United States
Environmental Protection
1=1 m m Agency
EPA/690/R-10/009F
Final
9-15-2010
Provisional Peer-Reviewed Toxicity Values for
Cyclohexanone
(CASRN 108-94-1)
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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AUTHORS, CONTRIBUTORS, AND REVIEWERS
CHEMICAL MANAGER:
Dan Petersen, National Center for Environmental Assessment, Cincinnati, OH
DRAFT DOCUMENT PREPARED BY:
SRC, Inc.
7502 Round Pond Road
North Syracuse, NY 13212
INTERNAL REVIEW PANEL:
National Center for Environmental Assessment, Cincinnati, OH
Dan Petersen
Jay Zhao
Jon Reid
National Center for Environmental Assessment, Research Triangle Park, NC
Anu Mudipalli
Geniece Lehmann
Nicole Hagan
Paul Reinhart
National Center for Environmental Assessment, Washington, D.C.
Audrey Galizia
Martin Gehlhaus
Sanjivani Diwan
Susan Makris
This document was externally peer-reviewed under contract to:
Eastern Research Group, Inc.
110 Hartwell Avenue
Lexington, MA 02421-3136
Questions regarding the contents of this document 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)
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TABLE OF CONTENTS
COMMONLY USED ABBREVIATIONS	iii
BACKGROUND	1
HISTORY	1
DISCLAIMERS	1
QUESTIONS REGARDING PPRTVS	2
INTRODUCTION	2
REVIEW 01 PERTINENT DATA	3
HUMAN STUDIES	3
ANIMAL STUDIES	5
Oral Exposure	5
Subchronic Studies	5
Chronic Studies	6
Reproductive/developmental Studies	9
Inhalation Exposure	10
Subchronic Studies	10
Reproductive/developmental Studies	11
OTHER STUDIES	17
Genotoxicity	17
DERIVATION OF PROVISIONAL SUBCHRONIC AND CHRONIC	19
ORAL RID VALUES FOR CYCLOHEXANONE	19
SUBCHRONIC p-RfD	19
CHRONIC p-RfD	22
DERIVATION OF PROVISIONAL SUBCHRONIC AND CHRONIC	22
INHALATION RfC VALUES FOR CYCLOHEXANONE	22
SUBCHRONIC p-RfC	25
CHRONIC p-RfC	26
PROVISIONAL CARCINOGENICITY ASSESSMENT	27
FOR CYCLOHEXANONE	27
WEIGHT-OF -E VIDEN CE DESCRIPTOR	27
QUANTITATIVE ESTIMATES OF CARCINOGENIC RISK	27
REFERENCES	28
ii

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COMMONLY USED ABBREVIATIONS
BMC
benchmark concentration
BMD
benchmark dose
BMCL
benchmark concentration lower bound 95% confidence interval
BMDL
benchmark dose lower bound 95% confidence interval
HEC
human equivalent concentration
HED
human equivalent dose
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
NOAELrec
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 reference concentration (inhalation)
p-RfD
provisional reference dose (oral)
POD
point of departure
RfC
reference concentration (inhalation)
RfD
reference dose (oral)
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
WOE
weight of evidence
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PROVISIONAL PEER-REVIEWED TOXICITY VALUES FOR
CYCLOHEXANONE (CASRN 108-94-1)
BACKGROUND
HISTORY
On December 5, 2003, the U.S. Environmental Protection Agency'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)	EPA's Integrated Risk Information System (IRIS)
2)	Provisional Peer-Reviewed Toxicity Values (PPRTVs) used in EPA's Superfund
Program
3)	Other (peer-reviewed) toxicity values, including
~	Minimal Risk Levels produced by the Agency for Toxic Substances and Disease
Registry (ATSDR);
~	California Environmental Protection Agency (CalEPA) values; and
~	EPA Health Effects Assessment Summary Table (HEAST) values.
A PPRTV is defined as a toxicity value derived for use in the Superfund Program when
such a value is not available in EPA's IRIS. PPRTVs are developed according to a Standard
Operating Procedure (SOP) and are derived after a review of the relevant scientific literature
using the same methods, sources of data, and Agency guidance for value derivation generally
used by the EPA IRIS Program. All provisional toxicity values receive internal review by a
panel of six 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 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 EPA Office of
Research and Development's National Center for Environmental Assessment, Superfund Health
Risk Technical Support Center for OSRTI. Other EPA programs or external parties who may
choose of their own initiative to use these PPRTVs are advised that Superfund resources will not
generally be used to respond to challenges of PPRTVs used in a context outside of the Superfund
Program.
QUESTIONS REGARDING PPRTVS
Questions regarding the contents of the PPRTVs and their appropriate use (e.g., on
chemicals not covered, or whether chemicals have pending IRIS toxicity values) may be directed
to the EPA Office of Research and Development's National Center for Environmental
Assessment, Superfund Health Risk Technical Support Center (513-569-7300), or OSRTI.
There is an RfD assessment but no RfC or carcinogenicity assessments for
cyclohexanone (chemical structure shown in Figure 1) on IRIS (U.S. EPA, 2009). The RfD of
5 mg/kg-day was derived based on a NOAEL for body-weight depression in rats exposed to
cyclohexanone in the drinking water for 2 years (Lijinsky and Kovatch, 1986). There are no
entries for cyclohexanone in the HEAST (U.S. EPA, 1997), Drinking Water Standards and
Health Advisories list (U.S. EPA, 2006), or Chemical Assessments and Related Activities
(CARA) database (U.S. EPA, 1994a, 1991).
Occupational health guidelines and standards are available for cyclohexanone. The
American Conference of Governmental Industrial Hygienists (ACGIH, 2007, 2003) recommends
a Threshold Limit Value-time-weighted average (TLV-TWA) of 20 ppm (80 mg/m3) and
TLV-Short-Term Exposure Limit (STEL) of 50 ppm (200 mg/m3), mainly to minimize the
potential for eye, nasal, and throat irritation. The TLV recommendations are accompanied by a
skin irritancy notation and an A4 carcinogenicity notation (Not Classifiable As a Human
Carcinogen). The National Institute for Occupational Safety and Health (NIOSH, 2005) lists a
INTRODUCTION
O
Figure 1. Chemical Structure of Cyclohexanone.
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Recommended Exposure Limit (REL) of 25 ppm (100 mg/m3) TWA with a skin notation, as well
as an Immediately Dangerous to Life or Health (IDLH) concentration of 700 ppm (2814 mg/m3).
The Occupational Safety and Health Administration (OSHA, 2009) has promulgated a
Permissible Exposure Limit of 50 ppm (200 mg/m3) TWA.
There is no ATSDR (2009) Toxicological Profile or World Health Organization (WHO,
2009) Environmental Health Criteria Document for cyclohexanone. CalEPA (2009a,b,c) has not
derived chronic oral or inhalation RELs or a cancer potency factor for cyclohexanone. The
National Toxicology Program (NTP) tested the oral carcinogenicity of cyclohexanone in rats and
mice with negative results (Lijinsky and Kovatch, 1986), and has not included the chemical in its
annual Report on Carcinogens (NTP, 2005), which lists known and likely human carcinogens.
The International Agency for Research on Cancer (IARC, 1999, 1989) classified the
carcinogenicity of cyclohexanone in Group 3 (Not Classifiable As to Human Carcinogenicity)
based on lack of human cancer data and inadequate evidence of carcinogenicity in animals.
Literature searches were conducted for studies relevant to the derivation of provisional
toxicity values for cyclohexanone. Databases searched include MEDLINE, TOXLINE (BIOSIS
and NTIS), TOXCENTER, CCRIS, DART/ETIC, DTIC, TSCATS/TSCATS 2, GENETOX,
HSDB, RTECS, and Current Contents. The time period covered by most of the searches ranged
from the 1960s through early January 2009, although some searches covered earlier years.
REVIEW OF PERTINENT DATA
HUMAN STUDIES
There are no available human data from oral exposure to cyclohexanone. Limited data
are available regarding inhalation exposure of humans to cyclohexanone. Nelson et al. (1943)
exposed volunteers to cyclohexanone at 25, 50, or 75 ppm (100, 201, or 301 mg/m3) in an
inhalation chamber for 3-5 minutes. No signs of discomfort were reported at 25 ppm
(100 mg/m3), but at 50 ppm (201 mg/m3), subjects reported throat irritation, and at 75 ppm
(301 mg/m3), subjects exhibited pronounced irritation of the eyes, nose, and throat. In another
study, volunteers exposed to cyclohexanone for 7 minutes reported marked eye irritation and
slight skin irritation at 160 ppm (642 mg/m3) (Esso Research and Engineering Co., 1965).
IARC (1989) reviewed a Russian study by Bereznyak (1984) that found no effect on
nervous system function, blood, or respiration in a group of 100 production workers exposed to
cyclohexanone by both inhalation (3.7 mg/m3) and skin contact (10"4 mg/m2 on the hands),
relative to 49 controls. The primary study was not available for this review, and the IARC
(1989) review does not provide any further detail on this study other than to mention that there
was some indication of liver disorders among a subgroup of workers, 30-39 years old, with more
than 5 years of exposure to cyclohexanone.
A more recent study found that clinical symptoms (i.e., ocular, upper respiratory tract,
and cutaneous irritation; mood disorders; irritability; memory difficulties; sleep disturbances;
headache; numbness; muscular pains; abdominal pains; and irregular bowel movements) were
more frequently reported in a group of 75 furniture factory workers with known exposure to a
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wood coating product containing cyclohexanone than in a group of 85 matched controls without
known chemical exposures (Mitran et al., 1997). It is unclear what other chemicals these
workers may have been exposed to in the wood coating product. Cyclohexanone exposure levels
over an 8-hour shift were reported to range from 162 to 368 mg/m3, and the duration of exposure
averaged 14 years. The basis of these air measurements is unclear, as Mitran et al. (1997) do not
provide any further information regarding this cohort, methodologies used to collect air
measurements in the factory, or conditions at the factory, which may have changed over time due
to the implementation of engineering controls or changes in processes. There were no significant
differences between the exposed and control groups in urinary excretion of cyclohexanone
sulfate metabolites or in serum chemistry tests. Motor nerve conduction velocity testing revealed
statistically significant (p < 0.05) differences between exposed and control groups—most notably
increased distal latency in peripheral nerves (see Table 1). Cyclohexanone-exposed workers also
showed significantly delayed reaction times to visual and auditory stimuli (data not shown).
Mitran et al. (1997) noted that although these results demonstrated peripheral nerve disturbances
following cyclohexanone exposure, the results should not be overinterpreted, given the
difficulties interpreting peripheral electrodiagnostic studies.
A possible neurological effect of cyclohexanone is also suggested by a worker with
known occupational exposure to a mixture of organic solvents (cyclohexanone, white spirit, and
isopropanol) who experienced a long history of temporal epileptic seizures (Jacobsen et al.,
1994). This worker also often complained of headache, nausea, and vertigo. The seizures
disappeared shortly after exposure ceased, but reappeared following short-term re-exposure to
high levels of cyclohexanone. Subsequent to this last seizure, the subject had no further
exposure to organic solvents and no further epileptic seizures.
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Table 1. Motor Nerve Conduction Velocity Among Furniture Factory Workers Exposed
to Cyclohexanone at 162-368 mg/m3 for an Average of 14 Years"
Parameter
Median Nerve
Ulnar Nerve
Peroneal Nerve
Proximal
Latency (msec)
Exposed
Controls
7.69 ±0.95
6.91 ± 1.20
9.30 ± 2.05b
7.01 ± 1.99
12.92 ± 1.59
11.64 ±2.69
Amplitude (mV)
Exposed
Controls
6.01 ± 1.20
7.15 ±2.24
6.75 ± 1.97
8.01 ± 1.01
3.55 ± 1.33b
5.99 ±2.01
Duration (msec)
Exposed
Controls
6.20 ± 1.44
5.70 ± 1.03
7.15 ±2.40
6.28 ± 1.03
6.08 ±2.56
5.10 ± 1.08
Distal
Latency (msec)
Exposed
Controls
4.12 ± 0.90b
2.90 ± 1.01
4.33 ± 1.0lb
3.01 ±0.93
5.96 ± 0.88°
2.11 ± 1.06
Amplitude (mV)
Exposed
Controls
7.01 ±2.61
9.05 ±2.33
7.99 ±0.90
8.95 ± 1.10
2.40 ± 2.03°
6.55 ±2.99
Duration (msec)
Exposed
Controls
5.90 ±0.80
5.05 ±0.99
6.02 ± 1.44
5.42 ± 1.75
6.02 ±3.02
5.86 ±2.77
Nerve conduction velocity (msec)
Exposed
Controls
52.01 ±8.03
55.12 ±7.91
46.10 ±0.93b
55.60 ±0.99
44.03 ±2.33
46.70 ± 1.45
aValues presented as mean ± SD.
Significantly different from controls by Student's /-test (p < 0.05).
°Significantly different from controls by Student's /-test (p < 0.01).
Source: Mitran et al. (1997).
ANIMAL STUDIES
Oral Exposure
Subchronic Studies—In a study by the National Cancer Institute (NCI), groups of five
male and five female F344 rats were treated with cyclohexanone (96% purity) in acidified (with
HC1 to pH 2.5) drinking water at 190, 400, 800, 1600, 3300, 4700, or 6500 ppm for 25 weeks
(Lijinsky and Kovatch, 1986; NCI, 1979). Water was acidified to suppress bacterial growth. An
additional group of 5 rats (number/sex not specified) served as the untreated control group.
Based on EPA (1988) reference values for body weight and water consumption for F344 rats in a
subchronic study, using water intake factors of 0.156 and 0.169 L/day for males and females
respectively, the estimated daily doses are 0, 30, 62, 124, 249, 513, 731, and 1010 mg/kg-day for
males, and 0, 32, 68, 135, 271, 559, 796, and 1100 mg/kg-day for females. Evaluations were
limited to survival, body weight, gross pathology, and histopathology (organs and tissues not
specified). No mortality occurred during the study. All rats, including controls, displayed signs
of moderate chronic respiratory disease, for which the study authors do not provide a reason or
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rationale. High-dose rats exhibited a 10% decrease in weight gain compared to controls (data not
provided). The only other effect reported was a mild degenerative change in the thyroid gland of
two male rats given 4700-ppm cyclohexanone. This pathological change was not observed in
female rats or high-dose male rats, indicating that it is not likely related to treatment with
cyclohexanone. No other treatment-related effects were reported. Based on decreased weight
gain in high-dose rats, the NOAEL values are identified as 731 and 796 mg/kg-day (4700 ppm)
for males and females, respectively, and the LOAEL values are identified as 1010 and
1100 mg/kg-day (6500 ppm), respectively.
In a companion mouse study, groups of 10 male and 10 female B6C3Fi mice were
administered cyclohexanone (96% purity) also in acidified drinking water at 0, 400, 2300, 6500,
13,000, 25,000, 34,000, or 47,000 ppm for 13 weeks (Lijinsky and Kovatch, 1986; NCI, 1979).
Mice were observed for the same endpoints as described above for the rat subchronic study:
survival, body weight, gross pathology, and histopathology. Based on EPA (1988) reference
values for body weight and water consumption for B6C3Fi mice in a subchronic study, the
estimated daily doses are 0, 99, 568, 1600, 3210, 6170, 8390, and 11,600 mg/kg-day for males
and 0, 106, 608, 1720, 3430, 6610, 8980, and 12,400 mg/kg-day for females. At the highest
dose, 3/10 females and 6/10 males died. One male died at the next highest dose (i.e.,
8390 mg/kg-day). Depression in weight gain was observed among females at 8980 mg/kg-day
(15%>), and among males at 6170 mg/kg-day (19%>) and 8390 mg/kg-day (24%). However,
specific body-weight data were not provided, and body-weight changes among high-dose
animals were not described. At the high-dose, the author reported that some mice showed
coagulative liver necrosis, and two female mice showed hyperplasia of the thymus (pathology
data not provided). NOAELs of 3210 mg/kg-day (13,000 ppm) and 6610 mg/kg-day
(25,000 ppm) and LOAELs of 6170 mg/kg-day (25,000 ppm) and 8980 mg/kg-day (34,000 ppm)
are identified for males and females, respectively, based on depression in body-weight gain.
Chronic Studies—Lijinsky and Kovatch (1986) also conducted 2-year drinking water
studies in rats and mice that are the principal studies used in the derivation of the EPA IRIS RfD
value (U.S. EPA, 2009). In the rat study, groups of 52 male and 52 female F344 rats were
treated with cyclohexanone (96% purity) in acidified drinking water at 0, 3300, or 6500 ppm for
2 years. Based on EPA (1988) reference values for body weight and water consumption of F344
rats in a chronic study, using water intake factors of 0.129 and 0.144 L/day for males and
females respectively, the estimated doses are 0, 426, and 838 mg/kg-day for males and 0, 476,
and 937 mg/kg-day for females. Evaluations included survival, body weight, gross pathology,
and histopathology. Survival among high-dose rats of both sexes was reported as >85% at
90 weeks and 70% at study termination (data plotted as probability of survival over time).
Survival among low-dose rats and controls of both sexes was >90% at 90 weeks and >70% at
termination. Lijinsky and Kovatch (1986) reported that high-dose rats exhibited significant
decreases in weight gain compared to controls. Based on the weight curves reported by the
authors, high-dose rats of both sexes experienced an estimated body-weight deficit of >30% (in
comparison to controls) at study termination. No change in weight gain was noted in the lower
dose group. No treatment-related nonneoplastic lesions were observed among either treatment
group. Based on the decreases in body-weight gain in male and female rats at the highest dose,
the EPA IRIS evaluation of this study identified the NOAEL as 462 mg/kg-day (3300 ppm) and
the LOAEL as 910 mg/kg-day (6500 ppm) (U.S. EPA, 2009).
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In the companion chronic mouse study, groups of B6C3Fi mice (group size was 41 and
47 for high-dose females and males, respectively, and 50 or 52 for all other groups) were treated
with cyclohexanone (96% purity) in acidified drinking water for 2 years at 0, 6500, or
13,000 ppm for males and at 0, 6500, 13,000, or 25,000 ppm for females (Lijinsky and Kovatch,
1986). Based on EPA (1988) reference values for body weight and water consumption for
B6C3Fi mice in a chronic study, the estimated daily doses are 0, 1530, and 3070 mg/kg-day for
males and 0, 1570, 3130, and 6020 mg/kg-day for females. The same endpoints that were
evaluated in the chronic rat study described above were evaluated in mice. Survival among
high-dose males was reported as 80% at 90 weeks and 70% at study termination. Among
females, however, survival was less than 20% in the high-dose group and only 40% in the
mid-dose group at 90 weeks. Survival among low-dose mice of both sexes was comparable to
controls, with approximately 90% survival at 90 weeks and >85%) survival at termination (data
plotted as probability of survival over time). Body weights of high-dose mice of both sexes were
decreased by approximately 15—20% compared to controls during most of the study. Body
weights were only slightly depressed among mid-dose female mice and were comparable to
controls among low-dose mice of both sexes. Lymphoid hyperplasia and lymphocytic infiltrates
were common in lymph nodes, spleen, salivary gland, kidneys, pancreas, lungs, and meninges of
the brain and spinal cord of most control and treated female mice in this study. Lijinsky and
Kovatch (1986) remarked that these changes often involved more than a single organ system,
and no cause was found histologically. Chronic effect levels for mice were not described in the
EPA evaluation of this study on IRIS (U.S. EPA, 2009). For this review, the lymphatic lesions
in control and treated females were considered a potentially confounding observation, and effect
levels for females were not defined. For male mice, the low dose of 1530 mg/kg-day
(6500 ppm) is identified as aNOAEL, and 3070 mg/kg-day (13,000 ppm) is identified as the
LOAEL based on a biologically significant decrease in body-weight gain.
The chronic studies in rats and mice conducted by Lijinsky and Kovatch (1986)
summarized above also evaluated the carcinogenic potential of cyclohexanone. Any lesions or
tissue masses among control and treated animals at the end of the 2-year study were examined
histologically. Table 2 contains the tumor incidence for rats and mice in this study. In rats, there
was an increased incidence of adenomas of the adrenal cortex among low-dose males compared
to concurrent and historical controls. However, there was no increased incidence in
adrenocortical adenomas among high-dose males. Survival of high-dose males in this study was
adequate to evaluate carcinogenicity, so this finding suggests that the tumors in the low-dose
group were not related to treatment. The only other finding in rats was a marginal increase in the
incidence of follicular cell adenoma-carcinomas of the thyroid gland among high-dose males
compared to concurrent controls. In mice, the incidence of combined benign and malignant
hepatocellular neoplasms was significantly higher among low-dose males compared to
concurrent controls, and the incidence of malignant lymphoma was significantly higher among
low-dose females compared to controls. However, no significant increases in tumor incidence
were observed among high-dose males or among mid- and high-dose females. Survival in
high-dose males was adequate to evaluate carcinogenicity, but poor survival in the mid- and
high-dose female mice may have compromised the study in female mice. Lijinsky and Kovatch
(1986) noted that although the results in low-dose animals are suggestive of a response to
cyclohexanone, the absence of a dose-related trend indicates that the evidence for carcinogenic
activity is marginal, and the effect, if any, is weak.
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Table 2. Neoplasms in Rats and Mice Given Cyclohexanone in Drinking Water for 2 Years
Neoplasm
Dose (mg/kg-d)
Male
Female
Rat
0
426
838
0
476
937
Adrenal cortex: adenoma
1/52
7/52a'b
1/51
8/52
4/51
4/52
Thyroid gland: follicular cell adenoma-carcinoma
1/52
0/51
6/51°
0/52
1/52
1/52
Mammary gland: fibroadenoma
2/52
1/52
0/52
13/52
10/52
4/52a
Uterus: endometrial stromal polyp
-
-
-
5/52
6/52
1/51
Liver:
Carcinoma
Carcinomas plus neoplastic nodules
2/52
6/52
0/52
5/52
0/51
4/51
0/52
3/52
0/52
4/52
0/52
5/52
Neoplasm
Dose (mg/kg-d)
Male
Female
Mouse
0
1530
3070
0
1570
3130
6020
Lung: alveolar-bronchiolar adenoma or carcinoma
13/52
7/51
3/47
3/52
2/50
2/50
1/41
Lymphoma or leukemia
6/52
2/52
4/47
8/52
17/50b
4/50
0/41
Liver: adenoma or carcinoma
16/52
25/5 r
13/46
3/52
6/50
3/50
2/41
Harderian gland: adenoma
0/52
4/52
0/47
0/52
1/50
1/50
0/41
Significantly different from control by incidental and/or life table tests (p < 0.05).
bHistorical control incidence of adrenocortical adenomas in male F344 rats = 1%.
°Marginally different from control (p = 0.053).
Source: Lijinsky and Kovatch (1986).
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Reproductive/developmental Studies—In a developmental study from the French
literature, pregnant mice (strains TB and MNRI, unknown group size) were fed a diet containing
1% cyclohexanone throughout pregnancy and lactation (Gondry, 1973). Based on EPA (1988)
reference values for body weight and food consumption of B6C3Fi mice (default strain used in
absence of values for TB and MNRI strains), using a food intake factor of 0.195 kg food/kg
BW/day, the estimated dose is 2000 mg/kg-day. A second group of pregnant mice fed a diet
containing no cyclohexanone served as a control group. The researchers evaluated the
developmental effects of treatment after the first generation and following continuous treatment
across multiple generations. Offspring were evaluated only for mortality and growth. No
evaluations of clinical signs, skeletal abnormalities, or other developmental endpoints were
conducted. Increased neonatal mortality was observed during the first 21 days of life among
offspring from treated dams compared to controls. A depression in growth was observed among
first-generation offspring from treated dams. This effect was more pronounced in female
offspring than male offspring. Following the discontinuation of treatment after the first
generation, growth was comparable to controls by the second generation. However, when
cyclohexanone was administered without interruption to multiple successive generations, the
inhibiting effect on growth was maintained. No numerical or statistical information is presented
for these findings. Based on these findings, a LOAEL of 2000 mg/kg-day is identified based on
increased neonatal mortality and depressed offspring growth.
Chernoff and Kavlock (1983) developed a screening system for identifying teratogens
and tested this system for a variety of chemicals. In this study, 24 pregnant CD-I mice were
administered cyclohexanone (purity not given) via gavage in corn oil at 800 mg/kg-day on
Gestation Days (GDs) 8-12. A second group receiving only corn oil served as a control.
Evaluations included maternal body weight, survival and clinical signs, as well as litter size,
offspring survival, birth weights, and body weights on Day 3. Evaluations of visceral and
skeletal abnormalities and other prenatal developmental endpoints were not conducted. Two of
24 treated mice died during the study, and one control mouse died. The data showed no
significant effects on maternal body-weight gain, litter size, offspring survival, or pup body
weights. Based on these findings, a NOAEL of 800 mg/kg-day is identified for both maternal
reproductive and developmental toxicity.
Gray and Kavlock (1984) followed the animals evaluated by Chernoff and Kavlock
(1983, described above) for 250 days to assess postnatal effects of cyclohexanone treatment.
Endpoints included body weights at 22 days of age (males and females) and at 57 days of age
(males only), behavioral testing for locomotor activity, reproductive function, gross
developmental abnormalities, organ weights (males only; liver, testes, seminal vesicles, and right
kidney), and gross pathology. There were no significant effects on viability, growth,
morphology, locomotor activity, reproductive function, organ weights, or gross pathology in
offspring (Gray and Kavlock, 1984; Gray et al., 1986). Based on these findings, a NOAEL of
800 mg/kg-day is identified.
Another teratogenic screening test conducted by Seidenberg et al. (1986) administered
cyclohexanone (purity not reported) to a group of 28 time-pregnant ICR/SIM mice via gavage in
corn oil at 2200 mg/kg-day on GDs 8-12. A second group receiving only corn oil served as the
control. Evaluations included maternal body weight, survival, and clinical signs, as well as litter
size and offspring survival, birth weights, and body weights on Day 3. Evaluations of visceral
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and skeletal abnormalities and other prenatal developmental endpoints were not conducted. Six
of the 28 treated dams died, and mean maternal body weight was significantly reduced compared
to controls. There was no effect on litter size or pup survival, but mean neonatal body weights
were significantly depressed both at birth and at Day 3 compared to controls. Based on these
findings, a LOAEL of 2200 mg/kg-day is identified for both maternal and developmental
toxicity.
Inhalation Exposure
Subchronic Studies—Groups of four rabbits (sex not specified) were exposed by
inhalation to 0 (untreated), 0 (sham-exposed), 190, 309, 773, 1414, or 3082 ppm (converted by
EPA to 0, 763, 1241, 3103, 5677, or 12,373 mg/m3 using the adjustment calculation of
mg/m3 = ppm x [molecular weight -^24.45]; the molecular weight of cyclohexanone =
98.15 g/mol) of cyclohexanone (purity not reported) vapor, 6 hours/days, 5 days/week, for
3 weeks (for the highest dose level only, 12,373 mg/m3) or 10 weeks (all other groups)
(Treon et al., 1943). In addition to the rabbits, one Rhesus monkey was exposed to 608 ppm
(2441 mg/m3) by the same schedule for 10 weeks. Evaluations during exposure and up to
2 months after exposure termination included clinical signs, survival, body weight, and
hematology (e.g. erythrocyte and leukocyte counts and hemoglobin concentration). Gross
pathology and histopathology (tissues not specified) were conducted following the 2-month
postexposure observation period. At the highest concentration, 2/4 rabbits died, and clinical
signs such as narcosis, labored breathing, loss of coordination, weight loss, and hypothermia
were observed after 3 weeks of exposure. At concentrations >773 ppm (3103 mg/m3), rabbits
exhibited salivation, conjunctival congestion and irritation, lacrimation, and lethargy. Rabbits
exposed to 309-ppm (1241-mg/m3) cyclohexanone only exhibited very slight conjunctival
congestion, and no clinical signs or effects on body weight were observed among rabbits
exposed at the lowest concentration. Incidence of clinical signs was not reported. The monkey
exhibited slight salivation and slight conjunctival congestion. No significant hematological
changes were observed at any concentration of the 10-week exposure protocol. Two months
after the end of exposure, pathology revealed "barely demonstrable" degenerative changes in
liver and kidneys (not further described, incidence not reported) of rabbits exposed to 190 ppm
(763 mg/m3). Histological observations in rabbits exposed at higher concentrations or in controls
were not described. Extensive injury to the heart, lungs, liver, and kidneys was found in the
treated monkey; however, these effects were confounded by a concurrent, chronic
broncho-pulmonary infection in this animal. Treon et al. (1943) concluded that the maximum
safe concentration of cyclohexanone was "slightly below" 190 ppm (763 mg/m3) based on the
liver and kidney lesions. However, due to the "barely demonstrable" nature of the undescribed
lesions, the 2-month separation between examination and the end of exposure, and the lack of
evidence of progression of these changes with exposure concentration (no discussion of
histopathology at higher exposures was provided), the reported liver and kidney lesions were not
used to identify effect levels for this review. Based on these findings, a NOAEL of 309 ppm
(1241 mg/m3) and a LOAEL of 773 ppm (3103 mg/m3) are identified based on clinical signs of
toxicity (i.e., salivation, conjunctival congestion and irritation, lacrimation, and lethargy)
following 10 weeks of inhalation exposure to cyclohexanone.
A series of subchronic studies examined the effects of cyclohexanone exposure on
olfactory bulb development in young rats. These studies showed that rat pups exposed to
cyclohexanone at low concentrations (ranging from 1-4 ppm [or 4-16 mg/m3]) over about
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1-4 months exhibit alterations of mitral cells (Panhuber and Laing, 1987; Laing et al., 1985;
Laing and Panhuber, 1980, 1978; Pinching and Doving, 1974) and spine density of granule cell
dendrites (Rehn et al., 1988) of the olfactory bulb. The alterations in mitral cells were
morphologically similar to transneuronal degeneration and appeared to occur in a
chemical-specific pattern. The affected cells are characterized by a darkening of the nucleus and
cytoplasm and are smaller than in normal rats. Panhuber et al. (1987) evaluated whether
prolonged exposure to cyclohexanone in adult rats would produce similar results to those seen in
rat pups. Significant shrinkage of mitral cells of the olfactory bulb was observed in adult rats
exposed to 8-ppm (32-mg/m3) cyclohexanone for 10 weeks. However, significant shrinkage of
mitral cells was also observed in adult rats exposed to deodorized air for 10 weeks (although the
severity of shrinkage was less than that seen in cyclohexanone-exposed rats). Experiments
performed by these researchers suggest that these effects are ultimately reversible and do not
alter learning rates or olfactory acuity for cyclohexanone. These studies did observe lowered
sensitivity to other similar but novel odorants following prolonged exposure to cyclohexanone.
These researchers acknowledge that the interpretation and functional significance of altered
mitral cells remain unclear. In addition, although epidemiology data on other chemicals (NRC,
1979) have shown evidence of olfactory sensitivity following prolonged exposure to an odorant,
there are no additional data to suggest that altered mitral cells result in decreased olfactory
sensitivity in humans.
Reproductive/developmental Studies—In a multigeneration study, groups of 30 male
and 30 female CD Sprague-Dawley rats were exposed by inhalation to 0, 250, 500, or
1000 [F0J/1400 [Fl] ppm (0, 1004, 2007, or 4015/5621 mg/m3) cyclohexanone (purity not
reported) vapor, 6 hours/day through two consecutive generations (ABC, 1986a). Evaluations
included clinical signs, growth, urinalysis (i.e., volume, glucose, pH, protein, ketone, bilirubin,
occult blood, and urobilinogen), mating and fertility indices, progeny survival and body weight,
pre- and postweaning neurologic performance and neuropathology, and histopathology
(reproductive organs, liver, kidneys, brain, and eyes). Additional information on the protocol
used, such as age of mating and number of pregnant dams is not reported in the ABC (1986a)
study. Lacrimation, ataxia, and irregular breathing were noted in high-dose F0 animals
following the first two exposures, but these clinical signs dissipated, and the animals appeared
normal thereafter. No significant effects were observed on growth or reproductive performance
in F0 animals. Six of 60 high-dose Fl animals died, including three during the first week of
increased exposure (from 1000 ppm [4015 mg/m3] in the parental animals to 1400 ppm
[5621 mg/m3] in the Fl animals). Only one other postweaning death occurred in the entire study,
suggesting that the Fl deaths at 1400 ppm (5621 mg/m3) were probably exposure related. Both
Fl males and females exposed to 1400 ppm (5621 mg/m3) and males exposed to 500 ppm
(2007 mg/m3) exhibited significant decreases in body weights compared to controls during the
first week of increased exposure. In addition, males demonstrated significant differences in body
weights during 31 of the 34 weeks of 1400-ppm (5621-mg/m3) exposure. Females only
demonstrated additional significant decreases in body weights through the first 3 weeks of
exposure to 1400 ppm (5621 mg/m3). Terminal body weights were comparable to controls
among both sexes at all exposure concentrations. Fl animals from the 1400-ppm (5621-mg/m3)
group also exhibited purportedly adverse clinical signs characterized by urine-soaked fur,
lacrimation, irregular breathing, ataxia, and lethargy. No significant depressions were observed
based on reproductive indices for any treatment group compared with controls—although male
fertility indices at 1400 ppm (5621 mg/m3) using all males paired were approximately 20% less
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than controls, and when using only males paired with fertile females, were 24 to 29% less than
controls. Table 3 shows that progeny weights among the Fla litter were significantly different
from controls during the lactation period. However, the dose-response pattern is not clearly
defined across all time points during the lactation period, and effects seen at the high-dose appear
to resolve by Lactation Day 28.
Table 3 summarizes significant findings observed in the F2a and F2b litters. The total
numbers of viable progeny born to F1 animals were not significantly different among exposure
groups compared to controls, but during the lactation period, there were significant decreases in
the mean numbers of viable progeny from 1400-ppm (5621-mg/m3) F1 animals compared to
controls in both the F2a and F2b generations (ABC, 1986a). Progeny survival was significantly
decreased at 1400 ppm (5621 mg/m3) during the first 4 days of the lactation period in both the
F2a and F2b generations. Mean litter weights were significantly reduced for most of the
lactation period at 1400 ppm for both F2a and F2b litters. In the F2a litters, pup body weights
were also significantly reduced at 250 and/or 500 ppm (1004 and/or 2007 mg/m3) during the
latter half of lactation. However, this was not seen with the F2b litters. The researchers did not
consider maternal exposure to 250- or 500-ppm (1004- and/or 2007-mg/m3) cyclohexanone to
adversely affect pup body weights because statistical weight differences noted for the 250- and
500-ppm (1004- and/or 2007-mg/m3) progeny were 'minimal' (5-17%) compared to controls,
and similar effects were not seen for the F2B progeny. Furthermore, effects on fetal weight in
developmental toxicity studies (described below) were observed only at 1400 ppm (5621 mg/m3)
and not at lower concentrations. For the purposes of this review, a LOAEL of 1400 ppm
(5621 mg/m3) and NOAEL of 500 ppm (2007 mg/m3) are identified for effects in F1 animals
including mortality, clinical signs, decreased body weights, and effects on reproduction (reduced
viability and body weight of F2 pups).
A subsequent study conducted during the postexposure recovery period evaluated the
reversibility of reproductive effects in the F1 male CD Sprague-Dawley rats treated with
1400-ppm (5621-mg/m3) cyclohexanone (ABC, 1986b). Males were rested (unexposed)
following the last exposure for 2 days prior to the start of mating trials that occurred for
4 consecutive weeks (Weeks 1-4), Week 6, and Week 8. ABC (1986b) determined that the
decrease in second-generation male fertility in the 1400-ppm group was reversible (ABC,
1986b).
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Table 3. Summary of Progeny Observations from F1 Generation Rats Exposed to Cyclohexanone Vapor
Exposure Concentration
(ppm)
Lactation Day
0 (birth)
1
4
7
14
21
28
F1A Litter
Mean progeny body weights (g)
Male
Female
0
5.4 ±0.8
-
7.8 ± 1.5
10.8 ±2.7
19 ±4.5
31.6 ±7.3
61.6 ± 13.1
55.7 ± 11.3
250
5.4 ±0.7
-
7.9 ± 1.3
10.3 ±2.1
17.8 ± 3.la
28.2 ± 5.4b
55.0 ± 9.8b
52.7 ±9.8
500
5.4 ±0.8
-
7.6 ± 1.6
9.7 ± 2.5b
16.0 ± 4.7b
26.9 ± 7.0b
50.9 ± 14.0b'°
48.8 ± 13.9b
1000
5.5 ± 1.0
-
8.3 ± 1.6b
10.8 ±2.3
17.6 ± 4.0a
28.9 ± 5.9b
56.5 ± 11.7
53.8 ± 10.2
F2A Litter
Mean number of viable progeny from F1 generation rats
0
11.8 ±2.83
11.7 ±2.83
11.5 ±2.74
7.6 ±0.99
7.2 ± 1.96
7.2 ± 1.95
7.2 ± 1.95
250
12.0 ±3.48
11.8 ±3.47
11.6 ±3.47
7.4 ± 1.27
6.9 ±2.05
6.6 ±2.52
6.6 ±2.52
500
11.8 ±2.89
11.2 ± 3.35
10.9 ±3.69
7.3 ± 1.81
7.3 ± 1.81
7.3 ± 1.80
7.3 ± 1.80
1400
9.1 ±4.76
7.1 ± 5.83b
6.8 ± 5.74b
4.7 ± 3.65b
3.9 ± 3.82b
3.9 ± 3.82b
3.5 ± 3.89b
Percent progeny survival (at birth: relative to total delivered; on Days 1 and 4: relative to number born alive; on Days 7-28: relative to number retained on Day 4)
0
96.7
99.6
97.4
98.7
93.5
92.9
92.9
250
99.3
98.9
97.1
98.8
91.3
87.9
87.9
500
95.2
95.4
92.7
99.4
99.4
98.8
98.8
1400
85.6b
77.9b
75.3b
96.4
79.5
79.5
72.3
Mean progeny body weights (g)
Male
Female
0
5.9 ±0.9
-
7.9 ± 1.5
11.7 ±2.4
19.7 ±3.7
32.2 ±6.2
62.7 ± 10.4
56.0 ± 11.4
250
5.6 ± 0.8b
-
8.2 ± 1.8
11.4 ±2.4
17.8 ± 3.9b
28.4 ± 6.0b'd
53.1 ± 13.lb
51.0 ± 12.2
500
5.5 ± 0.8b
-
8.1 ± 1.6
11.3 ±2.1
17.9 ± 3.5b
27.2 ± 5.7b'd
52.2 ± 12.8b'd
49.5 ± 10.7b
1400
5.3 ± 0.8b'd
-
7.8 ±1.1
9.1 ± 2.0b'd
14.3 ± 2.0b'd
20.8 ± 4.6b'd
38.8 ± 9.3b'd
37.5 ± 9.9b'd
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Table 3. Summary of Progeny Observations from F1 Generation Rats Exposed to Cyclohexanone Vapor
Exposure Concentration
(ppm)
Lactation Day
0 (birth)
1
4
7
14
21
28
F2B Litter
Mean number of viable progeny from F1 generation rats
0
12.3 ±3.33
12.3 ±3.33
12.2 ±3.26
7.6 ± 1.23
7.6 ± 1.23
7.6 ± 1.23
7.5 ± 1.28
250
12.8 ±2.67
12.1 ±3.95
12.0 ±3.92
7.4 ± 1.95
7.0 ±2.55
7.0 ±2.55
7.0 ±2.55
500
10.9 ±3.76
10.8 ±3.71
10.4 ±4.03
6.9 ±2.47
6.8 ±2.73
6.8 ±2.73
6.8 ±2.73
1400
9.3 ±5.58
7.0 ± 6.34a
6.4 ± 6.66b
3.9 ± 3.75b
3.8 ± 3.87b
3.8 ± 3.87b
3.8 ± 3.87b
Percent progeny survival (at birth: relative to total delivered; on Days 1 and 4: relative to number born alive; on Days 7-28: relative to number retained on Day 4)
0
98.1
100.0
99.0
99.2
99.2
99.2
98.5
250
99.6
98.5
94.1
99.3
94.6
94.6
94.6
500
99.5
95.0
94.9
96.9
95.3
95.3
95.3
1400
97.4
75.2b
69. lb
96.9
92.3
92.3
92.3
Mean progeny body weights (g)
Male
Female
0
5.9 ±0.8
-
8.9 ± 1.4
13.1 ± 1.5
22.5 ±3.7
35.5 ±6.9
67.8 ± 11.0
64.6 ±9.3
250
6.2 ± 0.9a
-
9.3 ± 1.3a
12.9 ±2.2
23.3 ±2.8
36.8 ±5.2
69.2 ±9.0
66.2 ±7.9
500
6.2 ± 1.0a
-
9.5 ± 1.5b
13.5 ± 1.8
22.0 ±3.4
34.6 ±6.5
69.2 ± 11.3
62.1 ± 10.9
1400
5.8 ± 1.2
-
8.3 ± 1.7a
10.8 ± 2.3b'°
17.3 ± 4.0b'°
26.0 ± 6.3b'°
51.9 ± 15.7b
47.9 ± 11.5b'°
Significantly different from controls (p < 0.05); progeny body-weight data based on individual weights analyzed by analysis of variance (ANOVA) and Scheffe's multiple comparison.
bSignificantly different from controls (p < 0.01); progeny body-weight data based on individual weights analyzed by ANOVA and Scheffe's multiple comparison.
Significantly different from controls based on mean litter weight data analyzed by ANOVA and Dunnett's f-test (p < 0.01).
Source: ABC (1986a).
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Groups of 10 pregnant Sprague-Dawley rats were exposed by inhalation to 0, 100, 250,
or 500 ppm (0, 402, 1004, or 2007 mg/m3) of cyclohexanone (99.8% purity) vapor, 7 hours/day
on GDs 5-20 (Samimi et al., 1989). Concurrent control groups of five pregnant rats exposed to
room air (negative controls) and five pregnant rats exposed to 2-ethoxyethanol (positive control)
were run with each exposure concentration. Dams were sacrificed on GD 21 and evaluated for
body weight, gravid uterus weights, mean number of corpora lutea per litter, mean number and
placement of early and late resorption sites per litter, mean percentage of resorption sites, and
placement of live and dead fetuses. Fetuses were examined for body weight and sex ratio as well
as for evidence of external, visceral, and skeletal malformations, and skeletal variations. No
mortality was reported. Dams exhibited only a slight reduction in body-weight gain compared to
controls. Gross examination revealed a grey mottling of the lungs in several dams exposed to
250 or 500 ppm (1004 or 2007 mg/m3) cyclohexanone (data not provided). The interpretation of
this finding is unclear because incidence was not provided, histology was not conducted, and no
other respiratory endpoints were evaluated. The data showed no treatment-related effects on the
number of corpora lutea per dam, number of implants, resorption percentage, fetal weight,
viability, or sex ratio. Table 4 shows developmental malformations and variations. Three rats
treated with >250 ppm (1004 mg/m3) exhibited visceral malformations in the form of a right
subclavian artery arising off of the aortic arch or absence of the innominate artery. Few external
or skeletal malformations or variations were observed among treated rats. Slight skeletal
developmental variations were noted in fetuses from rats treated with >250 ppm (1004 mg/m3)
characterized as rudimentary 14th ribs and incompletely ossified sternebrae numbers 5 and 6.
None of these findings were significantly different from controls. In the absence of other
conventional signs of embryotoxicity, Samimi et al. (1989) concluded that inhalation exposure to
up to 500-ppm (2007-mg/m3) cyclohexanone was not developmentally toxic in rats. In the
absence of significant systemic or developmental effects observed in rats in this study, a NOAEL
of 500 ppm (2007 mg/m3) is identified for maternal and developmental toxicity.
Table 4. Mean Percentages of Fetuses with External, Visceral, and Skeletal
Malformations and Skeletal Variations per Litter in Cyclohexanone-Exposed
Rats and Negative Controls3
Exposure
Concentration
(ppm)
External
Malformations
Visceral
Malformations
Skeletal
Malformations
Skeletal
Variations
NC
CH
NC
CH
NC
CH
NC
CH
100
1.5 ±3.4
0
0
0
0
0
13.7 ±6.4
24.2 ±23.4
250
0
0.8 ±2.2
0
5.0 ± 14.1
0
1.6 ±4.4
27.3 ±20.4
37.3 ±20.5
500
0
0
0
2.9 ±6.4
0
1.0 ±2.7
26.3 ±27.4
23.5 ±22.8
aValues reported as percentage ± standard deviation.
NC = negative control (exposed to room air); CH = cyclohexanone exposed.
Source: Samimi et al. (1989).
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Biodynamics (1984a), (range-finding data in Biodynamics 1983), exposed groups of
26 pregnant CD rats by inhalation to cyclohexanone (99.9% purity) vapor at mean measured
concentrations of 0 (sham-exposed), 303, 657, or 1410 ppm (0, 1216, 2638, or 5661 mg/m3),
6 hours/day, on GDs 6-19. Dams were evaluated for survival, body weight, clinical signs,
behavioral response, uterus weights, number of corpora lutea, and gross pathology.
Developmental endpoints included fetus viability, resorptions, implantations, fetal sex ratio, and
external, visceral, and skeletal malformations and skeletal variations. No mortality was observed
among treated dams, and no significant effects were observed on the number of corpora lutea or
pregnancy rate. Effects noted in dams were limited to the high-dose group and included adverse
clinical signs (lacrimation, lethargy, and nasal and vaginal discharge), abolition of a startle
response, and a significant decrease in body-weight gain during gestation (see Table 5).
Developmental effects were also limited to fetuses from high-dose dams and included decreased
fetal body weight and increased incidence of fetal skeletal alterations, including incompletely
ossified cranial bones, hyoid, sternebrae, metatarsals, and phalanges (see Table 5). The
increased incidence in skeletal variations was only significantly different from controls on a per
fetus-basis, not on a litter-basis, because 100% of the control and treated litters exhibited some
form of a skeletal variation. Based on clinical signs, abolition of startle response and decreased
body-weight gain, aNOAEL of 657 ppm (2638 mg/m3) and a LOAEL of 1410 ppm
(5661 mg/m3) are identified for maternal toxicity. Based on decreased fetal body weights and
increased skeletal variations at the high-dose, a NOAEL of 657 ppm (2638 mg/m3) and a
LOAEL of 1410 ppm (5661 mg/m3) are also identified for developmental toxicity.
Table 5. Maternal and Developmental Observations in Rats Exposed to Cyclohexanone
Vapor During Gestation
Endpoint
Exposure Concentration (ppm)
0
303
657
1410
Mean gestational body weights (g)
GD0
273 ± 20
274 ± 16
273 ± 15
274 ± 16
GD 6
307 ± 20
310 ±20
309 ± 19
310 ± 16
GD 15
350 ± 20
357 ±23
346 ±21
333 ± 19a
GD 20
420 ± 24
432 ±33
418 ±29
383 ± 29b
Mean corrected body weight gain (g)°
32.8 ± 12.3
35.5 ± 13.8
29.3 ± 13.7
13.2 ± 10.7b
Mean body weight of viable fetuses (g)
3.67 ±0.28
3.66 ±0.24
3.68 ±0.25
2.73 ± 0.43b
Males
3.77 ±0.28
3.8 ±0.26
3.76 ±0.27
2.83 ± 0.44b
Females
3.56 ±0.27
3.53 ±0.23
3.58 ±0.22
2.64 ± 0.45b
Incidence of fetal skeletal variationsd
Number of fetuses affected
133 (84.2%)
150 (89.3%)
138 (87.3%)
142 (97.9%f
Number of litters affected
24 (100%)
23 (100%)
24 (100%)
23 (100%)
aSignificantly different from controls (p < 0.05), test not specified.
bSignificantly different from controls (p < 0.01), test not specified.
°Corrected body weight = actual body weight - gravid uterine weight.
Values are presented as number (percent).
Source: Biodynamics (1984a).
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In a companion mouse study, groups of 30 pregnant CD-I mice were exposed to
cyclohexanone (99.9% purity) vapor at mean measured concentrations of 0 or 1380 ppm (0 or
5541 mg/m3), 6 hours/day, on GDs 6-17 (Biodynamics, 1984b). Evaluations were similar to
those described above for the rat study. Maternal effects included adverse clinical signs (shallow
breathing, lacrimation, lethargy), delayed response to stimulus, and reductions in gestational
body-weight gain and uterine weights (see Table 6). Based on corrected Day 18-body weights
(actual Day 18-body weight corrected by subtracting the weight of the gravid uterus), mean
weight gain during the exposure period was significantly higher than controls. Developmental
effects included an increase in resorptions, decreased fetal viability, decreased fetal body weight,
skeletal variations (retardation in ossification of the cranial bones, cervical vertebral centra, and
phalanges), and an increased incidence of visceral malformations (cleft palate and distended
renal pelvis) (see Table 6). Both cleft palate and distended renal pelvis have been noted at low
incidence in this strain of mouse based on historical controls. Biodynamics (1984b) concluded
that cyclohexanone was maternally toxic, fetotoxic, and embryotoxic at 1380 ppm
(5541 mg/m3)—but not teratogenic. Based on clinical signs in dams, decreased maternal body
weights and fetal effects including decreased weights and retarded ossification, a LOAEL of
1380 ppm (5541 mg/m3) is identified for both maternal and developmental effects.
OTHER STUDIES
Genotoxicity
In two independent reverse mutation assays, cyclohexanone tested negative for
mutagenicity in bacterial tests using Salmonella typhimurium strains TA1535, TA1537, TA98,
and TA100 (Haworth et al., 1983; Florin et al., 1980) in the presence and absence of metabolic
activation. However, Massoud et al. (1980) observed that cyclohexanone-induced genetic
mutations in Bacillus subtilis and produced a "large number of revertants" in S. typhimurium
strain TA98. This study is only available as an abstract and detailed information on the study
conditions or concentrations tested are not provided. There is no further elaboration on the
results for other S. typhimurium strains. A Danish review of this study indicates that the results
for S. typhimurium were ambiguous, as reversions were observed in controls, and there was no
evidence of dose dependency (Miljostyrelsen Institute, 2003). Cyclohexanone-induced DNA
damage in Escherichia coli in vitro (Rosenkranz and Leifer, 1980).
Cyclohexanone tested negative in a mouse lymphoma cell forward mutation assay in the
presence and absence of an exogenous metabolic system (McGregor et al., 1988). In Chinese
hamster ovary cells (CHO), cyclohexanone-induced gene mutations at the HGRPT locus and
sister chromatid exchanges in the absence of S9 metabolic activation but not in the presence of
metabolic activation (Aaron et al., 1985; DuPont, 1984). Under these same test conditions,
cyclohexanone did not induce chromosomal aberrations in CHO cells with or without metabolic
activation. However, chromosomal aberrations were induced by cyclohexanone in cultured
human leukocytes (Collin, 1971; Lederer et al., 1971), and an increased frequency in
chromosomal damage characterized by ploidy and structural changes was observed in human
lymphocytes (Dyshlovoi et al., 1981, as cited in IARC, 1989).
In vivo, de Hondt et al. (1983) observed an increase in the incidence of chromosomal
abnormalities in bone marrow cells of male rats characterized as chromatid gaps, breaks, centric
fusions, centromeric attenuation, chromatid exchanges, and polyploidy.
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Table 6. Maternal and Developmental Observations in Mice Exposed to Cyclohexanone
Vapor During Gestation
Endpoint
Exposure Concentration (ppm)
0
1380
Mean gestational body weights (g)
GD0
25 ± 1
25 ±2
GD 6
28 ±2
28 ±2
GD 12
35 ±2
34 ±2
GD 18
48 ±3
41 ± 6a
Mean corrected body weight gain (g)h
2.9 ±2.0
5.1 ± 2.7a
Number of resorptions
24
184
Mean ± SD
0.9 ± 1.1
6.3 ± 4.9°
Mean % ± SD
7.4 ± 10.1
51.4 ±38.6
Number of litters with resorptions (%)°
15 (53.6%)
26 (89.7%)a
Number of viable fetuses
301
170
Mean litter size ± SD
10.8 ± 1.8
5.9 ± 4.7a
Mean body weight of viable fetuses (g)
1.31 ±0.07
1.09 ± 0.08a
Males
1.33 ±0.09
1.09 ± 0.08a
Females
1.30 ±0.08
1.10 ± 0.06°
Incidence of fetal visceral malformations0
Number of fetuses affected
0
4 (4.4%)d
Number of litters affected
0
4 (19.0%)d
Incidence of fetal skeletal variations0
Number of fetuses affected
102 (70.8%)
79 (100%)a
Number of litters affected
26 (92.9%)
20 (100%)
Significantly different from controls (p < 0.01), test not specified.
bCorrected body weight = actual body weight - gravid uterine weight.
°Values are presented as number (percent).
Significantly different from controls (p < 0.05), test not specified.
Source: Biodynamics (1984b).
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DERIVATION OF PROVISIONAL SUBCHRONIC AND CHRONIC
ORAL RfD VALUES FOR CYCLOHEXANONE
Oral studies of cyclohexanone include subchronic and chronic drinking water studies in
rats and mice (Lijinsky and Kovatch, 1986; NCI, 1979), and screening level developmental
studies in mice (Gray et al., 1986; Seidenberg et al., 1986; Gray and Kavlock, 1984; Chernoff
and Kavlock, 1983; Gondry, 1973). Table 7 summarizes these data. Both the subchronic and
chronic studies indicate that cyclohexanone exposure results in decreased body weights in both
rats and mice. The developmental screening studies also observe decreased gestational body
weights and reduced fetal body weights among mice treated orally with cyclohexanone.
SUBCHRONIC p-RfD
The principal study of Lijinsky and Kavatch (1986) is based on an NCI cancer bioassay
oral study with adequate toxicologic endpoints using both mice and rats. It also provided the
lowest LOAEL among the subchronic and developmental toxicity studies; the LOAEL of
1010 mg/kg-day for decreased body-weight gain in male rats treated in the drinking water for
25 weeks (Lijinsky and Kavatch, 1986). The rat, rather than the mouse, is chosen as the species
to define the critical effect from the study because the rats were exposed for 25 weeks compared
to only 13 weeks for the mice, and descriptive details are lacking from the study author's
presentation of the mice data. The corresponding NOAEL of 731 mg/kg-day was chosen as the
point of departure (POD) for deriving the subchronic p-RfD (Lijinsky and Kovatch, 1986; NCI,
1979). Benchmark dose (BMD) modeling cannot be conducted because the body weight data
were not provided in the principal study.
A subchronic p-RfD was derived for cyclohexanone by dividing the NOAEL of
731 mg/kg-day for decreased body-weight gain in male rats by a UF of 300, as shown below:
Subchronic p-RfD = NOAEL UF
= 731 mg/kg-day -^300
= 2 mg/kg-day
The composite UF of 300 is composed of the following UFs:
•	UFh: A factor of 10 is applied for extrapolation to a potentially susceptible human
subpopulation because data for evaluating susceptible human response are
insufficient.
•	UFa: A factor of 10 is applied for animal-to-human extrapolation because data for
evaluating relative interspecies sensitivity are insufficient.
•	UFd: The database for oral exposure to cyclohexanone consists of limited subchronic
toxicity studies in two species and several screening level developmental toxicity
studies in mice. A factor of 3 (10°5) is applied for database inadequacies because
data for evaluating reproductive toxicity are inadequate and limited.
•	UFl: A NOAEL is identified from the database and used as the POD; therefore, a
factor of 1 is used.
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Table 7. Summary of Oral Noncancer Dose-Response Information for Cyclohexanone
Species and
Study Type
(w/sex/group)
Exposure
NOAEL
(mg/kg-day)a
LOAEL
(mg/kg-day)a
Responses at the
LOAEL
Comments
Reference
Rat (5/sex/group)
Continuously in the drinking
water for 25 weeks at 0, 30,
62, 124, 249, 513, 731, or
1010 mg/kg-day in males
andO, 32, 68, 135,271, 559,
796, or 1100 mg/kg-day in
females.
731 (males)
796 (females)
1010 (males)
1100 (females)
Decreased weight gain.
Weight-gain data were not
provided other than to state
that the magnitude of
difference from controls was
10%. Moderate chronic
respiratory infection among
control and treated animals.
Lijinsky and
Kovatch, 1986
Mouse
(10/sex/group)
Continuously in the drinking
water for 13 weeks at 0, 99,
568, 1600, 3210,6170, 8390,
or 11,600 mg/kg-day in
males and 0, 106, 608, 1720,
3430, 6610, 8980, or
12,400 mg/kg-day in
females.
3210 (males)
6610 (females)
6170 (males)
8980 (females)
Decreased body
weights.
Weight-gain data were not
provided other than the
percentage differences from
controls in selected groups.
Body-weight changes among
high-dose animals were not
described.
Lijinsky and
Kovatch, 1986
Rat (52/sex/group)
Continuously in the drinking
water for 2 years at 0, 426, or
838 mg/kg-day for males, 0,
476, or 937 for females, and
0, 462, or 910 mg/kg-day for
both sexes.
462
910
Depression in
body-weight gain.
Principal study used as the
basis of the EPA IRIS RfD
value. No significant
dose-related trend in tumor
incidence was observed.
Lijinsky and
Kovatch, 1986
Mouse
(41-52/sex/group)
Continuously in the drinking
water for 2 years at 0, 1530,
or 3070 mg/kg-day for males
and at 0, 1570, 3130, or
6020 mg/kg-day for females.
1530 (males)
NA (females)
3070 (males)
NA (females)
Decreased body
weights.
NOAEL/LOAEL was not
derived for females due to
confounding lymphatic
lesions in control and treated
mice. No significant dose-
related trend in tumor
incidence was observed.
Lijinsky and
Kovatch, 1986
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Table 7. Summary of Oral Noncancer Dose-Response Information for Cyclohexanone
Species and
Study Type
(«/sex/group)
Exposure
NOAEL
(mg/kg-day)a
LOAEL
(mg/kg-day)a
Responses at the
LOAEL
Comments
Reference
Mouse
(unknown group
size)
Diet containing
2000 mg/kg-day fed
continuously throughout
pregnancy and lactation for
multiple generations.
NA
2000
Neonatal mortality
within the first 21 days
of life and depressed
offspring growth.
Study report written in French
with English abstract.
Gondry, 1973
Mouse
(24/group)
Gavage treatment at 0 or
800 mg/kg-day on
GDs 8-12.
800
NA
No significant effects on
dams or offspring.

Chernoff and
Kavlock, 1983
Mouse
(10/group)
Gavage treatment at 0 or
800 mg/kg-day on
GDs 8-12.
800
NA
No significant effects on
dams or offspring.

Gray et al., 1986;
Gray and Kavlock,
1984
Mouse
(28/group)
Gavage treatment at 0 or
2200 mg/kg-day on
GDs 8-12.
NA
2200
Decreased gestational
body weights in dams;
decreased neonatal body
weights.

Seidenberg et al.,
1986
"NOAEL and LOAEL are based on continuous exposure in these studies.
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Confidence in the principal study (Lijinsky and Kovatch, 1986) is medium. The
subchronic study included multiple dose levels but only five rats per sex at each level. The
evaluations are limited to survival, body weight, and gross and microscopic examinations. The
results are reported briefly with few details, and no data were shown. All rats, including
controls, displayed signs of moderate chronic respiratory disease. However, the corresponding
chronic study (Lijinsky and Kovatch, 1986) demonstrated similar results and improved
evaluations with 52 rats per sex per dose level. Confidence in the database is medium. In
addition to the principal subchronic rat study, there is a subchronic mouse study reported by the
same researchers that provides supporting evidence for an effect on body weight but had some of
the same shortcomings. In addition, the database includes screening-level developmental studies
in mice, which were consistent in finding effects on offspring body weight at high doses and no
effects at doses near the POD. Reproductive toxicity has not been adequately studied by oral
exposure; Gondry (1973) only evaluated mortality and growth in the offspring and provided no
statistical or numerical information. The inhalation database suggests that developmental effects
may be an endpoint of concern for cyclohexanone. Overall confidence in the subchronic p-RfD
is medium.
The subchronic p-RfD derived herein is lower than the chronic RfD for cyclohexanone
on IRIS. This is because the derivation of the subchronic p-RfD includes application of a
database UF, which was not EPA practice at the time the chronic IRIS RfD was developed
(posted 09/02/1986).
CHRONIC p-RfD
A chronic RfD of 5 mg/kg-day based on the 2-year drinking water study in rats (Lijinsky
and Kovatch, 1986) was derived by EPA in 1986 and is available on IRIS (U.S. EPA, 2009).
The RfD was calculated from a NOAEL of 462 mg/kg-day for decreased body-weight gain
derived from combined male and female data and a composite UF of 100 (10 for interspecies
extrapolation and 10 for intraspecies variability among the human population).
DERIVATION OF PROVISIONAL SUBCHRONIC AND CHRONIC
INHALATION RfC VALUES FOR CYCLOHEXANONE
Human studies indicate that inhalation of cyclohexanone can cause eye, nose, and throat
irritation (Esso Research and Engineering Co., 1965; Nelson et al., 1943). Interpretation of
results suggestive of neurological effects at relatively low concentrations (Mitran et al., 1997) is
limited by methodological and reporting problems with the study. The human data are not
considered to be suitable for the derivation of the p-RfC values. Inhalation studies in animals
include a subchronic study that evaluated various endpoints in rabbits (Treon et al., 1943), a
series of subchronic studies that were specifically designed to evaluate the effects on the
olfactory bulb in rats (Rehn et al., 1988; Panhuber and Laing, 1987; Panhuber et al., 1987;
Laing et al., 1985; Laing and Panhuber, 1980, 1978; Pinching and Doving, 1974), a
multigenerational reproduction study in rats (ABC, 1986a,b), and developmental studies in rats
(Samimi et al., 1989; Biodynamics, 1984a) and mice (Biodynamics, 1984b). Table 8
summarizes these data. The primary findings were clinical signs indicative of eye irritation and
neurological effects and reduced body-weight gain in rabbits with subchronic exposure
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Table 8. Summary of Inhalation Noncancer Dose-Response Information for Cyclohexanone
Species and
Study Type
(«/sex/group)
Exposure
NOAEL
(mg/m3)
LOAEL
(mg/m3)
Responses at the LOAEL
Comments
Reference
Rabbit (4/group)
0, 763, 1241, 3103,
5677, or
12,373 mg/m3
6 hours/day,
5 days/week for
3 weeks
(12,373 mg/m3) for
10 weeks (all other
groups).
1241
HEC:
222
3103
HEC:
554
Clinical signs as salivation,
conjunctival congestion and
irritation, lacrimation and lethargy.
More severe clinical signs including
conjunctival irritation, lacrimation,
lethargy, narcosis, labored breathing,
loss of coordination, weight loss, and
hypothermia were observed at higher
concentrations.
Treonetal., 1943
Rat (various group
sizes)
0-32	mg/m3 for
1-4	months.
NA
NA
NA
Alterations in mitral cells and spine
density of granule cell dendrites were
seen at all concentrations tested, but
also in response to deodorized air.
Interpretation of these results is
unclear. There was no effect on
olfactory sensitivity.
Rehnetal., 1988;
Panhuber and
Laing, 1987;
Panhuber et al.,
1987; Laing etal.,
1985; Laing and
Panhuber, 1980,
1978; Pinching
andDoving, 1974
Rat (30/sex/group)
0, 1004, 2007, or
4015/5621 mg/m3
6 hours/day through
two consecutive
generations.
2007
HEC:
502
5621
HEC:
1405
Effects inFl animals included
mortality, adverse clinical signs,
decreases in body weights,
decreased fertility in males, and
decreases in progeny viability,
survival, and body weights.
A subsequent study determined that
the decrease in male fertility in this
study was reversible in the second
generation (ABC, 1986b).
ABC, 1986a
Rat (10/group)
0, 402, 1004, or
2007 mg/m3
7 hours/day on
GDs 5-20.
2007
HEC:
585
NA
NA
Grey mottling of the lungs was
observed in rats exposed to 1004, or
2007 mg/m3 upon gross examination.
No histology was performed, and
incidence was not reported.
Samimi et al.,
1989
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Table 8. Summary of Inhalation Noncancer Dose-Response Information for Cyclohexanone
Species and
Study Type
(«/sex/group)
Exposure
NOAEL
(mg/m3)
LOAEL
(mg/m3)
Responses at the LOAEL
Comments
Reference
Rat (26/group)
0, 1216, 2638, or
5661 mg/m3
6 hours/day on
GDs 6-19.
2638
HEC:
660
5661
HECa:
1415
Adverse clinical signs, abolition of
a startle response, and decreased
gestational body-weight gain in
dams. Decreased fetal body
weight and increased incidence of
fetal skeletal alterations.

Biodynamics,
1984a
Mouse (30/group)
0 or 5541 mg/m3
6 hours/day on
GDs 6-17.
NA
5541
HECa:
1385
Adverse clinical signs, delayed
response to stimulus, and
decreased gestational body-weight
gains in dams. Increased incidence
of resorptions, decreased fetal
viability and body weights, and
retarded bone ossification.

Biodynamics,
1984b
'HEC calculated as follows: NOAEL[hec] = NOAEL x exposure hours/24 hours x exposure days/7 days x dosimetric adjustment. For systemic effects, the dosimetric
adjustment is the ratio of the animal:human blood:gas partition coefficients for cyclohexanone (in the absence of experimental values, a default value of 1 was used).
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(Treon et al., 1943) and at higher concentrations in rats and mice exposed in reproductive and
developmental studies (ABC, 1986a; Biodynamics, 1984a,b). The latter studies also showed
effects on offspring viability and development (reduced body weights and increased skeletal
variations indicative of developmental delay) at the same concentrations.
The NOAELs and LOAELs in Table 8 were converted to human equivalent
concentrations (HECs) in order to facilitate comparisons across studies. First, the
concentrations, expressed originally in mg/m3, were adjusted to continuous exposure
(NOAELadj)- Current EPA practice is to include an adjustment to continuous exposure for
developmental effects, as is typically done for other endpoints (see U.S. EPA, 2002). The
NOAELadj values were calculated as follows:
NOAELadj = (NOAEL) (# hours ^ 24 hours) (# days ^ 7 days)
Because the observed effects of cyclohexanone were systemic in nature, the chemical
was treated as a Category 3 gas. The human equivalent concentration (NOAELHec) is calculated
for a Category 3 gas by multiplying the NOAELadj by the ratio of the blood:gas (air) partition
coefficients of cyclohexanone in animals and humans. However, partition coefficients for
cyclohexanone are not available in humans or animals. In accordance with "Methods of
derivation of inhalation reference concentrations and application of inhalation dosimetry"
(U.S. EPA, 1994b), the value of 1.0 is used for the ratio, and the NOAELrec values shown in
Table 8 are calculated as follows:
NOAELrec = NOAELadj x (Hb/g)A ^ (Hb/g)H
SUBCHRONIC p-RfC
The lowest LOAELrec across inhalation studies is 554 mg/m3 based on adverse clinical
signs in rabbits including salivation, conjunctival congestion and irritation, lacrimation, lethargy,
narcosis, labored breathing, loss of coordination, weight loss, and hypothermia (Treon et al.,
1943). The corresponding NOAELrec is 222 mg/m3. Similar clinical signs were seen at
LOAELrec values of approximately 1400 mg/m3 in parental rats and mice in the reproduction
(ABC, 1986a) and developmental (Biodynamics, 1984a,b) studies. Treon (1943) evaluated a
relatively wide array of clinical and histopathological effects, and the only other available data
are reproductive/developmental studies that demonstrated slightly higher NOAELs. The
NOAELrec of 222 mg/m3, based on clinical signs in rabbits at the next highest dose (described
above; Treon et al., 1943), is chosen as the POD for deriving the p-RfC. BMD modeling cannot
be conducted because quantitative data were not provided in the principal study.
A subchronic p-RfC for cyclohexanone, based on the NOAELrec of 222 mg/m3 for
adverse clinical signs in rabbits (Treon et al., 1943), was derived as follows:
Subchronic p-RfC = NOAELrec ^ UF
= 222 mg/m3 - 30
= 7 mg/m3
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The composite UF of 30 is composed of the following UFs:
•	UFa: A factor of 3 (10°5) is applied for animal-to-human extrapolation to account for
potential pharmacodynamic differences between rabbits and humans. The dosimetric
conversion to an HEC accounts for pharmacokinetic differences.
•	UFh: A factor of 10 is applied for extrapolation to a potentially susceptible human
subpopulation because data for evaluating susceptible human response are
insufficient.
•	UFd: The database for inhalation toxicity of cyclohexanone consists of a subchronic
toxicity study in rabbits, a series of subchronic studies in rats specifically evaluating
the effects on cells of the olfactory bulb, a multigenerational reproduction study in
rats, and developmental toxicity studies in rats and mice. A factor of 1 is applied for
database inadequacies.
•	UFl: A NOAEL is identified from the database and used as the POD; therefore, a
factor of 1 is used.
Confidence in the principal study (Treon et al., 1943) is low based on small group sizes
(four rabbits/group), examination of limited endpoints, inclusions of a 2-month recovery period
prior to histopathological examination, and insufficient detail in reporting of results. Confidence
in the database is medium. Aside from the subchronic study in rabbits used to derive the p-RfC,
a multigenerational reproduction study in rats and developmental toxicity studies in rats and
mice are also available that support the findings in rabbits. However, there are no supporting
systemic toxicity studies in other species. Overall confidence in the subchronic p-RfC is low.
CHRONIC p-RfC
To derive the chronic p-RfC in the absence of chronic data, the POD from the subchronic
p-RfC (adverse clinical signs in rabbits) is used along with a composite UF that includes the
same areas of uncertainty enumerated above for the subchronic p-RfC, as well as additional
10-fold UFs, as follows:
•	UFa: A factor of 3 (10°5) is applied for animal-to-human extrapolation to account for
potential pharmacodynamic differences between rabbits and humans. The dosimetric
conversion to an HEC accounts for pharmacokinetic differences.
•	UFh: A factor of 10 is applied for extrapolation to a potentially susceptible human
subpopulation because data for evaluating susceptible human response are
insufficient.
•	UFd: The database for inhalation toxicity of cyclohexanone consists of a subchronic
toxicity study in rabbits, a series of subchronic studies in rats specifically evaluating
the effects on cells of the olfactory bulb, a multigenerational reproduction study in
rats and developmental toxicity studies in rats and mice. A factor of 1 is applied for
database inadequacies.
•	UFl: A NOAEL is identified from the database and used as the POD; therefore, a
factor of 1 is used.
•	UFS: A factor of 10 is applied for using data from a subchronic study to assess
potential effects from chronic exposure because data for evaluating responses after
chronic exposure are not available.
This results in a total UF of 300 for derivation of the chronic p-RfC.
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A chronic p-RfC for cyclohexanone, based on the NOAELrec of 222 mg/m3 for adverse
clinical signs in rabbits (Treon et al., 1943), is derived as follows:
Chronic p-RfC = NOAELHEc-UF
= 222 mg/m3 - 300
= 0.7 mg/m3
As discussed for the subchronic p-RfC, confidence in the principal study is low.
However, unlike the characterization of the database in reference to the subchronic value
derivation, confidence in the database, in the context of a chronic value derivation, is reduced to
low for the chronic p-RfC due to the absence of a chronic study. Overall confidence in the
chronic p-RfC is low.
PROVISIONAL CARCINOGENICITY ASSESSMENT
FOR CYCLOHEXANONE
WEIGHT-OF-EVIDENCE DESCRIPTOR
Under the 2005 Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005), there is
"Inadequate Information to Assess [the] Carcinogenic Potential" of cyclohexanone. The only
available studies that evaluated the carcinogenic potential of cyclohexanone were the 2-year
drinking water studies in rats and mice conducted by Lijinsky and Kovatch (1986). These
studies found increased incidences of adrenocortical adenomas in low-dose male rats, increased
incidence of hepatocellular tumors in low-dose male mice, and increased incidence of malignant
lymphoma in low-dose female mice. None of these findings exhibited a dose-related trend, as
these neoplasms were not observed at higher doses. In addition, there was a marginal increase in
the incidence of follicular cell adenoma-carcinomas of the thyroid gland among high-dose males
compared to concurrent controls. Lijinsky and Kovatch (1986) concluded that the evidence of
carcinogenic activity is marginal, and the effect, if any, is weak. IARC (1999, 1989) reviewed
these studies and characterized cyclohexanone as Not Classifiable As to Its Carcinogenicity To
Humans based on inadequate data. ACGIH (2007) also considered cyclohexanone to be Not
Classifiable As a Human Carcinogen (A4). Genotoxicity data are mixed. Assays for
mutagenicity were largely negative in bacteria but mixed in mammalian cells, and there is some
evidence of clastogenicity in human cells in vitro and in rats in vivo.
QUANTITATIVE ESTIMATES OF CARCINOGENIC RISK
A lack of suitable data precludes derivation of quantitative estimates of cancer risk for
cyclohexanone.
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REFERENCES
Aaron, CS; Brewen, JG; Stetka, DG; et al. (1985) Comparative mutagenesis in mammalian
cells (CHO) Chinese hamster ovary in culture multiple genetic end point analysis of
cyclohexanone in-vitro. Environ Mutagen 7:60-61.
ABC (American Biogenics Corporation). (1986a) Two-generation reproduction study via
inhalation (with a neurotoxicology/pathology component) in albino rats using cyclohexanone.
Study No. 450-1587. Submitted under TSCA; EPA Document No. OTS40-8666137.
ABC (American Biogenics Corporation). (1986b) Assessment of male reproductive
performance during a post exposed recovery period of second generation males from a two
generation reproduction study (final report) with letter. Study No. 450-2326. Submitted under
TSCA; EPA Document No. 40-8666150.
ACGIH (American Conference of Governmental Industrial Hygienists). (2003) Documentation
of the threshold limit values for chemical substances. 7th Edition. Cincinnati, OH: ACGIH.
ACGIH (American Conference of Governmental Industrial Hygienists). (2007) Threshold limit
values for chemical substances and physical agents and biological exposure indices. Cincinnati,
OH: ACGIH.
ATSDR (Agency for Toxic Substances and Disease Registry). (2009) Toxicological profile
information sheet. U.S. Department of Health and Human Services, Public Health Service.
Available online at http://www.atsdr.cdc.gov/toxpro2.html.
Bereznyak (1984) - Cited in IARC (International Agency for Research on Cancer). (1989)
Cyclohexanone. In: IARC monographs on the evaluation of carcinogenic risks to humans.
Volume 47. Lyon, France: IARC; pp. 157-169.
Biodynamics. (1983) An inhalation dosage range-finding study in pregnant rats with
cyclohexanone. Project No. 83-2718. Submitted under TSCA; EPA Document No. 40-8466096;
NTIS No. OTS0507478.
Biodynamics. (1984a) Inhalation teratology study in rats with cyclohexanone. Project No.
83-2719. Submitted under TSCA; EPA Document No. #40-8466096; NTIS No. OTS0507478.
Biodynamics. (1984b) Inhalation teratogenicity study in the mouse with cyclohexanone.
Project No. 83-2766. Submitted under TSCA; EPA Document No. 40-8466096; NTIS No.
OTS0507478.
CalEPA (California Environmental Protection Agency). (2009a) OEHHA/ARB Approved
chronic reference exposure levels and target organs. Available online at
http ://www. arb. ca. gov/toxics/healthval/chronic.pdf.
28
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9-15-2010
CalEPA (California Environmental Protection Agency). (2009b) Air chronic reference exposure
levels adopted by OEHHA as of February 2005. Available online at
http://www.oehha.ca.gov/air/chronic rets/AllChrels.html.
CalEPA (California Environmental Protection Agency). (2009c) Hot spots unit risk and cancer
potency values. Available online at http://www.oehha.ca.gov/air/hot spots/pdf/TSDlookup20
02.pdf.
Chernoff, N; Kavlock, RJ. (1983) A teratology test system which utilizes postnatal growth and
viability in the mouse. Environ Sci Res 27:417-427.
Collin, JP. (1971) Cytogenetic effect of cyclamate, cyclohexanone and cyclohexanol. Le
Diabete 19(4):215-221.
de Hondt, HA; Temtamy, SA; Abd-Aziz, KB. (1983) Chromosomal studies on laboratory rats
(Rattus norvegicus) exposed to an organic solvent (cyclohexanone). Egypt J Genet Cytol
12:31-40.
DuPont de Nemours & Company Inc. (DUPONT). (1984) Evaluation of cyclohexanone in the
multiple genetic endpoint assay in cultured mammalian cells with cover letter. Submitted under
TSCA; NTIS No. OTS0206695.
Esso Research and Engineering Co. (1965) Acute inhalation and human sensory irritation
studies on cyclopentanone, isophorone, dihydroisophorone, cyclohexanone, and methyl isobutyl
ketone. Submitted under TSCA; NTIS No. OTS02206266.
Florin, I; Rutberg, L; Curvall, M; et al. (1980) Screening of tobacco smoke constituents for
mutagenicity using the Ames' test. Toxicology 15:219-232.
Gondry, E. (1973) Toxicity of cyclohexylamine, cyclohexanone, and cyclohexanol and
metabolites of cyclamate. J Environ Toxicol 5:227-238.
Gray, LE, Jr; Kavlock, RJ. (1984) An extended evaluation of an in vivo teratology screen
utilizing postnatal growth and viability in the mouse. Teratog Carcinog Mutagen 4:403-426.
Gray, LE, Jr; Kavlock, RJ; Ostby, J; et al. (1986) An evaluation of figure-eight maze activity
and general behavioral development following prenatal exposure to forty chemicals: effects of
cytosine arabinoside, dinocap, nitrofen, and vitamin A. Neurotoxicology 7:449-462.
Haworth, S; Lawlor, T; Mortelmans, K; et al. (1983) Salmonella mutagenicity test results for
250 chemicals. Environ Mutagen 5(Suppl 1):3—142.
IARC (International Agency for Research on Cancer). (1989) Cyclohexanone. In: IARC
monographs on the evaluation of carcinogenic risks to humans. Volume 47. Lyon, France:
IARC; pp. 157-169.
29
Cyclohexanone

-------
FINAL
9-15-2010
IARC (International Agency for Research on Cancer). (1999) Cyclohexanone. In: IARC
monographs on the evaluation of carcinogenic risks to humans. Volume 71, Part 3. Lyon,
France; pp. 1359-1364.
Jacobsen, M; Baelum, J; Bonde, JP. (1994) Temporal epileptic seizures and occupational
exposure to solvents. Occup Environ Med 51:429-430.
Laing, DG; Panhuber, H. (1978) Neural and behavioral changes in rats following continuous
exposure to an odor. J Comp Physiol 124:259-265.
Laing, DG; Panhuber, H. (1980) Olfactory sensitivity of rats reared in an odorous or deodorized
environment. Physiol Behav 25(4):555-558.
Laing, DG; Panhuber, H; Pittman, EA; et al. (1985) Prolonged exposure to an odor or
deodorized air alters the size of mitral cells in the olfactory bulb. Brain Res 336(1):81—88.
Lederer, J; Collin, JP; Pottier-Arnould, AM; et al. (1971) L'action cytogenetique et teratogene
du cyclamate et de ses metabolites. Therapeutique 47(4):357-363.
Lijinsky, W; Kovatch, RM. (1986) Chronic toxicity study of cyclohexanone in rats and mice
(NCI study). J Natl Cancer Inst 77: 941-949.
Massoud, A; Aly, A; Shafik, H. (1980) Mutagenicity and carcinogenicity of cyclohexanone.
MutatRes 74:174.
McGregor, DB; Brown, A; Cattanach, P; et al. (1988) Responses of the 15178y tk+/tk- mouse
lymphoma cell forward mutation assay. III.72 coded chemicals. Environ Mol Mutagen
12:85-154. Erratum in: Environ Mol Mutagen 112(153):345.
Miljostyrelsen Institute. (2003, Accessed 2010) Appendices 1-18 to: Report on the health
effects of selected pesticide coformulants. Working Report No. 51. Particularly Table 4.
Available online at http://www2.mst.dk/Udgiv/publications/2003/87-7614-055-
5/html/kapl21 eng.htm.
Mitran, E; Callender, T; Orha, B; et al. (1997) Neurotoxicity associated with occupational
exposure to acetone, methyl ethyl ketone, and cyclohexanone. Environ Res 73:181-188.
NCI (National Cancer Institute). (1979) Summary and experimental design of subchronic
studies of cyclohexanone. Final Report (as cited in U.S. EPA, 2009).
Nelson, KW; Ege, JF; Ross, M; et al. (1943) Sensory response to certain industrial solvent
vapors. Ind Hygiene Toxicol 25:282-285.
NIOSH (National Institute for Occupational Safety and Health). (2005) NIOSH pocket guide to
chemical hazards. Available online at http://www.cdc.gov/niosh/npe/.
30
Cyclohexanone

-------
FINAL
9-15-2010
NRC (National Research Council). (1979) Odors from stationary and mobile sources.
Committee on Odors from Stationary and Mobile Sources. Board of Toxicology and
Environmental Health Hazards. National Academy of Sciences, Washington, D.C.
NTP (National Toxicology Program). (2005) 11th report on carcinogens. Available online at
http://ntp. niehs.nih.gov/index.cfm? obi ectid=326A9724-F 1F6-975E-7FCE50709CB4C932.
OSHA (Occupational Safety and Health Administration). (2009) OSHA Standard 1910.1000
Table Z-l. Part Z, Toxic and Hazardous Substances. Available online at http://www.osha.gov/
pis/oshaweb/owadisp.show document?p table STANDARDS&p id=9992.
Panhuber, H; Laing, DG. (1987) The size of mitral cells is altered when rats are exposed to an
odor from their day of birth. Brain Res 431(1): 133-140.
Panhuber, H; Mackay-Sim, A; Laing, DG. (1987) Prolonged odor exposure causes severe cell
shrinkage in the adult rat olfactory bulb. Brain Res 428(2):307-311.
Pinching, AJ; Doving, KB. (1974) Selective degeneration in the rat olfactory bulb following
exposure to different odors. Brain Res 82(2): 195-204.
Rehn, B; Panhuber, H; Laing, DG; et al. (1988) Spine density on olfactory granule cell
dendrites is reduced in rats reared in a restricted olfactory environment. Brain Res
468(1):143-147.
Rosenkranz, HS; Leifer, Z. (1980) Determining the DNA-modifying activity of chemicals using
DNA polymerase-deficient Escherichia coli. Chem Mut 6:109-147.
Samimi, BS; Harris, SB; de Peyster, A. (1989) Fetal effects of inhalation exposure to
cyclohexanone vapor in pregnant rats. Toxicol Ind Health 5:103 5-1043.
Seidenberg, JM; Anderson, DG; Becker, RA. (1986) Validation of an in vivo developmental
toxicity screen in the mouse. Teratog Carcinog Mutagen 6:361-374.
Treon, JF; Crutchfield, WE, Jr; Kitzmiller, KV. (1943) The physiological response of animals
to cyclohexane, methylcyclohexane, and certain derivatives of these compounds. J Ind Hyg
Toxicol 25:323-346.
U.S. EPA (U.S. Environmental Protection Agency). (1988) Recommendations for and
documentation of biological values for risk assessment. Prepared by the Office of Health and
Environmental Assessment, Environmental Criteria and Assessment Office, Cincinnati, OH.
EPA/600/6-87/008.
U.S. EPA (U.S. Environmental Protection Agency). (1991) Chemical Assessments and Related
Activities (CARA). Office of Health and Environmental Assessment, Washington, DC.
U.S. EPA (U.S. Environmental Protection Agency). (1994a) Chemical Assessments and
Related Activities (CARA). Office of Health and Environmental Assessment, Washington, DC.
31
Cyclohexanone

-------
FINAL
9-15-2010
U.S. EPA (U.S. Environmental Protection Agency). (1994b) Methods of derivation of
inhalation reference concentrations and application of inhalation dosimetry. Office of Research
and Development, National Center for Environmental Assessment, Washington, DC. October
1994. EPA/600/8-90/066F.
U.S. EPA (U.S. Environmental Protection Agency). (1997) Health Effects Assessment
Summary Tables. FY-1997 Update. Prepared by the Office of Research and Development,
National Center for Environmental Assessment, Cincinnati OH for the Office of Emergency and
Remedial Response, Washington, DC. EPA/540/R-97/036. NTIS PB97-921199.
U.S. EPA (U.S. Environmental Protection Agency). (2002) A review of the reference dose and
reference concentration processes. Risk Assessment Forum, U.S. Environmental Protection
Agency, Washington, DC. EPA/630/P-02/002F. December 2002.
U.S. EPA (U.S. Environmental Protection Agency). (2005) Guidelines for carcinogen risk
assessment. Risk Assessment Forum, National Center for Environmental Assessment,
Washington, DC. EPA/630/P-03/001F. Available online at http://cfpub ena.gov/ncea/cfm/
recordi splay.ct'm?deid 116283.
U.S. EPA (U.S. Environmental Protection Agency). (2006) 2006 Edition of the drinking water
standards and health advisories. Office of Water, Washington, DC. EPA 822-R-02-038.
Washington, DC. Available online at http://www.epa.gov/waterscience/drinking/standards/
dwstandards.pdf.
U.S. EPA (U.S. Environmental Protection Agency). (2009) Integrated Risk Information System
(IRIS). Office of Research and Development, National Center for Environmental Assessment,
Washington, DC. Available online at http J/www, epa. gov/iri s/.
WHO (World Health Organization). (2009) Online catalogs for the Environmental Health
Criteria Series. Available online at http://www.who.int/ipcs/publications/ehc/ehc alphabetical/
en/index.html.
32
Cyclohexanone

-------