United States
Environmental Protection
1=1 m m Agency
EPA/690/R-06/019F
Final
9-20-2006
Provisional Peer Reviewed Toxicity Values for
Hexanedioic acid
(CASRN 124-04-9)
Superfund Health Risk Technical Support Center
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268

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Acronyms and Abbreviations
bw	body weight
cc	cubic centimeters
CD	Caesarean Delivered
CERCLA	Comprehensive Environmental Response, Compensation and
Liability Act of 1980
CNS	central nervous system
cu.m	cubic meter
DWEL	Drinking Water Equivalent Level
FEL	frank-effect level
FIFRA	Federal Insecticide, Fungicide, and Rodenticide Act
g	grams
GI	gastrointestinal
HEC	human equivalent concentration
Hgb	hemoglobin
i.m.	intramuscular
i.p.	intraperitoneal
IRIS	Integrated Risk Information System
IUR	inhalation unit risk
i.v.	intravenous
kg	kilogram
L	liter
LEL	lowest-effect level
LOAEL	lowest-observed-adverse-effect level
LOAEL(ADJ)	LOAEL adjusted to continuous exposure duration
LOAEL(HEC)	LOAEL adjusted for dosimetric differences across species to a human
m	meter
MCL	maximum contaminant level
MCLG	maximum contaminant level goal
MF	modifying factor
mg	milligram
mg/kg	milligrams per kilogram
mg/L	milligrams per liter
MRL	minimal risk level
MTD	maximum tolerated dose
MTL	median threshold limit
NAAQS	National Ambient Air Quality Standards
NOAEL	no-ob served-adverse-effect level
NOAEL(ADJ)	NOAEL adjusted to continuous exposure duration
NOAEL(HEC)	NOAEL adjusted for dosimetric differences across species to a human
NOEL	no-ob served-effect level
OSF	oral slope factor
p-IUR	provisional inhalation unit risk
p-OSF	provisional oral slope factor
p-RfC	provisional inhalation reference concentration
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p-RfD
provisional oral reference dose
PBPK
physiologically based pharmacokinetic
ppb
parts per billion
ppm
parts per million
PPRTV
Provisional Peer Reviewed Toxicity Value
RBC
red blood cell(s)
RCRA
Resource Conservation and Recovery Act
RDDR
Regional deposited dose ratio (for the indicated lung region)
REL
relative exposure level
RfC
inhalation reference concentration
RfD
oral reference dose
RGDR
Regional gas dose ratio (for the indicated lung region)
s.c.
subcutaneous
SCE
sister chromatid exchange
SDWA
Safe Drinking Water Act
sq.cm.
square centimeters
TSCA
Toxic Substances Control Act
UF
uncertainty factor
l^g
microgram
[j,mol
micromoles
voc
volatile organic compound
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PROVISIONAL PEER REVIEWED TOXICITY VALUES FOR
HEXANEDIOIC ACID (CASRN 124-04-9)
Background
On December 5, 2003, the U.S. Environmental Protection Agency's (EPA's) Office of
Superfund Remediation and Technology Innovation (OSRTI) revised its hierarchy of human
health toxicity values for Superfund risk assessments, establishing the following three tiers as the
new hierarchy:
1.	EPA's Integrated Risk Information System (IRIS).
2.	Provisional Peer-Reviewed Toxicity Values (PPRTV) used in EPA's Superfund
Program.
3.	Other (peer-reviewed) toxicity values, including:
~	Minimal Risk Levels produced by the Agency for Toxic Substances and Disease
Registry (ATSDR),
~	California Environmental Protection Agency (CalEPA) values, and
~	EPA Health Effects Assessment Summary Table (HEAST) values.
A PPRTV is defined as a toxicity value derived for use in the Superfund Program when
such a value is not available in EPA's Integrated Risk Information System (IRIS). PPRTVs are
developed according to a Standard Operating Procedure (SOP) and are derived after a review of
the relevant scientific literature using the same methods, sources of data, and Agency guidance
for value derivation generally used by the EPA IRIS Program. All provisional toxicity values
receive internal review by two EPA scientists and external peer review by three independently
selected scientific experts. PPRTVs differ from IRIS values in that PPRTVs do not receive the
multi-program consensus review provided for IRIS values. This is because IRIS values are
generally intended to be used in all EPA programs, while PPRTVs are developed specifically for
the Superfund Program.
Because science and available information evolve, PPRTVs are initially derived with a
three-year life-cycle. However, EPA Regions or the EPA Headquarters Superfund Program
sometimes request that a frequently used PPRTV be reassessed. Once an IRIS value for a
specific chemical becomes available for Agency review, the analogous PPRTV for that same
chemical is retired. It should also be noted that some PPRTV manuscripts conclude that a
PPRTV cannot be derived based on inadequate data.
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Disclaimers
Users of this document should first check to see if any IRIS values exist for the chemical
of concern before proceeding to use a PPRTV. If no IRIS value is available, staff in the regional
Superfund and RCRA program offices are advised to carefully review the information provided
in this document to ensure that the PPRTVs used are appropriate for the types of exposures and
circumstances at the Superfund site or RCRA facility in question. PPRTVs are periodically
updated; therefore, users should ensure that the values contained in the PPRTV are current at the
time of use.
It is important to remember that a provisional value alone tells very little about the
adverse effects of a chemical or the quality of evidence on which the value is based. Therefore,
users are strongly encouraged to read the entire PPRTV manuscript and understand the strengths
and limitations of the derived provisional values. PPRTVs are developed by the EPA Office of
Research and Development's National Center for Environmental Assessment, Superfund Health
Risk Technical Support Center for OSRTI. Other EPA programs or external parties who may
choose of their own initiative to use these PPRTVs are advised that Superfund resources will not
generally be used to respond to challenges of PPRTVs used in a context outside of the Superfund
Program.
Questions Regarding PPRTVs
Questions regarding the contents of the PPRTVs and their appropriate use (e.g., on
chemicals not covered, or whether chemicals have pending IRIS toxicity values) may be directed
to the EPA Office of Research and Development's National Center for Environmental
Assessment, Superfund Health Risk Technical Support Center (513-569-7300), or OSRTI.
INTRODUCTION
Neither IRIS (U.S. EPA, 2006) nor the HEAST (U.S. EPA, 1997) list an RfD, RfC, or
cancer assessment for hexanedioic acid (adipic acid). The Drinking Water Standards and Health
Advisories list (U.S. EPA, 2004) does not include an RFD or cancer assessment for hexanedioic
acid. The CARA list (U.S. EPA, 1991, 1994) does not include any documents for hexanedioic
acid. ACGIH (2001, 2006) established a time-weighted average - threshold limit value (TWA-
TLV) of 5 mg/m3 for hexanedioic acid based on a report from the Russian literature of irritant
and neurological effects in exposed workers (Krapotkina et al., 1981). No occupational exposure
limits have been proposed by NIOSH (2006) or OSHA (2006) for this compound. Hexanedioic
acid is considered a GRAS (Generally Recognized as Safe) substance by the U.S. Food and Drug
Administration (U.S. FDA, 2003) and is used as a food additive. Reviews for FDA were
performed by Informatics (1974) and FASEB (1976). WHO (1967, 1977, 1978, 2000) derived
an ADI of from 0 to 5 mg/kg-day for hexanedioic acid and salts based on a NOAEL of 1% in
feed in a two-year rat study. Toxicity data for hexanedioic acid were recently reviewed by
Kennedy (2002). ATSDR (2006), IARC (2006), and NTP (2006) have not published documents
for this compound. Literature searches were conducted for the period from 1965 to July, 2003 in
the following databases: TOXLINE (including NTIS and BIOSIS updates), CANCERLIT,
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MEDLINE, CCRIS, GENETOX, HSDB, EMIC/EMICBACK, DART/ETICBACK, RTECS, and
TSCATS. Additional literature searches from July 2003 through September 2004 were
conducted by NCEA-Cincinnati using MEDLINE, TOXLINE, Chemical and Biological
Abstracts databases.
REVIEW OF PERTINENT DATA
Human Studies
Oral Exposure. No relevant data were located regarding the toxicity or carcinogenicity of
hexanedioic acid to humans following oral exposure.
Inhalation Exposure. Functional disorders of the autonomic nervous system, gastrointestinal
tract, and upper respiratory tract were reported in a study of Russian workers exposed to
hexanedioic acid dust during its manufacture (Krapotkina et al., 1981, as cited in ACGIH, 2001).
No additional details regarding the observed effects are available.
No relevant data were located regarding the carcinogenicity of hexanedioic acid to
humans following inhalation exposure.
Animal Studies
Oral Exposure. Oral toxicity studies for hexanedioic acid in animals include several repeated-
dose studies in rats ranging in duration from 5 days to 33 weeks, a series of teratogenicity studies
in rats, mice, hamsters, and rabbits, and a chronic 2-year bioassay in rats. These data are
summarized below.
Lang and Bartsch (1953) conducted a series of studies designed to characterize the short-
term toxicity of hexanedioic acid in rats (unspecified strain). Four repeated-dose experiments
were conducted. In the first experiment, female rats (17-20 per group; average weight of 92
grams at study initiation) were fed diets providing 0, 10, 20, or 40 mg/day of hexanedioic acid
for 4 weeks. Based on body weight data reported in the article, the average daily doses are
estimated to have been 0, 85, 160, and 320 mg/kg-day, respectively. Weight gain and behavior
were monitored over the 4-week treatment period. No other toxicological parameters were
evaluated. No effects were observed in any treatment group. The high dose of 40 mg/day (320
mg/kg-day) is a NOAEL in this study; A LOAEL was not achieved.
In the second experiment (Lang and Bartsch, 1953), groups of 15-18 male rats
(unspecified strain, weighing 40-60 grams at study initiation) were fed diets providing 0, 200,
400, or 800 mg/day of hexanedioic acid neutralized with sodium hydroxide for five weeks.
Based on body weight data in the paper, these doses correspond to approximately 0, 1900, 4400,
and 11,000 mg/kg-day (rounded to 2 significant digits). Clinical signs of toxicity and body
weights were monitored over the treatment period. No other toxicological parameters were
evaluated. High-dose animals were observed with dull and ruffled fur and diarrhea for the first 2
to 3 weeks of treatment (incidence rates not reported). These signs of toxicity were not observed
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at the end of the study. Body weight gain was reduced in the two high-dose groups. After 5
weeks of treatment, mean body weights were 35% lower than controls in the high dose group
and 10% lower than controls in the mid-dose group. Statistical analysis was not conducted by
the researchers, but a Student's t-test on the body weight data performed for the current
evaluation determined that the differences from controls were statistically significant in both
dose groups. The NOAEL in this study was 200 mg/day (1900 mg/kg-day), and the LOAEL was
400 mg/day (4400 mg/kg-day) based on reduced body weight gain.
In the third experiment (Lang and Bartsch, 1953), groups of 13-15 rats (males and
females combined, unspecified strain, weighing 60-80 grams at study initiation) were exposed to
hexanedioic acid, neutralized with sodium hydroxide in the diet at doses of 0, 400, or 800
mg/day for up to 33 weeks. Based on time weighted average body weights, rats were dosed at
approximately 0, 1800, and 3900 mg/kg-day, respectively, over the 33-week treatment period.
Body weight and clinical signs of toxicity were recorded throughout the treatment period. After
33 weeks of treatment, an unspecified number of tissues were microscopically examined, and
hemoglobin, red and white blood cell count, and differential white blood cell counts were
determined (it is not clear whether these parameters were evaluated in animals that died prior to
scheduled sacrifice). A histopathology examination was also conducted on a group of animals
(unspecified number per dose) designated for interim sacrifice at Weeks 23 or 25. Clinical signs
of toxicity were similar to those observed in Experiment 2; diarrhea and dull and ruffled fur were
observed at 800 mg/day during the first 3 weeks of the study. In addition, high-dose animals
exhibited lethargy, which was not observed in high-dose animals of experiment 2. Mortality
incidence was higher in the 800 mg/day group (10 deaths) than in the control and low-dose
groups (4 deaths each). All deaths occurred within the first 4 treatment weeks. Body weight
gain was reduced in treated rats early in the study (after 8 weeks, average body weight was 26%
less than controls in the 800 mg/day group and 12% less than controls in the 400 mg/day group),
but reportedly recovered by Week 33 (control data for Week 33 were not shown).
Histopathology examinations revealed slight histological changes in the liver (including enlarged
cell nuclei, increased cell size and cell volume, and a decrease in Kupffer cells) and kidneys
(increased mitosis) and marked chronic inflammation in the intestinal mucosa at 400 and 800
mg/day. In addition to the animals discussed above, an unspecified number of pregnant female
rats were treated with 400 mg/day; the researchers reported that hexanedioic acid treatment did
not affect their ability to bear litters or nurse their young. The LOAEL in this study was 400
mg/day (1800 mg/kg-day), the lowest dose tested, based on reduced body weight gain and
lesions of the gastrointestinal mucosa, liver, and kidney. A NOAEL was not observed.
In the fourth experiment, male rats (unspecified number and strain, weighing 40-60
grams at study initiation) were maintained on protein restricted diets (11% protein, composed of
wheat and cod liver oil) supplemented with 0, 50, 100, 200, or 400 mg of hexanedioic acid daily
for 19 weeks (Lang and Bartsch, 1953). Based on body weight data reported in the paper, these
doses corresponded to approximately 0, 410, 880, 1600, and 4100 mg/kg-day (rounded to two
significant digits). Body weights and clinical signs of toxicity were monitored throughout the
treatment period. After 19 weeks of treatment, the animals were sacrificed, unspecified tissues
were microscopically examined, and hemoglobin, red and white blood cell count, and differential
white blood cell counts were determined (it is not clear whether these parameters were
determined for animals that died prior to the scheduled sacrifice). Also, three rats per group
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were designated for interim sacrifice after 8 weeks of treatment. These animals were subjected
to histological examinations (unspecified tissues). Mortality was similar in all groups. Clinical
signs of dull and ruffled fur and diarrhea (which were observed in previous experiments) were
not observed at any dose level in this experiment. Mean body weights of rats exposed at 400
mg/day were 22% less than controls after 6 weeks and 28% less after 19 weeks. Rats exposed at
400 mg/day were also observed to have slight histological changes in the liver and kidneys and
marked chronic inflammation in the intestinal mucosa. No treatment-related hematological
effects were noted. The NOAEL in this study was 200 mg/day (1600 mg/kg-day), and the
LOAEL was 400 mg/day (4100 mg/kg-day), based on reduced body weight gain and lesions of
the gastrointestinal mucosa, liver, and kidney.
Other subchronic studies also reported reduced body weight gain in rats fed hexanedioic
acid in the diet, but apparently did not examine other endpoints. In a 90-day feeding study
(Hazleton, 1950, as cited in FASEB, 1976), male albino rats (10 per dose, unspecified strain)
were maintained on a diet supplemented with hexanedioic acid at 0, 0.1%, 1.0%, or 5.0% (0,
100, 1000, or 5000 mg/kg-day, assuming a food factor of 10% for growing rats in a subchronic
study). Ten females were exposed at 0% or 1% (0 or 1000 mg/kg-day). Body weight and
survival were evaluated. Histopathology examinations were not conducted. Mean body weights
of rats exposed to 5% hexanedioic acid (5000 mg/kg-day) were substantially decreased
compared with controls throughout the exposure period. This effect was attributed to impaired
food utilization associated with high acid consumption by the study authors. No effects were
observed at the 1% hexanedioic acid concentration (1000 mg/kg-day). The original study report
was not available, and additional details were not reported in FASEB (1976). Similar results
were observed when male Carworth Farms albino rats (10 per dose) were maintained on a diet
supplemented with 5% (5000 mg/kg-day) sodium adipate (Informatics, 1974). Controls (N=5)
were fed untreated diet. Five treated rats were sacrificed after 14 weeks of treatment, and the
remaining five rats were fed untreated diet for an additional 8 weeks. Body weights were
recorded throughout the study, and all rats were subjected to a gross pathology examination at
terminal sacrifice. "Retardation of growth" occurred in sodium adipate treated rats. Rapid
growth occurred during the 8-week period after sodium adipate treatment stopped. The original
study report was not available, and additional details were not reported.
One chronic toxicity study in laboratory animals was located (Horn et al., 1957). Young
male Carworth Farms albino rats (20 per dose) were exposed to hexanedioic acid (unspecified
purity) in the diet at 0.1%, 1%, 3%, and 5% for two years (Horn et al., 1957). Based on time
weighted average body weights, these concentrations correspond to approximately 44, 470,
1500, and 2800 mg/kg-day (rounded to two significant digits). Nineteen female rats were also
exposed at 1% hexanedioic acid (-630 mg/kg-day). Controls (twenty males and 10 females)
were fed basal diet. The following parameters were recorded during the 2-year exposure period:
clinical signs of toxicity (weekly), body weights (recorded weekly, but reported for 8-week
intervals), food consumption, and survival. After two years of treatment, surviving animals were
sacrificed and the following parameters were evaluated: gross pathology (unspecified tissues),
organ weights (heart, liver, spleen, and kidneys [all surviving females and approximately half of
each male exposure group], brain, thyroid, lungs, adrenals, stomach, and testes [from
approximately half of each male exposure group]), and microscopic examination of 14 tissues
(thyroid, lungs, heart, liver, spleen, kidneys, adrenals, stomach, small intestine, large intestine,
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pancreas, bone marrow, testes or ovaries, and uterus). Animals that died prior to study
termination were subjected to gross pathology evaluations when possible. It is not clear if a
complete microscopic evaluation was conducted on animals that died prior to study termination;
however, the lungs of animals that died prior to terminal sacrifice were microscopically
examined. It does not appear that any of the data were subjected to statistical analyses.
Results are summarized in Table 1 and Figure 1 (Horn et al., 1957). Survival of rats was
not adversely affected by chronic dietary exposure to hexanedioic acid, and the occurrence of
clinical signs of toxicity was comparable in treated and control rats. Body weights of males
exposed to 3% or 5% hexanedioic acid were less than controls throughout the 2-year exposure
period (17% and 32% less than controls at Week 8, and remaining below controls throughout the
study, with terminal deficits of 9% and 18%, respectively). Food consumption of males exposed
to 5% was slightly (~6 %) lower than controls. Body weights and food intake of all other male
and female treated groups were similar to controls throughout the exposure period. There was no
effect on organ weights or the incidence of tumors or nonneoplastic gross or microscopic lesions
in any of the tissues examined at any dose. Based on the body weight changes, this study
identified a LOAEL of 3% (1500 mg/kg-day) and NOAEL of 1% (470 mg/kg-day). Although
this study appears to have been conducted at appropriate doses, with the high dose of 5%
approaching the MTD (maximum tolerated dose), it was limited as a carcinogenicity bioassay by
the small number of animals evaluated.
Table 1. Mean Body Weight, Survival, and Feed Consumption of Male and Female Rats
Exposed to Hexanedioic Acid in the Diet for 2 Years (Horn et al., 1957)
Concentration
Dose
(mg/day)
Dose
(mg/kg-day)a
Avg Body Wt
(Weeks 0-104)
Survival13
Mean Daily
Food Intake (g)
Males
0%
0
0
368.9
82.5%
16.8
0.1%
17
44
382.7
87.7%
17.0
1%
175
470
374.1
94.7%
17.5
3%
505
1500
332.5
94.5%
16.8
5%
814
2800
291.5
97.2%
15.8
Females
0%
0
0
259.6
98.9%
14.2
1%
158
630
251.1
96.3%
15.8
Calculated for the current review using body weight data reported in Table II of the publication.
''Reported by the researchers to account for both number of survivors and length of survival.
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Figure 1. Mean Body Weight of Male Rats Exposed to
Adipic Acid in the Diet for 2-years

0%
0.1%
1%
3%
5%
0 8 16 24 32 40 48 56 64 72 80 88 96 104
Treatment Week
There is evidence that toxicity of hexanedioic acid is enhanced by oral gavage
administration compared with dietary exposure. Short-term gavage studies found severe effects,
including death, at doses comparable to the subchronic/chronic dietary NOAELs/LOAELs. In a
range-finding study for their chronic study, Horn et al. (1957) gave male albino mice (13/dose) a
single dose of hexanedioic acid (6% suspension in methylcellulose) at 1500, 2000, or
2500mg/kg-day. Mortality was observed at all doses and increased with dose (3/13, 8/13, and
9/13 in the low- mid- and high-dose groups, respectively). Animals that died showed distention
of the stomach and small intestine, irritation and hemorrhage of the intestine, and spastic
contractions of the cecum. An LD50 of 1900 mg/kg-day was calculated. In another study,
treatment of male rats with 3600, 4000, 5000, or 5600 mg/kg-day by oral gavage for 5 days
resulted in mortality at all doses (3/6, 5/6, 6/6, 6/6, and 6/6, respectively), and a calculated LD50
of 3615 mg/kg-day (Litton Bionetics, 1974). Clinical signs of toxicity included depression,
labored respiration, ataxia, and convulsions in all dose groups. No lesions were found at
necropsy. Dietary exposure studies found no clinical signs of toxicity and no effect on survival,
but did find gastrointestinal lesions, at comparable doses.
Results from several short-term repeated-dose studies were reported in a review by
Informatics (1974). The original study reports were not available for review, and very few
details on the experimental designs or results were provided in these summaries; therefore, study
adequacy and reliability cannot be independently assessed. These data are summarized in Table
2.
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Table 2. Summary of Selected Repeated-Dose Studies Reported by Informatics, 1974
Study
Duration
Exposure
Route
Species
NOAEL /
LOAEL
Citation
4 Weeks
Adipic acid
by oral
gavage
(vehicle not
specified)
Rat (young)
NOAEL: 243 mg/d
(1350 mg/kg-d)
LOAEL: None
Enders, 1941, as cited
in Informatics, 1974
4 Weeks
Adipic acid
by oral
gavage
(vehicle not
specified)
Rat (adult)
NOAEL: 730 mg/d
(2433 mg/kg-d)
LOAEL: None
Enders, 1941, as cited
in Informatics, 1974
5 Weeks
Adipic acid
by oral
gavage in
ethanol
Rats (adult)
NOAEL: 200 mg/d
(610 - 922 mg/kg-d)
LOAEL: None
NAS, 1943 as cited in
Informatics, 1974
1 Week
Adipic acid
by oral
capsule
Guinea Pig
NOAEL: 400 mg/d
(682-942 mg/kg-d)
LOAEL: None
NAS, 1943 as cited in
Informatics, 1974
5 Weeks
Adipic acid
by oral
capsule
Guinea Pig
NOAEL: 600 mg/d
(1032-1739 mg/kg-d)
LOAEL: None
NAS, 1943 as cited in
Informatics, 1974
9 Weeks
Sodium
adipate by
unspecified
route
Rats (young)
NOAEL: None
LOAEL: 199 mg/d
(638-1332 mg/kg-d)
Reduced body weight
NAS, 1943 as cited in
Informatics, 1974
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Hexanedioic acid was the subject of a series of teratology studies in rats, mice, hamsters,
and rabbits conducted on behalf of the U.S. Food and Drug Administration (FDRL, 1972, 1974).
These studies followed similar protocols; hexanedioic acid was administered via oral gavage to
the following:
1.	25-31 mated CD-I mice/dose on gestation days (GD) 6-15 at 0, 2.6, 12.0, 56.0, or 263
mg/kg-day;
2.	24-28 mated Wistar rats/dose on GDs 6-15 at 0, 2.9, 13.0, 62.0, or 288 mg/kg-day;
3.	25-27 mated Golden hamsters/dose on GDs 6-10 at 0, 2.0, 9.5, 44.0, or 205 mg/kg-
day; and
4.	13-20 artificially inseminated Dutch-belted rabbits on GDs 6-18 at 0, 2.5, 12, 54, and
250 mg/kg-day.
In each study, body weights were determined on gestation day (GD) 0, at one or two
intervals before treatment ended, on the last day of hexanedioic acid treatment, and at terminal
sacrifice. Appearance, behavior, and food consumption were evaluated daily. Two to 11 days
after the final hexanedioic acid administration, pregnant animals were subjected to Caesarean
section and the following parameters were recorded: number of corpora lutea (rabbits only),
implantation sites, resorption sites, and live and dead fetuses. A detailed examination of the
urogenital tract of each pregnant female was performed, fetal weights of live pups were recorded,
and a gross examination for external congenital abnormalities was performed on all fetuses
(survival of neonatal rabbits was evaluated after placing live fetuses in an incubator for 24
hours). All surviving rabbit fetuses were dissected and examined for visceral abnormalities, then
cleared, stained, and examined for skeletal defects. For mice, rats, and hamsters, approximately
one-third of the fetuses of each litter were subjected to visceral examination, the rest were
stained and evaluated for skeletal defects. Aspirin or 6-aminonicotinamide was used as a
positive control in each study. Key parameters of each study design are summarized in Table 3.
Hexanedioic acid treatment did not affect implantation of the conceptus into the uterus,
maternal or fetal survival, or the incidence of soft tissue or skeletal tissue abnormalities. The
NOAEL in rats, mice, hamsters, and rabbits was 288, 263, 205, and 250 mg/kg-day, respectively,
the highest dose evaluated in each study. Although hexanedioic acid did not induce
developmental toxicity in any of the species tested, the positive control substance did not clearly
induce developmental toxicity in mice or hamsters; therefore, it is not clear that the assay was
adequately sensitive to detect positive responses in these species. Also, maternal toxicity was
not achieved in any of the studies; therefore, a definitive conclusion cannot be made regarding
the ability of hexanedioic acid to induce developmental toxicity. These data do, however,
support the conclusion that teratogenicity is not likely a sensitive toxicological endpoint for
hexanedioic acid.
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Table 3. Selected Study Design Parameters of FDRL, 1972,1974
Strain
Rat
Mouse
Hamster
Rabbit
Wistar
Albino CD-I
outbred
Golden
Dutch-belted
Number of
Animals Dosed
24 to 28 per
dose
25 to 31 per dose
25 to 27 per dose
13 to 20 per dose
Number of
Animals
Evaluated
(pregnant)
20-24 per dose
20-24 per dose
21-24 per dose
10-14 per dose
Doses
2.9, 13, 62,
288 mg/kg-
day
2.6, 12, 56, 263
mg/kg-day
2, 9.5, 44, 205
mg/kg-day
2.5, 12, 54, 250,
mg/kg-day
Dosing Volume
1-2 mL/kg
10 mL/kg
1 mL/kg
1 mL/kg
Dosing Schedule
GD 6-15
GD 6-15
GD 6-10
GD 6-18
Sacrifice Day
GD 20
GD 17
GD 14
GD 29
Parameters
Evaluated
Number of corpora lutea (rabbits only), implantation sites, resorption sites, and
live and dead fetuses, urogenital tract normality of dams or does, fetal weight,
external congenital abnormalities (gross examination), visceral abnormalities, and
skeletal defects.
Negative Control
Water
Water
Water
Water
Positive Control
Aspirin
(150 mg/kg)
Aspirin
(250 mg/kg-day)
Aspirin
(250 mg/kg-day)
6-
Aminonicotinamide
(2.5 mg/kg, Day 9)
Inhalation Exposure. Only one study was located regarding the inhalation toxicity of
hexanedioic acid in animals. Alderley Park specific-pathogen-free rats (two per sex, weighing
approximately 200 grams) were exposed to powdered hexanedioic acid at 126 mg/m3 via
inhalation in a dynamic chamber 6 hours/day, 5 days/week for three weeks (Gage, 1970). Body
weight and clinical signs of toxicity were monitored throughout the exposure period. Rats were
sacrificed after 3 weeks of exposure and the following parameters were evaluated: unspecified
urinalysis and hematology parameters, gross pathology, and microscopic pathology of 5 tissues
(lungs, liver, kidneys, spleen and adrenals). The study report also indicated that the heart,
jejunum, ileum, and thymus were "occasionally" examined. No effects on any endpoint were
observed, making the 126 mg/m3 concentration used in this study a free standing NOAEL.
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Other Studies
Hexanedioic acid was not mutagenic in Salmonella typhimurium strains TA98, TA100,
TA1535, TA1537, or TA1538 or in Escherichia coli (WP2wvrA), with or without addition of
exogenous metabolic activation (Prival et al., 1991). Results were also negative in tests for
mutagenicity in S. typhimurium strains TA1530 or G46 without activation in vitro and in host-
mediated assays with these strains in mice given either a single dose or five consecutive daily
doses of hexanedioic acid (Litton Bionetics, 1974). Hexanedioic acid did not cause a significant
increase in Saccharomyces cerevisiae D3 recombinants in vitro and produced little or no
response in host-mediated assays with this strain in mice (Litton Bionetics, 1974). Hexanedioic
acid did not induce chromosomal aberrations in human embryonic lung cultures (WI-38) in vitro
or in rat bone marrow in vivo, and did not induce dominant lethal mutations in an in vivo assay in
male rats (Litton Bionetics, 1974). The compound also did not induce chromosomal
nondisjunction in Drosophila (Ramel and Magnusson, 1979). A cell transformation assay in
Syrian hamster embryo (SA7/SHE) cells was negative (Heidelberger et al., 1983).
DERIVATION OF PROVISIONAL SUBCHRONIC AND CHRONIC RfDs
FOR HEXANEDIOIC ACID
No relevant data were located regarding the subchronic or chronic toxicity of hexanedioic
acid to humans following oral exposure. The most widely reported and most sensitive effect in
animal feeding studies was reduced body weight gain, which was observed at doses of 1500-
2800 mg/kg-day with chronic exposure (Horn et al., 1957) and 1800-5000 mg/kg-day with
subchronic exposure (Lang and Bartsch, 1953; Hazleton, 1950; Litton Bionetics, 1974). In one
of the subchronic studies, the reduction in weight gain was accompanied by marked chronic
inflammation of the intestinal mucosa, as well as slight liver and kidney lesions (Lang and
Bartsch, 1953). No intestinal (or other) lesions were seen in the chronic study (Horn et al.,
1957). Horn et al. (1957) did, however, observe intestinal irritation and hemorrhage at >1500
mg/kg in an acute range-finding study in mice exposed by oral gavage in methylcellulose. This
suggests that hexanedioic acid is irritating to the intestinal mucosa by concentrated bolus
exposure, and that differences in dietary composition or feeding regimen may explain the
different findings in the subchronic and chronic studies. However, gastrointestinal lesions were
not directly related to the effect on body weight gain, which was found with or without the
lesions. A series of gestational exposure studies found no evidence to suggest that the
developing fetus is a sensitive target for hexanedioic acid (FDRL, 1972, 1974).
The chronic study by Horn et al. (1957), which found both the lowest LOAEL for
hexanedioic acid and a corresponding NOAEL, is suitable for both subchronic and chronic RfD
derivation. In this study, male rats were maintained on a diet supplemented with 0, 0.1%, 1%,
3%, or 5% hexanedioic acid (approximately 0, 44, 470, 1500, or 2800 mg hexanedioic acid/kg-
day). Females were only exposed at 0% or 1% (approximately 0 or 630 mg/kg-day,
respectively). The only effect observed in this study was decreased body weight at the 3% and
5% dietary concentrations (-1500 and 2800 mg/kg-day). Body weights at these concentrations
were substantially reduced at all reported time intervals (> Week 8). The use of the 2-year study
as the basis for the subchronic study is justified because the observed effect (decreased body
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weight) occurred at subchronic durations (>8 weeks) and persisted for the entire 2-year study
period. The study included gross necropsy, weight determinations for the major organs, and
microscopic examination of 14 tissues. The NOAEL was 470 mg/kg-day. Application of a
composite uncertainty factor of 300 (10 for extrapolation from rats to humans, 10 to protect
sensitive individuals, and 3 for deficiencies in the database, including absence of reproduction
toxicity studies) to the NOAEL of 470 mg/kg-day yields provisional subchronic and chronic
RfDs of 2 mg/kg-day for hexanedioic acid, as follows:
subchronic p-RfD/p-RfD = NOAEL / UF
= 470 mg/kg-day / 300
= 2 mg/kg-day or 2E-0 mg/kg-day
Confidence in the critical study is medium. The study included investigations of key
systemic endpoints in multiple dose groups and identified a NOAEL and LOAEL. The
sensitivity of the test may have been somewhat compromised by the lack of clinical chemistry or
hematology evaluations and the limited number of tissues microscopically examined.
Confidence in the database is medium because of the lack of reproductive toxicity studies and
the lack of available data on some commonly evaluated toxicological parameters (e.g., clinical
chemistry). Confidence in the subchronic and chronic p-RfDs for hexanedioic acid is, therefore,
medium.
FASEB (1976) estimated per capita consumption of hexanedioic acid in the U.S. to be
about 0.8 mg/kg-day, based on the quantity of the chemical used in foods in 1970. This estimate
was considered to be high because wastage and other losses were not taken into account.
Estimated intake based on market surveys was as high as 8 mg/kg-day for chronic exposure, but
these data were considered to be less reliable (FASEB, 1976). On the basis of more recent
production data, WHO (2000) estimated per capita daily intake of about 18 mg (0.26 mg/kg-day)
for hexanedioic acid from its use as a flavoring agent in the U.S. Based on the most recent and
most reliable data available, intake of hexanedioic acid in the U.S. is less than 0.8 mg/kg-day,
and probably less than 0.26 mg/kg-day. The provisional RfD of 2 mg/kg-day, therefore, is at
least 2-10 fold higher than the best estimate of chronic intake of hexanedioic acid in the U.S.
FEASIBILITY OF DERIVING PROVISIONAL SUBCHRONIC AND CHRONIC
RfCs FOR HEXANEDIOIC ACID
Short-term (3 weeks) inhalation exposure to hexanedioic acid powder at a concentration
of 126 mg/m3 resulted in no observed toxic effects in male or female rats (Gage, 1970).
However, the limitations of this study (e.g., use of only 2 rats/sex, evaluation of few
toxicological endpoints, microscopic evaluation of only 5 tissues, inclusion of only a single dose
level, short exposure duration) preclude its use for p-RfC derivation. No other repeated-dose
inhalation toxicity studies were located. The lack of inhalation data precluded the derivation of
non-cancer inhalation toxicity values.
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PROVISIONAL CARCINOGENICITY ASSESSMENT FOR
HEXANEDIOIC ACID
Weight-of-evidence Classification
No evidence of carcinogenicity was observed in a 2-year dietary exposure study in rats
(Horn et al., 1957). The study, however, was limited by the small number of animals evaluated.
A fairly broad array of genotoxicity studies found no evidence that hexanedioic acid is a genetic
toxicant. Overall, there is inadequate information to assess carcinogenic potential of
hexanedioic acid under the Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005).
Quantitative Estimates of Carcinogenic Risk
Derivation of quantitative estimates of cancer risk for hexanedioic acid is precluded by
the lack of data demonstrating carcinogenicity associated with hexanedioic acid exposure.
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