United States
Environmental Protection
1=1 m m Agency
EPA/690/R-09/072F
Final
9-30-2009
Provisional Peer-Reviewed Toxicity Values for
Xylenes (CASRN 1330-20-7)
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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Commonly Used Abbreviations
BMD
Benchmark Dose
IRIS
Integrated Risk Information System
IUR
inhalation unit risk
LOAEL
lowest-observed-adverse-effect level
LOAELadj
LOAEL adjusted to continuous exposure duration
LOAELhec
LOAEL adjusted for dosimetric differences across species to a human
NOAEL
no-ob served-adverse-effect level
NOAELadj
NOAEL adjusted to continuous exposure duration
NOAELhec
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
p-RfD
provisional oral reference dose
RfC
inhalation reference concentration
RfD
oral reference dose
UF
uncertainty factor
UFa
animal to human uncertainty factor
UFC
composite uncertainty factor
UFd
incomplete to complete database uncertainty factor
UFh
interhuman uncertainty factor
UFl
LOAEL to NOAEL uncertainty factor
UFS
subchronic to chronic uncertainty factor
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PROVISIONAL PEER-REVIEWED TOXICITY VALUES FOR
XYLENES (CASRN 1330-20-7)
Background
On December 5, 2003, the U.S. Environmental Protection Agency's (U.S. EPA) 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)	U.S. EPA's Integrated Risk Information System (IRIS).
2)	Provisional Peer-Reviewed Toxicity Values (PPRTVs) used in U.S. 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 U.S. EPA's 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 U.S. EPA IRIS Program. All provisional toxicity values receive internal review by
two U.S. EPA scientists and external peer review by three independently selected scientific
experts. PPRTVs differ from IRIS values in that PPRTVs do not receive the multiprogram
consensus review provided for IRIS values. This is because IRIS values are generally intended
to be used in all U.S. EPA programs, while PPRTVs are developed specifically for the Superfund
Program.
Because new information becomes available and scientific methods improve over time,
PPRTVs are reviewed on a 5-year basis and updated into the active database. 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 documents conclude that
a PPRTV cannot be derived based on inadequate data.
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 Resource Conservation and Recovery Act (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.
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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 document and understand the strengths
and limitations of the derived provisional values. PPRTVs are developed by the U.S. EPA
Office of Research and Development's National Center for Environmental Assessment,
Superfund Health Risk Technical Support Center for OSRTI. Other U.S. 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 U.S. EPA Office of Research and Development's National Center for Environmental
Assessment, Superfund Health Risk Technical Support Center (513-569-7300), or OSRTI.
INTRODUCTION
A streamlined approach was used to derive provisional subchronic RfD and RfC values
for xylenes. Xylenes have a chronic RfD, a chronic RfC, and cancer assessment on IRIS
(U.S. EPA, 2008)—so only subchronic toxicity values are derived in this assessment. Xylenes
have recently been reassessed by the IRIS program and a Toxicological Review (U.S. EPA,
2003) is available. In addition, the Agency for Toxic Substances and Disease Registry (ATSDR)
Toxicological Profile for xylenes has been updated recently (ATSDR, 2007). Both the IRIS
Toxicological Review and the ATSDR Toxicological Profile contain comprehensive overviews
of the toxicology and toxicokinetics information available on xylenes. Although all of the
exposure duration-relevant studies reviewed by ATSDR (2007) are included in the 2003 IRIS
review of xylenes, updated literature searches were conducted from January 2002 to July 2007
for studies relevant to the derivation of subchronic toxicity values for xylenes; Appendix C
provides a description of the literature search process. No new studies with the ability to inform
the derivation of subchronic provisional toxicity values were identified. As such, given the
availability of the recent IRIS and ATSDR reviews, these reports were used to identify the
exposure duration-relevant critical studies and endpoints for use in deriving the subchronic
values.
The derivation of subchronic toxicity values for xylenes is discussed below. A brief
rationale is provided for the selection of the critical study and endpoint, a summary of the critical
study is presented, and the subchronic toxicity-value-derivation process is described. For further
information on the toxicology and toxicokinetics of xylenes, the reader may consult the IRIS
records (attached to this report as Appendix A), IRIS Toxicological Review document
(U.S. EPA, 2003), or ATSDR (2007) Toxicological Profile for xylenes.
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REVIEW OF PERTINENT DATA AND DERIVATION OF PROVISIONAL
SUBCHRONIC TOXICITY VALUES FOR XYLENES
Subchronic p-RfD
The chronic RfD for xylenes (0.2 mg/kg-day) on IRIS (consensus date January 2003) is
based on mortality and decreased body weight in a chronic rat study (NTP, 1986). The ATSDR
intermediate-duration oral minimal risk level (MRL) of 0.4 mg/kg-day for mixed xylenes was
derived in August 2007 based on neurological effects (hyperactivity) in a chronic mouse gavage
study (NTP, 1986). NTP (1986) reported that the effects were first observed beginning in
Week 4 of the study and were considered to result from intermediate-duration exposure
(e.g., ATSDR considers intermediate duration to span from >14 days to 1 year).
All of the exposure duration-relevant studies reviewed by ATSDR (2007) are included in
the 2003 IRIS review of xylenes (i.e., no new studies were published between 2003 and
July 2007). Thus, the IRIS Toxicological Review was consulted to identify studies that might be
relevant to the derivation of a subchronic p-RfD for xylenes (e.g., oral subchronic,
developmental or reproductive toxicity studies); these studies were compared with the
NTP (1986) chronic mouse study that served as the basis for the intermediate duration oral MRL
for xylenes (ATSDR, 2007). Neither body weight nor survival rates (critical effects for the
chronic study), used as the basis of the chronic RfD on IRIS, were affected in the first few
months of the chronic rat study (NTP, 1986); as such, this study was not considered suitable for
use in the derivation of a subchronic p-RfD. Table 1 compares the NOAELs, LOAELs, and
endpoints of the available studies. In the table, the NOAELs and LOAELs adjusted for
continuous exposure are also presented because several of the available studies (all reported by
NTP, 1986) used gavage administration on 5 days/week.
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Table 1. Available Studies for Subchronic p-RfD Derivation for Xylenes
Species
Sex
Doses
(mg/kg-day)
Exposure
NOAEL
(mg/kg-
day)
LOAEL
(mg/kg-
day)
Duration-
Adjusted
NOAEL3
(mg/kg-
day)
Duration-
Adjusted
LOAEL3
(mg/kg-
day)
Responses
Comments
Reference
Rat
Subchronic
Gavage
Study
M/F
0, 62.5, 125,
250, 500,
1,000
5 days/week for
13 weeks
500
1,000
357
714
Decreased body
weight in males
Mixed xylenes;
included 17%
ethylbenzene.
Females also
exhibited reduced
body weight gain
at the high dose
NTP, 1986
Rat
Subchronic
Gavage
Study
M/F
0, 150, 750,
1,500
Daily for 90
consecutive days
150
750
150
750
Increased kidney
weights in males
and early
appearance of
nephropathy in
females
Mixed xylenes
Condie et al.,
1988
Rat
Subchronic
Gavage
Study
M/F
0, 100, 200,
800
Daily for 90
consecutive days
200
800
200
800
Decreased body
weight in males
m-Xylene tested.
Female body
weight was also
reduced at the high
dose
Wolfe, 1988a
Rat
Subchronic
Gavage
Study
M/F
0, 100, 200,
800
Daily for 90
consecutive days
200
800
200
800
Early mortality in
males
£>-Xylene tested;
mortality may have
been related to
aspiration of test
material into lungs
Wolfe, 1988b
Mouse
Subchronic
Gavage
Study
M/F
0, 125, 250,
500, 1,000,
2,000
5 days/week for
13 weeks
1,000
2,000
714
1,430
Transient signs of
nervous system
depression
Mixed xylenes;
included 17%
ethylbenzene
NTP, 1986
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Table 1. Available Studies for Subchronic p-RfD Derivation for Xylenes
Species
Sex
Doses
(mg/kg-day)
Exposure
NOAEL
(mg/kg-
day)
LOAEL
(mg/kg-
day)
Duration-
Adjusted
NOAEL3
(mg/kg-
day)
Duration-
Adjusted
LOAEL3
(mg/kg-
day)
Responses
Comments
Reference
Mouse
Gavage
Develop-
mental
Toxicity
F
0, 520,
1,030, 2,060,
3,100, 4,130
Daily during
GD 6-15
1,030
(maternal
and
develop-
mental)
2,060
(maternal
and
develop-
mental)
1,030
2,060
Decreased gravid
uterine weight in
dams; decreased
fetal weight and
increased
incidence
malformations in
offspring
Mixed xylenes
Marks et al.,
1982
Mouse
Chronic
Gavage
Study
M/F
0, 500, 1,000
5 days/week for
103 weeks
500
1,000
357
714
Hyperactivity
beginning Week 4
Mixed xylenes;
included
17% ethylbenzene.
This study was
used as the basis
for the AT SDR
intermediate MRL
NTP, 1986
'Adjusted for continuous exposure where applicable, as follows: Adjusted NOAEL = NOAEL x 5/7 days/week. Adjusted
LOAEL = LOAEL x 5/7 days/week.
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As the table shows, several subchronic rat studies (NTP, 1986; Condie et al., 1988;
Wolfe, 1988a,b) and the chronic mouse study (NTP, 1986) identify adjusted LOAELs in the
same range (700-800 mg/kg-day). A LOAEL of 800 mg/kg-day was identified for the data in
Wolfe (1988b) based on early mortality; however, U.S. EPA (2003) indicated that some of the
mortality may have been related to aspiration of the test material into the lungs. Thus, this study
is not considered suitable for use in deriving the subchronic p-RfD. Condie et al. (1988)
identified a LOAEL of 750 mg/kg-day for increased kidney weights and early appearance of
nephropathy in rats. However, as noted by U.S. EPA (2003), kidney effects were not observed
in other subchronic studies in rats (NTP, 1986; Wolfe, 1988a, 1988b) nor in the chronic rat study
(NTP, 1986). Consequently, this endpoint is not considered for use in deriving the subchronic
p-RfD. NTP (1986) and Wolfe (1988a) identify LOAELs of 714 and 800 mg/kg-day,
respectively, for decreased body weight; this endpoint is also a critical effect observed in the
chronic rat study (NTP, 1986). The chronic mouse study (NTP, 1986) identifies a LOAEL of
714 mg/kg-day for transient hyperactivity beginning after 4 weeks of the study. ATSDR (2007)
considered this to be an effect of intermediate duration (<1 year) exposure and used it as the
basis for the intermediate oral MRL. While there are few data on the neurotoxicity of orally
administered xylenes, neurological effects are a known critical hazard of inhalation exposure (for
review, see Ritchie et al., 2001) and serve as the basis for the chronic RfC.
In the absence of a compelling reason to select any one of these three studies over the
other (subchronic rat studies by NTP, 1986 and Wolfe, 1988a and the chronic mouse study by
NTP, 1986), all three were considered as potential principal studies for the purpose of deriving a
subchronic p-RfD for xylenes.
Summaries of the these studies are excerpted from the U.S. EPA (2003) Toxicological
Review for Xylenes and reproduced here for the reader's convenience.
In the NTP (1986) subchronic rat study, groups of 10 male and 10 female Fischer
344 rats were administered mixed xylenes (60% m-xylene, 13.6% p-xylene,
9.1% o-xylene and 17.0% ethylbenzene) in corn oil by gavage at doses of 0, 62.5, 125,
250, 500 or 1000 mg/kg-day for 5 days per week for 13 weeks. At termination of the
study, necropsy was performed on all animals and comprehensive histologic
examinations were performed on vehicle and high dose-group animals. High-dose males
andfemales gained 15% and 8% less body weight, respectively, than did controls, with
final body weights being 89% and 97%, respectively, of those of controls (statistical
significance not reported). No signs of toxicity or treatment-related gross or microscopic
pathologic lesions were observed. The LOAEL is 1000 mg/kg-day, based on decreased
body weights in male rats and the NOAEL is 500 mg/kg-day. After adjustment for
continuous exposure, the LOAEL and NOAEL are 714 and 357 mg/kg-day, respectively.
In the study by Wolfe (1988a), groups of 20 male and 20 female Spr ague-Daw ley rats
were administered m-xylene (99% purity) by gavage in corn oil at doses of0, 100, 200 or
800 mg/kg-day for 90 consecutive days. Survival incidences were 20/20, 17/20, 15/20
and 18/20, respectively, for males and 20/20, 20/20, 16/20 and 16/20, respectively, for
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females. Mortality in the mid-dose males and mid- and high-dose females attained
statistical significance (p < 0.05), but a significant trend was observed only in females.
Mottled lungs and a failure of the lungs to collapse were observed in all mid- and
high-dose animals that died early and in 2/3 of the low-dose males that died early, but
were not evident in any of the animals that survived to study termination.
Histopathologic examination of the lungs from animals that died before study termination
revealedforeign material in the alveoli in all but one animal. Therefore, these deaths
were attributed to vehicle and/or compound aspiration. Clinical signs present
throughout the study were limited to high levels of salivation prior to dosing in high-dose
males and females. Body-weight gains over the entire study period were decreased
(p <0.05) in mid- and high-dose males (89% and 75%, respectively, of control weight
gain) and high-dose females (85%). Food consumption was likewise decreased
(p < 0.05) in high-dose males during weeks 15 (90% of control levels) and in mid- and
high-dose males during weeks 6 9 (92% of control levels for both groups). A thorough
histologic examination revealed no other abnormalities. Other effects noted were not
definitively related to treatment and/or were not biologically significant. The NOAEL
and LOAEL are identified as 200 and 800 mg/kg-day, respectively, based on decreased
body weight in males.
In the chronic mouse study (NTP, 1986), groups of 50 male and 50 female B6C3F1 mice
were administered mixed xylenes (60% m-xylene, 13.6% p-xylene, 9.1% o-xylene,
17.0% ethylbenzene) in corn oil by gavage at doses of 0, 500 or 1000 mg/kg-day for
5 days per week for 103 weeks. Necropsy and histologic examinations were performed
on all animals. Tissues were examinedfor gross lesions and masses. The tissues
examined included mandibular lymph nodes, salivary gland, femur (including marrow),
thyroid gland, parathyroids, small intestine, colon, liver, prostate/testis or ovaries/uterus,
heart, esophagus, stomach, brain, thymus, trachea, pancreas, spleen, skin, lungs and
mainstem bronchi, kidneys, adrenal glands, urinary bladder, pituitary gland, eyes (if
grossly abnormal) and mammary gland. Hematology and clinical chemistry analyses
were not conducted. No statistically significantly increased incidences of nonneoplastic
or neoplastic lesions were found in male or female exposed groups when compared with
controls. The only treatment-related effect observed was hyperactivity, which occurred
in all high-dose mice of each sex 5-30 minutes after dosing. This effect was observed
consistently beginning at week 4 and continued until study termination at 103 weeks. The
LOAEL is 1000 mg/kg-day and the NOAEL is 500 mg/kg-day for hyperactivity. After
adjustment for continuous exposure, the LOAEL and NOAEL are 714 and
357 mg/kg-day, respectively.
The potential principal studies (NTP, 1986; Wolfe, 1988a) identify LOAELs of 714 or
800 mg/kg-day and NOAELs of 200 or 357 mg/kg-day. The principal observation in the chronic
mouse study (NTP, 1986) is transient hyperactivity after dosing. The incidence of this effect was
100% in both male and female mice exposed for at least 4 weeks at the LOAEL. The incidence,
if any, of hyperactivity at lower doses was not reported, precluding the use of benchmark dose
(BMD) modeling on this endpoint. The critical effect in the subchronic rat studies is decreased
body weight (>10% difference from controls) in male rats. The body-weight decrement was
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observed in both male and female rats in two different subchronic studies (Wolfe et al., 1988a;
NTP, 1986), as well as in the chronic study (NTP, 1986), and the effect exhibited
dose-dependence in male rats. In addition, the chronic RfD on IRIS is based on decreased body
weight in rats in the chronic NTP (1986) study. Data on decreased body weight in male rats
were considered for benchmark dose modeling to derive a point of departure (POD) for the
subchronic p-RfD. Modeling of the final body weights of male rats in the subchronic study
reported by NTP (1986) did not result in model fit by U.S. EPA (2000) criteria (Appendix B
describes the modeling approach and provides results). In contrast, modeling of the data on final
body weights of male rats reported by Wolfe (1988a) using the linear model with constant
variance provided adequate fit to both the variance and means data. Appendix B describes the
modeling effort and results. The BMD and BMDL associated with a 10% decrease in body
weight (compared with the control mean) were 538 and 440 mg/kg-day, respectively. The
BMDL is selected as the POD for subchronic p-RfD derivation. A composite UF of 1000 is
applied to the BMDL of 440 mg/kg-day to derive a subchronic p-RfD for xylenes, as shown
below.
Subchronic p-RfD = BMDL UF
= 440 mg/kg-day ^ 1,000
= 0.4 or 4 x 10"1 mg/kg/-day
The composite UF includes a factor of 10 for interspecies extrapolation, a factor of 10 for human
variability and 10 for database limitations, as follows:
•	An UF of 10 is applied to account for laboratory animal-to-human interspecies
differences in toxicokinetics and toxicodynamics (UFA).
•	An UF of 10 is applied for intraspecies uncertainty to account for human variability
and sensitive populations (UFH). This factor accounts for humans who may be more
sensitive than the general population to exposure to xylenes.
•	An UF of 1 for extrapolation from a LOAEL to NOAEL (UFL) is applied because the
current approach is to address this extrapolation as one of the considerations in
selecting a BMR for BMD modeling. In this case, a BMR corresponding to a change
in body-weight equal to one control standard deviation from the control mean
body-weight was selected under an assumption that it represents a minimal
biologically significant change.
•	An UF of 10 is applied to account for database uncertainty (UFD). The available
subchronic oral database for xylenes includes subchronic gavage toxicity studies in
mice and rats, and it includes a developmental toxicity study. However, the database
lacks adequate studies of the oral neurotoxicity of xylenes, as well as
multigenerational reproductive toxicity and developmental neurotoxicity studies.
Since neurological impairment is a critical health hazard from inhalation exposure to
xylenes and clinical signs of neurotoxicity have been observed after subchronic and
chronic exposure in mice (NTP, 1986), the lack of comprehensive neurotoxicity
testing is of particular concern.
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Confidence in the principal and supporting studies is medium. The studies include two
subchronic studies in rats and a chronic study (with effects observed at 4 weeks) in mice. The rat
studies identified one of the same critical endpoints as was used for the chronic RfD derivation
(body-weight decreases). Comprehensive histologic examinations of tissues were performed in
all three studies. Confidence in the oral subchronic toxicity database is low-to-medium because
the database contains several subchronic studies and an evaluation of the developmental effects,
but it is lacking oral neurotoxicity studies, multigenerational reproductive toxicity studies, and
developmental neurotoxicity studies. Low-to-medium confidence in the subchronic p-RfD
follows.
Subchronic p-RfC
Review of the data supporting the chronic RfC for xylenes on IRIS indicate that
subchronic data were used to derive the chronic value, and, thus, are appropriate to serve as the
basis for the subchronic p-RfC. The chronic RfC for xylenes (0.1 mg/m3) on IRIS (consensus
date January 2003) is based on neurological effects in a subchronic rat inhalation study
(Korsak et al., 1994). The derivation included a UF of 3 for subchronic-to-chronic extrapolation,
which is justified because the effects did not increase with longer exposure durations. The
intermediate duration inhalation MRL (0.6 ppm or 2.6 mg/m3) was derived in August 2007 and
was also based on neurological effects in the Korsak et al. (1994) study. Because
U.S. EPA (2003) used a subchronic study as the basis for the chronic RfC for xylenes and
ATSDR (2007), and an updated literature search did not identify any newer subchronic
inhalation studies, the subchronic p-RfC is based on the same critical study (Korsak et al., 1994),
endpoint (neurological effects), and POD (NOAELrec) as the chronic RfC—but without the UF
for subchronic-to-chronic extrapolation.
A summary of the critical study is excerpted from the U.S. EPA (2003) IRIS record for
xylenes and reproduced below. Additional study details are available in the U.S. EPA (2003)
Toxicological Review for xylenes.
Korsak et al. (1994) exposed groups of 12 male Wistar rats by inhalation to 0, 50 or
100 ppm m-xylene or n-butyl alcohol or a 1:1 mixture (purity of chemicals not provided)
for 6 hours per day, 5 days per week for 3 months and evaluated similar endpoints as in
the earlier study (Korsak et al., 1992). Rotarodperformance and spontaneous motor
activity were assayed. The report does not specify the timing of the neurologic
examinations; however, given that the 1994 study was conducted by the same group of
investigators as a 1992 study (Korsak et al., 1992) and that one of the tests (rotarod
performance) was the same in both studies, it appears reasonable to assume that the tests
were administered 24 hours after termination of exposure. The rotarod test was used as
a measure of motor coordination disturbances from exposure to m-xylene. The rotarod
test involves placing the subject animals on a rotating rod and evaluating their ability to
remain on the rod for a period of 2 minutes. The animals were trained to perform the
task, exposed to chemical or control gas and evaluated at defined intervals. By the time
interval after exposure, considerable proportions of absorbed xylenes are expected to
have been eliminatedfrom the body (see Toxicological Review, U.S. EPA, 2003). Body
weights and weights of seven organs were measured. Bloodfor clinical biochemistry
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(e.g., alanine aminotransferase, aspartate aminotransferase, sorbitol dehydrogenase,
alkaline phosphatase and total protein) and hematologic analysis (erythrocyte counts,
hemoglobin concentration, hematocrit, leukocyte count and differential leukocyte counts)
was collected 24 hours after termination of exposure. Statistical evaluations (using a
p = 0.05 level of significance) of the collected data included analysis of variance,
Dunnett's test and Fisher's exact test.
No statistically significant exposure-related changes were noted in body-weight gain,
absolute or relative organ weights, hepatic activities of microsomal monooxygenases,
lipid peroxidation or levels of triglycerides in the liver (Korsak et al., 1994). Statistically
significant decreases in erythrocyte number were seen in animals exposed to 50 ppm
(93% of controls) or 100 ppm (80.5% of controls) of m-xylene alone. Similarly,
decreased levels of hemoglobin were reported in both groups (92% of controls for both
groups). At 100ppm, a statistically significant increase in leukocyte number
(35% increase over controls) was reported. Exposure to 50 or 100ppm m-xylene alone
also resulted in decreased rotarod performance starting at 1 month of exposure, which
remained at the same level until the end of the 3-month exposure. Decreases were
statistically significant in the 100 ppm group when compared with the controls. The
results were presented in graphical form; the actual numerical data are not provided.
The decreases in performance were roughly 8% and 33% for the 50 and 100 ppm groups,
respectively, versus 0% for the controls.
Sensitivity to pain was assessed using the hot plate behavior test, in which the animals
are placed on a hot (54°C) surface and the time interval between being placed on the
plate and licking of the paws is measured (Korsak et al., 1994). Rats exposed to 50 or
100 ppm m-xylene alone had statistically significantly increased sensitivity to pain at the
end of the 3-month exposure (latency of the paw-lick response was 8.7 and 8.6 seconds,
respectively, us. 12.2 seconds for the controls). The LOAEL is 100 ppm, based on
decreased rotarod performance and decreased latency in the paw-lick response in the
hot-plate test and the NOAEL is 50 ppm.
As noted above, the subchronic p-RfC is based on the same critical study, effect, and
POD as the IRIS chronic RfC. For the chronic RfC, U.S. EPA (2003) used the NOAELrec (39
mg/m3) calculated from the data reported by Korsak et al. (1994) as the POD. The NOAELHec
was divided by a composite UF of 300 that includes a 3-fold UF for interspecies extrapolation
(dosimetric adjustments were used to extrapolate the toxicokinetic portion), a 10-fold UF for
intraspecies variation, a 3-fold UF for extrapolation from subchronic-to-chronic exposure
duration and a 3-fold UF for database deficiencies (reflecting a lack of multigeneration
reproductive toxicity study). Further detail on the UF selections is available in the IRIS record
(see Appendix A).
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For the derivation of a subchronic p-RfC, the NOAELrec of 39 mg/m3 was divided by a
composite UF of 100, resulting in a subchronic p-RfC calculated as follows:
Subchronic p-RfC = NOAELHec UF
= 39 mg/m3 - 100
= 0.4 or 4 x 10"1 mg/m3
The uncertainty factors included in the composite UF are the same as those used for the
chronic RfC—but without the UF for sub chronic-to-chronic extrapolation. The composite UF of
100 applied here includes a 3-fold UF for interspecies extrapolation (toxic dynamic portion
only), a 10-fold UF for intraspecies variation, and a 3-fold UF for database deficiencies as
follows:
•	An UF of 3 is applied to account for laboratory animal-to-human interspecies
differences (UFA). A factor of 3 is applied because default NOAELHec dosimetric
adjustments were used to calculate a human equivalent concentration (HEC),
reducing the uncertainty involved with the extrapolation from the results of an animal
study to a human exposure scenario (i.e., the toxicokinetic portion of the UF is 1; the
toxicodynamic portion of the UF is 3).
•	An UF of 10 is applied for intraspecies uncertainty to account for human variability
and sensitive populations (UFH). The degree of human variance in abilities to absorb
or dispose of xylenes is unknown, as is the degree of human variance in responding to
xylenes neurotoxicity. Results from developmental toxicity studies of rats exposed
by inhalation during gestation indicate that untoward developmental effects occur
only at higher doses than chronic doses producing the critical effects observed in
adult male rats in the principal and supporting studies. This suggest that the
developing fetus is not at special risk from low-level exposure to xylenes (please refer
to the IRIS Toxicological Review of Xylenes for details). However, as with oral
exposure, the effects of inhaled xylenes in other potentially sensitive populations such
as newborns or young children or animals have not been assessed.
•	An UF of 3 is applied for uncertainties in the database (UFD). The inhalation
database includes some human studies, subchronic studies in rats and dogs,
neurotoxicity studies, a one-generation reproductive toxicity study, developmental
toxicity studies, and developmental neurotoxicity studies. Although the available
developmental toxicity studies are confounded by a lack of litter incidence reporting,
the data reported for fetal incidences do not indicate effects at levels lower than that
found to induce neurologic impairment in several endpoints in male rats (please refer
to the IRIS Toxicological Review of Xylenes for details). The database is lacking a
two-generation reproductive toxicity study.
As discussed further in the IRIS Summary and Toxicological Review for xylenes,
confidence in the principal study is medium because the study examined a broad array of
endpoints including the critical effect of xylenes. However, only one sex of a single species was
examined and histologic examination of the animals was not performed. Confidence in the
database is medium, as it contains several subchronic studies as well as several developmental
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studies, developmental neurotoxocity studies, and a one-generation reproductive toxicity study;
however, a two-generation reproduction study is not available. As such, confidence in the
subchronic p-RfC is medium.
REFERENCES
ATSDR (Agency for Toxic Substances and Disease Registry). 2007. Toxicological Profile for
Xylenes. Agency for Toxic Substances and Disease Registry, Public Health Service,
U.S. Department of Health and Human Services. Draft for Public Comment. Online.
http://www.atsdr.cdc.gov/toxprofiles/tp71.html.
Condie, L.W., J.R. Hill and J.F. Borzelleca. 1988. Oral toxicology studies with xylene isomers
and mixed xylene. Drug Chem. Toxicol. 11:329-354.
Korsak, Z., J. A. Sokal and R. Gomy. 1992. Toxic effects of combined exposure to toluene and
m-xylene in animals. III. Subchronic inhalation study. Pol. J. Occup. Med. Environ. Health.
5(1 ):27—33.
Korsak, Z., J. Wisniewska-Knypl and R. Swiercz. 1994. Toxic effects of subchronic combined
exposure to n-butyl alcohol and m-xylene in rats. Int. J. Occup. Med. Environ. Health.
7:155-166.
NTP (National Toxicology Program). 1986. NTP technical report on the toxicology and
carcinogenesis of xylenes (mixed) (60% m-xylene, 13.6% p-xylene, 17.0% ethylbenzene, and
9.1% o-xylene) in F344/N rats and B6C3F1 mice (gavage studies). Research Triangle Park, NC.
NTP TR-327, NMPubl. No. 86-2583.
Ritchie, G.D., K.R. Still, W.K. Alexander, A.F. Nordholm, C.L. Wilson, J. Rossi 3rd and D.R.
Mattie. 2001. A review of the neurotoxicity risk of selected hydrocarbon fuels. J. Toxicol.
Environ. Health. B Crit Rev 4:223-312.
U.S. EPA. 2000. Benchmark Dose Technical Guidance Document [external review draft],
EPA/63O/R-00/001. Online, http://www.epa.gov/iris/backgr-d.htm.
U.S. EPA. 2003. Toxicological Review of Xylenes (CAS No. 1330-20-7) in Support of
Summary Information on the Integrated Risk Information System (IRIS). U.S. Environmental
Protection Agency, Washington, DC. EPA/635/R-03/001. Online.
http://www.epa.gov/iris/toxreviews/0270-tr.pdf.
U.S. EPA. 2008. Integrated Risk Information System (IRIS). Online. Office of Research and
Development, National Center for Environmental Assessment, Washington, DC. Online.
http://www.epa.gov/iris/.
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Wolfe, G.W. 1988a. Subchronic toxicity study in rats with m-xylene. Report by Hazleton
Laboratories America, Inc. Sponsored by Dynamac Corporation, Rockville, MD.
Project No. 2399-108.
Wolfe, G.W. 1988b. Subchronic toxicity study in rats with p-xylene. Report by Hazleton
Laboratories America, Inc. Sponsored by Dynamac Corporation, Rockville, MD.
Project No. 2399-110.
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APPENDIX A: PERTINENT SECTIONS FROM IRIS SUMMARY FOR
XYLENES: CHRONIC HEALTH HAZARD ASSESSMENTS FOR
NONCARCINOGENIC EFFECTS
Xylenes; CASRN 1330-20-7
Health assessment information on a chemical substance is included in IRIS only after a
comprehensive review of chronic toxicity data by U.S. EPA health scientists from several
Program Offices and the Office of Research and Development. The summaries presented in
Sections I and II represent a consensus reached in the review process. Background information
and explanations of the methods used to derive the values given in IRIS are provided in the
Background Documents.
STATUS OF DATA FOR Xylenes
File First On-Line 09/30/1987
M'Clllllll
SliilMS
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on-line
on-line
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02/21/2003
02/21/2003
02/21/2003
Oral RfD Assessment (I. A.)
Inhalation RfC Assessment (I.B.)
Carcinogenicity Assessment (II.)
I. Chronic Health Hazard Assessments for Noncarcinogenic Effects
I.A. Reference Dose for Chronic Oral Exposure (RfD)
Substance Name — Xylenes
CASRN — 1330-20-7
Last Revised — 02/21/2003
The oral Reference Dose (RfD) is based on the assumption that thresholds generally exist for
certain toxic effects such as cellular necrosis. It is expressed in units of mg/kg-day. In general,
the RfD is an estimate (with uncertainty spanning perhaps an order of magnitude) of a daily
exposure to the human population (including sensitive subgroups) that is likely to be without an
appreciable risk of deleterious effects during a lifetime. Please refer to the Background
Document for an elaboration of these concepts. RfDs can also be derived for the noncarcinogenic
health effects of substances that are also carcinogens. Therefore, it is essential to refer to other
sources of information concerning the carcinogenicity of this substance. If the U.S. EPA has
evaluated this substance for potential human carcinogenicity, a summary of that evaluation will
be contained in Section II of this file.
The RfD in this updated assessment replaces a previous RfD value of 2 mg/kg-day. The previous
and new RfD values are based on the same principal study (NTP, 1986). A database uncertainty
factor (UF) was not considered in the derivation of the previous RfD.
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The term xylenes refers to mixtures of the three xylene isomers (o-, m-, p-) and ethylbenzene. m-
Xylene is commonly the predominant component (40-77%) in commercial preparations of
xylenes (also referred to as mixed xylenes), with the other components each comprising roughly
up to 20% of the mass. The use of xylenes as a solvent, in paints and coatings, and in gasoline is
widespread. For the most part, studies cited in this assessment are conducted on mixed xylenes.
Results from studies comparing the toxicity of individual xylene isomers indicate that
differences, when they occur, are specific to the endpoint under consideration (see Section 4.4.3
of the Toxicological Review for more information).
	I.A.1. Oral RfD Summary
&
Decreased body weight,	NOAEL: 250 mg/kg-day	* 0.2 mg/kg-day
increased mortality	(179 mg/kg-day)*
Chronic F344/N rat study	LOAEL: 500 mg/kg-day
Oral gavage exposure
(NTP, 1986)
*Conversion Factors and Assumptions — 250 mg/kg-day x 5 days/7 days =179 mg/kg-day.
	I.A.2. Principal and Supporting Studies (Oral RfD)
The National Toxicology Program's 2-year study in rats was selected as the principal study and
the subchronic toxicity studies in rats by Wolfe (1988a, b) as supporting studies. In the
NTP (1986) study, groups of 50 male and 50 female Fischer 344 rats and 50 male and 50 female
B6C3F1 mice were administered mixed xylenes (60% m-xylene, 13.6% p-xylene,
9.1%) o-xylene, 17.0% ethylbenzene) in corn oil by gavage at doses of 0, 250, or 500 mg/kg-day
(rats) and 0, 500, or 1000 mg/kg-day (mice) for 5 days per week for 103 weeks. Necropsy and
histologic examinations were performed on all animals. Tissues were examined for gross lesions
and masses. The tissues examined included mandibular lymph nodes, salivary gland, femur
(including marrow), thyroid gland, parathyroids, small intestine, colon, liver, prostate/testis or
ovaries/uterus, heart, esophagus, stomach, brain, thymus, trachea, pancreas, spleen, skin, lungs
and mainstem bronchi, kidneys, adrenal glands, urinary bladder, pituitary gland, eyes (if grossly
abnormal), and mammary gland. Hematology and clinical chemistry analyses were not
conducted.
Effects of exposure in rats were limited to decreased body weight and decreased survival in
high-dose (500 mg/kg-day) males. Mean body weights were 5-8% lower in high-dose male rats
than in controls from week 59 to week 97, with body weights at 103 weeks being 4% less in
high-dose males than in controls (statistical significance not reported). Male rat survival rates
after 103 weeks showed a dose-related decrease (36/50, 25/50, and 20/50 for the control, low-,
and high-dose males, respectively). A life-table trend test for decreased survival incidence with
increasing dose was statistically significant (p=0.033). Pair-wise comparisons with control
survival incidence indicated that only the high-dose male rat incidence was significantly
decreased (p=0.04). A number of the deaths were attributed to gavage error (3/50, 8/50, and
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11/50, respectively, for the control, low-, and high-dose groups). The authors did not record
observations of rat behavior during dosing. Based on the available observations, the incidence of
treatment-related deaths demonstrated a dose-related increase (11/50, 17/50, and 19/50,
respectively [22%, 34%, and 38%]). The LOAEL is 500 mg/kg-day and the NOAEL is
250 mg/kg-day for decreased body weight and decreased survival. There was no evidence of
carcinogenicity in male or female rats exposed to doses up to 500 mg/kg-day.
In mice, the only treatment-related effect observed was hyperactivity, which occurred in all
high-dose mice of each sex, 5-30 minutes after dosing. This effect was observed consistently
beginning at week 4, and it continued until study termination at 103 weeks. The LOAEL is
1000 mg/kg-day and the NOAEL is 500 mg/kg-day for hyperactivity.
In a study by Wolfe (1988a), groups of 20 male and 20 female Sprague-Dawley rats were
administered m-xylene (99% purity) by gavage in corn oil at doses of 0, 100, 200, or
800 mg/kg-day for 90 consecutive days. Survival incidences were 20/20, 17/20, 15/20, and
18/20, respectively, for males, and 20/20, 20/20, 16/20, and 16/20 for females. Mortality in the
mid-dose males and mid- and high-dose females attained statistical significance (p<= 0.05), but a
significant trend was observed only in females. Mottled lungs and a failure of the lungs to
collapse were observed in all mid- and high-dose animals that died early and in 2/3 of the
low-dose males that died early but was not evident in any of the animals that survived to study
termination. Histopathologic examination of the lungs from animals that died before study
termination revealed foreign material in the alveoli in all but one animal. Therefore, these deaths
were attributed to vehicle and/or compound aspiration.
Clinical signs present throughout the study were limited to high levels of salivation prior to
dosing in high-dose males and females. Body weight gains over the entire study period were
decreased (p<= 0.05) in mid- and high-dose males (89% and 75% of controls', respectively) and
high-dose females (85% of controls'). Food consumption was likewise decreased (p<= 0.05) in
high-dose males during weeks 1-5 (90% of control levels) and in mid- and high-dose males
during weeks 6-9 (92% of control levels for both groups). A thorough histologic examination
revealed no other abnormal findings. Other effects noted were not definitively related to
treatment and/or were not biologically significant. The NOAEL and LOAEL are identified as
200 and 800 mg/kg-day, respectively, based on decreased body weight.
In a second study by Wolfe (1988b), groups of 20 male and 20 female Sprague-Dawley rats were
administered p-xylene (99% purity) by gavage in corn oil at doses of 0, 100, 200, or
800 mg/kg-day for 90 consecutive days. Survival incidences were 20/20, 19/20, 17/20, and
16/20, respectively, for males, and 20/20, 18/20, 18/20, and 17/20 for females. Mortality in
high-dose males attained statistical significance, and a statistically significant trend was present
in the male groups. As in the Wolfe (1988a) study, mottled lungs and/or a failure of the lungs to
collapse was observed in nearly all treated animals that died early but was not evident in any of
the animals that survived to study termination. It was determined that most of the unscheduled
deaths were the result of test material aspiration, as indicated by the presence of intra-alveolar
foreign material in the lungs that was generally associated with pulmonary congestion.
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Treatment-related clinical signs were limited to increased salivation occurring just prior to
dosing that was resolved by 1-hour post-dosing in both high-dose males and females. Body
weight gains at 13 weeks were slightly reduced (89% of control levels, not statistically
significant) in high-dose males and females, and high-dose females had significantly increased
food consumption for weeks 10-13 (110%). No treatment-related effects were observed in
hematology or clinical chemistry parameters, ophthalmologic examination, or organ weights.
Histopathology revealed no abnormal findings in any tissue or organ. The NOAEL and LOAEL
are identified as 200 and 800 mg/kg-day, respectively, based on early mortality in male rats that
showed signs of test material aspiration into the lungs.
The NTP (1986) 2-year study in rats was selected as the principal study for the derivation of the
RfD for xylenes because it is the only oral animal study of chronic duration, and some effects
(decreased body weight and possible increased mortality) were evident at doses lower than those
for effects seen in other studies. The body weight decrease (5-8% of controls') is considered to be
of marginal biological significance, but there was a statistically significant trend for decreased
survival in male rats with increasing exposure levels, and survival in the high-dose males was
statistically significantly decreased when compared with controls. Given the possibility of
treatment-related frank toxicity, it is not considered prudent to discount the only other observed
effect, i.e., decreased body weight. Thus, the highest dose in the study, 500 mg/kg-day, is
considered a LOAEL for changes in body weight and mortality.
	I.A.3. Uncertainty and Modifying Factors (Oral RfD)
UF =1000
A UF of 10 was applied to account for laboratory animal-to-human interspecies differences. No
information is available to support a change from default.
A UF of 10 was applied for intraspecies uncertainty to account for human variability and
sensitive populations. This factor accounts for humans who may be more sensitive than the
general population to exposure to xylenes.
A UF of 10 was used to account for database uncertainty. The available oral database for xylenes
includes chronic and subchronic gavage toxicity studies in mice and rats and a developmental
toxicity study. None of these studies indicate that additional data would result in a lower RfD.
However, the database lacks adequate studies of the oral neurotoxicity of xylenes as well as
multigenerational reproductive toxicity and developmental neurotoxicity studies. Given the
identification of neurological impairment as a critical health hazard from inhalation exposure to
xylenes, the lack of comprehensive neurotoxicity testing following chronic oral exposure is of
particular concern. It should be noted that transient neurotoxic effects (e.g., lethargy, tremors and
unsteadiness) were reported in mice following oral exposure to xylenes for 13 weeks
(NTP, 1986). There are no toxicokinetic data identifying oral dose levels at which first-pass
hepatic metabolism of xylenes becomes saturated in animals or humans; such data could
decrease uncertainty regarding whether neurological impairment may occur at dose levels below
those causing body weight decreases and mortality in rats. It is uncertain whether the availability
of comprehensive oral neurotoxicity data would result in a lower RfD.
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An additional uncertainty associated with the oral database is that the majority of studies
examined mixed xylenes, which are known to contain ethylbenzene. The IRIS assessment for
ethylbenzene (U.S. EPA, 2002a), which was entered on the database in 1987, cites effects on
liver and kidney as the most sensitive endpoints following oral exposure. As discussed below,
effects on the liver and kidney have been reported following oral exposure to mixed xylenes, but
the most sensitive effect reported in animal bioassays is decreased body weight and increased
mortality, as identified by the principal study (NTP, 1986). However, because the mechanism
behind the critical effect has not been clearly elucidated, a possible contribution of ethylbenzene
to the toxicity of mixed xylenes cannot be entirely eliminated. Additional studies comparing the
toxicity of mixed xylenes with that of the individual isomers would better inform the database.
The RfD is based on a NOAEL from a chronic study, which obviates the need for a UF due to
LOAEL to NOAEL extrapolation or subchronic extrapolation.
MF = 1
	I.A.4. Additional Studies/Comments (Oral RfD)
In a NTP (1986) study, groups of 10 male and 10 female Fischer 344 rats were administered
mixed xylenes (60% m-xylene, 13.6% p-xylene, 17.0% ethylbenzene, 9.1% o-xylene) in corn oil
by gavage at doses of 0, 62.5, 125, 250, 500, or 1000 mg/kg-day for 5 days per week for
13 weeks. At termination of the study, necropsy was performed on all animals and
comprehensive histologic examinations were performed on vehicle and high-dose group animals.
High-dose males and females gained 15% and 8% less body weight, respectively, than did
controls, with final body weights being 89% and 97%, respectively, of those of controls
(statistical significance not reported). No signs of toxicity or treatment-related gross or
microscopic pathologic lesions were observed. The LOAEL is 1000 mg/kg-day and the NOAEL
is 500 mg/kg-day based on decreased body weight in male rats without tissue lesions.
In the same study, male and female B6C3Fi mice were treated with mixed xylenes. Groups of
10 mice of each sex were administered 0, 125, 250, 500, 1000, and 2000 mg/kg-day in corn oil
by gavage for 5 days per week for 13 weeks. Two female mice in the high-dose group died
prematurely, although gavage error could not be ruled out as the cause. At 2000 mg/kg-day,
starting 5-10 minutes after dosing and lasting for 15-60 minutes, the animals exhibited lethargy,
short and shallow breathing, unsteadiness, tremors, and paresis. In the high-dose group, mean
body weight was 7% lower for males and 17% lower for females than in the vehicle control.
Although not stated explicitly, the text implies that this was a common finding among the
animals dosed at this level. No treatment-related gross or microscopic pathologic lesions were
seen in this study. The NOAEL is 1000 mg/kg-day and the LOAEL is 2000 mg/kg-day for
transient signs of nervous system depression in mice without tissue lesions.
In a study by Condie et al. (1988) groups of 10 male and 10 female Sprague-Dawley rats were
administered mixed xylenes (17.6% o-xylene, 62.3% m-xylene and p-xylene [which coeluted],
20% ethyl benzene) by gavage in corn oil for 90 consecutive days at doses of 0, 150, 750, or
1500 mg/kg-day. Effects of exposure included decreased body weights in high-dose males
(94%) of controls'), dose-related increased liver weights and liver-to-body weight ratios in all
exposed groups of males (8, 18, and 29% increase in absolute weight above controls' in the low-,
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mid-, and high-dose animals, respectively) and in mid- and high-dose females (14 and 30%,
respectively), and increased kidney weights and kidney-to-body weight ratios in mid- and
high-dose males (16 and 19% increase in absolute weight relative to controls', respectively) and
high-dose females (18% increase in absolute weight relative to controls). The authors postulated
that the modest increases in aspartate aminotransferase seen in high-dose females and increases
in alanine aminotransferase in high-dose males and in mid- and high-dose females, combined
with the lack of significant histopathologic findings in the liver, suggest that the enlargement of
the liver was an adaptation response to xylenes treatment rather than an adverse toxicological
effect.
Hematology analysis revealed a mild polycythemia and leukocytosis in the high-dose males and
females in the absence of any observable changes in the health of the rats. Microscopic
evaluation of the kidneys revealed a dose-related increase in hyaline droplet formation in male
rats (0/9, 3/9, 5/10, 8/10, respectively) and a dose-related increase in the early appearance of
minimal chronic nephropathy in female rats (1/10, 3/10, 6/10, 7/10, respectively). Compared
with controls, the incidence of minimal nephropathy was statistically significantly elevated
(p<0.05) in the 750 and 1500 mg/kg-day female groups but not in the 150 mg/kg-day group
(Fishers exact test performed by Syracuse Research Corporation). The hyaline droplet formation
in male rats was assumed by the authors to be related to male rat-specific a-2|i-globulin
accumulation and not to be relevant to humans. The LOAEL is 750 mg/kg-day, based on
increased kidney weights and early appearance of mild nephropathy in female rats, and the
NOAEL is 150 mg/kg-day.
Kidney effects were not found in the NTP (1986) bioassay with Fisher 344/N rats or
B6C3F1 mice exposed to xylenes for 13 weeks or 2 years. Likewise, no nephropathy was
reported in a nephrotoxicity screening assay in male Fischer 344/N rats exposed to 2000 mg/kg
m-xylene for 5 days per week for 4 weeks (Borriston Laboratories, Inc., 1983). In addition, no
kidney effects were found in Sprague-Dawley rats exposed for 90 days to m-xylene or p-xylene
at doses as high as 800 mg/kg-day (Wolfe et al., 1988a, b). Thus, the available data do not
consistently identify the kidney as a sensitive target of xylenes in animals. Likewise, the
available data do not consistently identify the liver as a sensitive target of xylenes in animals
(NTP, 1986; Wolfe et al., 1988a, b; Condie et al., 1988).
A developmental toxicity study in CD-I mice (Nawrot and Staples, 1980) indicates that
developmental effects may occur following exposure to xylenes. However, the study was
reported as an abstract with incomplete documentation of exposure protocols and results; it does
not identify reliable NOAELs and LOAELs for maternal and developmental toxicity.
Nevertheless, information in the abstract indicates that exposure on gestation days 6-15 to daily
doses of o-, m-, or p-xylene at 1935 or 2580 mg/kg-day-but not at 774 mg/kg-day-resulted in
overt maternal toxicity and increased incidences of cleft palate in the fetuses.
For more detail on Susceptible Populations, exit to the toxicolosical review. Section 4.7
(PDF).
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	I.A.5. Confidence in the Oral RfD
Study — Medium
Database — Medium
RfD — Medium
Confidence in the principal study is medium. The study was a 2-year toxicology and
carcinogenesis assay that evaluated the critical endpoint for RfD derivation (body weight and
mortality) and included comprehensive histologic examination of tissues for nonneoplastic and
neoplastic lesions. Some gavage errors occurred during the study, limiting the confidence
assessment to medium. Confidence in the oral exposure database is medium because the database
contains chronic animal studies in two species (rats and mice), numerous subchronic studies, and
an evaluation of the developmental effects of oral xylenes, but it is lacking oral neurotoxicity
studies as well as multigenerational reproductive toxicity and developmental neurotoxicity
studies. Medium confidence in the RfD follows.
For more detail on Characterization of Hazard and Dose Response, exit to the toxicological
review. Section 6 (PDF).
	I.A.6. EPA Documentation and Review of the Oral RfD
Source Document - U.S. EPA, 2002a
This assessment was peer reviewed by external scientists. Their comments have been evaluated
carefully and incorporated in finalization of this IRIS summary. A record of these comments is
included as an appendix to U.S. EPA (2002a). To review this appendix, exit to the toxicological
review. Appendix A. Summary of and Response to External Peer Review Comments (PDF)
Agency Consensus Date - 01/30/2003
	I.A.7. EPA Contacts (Oral RfD)
Please contact the IRIS Hotline for all questions concerning this assessment or IRIS, in general,
at (202)566-1676 (phone), (202)566-1749 (FAX) or hotline.iris@epa.gov (internet address).
Top of page
I.B. Reference Concentration for Chronic Inhalation Exposure (RfC)
Substance Name — Xylenes
CASRN — 1330-20-7
Last Revised — 02/21/2003
The inhalation Reference Concentration (RfC) is analogous to the oral RfD and is likewise based
on the assumption that thresholds exist for certain toxic effects such as cellular necrosis. The
inhalation RfC considers toxic effects for both the respiratory system (portal-of-entry) and for
effects peripheral to the respiratory system (extrarespiratory effects). It is generally expressed in
units of mg/cu.m. In general, the RfC is an estimate (with uncertainty spanning perhaps an order
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of magnitude) of a daily inhalation exposure of the human population (including sensitive
subgroups) that is likely to be without an appreciable risk of deleterious effects during a lifetime.
Inhalation RfCs are derived according to Methods for Derivation of Inhalation Reference
Concentrations and Application of Inhalation Dosimetry (EPA/600/8-90/066F October 1994).
RfCs can also be derived for the noncarcinogenic health effects of substances that are
carcinogens. Therefore, it is essential to refer to other sources of information concerning the
carcinogenicity of this substance. If the U.S. EPA has evaluated this substance for potential
human carcinogenicity, a summary of that evaluation will be contained in Section II of this file.
As noted in Section I. A. of this file, xylenes refers to mixtures of all three xylene isomers and
ethylbenzene. The inhalation RfC for xylenes presented herein is based on a principal study
(Korsak et al., 1994) in which rats were exposed by inhalation to m-xylene. There is some
uncertainty associated with selecting a principal study for xylenes that involved exposure to
m-xylene alone, but this isomer is generally predominant in commercial mixtures. In addition,
although there are no studies comparing xylene isomers in affecting critical neurological
endpoints following subchronic or chronic inhalation exposure, the potencies of individual
xylene isomers were similar in affecting neurobehavior, as shown in a study of rats following
acute exposures (Moser et al., 1985) (see Section 4.4.3 of the Toxicological Review for more
information).
No inhalation RfC for xylenes has previously been on IRIS.
	I.B.1. Inhalation RfC Summary
Critical Effect	[Experimental Doses*	|UF |MF [RfC
Impaired motor	NOAEL: 50 ppm	300 1 0.1 mg/m3
coordination (decreased	NOAEL(Hec): 39 mg/m3
rotarod performance)
LOAEL: 100 ppm
Subchronic inhalation	LOAEL(Hec): 78 mg/m3
study in male rats
(Korsak et al., 1994)
*Conversion Factors and Assumptions - MW = 106.17. Assuming 25C and 760 mmHg,
NOAEL(mg/m3) = 50 ppm x 106.17/24.45 = 217 mg/m3. NOAEL[adj] = 217 mg/m3 x 6 hrs/day
x 5 days/7 days = 39 mg/m3. The NOAEL*Hec was calculated for extrarespiratory effects of a
Category 3 gas (U.S. EPA, 1994). Blood/gas partition coefficients: H(b/g)rat = 46.0;
H(b/g)human=26.4 (Tardif et al., 1995). (Hb/g)rat/(Hb/g)human =1.7; value of 1 is used when the ratio
is >1 (U.S. EPA, 1994). NOAEL*hec =NOAEL[ADj] x (Hb/g)rat/(Hb/g)human = 39 mg/m3.
	I.B.2. Principal and Supporting Studies (Inhalation RfC)
Korsak et al. (1992) exposed groups of 12 male Wistar rats to toluene, m-xylene, or a
1:1 mixture for 6 hours per day, 5 days per week at a concentration of 0 or 100 ppm for 6 months
or 1000 ppm for 3 months. Rotarod performance and spontaneous motor activity were assayed
24 hours after termination of the exposure periods. The rotarod test was used as a measure of
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motor coordination disturbances from exposure to m-xylene. The rotarod test involves placing
the subject animals on a rotating rod and evaluating their ability to remain on the rod for a period
of 2 minutes. The animals were trained to perform the task, exposed to chemical or control gas,
and evaluated at defined intervals. By the time interval after exposure, considerable proportions
of absorbed xylenes are expected to have been eliminated from the body (see Section 3.4 and
Appendix B of the Toxicological Review).
Body weights and weights of seven organs were measured; only data for animals sacrificed after
3 months of exposure was reported (controls and 1000 ppm rats). At 3 and 6 months, blood
samples were collected 24 hours after termination of exposure for measurement of serum
chemistry variables (e.g., alanine aminotransferase, aspartate aminotransferase, sorbitol
dehydrogenase, alkaline phosphatase, and total protein) and hematologic variables (erythrocyte
counts, hemoglobin concentration, hematocrit, leukocyte count, and differential leukocyte
counts). Serum chemistry and hematologic results were reported only for rats exposed to
1000 ppm for 3 months. Statistical evaluations (using ap=0.05 level of significance) of collected
data included analysis of variance, Dunnet's test, and Fishers exact test.
Rats exposed to m-xylene alone exhibited statistically significantly decreased rotarod
performance and decreased spontaneous activity, as measured 24 hours after termination of the
exposures, when compared with controls. The percentages of failures in the rotarod test were
roughly 60% in rats exposed to 1000 ppm for 3 months, 35% in rats exposed to 100 ppm for
6 months, and 0% for controls at either time period. The mean spontaneous motor activity in rats
exposed to 100 ppm for 6 months was about 400 movements per hour, compared with about
800 movements per hour for controls. Spontaneous motor activity data for rats exposed to
1000 ppm m-xylene for 3 months were not presented in the report. No statistically significant
exposure-related changes in body weight, absolute or relative organ weights, or clinical
chemistry or hematology variables were noted in rats exposed to 1000 ppm m-xylene for
3 months, with the exception of decreased differential counts (percentage of white blood cells
counted) of lymphocytes (45.5 ± 9.5 vs. 60.8 ± 6 .4 for controls; 25% decrease) and increased
counts of monocytes (16.3 ± 8.9 vs. 8.3 ± 4.2 for controls; 96% increase). Total counts of white
blood cells (in units of cells per mm3 of blood), however, were not statistically significantly
changed by exposure. The LOAEL is 100 ppm, based on decreased rotarod performance and
decreased spontaneous motor activity. No NOAEL was identified.
In a second study, Korsak et al. (1994) exposed groups of 12 male Wistar rats by inhalation to 0,
50, or 100 ppm m-xylene or n-butyl alcohol or a 1:1 mixture (purity of chemicals not provided)
for 6 hours per day, 5 days per week, for 3 months and evaluated similar endpoints as in the
earlier study (Korsak et al., 1992). Blood for clinical biochemistry and hematologic analysis was
collected 24 hours after termination of exposure. The report does not specify the timing of the
neurologic examinations; however, given that the 1994 study was conducted by the same group
of investigators as the 1992 study and that one of the tests (rotarod performance) was the same in
both studies, it appears reasonable to assume that the tests were administered 24 hours after
termination of exposure. Statistical evaluations (using a p=0.05 level of significance) of the
collected data included analysis of variance, Dunnet's test, and Fishers exact test.
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No statistically significant exposure-related changes were noted in body weight gain, absolute or
relative organ weights, hepatic activities of microsomal monooxygenases, lipid peroxidation, or
levels of triglycerides in the liver. Statistically significant decreases in erythrocyte number were
seen in animals exposed to 50 ppm (93% of controls') or 100 ppm (80.5% of controls') of
m-xylene alone. Similarly, decreased levels of hemoglobin were reported in both groups (92% of
controls' for both groups). At 100 ppm, a statistically significant increase in leukocyte number
(35%) increase over controls') was reported. Exposure to 50 or 100 ppm m-xylene alone also
resulted in decreased rotarod performance starting at 1 month of exposure, which remained at the
same level until the end of the 3-month exposure. Decreases were statistically significant in the
100 ppm group when compared with the controls. The results were presented in graphical form;
the actual numerical data are not provided. The decreases in performance were roughly 8% and
33%) for the 50 and 100 ppm groups, respectively, versus 0% for the controls.
Sensitivity to pain was assessed using the hot plate behavior test, in which the animals are placed
on a hot (54°C) surface and the time interval between being placed on the plate and licking of the
paws is measured. Rats exposed to 50 or 100 ppm m-xylene alone had statistically significantly
increased sensitivity to pain at the end of the 3-month exposure (latency of the paw-lick response
was 8.7 and 8.6 seconds, respectively, vs. 12.2 seconds for the controls). The LOAEL is
100 ppm, based on decreased rotarod performance and decreased latency in the paw-lick
response in the hot-plate test, and the NOAEL is 50 ppm.
To evaluate whether xylenes influence aging of the central nervous system or induces persistent
changes in radial maze performance, Gralewicz et al. (1995) exposed 8-month-old, male
LOD-Wistar rats (20 per dose level) to air containing 0, 100, or 1000 ppm "pure" m-xylene
(exact purity not provided) for 6 hours per day, 5 days per week, for 3 months. One-hour
electroencephalograph (EEG) recordings were performed on days 28 and 56 of exposure and on
days 14, 28, 56, and 84 after exposure. The number and duration of spontaneous neocortical
spike and wave discharges (SWD) from the EEG were taken as electrophysiological indices of
the biological age of the brain. As rats age, SWDs increase in number and become longer.
Because of large interindividual variation in number and duration of SWDs within each group,
these variables were normalized to a percentage of the initial values. Exposed rats were not
subjected to the daily exposure protocol when EEG recordings were made on days 28 and
56 during the exposure period. Tests of spatial learning in an 8-arm radial maze were also
conducted for a 2-week period starting from day 70 after exposure to day 83.
During the first adaptation stage of the test (five consecutive daily training periods), rats were
familiarized with the maze. The second stage (five consecutive daily trials) measured
effectiveness of finding water in the maze (e.g., duration of trial, number of entries into the arms,
number of omission and preservation errors). One-way or two-way parametric analysis of
variance was applied to the collected data, and effects were regarded as statistically significant at
p<0.05. Body weights were also measured during and after the exposure period at various
intervals, but statistically significant differences were not found among the groups.
The analysis of variance indicated no group effect on the normalized number and
cumulative-duration SWD variables. However, a statistically significant group x successive
recording period effect was indicated. In control rats, these variables were increased to a
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statistically significant degree, compared with those of the exposed groups, only on day 84 after
exposure. The mean cumulative SWD duration (expressed in percentage) on day 84 was about
300 for the control compared with means of about 150 in each of the exposed groups. The
authors hypothesized that these exposure-related changes in the spontaneous, age-related
changes in cortical SWD activity may be related to cortical excitability or to an increase in
catecholaminergic transmissions.
Unlike the controls, rats exposed to 100 or 1000 ppm m-xylene did not exhibit a statistically
significant shortening of the time needed to complete a trial in the radial maze with successive
daily trials. These results indicate a learning deficit in the exposed rats. For example, on the fifth
consecutive trial, the mean trial durations in each of the exposed groups were about
240-250 seconds, compared with a mean of about 150 seconds for the control group. In addition,
the exposed groups did not exhibit the statistically significant decrease in omission errors with
successive days in the radial arm maze test that was exhibited by the control group (number of
arms in the maze omitted during a 5-minute period when the rats explored the maze). The mean
number of omission errors in control rats showed a progressive decrease from about 2.75 on the
first trial to 0 on the fourth and fifth successive trials. In contrast, the means on the fifth
consecutive trial were about 1.5 and 2.5 for the 100 ppm and 1000 ppm groups, respectively. The
lowest exposure level in this study, 100 ppm, is designated as a LOAEL for deficits in radial
maze performance.
Gralewicz and Wiaderna (2001) exposed groups of male Wistar rats (10-11 animals/group) to
0 or 100 ppm of m-xylene for 6 hours per day, 5 days per week for 4 weeks. Behavioral testing
was performed at various intervals before (radial maze and open-field evaluations) and after
exposure (radial maze [days 14-18], open-field activity [day 25], passive avoidance [days 39-48],
hot plate test [days 50-51], and active avoidance [days 54-60]). The radial maze and hot plate test
protocols are described in previous studies from this group (Gralewicz and Wiaderna, 1995;
Korsak et al., 1992).
In the open-field activity test, animals were placed in a 100 cm x 100 cm arena that was
surrounded by 20 cm high walls and divided into 49 equal squares. The number of square
borders crossed (locomotor activity), number of rearings (exploratory activity), and number of
grooming episodes were recorded. In the passive avoidance test, animals were placed on a
platform above the floor of the cage, and the time until the animal stepped off the platform was
recorded in a series of six trials. In the first two trials, the animals were allowed to explore the
cage for 60 seconds after stepping down; in the third trial, the animals received a series of
footshocks after stepping off the platform. In trials 4, 5, and 6 the animals received no shocks
and were allowed to stay on the floor for 1 minute after stepping off the platform. In the active
avoidance test, animals were trained to avoid an electric footshock by moving from one
compartment of the cage to another when a sound is played. After successfully displaying
avoidance behavior in four of five trials, the animals were considered to be trained.
Post-exposure evaluations determined the frequency of avoidance behavior in response to the
same stimulus.
No differences between control and exposed rats were seen in radial maze parameters (number of
arm entries, arms omitted, or arms entered multiple times) either before exposure (7 days prior to
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exposure) or at 14-18 days after the termination of exposure. Similarly, no differences in open-
field activity were seen between groups examined on day 8 prior to exposure or day 25
postexposure or in active avoidance (number of trials to avoidance criterion), examined on days
54 and 60 post-exposure. Xylene-exposed rats showed a significantly shorter step-down time
(trial 6 only; no difference in trials 1-5) in the passive avoidance test (examined on days
39-48 postexposure) and a significantly greater paw-lick latency in the hot plate behavior test
(examined on days 50-51 postexposure), identifying 100 ppm as a LOAEL for neurobehavioral
effects.
Because available human data are insufficient for deriving an RfC and chronic animal inhalation
data are lacking, the subchronic study of Korsak et al. (1994) was selected as the principal study
and Korsak et al. (1992), Gralewicz et al. (1995), and Gralewicz and Wiaderna (2001) as the
supporting studies. Neurological effects (impaired motor coordination) are selected as the critical
effect for deriving the RfC. Two neurological endpoints were evaluated in this study. Rotarod
performance was statistically significantly decreased (33% from controls') at 100 ppm, and a
statistically significant decreased sensitivity to pain was observed at 50 and 100 ppm (8.6 and
8.7 seconds, respectively, vs. 12.2 seconds for controls; measurements made 24 hours
postexposure). Gralewicz and Wiaderna (2001) also measured the effect of m-xylene exposure
(6 hrs/day, 5 days/wk for 4 weeks; neurological endpoints measured postexposure day 50) on
pain sensitivity. In this study, a statistically significant increase in pain sensitivity (35 seconds
vs. 10 seconds in control) was found at the 100 ppm dose, the lowest dose tested. The variation
in the response to m-xylene in these two studies decreases the confidence in using the pain
sensitivity endpoint as the critical effect.
A number of statistically significant neurological effects have been noted in male rats at a dose
of 100 ppm m-xylene in other supporting studies: decreased rotarod performance and
spontaneous movement activity following exposure for 6 hours per day, 5 days per week for
6 months (Korsak et al., 1992), decreased radial maze performance following exposure for
6 hours per day, 5 days per week for 3 months (Gralewicz et al., 1995); and shortened step-down
time in the passive avoidance test following exposure for 6 hours per day, 5 days per week for
4 weeks. All studies measured neurological endpoints 24 hours postexposure with the exception
of Gralewicz and Wiaderna (2001), which measured effects at postexposure day 50. For these
reasons, a NOAEL of 50 ppm and a LOAEL of 100 ppm is identified for neurological effects
(impaired motor coordination).
The principal study (Korsak et al., 1994) reported no statistically significant exposure-related
changes in body weight gain, absolute or relative organ weights, hepatic activities of
monoxygenases or lipid peroxidation, or levels of triglycerides in the liver. Compared with
controls, exposed rats showed statistically significant changes in red blood cell counts
(7-20% decreased), hemoglobin levels (-8%) decreased), and white blood cell counts (35%
increased). Effects in red blood cell counts and hemoglobin levels were observed at 50 ppm.
However, these changes were not observed in another study from the same laboratory
(Korsak et al.,1992) in rats exposed to 1000 ppm m-xylene. Furthermore, effects on erythrocytes
were not found at concentrations of 78-810 ppm in other studies (Carpenter et al., 1975;
Jenkins et al., 1970).
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	I.B.3. Uncertainty and Modifying Factors (Inhalation RfC)
UF = 300
A UF of 3 was applied to account for laboratory animal-to-human interspecies differences. A
factor of 3 was applied because default NOAELrec dosimetric adjustments were used to
calculate a human equivalent concentration (HEC), reducing the uncertainty involved with the
extrapolation from the results of an animal study to a human exposure scenario (i.e., the
toxicokinetic portion of the UF is 1; the toxicodynamic portion of the UF is 3).
A uncertainty factor of 10 was applied for intraspecies uncertainty to account for human
variability and sensitive populations. The degree of human variance in abilities to absorb or
dispose of xylenes is unknown, as is the degree of human variance in responding to xylenes
neurotoxicity. Results from developmental toxicity studies of rats exposed by inhalation during
gestation indicate that adverse developmental effects occur only at higher doses than chronic
doses producing the critical effects observed in adult male rats in the principal and supporting
studies, suggesting that the developing fetus is not at special risk from low-level exposure to
xylenes. However, as with oral exposure, the effects of inhaled xylenes in other potentially
sensitive populations such as newborns or young children or animals have not been assessed.
A UF of 3 was applied for extrapolation from subchronic to chronic duration. A factor of 10 was
not used because the changes in rotarod performance did not increase with time from 1 to 3
months and were similar to those described in a separate study of 6-months duration (Korsak et
al., 1992).
A UF of 3 was applied for uncertainties in the database. The inhalation database includes some
human studies, subchronic studies in rats and dogs, neurotoxicity studies, a one-generation
reproductive toxicity study, developmental toxicity studies, and developmental neurotoxicity
studies. Although the available developmental toxicity studies are confounded by a lack of litter
incidence reporting, the data reported for fetal incidences do not indicate effects at levels lower
than that found to induce neurologic impairment in several endpoints in male rats. The database
is lacking a two-generation reproductive toxicity study.
MF = 1
	I.B.4. Additional Studies/Comments (Inhalation RfC)
The weight of evidence from limited human data and more extensive animal data identify mild
neurological impairment and possible developmental effects as potential health hazards from
repeated inhalation exposure to xylenes. The animal inhalation exposure database contains no
chronic toxicity studies, but there are a number of subchronic toxicity studies (of which several
focused on neurological endpoints), a one-generation reproduction study in rats, and several
developmental toxicity studies, some of which evaluated offspring for performance in
neurobehavioral tests. Subchronic toxicity assays in animals have not found consistent evidence
for other noncancer effects, such as changes in body weight or in hepatic, hematologic, or renal
toxicity endpoints, following exposure to concentrations of xylenes as high as 800-1000 ppm for
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6 hours per day, 5 days per week (e.g., Carpenter et al., 1975; Jenkins et al., 1970;
Korsaketal., 1992, 1994).
Reversible symptoms of neurological impairment and irritation of the eyes and throat are well-
known health hazards from acute inhalation exposure to xylenes and other aromatic solvents. In
general, these acute effects are expected to involve reversible molecular interactions of the
solvent itself (not metabolites) with membranes of the affected tissues, including neuronal
membranes, and are most pronounced at high exposure levels in excess of 1000 ppm. At lower
concentrations, more subtle effects may occur. Human volunteers exposed under controlled
conditions to xylenes concentrations in the range of 200-400 ppm for short time periods
(15 minutes to 4 hours) have reported symptoms of irritation (e.g., watering eyes and sore throat)
or neurological impairment (e.g., mild nausea, headache) (Carpenter et al., 1975;
Gamberale et al., 1978).
In other studies involving single or multiple 4-hour exposures of human volunteers to 200 ppm
xylenes, reversible effects on balance and reaction times have been reported (Laine et al., 1993;
Savolainen and Linnavuo, 1979; Savolainen et al., 1984); however, other studies of 4-hour
exposures to 200 ppm have not found impaired performance in tests of simple reaction time,
short-term memory, and choice reaction time (Olson et al., 1985) or changes in visually evoked
brain potentials (Seppalainen et al., 1983) or electroencephalographic patterns (Seppalainen et
al., 1991). Impaired performance on tests of memory and reaction times was also reported for
subjects exposed to 100 ppm xylenes for 4 hours (Dudek et al., 1990). The available controlled-
exposure human studies indicate that concentrations around 100-200 ppm are close to the
threshold level for short-term reversible neurological and irritation effects from xylenes.
The available human data alone do not provide adequate evidence for neurological impairment
from repeated exposure to xylenes concentrations less than or equal to 200 ppm. Aside from the
controlled-exposure studies reviewed above, most of the human data associating xylenes
exposure to neurological impairment are case reports involving acute high-level exposures
(800-10,000 ppm) (e.g., Goldie, 1960; Hipolito, 1980; Klaucke et al., 1982). Epidemiologic
studies are restricted to a cross-sectional health evaluation study (Uchida et al., 1993) that
reported increased prevalence of self-reported neurological symptoms and irritation, but no
apparent changes in serum enzymes indicative of liver or kidney damage in a group of Chinese
workers. The workers were from a boot manufacturing plant that used a xylene-containing glue
and two other plants that used mixed xylenes as a solvent in wire production or printing. The
measured time-weighted-average mean concentration of airborne xylenes in these workplaces
was 21 (± 21) ppm. The study has several limitations, including a lack of reporting on the
duration of exposure, co-exposure to other chemicals, no clear demonstration of relationships
between response and dose or duration, and the inherent bias presented by self-reporting of
symptoms.
Although the human evidence for persistent effects on the nervous system or other persistent
effects from repeated inhalation exposure to xylenes is inadequate, results from animal studies
more clearly identify potential persistent neurological impairment and possible developmental
effects as potential health hazards from repeated inhalation exposure.
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Overall results from rat studies described in Section I.B.2 provide evidence that repeated
exposure to m-xylene at concentrations >= 100 ppm (6 hrs/day, 5 days/wk) may produce
persistent changes in several neurologic endpoints in adult rats. Supporting evidence for potential
persistent neurologic effects from xylenes includes reports of changes in indices of hearing loss
in rats exposed to >= 800 ppm mixed xylenes for 14 hours per day for 6 weeks (Pryor et al.,
1987) and in rats exposed to 1000 ppm mixed xylenes for 18 hours per day, 7 days per week, for
61 days (Nylen and Hagman, 1994).
There are no studies of the possible developmental toxicity of inhaled xylenes in humans, but
there are a number of studies examining standard developmental toxicity endpoints and
neurobehavioral endpoints in offspring of animals exposed to mixed xylenes or individual xylene
isomers. Evidence for impaired neurological development in rat offspring following gestational
exposure to inhaled xylenes is not strong or consistent. Changes in neurobehavioral variables
reported for offspring of animals exposed during gestation are restricted to impaired cognitive
but not motor performance in the Morris water maze test in female but not male offspring of rats
exposed to 500 ppm mixed xylenes for 6 hours per day on gestation days 7-20 (Hass et al., 1995,
1997) and decreased rotarod performance in offspring of rats exposed to 200 ppm "technical"
xylenes for 6 hours per day on gestation days 6-20 (Hass and Jakobsen, 1993). Deficits in the
water maze test were only observed in female rat offspring raised in standard housing and not in
female rats raised in "enriched" housing with various toys (Hass et al., 1995).
Although decreased rotarod performance by offspring was observed in the study by Hass and
Jakobsen (1993), it was not observed in the later study by the same group of investigators
(Hass et al., 1995). The reported effect on rotarod performance in the earlier study was
questioned by Hass et al. (1995) because the test was not conducted by experimenters who were
blind to the exposure status of the rats. In addition, offspring of rats exposed to 800 or 1600 ppm
p-xylene for 6 hours per day on gestation days 7-16 performed similarly to offspring of
nonexposed rats in tests of central nervous system development: an acoustic startle response test
on postnatal days 13, 17, 21, and 63 and a figure-8 maze activity test on postnatal days 22 and
65 (Rosen et al., 1986).
Several other inhalation developmental toxicity studies have examined standard developmental
toxicity endpoints in rats (Litton Bionetics, 1978; Bio/dynamics Inc., 1983; Rosen et al., 1986;
Ungvary et al., 1980; Ungvary and Tatrai, 1985), mice (Ungvary and Tatrai, 1985) and rabbits
(Ungvary and Tatrai, 1985) following gestational exposure to xylenes. These studies have most
clearly identified maternally toxic levels for decreased body weight gain in pregnant rats at
concentrations greater than or equal to 700 ppm o-, p-, or m-xylene for 24 hours per day
(Ungvary et al., 1980) or 1600 ppm p-xylene for 6 hours per day (Rosen et al., 1986) and for
maternal death and abortions in pregnant rabbits exposed to 230 ppm (but not 115 ppm) mixed
xylenes or p-xylene for 24 hours per day (Ungvary and Tatrai, 1985). In rats, effects on fetal
skeletal and visceral malformations (such as cleft palate) and variations (such as retarded skeletal
ossification or extra ribs) were reported at concentrations of up to 700 ppm o-, m-, or p-xylene
for 24 hours per day (Ungvary et al., 1980) or 780 ppm mixed xylenes for 24 hours per day
(Bio/dynamics Inc., 1983; Litton Bionetics, 1978; Ungvary and Tatrai, 1985). Likewise, effects
on skeletal and visceral malformations and variations were reported in mice at concentrations of
up to 230 ppm mixed xylenes (12 hrs/day in three 4-hr periods) or 115 ppm o-, p-, or m-xylene
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by the same protocol (Ungvary and Tatrai, 1985) or in rabbits exposed to 115 ppm mixed
xylenes or o-, p-, or m-xylene for 24 hours per day (Ungvary and Tatrai, 1985).
Statistically significant increased incidences of fetuses with retarded skeletal ossification or extra
ribs were reported in these studies, but the incidences were reported on an exposure-group basis
in all but one of the studies. No litter-specific information was provided except in the
Litton Bionetics (1978) study, which reported that, after adjustment for covariance with litter
size, incidences of fetuses with delayed ossification in rats exposed to 400 ppm were no longer
statistically significantly different from control values.
The most significant effects on developmental endpoints were decreased fetal body weight or
fetal survival in rats at xylene isomer concentrations of 350 or 700 ppm for 24 hours per day
(Ungvary et al., 1980) or a mixed xylenes concentration of 780 ppm for 24 hours per day
(Ungvary and Tatrai, 1985) and increased abortions in rabbits exposed to 230 ppm for 24 hours
per day (Ungvary and Tatrai, 1985). These effects, although of concern, occurred at
concentrations above those at which neurobehavioral effects were found in adult male rats
following subchronic exposure (see Section I.B.2.).
Information regarding the potential reproductive toxicity of xylenes in humans is restricted to
case-control studies reporting possible associations between occupational exposure to xylenes
and other solvents and spontaneous abortions (e.g., Taskinen et al., 1986, 1994). However, these
studies are of limited usefulness in assessing the potential reproductive toxicity of xylenes,
because the numbers of cases of spontaneous abortions were small, and the women had been
exposed to a number of chemicals.
Two reproductive toxicity studies in rats exposed to xylenes by inhalation are available
(Bio/dynamics Inc., 1983; Nylen and Hagman, 1994). In a one-generation
reproductive/developmental toxicity study (Bio/dynamics Inc., 1983), male and female CD rats
were exposed to 0, 60, 250, or 500 ppm xylenes (technical grade xylene: 2.4% toluene,
12.8% ethylbenzene, 20.3% p-xylene, 44.2% m-xylene, 20.4% o-xylene) by inhalation for
6 hours per day, 5 days per week, for 131 days prior to mating, with exposure continued in the
females during gestation days 1-20 and lactation days 5-20. Two additional 500-ppm groups
used the same exposure protocol, except that only the F0 males were exposed in one, and only
the F0 females were exposed in the other. The highest exposure level in this study, 500 ppm, was
a NOAEL for reproductive performance in the parental generation. Likewise, a study of male
Sprague-Dawley rats exposed to 0 or 1000 ppm xylene solvent for 18 hours per day, 7 days per
week, for 61 days reported no differences between control and exposed rats in several testicular
endpoints and fertility (Nylen and Hagman, 1994).
In summary, human data are suggestive of neurological effects and irritation of the eyes and
respiratory tract following inhalation exposure to xylenes. Animal studies have demonstrated that
neurological effects are the most sensitive effect of xylenes inhalation, with measurable effects in
several neurobehavioral endpoints beginning at concentrations as low as 100 ppm following
subchronic exposure (Gralewicz et al., 1995; Korsak et al., 1992, 1994; Nylen and Hagman,
1994; Pry or et al., 1987). At higher exposure levels, changes in body weight have been reported
by some studies (Tatrai and Ungvary, 1980; Tatrai et al., 1981) but not by others
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(Carpenter et al., 1975; Jenkins et al., 1970; Ungvary, 1990). Similarly, high-level exposure to
xylenes has resulted in changes in liver morphology, weight, and enzymatic functions
(Tatrai and Ungvary, 1980; Tatrai et al., 1981; Ungvary, 1990). Gestational exposure of animals
to xylenes has resulted in neurodevelopmental effects (Hass et al., 1995, 1997;
Hass and Jakobsen, 1993) and other possible developmental effects (Ungvary et al., 1980;
Ungvary and Tatrai, 1985), but only at levels above those at which neurobehavioral effects in
adult male rats were reported. Finally, no reproductive effects were found in a one-generation
reproductive/developmental study of male and female rats exposed to 500 ppm xylenes
(Bio/dynamics, Inc., 1983) or in male rats exposed to 1000 ppm xylenes for 61 days
(Nylen and Hagman, 1994).
For more detail on Susceptible Populations, exit to the toxicolosical review. Section 4.7
(PDF).
	I.B.5. Confidence in the Inhalation RfC
Study — Medium
Database — Medium
RfC — Medium
Confidence in the principal study is medium, because the study was an examination of a critical
effect of xylenes toxicity that also examined organ weights, body weights, and hematological
parameters but was of subchronic duration, examined only one sex of a single species, and did
not conduct histologic examination of the animals. Confidence in the database is medium; the
database contains several subchronic studies as well as several developmental studies,
developmental neurotoxocity studies, and a one-generation reproductive toxicity study.
However, a two-generation reproduction study and chronic animal data are lacking. Medium
confidence in the RfC results.
For more detail on Characterization of Hazard and Dose Response, exit to the toxicolosical
review. Section 6 (PDF)
	I.B.6. EPA Documentation and Review of the Inhalation RfC
Source Document - U.S. EPA, 2002
This assessment was peer reviewed by external scientists. Their comments have been evaluated
carefully and incorporated in finalization of this IRIS summary. A record of these comments is
included as an appendix to U.S. EPA, 2002. To review this appendix. exit to the toxicolosical
review. Appendix A, Summary of and Response to External Peer Review Comments (PDF)
Agency Consensus Date - 01/30/2003
	I.B.7. EPA Contacts (Inhalation RfC)
Please contact the IRIS Hotline for all questions concerning this assessment or IRIS, in general,
at (202)566-1676 (phone), (202)566-1749 (fax), or hotline.iris@epa.gov (Internet address).
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Top of page
VI. Bibliography
Substance Name — Xylenes
CASRN — 1330-20-7
Last Revised — 02/21/2003
VI.A. Oral RfD References
Borriston Laboratories, Inc. (1983) Four-week oral nephrotoxicity screening study in male F-344
rats phases I and II pathology report. FYI submission AX-1283-0280. Submitted by the
American Petroleum Institute to U.S. Environmental Protection Agency, Washington, DC.
Condie, LW; Hill, JR; Borzelleca, JF. (1988) Oral toxicology studies with xylene isomers and
mixed xylene. Drug Chem Toxicol 11:329-354.
Nawrot, PS; Staples, RE. (1980) Embryotoxicity and teratogenicity of isomers of xylene in the
mouse. Soc Toxicol Abst. PAP 19th: A22, 65.
NTP (National Toxicology Program). (1986) NTP technical report on the toxicology and
carcinogenesis of xylenes (mixed) (60% m-xylene, 13.6% p-xylene, 17.0% ethylbenzene, and
9.1%) o-xylene) in F344/N rats and B6C3F1 mice (gavage studies). Research Triangle Park, NC.
NTP TR 327, NIH Publ. No. 86-2583.
U.S. EPA (Environmental Protection Agency). (2002a) Integrated Risk Information System
(IRIS). Online. Office of Research and Development. National Center for Environmental
Assessment, Washington, DC. Examined September, 2001. Online, http://www.epa.gov/iris
U.S. EPA. (2002b) Toxicological review of xylenes (CAS No. 1330-20-7). National Center for
Environmental Assessment, Washington, DC. Available online at: http://www.epa.gov/iris.
Wolfe, GW. (1988a) Subchronic toxicity study in rats with m-xylene. Report by Hazleton
Laboratories America, Inc. Sponsored by Dynamac Corporation, Rockville, MD. Project No.
2399-108.
Wolfe, GW. (1988b) Subchronic toxicity study in rats with p-xylene. Report by Hazleton
Laboratories America, Inc. Sponsored by Dynamac Corporation, Rockville, MD. Project No.
2399-110.
VLB. Inhalation RfD References
Bio/dynamics Inc. (1983) Parental and fetal reproduction toxicity study in rats with mixed
xylene. EPA/OTS public files. Bio/dynamics Inc., East Millstone, NJ;
Document # FYI-AX-0983-0209.
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Carpenter, CP; Kinkead, ER; Geary, DL Jr; et al. (1975) Petroleum hydrocarbon toxicity studies.
V. Animal and human response to vapors of mixed xylene. Toxicol Appl Pharmacol 33:543-58.
Dudek, B; Gralewicz, K; Jakubowski, M; et al. (1990) Neurobehavioral effects of experimental
exposure to toluene, xylene and their mixture. Polish J Occup Med 3:109-116.
Gamberale, F; Annwall, G; Hultengren, M. (1978) Exposure to xylene and ethylbenzene. III.
Effects on central nervous functions. Scand J Work Environ Health 4:204-211.
Goldie, I. (1960) Can xylene (xylol) provoke convulsive seizures? Ind Med Surg 29:33-35.
Gralewicz, S; Wiaderna, D. (2001) Behavioral effects following subacute inhalation exposure to
m-xylene or trimethylbenzene in the rat. A comparative study. NeuroToxicology 22: 79-89.
Gralewicz, S; Wiaderna, D; Tomas, T. (1995) Development of spontaneous, age-related
nonconvulsive seizure electrocortical activity and radial-maze learning after exposure to
m-xylene in rats. Int J Occup Med Environ Health 8:347-360.
Hass, U; Jakobsen, BM. (1993) Prenatal toxicity of xylene inhalation in the rat: a teratogenicity
and postnatal study. Pharmacol Toxicol 73:20-23.
Hass, U; Lund, SP; Simonsen, L; et al. (1995) Effects of prenatal exposure to xylene on postnatal
development and behavior in rats. Neurotoxicol Teratol 17:341-349.
Hass, U; Lund, SP; Simonsen, L. (1997) Long-lasting neurobehavioral effects of prenatal
exposure to xylene in rats. Neurotoxicology 18:547-551.
Hipolito, RN. (1980) Xylene poisoning in laboratory workers: case reports and discussion. Lab
Med 11:593-595.
Jenkins, L J Jr.; Jones, R A; Siegel, J. (1970) Long-term inhalation screening studies of benzene,
toluene, o-xylene, and cumene on experimental animals. Toxicol Appl Pharmacol 16:818-823.
Klaucke, DN; Johansen, M; Vogt, R. (1982) An outbreak of xylene intoxication in a hospital.
Am J Ind Med 3:173-178.
Korsak, Z; Sokal, JA; Gorny, R. (1992) Toxic effects of combined exposure to toluene and m-
xylene in animals. III. Subchronic inhalation study. Polish J Occup Med Environ Health 5:27-33.
Korsak, Z; Wisniewska-Knypl, J; Swiercz, R. (1994) Toxic effects of subchronic combined
exposure to n-butyl alcohol and m-xylene in rats. Int J Occup Med Environ Health 7:155-166.
Laine, A; Savolainen, K; Riihimaki, V; et al. (1993) Acute effects of m-xylene inhalation on
body sway, reaction times, and sleep in man. Int Arch Occup Environ Health 65:179-188.
Litton Bionetics. (1978) Teratology study in rats - xylene. Final report EPA/OTS Public Files.
Litton Bionetics, Kensington, MD; Document 878210350.
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Moser, V C; Coggeshall, EM; Balster, RL. (1985) Effects of xylene isomers on operant
responding and motor performance in mice. Toxicol Appl Pharmacol 80:293-298.
Nylen, P; Hagman, M. (1994) Function of the auditory and visual systems, and of peripheral
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derivatives. II. Xylene. Pharmacol Toxicol 74:124-129.
Olson, BA; Gamberale, F; Inegren, A. (1985) Coexposure to toluene and p-xylene in man:
central nervous functions. Br J Ind Med 42:117-122.
Pryor, GT; Rebert, CS; Howd, RA. (1987) Hearing loss in rats caused by inhalation of mixed
xylene and styrene. J Appl Toxicol 7:55-61.
Rosen, MB; Crofton, KM; Chernoff, N. (1986) Postnatal evaluation of prenatal exposure to
p-xylene in the rat. Toxicol Let 34:223-229.
Savolainen, K; Linnavuo, M. (1979) Effects of m-xylene on human equilibrium measured with a
quantitative method. Acta Pharmacol Toxicol 44:315-318.
Savolainen, K; Kekoni, J; Riihimaki, V; et al. (1984) Immediate effects of m-xylene on the
human central nervous system. Arch Toxicol Suppl 7:412-417.
Seppalainen, AM; Salmi, T; Savolainen, K; et al. (1983) Visual evoked potentials in short-term
exposure of human subjects to m-xylene andl,l,l-trichloroethane. Appl Behav Pharmacol
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modeling of a mixture of toluene and xylene in humans. Risk Anal 15:335-342.
Tatrai, E; Ungvary, G. (1980) Changes induced by o-xylene inhalation in the rat liver. Acta Med
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Ungvary, G. (1990) The effect of xylene exposure on the liver. Acta Morphol Hung 38:245-258.
Ungvary, G; Tatrai, E. (1985) On the embryotoxic effects of benzene and its alkyl derivatives in
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meta- and para-xylene. Toxicology 18:61-74.
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U.S. EPA (Environmental Protection Agency). (1994) Methods for derivation of inhalation
reference concentrations and application of inhalation dosimetry. EPA/600/8-90/066F.
October 1994.
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Environmental Assessment, Washington, DC. Available online at: http://www.epa.gov/iris.
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APPENDIX B: DETAILS OF BENCHMARK DOSE MODELING
FOR SUBCHRONIC ORAL p-RfD
Description of Model-Fitting Procedure for Continuous Data
The model-fitting procedure for continuous data is as follows. When a biologically
defined BMR is not available, the default BMR of 1 standard deviation from the control mean
response is used (U.S. EPA, 2000). The simplest model (linear) is first applied to the data while
assuming constant variance. If the data are consistent with the assumption of constant variance
(p> 0.1), then the fit of the linear model to the means is evaluated. If the linear model
adequately fits the means (p> 0.1), then it is selected as the model for BMD derivation. If the
linear model does not adequately fit the means, then the more complex models are fit to the data
while assuming constant variance. Among the models providing adequate fit to the means
(p> 0.1), the one with the lowest AIC for the fitted model is selected for BMD derivation. If the
test for constant variance is negative, the linear model is run again while applying the power
model integrated into the BMDS to account for nonhomogenous variance. If the
nonhomogenous variance model provides an adequate fit (p> 0.1) to the variance data, then the
fit of the linear model to the means is evaluated. If the linear model does not provide adequate
fit to the means while the nonhomogenous variance model is applied, then the polynomial,
power, and Hill models are fit to the data and evaluated while the variance model is applied.
Among those providing adequate fit to the means (p> 0.1), the one with the lowest AIC for the
fitted model is selected for BMD derivation. If the test for constant variance is negative and the
nonhomogenous variance model does not provide an adequate fit to the variance data, then the
dataset is considered unsuitable for modeling.
Modeling of Data on Terminal Body Weight in Male Rats (NTP, 1986)
Following the above procedure, continuous-variable models in the U.S. EPA BMDS
(version 1.4. lc) were fit to the data shown in Table B-l (below) for decreased body weight in
male rats exposed for 13 weeks (NTP, 1986) using a biologically based BMR of 10% decrease
from the control mean. The constant variance model did not provide adequate fit to the variance
data. Further, the variance model included in the BMDS did not provide an adequate fit to the
variance, as shown in Table B-2. Because body weight was significantly different from control
only in the high-dose group, no attempt was made to exclude this group and model the
lower-dose groups. Thus, this dataset was not suitable for BMD analysis.

Table B-l. Final Body Weights of Male Rats


Exposed Orally to Mixed Xylenes for 13 Weeks3

Dose
Adjusted Doseb
Number of
Mean Final
Standard
(mg/kg-day)
(mg/kg-day)
Rats
Body Weight (g)
Deviation
0
0
10
328
15.8
62.5
44.6
10
323
12.6
125
89.3
10
327
25.3
250
179
10
315
28.5
500
357
10
330
28.5
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1000
714
10
291
22.1
aNTP, 1986
bAdjusted for continuous exposure (adjusted dose = dose x 5/7 days/week)
Table B-2. Model Predictions for Decreased Body
Weight in Male Rats Exposed Orally to Xylenes for 13 Weeks3
Model
Variance
/7-value b
Means
/7-value b
BMD0.1
(mg/kg-day)
BMDL01
(mg/kg-
day)
All dose groups
Linear (constant
variance)c
0.08666
0.11
745.97
517.78
Linear (modeled
variance)c
0.04729
0.208
849.17
532.93
aNTP, 1986
bValues <0.10 fail to meet conventional goodness-of-fit criteria (U.S. EPA, 2000)
Coefficients restricted to be negative
Modeling of Data on Terminal Body Weight in male rats (Wolfe, 1988a)
Following the above procedure, continuous-variable models in the U.S. EPA BMDS
(version 1.4. lc) were fit to the data shown in Table B-3 (below) for decreased body weight in
male rats exposed for 13 weeks (Wolfe, 1988a) using a biologically-based BMR of
10% decrease from the control mean. Using these data, the linear model with constant variance
model provided adequate fit to both the variance and means data, as shown in Table B-4.
Figure B-l shows the fit of the linear model with constant variance to the data.
Table B-3. Final Body Weights of Male Rats Exposed
Orally to Mixed Xylenes for 13 Weeks"
Dose
(mg/kg-day)
Number of Rats
Mean Final Body
Weight (g)
Standard Deviation
0
20
527.8
46.28
100
17
518.1
37.55
200
15
492.1
31.26
800
18
448.1
30.37
aWolfe, 1988a
Table B-4. Model Predictions for Decreased Body
Weight in Male Rats Exposed Orally to Xylenes for 13 Weeks3
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Model
Variance
p-valueb
Means
p-v alueb
BMD0.1
(mg/kg-day)
BMDL01
(mg/kg-
day)
All dose groups
Linear (constant
variance)0
0.2173
0.3367
538.39
440.22
"Wolfe, 1988a
bValues <0.10 fail to meet conventional goodness-of-fit criteria (U.S. EPA, 2000)
Coefficients restricted to be negative
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Linear Model with 0.95 Confidence Level
560
540
520
o 500
480
460
440
Linear
Dose
13:55 05/06 2008
BMDs and BMDLs indicated are associated with a 10% decrease from control body weight and are in units of
mg/kg-day.
Figure B-l. Fit of Linear Model (Constant Variance) to Data on Final Body Weight in
Male Rats (Wolfe, 1988a)
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APPENDIX C: DESCRIPTION OF LITERATURE
SEARCH PROCESS FOR XYLENE
The IRIS Toxicological Review (U.S. EPA, 2003) contained a thorough review of oral
toxicity data on xylene, so searches were limited to studies published since 2002. The search for
additional studies of xylene included terms to identify human exposure studies (epidemiologic,
occupational) and animal studies for all relevant noncancer endpoints. The search included
health effects and toxicity information available from the U.S. EPA (IRIS), ATSDR and other
relevant federal, state or international governmental or quasi-governmental agencies, including,
but not limited to ACGM, NIOSH, OSHA, NTP, IARC, WHO, and CalEPA. In addition,
electronic databases, including: CURRENT CONTENTS, MEDLINE, TOXLINE,
BIOSIS/TOXCENTER, TSCATS/TSCATS2, CCRIS, DART/ETIC, GENETOX, HSDB, and
RTECS, were searched. An electronic listing of all results of the gross literature review
(including titles, references and abstracts) and a tabular summary of the search results were
provided to EPA.
A toxicologist screened the literature searches based on review of abstracts and titles for
studies pertaining to the health effects from subchronic oral exposure to xylenes in humans and
animals. Decisions about whether to further consider a particular citation were based on the
scientific judgment of the toxicologist, based on reading the abstract provided in the literature
search output. Studies that were not considered pertinent were not retrieved. Citations may also
have been excluded after retrieval and review of the article by the toxicologist. A study may
have been excluded if its scope was outside the scope of the use under consideration, if it was not
relevant or appropriate, if its study design was inadequate, or if the study showed inadequacy of
quality control or flaws in the interpretation of results.
No new pertinent studies were identified.
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