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Review
Occurrence, genotoxicity, and carcinogenicity of regulated and
emerging disinfection by-products in drinking water:
A review and roadmap for research
Susan D. Richardson
a,
*, Michael J. Plewa
b
, Elizabeth D. Wagner
b
,
Rita Schoeny
c
, David M. DeMarini
d
a
National Exposure Research Laboratory, U.S. Environmental Protection Agency, Athens, GA 30605, USA
b
Department of Crop Sciences, College of Agricultural, Consumer, and Environmental Sciences,
University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA
c
Office of Water, U.S. Environmental Protection Agency, ML4301, 1200 Pennsylvania Avenue NW, Washington, DC 20460, USA
d
National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency,
Research Triangle Park, NC 27711, USA
Received 7 May 2007; received in revised form 5 September 2007; accepted 6 September 2007
Available online 12 September 2007
Abstract
Disinfection by-products (DBPs) are formed when disinfectants (chlorine, ozone, chlorine dioxide, or chloramines) react with
naturally occurring organic matter, anthropogenic contaminants, bromide, and iodide during the production of drinking water. Here
we review 30 years of research on the occurrence, genotoxicity, and carcinogenicity of 85 DBPs, 11 of which are currently regulated
by the U.S., and 74 of which are considered emerging DBPs due to their moderate occurrence levels and/or toxicological properties.
These 74 include halonitromethanes, iodo-acids and other unregulated halo-acids, iodo-trihalomethanes (THMs), and other
unregulated halomethanes, halofuranones (MX [3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone] and brominated MX
DBPs), haloamides, haloacetonitriles, tribromopyrrole, aldehydes, and N-nitrosodimethylamine (NDMA) and other nitrosamines.
Alternative disinfection practices result in drinking water from which extracted organic material is less mutagenic than extracts of
chlorinated water. However, the levels of many emerging DBPs are increased by alternative disinfectants (primarily ozone or
chloramines) compared to chlorination, and many emerging DBPs are more genotoxic than some of the regulated DBPs. Our
analysis identified three categories of DBPs of particular interest. Category 1 contains eight DBPs with some or all of the toxicologic
characteristics of human carcinogens: four regulated (bromodichloromethane, dichloroacetic acid, dibromoacetic acid, and
bromate) and four unregulated DBPs (formaldehyde, acetaldehyde, MX, and NDMA). Categories 2 and 3 contain 43 emerging
DBPs that are present at moderate levels (sub- to low-mg/L): category 2 contains 29 of these that are genotoxic (including chloral
hydrate and chloroacetaldehyde, which are also a rodent carcinogens); category 3 contains the remaining 14 for which little or no
toxicological data are available. In general, the brominated DBPs are both more genotoxic and carcinogenic than are chlorinated
compounds, and iodinated DBPs were the most genotoxic of all but have not been tested for carcinogenicity. There were
toxicological data gaps for even some of the 11 regulated DBPs, as well as for most of the 74 emerging DBPs. A systematic
assessment of DBPs for genotoxicity has been performed for 60 DBPs for DNA damage in mammalian cells and 16 for
mutagenicity in Salmonella. A recent epidemiologic study found that much of the risk for bladder cancer associated with drinking
water was associated with three factors: THM levels, showering/bathing/swimming (i.e., dermal/inhalation exposure), and
www.elsevier.com/locate/reviewsmr
Community address: www.elsevier.com/locate/mutres
A
vailable online at www.sciencedirect.com
Mutation Research 636 (2007) 178–242
* Corresponding author. Tel.: +1 706 355 8304; fax: +1 706 355 8302.
E-mail address: richardson.susan@epa.gov (S.D. Richardson).
1383-5742/$ – see front matter #2007 Elsevier B.V. All rights reserved.
doi:10.1016/j.mrrev.2007.09.001
genotype (having the GSTT1-1 gene). This finding, along with mechanistic studies, highlights the emerging importance of dermal/
inhalation exposure to the THMs, or possibly other DBPs, and the role of genotype for risk for drinking-water-associated bladder
cancer. More than 50% of the total organic halogen (TOX) formed by chlorination and more than 50% of the assimilable organic
carbon (AOC) formed by ozonation has not been identified chemically. The potential interactions among the 600 identified DBPs in
the complex mixture of drinking water to which we are exposed by various routes is not reflected in any of the toxicology studies of
individual DBPs. The categories of DBPs described here, the identified data gaps, and the emerging role of dermal/inhalation
exposure provide guidance for drinking water and public health research.
#2007 Elsevier B.V. All rights reserved.
Keywords: N-Nitrosodimethylamine; Regulated and unregulated DBPs; Total organic halogen; Total organic carbon
Contents
1. Introduction . ...................................................................... 180
2. Overview of DBP regulations in the United States............................................. 183
3. Summary of epidemiology studies of cancer and drinking water . . . ................................ 185
4. Occurrence, genotoxicity, and carcinogenicity of the regulated DBPs................................ 185
4.1. Trihalomethanes (THMs). ......................................................... 185
4.1.1. Occurrence ............................................................. 185
4.1.2. Genotoxicity . . . ......................................................... 186
4.1.3. Carcinogenicity . ......................................................... 188
4.2. Haloacetic acids (HAAs) . ......................................................... 191
4.2.1. Occurrence ............................................................. 191
4.2.2. Genotoxicity . . . ......................................................... 191
4.2.3. Carcinogenicity . ......................................................... 193
4.3. Bromate . . . .................................................................. 193
4.3.1. Occurrence ............................................................. 193
4.3.2. Genotoxicity . . . ......................................................... 193
4.3.3. Carcinogenicity . ......................................................... 194
4.4. Chlorite...................................................................... 194
4.4.1. Occurrence ............................................................. 194
4.4.2. Genotoxicity . . . ......................................................... 195
4.4.3. Carcinogenicity . ......................................................... 195
5. Summary of the occurrence, genotoxicity, and carcinogenicity of the regulated DBPs .................... 195
5.1. Summary of the occurrence of the regulated DBPs. . . ..................................... 195
5.2. Summary of the genotoxicity of the regulated DBPs. . ..................................... 195
5.3. Summary of the carcinogenicity of the regulated DBPs..................................... 195
5.4. Overall summary of the regulated DBPs . . ............................................. 196
6. Emerging unregulated DBPs . . . ......................................................... 197
6.1. Halonitromethanes .............................................................. 197
6.1.1. Occurrence ............................................................. 197
6.1.2. Genotoxicity . . . ......................................................... 198
6.1.3. Carcinogenicity . ......................................................... 199
6.2. Iodo-acids and other unregulated halo-acids............................................. 199
6.2.1. Occurrence ............................................................. 199
6.2.2. Genotoxicity . . . ......................................................... 202
6.2.3. Carcinogencity . . ......................................................... 202
6.3. Iodo-THMs and other unregulated halomethanes ......................................... 202
6.3.1. Occurrence ............................................................. 202
6.3.2. Genotoxicity . . . ......................................................... 203
6.3.3. Carcinogenicity . ......................................................... 204
6.4. MX and BMX compounds (halofuranones) ............................................. 205
6.4.1. Occurrence ............................................................. 205
6.4.2. Genotoxicity . . . ......................................................... 205
6.4.3. Carcinogenicity . ......................................................... 207
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242 179
6.5. Haloamides . . . ................................................................ 208
6.5.1. Occurrence . ............................................................ 208
6.5.2. Genotoxicity ............................................................ 208
6.5.3. Carcinogenicity . . . ....................................................... 209
6.6. Haloacetonitriles................................................................ 209
6.6.1. Occurrence . ............................................................ 209
6.6.2. Genotoxicity ............................................................ 209
6.6.3. Carcinogenicity . . . ....................................................... 211
6.7. Tribromopyrrole ................................................................ 211
6.7.1. Occurrence . ............................................................ 211
6.7.2. Genotoxicity ............................................................ 211
6.7.3. Carcinogenicity . . . ....................................................... 211
6.8. Nitrosodimethylamine (NDMA) and other nitrosamines . ................................... 211
6.8.1. Occurrence . ............................................................ 211
6.8.2. Genotoxicity ............................................................ 213
6.8.3. Carcinogenicity . . . ....................................................... 216
6.9. Aldehydes .................................................................... 217
6.9.1. Occurrence . ............................................................ 217
6.9.2. Genotoxicity ............................................................ 217
6.9.3. Carcinogenicity . . . ....................................................... 218
6.10. Chlorate . .................................................................... 219
6.10.1. Occurrence . ............................................................ 219
6.10.2. Genotoxicity ............................................................ 219
6.10.3. Carcinogenicity . . . ....................................................... 219
7. Summary of the occurrence, genotoxicity, and carcinogenicity of the emerging unregulated DBPs . .......... 219
7.1. Summary of the occurrence of the emerging unregulated DBPs . .............................. 219
7.2. Summary of the genotoxicity of the emerging unregulated DBPs .............................. 220
7.3. Summary of the carcinogenicity of the emerging unregulated DBPs . . .......................... 221
8. DBPs formed from anthropogenic contaminants . . . ........................................... 223
9. Mutagenicity of organic extracts or concentrates of drinking water . . . .............................. 224
10. Carcinogenicity of raw waters or organic extracts of drinking water or mixtures of DBPs . . . .............. 224
11. Risk assessment of DBPs . . ............................................................ 225
12. Conclusions and research needs from current analysis . . . ....................................... 226
12.1. Categories of DBPs to prioritize testing and aid in decision-making . . .......................... 226
12.1.1. DBPs that have some or all of the toxicologic characteristics of human carcinogens .......... 227
12.1.2. Emerging DBPs with moderate occurrence that are genotoxic .......................... 228
12.1.3. Emerging DBPs with moderate occurrence and no toxicology data ...................... 228
12.2. Systematic generation of quantitative genotoxicity data for classes of DBPs ...................... 229
12.3. Studies on the route of exposure and the role of genotype................................... 229
12.4. Chemical identification of the unknown fraction of drinking water . . . .......................... 231
12.5. Evaluate DBPs from alternative disinfection methods . . . ................................... 231
12.6. Evaluate source-water contamination . . ............................................... 231
12.7. Complex mixture studies . . . ....................................................... 231
Acknowledgement . . . ................................................................ 232
References ........................................................................ 232
1. Introduction
Water disinfection is one of the most important
public health advances of the last century; its
introduction in the U.S. reduced cholera incidence by
90%, typhoid by 80%, and amoebic dysentery by 50%
[1]. Millions of people worldwide receive quality
drinking water every day from their public water
systems. However, chemical disinfection has also raised
a public health issue: the potential for cancer and
reproductive/developmental effects associated with
chemical disinfection by-products (DBPs).
Chemical disinfectants are effective for killing
harmful microorganisms in drinking water, but they
are also powerful oxidants, oxidizing the organic
matter, anthropogenic contaminants, and bromide/
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242180
iodide naturally present in most source waters (rivers,
lakes, and many groundwaters). Chlorine, ozone,
chlorine dioxide, and chloramines are the most common
disinfectants in use today; each produces its own suite
of DBPs in drinking water, with overlapping constitu-
ents [2]. Most developed nations have published
regulations or guidelines to control DBPs and minimize
consumers’ exposure to potentially hazardous chemi-
cals while maintaining adequate disinfection and
control of targeted pathogens.
Scientists first became aware of DBPs only in the
early 1970s. In 1974, Rook and others reported the
identification of the first DBPs in chlorinated drinking
water: chloroform and other trihalomethanes (THMs)
[3,4]. In 1976, the U.S. Environmental Protection
Agency (U.S. EPA) published the results of a national
survey that showed that chloroform and the other THMs
were ubiquitous in chlorinated drinking water [5]. In the
same year, the National Cancer Institute published
results showing that chloroform was carcinogenic in
laboratory animals [6]. In addition, the first reports
appeared in the late 1970s showing that organic extracts
of drinking water were mutagenic in the Salmonella
mutagenicity assay [7]. As a result of these observa-
tions, an important public health issue was recognized.
In the 30 years since the THMs were identified as
DBPs in drinking water, significant research efforts have
been directed toward increasing our understanding of
DBP formation, occurrence, and health effects [2,8–17].
Although more than 600 DBPs have been reported in the
literature [2,18], only a small number has been assessed
either in quantitative occurrence or health-effects studies.
The DBPs that have been quantified in drinking water
are generally present at sub-mg/L (ppb) or low- to mid-
mg/L levels. However, more than 50% of the total organic
halide (TOX) formed during the chlorination of drinking
water [19] and more than 50% of the assimilable organic
carbon (AOC) formed during ozonation of drinking water
has not been accounted for as identified DBPs [20];
furthermore, nothing is known about the potential
toxicity of many of the DBPs present in drinking water.
Here we review 30 years of results of occurrence,
genotoxicity, and carcinogenicity studies of DBPs
regulated by the U.S. Government and those that are
not named specifically in regulations. The compounds in
these two categories, and a qualitative assessment of the
results, are shown in Table 1. Although most of the
research has been performed on the regulated DBPs,
there is a growing literature on the unregulated DBPs.
The results of our analyses in this paper offer an
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242 181
Table 1
Summary of occurrence, genotoxicity, and carcinogenicity of regulated and unregulated DBPs
DBP Occurrence
a
Genotoxicity
b
Carcinogenicity
Regulated DBPs
THMs
Chloroform ***** +
Bromodichloromethane **** + +
Chlorodibromomethane **** + +
Bromoform **** + +
HAAs
Chloroacetic acid *** +
Bromoacetic acid *** +
Dichloroacetic acid ***** + +
Dibromoacetic acid ***** + +
Trichloroacetic acid ***** +
Oxyhalides
Bromate *** + +
Chlorite ******
c
Unregulated DBPs
Halonitromethanes
Chloronitromethane ** +
Bromonitromethane ** +
Dichloronitromethane ** +
Dibromonitromethane *** +
Bromochloronitromethane ** +
Trichloronitromethane (chloropicrin) **** +
Bromodichloronitromethane *** +
Dibromochloronitromethane *** +
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242182
Table 1 (Continued )
DBP Occurrence
a
Genotoxicity
b
Carcinogenicity
Tribromonitromethane *** +
Iodo-acids
Iodoacetic acid *** +
Bromoiodoacetic acid *** +
(Z)-3-Bromo-3-iodopropenoic acid **
(E)-3-Bromo-3-iodopropenoic acid **
2-Iodo-3-methylbutenedioic acid *** +
d
Other halo-acids
Bromochloroacetic acid **** +
e
Bromodichloroacetic acid **** +
e
Dibromochloroacetic acid **** +
e
Tribromoacetic acid **** +
Iodo-THMs and other unregulated THMs
Dichloroiodomethane ***
Bromochloroiodomethane ***
Dibromoiodomethane ***
Chlorodiiodomethane ***
Bromodiiodomethane ***
Iodoform *** +
Dichloromethane *** +
Bromochloromethane ND +
Dibromomethane ND/** +
MX compounds
MX ** + +
Red-MX * +
Ox-MX * +
EMX * +
ZMX * +
Mucochloric acid ** +
BMX-1 ** +
BMX-2 * +
BMX-3 * +
BEMX-1 ** +
BEMX-2 ** +
BEMX-3 ** +
Haloamides
Chloroacetamide *** +
Bromoacetamide *** +
Iodoacetamide +
Dichloroacetamide *** +
Bromochloroacetamide *** +
Dibromoacetamide *** +
Bromoiodoacetamide *** +
Trichloroacetamide *** +
Bromodichloracetamide *** +
Dibromochloroacetamide *** +
Tribromoacetamide *** +
Diiodoacetamide +
Chloroiodoacetamide +
Haloacetonitriles
Chloroacetonitrile *** +
Bromoacetonitrile *** +
Iodoacetonitrile +
Dichloroacetonitrile *** +
Bromochloroacetonitrile *** +
Dibromoacetonitrile *** + On test
opportunity to assess the value and completeness of the
current literature on the regulated DBPs and to consider
how the emerging literature on the unregulated DBPs
might inform future research needs and assessments of
drinking water.
To provide a historical context for this work, we
begin with an overview of U.S. DBP regulations,
followed by a brief summary of the epidemiology of
drinking water and cancer. We have not reviewed the
literature on reproductive/developmental effects asso-
ciated with DBPs or drinking water. We then review the
occurrence, genotoxicity, and carcinogenicity literature
for the regulated and then the unregulated DBPs, ending
with our conclusions regarding research needs.
2. Overview of DBP regulations in the United
States
Based on the discoveries of DBPs described in the
Introduction, the U.S. EPA issued a regulation in 1979
to control total THMs at an annual average of 100 mg/L
(ppb) in drinking water; THMs here are defined as
chloroform, bromodichloromethane, dibromochloro-
methane, and bromoform [21]. In 1998, the U.S. EPA
issued the Stage 1 Disinfectants (D)/DBP Rule, which
lowered permissible levels of total THMs to 80 mg/L
and regulated for the first time five haloacetic acids
(HAAs) (60 mg/L), bromate (10 mg/L), and chlorite
(1000 mg/L) (Table 2)[22]. Stage 1 regulations required
monitoring based on running annual averages, which
represented averages of all samples collected in a
utility’s distribution system over a 1-year period. This
Rule became effective on 1 January 2002 [23].
The Stage 2 D/DBP Rule (published in January
2006) maintained the Stage 1 Rule maximum con-
taminant levels (MCLs) for THMs and HAAs (Table 1)
and required that MCLs be based on locational running
annual averages; that is, each location in the distribution
system needs to comply on a running annual average
basis [24]. The reason for this change was that the
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242 183
Table 1 (Continued )
DBP Occurrence
a
Genotoxicity
b
Carcinogenicity
Trichloroacetonitrile *** +
Bromodichloroacetonitrile ***
Dibromochloroacetonitrile ***
Tribromoacetonitrile ***
Halopyrroles
2,3,5-Tribromopyrrole ** +
Nitrosamines
NDMA ** + +
f
N-Nitrosopyrrolidine * + +
f
N-Nitrosomorpholine * + +
f
N-Nitrosopiperidine * + +
f
N-Nitrosodiphenylamine * + +
f
Aldehydes
Formaldehyde *** + +
Acetaldehyde *** + +
Chloroacetaldehyde *** +
Dichloroacetaldehyde ***
Bromochloroacetaldehyde ***
Trichloroacetaldehyde (chloral hydrate) **** + +
Tribromoacetaldehyde ***
Other DBPs
Chlorate ****** + +
a
Key to occurrence symbols: *low-ng/L levels; **ng/L to sub-mg/L levels; ***sub- to low-mg/L levels; ****low-mg/L levels; *****low- to mid-
mg/L levels; ******high mg/L levels; ND, non-detect; entries left blank have no occurrence data available; bromine-containing DBPs formed only
when source waters contain natural bromide (occurrence lower than shown if low bromide levels in source waters).
b
Symbols represent weight of evidence for the genotoxicity data. In general, where a compound was genotoxic in several studies in the same assay
or was genotoxic in several different assays, it was declared ‘‘+’’ in the table even if the compound was negative in other assays.
c
Based on 85-week studies.
d
M.J. Plewa, in preparation, personal communication.
e
A.B. DeAngelo, in preparation, personal communication.
f
Details of these studies are not given in the following tables because they have been reviewed extensively [230]. As noted in the text, most of
these compounds are rodent carcinogens by various routes of exposure, including via the drinking water.
running annual averages (used with the Stage 1 D/DBP
Rule) permitted some locations within a water
distribution system to exceed the MCLs as long as
the average of all sampling points did not exceed the
MCLs. As a result, consumers served by a particular
section of the distribution system could receive water
that regularly exceeded the MCLs. The Stage 2 D/DBP
Rule maintains the MCLs for bromate and chlorite;
however, the U.S. EPA plans to review the bromate
MCL as part of their 6-year review process (additional
details are available at http://www.epa.gov/safewater/
stage2/index.html). Other countries besides the United
States have regulated DBPs, and there are World Health
Organization (WHO) guidelines for DBPs as well as
European Union DBP standards (Table 2).
With stricter regulations for THMs and new
regulations for HAAs, many drinking-water utilities
have changed their disinfection practices to meet the
new regulations. Often, the primary disinfectant is
changed from chlorine to so-called alternative disin-
fectants, including ozone, chlorine dioxide, and
chloramines. In some cases, chlorine is used as a
secondary disinfectant following primary treatment
with an alternative disinfectant, particularly for ozone
and chlorine dioxide. However, new issues and
problems can result with changes in disinfection
practice.
For example, the use of ozone can significantly
reduce or eliminate the formation of THMs and HAAs,
but it can result in the formation of bromate, especially
when elevated levels of bromide are present in the
source waters. Bromate is a concern because it has been
shown to be a carcinogen in laboratory animals [25].As
a result, the U.S. EPA regulated bromate under the Stage
1 D/DBP Rule at an MCL of 10 mg/L to limit its
occurrence [22]. Nitrosodimethylamine (NDMA),
which can form at higher levels with chloramination,
is also a concern because there are data indicating that it
is a carcinogen in several animal species. Under its 1986
Guidelines for Carcinogen Risk Assessment (http://
www.epa.gov/ncea/raf/car2sab/guidelines_1986.pdf),
the U.S. EPA classified NDMA as a probable human
carcinogen [26].
Likewise, a recent U.S. Nationwide DBP Occurrence
Study, which included drinking waters from source
waters containing high bromide/iodide and natural
organic matter levels, revealed that iodo-THMs and
newly identified iodo-acids were increased in formation
with chloramination; moreover, bromonitromethanes
were increased with preozonation followed by post-
chlorination or chloramination [9,27]. Differences in
source water conditions, including concentrations of
bromide or iodide, concentrations of natural organic
matter, and pH, can have a dramatic effect on the
formation of various DBPs (chlorine-, bromine-, or
iodine-containing) and the levels formed [9,28,29].
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242184
Table 2
DBP regulations and guidelines
DBP
U.S. EPA regulations MCL
a
(mg/L)
Total THMs 0.080
Five haloacetic acids 0.060
Bromate 0.010
Chlorite 1.0
World Health Organization (WHO) guidelines
DBP Guideline
value (mg/L)
Chloroform 0.2
Bromodichloromethane 0.06
Chlorodibromomethane 0.1
Bromoform 0.1
Dichloroacetic acid 0.05
b
Trichloroacetic acid 0.2
Bromate 0.01
b
Chlorite 0.7
b
Chloral hydrate (trichloroacetaldehyde) 0.01
b
Dichloroacetonitrile 0.02
b
Dibromoacetonitrile 0.07
Cyanogen chloride 0.07
2,4,6-Trichlorophenol 0.2
Formaldehyde 0.9
European Union Standards
DBP Standard
value (mg/L)
Total THMs 0.1
Bromate 0.01
c
a
The total THMs represent the sum of the concentrations of four
trihalomethanes: chloroform, bromoform, bromodichloromethane,
and chlorodibromomethane. They have been regulated in the United
States since 1979 [21], but the maximum contaminant level (MCL)
was lowered from 100 to 80 mg/L under the Stage 1 Disinfectants/
DBP (D/DBP) Rule [22]. World Health Organization (WHO) guide-
lines on THMs state that the sum of the ratio of the concentration of
each THM to its respective guideline value should not exceed unity.
The five haloacetic acids represent the sum of monochloro-, dichloro-,
trichloro-, monobromo-, and dibromoacetic acid. These haloacetic
acids, together with bromate and chlorite, were regulated for the first
time in the United States under the Stage 1 D/DBP Rule [22]. WHO
guidelines can be found at http://www.who.int/water_sanitation_-
health/dwq/gdwq3/en. European Union drinking-water standards
can be found at http://www.nucfilm.com/eu_water_directive.pdf.
b
Provisional guideline value.
c
Where possible, without compromising disinfection, EU member
states should strive for a lower value. This value must be met, at the
latest, 10 calendar years after the issue of Directive (3 November
1998); within 5 years of the Directive, a value of 25 mg/L must be met.
3. Summary of epidemiology studies of cancer
and drinking water
Some epidemiologic studies have shown that a life-
time exposure to chlorinated water is associated with an
increased risk for cancer, especially of the urinary bladder
and colorectum [17,30]. Besides DBPs, drinking water
may contain other potential carcinogens, such as arsenic
and radionuclides; however, the bladder cancer risk has
generally been associated with THM levels [31,32].One
study showed that both bladder and kidney cancer risks
were associated withthe mutagenicity of the water, which
may be related to levels of the chlorinated furanone, MX
[33] or possibly other mutagenic DBPs. Risk for rectal
cancer has recently been shown to be associated
specifically with levels of the THM bromoform [34].
The first and only epidemiologic study to stratify risk
by route of exposure has found that much of the bladder
cancer risk associated with chlorinated water appears to
be due to showering, bathing, and swimming rather than
to drinking the water [32] and that the risk may be
highest for people having the GSTT1-1 gene [35]. Such
observations indicate that genetic susceptibility may
play a role in the cancer risk and that the risk may be
especially related to dermal and inhalation exposure.
One study has shown that the risk for bladder cancer
decreased as the duration of exposure to ozonated water
increased [36]. Such an observation supports the shift
from chlorination to modified treatments such as
ozonation. Earlier studies had found that organic
extracts of ozonated water were far less mutagenic
than those of chlorinated water [37–39]; this has been
confirmed recently for organic concentrates of ozonated
water [40]. However, studies of water treated with
alternative disinfectants are limited, and there has not
been a systematic analysis carried out on drinking water
prepared from various types of source waters, including
high-bromide/iodide source waters.
Most of the DBPs tested for carcinogenicity in rodents
cause primarily liver cancer rather than bladder or
colorectal cancer [17,30]. As reviewed here, exceptions
include renal tumors induced by bromodichloromethane,
chloroform, and bromate; intestinal tumors induced by
bromodichloromethane and bromoform; and thyroid
tumors induced by bromate. The most striking exception
is the variety of organ sites at which MX induced tumors
in the rat, as well as the low doses at which these tumors
were induced (relative to the doses of the other DBPs).
This general lack of correlation between site of tumors
in animal cancer studies for individual DBPs and human
epidemiological studies for drinking water has not yet
been explained. However, in addressing the potential for
animal carcinogens to be hazardous to humans, most
regulatory agencies do not presume that there is tumor
site concordance between rodents and humans. Possible
areas for exploration involve route of exposure. Most of
the carcinogenicity studies of DBPs have involved
administration of the DBP in the drinking water (oral
exposures). However, the recent route-of-exposure study
[32,41] indicated that much of the bladder cancer
associated with chlorinated water may be due to
showering, bathing, and swimming (dermal and inhala-
tion exposures) rather than oral exposures. In addition,
only a few of the newly identified DBPs discussed in this
review have been tested for carcinogenicity, and perhaps
some of these will cause bladder or colorectal cancer.
Although not reviewed here, recent epidemiologic
studies have raised the issue of potential adverse
reproductive and developmental effects, such as low
birth weight, intrauterine growth retardation, and
spontaneous abortion [8,42–56].
4. Occurrence, genotoxicity, and carcinogenicity
of the regulated DBPs
4.1. Trihalomethanes (THMs)
4.1.1. Occurrence
The halomethanes make up one class of the
approximately 600 drinking-water DBPs that have
been identified. Within the halomethane class are the
THMs (chloroform, bromoform, bromodichloro-
methane, and chlorodibromomethane), which are
currently regulated by the U.S. EPA at a level of
80 mg/L for total trihalomethanes [24]. The THMs were
the first DBPs identified [3,4]. Together, the THMs and
HAAs are the two most prevalent classes of DBPs
formed in chlorinated drinking water, accounting for
approximately 25% of the halogenated DBPs [9]. They
are also formed at significantly lower levels in
chloraminated drinking water, and bromoform can be
formed in high-bromide source waters treated with
ozone [2,57]. Disinfection with chlorine dioxide does
not form THMs; however, low THM levels can be
present due to chlorine impurities in chlorine dioxide.
A National Organics Reconnaissance Survey (NORS)
and National Organics Monitoring Survey (NOMS)
conducted in the mid- to late-1970s collected the first
substantial information on THMs in the United States
[58]. Later, the U.S. EPA Information Collection Rule
(ICR), which involved 500 large drinking-water plants in
the United States, reported mean levelsin the distribution
system of 38 mg/L and 90th percentage levels of 78 mg/L
for THM4 (the four regulated THMs summed together)
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242 185
[23]. Chloroform was by far the most prevalent of the
THMs measured, and it had the highest mean concentra-
tion of 23 mg/L. Brominated THMs (bromodichloro-
methane, chlorodibromomethane, and bromoform) can
increase in formation relative to chloroform when eleva-
ted levels of natural bromide are present in source waters
(often due to salt water intrusion). THM levels observed
in the ICR were substantially lower (reduced by 50–60%)
than levels observed in the earlier NORS study [23].
4.1.2. Genotoxicity
The THMs have been studied intensively over the past
30 years, and many in vitro techniques have been used to
investigate their mutagenic and genotoxic properties [59]
(Table 3). We have used the term ‘‘mutagenicity’’ to refer
to assays that measure a change in DNA sequence (either
gene or chromosomal mutation); we have used the term
‘‘genotoxicity’’ to refer to mutagenicity as well as DNA
damage (DNA adducts, DNA strand breaks, etc.).
Although many of the initial genotoxicity tests of the
THMs resulted in negative responses, later studies
(discussed below) showed that the brominated THMs
were mutagenic after activation by glutathione S-
transferase-theta (GSTT1-1).
The genotoxicity of chloroform (trichloromethane)
has been reviewed extensively [59], and those reports
not included in the IARC review are shown in Table 3.
With few exceptions, chloroform is not mutagenic or
genotoxic in a wide array of systems and endpoints in
vivo and in vitro. Although some weak positive
responses have been observed, these are either in
single studies, or the results have not been highly
repeatable. Unlike some of the THMs, chloroform is not
activated by GSTT1-1 to a mutagen in Salmonella [60].
As discussed in the carcinogenicity section below,
chloroform is generally considered to be a nongeno-
toxic carcinogen whose mechanism of action involves
cytotoxicity and regenerative cell proliferation [59]
(http://www.epa.gov/iris/subst/0025.htm).
Bromodichloromethane, chlorodibromomethane,
and bromoform have generally not induced gene
mutations in the standard test systems; the few positive
results are either in single studies or were not found in
repeated studies [59] (Table 3). Nonetheless, some
studies have found that chlorodibromomethane induced
chromosomal aberrations or sister chromatid exchanges
(SCEs) and that bromoform induced SCEs and
micronuclei [61]. Recently these DBPs were evaluated
for genotoxicity in CHO cells; they were refractory
to concentrations of 5 mM. The rank order of
chronic CHO cell cytotoxicity was bromoform >
chlorodibromomethane >chloroform >bromodichlor-
omethane [62]. However, unlike chloroform, these bro-
minated THMs are activated to mutagens by GSTT1-1
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242186
Table 3
Comparative genotoxicity of halomethane DBPs
Chemical Biosystem Genetic endpoint Concentration range of
positive response or highest
genotoxic potency
References
Dibromomethane Salmonella TA100 his reversion [121]
Preincubation
S9 279 revertants/mmol
+S9 551 revertants/mmol
E. coli TRG8 his reversion 0.02–0.1 mM [140]
Salmonella TA1535 (+)GST5-5 his reversion 0.1–1 mM [139]
Bromoform Review [61]
Salmonella TA100 his reversion Negative [121]
Preincubation
S9, +S9
Human lymphocytes SCGE Weakly + [283]
S. typhimurium his reversion [77]
RSJ100 S9 44 revertants/mmol
+S9 Negative
TA98 S9 Negative
+S9 237 revertants/mmol
TA100 S9 Negative
+S9 83 revertants/mmol
Human lung epithelial cells SCGE 100–1000 mM[138]
Salmonella RSJ100 his reversion 1798 revertants/1600 ppm [63]
Dichloromethane Review [61]
Salmonella TA100 his reversion [121]
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242 187
Table 3 (Continued )
Chemical Biosystem Genetic endpoint Concentration range of
positive response or highest
genotoxic potency
References
Preincubation
–S9 Negative
+S9 7.9 revertants/mmol
Human lung epithelial cells SCGE Weakly + [138]
Salmonella RSJ100 his reversion 140 revertants/400 ppm [63]
Salmonella TA100 his reversion 976 revertants/24,000 ppm [63]
Chloroform Review [59]
Salmonella TA100 his reversion Preincubation Negative [121]
S9, +S9
Saccharomyces cerevisiae Deletion recombination 5.59 mg/mL [284]
Assay
Female B6C3F1 lacI transgenic mice lacI mutation Negative [285]
Salmonella his reversion Negative [77]
RSJ100 S9, +S9
TA98
TA100
Human lung epithelial cells SCGE Weakly + [138]
Salmonella his reversion 19,200 and 25,600 ppm [60]
TA1535 Plate-incorporation
Salmonella S9, +S9, Negative [286]
TA98, TA100, TA1535, TA1537 Glutathione suppl. S9
E. coli S9, Negative [286]
WP2uvrA/pKM101 Glutathione supplemented S9 Negative
E. coli WP2/pKM101 +Glutathione supplemented S9 Negative
0.5–2%
Bromochloromethane Salmonella TA100 his reversion [121]
Preincubation
S9 75 revertants/mmol
+S9 424.4 revertants/mmol
Salmonella TA1535 (+)GST5-5 his reversion 0.2–1.75 mM [139]
Chlorodibromomethane Review [61]
Salmonella TA100 his reversion Negative [121]
Preincubation
S9, +S9
Human lung epithelial cells SCGE Negative [138]
Salmonella RSJ100 his reversion 1364 revertants/400 ppm [63]
Salmonella RSJ100 his reversion [283]
S9 1110 revertants/800 ppm
+S9 1018 revertants/800 ppm
Salmonella TA1535 umuDC-lacZ [287]
S9 Positive
+S9 Negative
Bromodichloromethane Review [59]
Salmonella TA100 his reversion Negative [121]
Preincubation
S9, +S9
Human lung epithelial cells SCGE 10–1000 mM[138]
Salmonella RSJ100 his reversion 831 revertants/plate [60,63]
Salmonella TA1535 umuDC-lacZ Negative [287]
S9,+S9
Iodoform Salmonella BA13 Ara 7371 mut/mmol S9 [306]
1782 mut/mmol +S9
SHE cells Chrom. Ab. Negative [307]
in a transgenic strain of Salmonella (RSJ100); their
rank order of mutagenic potency was bromoform >
bromodichloromethane >chlorodibromomethane
[60,63]. Thus, the likely absence of GSTT1-1 in most
(if not all) of the studies in which these compounds were
not genotoxic may account for the general negative
results in the standard test systems. The dependence of
these compounds on GSTT1-1 to be activated to
mutagens raises important limitations regarding the
standard test systems and emphasizes the need for basic
research of the sort that has been applied to these
brominated THMs.
DeMarini et al. [63] proposed two possible pathways
of metabolism of THMs that would result in the
GC !AT transitions identified as the sole class of base
substitutions induced by these THMs in strain RSJ100
of Salmonella. The authors demonstrated that GSTT1-1
had the ability to mediate the mutagenicity of bromine-
containing THMs but not chloroform. They suggested
that the difference in mutational mechanisms between
the brominated THMs and chloroform is likely due to
initial metabolism in which the bromine is removed via
nucleophilic displacement of bromine or reductive
dehalogenation. Data in humans and animals indicate
that chloroform is metabolized chiefly to phosgene
except at high doses [63]. Pegram et al. [60]
demonstrated that brominated THMs could be activated
by GST-mediated transformation into mutagenic inter-
mediates. Also, chloroform displayed a low affinity for
the same pathway, indicating that the THMs as a
chemical class do not share the same mode of action.
More recently, the biotransformation and genotoxi-
city of
14
C-bromodichloromethane were studied. These
in vitro experiments demonstrated that GSTT1-1
catalyzed the covalent binding of bromodichloro-
methane to DNA and the formation of guanine adducts
[64]. The cancer target tissues in the rat had greater
potential formation of bromodichloromethane-derived
DNA adducts compared to the rat liver due to greater
flux through the GSTT1-1 pathway [64].
4.1.3. Carcinogenicity
All four of the regulated THMs are carcinogenic in
rodents (Table 4)[59,61,65]. Only two have been
administered in the drinking water, bromodichloro-
methane and chloroform, and both were negative in the
mouse via this route. However, in the rat, bromodi-
chloromethane produced liver tumors, and chloroform
produced renal tumors when exposure was via the
drinking water (Table 4). When administered by
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242188
Table 4
Carcinogenicity of regulated disinfection by-products in rodents based on 2-year dosing studies
Chemical (RfD) Species Route and dose Tumor diagnoses References
Trihalomethanes
Bromodichloromethane
(20 mg/(kg day))
Mouse Drinking water: 0, 9, 18,
36 mg/(kg day)
Drinking water: no evidence of
carcinogenicity
[288]
Gavage male mice: 0,25,
50 mg/(kg day)
Gavage: male mice renal tumors 1/49,
2/50, 10/50
Gavage female mice: 0, 75,
150 mg/(kg day)
Gavage: female mice hepatocellular
tumors 3/50, 18/48, 33/50
Drinking water: 8.1, 27.2,
43.4 mg/(kg day)
Drinking water: no evidence of
carcinogenicity
[290]
Rat Drinking water: 0, 6, 12,
25 mg/(kg day)
Drinking water: no evidence of
carcinogenicity
[288–290]
Gavage: 0, 50, 100 mg/(kg day) Gavage: male rats renal tumors 0/50,
1/50, 13/50; intestinal carcinoma 0/50,
11/50, 38/50
Gavage: female rats renal tumors
0/50, 1/50, 15/50; intestinal
carcinoma 0/46, 0/50, 6/47
Feed: 0, 6.1, 25.5, 138
mg/(kg day)
Feed: no evidence of carcinogenicity
Drinking water 2: 0, 8.1, 27.2,
43.4 mg/(kg day)
Drinking water 2: male rat liver
tumors 2/45, 8/45, 7/48, 4/49
Bromoform (20 mg/(kg day)) Mouse Gavage: 0, 50, 100 mg/(kg day) Gavage: no evidence of carcinogenicity [291]
Rat Gavage: 0, 100, 200 mg/(kg day) Gavage: male rats intestinal tumors
0/50, 0/50, 3/50
[291]
Gavage: female rats intestinal
tumors 0/50, 1/50, 8/50
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242 189
Table 4 (Continued )
Chemical (RfD) Species Route and dose Tumor diagnoses References
Chlorodibromomethane
(20 mg/(kg day))
Mouse Gavage: 0, 50, 100 mg/(kg day) Gavage: male mice hepatocellular
tumors 23/50, 27/50
[292]
Gavage: female mice hepatocellular
tumors 6/50, 10/49, 19/50
Rat Gavage: 0, 40, 80 mg/(kg day) Gavage: no evidence of
carcinogenicity
[292]
Chloroform (10 mg/(kg day)) Mouse Gavage: males 0, 138, 277 mg/
(kg day); females 0, 238, 477
mg/(kg day)
Gavage: male mice hepatocellular
tumors 3/50, 18/50, 49/50
[6,293]
Gavage: female mice hepatocellular
tumors 0/50, 40/50, 48/50
Drinking water: 0, 34, 65, 130,
263 mg/(kg day)
Drinking water: no evidence of
carcinogenicity
Rat Gavage: males 0, 90, 180 mg/
(kg day); females 0, 100, 200
mg/(kg day)
Gavage: male rats renal tumors
0/50, 4/50, 12/50
[6,293]
Drinking water: 0, 19, 38, 81,
160 mg/(kg day)
Drinking water: male rat renal
tumors 4/301, 4/313, 4/148, 3/48,
7/50
[66]
Inhalation males (0, 25, 50,
100 ppm, 6 h/day, 5 day/week)
combined with drinking water
(1000 ppm): total dose was 0,
73, 93, 135, mg/(kg day)
Combined exposure: renal
tumors 0/50, 4/50, 4/50, 18/50
Haloacetic acids
Chloroacetic acid
(not listed on IRIS)
Mouse Gavage: 0, 50, 100 mg/(kg day) Gavage: no evidence of
carcinogenicity
[87]
Rat Gavage: 0, 15, 30 mg/(kg day) Gavage: no evidence of
carcinogenicity
[86,87]
Drinking water: 0, 3.5, 26.1,
59.9 mg/(kg day)
Drinking water: no evidence of
carcinogenicity
Bromoacetic acid
(not listed on IRIS)
Mouse No data No data No cancer
studies
performed
Rat No data No data No cancer
studies
performed
Dibromoacetic acid
(not listed on IRIS)
Mouse Drinking water: 0, 50,
500, 1000 mg/L
Male hepatocellular tumors
28/49, 41/50, 42/50, 47/50; male
lung tumors 12/49, 12/50, 22/50,
47/50
[88]
Female hepatocellular tumors
22/49, 28/50, 37/50, 37/49
Rat Drinking water: 0, 50,
500, 1000 mg/L
Male mesothelioma 3/50, 1/50,
0/50, 10/50; male leukemia
17/50, 31/50, 24/50, 13/50
[88]
Female mesothelioma 11/50,
13/50, 16/50, 22/50
Dichloroacetic acid
(4 mg/(kg day))
Mouse Drinking water 52 weeks: 0,
1, 2 g/L
Drinking water 52 weeks: male
mouse liver tumors 0/35, 0/11, 7/24
[294,295]
Drinking water: 0, 8, 84, 168,
315, 429 mg/(kg day)
Drinking water: male mouse
hepatocellular tumors 13/50,
11/33, 12/24, 23/32, 13/14, 8/8
Rat Drinking water: 0, 3.6, 40.2,
139.1 mg/(kg day)
Drinking water: male rat
hepatocellular tumors 1/33,
0/26, 7/29, 8/28
[296]
gavage, bromodichloromethane produced renal and
liver tumors in the mouse, and renal and intestinal
tumors in the rat. Chloroform also produced liver
tumors in the mouse and renal tumors in the rat
(Table 4). A combined exposure of rats to chloroform
via both the drinking water and inhalation produced
renal tumors [66].
The other two regulated THMs, bromoform and
chlorodibromomethane, have been administered only
by gavage, and both were negative in one species
(bromoform in mouse and chlorodibromomethane in
rat) (Table 4). However, bromoform induced intestinal
tumors in the rat, and chlorodibromomethane induced
liver tumors in the mouse (Table 4). All but bromoform
produced liver tumors. Chloroform and bromodichlor-
omethane also produced renal tumors, and bromodi-
chloromethane and bromoform produced intestinal
tumors. Only two of the four regulated THMs produced
tumors at multiple organ sites (chloroform and
bromodichloromethane), and these same two are the
only ones that are carcinogenic in both mouse and rats,
i.e., are trans-species carcinogens.
With two notable exceptions, the regulated THMs
did not produce urinary bladder or colorectal tumors,
which are the primary tumors associated with drinking-
water exposure in epidemiological studies (Section 3).
The exceptions were bromodichloromethane and
bromoform, which produced tumors of the large
intestine in the rat, and these tumors are anatomically
and functionally analogous to the colon in humans.
Mechanistic studies have also shown that bromoform
and bromodichloromethane induce aberrant crypt foci
(ACF) primarily in the rectal segment of the colon of
rats (not in mice) when administered either via drinking
water or gavage [66a,66b]. A high-fat diet had no
influence on the ACF frequency induced by bromodi-
chloromethane; however, it increased by twofold the
frequency of ACF induced by bromoform [66c]. A diet
lacking folate significantly increased the frequency of
ACF induced by bromoform relative to that of a normal
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242190
Table 4 (Continued )
Chemical (RfD) Species Route and dose Tumor diagnoses References
Trichloroacetic acid
(no RfD)
Mouse Drinking water 52 weeks: 0,
1, 2 g/L
Drinking water 52 weeks: male
mouse hepatocellular tumors
0/35, 4/11, 5/24
[294]
Rat Drinking water: 0, 3.6, 32.5,
363.8 mg/(kg day)
Drinking water: no evidence
of carcinogenicity
[86]
Other
Bromate (4 mg/(kg day)) Mouse Drinking water: 0, 9.1, 42.4,
77.8 mg/(kg day)
Drinking water: mouse renal
tumors 0/40, 5/38, 3/41, 1/44
[101]
Rat Drinking water: males 0, 12.5,
27.5; females 0, 12.5, 25.5
mg/(kg day)
Drinking water: male rat renal
tumors 3/53, 32/53, 46/52;
male rat mesothelioma 6/53,
17/52, 28/46; female rat renal
tumors 0/47, 28/50, 39/49
[25,100,
101,297]
Drinking water 2: 0, 0.9, 1.7, 3.3,
7.3, 16.0, 43.4 mg/(kg day)
Drinking water 2: male rat renal
tumors 0/19, 0/19, 0/20, 1/24,
5/24, 5/20, 9/20; male rat thyroid
follicular cell tumor 0/16, 0/19, 3/20,
4/24, 2/24, 3/20, 15/19; male rat
mesothelioma 0/19, 0/20, 3/20,
4/24, 2/24, 3/20, 15/20
Drinking water 3: 0, 1.5, 7.9,
16.9, 37.5 mg/(kg day)
Drinking water 3: male rat renal
tumors 1/45, 1/43, 6/47, 3/39, 12/32;
male rat thyroid follicular cell tumor
0/36, 4/39, 1/43, 4/35, 14/30; male
rat mesothelioma 0/47, 4/49, 5/49,
10/47, 27/43
Chlorite (30 mg/(kg day))
(85-week studies)
Mouse Drinking water: 0, 0.025, 0.05% Drinking water: no evidence of
carcinogenicity
[100,298]
Drinking water 2: 0, 250, 500 ppm Drinking water 2: no evidence
of carcinogenicity
Chlorite (30 mg/(kg day))
(85-week study)
Rat Drinking water: 0, 300, 600 ppm Drinking water: no evidence
of carcinogenicity
[100]
diet in rats [66d]. These studies provide an important
mechanistic link to a type of cancer associated with
drinking-water exposure in humans.
IARC has found bromoform [61] and chlorodibro-
momethane [65] to be group 3, which is not classifiable
as to their human carcinogenicity. In contrast, both
chloroform [59] and bromodichloromethane [61] have
been classified by IARC as 2B, possibly carcinogenic to
humans. The U.S. EPA’s Integrated Risk Information
System (IRIS) describes bromodichloromethane as B2,
probable human carcinogen (http://www.epa.gov/iris/
subst/0213.htm).
Chloroform is the only regulated THM for which there
is enough evidence to develop a risk assessment based on
its mode of action [67]. Numerous studies have shown
that chloroform is not genotoxic and that tumors, when
they arise, develop only at doses that produce significant
cellular toxicity, cell death, and regenerative proliferation
[68–70]. The IRIS discussion of chloroform (http://
www.epa.gov/iris/subst/0025.htm) indicates that three
different types of quantitative assessments are possible.
The weight-of-evidence assessment concludes that
‘‘chloroform is likely to be carcinogenic to humans by
all routes of exposure under high-exposure conditions
that lead to cytotoxicity and regenerative hyperplasia in
susceptible tissues. However, chloroform is not likely to
be carcinogenic to humans by any route of exposure
under exposure conditions that do not cause cytotoxicity
and cell regeneration.’’
Chloroform has induced kidney tumors in male rats
and liver tumors in male and female mice only at doses
that resulted in cytotoxicity. The tumors were postulated
to be secondary to sustained or repeated oxidative
metabolism-mediated cytotoxicity and secondary regen-
erative hyperplasia. This oxidative pathway can produce
the electrophilic metabolite phosgene, which can lead to
tissue injury andcell death by reaction with tissue proteins
and cellular macromolecules as well as phospholipids,
glutathione, free cysteine, histidine, methionine, and
tyrosine. Persistent cell proliferation could lead to
increased mutation,increased conversion of spontaneous
DNA damage into mutations, and subsequent cancer. The
weight of the evidence indicates thata mutagenic mode of
action via DNA reactivity is not significant.
Although there is insufficient information for the
other regulated THMs to develop a specific mode of
action, mutational events and cellular death and
regeneration may be necessary for the carcinogenicity
of the brominated THMs. Recent data on the
pharmacokinetics of bromodichloromethane in humans
showed that the maximum blood concentrations of
bromodichloromethane were 25–130 times higher from
dermal exposure compared to oral exposure [71],
emphasizing the importance of route of exposure in risk
assessment of the brominated THMs [64,72,73].
4.2. Haloacetic acids (HAAs)
4.2.1. Occurrence
Currently, five haloacetic acids are regulated by the
U.S. EPA. The maximum contaminant level (MCL) is
60 mg/L for the sum of bromoacetic acid, dibromoacetic
acid, chloroacetic acid, dichloroacetic acid, and trichlor-
oacetic acid. HAAs can be formed by disinfection with
chlorine, chloramines, chlorine dioxide, and ozone, but
they are generally formed at highest levels with
chlorination [2]. Chloramines form substantially lower
levels of HAAs, which is one of the reasons it has become
a popular alternative disinfectant for public water
systems that cannot meet the regulation with chlorination
[74]. Because chlorine dioxide disinfection significantly
reduces the levels of THMs and HAAs relative to chlo-
rine, it is not generally well known that chlorine dioxide
can form HAAs. However, studies have shown that
chlorine dioxide can form HAAs, primarily dichloro-,
bromochloro-, and dibromoacetic acid [9,23,29,74–76].
The Information Collection Rule (ICR) data revealed
that water-treatment systems using chlorine dioxide had
higher haloacetic acid levels for the nine bromo-chloro-
HAAs than those using chlorine or chloramine only [23].
Water-treatment systems using chlorine dioxide also
used chlorine or chloramines (mostly as post-disin-
fectants), but this is further evidence that chlorine dioxide
can contribute to the formation of HAAs. Increased
formation of dihaloacetic acids was also observed in a
recently conducted Nationwide Occurrence Study [9,27].
Overall, the ICR found mean concentrations of the five
regulated HAAs at 23 mg/L and a 90th percentile of
47.5 mg/L at all water-treatment systems measured [23].
Like chloramines and chlorine dioxide, ozone used
in water treatment is well known for lowering the levels
of THMs and HAAs formed, relative to chlorine.
However, when source waters contain elevated levels of
natural bromide, dibromoacetic acid has been shown to
form [2,57].
4.2.2. Genotoxicity
The genotoxicity data for the HAAs are summarized
in [17] and Table 5. As shown in Table 5, limited data
are available for iodoacetic acid, bromoacetic acid,
dibromoacetic acid, tribromoacetic acid, and chloroa-
cetic acid. However, in general, all five were mutagenic
in Salmonella and induced DNA damage (SCGE assay)
in CHO cells in the absence of S9. Thus, these HAAs
S.D. Richardson et al. / Mutation Research 636 (2007) 178–242 191