bioaccumulation and Retention . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 21 F. Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 24 IV. Human Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 1 A. Drinking Water Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 1 A. 1 National Occurrence Data for MBA, BCA, and DBA . . . . . . . . . . . . IV 1 A. 0.1 ICR Plants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 2 A. 0.2 Quarterly Distribution System Average and Highest Value for the MBA, BCA, and DBA . . . . . . . . . . IV 2 A. 2 Factors Affecting the Relative Concentrations of MCA, BCA, and DBA in Drinking Water . . . . . . . . . . . . . . . . . . . . . IV 5 A. 2.1 Disinfection Treatment . . . . . . . . . . . . . . . . . . . . . . . . . . IV 6 A. 2.1.1 Disinfection Treatment in ICR Database . . . . . IV 9 A. 2.2 Bromide Concentration . . . . . . . . . . . . . . . . . . . . . . . . . IV 15 A. 2.2.1 Bromide Concentration in the ICR Database . IV 16 A. 2.3 Total Organic Carbon ( TOC) Concentration in ICR Database . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 22 A. 2.4 Seasonal Shifts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 27 A. 2.4.1 Seasonal Shifts in ICR Database . . . . . . . . . . IV 28 Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD iii Draft, do not cite or quote B. Exposure to Sources Other Than Drinking Water . . . . . . . . . . . . . . . . . . . . IV 32 C. Overall Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 33 D. Body Burden . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 33 E. Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 34 V. Health Effects in Animals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 1 A. Short Term Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 1 B. Long Term Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 19 C. Reproductive and Developmental Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . V 22 D. Mutagenicity and Genotoxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 70 E. Carcinogenicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 84 F. Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 85 VI. Health Effects in Humans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VI 1 VII. Mechanisms of Toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII 1 A. Mechanisms of Noncancer Toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII 1 B. Cancer Mechanisms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII 15 C. Sensitive Subpopulations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII 16 D. Interactions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII 21 E. Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VII 21 VIII. Quantification of Toxicological Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 1 A. Introduction to Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 1 A. 2 Quantification of Noncarcinogenic Effects . . . . . . . . . . . . . . . . . . . . VIII 1 A. 1.1 Reference Dose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 1 A. 1.2 Drinking Water Equivalent Level . . . . . . . . . . . . . . . . . . VIII 4 A. 1.3 Health Advisory Values . . . . . . . . . . . . . . . . . . . . . . . . . VIII 5 A. 2 Quantification of Carcinogenic Effects . . . . . . . . . . . . . . . . . . . . . . VIII 7 B. Noncarcinogenic Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 11 Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD iv Draft, do not cite or quote B. 1 Monobromoacetic acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 11 B. 1.1 One Day Health Advisory for MBA . . . . . . . . . . . . . . . VIII 12 B. 1.2 Ten Day Health Advisory for MBA . . . . . . . . . . . . . . . VIII 12 B. 1.3 Longer Term Health Advisory for MBA . . . . . . . . . . . VIII 13 B. 1.4 Reference Dose and Drinking Water Equivalent Level for MBA . . . . . . . . . . . . . . . . . . . . . . VIII 14 B. 2 Bromochloroacetic acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 14 B. 2.1 One Day Health Advisory for BCA . . . . . . . . . . . . . . . VIII 18 B. 2.2 Ten Day Health Advisory for BCA . . . . . . . . . . . . . . . VIII 18 B. 2.3 Longer Term Health Advisory for BCA . . . . . . . . . . . . VIII 19 B. 2.4 Reference Dose and Drinking Water Equivalent for BCA . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 19 B. 3 Dibromoacetic acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 20 B. 3.1 One Day Health Advisory for DBA . . . . . . . . . . . . . . . VIII 25 B. 3.2 Ten Day Health Advisory for DBA . . . . . . . . . . . . . . . VIII 26 B. 3.3 Longer Term Health Advisory for DBA. . . . . . . . . . . . VIII 28 B. 3.4 Reference Dose and Drinking Water Equivalent Level for DBA . . . . . . . . . . . . . . . . . . . . . . VIII 31 C. Carcinogenic Effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 32 C. 1 Monobromoacetic acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 32 C. 2 Bromochloroacetic acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 33 C. 3 Dibromoacetic acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 34 D. Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . VIII 36 IX. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IX 1 Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD v Draft, do not cite or quote List of Figures Figure II 1. The Chemical Structures of MBA, BCA, and DBA . . . . . . . . . . . . . . . . . . . . . II 3 Figure III 1. Proposed Metabolism of DBA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III 14 Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD vi Draft, do not cite or quote List of Tables Table II 1 Physical and Chemical Properties of Brominated Acetic Acids . . . . . . . . . . . . II 3 Table III 1 Toxicokinetic Data for BCA and DBA in F344 Rats . . . . . . . . . . . . . . . . . . . III 2 Table IV 1 Bromoacetic Acids Quarterly Distribution System Average and Highest Value . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 4 Table IV 2 MBA by Disinfection Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 11 Table IV 3 BCA by Disinfection Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 12 Table IV 4 DBA by Disinfection Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 13 Table IV 5 MBA by Influent Bromide Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . IV 18 Table IV 6 BCA by Influent Bromide Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . IV 19 Table IV 7 DBA by Influent Bromide Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . IV 20 Table IV 8 MBA by Influent Total Organic Carbon ( TOC) Concentration . . . . . . . . . . IV 24 Table IV 9 BCA by Influent Total Organic Carbon ( TOC) Concentration . . . . . . . . . . . IV 25 Table IV 10 DBA by Influent Total Organic Carbon ( TOC) Concentration . . . . . . . . . . IV 26 Table IV 11 MBA by Sample Quarter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 29 Table IV 12 BCA by Sample Quarter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 30 Table IV 13 DBA by Sample Quarter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IV 31 Table V 1 Body and Liver Weight Changes Induced by BCA and DBA . . . . . . . . . . . . . V 8 Table V 2 Immunotoxicity of DBA in Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . V 13 Table V 3 General Toxicity of DBA in Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . V 14 Table V 4 Reproductive and Developmental Toxicity of BCA Following Peri conception Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 28 Table V 5 Sperm Quality Parameters in Rats Given 14 Daily Doses of DBA . . . . . . . . . V 41 Table V 6 Reproductive Outcomes in Rats Following Oral Dosing with DBA . . . . . . . . V 43 Table V 7 Outcome of Artificial Insemination of Sperm from Rats Dosed with DBA . . . V 44 Table V 8 Reproductive Organ Weights and Sperm Counts in Rats Given Daily Doses of DBA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 45 Table V 9 Sperm Quality Parameters in Rats Given Daily Doses of DBA . . . . . . . . . . . V 46 Table V 10 Average Consumed Daily Doses ( mg/ kg/ day) for Male and Female Sprague Dawley Rats in the Two Generation Reproductive/ Developmental Toxicity Study . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 60 Table V 11 Incidences of Exposure Related Histopathologic Findings in the Testes of Rats Consuming DBA in Drinking Water . . . . . . . . . . . . . . . . . . . . . . . . V 62 Table V 12 Genotoxicity Studies of MBA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 74 Table V 13 Genotoxicity Studies of DBA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . V 83 Table V 14 Summary of Genotoxicity Data for Brominated Acetic Acids . . . . . . . . . . . . V 84 Table VIII 1 Summary of Oral Studies of MBA Toxicity . . . . . . . . . . . . . . . . . . . . . . . VIII 11 Table VIII 2 Summary of Oral Studies of BCA Toxicity . . . . . . . . . . . . . . . . . . . . . . . . VIII 15 Table VIII 3 Summary of Oral Studies of DBA Toxicity . . . . . . . . . . . . . . . . . . . . . . . . VIII 20 Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 1 Draft, do not cite or quote Chapter I. Executive Summary Three brominated acetic acids, monobromoacetic acid ( MBA), dibromoacetic acid ( DBA),

and bromochloroacetic acid ( BCA), have been selected for evaluation in this drinking water criteria document on the basis of ( 1) their occurrence in drinking water as chlorine disinfection byproducts, and ( 2) the availability of toxicological data on their potential human health effects.

MBA, BCA, and DBA are water soluble hygroscopic crystals in pure form, and are very soluble in water. Brominated acetic acids are formed during ozonation or chlorination of water that contains bromide ions and organic matter, primarily humic and fulvic acids. Formation of chlorinated acetic acids is higher in the presence of humic acid fractions of water than in the presence of fulvic acid, suggesting that a similar relationship may hold for brominated acetic acids.

Bromide ions occur naturally in surface water and ground water, with seasonal fluctuations, and may increase due to saltwater intrusions under conditions of drought or as a result of pollution. In the presence of sufficient concentrations of bromide ion, the formation of brominated compounds may be favored over formation of chlorinated compounds. Brominated acetic acid concentrations in drinking water are typically in the order of BCA> DBA> MBA. No toxicokinetic studies of MBA have been identified in the literature. Although quantitative information on BCA and DBA toxicokinetics is limited to the findings in a single comparative toxicokinetic study with rats, the data demonstrate that both compounds are rapidly absorbed from the gastrointestinal tract, almost completely metabolized, and minimally excreted in the urine and feces. Following a single intravenous dose, neither BCA nor DBA appeared to bind significantly to plasma proteins or Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 2 Draft, do not cite or quote accumulate in blood cells, and the unbound fraction in plasma and the plasma: blood concentrations was close to unity. Further, the apparent volume of distribution was similar to the total body water volume for rats, leading the authors to conclude that both compounds were uniformly distributed outside the vascular system and did not sequester in peripheral tissues.

However, in the absence of specific tissue measurements, the distribution of BCA and DBA cannot be ascertained. The mechanisms by which brominated acetic acids are metabolized remains unclear. Potential pathways of brominated acetic acid metabolism to glyoxylic acid have been proposed based on the observed metabolism of 1,1,2,2 tetrabromoethane, and based on analogy to chlorinated acetic acids Metabolic data from a number of studies demonstrate that chlorinated acetic acids undergo oxidative dehalogenation by glutathione transferase zeta ( GST Zeta) activity and preliminary data indicate that a similar metabolic pathway is likely to occur for the brominated acetic acids. It is not clear whether the toxicologically effective moiety is the parent compound or an active metabolite. Both BCA and DBA are rapidly cleared from the blood, following single oral or intravenous dosing, although these data are inconsistent with the results of repeatedexposure drinking water studies. Based on current information, brominated acetic acids appear to be rapidly excreted and to have little propensity for bioaccumulation. DBA administered at high concentrations to pregnant Sprague Dawley females was reliably measured in placental tissue and in fetal plasma at concentrations that were generally similar to those measured in maternal plasma.

However, quantifiable levels of DBA in the milk of the lactating rats were not detected, leading to the conclusion that DBA freely crosses the placenta and distributes to the fetus during gestation,

but does not appear to bioaccumulate. In contrast, preliminary data from a published abstract Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 3 Draft, do not cite or quote reported the presence of DBA in the milk of lactating female rats at concentrations higher than those in blood serum, suggesting to the authors that DBA might accumulate in milk.

EPA's Information Collection Rule ( ICR) database contains extensive information on concentrations of MBA, BCA, and DBA in drinking water systems, and on how those concentrations vary with input water characteristics and treatment methods. The database contains information from six quarterly samples from 7/ 97 to 12/ 98, from approximately 300 large systems covering approximately 500 plants. The mean concentrations of BCA were 1.47 and 3.61 µ g/ L from groundwater and surface water respectively. The mean concentrations of DBA were 0.82 and 1.09 µ g/ L in groundwater and surface water, respectively. Statistical analysis of these data indicated that the mean concentrations of MBA, BCA, and DBA in surface water were significantly higher than the mean concentrations of these chemicals in groundwater, with BCA >

DBA > MBA in both surface water and groundwater.

The concentrations of MBA in surface water treated with chlorine were similar to those treated with chlorine followed by chloramine. BCA and DBA concentrations were lower when free chlorine was used both in the treatment plant and the distribution system. Although ozonation appeared to significantly reduce the formation of BCA, there were no significant differences in MBA or DBA concentrations with the use of ozone in treating surface water as compared to the common ( non ozonation) chemical disinfection processes. In addition there were no significant differences between the two treatments using ozonation in treating surface water for MBA, BCA and DBA. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 4 Draft, do not cite or quote Consistent with the findings of other investigators, and the chemistry of the formation of bromoacetic acids, a regression analysis of the ICR data indicated that, with the exception of MBA in surface water, there was a significant correlation between influent bromide concentration and the mean concentrations of BCA and DBA in surface water and groundwater.

In addition, for a given influent bromide concentration range, the mean concentrations of BCA were generally significantly higher that the mean concentrations of DBA and MBA in both surface water and groundwater.

A regression analysis of the ICR data indicated that there was a significant correlation between influent total organic carbon ( TOC) concentration and the mean concentrations of MBA,

BCA, and DBA in surface water. This is consistent with the formation of brominated acetic acids from the reaction of humic acid and hypobromous acid, a compound formed by the reaction of bromide ion with ozone and/ or chlorine in the disinfection process. In addition, for a given influent TOC concentration range in surface water, the mean concentrations of BCA were significantly higher than the mean concentrations of DBA, which were significantly higher than the MBA mean concentrations.

Based on only two seasons of monitoring, statistical analysis indicated that the mean concentrations of MBA in surface water were significantly higher in the summer than in the spring Also, based on only two seasons of monitoring, the mean concentrations of BCA in surface water were higher in summer than in winter. Aside from these exceptions, there were no consistently significant differences in the mean concentrations of MBA, BCA or DBA between one season and another in either surface water or groundwater. Seasonal variations in brominated acetic acids Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 5 Draft, do not cite or quote may be dependent on seasonal fluctuations in bromide ion concentration, which were not monitored.

The data on exposure to sources other than drinking water are limited, but MBA has been used in industry and in hospitals. Between 1981 to 1983, approximately 5000 workers were potentially exposed to MBA.. No data were located on exposure to MBA, BCA, or DBA in food,

air, or via dermal exposure. No data could be located on body burden levels of MBA, BCA, or DBA.

The available toxicity database for the brominated acetic acids is limited and many toxicity endpoints have not been fully explored. However, there is a large body of ongoing work,

particularly for BCA and DBA. Preliminary results for many studies have been reported in published abstracts and are included in this document to provide a sense of the spectrum of effects induced by the brominated acetic acids.

Monobromoacetic acid The toxicity data for MBA are very limited. The oral LD 50 for MBA was reported as 177 mg/ kg in male rats. Oral gavage single dose ( 0 or 100 mg/ kg) and 14 day studies ( 0 or 25 mg/ kg/ day) have been conducted to assess the spermatotoxicity of MBA. Neither general toxicity nor spermatotoxicity were observed with either dosing regimen. In a published abstract on the developmental toxicity of MBA, decreased maternal weight gain, decreased live fetus size, and increased incidence of soft tissue malformations were reported at gavage doses of

50 mg/ kg/ day Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 6 Draft, do not cite or quote administered during gestation days ( GD) 6 15. No multi generation reproductive toxicity,

subchronic or chronic studies have been conducted with MBA.

MBA is a dermal irritant, with a lowest observed effect concentration ( LOEC) of 0.2 M following a one hour occluded dermal exposure in rabbits. No data were identified for the toxicity of MBA following exposure by the inhalation route.

No data were identified on the carcinogenicity of MBA. The genotoxicity data for MBA have provided mixed results. MBA was mutagenic in Salmonella typhimurium and induced DNA single strand breaks in vitro, but did not induce SOS DNA repair ( a DNA repair system induced in response to DNA damage) in bacteria or micronuclei in a newt larvae system.

Bromochloroacetic acid Oral toxicity studies of BCA have identified the kidney, liver, and developmental organs as potential targets of toxicity. Increased liver weight was observed at the highest drinking water dose tested ( 500 mg/ kg/ day) in B6C3F1 male mouse given BCA in drinking water in a 21 day study evaluating peroxisomal proliferation and oxidative damage. Marginal increases in liver weight were induced at the highest dose tested ( 39 mg/ kg/ day in drinking water) in rats evaluated for target organ toxicity as part of a 26 or 30 day reproductive and developmental screening assay. In this assay, treatment related liver histopathological changes ( cytoplasmic vacuolization)

were observed beginning at 5 mg/ kg/ day, and became more prominent at 39 mg/ kg/ day in rats given BCA for 30 days. The biological significance of these changes was unclear, as control males in the parallel 26 day study exhibited the same lesion and there was no dose response. Overall, the Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 7 Draft, do not cite or quote authors concluded that the high dose of 39 mg/ kg/ day was sufficient to cause systemic toxicity,

and this value is considered to be a marginal LOAEL.

No standard developmental toxicity studies have been conducted with BCA. The reproductive toxicity of BCA has been assessed in females and male rats exposed to BCA in drinking water for 30 to 34 days during the peri conception period, which included a 12 day premating period of exposure, exposure during cohabitation on days 13 18, and/ or gestational exposures of varying duration ( e. g., GD 1 12, GD 1 21). In females exposed for 30 days, the NOAEL for reproductive and developmental effects ( decreased live fetuses/ litter and decreased total implants/ litter) was 19 mg/ kg/ day, and the NOAEL for maternal toxicity was also 19 mg/ kg/ day, based on kidney toxicity in the pregnant dam. In females exposed only from GD 6 to parturition, no dose dependent increases in either maternal or fetal toxicity were observed. No effects of BCA on male fertility and sperm quality were noted. In another study ( reported in a published abstract), male Sprague Dawley rats administered BCA by gavage for 14 days exhibited impaired sperm motility, abnormalities in sperm morphology, altered spermiation, and reduced fertility ( evaluated by in utero insemination of untreated females). The LOAEL was 8 mg/ kg/ day,

and a NOAEL could not be determined. Adverse effects on sperm quality and male fertility were also reported ( in a published abstract) in male mice exposed to BCA for 14 days. A decrease in the mean number of litters per male and a decrease in the percent of live litters per mated female were observed, with a NOAEL of 24 mg/ kg/ day. No multigeneration reproductive toxicity study has been conducted for BCA. However, BCA is currently undergoing 90 day subchronic and 2

year chronic bioassays. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 8 Draft, do not cite or quote QSAR modeling predicted a LOEC for skin corrosion of 0.7 M for BCA, indicating a potential for dermal irritation. No data were identified for the toxicity of BCA following exposure by the inhalation route.

In a published abstract, BCA was reported to induces liver tumors in mice, but there are no published reports of a full bioassay with BCA. BCA was mutagenic in Salmonella typhimurium and induced oxidative DNA damage, as measured by an increase in the DNA adduct 8 hydroxydeoxyguanisine ( 8 OH dG), in the livers of mice given BCA in drinking water. The data are insufficient to evaluate either the genotoxicity or the potential carcinogenicity of BCA.

Dibromoacetic acid In a range finding study, the reproductive and developmental toxicity of DBA,

administered in deionized drinking water to male and female Sprague Dawley rats, was evaluated.

Animals were exposed beginning 14 days prior to cohabitation and continuing through gestation and lactation ( a total of 63 to 70 days of treatment). Apparent taste aversion was associated with an exposure dependent reduction in water consumption, which was paralleled by a reduction in food intake at all concentrations, resulting in decreased body weights in parental animals and postweanling pups at the two highest doses tested. The only observed adverse reproductive effect was a possible reduction in mating performance in the highest dose group ( 1000 ppm), as evidenced by a slight, but nonsignificant, increase in the number of days of cohabitation and a decrease in the number of mated pairs ( 6/ 10 in the 1000 ppm group versus 9 10/ 10 in all other groups). No effects were observed on the incidence of pre and post implantation losses, live litter Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 9 Draft, do not cite or quote sizes, and gross external morphology or sex ratios in the pups. Based on a lack of reproductive effects, the parental and reproductive/ developmental NOAEL for this study was 1000 ppm.

In a recently completed a two generation reproductive follow up study, male and female Sprague Dawley rats were administered DBA in drinking water, continuously from initiation of exposure of the P generation through weaning of the F2 offspring. Decreased body weight gains were observed in high dose P males and females, and at all exposure levels for F1 male and females, attributed to a general retardation in growth caused by decreased water and food consumption. Observed delays in sexual maturation in the F1 high dose group were also considered to be due to growth retardation. No adverse treatment related effects were observed on any reproductive index or developmental parameters, except for statistically significant,

doserelated increase in the number of males exhibiting altered spermatogenesis ( i. e., retained Step 19 spermatids in Stage IX and X tubules, and increased or abnormal residual bodies in affected seminiferous tubules) in the P and F1 groups, and testicular malformation in four males in the high dose F1 group. The NOAEL was 50 ppm for both P and F1 generations.

A series of rat oral gavage studies on the effects of DBA on spermatogenesis and male fertility, have been conducted using a number of different experimental protocols, including a single high dose study, a 14 day study, and several longer term studies. The results indicated that DBA is clearly spermatotoxic, as demonstrated by histopathology indicative of altered spermiation. In the 14 day study, the LOAEL was 10 mg/ kg/ day, based on mild histopathologic changes ( retention of Step 19 spermatids) and a NOAEL could not be determined. In the longerterm studies, male rats were treated for up to 79 days, with the highest dose group ( 250 Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 10 Draft, do not cite or quote mg/ kg/ day) being treated for only 42 days due to the onset of overt toxicity. However, fertility was assessed at various time points up to 213 days by mating treated males with untreated females. The severity of male reproductive tract toxicity was both dose and duration dependent.

In the group of males given 250 mg/ kg/ day, fertility was impaired throughout the 6 month recovery period following cessation of treatment, indicating that damage was structural and likely permanent. Mild histopathologic changes ( retention of Step 19 spermatids) were observed beginning at 10 mg/ kg/ day while adverse effects on sperm quality were reported beginning at 50 mg/ kg/ day. The NOAEL for spermatotoxicity was 2 mg/ kg/ day. In a published abstract, rats were exposed in utero from GD 15 through postnatal day ( PND) 98 to DBA in drinking water and reproductive development and adult reproductive function were assessed. Male reproductive tract development ( as indicated by delayed preputial separation), as well as spermatogenic and fertility endpoints, were adversely affected at the lowest dose tested ( 50 mg/ kg/ day). In another published abstract, exposure of male rabbits to DBA in drinking water, for a period beginning in utero on GD 15 and continuing to 24 weeks of age, reduced the conception rates of females artificially inseminated with sperm from these treated males. The LOAEL was 0.97 mg/ kg/ day and a NOAEL could not be determined. Full reports of these studies have not yet been published.

A recently published reproductive toxicity study did not demonstrate significant spermatotoxic effects in male rats treated with single oral gavage doses of DBA, as evidenced by the absence of treatment related effects on sperm motility, morphology, and membrane permeability at doses

600 mg/ kg, although mild testes histopathology was reported. The LOAEL histopathology was 600 mg/ kg and a NOAEL could not be determined. The lack of Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 11 Draft, do not cite or quote reported spermatotoxicity in this study is in contrast to an earlier single dose gavage study, in which significantly adverse effects of DBA on sperm count, motility, and morphology at doses

1250 mg/ kg were observed in a different strain of rats. Variation among studies in DBA spermatotoxic effects may have been associated with differences in rat strain, experimental design,

end point measurements, and other study variables.

DBA administered to Holtzman rats via oral gavage on GD 1 8 did not impair female fertility although a 170 % increase in serum 17  estradiol was observed at 250, but not 500,

mg/ kg/ day. In two published abstracts, DBA was reported to induce reproductive and developmental toxicity in pregnant CD 1 mice administered DBA on GD 6 15. Delayed parturition was observed at doses

24 mg/ kg/ day, but the toxicological significance of this effect is unclear. Increased postnatal mortality, decreased pup weight, and tail defects were observed at

610 mg/ kg/ day. In the second abstract by the same authors, the ability of DBA to induce fetal malformations was examined in pregnant CD 1 mice administered DBA by oral gavage on GD 6

15. The NOAEL for renal malformations ( hydronephrosis) was 50 mg/ kg/ day. In contrast, no treatment related effects on litter viability, postnatal mortality, gross malformations and a wide array of other developmental end points were observed in a recent two generation drinking water reproductive/ developmental toxicity study, conducted according to currently accepted standard test guidelines. Differences in findings between the two generation study and those reported in published abstracts may have been due to differences in internal doses associated with gavage versus drinking water DBA administration, species differences in susceptibility to DBA toxicity,

the lower mean doses tested in the two generation study, and/ or other factors. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 12 Draft, do not cite or quote Among the three brominated acetic acids, MBA, BCA, and DBA, unequivocal evidence of adverse effects on spermatogenesis in rats is available for DBA. Although an adverse effect on the mating performance in male rats treated with high doses of DBA was reported , reproductive function was unaffected by altered spermiation in the two generation reproductive/ developmental toxicity study. In the single study identified in the literature, MBA had no effect on spermatogenesis under treatment conditions similar to those that yielded positive indications of spermatotoxicity for DBA. For BCA, no effects on sperm quality were reported in a reproductive and developmental toxicity screening assay. Although a decrease in total implants per litter and live fetuses per litter was reported for BCA in this study, both males and females were exposed to treated drinking water, and thus these reproductive effects might have been due to female exposure. However, adverse effects on male fertility have been reported in two other studies in which only males were treated.

In addition to reproductive and developmental endpoints, the liver, immunotoxicity, and neurotoxicity of DBA have been evaluated. In male mice treated with DBA in drinking water for 21 days, increased liver weight was observed beginning at 125 mg/ kg/ day and was accompanied by oxidative stress, as indicated by increases in hepatic cyanide insensitive Acyl CoA activity and 8 OHdG levels. The NOAEL was 125 mg/ kg/ day.

In an immunotoxicity assay , female mice were given DBA in their drinking water for 28 days. Four independent studies were conducted and different endpoints were examined in each study. Studies 1 3 investigated selected immunologic parameters, body weight changes, and selected organ weights; in Study 4, body weight, organ weights, hematology, and gross pathology Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 13 Draft, do not cite or quote were examined. Overall, the results of Study 1 demonstrated an increase in several measures of cellular immunity, including an increase in the total number of spleen cells and an increase in the percent of spleen cells as macrophages, with statistically significant effects generally occurring at doses

73 mg/ kg/ day. However, the toxicological significance of these findings was unclear. In Study 2, the spleen IgM antibody forming cell response was significantly decreased at

70 mg/ kg/ day, but no change was observed in serum IgM titer ( a more generalized measure of immune function, encompassing immune activity in the bone marrow and lymph nodes as well as the spleen). No change in macrophage activation was observed when tested for in Study 3. In Study 4, the authors reported decreased body weight; decreased thymus gland weight; increased liver, kidney, and spleen weights; and increased reticulocyte counts. For this group of studies,

spleen IgM antibody forming cell response was chosen as the critical effect because it represented a clear decrease in spleen immune system function. The NOAEL for this end point was 38 mg/ kg/ day. Changes in body weight, spleen and thymus weights, and reticulocyte counts occurred at the same or higher doses than the critical effect; changes in liver and kidney weights were observed at lower doses, but were not considered to be toxicologically significant in the absence of supporting clinical chemistry and histopathology data.

In a neurotoxicity study published as an abstract, male and female adolescent F344 rats were exposed to DBA in drinking water for 6 months, and a neurobehavioral test battery was administered to all animals at 1, 2, 4, and 6 months. Dose dependent neuromuscular effects included mild gait abnormalities, decreased forelimb and hindlimb strength, hypotonia, decreased sensorimotor responsiveness ( as measured by responses to a tail pinch and auditory click), Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 14 Draft, do not cite or quote decreased motor activity, and a chest clasping response that was only observed in high dose females. Sensorimotor responsiveness did not progress with continued exposure. Neuropathologic examination showed significant myelin fragmentation, axonal swelling, and axonal degeneration in the white matter of the spinal cord, and eosinophilic or faintly basophilic, occasionally vacuolated swelling, indicative of degenerating axons, in the spinal cord gray matter. Histological evidence of neuropathology was observed in the mid and high dose, and was not evaluated in the low dose group. The LOAEL for this study was 20 mg/ kg/ day, and a NOAEL could not be determined.

No long term systemic toxicity studies for any exposure route were identified in the peerreviewed literature. However, DBA is currently undergoing 90 day subchronic and 2 year chronic bioassays.

Although the genotoxicity database is limited, DBA is mutagenic in Salmonella typhimurium assays and tests for DNA damage repair, and has increased the DNA adduct, 8

OHdG, in hepatic DNA of mice exposed via drinking water.. On the other hand, no induction of micronuclei was reported in a newt larvae system, suggesting that DBA was not clastogenic in the newt test system. The clastogenicity of DBA has not been reported in other assays using a standard test protocol, although DBA has been reported to be co clastogenic. A standard mousemicronucleus assay has not been conducted. These data support the conclusion that DBA is mutagenic and genotoxic, although the nature of the DNA damage induced by DBA remains unclear. In published abstracts, DBA was reported to induce aberrant crypt foci in the colon of rats and liver tumors in mice. However, complete reports of these bioassays have not been published, limiting the utility of these data in assessing the potential carcinogenicity of DBA. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 15 Draft, do not cite or quote No studies were identified that directly evaluated human health effects of exposure to MBA, BCA, or DBA via any route MBA is more toxic than DBA in acute toxicity studies. One proposed cellular basis for the toxicity of MBA is its ability to inhibit enzyme activity through direct alkylation of sulfhydryl and amino groups. This hypothesis is supported by in vitro studies using purified human enzymes and by evidence for DNA alkylation, but a direct relationship between these reactions with cellular macromolecules in vivo and the observed toxic effects of MBA has not yet been established.

DBA and BCA have been associated with liver, kidney, and reproductive and developmental toxicity in a variety of toxicity studies. Potential mechanisms for the induction of adverse liver effects include perturbations of carbohydrate homeostasis or toxicity due to the formation of reactive metabolites from haloacetic acid or tyrosine metabolism pathways. The kidney may also be a target for brominated acetic acids, possibly reflecting direct toxicity associated with the formation of reactive metabolites, or toxicity secondary to oxalate formation although it appears unlikely that sufficient oxalate is formed during brominated acetic acid metabolism to adversely affect the kidney.

Differences in the spermatotoxicity of these brominated compounds are also apparent.

DBA, but not MBA, induced spermatotoxicity at gavage doses that also induced overt toxicity in an acute toxicity study, but effects on reproductive function were not observed in a twogeneration reproductive toxicity drinking water study in which animals were administered spermatotoxic doses of DBA. No evidence of BCA induced spermatotoxicity was found in a reproductive and developmental toxicity screening assay . However, a published abstract reported Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 16 Draft, do not cite or quote impaired sperm quality and spermiation, and reduced male fertility as assessed by in utero insemination of untreated females with the sperm of males treated with BCA for 14 days. In another published abstract, BCA was also reported to decrease male fertility in mice. The weightof evidence suggests that both DBA and BCA are male reproductive tract toxicants. One hypothesized target for observed spermatotoxicity is the Sertoli cells. Although the cellular mechanisms of brominated acetic acid spermatotoxicity have not been identified, the modification of key proteins necessary for Sertoli cell function or direct cytotoxicity by DBA and/ or its reactive metabolites have been suggested as possible mechanisms. Another potential mechanism of spermatotoxicity is haloacetic acid mediated disruption of the early stages of steroidogenesis,

possibly by interfering with the steroidogenic acute regulatory protein ( StAR) mediated transport of cholesterol within the mitochondrial membrane and thereby affecting the synthesis of pregnenolone, the precursor of progesterone. Other studies suggest that brominated acetic acids may interfere with spermatogenesis by altering sperm proteins ( most notably SP22) that play an important role in the fertilization process, possibly by regulating the androgen receptor. It has also been suggested that haloacetic acids acting on SP22 and other sperm proteins may indirectly compromise androgen dependent maintenance of spermatogenesis.

All three brominated acetic acids have been reported to induce developmental effects.

Although the spectrum of developmental endpoints affected by in vivo treatment does not implicate a common mode of action, the results of whole embryo culture ( WEC) testing have suggested the mechanisms of developmental toxicity among haloacetic acids are similar. A QSAR Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 17 Draft, do not cite or quote model using WEC data was able to adequately describe the rank order potency of a series of haloacetic acids. The results of a WEC study testing mixtures of haloacetic acids were consistent with the QSAR model predictions of dose additivity. Brominated acetic acids also induced dysmorphogenesis in WEC at doses lower than the doses of known metabolites, suggesting that either the parent compound or other unidentified metabolites are responsible for these developmental effects. Apoptosis induction has been proposed as having a role in the mechanism of onset of in vivo developmental toxicity based on the results of WEC testing, but this hypothesis has not been confirmed in vivo No data are available for identifying susceptible populations. In addition, no data on agedependent changes in the expression of genes involved in brominated acetic acid were located.

Based on the results of some in vivo developmental toxicity studies reported in abstracts, DBA,

but not MBA or BCA, induced fetal toxicity at lower doses than those associated with maternal effects, suggesting that, at least for DBA, the fetus might be more susceptible than the adult.

However, these preliminary developmental studies found fetal and maternal effects only at doses well above those causing effects on spermiation, indicating that protection against the latter effect would also provide adequate protection to children and fetuses. Additionally, DBA was administered by gavage in these studies and observed fetal toxicity might have been routedependent because increased fetal susceptibility is not supported by the results of the twogeneration drinking water reproductive toxicity study with DBA.

There are also inadequate data on potentially susceptible subpopulations based on genetic variability. Human polymorphisms in GST Zeta, the enzyme that metabolizes DBA and BCA to Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 18 Draft, do not cite or quote glyoxylate, have been characterized by several investigators. However, in the absence of data on whether the parent compound or a metabolite is the active moiety, the functional consequences of this polymorphism are not clear. Individuals having underlying defects in glycogen storage may be susceptible to liver effects induced by brominated acetic acids, and individuals lacking certain enzymes for glyoxylate metabolism may be at risk for BCA or DBA induced kidney toxicity. If the formation of reactive oxygen or lipid intermediates is responsible for the toxicity of brominated acetic acids, then deficits in the activity of anti oxidant enzymes might also represent a source of increased susceptibility. Another potentially susceptible population to DBA are individuals with hereditary tryosinemia II ( a disease involving a deficit in tyrosine metabolism); its chlorinated analog, DCA, has been shown to alter tyrosine metabolism as a consequence of its inhibitory effects on GST Zeta. None of the possibilities has been examined directly in in vivo studies, and potentially susceptible populations have not been identified.

No suitable studies for the derivation of drinking water health advisories ( HA) were identified in the literature for MBA, BCA, or DBA. Neither subchronic nor chronic toxicity studies have been conducted with any of these compounds, although both subchronic and chronic toxicity testing of BCA and DBA is planned or in progress. A number of additional studies are currently ongoing.

There are no human epidemiology studies or full animal cancer bioassays for MBA, BCA,

or DBA, although both BCA and DBA are slated for full testing ( NTP, 2000b; NTP, 2000c).

Under the 1999 Draft Guidelines for Cancer Risk Assessment, the data are inadequate for an assessment of human carcinogenic potential of MBA and BCA. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD I 19 Draft, do not cite or quote There is concern for the potential carcinogenicity of DBA based on preliminary findings reported in published abstracts,, and analogy to DCA, a known high dose rodent liver carcinogen.

However, insufficient data are available to assess DBA carcinogenic hazard. Under the 1999 Proposed Guidelines for Cancer Risk Assessment, the data are inadequate for an assessment of human carcinogenic potential of DBA. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD II 1 Draft, do not cite or quote Chapter II. Physical and Chemical Properties Three brominated acetic acids have been selected for consideration in this document.

These are monobromoacetic acid ( MBA), dibromoacetic acid ( DBA), and bromochloroacetic acid ( BCA). Brominated acetic acids are formed during ozonation or chlorination of water containing bromide ions ( Jacangelo et al., 1989; Pourmoghaddas et al., 1993) and organic matter, primarily humic and fulvic acids. Formation of chlorinated acetic acids is higher in the presence of humic acid than in the presence of fulvic acid ( WHO, 2000), suggesting that a similar relationship may hold for brominated acetic acids. Bromide ions occur naturally in surface and ground water.

However, seasonal fluctuations in bromide ion levels can occur. In addition, bromide ion levels can increase due to saltwater intrusion resulting from drought conditions, or due to pollution ( WHO, 2000).

The bromide ion concentration is an important determinant of the spectrum of haloacetic acids formed from the reaction of disinfectants with organic material. In the presence of sufficient concentrations of bromide ion, the formation of brominated compounds may be favored over formation of chlorinated compounds ( Pourmoghaddas et al., 1993). Brominated acetic acid concentrations in drinking water are typically in the order of BCA> DBA> MBA ( Jacangelo et al.,

1989; Krasner et al., 1989; Boorman et al., 1999; U. S. EPA, 2000a). The occurrence of these compounds is discussed more fully in Chapter IV. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD II 2 Draft, do not cite or quote The chemical reactions resulting in the formation of brominated acetic acids have been described in detail for chlorinated water ( WHO, 2000). When chlorine gas is added to water ( e. g.,

as a disinfectant), it hydrolyzes almost immediately to form hypochlorous acid ( HOCl):

Cl 2 + H 2 O

HOCl + H+ + Cl

Hypochlorous acid can then dissociate into the hydrogen ion and hypochlorite in a reversible reaction:

HOCl  H+ + OCl

In the presence of bromide ion, hypobromous acid ( HOBr) is formed from hypochlorous acid in the following irreversible reaction:

HOCl + Br

HOBr + Cl

Similar reactions occur to form HOBr from other drinking water disinfectants, and the resulting HOBr reacts with organic material to form brominated acetic acids, as shown below ( Jacangelo et al., 1989; Pourmoghaddas et al., 1993). In summary, brominated acetic acids are formed from the following reactions:

Bromide ion + Ozone or HOCl

HOBr HOBr + Organic acid ( e. g., humic acid)

Brominated acetic acid Figure II 1 shows the structure of monobromoacetic acid ( MBA), bromochloroacetic acid ( BCA), and dibromoacetic acid ( DBA), and Table II 1 summarizes key physical and chemical properties of these compounds. The data contained in Table II 1 apply to the pure form of the selected chemicals. These chemicals exist in the environment in a dissolved form. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD II 3 Draft, do not cite or quote DBA Br C OH O Br H C MBA Br H H C C OH O BCA Br C OH O Cl H C Various synonyms exist for the selected chemicals addressed in the document. Some common synonyms for MBA are bromoacetic acid, 2 bromoacetic acid,

 bromoacetic acid,

bromoacetate ion, bromoethanoic acid, carboxymethyl bromide, and acetic acid, bromo.

Synonyms for DBA are dibromoacetate and acetic acid, dibromo. Likewise for BCA, the known synonyms are acetic acid, bromochloro, and chlorobromoacetic acid.

Figure II 1. The Chemical Structures of MBA, BCA, and DBA Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD II 4 Draft, do not cite or quote Table II 1. Physical and Chemical Properties of Brominated Acetic Acids. a Property Monobromoacetic acid ( MBA) Dibromoacetic acid ( DBA) Bromochloroacetic acid ( BCA)

Chemical Abstracts Registry Services No. 79 08 3 631 64 1 5589 96 8 Formula BrCH 2 COOH Br 2 CHCOOH BrClCHCOOH Molecular weight 138.95 217.84 173.39 Appearance hygroscopic crystal hygroscopic crystal hygroscopic crystal Density ( g/ mL) 1.93   1.98 Melting point (  C) 49 51 49 27.5 31.5 Boiling point (  C) 208 218 210 215 Solubility Water Alcohol Log p Log De miscible miscible 0.41c nd very soluble very soluble 1.22d  1.69 ndb nd 1.08d  1.77 a. Adapted from the CRC Handbook of Chemistry and Physics ( 1999), The Merck Index ( 1996), and Sigma Aldrich ( 2000).

b. nd: no data c. Log p is the value derived experimentally as presented in Hansch et al. ( 1995).

d. Log p is the calculated octanol  water partition coefficient in the un ionized form as presented in Schultz et al. ( 1999).

e. Log D is the distribution coefficient between n octanol and buffer at pH 7.4, as presented in Schultz et al. ( 1999).

PK a values were not identified for these compounds. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 1 Draft, do not cite or quote Chapter III. Toxicokinetics Although quantitative toxicokinetic data for brominated acetic acids are limited, the available information suggests that DBA and BCA are well absorbed, almost completely metabolized, and minimally excreted in the urine and feces. Most of these quantitative data come from a single, preliminary study conducted by Schultz et al. ( 1999).

A. Absorption Monobromoacetic acid No studies investigating the quantitative parameters of MBA absorption were identified for any route of exposure. Adverse target organ effects observed in short term toxicity studies ( described in detail in Section V) show that MBA is absorbed following exposure by the oral route; however, the kinetics of absorption are not currently known.

Bromochloroacetic acid BCA is systemically absorbed following oral dosing ( Table III 1). Schultz et al. ( 1999)

administered a single oral gavage or intravenous ( IV) doses of 500 µ mol/ kg ( 87 mg/ kg ) to male F344 rats. Following dosing, BCA venous blood concentrations were measured at 0, 5, 10, 20, Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 2 Draft, do not cite or quote Table III 1. Toxicokinetic Data for BCA and DBA in F344 Ratsa Parameters determined following IV dosing with 500 µ mol/ kg BCA ( 87 mg/ kg) DBA ( 109 mg/ kg)

Area under blood concentration time curve AUC ( µ M h) 576 ± 286b 1120 ± 362 Amount excreted in urine in 24 h (% Dose) 2.16 ± 1.07 2.67 ± 1.09 Steady state apparent volume of distribution ( mL/ kg) 881 ± 373 400 ± 112 Total body clearance ( mL/ h kg) 1037 ± 453 491 ± 116 Renal clearance ( mL/ h kg) 36.9 ± 20.8 12.9 ± 4.0 Mean residence time ( h) 0.92 ± 0.41 0.93 ± 0.50 Elimination half life ( h) 3.93 ± 1.50 0.72 ± 0.12 Unbound fraction in plasma ( f u) 0.93 ± 0.07 0.89 ± 0.05 Blood/ plasma concentration ratio 0.98 ± 0.12 0.91 ± 0.05 Parameters determined following oral dosing with 500 µ mol/ kg BCA ( 87 mg/ kg) DBA ( 109 mg/ kg)

Area under blood concentration time curve AUC ( µ M h) 270 ± 38 333 ± 70 Mean residence time ( h) 2.12 ± 0.81 2.10 ± 0.70 Time to peak blood concentration ( h) 1.5 1 Mean absorption time ( h) c 1.20 1.17 Oral Bioavailability (%) d > 47e > 30e Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 3 Draft, do not cite or quote Notes: a. Adapted from Schultz et al., 1999 b. Mean ± standard deviation c. Calculated as the difference between the mean residence time following IV versus oral dosing. d. The ratio of the blood concentration AUC for oral versus IV dosing x 100%. e. Although the study authors reported estimated values of 47% and 30% for BCA and DBA, respectively, the oral bioavailability is likely to be underestimated, based on first pass metabolism considerations associated with oral and not IV dosing.

30, 60 and 90 minutes, and 3, 4, 6, 8 and 12 hours; concentrations of BCA in the urine and feces were measured in samples collected for 24 hours after dosing. The oral bioavailability  the ratio of the averaged values for the area under the curve for the oral and i. v. doses  was estimated as 47% for BCA. However the oral bioavailability of BCA might be higher than indicated due to more extensive first pass metabolism via this route than via the intraveneous route.

To measure the absorption rate, the time to peak blood concentration was determined.

The peak concentration of BCA was observed 1.5 hours following oral dosing. Rapid absorption of BCA was confirmed by the mean absorption time, which was determined by measuring the difference in the mean residence time in blood between oral and IV dosing. The mean absorption time was reported as 1.2 hours. These data show that BCA is readily absorbed following a single bolus dose. However, no quantitative data are available to assess whether the degree of absorption would be different under ingestion conditions more closely resembling human exposure conditions ( i. e., temporally dispersed and at much lower doses).

No studies on the absorption of BCA were identified following exposure by the inhalation or dermal routes.

Dibromoacetic acid Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 4 Draft, do not cite or quote Shultz et al. ( 1999) also studied the systemic absorption of DBA following oral dosing, as described above for BCA ( Table III 1). Male F344 rats were given single oral gavage or IV doses of 500 µ mol/ kg ( 109 mg/ kg) DBA. Following dosing, the DBA venous blood concentrations were measured at 0, 5, 10, 20, 30, 60 and 90 minutes, and 4, 6, and 8 hours; concentrations of DBA in the urine and feces were measured in samples collected 24 hours after dosing. The oral bioavailability of DBA was estimated to be 30%. However, this value is likely to be an underestimate because it probably reflects first pass metabolism following oral dosing and, thus,

underestimates the degree of oral absorption. The peak blood concentration was observed 1 hour following oral dosing and the mean absorption time ( the difference in the mean residence time in blood following dosing via oral and IV routes) was 1.17 hours, indicating rapid absorption.

However, no quantitative data were provided on the degree of absorption under conditions of low dose repeated oral dosing, a regimen which would more closely resemble human exposure conditions.

Blood level measurements for brominated acetic acids following oral dosing have been reported in other toxicity studies, and provide some additional information on DBA absorption.

As part of a study on the effects of DBA on pubertal development and adult reproductive function, Klinefelter et al. ( 2000, abstract only) reported blood serum and milk concentrations in Sprague Dawley rats ( 3 litters/ dose) administered drinking water containing 0, 400, 600, or 800 ppm DBA from gestation day ( GD) 15 through postnatal day ( PND) 98. Estimated doses resulting from these treatments were 0, 50, 75, and 100 mg/ kg/ day, respectively ( personal communication). DBA levels were also assayed in dams' milk and blood serum, and in the serum Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 5 Draft, do not cite or quote of suckling males on PND 20 ( personal communication). Only data for the 800 ppm treatment group were presented in the abstract. In this group, DBA concentrations were 5.2 14.1 µ g/ mL in dams' milk, 3.0 6.9 µ g/ mL in dams' serum, and 0.01 0.24 µ g/ mL in the serum of male offspring.

The limited amount of information in the abstract precluded quantification of the degree of absorption. However, the data demonstrate that DBA was absorbed following drinking water exposure. In contrast, no detectable levels of DBA were observed in the plasma of female B6C3F1 mice following 28 days of exposure to DBA treated drinking water at concentrations up to 2000 mg/ L, corresponding to estimated doses of 229 to 285 mg/ kg/ day depending on the substudy ( NTP, 1999). The reasons for the differences in findings between the Klinefelter ( 2000) and NTP ( 1999) studies are unclear, and may have been due to differences in analytical methods. No details on the post dosing sampling schedules for blood and/ or milk were presented in either study. The absence of measurable DBA in plasma in the NTP ( 1999) study might reflect extensive metabolism and rapid excretion, rather than limited absorption. Alternately, species differences and/ or differences in hormonal status might have affected the kinetics of DBA absorption,

distribution, metabolism and/ or excretion in these studies. Christian et al. ( 1999) reported the detection of DBA in the plasma of male and female rats exposed to DBA in drinking water concentrations for 16 hours, and following 14 days of treatment, also demonstrating that gastrointestinal absorption occurred.

No studies on the absorption of DBA were identified following exposure by the inhalation or dermal routes. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 6 Draft, do not cite or quote Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 7 Draft, do not cite or quote B. Distribution Monobromoacetic acid No studies of MBA tissue distribution following dosing by any route were identified.

Bromochloroacetic acid Schultz et al. ( 1999) administered male F344 rats single oral gavage or IV doses of 500 µ mol/ kg ( 87 mg/ kg) BCA. BCA venous blood concentrations were measured at 0, 5, 10, 20, 30,

60 and 90 minutes, and at 3, 4, 6, 8 and 12 hours post dosing. No concentrations in tissues other than blood were assessed. Following intravenous dosing, BCA did not appear to bind significantly to plasma proteins or accumulate in blood cells. The unbound fraction in plasma was 0.93, and the ratio between plasma and blood concentrations was close to unity ( i. e., 0.98). Tissue concentrations were not measured directly. However, following intravenous dosing, the apparent volume of distribution ( 881 mL/ kg) was similar to the total body water volume for rats ( approximately 660 mL/ kg) ( Reinoso et al., 1997). This similarity suggested to the study authors that BCA distributed uniformly outside the vascular system and was unlikely to sequester significantly in peripheral tissues. The octanol buffer partition coefficient ( Log D), considered to be a reasonable predictor of lipophilicity ( Schultz et al., 1999) was reported to be  1.77 at pH of 7.4. This low value also suggested that at physiological pH values, BCA has little propensity to Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 8 Draft, do not cite or quote accumulate in fat tissue. However, in the absence of direct tissue measurements, the distribution of BCA cannot be ascertained. Plasma binding, plasma: blood concentrations, and the apparent volume of distribution ( V ss) were only measured following intravenous dosing and, thus, the effects of first pass metabolism are not known. Further, the high dose employed in this study might have resulted in metabolic saturation which could have led to a wider distribution of BCA than would have occurred at lower doses where metabolism was not saturated.

Short term animal studies suggest that oral administration of high doses of BCA results in liver, reproductive, and developmental toxicity ( NTP, 1998, Parrish et al., 1996; see Chapter 5 for more details), indicating that BCA does distribute to the liver, the reproductive organs, and the fetus under the conditions of these studies. The low protein binding capacity of BCA suggests that the potential for distribution across the placenta to the fetus may be significant at high maternal doses administered during pregnancy.

No data on tissue distribution of BCA following exposure by the inhalation or dermal routes were identified.

Dibromoacetic acid The systemic distribution of DBA has also been studied by Schultz et al. ( 1999). Male F344 rats were given single oral gavage or IV doses of 500 µ mol/ kg ( 109 mg/ kg) DBA. DBA venous blood concentrations were measured at 0, 5, 10, 20, 30, 60 and 90 minutes, and at 4, 6,

and 8 hours post dosing. No tissue concentrations other than blood were measured. Following Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 9 Draft, do not cite or quote intravenous dosing, DBA did not bind significantly to plasma proteins or accumulate in blood cells. The unbound fraction in plasma was 0.89 for DBA, and the ratio between plasma and blood levels of DBA was close to unity ( i. e., 0.91). Tissue concentrations were not measured directly,

but the apparent volume of distribution and the total body water volume in rats were similar,

suggesting to the study authors that DBA was widely and uniformly distributed outside the vascular system and was unlikely to sequester significantly in peripheral tissues. The octanolbuffer partition coefficient ( Log D), considered to be a reasonable predictor of lipophilicity ( Schultz et al., 1999), was reported to be  1.69 at pH of 7.4. This low value suggested that at physiological pH values, DBA has little propensity to accumulate in fat tissue. However, in the absence of direct tissue measurements, the distribution of DBA is not known. Plasma binding,

plasma: blood concentrations, and the apparent volume of distribution ( V ss) were only measured following intravenous dosing and, thus, the effects of first pass metabolism cannot be ascertained.

Further, the high dose employed in this study might have resulted in metabolic saturation, which could have led to a wider distribution of DBA than would have occurred at lower doses where metabolism was not saturated.

Short term animal studies suggest that oral administration of DBA results in toxicity to the liver, kidney, spleen, and male reproductive system.( Linder et al., 1994a; Linder et al., 1994b;

Linder et al., 1995; Parrish et al., 1996; Linder et al., 1997; Cummings and Hedge, 1998; Vetter et al., 1998; NTP, 1999; more details in Chapter 5), indicating that DBA does distribute to these organs under the dosing conditions of these studies. Further, the study by Klinefelter et al. ( 2000,

abstract), described in the previous section on DBA absorption, showed that the milk of lactating Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 10 Draft, do not cite or quote Sprague Dawley females treated with 800 ppm ( 100 mg/ kg/ day) of DBA in drinking water contained elevated levels of DBA relative to the females' blood serum ( 5.2 14.1 µ g/ mL in milk,

3.0 6.9 µ g/ mL in serum). Thus, lactational distribution to nursing pups may be significant at high maternal doses.

As part of a series of range finding reproductive and developmental toxicity studies,

Christian et al. ( 2001) evaluated the distribution of DBA in adult Sprague Dawley rats, and in pregnant females and their fetuses. Male and female rats ( 10/ sex/ group) were given DBA in deionized drinking water at concentrations of 0, 125, 250, 500 or 1000 ppm, beginning 14 days prior to cohabitation and continuing through gestation and lactation ( 63 70 days of treatment).

The average daily doses ( based on measured water consumption and body weights) varied,

depending on the phase of reproduction. For parental males throughout the study ( SD 1 70),

equivalent mean daily doses were 10.2, 20.4, 35.7, and 66.1 mg/ kg/ day, respectively. For females on SD 1 15, equivalent mean daily doses were 13.3, 26.2, 41.8 and 60.2 mg/ kg/ day, respectively;

and 14.8, 30.3, 48.5 and 81.6 mg/ kg/ day, respectively, on gestation day ( GD) 0 21. During lactation ( LD 1 29), the estimated doses were was 43.5, 86.6, 150.7 and 211.7 mg/ kg/ day for the 0, 125, 250, 500, and 1000 ppm groups, respectively; however, these doses included consumption of water by the pups and thus overestimated the mean daily intake for lactating females.

An additional 6 male and 17 female rats/ group were used for collecting bioanalytical samples. Blood plasma levels of DBA in parental male and female rats were taken on SD 1 and 14, in mated females on GD 20 and LD 15, and in weanling male and female rats on LD 29 ( which was also postweaning day 1). Tissue levels of DBA in placenta and amniotic fluid were Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 11 Draft, do not cite or quote assessed on GD 21 and in milk on LD 15. During the designated collection days, three plasma samples were collected from male and female rats at approximately 8 hour intervals in order to assess the potential differences in tissue concentrations associated with diurnal rhythms of water consumption by animals.

In analyzing biodisposition samples, numerous values were measured at concentrations below the limit of detection ( LOD) of the methodology used. These values were quantified but were not considered by the study authors to be reliable. In blood plasma, DBA was detected in male rats at 125 ppm and reliably quantified at

250 ppm after 24 hours of continual access to DBA in drinking water. Blood plasma levels in males exposed to 125 ppm did not exceed the level of detection even after 14 days of exposure. The overall pattern of DBA detection in plasma in female rats was similar but more variable; however, quantifiable DBA was increased in a dosedependent fashion at all exposure concentrations on SD 14, and was observed at concentrations of 4.4 6.7 µ g/ g at the end of the dark period. The higher plasma levels at this time were attributed to nocturnal drinking habits and not to accumulation of DBA in plasma. During gestation, the calculated daily dose of DBA was increased, and was reflected in increased plasma concentrations at all DBA exposure levels on GD 20 ( ranging from 4.2 18.0 µ g/ g), but not in an exposuredependent manner. During lactation, DBA could be quantified in the maternal plasma at all doses when measured on LD 15. Measurements ranged from 1.9 to 26.2 µ g/ g and varied, depending on time of sample; a dose response was only observed at one time point. Analysis of milk samples collected from three lactating dams/ group during the light period on LD 15 indicated that DBA was not detected in milk, in contrast to the findings of Klinefelter et al ( 2000, abstract). Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 12 Draft, do not cite or quote Pooled fetal blood, amniotic fluid, and placentas were collected on GD 21 from three litters/ exposure group and analyzed for DBA. In the placenta, DBA was detected at all exposure levels but could only be reliably quantified in the 1000 ppm group ( 8.1 µ g/ g at a mean maternal daily dose of 8.16 mg/ kg/ day). In fetal plasma, DBA was detected in a dose dependent manner in all groups; however, levels in the 125 and 250 ppm groups were below the limit of detection and reliable quantitation was only possible in the 500 and 1000 ppm groups ( 2.4 and 9.2 µ g/ g at maternal doses of 48.5 and 81.6 mg/ kg/ day, respectively). In contrast, quantifiable amounts of DBA were noted in the amniotic fluid ( 3.9, 5.3, 3.0, and 5.8 µ g/ g at 14.8, 30.3, 48.5, and 81.6 mg/ kg/ day, respectively) and levels were comparable to those observed in the plasma of maternal rats on GD 20 at the start of the dark period. demonstrated a dose dependent distribution of DBA in amniotic fluid ( 3.9 5.8 µ g/ g) on GD 21 at DBA drinking water concentrations of 125 1000 ppm ( estimated mean daily doses of 14.8 81.6 mg/ kg/ day). Equivalent concentrations of serum plasma DBA in pregnant females on GD 20, taken at the same time during the dark cycle as the amniotic fluid samples, ranged from 3.6 to 6.4 µ g/ g. These results demonstrate that at high drinking water concentrations, DBA can cross the placenta and distribute to fetal tissue.

No data on tissue distribution of DBA following exposure by the inhalation or dermal routes were identified. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 13 Draft, do not cite or quote C. Metabolism Monobromoacetic acid No studies were identified that described the metabolism of MBA following exposure by any route.

Bromochloroacetic acid Most of the data on the metabolism of BCA are indirect or based on analogy to the chlorinated acetic acids. These indirect data suggest that dihalogenated acetic acids are rapidly metabolized. Schultz et al. ( 1999) did not directly measure the metabolism of BCA. However,

comparisons of renal and blood clearance following IV administration suggested that metabolism was likely to be the major contributor to BCA removal from the blood. Only 2% of BCA blood clearance was accounted for by renal clearance and excretion in the feces was negligible. Thus,

non renal clearance ( e. g., through metabolism) accounted for most of the removal of the parent compound from the blood. It should be noted that these data were obtained following IV administration of the test compound and, thus, the degree of metabolism in different tissues could not be determined. The relatively low blood concentrations of BCA following oral dosing,

compared to the blood concentrations following IV dosing, suggest that the liver may be an important site for first pass metabolism. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 14 Draft, do not cite or quote Shultz et al. ( 1999) suggested that the similarities between the toxicokinetics of BCA and dichloroacetic acid ( DCA), coupled with limited in vitro metabolism data ( Schultz, et al., 1998;

Tong et al., 1998a), support the hypothesis that metabolism of BCA is similar to that of DCA.

However, while the metabolic pathways for BCA and DCA might be similar, the rate of metabolism seems to be greater for BCA than DCA based on area under the curve plasma data.

In a review of DCA metabolism, Stacpoole et al. ( 1998) described several potential mechanisms for the dehalogenation of DCA, but evidence for any single pathway was limited.

Recent evidence suggests that DCA is metabolized to glyoxylic acid through a glutathione

Stransferase dependent mechanism by a novel GST isozyme, GST Zeta ( Tong et al., 1998b); in vitro studies have demonstrated that this enzyme can also catalyze the metabolism of BCA to glyoxylic acid ( Tong et al., 1998a). In comparative studies using DCA, Tong et al. ( 1998a) found that GST Zeta is expressed in mouse, rat and human liver cytosol. For DCA, the relative biotransformation rates were mouse> rat> human. Although glutathione was a required cofactor, it was not consumed or oxidized during the conversion of DCA to glyoxylate. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 15 Draft, do not cite or quote Br Br O H Br Br O OH OH O OH O Br O O H OH O O OH OH CO 2 OH HAOX Glycolate GST Zeta OGC H H O OH N H H Glycine OH H H H O LDH Dibromoacetic acid Glyoxylic acid Oxalic acid C AGT Figure III 1. Proposed Metabolism of DBAa AGT = alanine: glyoxylate aminotransferase GR = glyoxylate reductase GSTZ = glutathione S transferase Zeta HAOX = glycolate oxidase ( 2 hydroxyacid oxidase) LDH = lactate dehydrogenase OGC = 2 oxoglutarate: gloxylate carboligase a. Adapted largely from Kennedy et al. ( 1993), Stacpoole et al. ( 1998) Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 16 Draft, do not cite or quote In addition to glyoxylate, DCA metabolites detected in vivo included monochloroacetic acid and a series of downstream metabolites of glyoxylate ( Stacpoole et al., 1998). Thus, the formation of glyoxylate metabolites may be of toxicologic importance in BCA metabolism and is described here briefly. Glyoxylate may be metabolized through competing pathways to form glycine, glycolate, oxalate, and CO 2 ( Stacpoole et al., 1998).. Glyoxylate can also be converted to glycolate by glyoxylate reductase ( Michal, 1999). Glycine may be formed through the activity of glycolate aminotransferase ( formally known as alanine: glyoxylate aminotransferase). Glycine,

which can be incorporated into proteins, used in the synthesis of serine, or degraded releasing carbon dioxide ( Michal, 1999). Conversion to oxalate may occur via a ( S) 2 hydroxyacid dehydrogenase such as lactate dehydrogenase Few data are available regarding the kinetics of brominated acetic acid metabolism. In the single toxicokinetics study identified in the literature that examined absorption, distribution,

metabolism, and excretion ( Schultz et al., 1999), only one dose of BCA was used and analysis of metabolic saturation could not be conducted. Gonzalez Leon et al. ( 1999, published abstract)

used microsomes to study the effect of BCA pre treatment on metabolic inhibition following administration of subsequent BCA doses. Microsomal fractions were prepared from the livers of male F344 rats given 2000 mg/ L BCA in drinking water for 2 weeks and in vitro metabolism was assessed. Pretreatment reduced the V max by 50% to 75%, while the K m remained unchanged,

indicating possible noncompetitive inhibition of metabolism. However, in the absence of additional detail on laboratory and analytic methods, it is not clear whether metabolism actually occurred in the microsomes or whether the microsomal samples were contaminated with cytosol, and Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 17 Draft, do not cite or quote metabolism occurred in the cytosol. Thus, an alternative interpretation of the data is that the observed metabolism was due to cytosolic contamination of the microsomal fraction, and that pretreatment with BCA inhibited the cytosolic enzyme. Anderson et al. ( 1999) administered i. p.

injections of 0.3 mmol BCA to male F344 rats ( 3/ dose) and measured GST Zeta activity in liver cytosol 12 hours later. BCA reduced GST Zeta activity to 19% of that in saline treated controls,

indicating a possible mechanism for auto inhibition of metabolism. The toxicologic implications of these findings are not yet clear because it is not known whether the toxic moiety is the parent compound or a metabolite. However, this finding is of interest because DCA is known to inhibit its own metabolism through inhibition in both rodents ( Line et al., 1993) and humans ( Stacpoole et al., 1998), and BCA may also exhibit similar metabolic activity. In the case of DCA inhibition is the result of DCA reaction with and subsequent modification of GST zeta.( Tong et al., 1998a).

Dibromoacetic acid As for BCA, most of the information on the metabolism of DBA is indirect or based on analogy to chlorinated acetic acids. These indirect data suggest that DBA is likely to be rapidly metabolized. Comparisons of DBA renal and blood clearance in the Schultz et al. ( 1999) study revealed that less than 1% of blood clearance of the parent compound was accounted for by urinary excretion, suggesting that metabolism is the major contributor to DBA removal from the blood. However, these data were obtained following IV administration of the test compound and,

thus, the degree of metabolism in different tissues cannot be determined from the data provided. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 18 Draft, do not cite or quote The limited oral bioavailability of DBA suggests that the liver may be an important site for firstpass metabolism. The proposed metabolic pathway for DBA, based on the data presented in the following paragraphs, is shown in Figure III 1.

Shultz et al. ( 1999) proposed that similarities between the toxicokinetics of DCA and those of DBA and BCA indicate that these compounds are likely to share metabolic pathways.

This hypothesis is supported by several comparative in vitro metabolic studies ( Schultz et al.,

1998; Tong et al., 1998a) that demonstrated that GST Zeta can catalyze the metabolism of DBA,

as well as BCA and DCA, to glyoxylic acid ( Tong et al., 1998a)

As described in the previous section for BCA, very little data are available regarding the kinetics of brominated acetic acid metabolism. In the single toxicokinetics study identified in the literature that examined absorption distribution, metabolism, and excretion ( Schultz et al., 1999),

only one dose of DBA was used. Therefore, no conclusions regarding metabolic saturation can be made. In a study similar to that reported in the previous section for BCA, Gonzalez Leon et al.

( 1999, published abstract) studied the effects of DBA pre treatment on in vitro metabolic inhibition in the liver microsomes of male F344 rat liver following in vivo administration of 2000 mg/ L DBA in drinking water for 2 weeks. Similar to BCA, pretreatment reduced the V max by 50%

to 75%, while the K m remained unchanged, suggesting metabolic inhibition; however it was not clear from the abstract whether metabolism occurred in the microsomes or in the cytosol.

Anderson et al. ( 1999) administered i. p. injections of 0.3 mmol DBA to male F344 rats ( 3/ dose)

and measured GST Zeta activity in liver cytosol preparations 12 hours later. DBA administration reduced GST Zeta activity to 17% of that of saline treated controls, indicating a possible Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 19 Draft, do not cite or quote mechanism for auto inhibition of metabolism. The toxicologic implications of these findings are not yet clear because it is not known whether the toxic moiety is the parent compound or a metabolite. However, it is of interest that similar auto inhibition was seen for DCA metabolism ( Tong et al., 1998a).

D. Excretion Monobromoacetic acid No studies on the excretion of MBA following exposure by any route were identified.

Bromochloroacetic acid Schultz et al. ( 1999) measured parent compound concentrations in the blood, urine, and feces after oral or i. v. dosing of male F344 rats with 500 µ mol/ kg ( 87 mg/ kg) BCA. Blood measurements were taken at 0, 5, 10, 20, 30, 60 and 90 minutes, and at 3, 4, 6, 8 and 12 ( i. v.

only) hours post dosing; urine and feces were collected 24 hours following dosing. BCA was rapidly cleared from the blood, with apparent bi exponential elimination following i. v.

administration. There was an initial, rapid decline in blood concentration, corresponding to a short distributive phase, followed by a long linear decline. In the concentration time profile, peculiarities were noted in the profile which suggested that physiological mechanisms or processes were Drinking Water Criteria Document for Brominated Acetic Acids 1 Personal communication, I. Schultz, Battelle Laboratories, Washington EPA/ OW/ OST/ HECD III 20 Draft, do not cite or quote involved other than multiple distribution phases ( e. g., 2 3 compartments or distribution phases). 1 As a result, the authors were doubtful that the unique appearance of the profiles was due to a prolonged distribution phase( s), and chose to analyze the data using simple non compartmental methods, which require fewer assumptions than compartmental models with regard to distribution within the animal. Therefore, they provided two estimates of half life ( t 1/ 2): one relying on the initial decline in the profiles ( 0 4 hours) and another using the full or complete profile. Truncating the concentration time profiles had no significant effect on the AUC; however, the elimination half life was markedly altered by more than five fold; t ½ was 3.93 hours for the complete profile versus 0.74 hours for the truncated profile. After oral administration, blood levels reached a maximum at 1.5 hours following dosing and declined rapidly during the next 6 hours; t max was 1.5 hours.

Removal of parent compound from the blood appeared to be rapid, due mainly to biotransformation ( Schultz et al., 1999). The urine and feces were minimal contributors to overall blood clearance. Urinary clearance of the parent compound accounted for 2% of the total clearance and feces contained negligible amounts of BCA. The study authors did not measure either putative metabolites or expired CO 2. Therefore, it was not possible to determine the contribution of each route of excretion to the total administered dose of the parent compound.

No data on the excretion of BCA following exposure by the inhalation or dermal routes were identified. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 21 Draft, do not cite or quote Dibromoacetic acid Schultz et al. ( 1999) also measured DBA concentrations in the blood, urine, and feces 24 hours after oral or i. v. dosing of male F344 rats with 500 µ mol/ kg DBA ( 109 mg/ kg). Blood measurements were taken at 0, 5, 10, 20, 30, 60 and 90 minutes, and at 3, 4, 6 and 8 hours postdosing urine and feces were collected 24 hours following dosing. DBA was rapidly cleared from the blood, with apparent bi exponential elimination following i. v. administration. After oral administration, blood levels reached a maximum about one hour following dosing and declined rapidly during the next six hours. Similar to BCA, features were noted in the DBA concentrationtime profile which suggested physiological mechanisms or processes were involved other than multiple distribution phases ( e. g., 2 3 compartments or distribution phases), and the study authors chose to analyze the data using simple non compartmental methods, which require fewer assumptions than compartmental models with regard to distribution within the animal. As with BCA, they provided two estimates of t 1/ 2: one relying on the initial decline in the profiles ( 0 4 hours) and another using the full or complete profile. Unlike BCA, DBA elimination half lives were similar for both the complete and truncated profiles ( 0.72 versus 0.52 hours). The urine and feces were minimal contributors to overall blood clearance. Urinary clearance of DBA accounted for only a small fraction of the total clearance and was less than 1% of total clearance; negligible amounts of DBA were found in the feces. The study authors did not measure either putative metabolites or expired CO 2. Therefore, it was not possible to determine the contribution of each route of excretion to the total administered dose of the parent compound. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 22 Draft, do not cite or quote No data on excretion of DBA following exposure by the inhalation or dermal routes were identified.

E. Bioaccumulation and Retention Monobromoacetic acid No studies on the bioaccumulation and retention of MBA following exposure by any route were identified.

Bromochloroacetic acid The available data on the potential for bioaccumulation or retention of BCA are very limited. Schultz et al. ( 1999) demonstrated that a single dose of BCA is rapidly eliminated from the blood. Following intravenous dosing, BCA did not appear to bind significantly to plasma proteins or accumulate in blood cells. The unbound fraction in plasma and the plasma to blood concentration approached unity, and the apparent volume of distribution was similar to the total body water volume for rats ( Reinoso et al., 1997), suggesting that BCA distributed uniformly outside the vascular system and was unlikely to accumulate significantly in peripheral tissues.

However, only a single dose was administered and no measurements were collected in tissues other than blood. Therefore, the extent of bioaccumulation or retention cannot be determined. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 23 Draft, do not cite or quote BCA at physiologic pH is not lipophilic ( Schultz et al., 1999), suggesting little affinity for accumulation in adipose tissue.

No studies were identified on the bioaccumulation and retention of BCA following exposure by the inhalation or dermal routes.

Dibromoacetic acid Similar to BCA, DBA was rapidly eliminated from the blood in the toxicokinetic study by Schultz et al. ( 1999), and did not bind significantly to plasma proteins or accumulate in blood cells. Both the unbound fraction in plasma and the plasma: blood concentrations were close to unity and the apparent volume of distribution was similar to the total body water volume for rats ( Reinoso et al., 1997), suggesting uniform distribution outside the vascular system and little affinity for accumulation in peripheral tissues. However, as with BCA, only a single dose was administered and no measurements were collected in tissues other than blood. Rapid blood clearance of DBA was also suggested by the absence of detectable blood levels in an NTP ( 1999)

immunotoxicity study. Kennedy et al. ( 1993) reported on the tissue retention of radiolabel following oral dosing of rats with [ 14 C] 1,1,2,2 tetrabromoethane, a compound whose major urinary metabolite is DBA. After 96 hours, the percent of administered dose retained in the body ranged from 14% to 22% and was not dose dependent. The largest percentage of dose was in the carcass, followed by the liver, blood, and gastrointestinal tract, with lesser amounts found in the kidney and fat. These data suggest that 1,1,2,2 tetrabromoethane and/ or one ( or more) of its Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 24 Draft, do not cite or quote metabolites were widely distributed; however, it is not known whether the observed tissuedistribution pattern would be similar following direct oral dosing with DBA. Further, the results of total [ 14 C] distribution did not identify whether the tissue distribution represented the parent compound or its metabolites, and the specific metabolites were not identified.

DBA at physiologic pH is not lipophilic ( Schultz et al., 1999), suggesting little propensity for retention or accumulation in adipose tissue. However, Klinefelter et al. ( 2000, abstract)

reported the presence of DBA in the milk of Sprague Dawley females, following high dose exposure during pregnancy and lactation, at concentrations greater than those measured in females' blood serum ( Klinefelter et al., 2000, abstract), suggesting that retention or accumulation may be possible under certain physiologic conditions. A full report of this study has not been published and thus these findings cannot be comprehensively evaluated. In contrast, Christian et al. ( 2001) did not detect levels of DBA in the milk of lactating Sprague Dawley rats at drinking water concentrations up to 1000 ppm ( estimation of daily doses to lactating dams was confounded by concomitant pup water consumption). DBA was reliably detected on GD 21 in placental tissue at 1000 ppm ( 81.6 mg/ kg/ day), and in fetal plasma at 500 and 1000 ppm ( maternal doses of 48.5 and 81.6 mg/ kg/ day, respectively), at concentrations which were generally similar to those measured in the plasma of pregnant females on GD 20. Higher concentrations of DBA in the plasma of male and female rats noted on SD 14 as compared with SD 1 were attributed by the authors to variability and not to accumulation. Placental DBA levels on GD 21 were lower than those observed in maternal serum plasma on GD 20 except at 1000 Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 25 Draft, do not cite or quote ppm. Christian et al. ( 2001) concluded that although DBA freely crossed the placenta and distributed to the fetus during gestation, it did not appear to bioaccumulate.

No studies were identified on the bioaccumulation and retention of DBA following exposure by the inhalation or dermal routes.

F. Summary No toxicokinetic studies of MBA have been identified in the literature. Brominated acetic acids appear to be rapidly absorbed from the gastrointestinal tract ( Schultz et al., 1999). Key data from this study are summarized in Table III 1. Following single dose intravenous or oral gavage exposure, both BCA and DBA were rapidly cleared from blood and had short plasma elimination half lives ( Schultz et al., 1999). However, the extent of tissue distribution is not known because tissue distribution studies of these compounds have not been conducted. Neither BCA nor DBA are lipophilic at physiologic pH ( Schultz et al., 1999), suggesting a low propensity to accumulate in fat. Following repeated exposure in drinking water, DBA was detected in the blood serum and milk of lactating female Sprague Dawley rats exposed from gestation day 15 to postnatal day 20 ( Klinefelter, 2000, abstract), but was not detected in the blood plasma of nonlactating female B6C3F1 mice after 28 days of exposure ( NTP, 1999). The discrepancy in these findings might be due to sampling differences, species differences, and/ or differences in physiologic status.

The metabolism of BCA and DBA has also not been thoroughly investigated. Limited in vitro data ( Schultz et al., 1998; Tong et al., 1998a, 1998b) and the single comparative Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD III 26 Draft, do not cite or quote toxicokinetics study by Schultz et al. ( 1999) suggest that both BCA and DBA are metabolized in a manner similar to DCA. Potential pathways of brominated acetic acid metabolism to glyoxylic acid have been proposed based on analogy to chlorinated acetic acids ( reviewed in Stacpoole et al., 1998); these pathways are mediated through a recently identified class of GST isoenzymes,

GST Zeta. It is not clear whether the effective toxicologic moiety is the parent compound or an active metabolite. Overall, the data are consistent with DBA and DCA being rapidly excreted and having little propensity for bioaccumulation. Lactational exposure may be a route of concern because of the presence of DBA in the milk of lactating Sprague Dawley females at concentrations greater than those measured in the females' blood serum ( Klinefelter et al., 2000,

abstract). In contrast, Christian et al. ( 2001) did not detect DBA in the milk samples of lactating Sprague Dawley rats exposed to high DBA drinking water concentrations, although DBA was measurable in fetal plasma on gestation day 21. Christian et al. ( 2001) observed that, although DBA freely crossed the placenta in pregnant Sprague Dawley rats, the attained maternal and fetal plasma levels were associated with the amount and timing of water consumption and did not appear to accumulate. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 1 Draft, do not cite or quote Chapter IV. Human Exposure A. Drinking Water Exposure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

MBA, BCA, and DBA have been identified as drinking water disinfection byproducts under the Information Collection Rule ( U. S. EPA, 1994) and are being assessed for regulatory consideration in the Stage 2 Disinfectants/ Disinfection Byproducts Rule to be promulgated.

Therefore, this section will examine the occurrence of these compounds in drinking water.

A. 1 National Occurrence Data for MBA, BCA, and DBA This section presents the data collected from the Information Collection Rule ( ICR)

databases, which provide data from surface and ground water systems serving at least 100,000 persons. This data base includes information gathered for 18 months from July 1997 to December 1998.

Section A. 1.1 describes the ICR data set and analysis techniques used to present the data for the plants that submitted data under the ICR. The data in Sections A. 1 and A. 2 were taken from the online version of the ICR database ( U. S. EPA, 2000a), and the explanation of the methods used was taken from the Draft EPA Document on Stage 2 Occurrence and Exposure Assessment for Disinfectants and Disinfection Byproducts ( D/ DBPs) in Public Drinking Water ( U. S. EPA, 2000b). Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 2 Draft, do not cite or quote A. 1.1 ICR Plants The ICR generated plant level sets of data that link water quality and treatment from source to tap, and aid in understanding the seasonal variability in these relationships. The database contains information from 18 monthly or 6 quarterly samples from 7/ 97 to 12/ 98 from approximately 300 large systems covering approximately 500 plants. These samples were tested for influent and finished water quality parameters ( e. g., TOC, temperature, pH, alkalinity), DBP levels, and disinfectant residuals. Samples were collected at several locations throughout the distribution system to cover the entire range of residence times during which DBPs can form in the finished water. Over the 18 month period, approximately 1470 samples were taken from 305 plants with surface water as their source, and approximately 580 samples were taken from 123 plants with groundwater as their source. For more detailed information, such as sampling locations and frequencies, refer to the ICR Data Analysis Plan ( U. S. EPA, 2000c).

A. 1.2 Quarterly Distribution System Average and Highest Value for MBA, BCA, and DBA This section describes the data analysis techniques employed for the analysis of observed data for water quality parameters, and for MBA, BCA, and DBA concentrations. All data are categorized according to the types of source water  surface or ground. Plants having both surface and ground water sources ( mixed) or that purchase water are included in the surface Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 3 Draft, do not cite or quote water category. Quarterly Distribution System Average and Highest Value for the brominated acetic acids are presented in Table IV 1.

The quarterly distribution system average is an average of the following four distinct locations in the distribution system.

 Distribution System Equivalent ( DSE) location;

 Average 1 ( AVG 1) and Average 2 ( AVG 2) locations: Two sample points in the distribution system representing the approximate average residence time as designated by the water system; and  Distribution System Maximum: Sample point in the distribution system having the highest residence time ( or approaching the longest time) as designated by the water system The quarterly distribution system highest value is the highest of the four distributionsystem samples collected by a plant in a given quarter. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 4 Draft, do not cite or quote Table IV 1. Bromoacetic Acids Quarterly Distribution System Average and Highest Value Source Quarterly Dist. Sys. Plants N PctND % Mean µ g/ L Median µ g/ L STD µ g/ L Min µ g/ L Max µ g/ L p10 µ g/ L p90 µ g/ L MBA SW Average 305 1470 83.06 0.25 0.00 0.98 ND 26.00 0.00 0.96 High 305 1470 83.06 0.41 0.00 1.84 ND 58.10 0.00 1.50 GW Average 123 581 86.57 0.16 0.00 0.54 ND 6.68 0.00 0.38 High 123 581 86.57 0.27 0.00 0.92 ND 12.00 0.00 1.30 BCA SW Average 305 1474 9.70 3.61 2.88 3.08 ND 24.18 0.25 7.70 High 305 1474 9.70 4.45 3.50 3.86 ND 41.90 1.00 9.00 GW Average 123 584 48.63 1.47 0.28 2.15 ND 11.28 0.00 4.50 High 123 584 48.63 2.18 1.10 3.40 ND 41.00 0.00 6.40 DBA SW Average 305 1484 60.58 1.09 0.00 2.08 ND 14.25 0.00 3.68 High 305 1484 60.58 1.39 0.00 2.52 ND 19.00 0.00 4.30 GW Average 123 584 56.16 0.82 0.00 1.48 ND 13.00 0.00 2.58 High 123 584 56.16 1.20 0.00 1.88 ND 16.00 0.00 3.70 1 Nondetects are treated as zero. Source: SW  Surface Water, GW  Groundwater Quarterly Dist. Sys: Quarterly Distribution System Average Plants: Number of plants sampled N: Number of samples PctND: Percent samples nondetect Mean: Arithmetic mean of all samples Median: Median value of all samples STD: Standard deviation Min: Minimum Value Max: Maximum Value p10: 10th percentile p90: 90th percentile ND: Nondetected Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 5 Draft, do not cite or quote The mean concentrations of MBA ( averaged across the four sampling locations) were 0.16 and 0.25  g/ L in groundwater and surface water, respectively. The mean concentrations of BCA ( averaged across the four sampling locations) were 1.47 and 3.61  g/ L in groundwater and surface water, respectively. The mean concentrations of DBA ( averaged across the four sampling locations) were 0.82 and 1.09  g/ L in groundwater and surface water, respectively. Examination of the data using the Student's t test indicates that the mean concentrations of MBA, BCA, and DBA in surface water was significantly higher that the mean concentrations of these chemicals in ground water. The mean concentrations of BCA were statistically significantly higher ( p = 0.05)

than the mean concentrations of DBA, which were significantly higher ( p = 0.05), that the mean MBA concentrations in both surface and ground water. The lowest mean concentrations are associated with the highest percentage of nondetects, which are treated as 0 in the calculation of the mean, median, standard deviation, and p10 values ( U. S. EPA, 2000a).

A. 2 Factors Affecting the Relative Concentrations of MBA, BCA, and DBA in Drinking Water Sections A. 2.1  A. 2.4 contain investigational information and ICR data on the effects of disinfection chemicals, influent bromide concentration, influent total organic carbon ( TOC)

concentration, and seasonal shifts, respectively in MBA, BCA, and DBA concentrations. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 6 Draft, do not cite or quote A. 2.1 Disinfection Treatment Chlorination has been the predominant water disinfection method in the United States.

However, water utilities are considering a shift to alternative disinfectants. Therefore, there is a need to understand the occurrence of DBPs in drinking water and the factors that may influence their formation. Several published studies ( Boorman et al., 1999; Richardson, 1998; Lykins et al.,

1994; Jacangelo et al., 1989) reported on the formation of brominated acetic acids and other DBPs under different disinfection conditions.

In a review on drinking water disinfection byproducts, Boorman et al. ( 1999) compared the concentrations of different drinking water disinfection byproducts, including MBA, BCA, and DBA, formed by chlorination, ozonation, chlorine dioxide, and chloramination. Most of the data were available for surface water systems that used chlorination. For the systems using chlorination, BCA, with a median and a maximum concentration of 3.2 and 49 µ g/ L, respectively,

was present at the highest concentrations. The median values of both MBA and DBA in chlorinated water were less than 0.5 µ g/ L, with maximum values of 1.7 and 7.4 µ g/ L,

respectively. The principal products formed by chloramination were similar to those formed by chlorination; additional information was not provided. Ozonation of water containing bromide may produce DBPs such as bromoform, dibromoacetic acid, cyanogen bromide, and bromate. The total concentration of brominated acetic acids formed by the use of ozonation ranged from 1 to 50 µ g/ L; concentrations for individual compounds were not provided. Chlorine dioxide formed oxidation by products similar to those formed by ozonation; additional details were not provided. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 7 Draft, do not cite or quote Richardson ( 1998) compared the relative concentrations of DBPs in drinking water using different treatment methods, and found that chlorination produced the highest concentration of DBPs, including MBA, BCA, and DBA. Chlorine dioxide and chloramine, when compared to chlorine, produced fewer and lower concentrations of DBPs. MBA, BCA, and DBA were not produced by chlorine dioxide in measurable quantities. Compared to chlorine treatment,

chloramine produced 3% to 20% lower levels of by products, including haloacetic acids.

Ozonation produced insignificant levels of trihalomethanes ( THMs). However, when elevated levels of bromide ion were in the raw water, MBA and DBA were detected following ozonation.

When ozone was the primary disinfectant followed by chloramine, the levels of most DBPs,

including haloacetic acids, were lower than when chloramine was used solely. However, there was an observed shift to more brominated species of THMs and haloacetic acids when ozone was followed by chlorine, than when chorine was used solely.

Lykins et al. ( 1994) investigated the formation of halogenated DBPs in the waterdistribution system, by predisinfecting and postdisinfecting the water with either chlorine or chloramine and holding the water for five days. They found that the use of chlorine produced the highest concentration of halogenated DBPs and that, in general, the concentrations could be reduced by adding ozone as a predisinfectant with postchlorination. Lykins et al. ( 1994) also found that the highest average concentrations of MBA ( 1.2  g/ L) and BCA ( 18  g/ L) were formed when chlorine was the sole treatment method. The average DBA concentration was 0.6  g/ L when chlorine was the sole treatment method. The next highest concentrations of MBA ( 1.0  g/ L) and BCA ( 14  g/ L) were observed with ozone treatment followed by chlorine. In contrast, Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 8 Draft, do not cite or quote the average DBA concentration ( 1.0  g/ L) was slightly higher with ozone treatment followed by chlorine than when chlorine was the sole treatment method. Chloramine treatment, and ozone treatment followed by chloramine, resulted in the lowest concentrations of MBA and DBA, with both treatment methods resulting in 0.1  g/ L MBA and 0.1  g/ L DBA. BCA concentrations were lower with ozone treatment followed by chloramine ( 1.0  g/ L BCA) than when chloramine was the sole treatment method ( 1.9  g/ L).

Jacangelo et al. ( 1989) examined the impact of ozonation on the formation and control of DBPs in drinking water at four utilities. Treatment modifications were made on the process train at each full or pilot scale plant to incorporate ozone in the treatment process. For two of the utilities in the Jacangelo et al. ( 1989) study, only total haloacetic acids ( HAAs) were measured,

and no measurements were made of individual HAAs. The disinfection schemes that employed ozonation followed by chloramines as a disinfectant resulted in large decreases in HAAs relative to chlorination. However, the sample size did not allow for statistical analysis of the data ( Jacangelo et al., 1989). For two other utilities that measured individual HAAs, preozonation followed by chlorination decreased the total HAAs by 14 to 50%, when compared with chlorination only. The concentration of MBA was essentially the same with and without preozonation, while DBA increased with ozonation. BCA was not measured in this study. The authors suggested that ozone reacts with bromide ions in the source water, resulting in the formation of hypobromous acid. Reaction of hypobromous acid and natural organic matter can produce brominated HAAs. When preozonation and postchlorination are practiced, competition exists between hypochlorous acid and hypobromous acid for organic matter, leading to varying Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 9 Draft, do not cite or quote concentrations of chlorinated and brominated HAAs ( Jacandgelo et al., 1989). In addition,

Jacangelo et al. ( 1989) noted that the concentrations of brominated acetic acids increased with increasing bromide concentrations in the source water.

Miltner et al. ( 199) studied DBP formation and control in three surface water pilot plants employing three different disinfectant methods ( chlorine, ozone followed by chlorine, and ozone followed by chloramine). In an examination of the data using the Student's t test, the authors found that the amount of BCA measured in finished water and in simulated distribution waters was lower ( p = 0.05) when ozonation was combined with chlorination or with cloramination than when chlorination was used a lone. However, ozonation had no effect ( p = 0.05) on the formation of MBA, and the formation of DBA was higher ( p = 0.05) when ozonation was followed by chlorination than when chlorination alone was used.

A. 2.1.1 Disinfection Treatment in ICR Database Data on the concentrations of MBA, BCA, and DBA were gathered from plants using several disinfection treatments. Those chemical disinfection treatments most commonly used ( used by 10% or more of the plants evaluated), along with the ozonation treatments, are presented in Tables IV 2, IV 3, and IV 4 for MBA, BCA, and DBA, respectively.

Examination of the data using the Student's t test indicates that, for all chemicaldisinfection treatments used for surface water, the mean concentrations of BCA were statistically significantly higher ( p = 0.05) than the mean concentrations of DBA which were significantly Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 10 Draft, do not cite or quote higher ( p = 0.05) than the mean concentrations ( Tables IV 2 and IV 3). For all chemicaldisinfection treatments used for ground water, the mean concentrations of BCA were significantly higher ( p = 0.05) than the mean concentrations of MBA. The mean concentrations of BCA in groundwater were significantly higher ( p = 0.05) than the mean DBA concentrations only in plants using ozone and choramine. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 11 Draft, do not cite or quote Table IV 2. MBA by Disinfection Method ( Quarterly Distribution System Average)

Source Disinfection Chemicals Plants N PctND % Mean µ g/ L Median µ g/ L STD µ g/ L Min µ g/ L Max µ g/ L p10 µ g/ L p90 µ g/ L SW Cl 2/ Cl 2 180 814 85.63 0.24 0.00 1.14 ND 26.00 0.00 0.75 Cl 2_ CLM/ CLM 66 307 80.78 0.27 0.00 0.72 ND 5.73 0.00 1.10 O 3/ CL 2 7 25 88.00 0.11 0.00 0.33 0.00 1.28 0.00 0.28 O 3/ CLM 10 49 85.71 0.10 0.00 0.30 0.00 1.48 0.00 0.58 GW / Cl 2 67 299 87.29 0.18 0.00 0.64 ND 6.68 0.00 0.35 Cl 2/ Cl 2 39 170 85.88 0.15 0.00 0.45 ND 2.53 0.00 0.51 O 3/ CLM 1 6 100.0 0 0.00 0.00 0.00 0.00 0.00 0.00 0.00 1 Nondetects are treated as zero. Source: SW  Surface Water, GW  Groundwater Cl2/ Cl2: Free chlorine in Water Treatment Plant ( WTP) and Distribution System ( DS).

Cl2_ CLM/ CLM: Free chlorine followed by chloramine in WTP and chloramine in DS. / Cl2: No disinfectant in WTP and free chlorine in DS.

O3/ Cl2: Ozone in WTP and free chlorine in DS.

O3/ CLM: Ozone in WTP and chloramine in DS. Plants: Number of plants sampled N: Number of samples PctND: Percent samples nondetect Mean: Arithmetic mean of all samples Median: Median value of all samples STD: Standard deviation Min: Minimum Value Max: Maximum Value p10: 10th percentile p90: 90th percentile Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 12 Draft, do not cite or quote Table IV 3. BCA Acid by Disinfection Method ( Quarterly Distribution System Average)

Source Disinfection Chemicals Plants N PctND % Mean µ g/ L Median µ g/ L STD µ g/ L Min µ g/ L Max µ g/ L p10 µ g/ L p90 µ g/ L SW Cl 2/ Cl 2 180 816 11.40 3.14 2.54 2.77 ND 18.73 0.00 6.83 Cl 2_ CLM/ CLM 66 306 1.63 4.67 4.01 3.28 ND 23.80 1.35 8.83 O 3/ CL 2 7 25 4.00 2.54 2.13 1.77 0.00 5.68 0.33 4.90 O 3/ CLM 10 49 22.45 2.19 2.15 1.70 0.00 6.60 0.00 4.55 GW / Cl 2 67 301 65.78 0.87 0.00 1.77 ND 10.70 0.00 2.70 Cl 2/ Cl 2 39 170 45.53 1.27 0.31 1.80 ND 7.88 0.00 3.78 O 3/ CLM 1 6 0.00 2.68 2.55 0.69 2.05 3.88 2.05 3.88 1 Nondetects are treated as zero. Source: SW  Surface Water, GW  Groundwater Cl2/ Cl2: Free chlorine in Water Treatment Plant ( WTP) and Distribution System ( DS).

Cl2_ CLM/ CLM: Free chlorine followed by chloramine in WTP and chloramine in DS. / Cl2: No disinfectant in WTP and free chlorine in DS.

O3/ Cl2: Ozone in WTP and free chlorine in DS.

O3/ CLM: Ozone in WTP and chloramine in DS. Plants: Number of plants sampled N: Number of samples PctND: Percent samples nondetect Mean: Arithmetic mean of all samples Median: Median value of all samples STD: Standard deviation Min: Minimum Value Max: Maximum Value p10: 10th percentile p90: 90th percentile Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 13 Draft, do not cite or quote Table IV 4. DBA by Disinfection Method ( Quarterly Distribution System Average)

Source Disinfection Chemicals Plants N PctND % Mean µ g/ L Median µ g/ L STD µ g/ L Min µ g/ L Max µ g/ L p10 µ g/ L p90 µ g/ L SW Cl 2/ Cl 2 180 823 68.53 0.68 0.00 1.55 ND 12.50 0.00 2.20 Cl 2_ CLM/ CLM 66 308 51.62 1.36 0.00 2.11 ND 14.25 0.00 4.83 O 3/ CL 2 7 25 68.00 1.02 0.00 2.27 0.00 8.15 0.00 2.83 O 3/ CLM 10 49 46.94 1.01 0.25 1.50 0.00 7.30 0.00 2.80 GW / Cl 2 67 303 67.66 0.68 0.00 1.52 ND 13.00 0.00 2.53 Cl 2/ Cl 2 39 169 46.15 1.01 0.25 1.62 ND 7.25 0.00 3.15 O 3/ CLM 1 6 50.00 0.41 0.14 0.55 0.00 1.25 0.00 1.25 1 Nondetects are treated as zero. Source: SW  Surface Water, GW  Groundwater Cl2/ Cl2: Free chlorine in Water Treatment Plant ( WTP) and Distribution System ( DS).

Cl2_ CLM/ CLM: Free chlorine followed by chloramine in WTP and chloramine in DS. / Cl2: No disinfectant in WTP and free chlorine in Distribution System.

O3/ Cl2: Ozone in WTP and free chlorine in DS.

O3/ CLM: Ozone in WTP and chloramine in DS. Plants: Number of plants sampled N: Number of samples PctND: Percent samples nondetect Mean: Arithmetic mean of all samples Median: Median value of all samples STD: Standard deviation Min: Minimum Value Max: Maximum Value p10: 10th percentile p90: 90th percentile Examination of the ICR data using the Student's t test indicates that the mean concentrations of BCA and DBA were significantly higher ( p = 0.05) in surface water plants using chlorine followed by chloramine than in those using free chlorine alone. There were no significant Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 14 Draft, do not cite or quote differences ( p = 0.05) between the mean concentrationof MBA in surface water plants using chlorine followed by chloramine and the concentration those solely using chlorine.

In plants with groundwater as a source, the mean concentrtions of BCA and DBA in plants with no disinfectant in the treatment plant and with free chlorine in the distribution system were significantly lower ( p = 0.05) than the mean concentrations of the same chemicals in plants with free chlorine in both the treatment plant and the distribution system. There were no significant differences ( p = 0.05) between the mean concentrtions of MBA in plants with no disinfectant in the treatment plant and with free chlorine in the distribution system and those with free chlorine in both the treatment plant and the distribution system.

Only a very limited number of plants used ozonation in combination with either chlorine or chloramine in the distribution system. An examination of the ICR data in Tables IV 2, IV 3 and IV 4 using the Student's t test indicates that, with one exception, the mean concentrations of BCA were significantly lower in surface water plants that use ozone in the water treatment plant and free chlorine or free chloramine in the distribution system than in plants using non ozonation disinfection processes. This finding was also presented by Lykins et al. ( 1994). However , there were no significant differences ( p = 0.05) between the mean concentrations of BCA, in surfacewater plants when free chlorine was used solely, and the BCA concentrtions when both ozone and chlorine were used. There were no significant differences ( p = 0.05) between the concentrations of MBA and DBA in surface water plants using common ( non ozonation) disinfection processes and the concentrations of the same chemicals pin plants with ozone in the water treatment plant and free chlorine or free chloramine in the distribution system. In addition, there were no Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 15 Draft, do not cite or quote significant differences ( p = 0.05) for MBA, BCA, or DBA between the two treatments using ozonation in treating surface water.

There was only 1 groundwater plant that used the ozone/ chloramine disinfection method and statistical analysis was not conducted.

In summary, an analysis of the ICR data suggest that although BCA concentrations in surface water treated with chlorine are similar to those treated with ozone and chlorine, surface water plants using ozonation had lower BCA concentrations than those using most common (

nonozonation disinfection processes. In addition, ozonation appeared to have no effect on the formation of MBA and DBA.

A. 2.2 Bromide Concentration Pourmoghaddas et al. ( 1993) examined the effects of source water and treatment characteristics, such as pH, reaction time, chlorine dosage, and bromide ion concentration, on the formation of HAAs.

The study quantified nine HAA species in the presence of bromide ion at low, neutral, and high pH over time at two chlorine dosages. This study found a shift in the distribution of HAAs from chlorinated to brominated and mixed ( bromochlorinated) halogenated species with increased bromide ion concentration. Chloride ion concentration had no observed effect on the formation of brominated HAAs. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 16 Draft, do not cite or quote At the low chlorine dose of 11.5 mg/ L, MBA showed a consistent trend toward higher concentrations as reaction time and bromide ion concentrations increased. The only apparent effect of pH was to increase the amount of MBA at the highest bromide concentration ( 4.5 mg/ L). The highest measured concentration of MBA was at a pH of 5 and a bromide concentration of 4.5 mg/ L ( Pourmoghaddas et al., 1993). DBA concentrations increased with increasing bromide ion concentration and reaction time. Changes in pH had little influence on the formation of DBA ( Pourmoghaddas et al., 1993).

BCA was formed only if the bromide ion was present. BCA concentrations increased with reaction time and were significantly lower at pH 9.4. The highest observed concentration of BCA was at 1.5 mg/ L bromide ion concentration, while BCA levels decreased at the highest bromide concentration of 4.5 mg/ L ( Pourmoghaddas et al., 1993). It is possible that the decreased BCA levels at high bromide concentrations reflect the preferential formation of DBA over BCA under such conditions, but the data provided are insufficient to test this hypothesis.

A. 2.2.1 Bromide Concentration in ICR Database Tables IV 5, IV 6, and IV 7 present the formation of MBA, BCA, and DBA, respectively,

as a function of influent bromide concentrations.

Bromide concentrations tended to be lower in plants using surface water as a source than in those using groundwater as a source. For example, 114 of the 305 plants using surface water as the source ( 37%) had influent bromide levels below the minimal reporting limit ( MRL) of 20 ppb, Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 17 Draft, do not cite or quote while only 13 of the 123 plants using groundwater as the source ( 11%) had influent bromide levels below the MRL. Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 18 Draft, do not cite or quote Table IV 5. MBA by Influent Bromide Concentration ( Quarterly Distribution System Average)

Source Influent Bromide Conc. ( ppb) Plants N PctND % Mean µ g/ L Median µ g/ L STD µ g/ L Min µ g/ L Max µ g/ L p10 µ g/ L p90 µ g/ L SW < MRL ( 20) 114 556 90.83 0.14 0.00 0.59 ND 6.63 0.00 0.00 20  < 30 41 200 88.50 0.21 0.00 0.72 ND 4.03 0.00 0.38 30  < 50 48 221 83.26 0.45 0.00 2.02 ND 26.00 0.00 1.13 50  < 100 59 282 77.30 0.24 0.00 0.57 ND 3.95 0.00 1.03

100 39 192 64.06 0.41 0.00 1.12 ND 7.20 0.00 1.90 GW < MRL ( 20) 13 66 93.94 0.07 0.00 0.30 ND 1.60 0.00 0.00 20  < 30 11 50 98.00 0.01 0.00 0.04 ND 0.30 0.00 0.00 30  < 50 26 109 96.33 0.03 0.00 0.18 ND 1.45 0.00 0.00 50  < 100 32 148 83.11 0.18 0.00 0.67 ND 6.68 0.00 0.63

100 41 208 78.85 0.27 0.00 0.66 ND 3.65 0.00 1.20 1 Nondetects are treated as zero. Source: SW  Surface Water, GW  Groundwater Plants: Number of plants sampled N: Number of samples PctND: Percent samples nondetect Mean: Arithmetic mean of all samples Median: Median value of all samples STD: Standard deviation Min: Minimum Value Max: Maximum Value p10: 10th percentile p90: 90th percentile MRL: Minimum Reporting Limit ND: Nondetect Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 19 Draft, do not cite or quote Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 20 Draft, do not cite or quote Table IV 6. BCA by Influent Bromide Concentration ( Quarterly Distribution System Average)

Source Influent Bromide Conc. ( ppb) Plants N PctND % Mean µ g/ L Median µ g/ L STD µ g/ L Min µ g/ L Max µ g/ L p10 µ g/ L p90 µ g/ L SW < MRL ( 20) 114 559 22.00 1.76 1.65 1.79 ND 23.80 0.00 3.43 20  < 30 41 197 0.51 3.32 3.03 1.67 ND 12.28 1.63 5.30 30  < 50 48 225 3.11 3.88 3.40 2.94 ND 22.50 0.93 7.18 50  < 100 59 282 3.90 5.43 4.96 3.26 ND 15.50 1.48 9.68

100 39 192 0.52 6.29 6.01 3.52 ND 24.18 2.60 11.20 GW < MRL ( 20) 13 66 78.79 0.35 0.00 1.34 ND 10.25 0.00 0.75 20  < 30 11 51 58.52 0.69 0.00 1.19 ND 6.00 0.00 2.25 30  < 50 26 108 49.07 1.16 0.25 1.61 ND 7.05 0.00 3.58 50  < 100 32 150 48.67 1.22 0.26 1.86 ND 7.88 0.00 4.23

100 41 209 36.36 2.35 1.83 2.60 ND 11.28 0.00 6.25 1 Nondetects are treated as zero. Source: SW  Surface Water, GW  Groundwater Plants: Number of plants sampled N: Number of samples PctND: Percent samples nondetect Mean: Arithmetic mean of all samples Median: Median value of all samples STD: Standard deviation Min: Minimum Value Max: Maximum Value p10: 10th percentile p90: 90th percentile MRL: Minimum Reporting Limit ND: Nondetect Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 21 Draft, do not cite or quote Table IV 7. DBA by Influent Bromide Concentration ( Quarterly Distribution System Average)

Source Influent Bromide Conc. ( ppb) Plants N PctND % Mean µ g/ L Median µ g/ L STD µ g/ L Min µ g/ L Max µ g/ L p10 µ g/ L p90 µ g/ L SW < MRL ( 20) 114 561 91.62 0.08 0.00 0.32 ND 3.00 0.00 0.00 20  < 30 41 201 67.66 0.29 0.00 0.59 ND 3.48 0.00 1.10 30  < 50 48 226 58.41 0.66 0.00 1.10 ND 9.58 0.00 2.03 50  < 100 59 284 32.04 1.70 1.33 1.84 ND 7.65 0.00 4.30

100 39 193 6.74 4.37 3.78 3.22 ND 14.25 0.58 8.48 GW < MRL ( 20) 13 67 88.06 0.09 0.00 0.31 ND 1.68 0.00 0.28 20  < 30 11 50 90.00 0.12 0.00 0.49 ND 2.45 0.00 0.13 30  < 50 26 109 69.72 0.27 0.00 0.54 ND 2.15 0.00 1.10 50  < 100 32 150 42.00 1.00 0.40 1.27 ND 5.83 0.00 3.10

100 41 208 40.87 1.38 0.60 2.00 ND 13.00 0.00 3.73 1 Nondetects are treated as zero. Source: SW  Surface Water, GW  Groundwater Plants: Number of plants sampled N: Number of samples PctND: Percent samples nondetect Mean: Arithmetic mean of all samples Median: Median value of all samples STD: Standard deviation Min: Minimum Value Max: Maximum Value p10: 10th percentile p90: 90th percentile MRL: Minimum Reporting Limit ND: Nondetect Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 22 Draft, do not cite or quote A regression analysis of the ICR data indicates that, with the exception of MBA in surface water, there was a significant correlation (   = 0.05) between influent bromide concentration and the mean concentrations of BCA and DBA in surface water and groundwater.( Tables IV 5, IV 6 and IV 7).

Tan examination of th data using the Student's t test indicates that in plants treating surface water, the mean concentrations of BCA were significantly higher ( p = 0.05_ than the mean MBA and DBA concentrtions, for a given bromide concentration range ( Tables IV 5, IV 6,

and IV 7). In addition, the mean concentrations of DBA were significantly lower ( p = 0.05) than the mean MBA concentrations at the lowest influent bromide concentration (< minimum reporting limit of 20 ppb) and higher than MBA concentrations at the two highest influent bromide concentrations ( ranging from 50 to > 100 ppb ( Tables IV 5 and IV 7).

The mean concentrations of BCA in surface water were statistically significantly higher ( at p = 0.05) than the mean concentrations of BCA in groundwater at influent bromide concentrations of 20  < 30 ppb and at 50  < 100 ppb. There were no statistically significant differences ( at p = 0.05) between the mean concentrations of DBA and MBA in surface water and the mean concentrations of these chemicals in groundwater.

The mean concentrations of BCA and DBA are significantly higher in surface water than their mean concentrations in groundwater for a given influent bromide concentration ( with the exception of DBA at influent bromide concentrations at < 20ppb). However, the mean MBA concentrations in surface water were significantly higher ( p = 0.05) than the mean concentrations Drinking Water Criteria Document for Brominated Acetic Acids EPA/ OW/ OST/ HECD IV 23 Draft, do not cite or quote of MBA in groundwater only for the influent bromide concentrations ranging from 20 ppb to < 50 ppb.

A. 2.3 Total Organic Carbon ( TOC) Concentration in ICR Database Many researchers have documented that chlorine reacts with natural organic matter in water to produce a variety of DBPs, including trihalomethanes and haloacetic acids ( Reckhow and Singer, 1990; Reckhow et al., 1990; Marhaba and Van, 2000). Natural organic matter in source water is generally monitored as total organic carbon ( TOC). Arora et al. ( 1997) analyzed results of a DBP survey and a two year DBP monitoring study of more than 100 treatment plants of the American Water System from 1989 to 1991, and reported no correlation between rawwater TOC and the total of 5 haloacetic acid ( HAA5) concentrations in finished and distributedwater samples. A significant correlation ( p < 0.01) was found between TOC and HAA5 in plant effluent and distributed water samples. However, only 11 and 15 percent of the variation in HAA5 was explained by TOC for the distributed water samples and plant effluent, respectively. No published studies were located that examined the effect of TOC on the concentration of brominated acids.