% jT%J^ United States
I^bTSp^ Environmental Protection
Jf % Agency
EPA/690/R-05/017F
Final
8-03-2005
Provisional Peer Reviewed Toxicity Values for
N-Methyl aniline
(CASRN 100-61-8)
Derivation of Subchronic and Chronic Oral RfDs
Superfund Health Risk Technical Support Center
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268

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Acronyms and Abbreviations
bw	body weight
cc	cubic centimeters
CD	Caesarean Delivered
CERCLA	Comprehensive Environmental Response, Compensation and Liability Act
of 1980
CNS	central nervous system
cu.m	cubic meter
DWEL	Drinking Water Equivalent Level
FEL	frank-effect level
FIFRA	Federal Insecticide, Fungicide, and Rodenticide Act
g	grams
GI	gastrointestinal
HEC	human equivalent concentration
Hgb	hemoglobin
i.m.	intramuscular
i.p.	intraperitoneal
i.v.	intravenous
IRIS	Integrated Risk Information System
IUR	inhalation unit risk
kg	kilogram
L	liter
LEL	lowest-effect level
LOAEL	lowest-observed-adverse-effect level
LOAEL(ADJ)	LOAEL adjusted to continuous exposure duration
LOAEL(HEC)	LOAEL adjusted for dosimetric differences across species to a human
m	meter
MCL	maximum contaminant level
MCLG	maximum contaminant level goal
MF	modifying factor
mg	milligram
mg/kg	milligrams per kilogram
mg/L	milligrams per liter
MRL	minimal risk level
1

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MTD
maximum tolerated dose
MTL
median threshold limit
NAAQS
National Ambient Air Quality Standards
NOAEL
no-observed-adverse-effect level
NOAEL(ADJ)
NOAEL adjusted to continuous exposure duration
NOAEL(HEC)
NOAEL adjusted for dosimetric differences across species to a human
NOEL
no-observed-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
PBPK
physiologically based pharmacokinetic
PPb
parts per billion
ppm
parts per million
PPRTV
Provisional Peer Reviewed Toxicity Value
RBC
red blood cell(s)
RCRA
Resource Conservation and Recovery Act
RDDR
Regional deposited dose ratio (for the indicated lung region)
REL
relative exposure level
RfC
inhalation reference concentration
RfD
oral reference dose
RGDR
Regional gas dose ratio (for the indicated lung region)
s.c.
subcutaneous
SCE
sister chromatid exchange
SDWA
Safe Drinking Water Act
sq.cm.
square centimeters
TSCA
Toxic Substances Control Act
UF
uncertainty factor
Hg
microgram
|j,mol
micromoles
voc
volatile organic compound
11

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8-3-05
PROVISIONAL PEER REVIEWED TOXICITY VALUES FOR
N-METHYLANILINE (CASRN 100-61-8)
Derivation of Subchronic and Chronic Oral RfDs
Background
On December 5, 2003, the U.S. Environmental Protection Agency's (EPA's) Office of
Superfund Remediation and Technology Innovation (OSRTI) revised its hierarchy of human
health toxicity values for Superfund risk assessments, establishing the following three tiers as the
new hierarchy:
1.	EPA's Integrated Risk Information System (IRIS).
2.	Provisional Peer-Reviewed Toxicity Values (PPRTV) used in EPA's Superfund
Program.
3.	Other (peer-reviewed) toxicity values, including:
~	Minimal Risk Levels produced by the Agency for Toxic Substances and Disease
Registry (ATSDR),
~	California Environmental Protection Agency (CalEPA) values, and
~	EPA Health Effects Assessment Summary Table (HEAST) values.
A PPRTV is defined as a toxicity value derived for use in the Superfund Program when
such a value is not available in EPA's Integrated Risk Information System (IRIS). PPRTVs are
developed according to a Standard Operating Procedure (SOP) and are derived after a review of
the relevant scientific literature using the same methods, sources of data, and Agency guidance
for value derivation generally used by the EPA IRIS Program. All provisional toxicity values
receive internal review by two EPA scientists and external peer review by three independently
selected scientific experts. PPRTVs differ from IRIS values in that PPRTVs do not receive the
multi-program consensus review provided for IRIS values. This is because IRIS values are
generally intended to be used in all EPA programs, while PPRTVs are developed specifically for
the Superfund Program.
Because science and available information evolve, PPRTVs are initially derived with a
three-year life-cycle. However, EPA Regions (or the EPA HQ Superfund Program) sometimes
request that a frequently used PPRTV be reassessed. Once an IRIS value for a specific chemical
becomes available for Agency review, the analogous PPRTV for that same chemical is retired. It
should also be noted that some PPRTV manuscripts conclude that a PPRTV cannot be derived
based on inadequate data.
1

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8-3-05
Disclaimers
Users of this document should first check to see if any IRIS values exist for the chemical
of concern before proceeding to use a PPRTV. If no IRIS value is available, staff in the regional
Superfund and RCRA program offices are advised to carefully review the information provided
in this document to ensure that the PPRTVs used are appropriate for the types of exposures and
circumstances at the Superfund site or RCRA facility in question. PPRTVs are periodically
updated; therefore, users should ensure that the values contained in the PPRTV are current at the
time of use.
It is important to remember that a provisional value alone tells very little about the
adverse effects of a chemical or the quality of evidence on which the value is based. Therefore,
users are strongly encouraged to read the entire PPRTV manuscript and understand the strengths
and limitations of the derived provisional values. PPRTVs are developed by the EPA Office of
Research and Development's National Center for Environmental Assessment, Superfund Health
Risk Technical Support Center for OSRTI. Other EPA programs or external parties who may
choose of their own initiative to use these PPRTVs are advised that Superfund resources will not
generally be used to respond to challenges of PPRTVs used in a context outside of the Superfund
Program.
Questions Regarding PPRTVs
Questions regarding the contents of the PPRTVs and their appropriate use (e.g., on
chemicals not covered, or whether chemicals have pending IRIS toxicity values) may be directed
to the EPA Office of Research and Development's National Center for Environmental
Assessment, Superfund Health Risk Technical Support Center (513-569-7300), or OSRTI.
INTRODUCTION
An RfD for N-methylaniline is not listed on IRIS (U.S. EPA, 2005), the HEAST (U.S.
EPA, 1997), or the Drinking Water Standards and Health Advisories list (U.S. EPA, 2002). The
CARA list (U.S. EPA, 1991a, 1994) does not include any documents for N-methylaniline.
ATSDR (2003), IARC (2003) and WHO (2003) have not published toxicological reviews of this
compound. A toxicity review on aromatic amines (Weisburger and Hudson, 2001) and the NTP
(2003a,b) management status and health and safety reports were consulted for relevant
information. Literature searches were conducted in TOXLINE (1965-1992), CANCERLINE
(1963-1992), CHEM ID, HSDB, RTECS and TSCATS in May, 1992. Update computer
literature searches were conducted in TOXLINE (1992-1994), MEDLINE (1992-1994),
CANCERLINE (1992-1994), TSCATS and RTECS in March, 1994. Finally, update literature
searches were screened for the period from 1994 to January, 2003 in the following databases:
2

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8-3-05
TOXLINE (including NTIS and BIOSIS updates), CANCERLIT, MEDLINE, CCRIS,
GENETOX, HSDB, EMIC/EMICBACK, DART/ETICBACK, RTECS and TSCATS.
Additional literature searches from January 2003 through October 2004 were conducted by
NCEA-Cincinnati using MEDLINE, TOXLINE, Chemical and Biological Abstracts databases
REVIEW OF PERTINENT LITERATURE
Human Studies
No relevant data were located regarding the toxicity of N-methylaniline to humans
following oral exposure.
Animal Studies
Oral toxicity studies for N-methylaniline in animals include two carcinogenicity assays
and one 28-day gavage assay in rats. Chronic studies (Sander, 1971; Greenblatt et al., 1971) had
several experimental uncertainties and provided no information useful for deriving a p-RfD for
N -methy laniline.
Sander (1971) administered N-methylaniline in feed to 16 female rats (strain SIV 50) at a
dietary concentration of 0.09% for 114 days; using U.S. EPA (1988) reference values for body
weight and food consumption, the dose is calculated as 102 mg/kg-day. Control rats (n=36)
received standard diets. Both groups were maintained on standard diets for an additional 669
days. All rats were necropsied and suspected neoplastic tissues were examined histologically.
Treatment with N-methylaniline had no adverse effect on survival or tumor incidence. No
information was provided about non-neoplastic effects in tissues.
In a preliminary study for a pulmonary adenoma assay, Greenblatt et al. (1971)
administered N-methylaniline at a dietary concentration of 7.8 g per kg food to Swiss mice.
Using reference values for body weight and food consumption in U.S. EPA (1988), the dose is
calculated as 1457 mg/kg-day. In the main study, N-methylaniline was administered at a dietary
concentration of 1.95 g per kg food to Swiss mice (20/sex) for 28 weeks, after which the mice
received control diets for 12 weeks. As above, the dose in the main study is calculated as
approximately 364 mg/kg-day for male and female mice. Controls received standard diets for 40
weeks. Aside from a comment that the high dose of 1457 mg/kg-day in the preliminary study
produced severe cyanosis, presumably due to methemoglobinemia, no information was provided
about non-neoplastic effects in this study.
A description of a 4-week gavage assay conducted in Japan was available as a brief
summary in English (GINC, 2003); the more complete report in Japanese includes data tables in
3

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8-3-05
English (Biosafety Research Center, undated). In addition, the complete report of the Japanese
study was translated in English and evaluated by the NCEA-Cincinnati scientists. In this
Japanese study, N-methylaniline at doses of 0, 5, 25 or 125 mg/kg-day was administered by
gavage in corn oil to groups of CD rats (5/sex) for 28 days. Hematology, urinalysis and clinical
chemistry data were collected before terminal sacrifice on day 29. At termination, all rats were
necropsied, and data were collected for body weights and organ weights (brain, liver, kidneys,
spleen, adrenals, and testes or ovaries). The liver, kidney, spleen, bone marrow, adrenal and
femur were evaluated for histopathology.
Treatment with N-methylaniline had no adverse effect on survival or body weights.
Dose-related changes in hematology, serum chemistry and organ weights are shown in Table 1.
Alterations in indicators of erythrocyte toxicity (reduced hematocrit, hemoglobin and erythrocyte
counts) and compensatory hematopoiesis (elevated reticulocyte counts) were observed in both
sexes at >25 mg/kg-day in a dose-related fashion; in addition, hemoglobin was significantly
reduced in females treated at 5 mg/kg-day. Although methemoglobin levels were not reported in
the Japanese study (Biosafety Research Center, undated), evidence from the inhalation study
(Markosyan, 1969), as well as chronic oral toxicity data on the related compound aniline (CUT,
1982), implicate methemoglobin production as the primary effect of N-methylaniline exposure.
Statistically significant (p<0.05) serum chemistry changes included increased total bilirubin in
both sexes at 125 mg/kg-day and slight increases in AST in males at 125 mg/kg-day and
creatinine in females at >5 mg/kg-day. No significant urinalysis parameters were identified,
except that urine volume was higher and urine color was darker in the groups treated at 125
mg/kg-day. Absolute and relative spleen weights were significantly elevated in males at 125
mg/kg-day and females at >25 mg/kg-day. Significant gross necropsy findings included
splenomegaly (all males at >25 mg/kg-day; 3/5 females at 25 mg/kg-day and 5/5 females at 125
mg/kg-day) and black coloration (presumably hemosiderin, possibly methemoglobin, deposition)
of the spleen (all rats at >25 mg/kg-day), liver (all rats at 125 mg/kg-day), and kidney (4/5 males
and 5/5 females at 125 mg/kg-day). No gross findings were observed in the controls.
Histopathological changes were observed in the spleen (congestion in all males at >5 mg/kg-day
and all females at >25 mg/kg-day; pigment deposition and increased hematopoiesis in all rats at
>25 mg/kg-day), bone marrow (increased hematopoiesis in 4/5 females at 25 mg/kg-day and all
males and females at 125 mg/kg-day), liver (pigment deposition and extramedullary
hematopoiesis in all rats at 125 mg/kg-day), and kidney (pigment deposition in all rats at 125
mg/kg-day). These changes were all considered to be of slight severity, even at the highest dose
group, and, with the exception of pigment deposition in the spleen of 2/5 females, were not seen
in controls at all. In addition to these lesions, male rats showed a dose-related increase in
incidence and severity of hyaline droplet formation in the kidney. This lesion, however, is
characteristic of a male-rat-specific nephropathy associated with the presence of a low molecular
weight protein, a2[1-globulin, in male rats, and is not considered to be predictive of a renal effect
in humans (U.S. EPA, 1991b).
4

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8-3-05
Table 1. Dose-Related Changes in Rats Exposed to N-Methylaniline by Gavage for 28 Days:
A) Hematology, B) Serum Chemistry, C) Organ Weight (Japanese Study, undated; GINC, 2003)

Male rats
Female rats
Dose mg/kg-day
Control
5
25
125
Control
5
25
125
A) Hematology

Hematocrit (%)
45.3
44.2
40.7*
33.7**
44.6
43.3
39.9**
35.6**
Hemoglobin (g/dL)
15.0
14.6
13.2*
11.8**
14.5
13.9*
12.7**
12.3**
Erythrocytes (x 106/mm3)
7.30
7.10
6.37*
4.37**
7.02
7.12
6.20**
4.56**
Reticulocytes (%)
43
58
113**
943**
26
36
125*
639**
B) Serum chemistry

AST (U/L)
54
50
46
75 **
59
55
71
67
Creatinine (mg/dL)
0.52
0.51
0.50
0.53
0.52
0.68**
0.66**
0.71**
Total bilirubin (mg/dL)
0.11
0.12
0.16
0.77**
0.14
0.15
0.20
0.44**
C) Organ weight

Spleen absolute weight (g)
0.54
0.56
0.84
3.37**
0.36
0.39
0.67*
1.60**
Spleen relative weight (%)
0.193
0.172
0.261
1.088**
0.183
0.203
0.320*
0.804**
Significant difference from control group: * P<0.05; ** P<0.01
Although some minor effects were noted at 5 mg/kg-day (Biosafety Research Center,
undated), the weight of evidence of the hematology, organ weight, and pathology data suggests
that adverse effects occurred at 25 mg/kg-day and above. Hemoglobin was slightly reduced in
females at 5 mg/kg-day, but without concommitant changes in hematocrit and erythrocyte count.
By contrast, all of these measures were significantly reduced in both males and females at 25
mg/kg-day. Creatinine levels were elevated in all dosed female rats. Slight splenic congestion
was observed in males at 5 mg/kg-day, but other lesions, more clearly associated with the
hematological effects of the chemical (splenomegaly; pigment deposition and increased
hematopoiesis in the spleen, bone marrow, liver and kidney) were increased only at 25 mg/kg-
day and above. Therefore, this study identified a LOAEL of 25 mg/kg-day and NOAEL of 5
mg/kg-day for splenic and hematological effects.
Other Studies
Systemic effects of oral exposure to N-methylaniline are similar to those following
inhalation exposure. Rats continuously exposed by inhalation at a concentration of 0.3 mg/m3 for
5

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8-3-05
100 days exhibited increased methemoglobin levels, decreased numbers of erythrocytes, and
secondary effects in the spleen (hematopoiesis, splenomegaly) (Markosyan, 1969).
DERIVATION OF PROVISIONAL SUBCHRONIC AND
CHRONIC RfDs FOR N-METHYLANILINE
Based on the available data for N-methylaniline, the primary targets of toxicity appear to
be the erythrocyte and the spleen (Biosafety Research Center, undated; GINC, 2003). Although
methemoglobin levels were not measured in this study, evidence from the subchronic inhalation
test on N-methylaniline (Markosyan, 1969), as well as chronic oral toxicity data on the related
compound aniline (CUT, 1982), implicate methemoglobin production as the primary effect of N-
methylaniline exposure. In rats exposed to N-methylaniline by gavage for 28 days, minimal
effects on erythrocytes (significantly reduced hemoglobin in females) and slight congestion of the
spleen in males were observed at the NOAEL of 5 mg/kg-day. At the LOAEL of 25 mg/kg-day,
significant evidence of anemia (reduced hematocrit, hemoglobin and erythrocyte counts), as well
as compensatory hematopoiesis was observed in both sexes. Pigment deposition in several
tissues at 125 mg/kg-day was presumably related to erythrocyte destruction.
The NOAEL of 5 mg/kg-day for hematological and splenic effects in rats identified in the
study by the Biosafety Research Center (undated; GINC, 2003) was selected as the basis for the
subchronic RfD for N-methylaniline. An uncertainty factor of 300 was derived, consisting of
factors of 10 for extrapolation from rats to humans, 10 to protect sensitive individuals, and 3 for
deficiencies in the database, including absence of supporting chronic or subchronic studies and
any reproduction or developmental studies). Application of the composite uncertainty factor of
300 to the NOAEL of 5 mg/kg-day yields a provisional subchronic RfD of 2E-2 mg/kg-day for
N-methy laniline.
subchronic p-RfD = NOAEL / UF
= 5 mg/kg-day / 300
= 0.02 mg/kg-day or 2E-2 mg/kg-day
The chronic p-RfD is similarly derived by applying a composite uncertainty factor of
3000 to the NOAEL of 5 mg/kg-day. The composite uncertainty factor includes a factor of 300,
as derived above for the subchronic p-RfD, and an additional factor of 10 for use of a subchronic
study. Application of the uncertainty factor of 3000 to the NOAEL of 5 mg/kg-day yields a
provisional chronic RfD of 2E-3 mg/kg-day for N-methylaniline.
p-RfD = NOAEL / UF
= 5 mg/kg-day / 3000
= 0.002 mg/kg-day or 2E-3 mg/kg-day
6

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8-3-05
Confidence in the critical study is low. The study included investigation of a broad array
of systemic endpoints in multiple dose groups and identified a NOAEL and LOAEL. Tabular
results were well reported. The study was limited by small group sizes, short duration, and
failure to analyze for methemoglobin. Confidence in the database is low because of the lack of
supporting systemic toxicity data and developmental and reproductive toxicity studies.
Confidence in the subchronic and chronic p-RfDs for N-methylaniline is, therefore, low.
REFERENCES
ATSDR (Agency for Toxic Substances and Disease Registry). 2003. Toxicological Profile
Information Sheet. Online, http://www.atsdr.cdc.gov/toxpro2.html
Biosafety Research Center. Undated. Twenty-eight-day repeat dose oral toxicity test ofN-
Methylaniline in rats. (Jpn.; Eng. data tables). Online.
http://wwwdb.mhlw.go.ip/ginc/dbfilel/paper/paperl00-61-8B.html
CUT (Chemical Industry Institute of Toxicology). 1982. 104-Week chronic toxicity study in
rats. Aniline hydrochloride. Prepared for CUT, Research Triangle Park, NC by Hazleton
Laboratories America, Inc, Vienna, VA. Submitted to U.S. EPA under TSCA Section 8D. EPA
Doc. No. 878212078. Fiche No. OTS 205944. TSCATS 16580.
GINC (Global Information Network on Chemicals). 2003. N-Methylaniline. CAS No. 100-61-
8. Summary of Toxicity Data. (Eng.). Chemical Toxicity Database. Ministry of Health, Labour
and Welfare, Japan. Online, http://wwwdb.mhlw.go.ip/ginc/dbfilel/file/filel00-61-8.html
Greenblatt, M., S. Mirvish and B.T. So. 1971. Nitrosamine studies: induction of lung adenomas
by concurrent administration of sodium nitrite and secondary amines in Swiss mice. J. Natl.
Cancer Inst. 46: 1029-1034.
IARC (International Agency for Research on Cancer). 2003. IARC Agents and Summary
Evaluations. Online, http://www-cie.iarc.fr/htdig/search.html
Markosyan, T.M. 1969. On the comparative toxicity of monomethylaniline and dimethylaniline
in chronic experiments. Hyg. Sanit. 34: 328-332.
Merck. 1989. Merck Index. 11th edition. S. Budavari, M.J. O'Neil, A. Smith and P.E.
Heckelman, Ed. Rahway, NJ. p. 5944.
http://ntp
gov/htdocs/CHEM
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NTP (National Toxicology Program). 2003b. N-Methylaniline. Management Status Report.
Online. http://ntp-server.niehs.nih.gov/htdocs/Results Status/Resstatm/11419-J.Html
Sander, J. 1971. Untersuchungen iiber die Entstehung cancerogener Nitrosoverbindungen im
Magen von Versuchstieren und ihre Bedeutung fur den Menschen. [Studies on the origin of
carcinogenic nitrosylation in the stomach of laboratory animals and its implication for man.]
Arzneimittel Forschung. 21: 1572-1580.
U.S. EPA. 1988. Recommendations for and Documentation of Biological Values for Use in
Risk Assessment. Office of Health and Environmental Assessment, Environmental Criteria and
Assessment Office, Cincinnati, OH. PB88-17874. EPA/600/6-87/008.
U.S. EPA. 1991a. Chemical Assessments and Related Activities (CARA). Office of Health and
Environmental Assessment, Washington, DC. April.
U.S. EPA. 1991b. Alpha 2li-Globulin: Association with Chemically Induced Renal Toxicity and
Neoplasia in the Male Rat. Risk Assessment Forum, National Center for Environmental
Assessment, Office of Research and Development, Washington, DC. EPA/625/3-91/019F.
U.S. EPA. 1994. Chemical Assessments and Related Activities (CARA). Office of Health and
Environmental Assessment, Washington, DC. December.
U.S. EPA. 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. July.
EPA/540/R-97/036. NTIS 97-921199.
U.S. EPA. 2002. 2002 Edition of the Drinking Water Standards and Health Advisories. Office
of Water, Washington, DC. EPA 822-R-02-038.
http ://www.epa. gov/waterscience/drinking/ standards/dwstandards .pdf
U.S. EPA. 2005. Integrated Risk Information System (IRIS). Office of Research and
Development, National Center for Environmental Assessment, Washington, DC. Online.
http://www.epa.gov/iris/
Weisburger, E.K. and V.W. Hudson. 2001. Aromatic nitro and amino compounds. In: Patty's
Toxicology, Volume 4, 5th ed., E. Bingham, B. Cohrssen and C.H. Powell, Ed. John Wiley and
Sons, New York. p. 817-968.
WHO (World Health Organization). 2002. Online Catalogs for the Environmental Criteria
Series. Online, http://www.who.int/dsa/cat98/zehc.htm
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10-04-2005
Provisional Peer Reviewed Toxicity Values for
N-Methylaniline
(CASRN 100-61-8)
Derivation of Subchronic and Chronic Inhalation RfCs
Superfund Health Risk Technical Support Center
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268

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Acronyms and Abbreviations
bw	body weight
cc	cubic centimeters
CD	Caesarean Delivered
CERCLA	Comprehensive Environmental Response, Compensation and Liability Act
of 1980
CNS	central nervous system
cu.m	cubic meter
DWEL	Drinking Water Equivalent Level
FEL	frank-effect level
FIFRA	Federal Insecticide, Fungicide, and Rodenticide Act
g	grams
GI	gastrointestinal
HEC	human equivalent concentration
Hgb	hemoglobin
i.m.	intramuscular
i.p.	intraperitoneal
i.v.	intravenous
IRIS	Integrated Risk Information System
IUR	inhalation unit risk
kg	kilogram
L	liter
LEL	lowest-effect level
LOAEL	lowest-observed-adverse-effect level
LOAEL(ADJ)	LOAEL adjusted to continuous exposure duration
LOAEL(HEC)	LOAEL adjusted for dosimetric differences across species to a human
m	meter
MCL	maximum contaminant level
MCLG	maximum contaminant level goal
MF	modifying factor
mg	milligram
mg/kg	milligrams per kilogram
mg/L	milligrams per liter
MRL	minimal risk level
1

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MTD
maximum tolerated dose
MTL
median threshold limit
NAAQS
National Ambient Air Quality Standards
NOAEL
no-observed-adverse-effect level
NOAEL(ADJ)
NOAEL adjusted to continuous exposure duration
NOAEL(HEC)
NOAEL adjusted for dosimetric differences across species to a human
NOEL
no-observed-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
PBPK
physiologically based pharmacokinetic
PPb
parts per billion
ppm
parts per million
PPRTV
Provisional Peer Reviewed Toxicity Value
RBC
red blood cell(s)
RCRA
Resource Conservation and Recovery Act
RDDR
Regional deposited dose ratio (for the indicated lung region)
REL
relative exposure level
RfC
inhalation reference concentration
RfD
oral reference dose
RGDR
Regional gas dose ratio (for the indicated lung region)
s.c.
subcutaneous
SCE
sister chromatid exchange
SDWA
Safe Drinking Water Act
sq.cm.
square centimeters
TSCA
Toxic Substances Control Act
UF
uncertainty factor
Hg
microgram
|imol
micromoles
VOC
volatile organic compound
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10-04-2005
PROVISIONAL PEER REVIEWED TOXICITY VALUES FOR
N-METHYLANILINE (CASRN 100-61-8)
Derivation of Subchronic and Chronic Inhalation RfCs
Background
On December 5, 2003, the U.S. Environmental Protection Agency's (EPA's) Office of
Superfund Remediation and Technology Innovation (OSRTI) revised its hierarchy of human
health toxicity values for Superfund risk assessments, establishing the following three tiers as the
new hierarchy:
1.	EPA's Integrated Risk Information System (IRIS).
2.	Provisional Peer-Reviewed Toxicity Values (PPRTV) used in EPA's Superfund
Program.
3.	Other (peer-reviewed) toxicity values, including:
~	Minimal Risk Levels produced by the Agency for Toxic Substances and Disease
Registry (ATSDR),
~	California Environmental Protection Agency (CalEPA) values, and
~	EPA Health Effects Assessment Summary Table (HEAST) values.
A PPRTV is defined as a toxicity value derived for use in the Superfund Program when
such a value is not available in EPA's Integrated Risk Information System (IRIS). PPRTVs are
developed according to a Standard Operating Procedure (SOP) and are derived after a review of
the relevant scientific literature using the same methods, sources of data, and Agency guidance
for value derivation generally used by the EPA IRIS Program. All provisional toxicity values
receive internal review by two EPA scientists and external peer review by three independently
selected scientific experts. PPRTVs differ from IRIS values in that PPRTVs do not receive the
multi-program consensus review provided for IRIS values. This is because IRIS values are
generally intended to be used in all EPA programs, while PPRTVs are developed specifically for
the Superfund Program.
Because science and available information evolve, PPRTVs are initially derived with a
three-year life-cycle. However, EPA Regions or the EPA Headquarters Superfund Program
sometimes request that a frequently used PPRTV be reassessed. Once an IRIS value for a
specific chemical becomes available for Agency review, the analogous PPRTV for that same
chemical is retired. It should also be noted that some PPRTV manuscripts conclude that a
PPRTV cannot be derived based on inadequate data.
1

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10-04-2005
Disclaimers
Users of this document should first check to see if any IRIS values exist for the chemical
of concern before proceeding to use a PPRTV. If no IRIS value is available, staff in the regional
Superfund and RCRA program offices are advised to carefully review the information provided
in this document to ensure that the PPRTVs used are appropriate for the types of exposures and
circumstances at the Superfund site or RCRA facility in question. PPRTVs are periodically
updated; therefore, users should ensure that the values contained in the PPRTV are current at the
time of use.
It is important to remember that a provisional value alone tells very little about the
adverse effects of a chemical or the quality of evidence on which the value is based. Therefore,
users are strongly encouraged to read the entire PPRTV manuscript and understand the strengths
and limitations of the derived provisional values. PPRTVs are developed by the EPA Office of
Research and Development's National Center for Environmental Assessment, Superfund Health
Risk Technical Support Center for OSRTI. Other EPA programs or external parties who may
choose of their own initiative to use these PPRTVs are advised that Superfund resources will not
generally be used to respond to challenges of PPRTVs used in a context outside of the Superfund
Program.
Questions Regarding PPRTVs
Questions regarding the contents of the PPRTVs and their appropriate use (e.g., on
chemicals not covered, or whether chemicals have pending IRIS toxicity values) may be directed
to the EPA Office of Research and Development's National Center for Environmental
Assessment, Superfund Health Risk Technical Support Center (513-569-7300), or OSRTI.
INTRODUCTION
An RfC for N-methylaniline is not listed on IRIS (U.S. EPA, 2005) or the HEAST (U.S.
EPA, 1997). ACGIH (2001, 2002) andNIOSH (2003) established occupational exposure limits
(8-hour TWA) of 0.5 ppm (2.2 mg/m3) for N-methylaniline to protect against
methemoglobinemia and its sequelae (anoxia, cyanosis, dizziness, weakness and headache).
OSHA (2003a,b) had proposed an identical PEL in its final rule of 1989, but after this rule was
revoked in 1993, the PEL reverted to its former value of 2 ppm (9 mg/m3). All three agency limit
values include a skin notation to indicate that dermal absorption of N-methylaniline can
contribute significantly to systemic toxicity. The CARA list (U.S. EPA, 1991, 1994) does not
include any documents for N-methylaniline. ATSDR (2003), IARC (2003) and WHO (2003)
have not published toxicological reviews of this compound. A toxicity review on aromatic
amines (Weisburger and Hudson, 2001) and the NTP (2003a,b) management status and health
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10-04-2005
and safety reports were consulted for relevant information. Literature searches were conducted in
TOXLINE (1965-1992), CANCERLINE (1963-1992), CHEM ID, HSDB, RTECS and TSCATS
in May, 1992. Updated computer literature searches were also conducted in TOXLINE (1992-
1994), MEDLINE (1992-1994), CANCERLINE (1992-1994), TSCATS and RTECS in March,
1994. Additional literature searches were screened for the period from 1994 to September 2005.
REVIEW OF PERTINENT LITERATURE
Human Studies
No relevant data were located regarding the toxicity N-methylaniline to humans
following inhalation exposure.
Animal Studies
No chronic inhalation studies in animals were located for N-methylaniline. The literature
search uncovered one subchronic study in rats.
In the subchronic inhalation study, rats were exposed to 0.04 or 0.3 mg/m3 of N-
methylaniline or to 0.005 or 0.3 mg/m3 of the related compound N,N-dimethylaniline
continuously for 100 days (Markosyan, 1969). No information on the number or strain of
animals, treatment of controls, purity of the compounds, generation of the exposure atmosphere
or monitoring of the exposure level was provided. Although statistical significance is mentioned
in the report, no information on statistical methods was provided. For both compounds, effects
at the highest exposure level included increased methemoglobin levels, reduced erythrocyte
counts and hemoglobin levels, reticulocytosis, leukopenia, splenomegaly and splenic
hemosiderosis. For the most part, no documentation was provided for these effects. A number
of other changes in serum or tissue constituents were also reported. Some histological changes
were reported in the brain, but these could reflect autolysis during the interval prior to fixation.
Histological changes in the livers and lungs were also reported. No effects were seen at the
lower exposure levels. Given the lack of information regarding experimental methods and
results, and the lack of corroborating data from the literature, this study is not a suitable basis for
deriving an RfC. Single dose exposure studies conducted in dogs, cats, rabbits, rats and guinea
pigs indicated a range of exposure (2.3 to 7.6 ppm) considered safe for exposure to
N-methylaniline in most sensitive species (ACGIH, 2001). However, methemoglobinemia and
formation of Heinz bodies have been reported at 7.6 ppm and 2.4 ppm, respectively (U.S. EPA,
2005).
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Other Studies
No adequate compound-specific data are available for the inhalation toxicity of
N-methylaniline to derive a p-RfC.
The conversion of hemoglobin to methemoglobin is one of the characteristic effects
caused by metabolites of aromatic amines (Weisburger and Hudson, 2001). N-methylaniline
(U.S. EPA, 2005) has been shown to induce methemoglobin formation in erythrocytes of
exposed animals (Markosyan,1969; Oberst et al., 1956). The sequelae of methemoglobin
formation are: erythrocyte effects (reduced hemoglobin, hematocrit and erythrocyte counts),
cyanosis from the reduced oxygen-carrying capacity of the blood, and secondary splenic effects
(hemosiderosis, hematopoiesis, splenomegaly) (Markosyan, 1969; Greenblatt et al., 1971; GINC,
2003; Biosafety Research Center, undated a; U.S. EPA, 1992, 2005). In a 4-week gavage study
in rats exposed to N-methylaniline, the NOAEL was 5 mg/kg-day and the LOAEL was 25
mg/kg-day for blood and spleen effects (GINC, 2003; Biosafety Research Center, undated b).
Several different phase I metabolic pathways in the liver have been identified for aniline
(U.S. EPA, 1992): N-acetylation, aromatic hydroxylation and N-hydroxylation. The acetylation
of aniline by N-acetyltransferase results in the formation of acetanilide, which is further
metabolized to products excreted in urine. In organisms with efficient acetylation processes
(mice and humans with a 'fast acetylator' phenotype), a smaller percentage of an administered
dose of aniline is metabolized by the other two pathways, which are known to generate reactive
intermediates (Weisburger and Hudson, 2001). The most toxicologically significant pathway is
N-hydroxylation of aniline by cytochrome P450 (CYP450) mixed function oxidases to form
phenylhydroxylamine. In erythrocytes, the metabolism of phenylhydroxylamine to
nitrosobenzene results in the conversion of hemoglobin to methemoglobin (U.S. EPA, 1992).
CYP450 enzymes also catalyze ring hydroxylation of aniline to aminophenols, which may be
subsequently conjugated to form glucuronide or sulfate metabolites typically excreted in urine.
The metabolism of N-methylaniline is expected to be similar to aniline following an N-
demethylation reaction, which is a function of heme-containing enzymes such as CYP450 and
peroxidases (Hover and Kalkami, 2000). N-demethylation of N-methylaniline has been
demonstrated in erythrocytes (Stecca et al., 1992). N-glucuronidation has been reported in
primary cultures of rat hepatocytes treated with N-methylaniline (Sherrat and Damani, 1989). In
the acidic conditions of the stomach, N-methylaniline reacts with nitrite to form N-nitroso
derivatives (Greenblatt et al., 1971; Sander, 1973). However, because of considerable difference
in solubility in water, the metabolic pathway for N-methylaniline may not be similar to aniline in
assessing critical effects via inhalation route of exposure.
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DERIVING PROVISIONAL SUBCHRONIC AND CHRONIC
RFCs FOR N-METHYLANILINE
Inhalation data for N-methylaniline are limited to one study (Markosyan, 1969), which
has several uncertainties in study design, data collection and data analysis; information pertinent
to mode of action and route-specific toxicodynamics are not available. Lack of route-specific
pharmacokinetic/pharmacodynamic information precludes derivation of either subchronic or
chronic p-RfCs. The Markosyan (1969) study did not provide adequate dose-response data on
methemoglobinemia. Therefore, additional studies are needed in support of this critical effect.
REFERENCES
ACGIH (American Conference of Governmental Industrial Hygienists). 2001. N-Methyl
Aniline. CAS Number 100-61-8. Documentation of the Threshold Limit Values and Biological
Exposure Indices, Sixth ed. Cincinnati, OH.
ACGIH (American Conference of Governmental Industrial Hygienists). 2002. 2002 Threshold
Limit Values for Chemical Substances and Physical Agents and Biological Exposure Indices.
Cincinnati, OH. p. 39.
ATSDR (Agency for Toxic Substances and Disease Registry). 2003. Toxicological Profile
Information Sheet. Online, http://www.atsdr.cdc.gov/toxpro2.html
Biosafety Research Center. Undated a. Single dose toxicity test of N-methylaniline in rats.
(Jpn.; Eng. data tables). Online.
http://wwwdb.mhlw. go ,jp/ginc/ dbfile 1 /paper/paper 100-61 -8A.html
Biosafety Research Center. Undated b. Twenty-eight-day repeat dose oral toxicity test of N-
methylaniline in rats. (Jpn.; Eng. data tables). Online.
http://wwwdb.mhlw.go.ip/ginc/dbfilel/paper/paperl00-61-8B.html
CUT (Chemical Industry Institute of Toxicology). 1982. 104-Week Chronic Toxicity Study in
Rats. Aniline Hydrochloride. Prepared for CUT, Research Triangle Park, NC by Hazleton
Laboratories America, Inc, Vienna, VA. Submitted to U.S. EPA under TSCA Section 8D. EPA
Doc. No. 878212078. Fiche No. OTS 205944. TSCATS 16580.
duPont (du Pont deNemours and Company, Inc). 1982. Subacute inhalation toxicity study of
aniline in rats. OTS No. 878220240. Fiche No. 0215025.
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10-04-2005
GINC (Global Information Network on Chemicals). 2003. N-Methylaniline. CAS No. 100-61-
8. Summary of Toxicity Data. (Eng.). Chemical Toxicity Database. Ministry of Health,
Labour and Welfare, Japan. Online, http://wwwdb.mhlw.go.ip/ginc/dbfilel/file/filel00-61-
8.html.
Greenblatt, M., S. Mirvish and B.T. So. 1971. Nitrosamine studies: induction of lung adenomas
by concurrent administration of sodium nitrite and secondary amines in Swiss mice. J. Natl.
Cancer Inst. 46:1029-1034.
Hover, C.G. and A.P. Kalkami. 2000. Lipoxygenase-mediated hydrogen peroxide-dependent N-
demethylation of N,N-dimethylaniline and related compounds. Chem. Biol. Interact. 124: 191-
203.
IARC (International Agency for Research on Cancer). 2003. IARC Agents and Summary
Evaluations. Online, http://www-cie.iarc.fr/htdig/search.html
Markosyan, T.M. 1969. On the comparative toxicity of monomethylaniline and dimethylaniline
in chronic experiments. Hyg. Sanit. 34: 328-332.
Merck. 1989. Merck Index. 11th edition. S. Budavari, M.J. O'Neil, A. Smith and P.E.
Heckelman, Ed. Rahway, NJ. p. 5944.
NIOSH (National Institute for Occupational Safety and Health). 2003. Monomethyl Aniline.
CAS 100-61-8. NIOSH Pocket Guide to Chemical Hazards. Online.
http://www.cdc.gov/niosh/npg/npgd0436.html
NTP (National Toxicology Program). 2003a. N-Methylaniline. Health and Safety Report.
Online. http://ntp-server.niehs.nih.gov/htdocs/CHEM H&S/NTP Cheml/Radianl00-61 -8.html
NTP (National Toxicology Program). 2003b. N-Methylaniline. Management Status Report.
Online. http://ntp-server.niehs.nih.gov/htdocs/Results Status/Resstatm/11419-J.Html
Oberst, F.W., E. Hackley and C. Comstock. 1956. Chronic toxicity of aniline vapor (5 ppm) by
inhalation. Arch. Ind. Health. 13: 379-384.
OSHA (Occupational Safety and Health Administration). 2002a. OSHA Standard 1910.1000
Table Z-l. Part Z, Toxic and Hazardous Substances. Online.
http://www.osha-slc.gov/OshStd data/1910 1000 TABLE Z-l.html
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10-04-2005
OSHA (Occupational Safety and Health Administration). 2002b. OSHA Standard 1915.1000
for Air Contaminants. Part Z, Toxic and Hazardous Substances. Online.
http://www.osha-slc.gov/OsliStd data/1915 1000.html
Sander, J. 1973. The formation of N-nitroso compounds in the stomach of animals and man in
the diet. IARC Sci. Pub. 4: 159-163.
Sherrat, A.J. and L.A. Damani. 1989. The metabolism of N,N-dimethylaniline by rat
hepatocytes: identification of a novel N-conjugate. Xenobiotica. 19: 379-388.
SRC (Syracuse Research Corporation). 2003. Interactive PhysProp Database Demo. Online.
http://esc.syrres.com/interkow/phvsdemo .htm
Stecca, C., J. Cumps and M. Duverger-Van Bogaert. 1992. Enzymic N-demethylation reaction
catalysed by red blood cell cytosol. Biochem. Pharmacol. 43: 207-211.
U.S. EPA. 1985. Health and Environmental Effects Profile for Aniline. Prepared by the Office
of Health and Environmental Assessment, Environmental Criteria and Assessment Office,
Cincinnati, OH for the Office of Solid Waste and Emergency Response, Washington, DC.
U.S. EPA. 1988. Recommendations for and Documentation of Biological Values for Use in
Risk Assessment. Office of Health and Environmental Assessment, Environmental Criteria and
Assessment Office, Cincinnati, OH. PB88-17874. EPA/600/6-87/008.
U.S. EPA. 1991. Chemical Assessments and Related Activities (CARA). Office of Health and
Environmental Assessment, Washington, DC. April.
U.S. EPA. 1992. Health and Environmental Effects Document for Aniline. Prepared by the
Office of Health and Environmental Assessment, Environmental Criteria and Assessment Office,
Cincinnati, OH for the Office of Solid Waste and Emergency Response, Washington, DC.
U.S. EPA. 1994. Chemical Assessments and Related Activities (CARA). Office of Health and
Environmental Assessment, Washington, DC. December.
U.S. EPA. 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. July.
EPA/540/R-97/036. NTIS 97-921199.
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U.S. EPA. 2005. Integrated Risk Information System (IRIS). Office of Research and
Development, National Center for Environmental Assessment, Washington, DC. Online.
http://www.epa. gov/ iris/
Weisburger, E.K. and V.W. Hudson. 2001. Aromatic nitro and amino compounds. In: Patty's
Toxicology, Volume 4, 5th ed., E. Bingham, B. Cohrssen and C.H. Powell, Ed. John Wiley and
Sons, New York. p. 817-968.
WHO (World Health Organization). 2002. Online Catalogs for the Environmental Criteria
Series. Online, http://www.who.int/dsa/cat98/zehc.htm
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Provisional Peer Reviewed Toxicity Values for
N-Methylaniline
(CASRN 100-61-8)
Derivation of a Carcinogenicity Assessment
Superfund Health Risk Technical Support Center
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268

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Acronyms and Abbreviations
bw	body weight
cc	cubic centimeters
CD	Caesarean Delivered
CERCLA	Comprehensive Environmental Response, Compensation and Liability Act
of 1980
CNS	central nervous system
cu.m	cubic meter
DWEL	Drinking Water Equivalent Level
FEL	frank-effect level
FIFRA	Federal Insecticide, Fungicide, and Rodenticide Act
g	grams
GI	gastrointestinal
HEC	human equivalent concentration
Hgb	hemoglobin
i.m.	intramuscular
i.p.	intraperitoneal
i.v.	intravenous
IRIS	Integrated Risk Information System
IUR	inhalation unit risk
kg	kilogram
L	liter
LEL	lowest-effect level
LOAEL	lowest-observed-adverse-effect level
LOAEL(ADJ)	LOAEL adjusted to continuous exposure duration
LOAEL(HEC)	LOAEL adjusted for dosimetric differences across species to a human
m	meter
MCL	maximum contaminant level
MCLG	maximum contaminant level goal
MF	modifying factor
mg	milligram
mg/kg	milligrams per kilogram
mg/L	milligrams per liter
MRL	minimal risk level
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MTD
maximum tolerated dose
MTL
median threshold limit
NAAQS
National Ambient Air Quality Standards
NOAEL
no-observed-adverse-effect level
NOAEL(ADJ)
NOAEL adjusted to continuous exposure duration
NOAEL(HEC)
NOAEL adjusted for dosimetric differences across species to a human
NOEL
no-observed-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
PBPK
physiologically based pharmacokinetic
PPb
parts per billion
ppm
parts per million
PPRTV
Provisional Peer Reviewed Toxicity Value
RBC
red blood cell(s)
RCRA
Resource Conservation and Recovery Act
RDDR
Regional deposited dose ratio (for the indicated lung region)
REL
relative exposure level
RfC
inhalation reference concentration
RfD
oral reference dose
RGDR
Regional gas dose ratio (for the indicated lung region)
s.c.
subcutaneous
SCE
sister chromatid exchange
SDWA
Safe Drinking Water Act
sq.cm.
square centimeters
TSCA
Toxic Substances Control Act
UF
uncertainty factor
Hg
microgram
|imol
micromoles
VOC
volatile organic compound
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PROVISIONAL PEER REVIEWED TOXICITY VALUES FOR
N-METHYLANILINE (CASRN 100-61-8)
Derivation of a Carcinogenicity Assessment
Background
On December 5, 2003, the U.S. Environmental Protection Agency's (EPA's) Office of
Superfund Remediation and Technology Innovation (OSRTI) revised its hierarchy of human
health toxicity values for Superfund risk assessments, establishing the following three tiers as the
new hierarchy:
1.	EPA's Integrated Risk Information System (IRIS).
2.	Provisional Peer-Reviewed Toxicity Values (PPRTV) used in EPA's Superfund
Program.
3.	Other (peer-reviewed) toxicity values, including:
~	Minimal Risk Levels produced by the Agency for Toxic Substances and Disease
Registry (ATSDR),
~	California Environmental Protection Agency (CalEPA) values, and
~	EPA Health Effects Assessment Summary Table (HEAST) values.
A PPRTV is defined as a toxicity value derived for use in the Superfund Program when
such a value is not available in EPA's Integrated Risk Information System (IRIS). PPRTVs are
developed according to a Standard Operating Procedure (SOP) and are derived after a review of
the relevant scientific literature using the same methods, sources of data, and Agency guidance
for value derivation generally used by the EPA IRIS Program. All provisional toxicity values
receive internal review by two EPA scientists and external peer review by three independently
selected scientific experts. PPRTVs differ from IRIS values in that PPRTVs do not receive the
multi-program consensus review provided for IRIS values. This is because IRIS values are
generally intended to be used in all EPA programs, while PPRTVs are developed specifically for
the Superfund Program.
Because science and available information evolve, PPRTVs are initially derived with a
three-year life-cycle. However, EPA Regions or the EPA Headquarters Superfund Program
sometimes request that a frequently used PPRTV be reassessed. Once an IRIS value for a
specific chemical becomes available for Agency review, the analogous PPRTV for that same
chemical is retired. It should also be noted that some PPRTV manuscripts conclude that a
PPRTV cannot be derived based on inadequate data.
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Disclaimers
Users of this document should first check to see if any IRIS values exist for the chemical
of concern before proceeding to use a PPRTV. If no IRIS value is available, staff in the regional
Superfund and RCRA program offices are advised to carefully review the information provided
in this document to ensure that the PPRTVs used are appropriate for the types of exposures and
circumstances at the Superfund site or RCRA facility in question. PPRTVs are periodically
updated; therefore, users should ensure that the values contained in the PPRTV are current at the
time of use.
It is important to remember that a provisional value alone tells very little about the
adverse effects of a chemical or the quality of evidence on which the value is based. Therefore,
users are strongly encouraged to read the entire PPRTV manuscript and understand the strengths
and limitations of the derived provisional values. PPRTVs are developed by the EPA Office of
Research and Development's National Center for Environmental Assessment, Superfund Health
Risk Technical Support Center for OSRTI. Other EPA programs or external parties who may
choose of their own initiative to use these PPRTVs are advised that Superfund resources will not
generally be used to respond to challenges of PPRTVs used in a context outside of the Superfund
Program.
Questions Regarding PPRTVs
Questions regarding the contents of the PPRTVs and their appropriate use (e.g., on
chemicals not covered, or whether chemicals have pending IRIS toxicity values) may be directed
to the EPA Office of Research and Development's National Center for Environmental
Assessment, Superfund Health Risk Technical Support Center (513-569-7300), or OSRTI.
INTRODUCTION
A cancer assessment for N-methylaniline is not listed on IRIS (U.S. EPA, 2005a), the
HE AST (U.S. EPA, 1997), or the Drinking Water Standards and Health Advisories list (U.S.
EPA, 2002). The CARA list (U.S. EPA, 1991, 1994) does not list any documents for N-
methylaniline. ATSDR (2003), IARC (2003) and WHO (2003) have not published toxicological
reviews of this compound. A toxicity review on aromatic amines (Weisburger and Hudson,
2001) and the NTP (2003a,b) management status and health and safety reports were consulted for
relevant information. Literature searches were conducted in TOXLINE (1965-1992),
CANCERLINE (1963-1992), CHEM ID, HSDB, RTECS and TSCATS in May, 1992. Updated
computer literature searches were also conducted in TOXLINE (1992-1994), MEDLINE (1992-
1994), CANCERLINE (1992-1994), TSCATS and RTECS in March, 1994. Additional literature
searches were screened for the period from 1994 to September 2005.
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REVIEW OF PERTINENT LITERATURE
Human Studies
No studies were located regarding the carcinogenicity of N-methylaniline to humans by
any route of exposure.
Animal Studies
Two oral carcinogenicity studies in rodents are available for N-methylaniline, but neither
is adequate by current standards. No inhalation carcinogenicity studies in animals were located.
Sander (1971) evaluated the carcinogenicity of N-methylaniline and related compounds in
rodents to evaluate the effect of nitrosamine formation. A group of 16 female rats (strain SIV 50)
were given 0.09% N-methylaniline in the diet for 114 days; using reference values for body
weight and food consumption in U.S. EPA (1988), the dose is calculated as 102 mg/kg-day. A
control group of 32 rats received a standard diet and untreated drinking water. Three other
groups received drinking water containing 1.0% sodium nitrite and diets with or without
N-methylaniline (0.015 or 0.03%) for 114-117 days; as above, the doses of N-methylaniline are
calculated as 17 or 24 mg/kg-day. All groups received standard diets and untreated drinking
water for the remaining study period. The study was terminated on day 783, by which time all
rats in the dually-treated groups had died. All rats were necropsied and suspect tissues were
examined histologically. Treatment with N-methylaniline alone or sodium nitrite alone had no
adverse effect on survival or the incidence of tumors. However, simultaneous administration of
sodium nitrite and N-methylaniline significantly increased mortality and the incidence of tumors,
notably in the esophagus (adenomas and carcinomas) and nasal cavity
(aesthesioneuroepithelioma and carcinoma). Carcinogenicity was attributed to the formation of
the N-nitroso derivative of N-methylaniline (Sander, 1973). This study is not an adequate test of
the carcinogenicity of N-methylaniline alone because the period of exposure was relatively short,
the group sizes were small, a single dose level was tested (without co-exposure to nitrite), and the
histological examination appears to have been limited to 'interesting' tissues.
Similar results were reported in a 40-week pulmonary adenoma assay by Greenblatt et al.
(1971). N-methylaniline at a dietary concentration of 1.95 g per kg was administered to Swiss
mice (20/sex) for 28 weeks, after which the mice received standard diets for 12 weeks. Using
reference values for food consumption and body weight in U.S. EPA (1988), the dose is
calculated as 364 mg/kg-day for male and female mice. A control group of 80/sex received
standard diets for 40 weeks. At termination, mice were necropsied and examined for lung
adenomas exceeding 0.5 mm; the diagnosis was confirmed microscopically in every fifth animal.
Treatment with N-methylaniline alone did not increase the incidence of pulmonary adenoma.
The incidence of malignant melanoma at 40 weeks was higher in the treated group compared to
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the control group (14% versus 7%). A third group that was given 1.95 g N-methylaniline per kg
food and drinking water containing 0.1% sodium nitrite for 28 weeks showed a significant
increase in the incidence of pulmonary adenoma; this result was attributed to the production of
nitrosamine. This study is not an adequate test of the carcinogenicity of N-methylaniline alone
because histopathology analysis was limited to the lung and because the group sizes were small,
a single dose level was tested (without co-exposure to nitrite), and the study duration was
intentionally truncated to facilitate the counting of individual lung tumors.
Other Studies
Whereas little information is available on the metabolism of N-methylaniline, several
different phase I metabolic pathways in the liver have been identified for aniline (U.S. EPA,
1992): N-acetylation, aromatic hydroxylation and N-hydroxylation. The acetylation of aniline by
N-acetyltransferase results in the formation of acetanilide, which is further metabolized to
products excreted in urine. In organisms with efficient acetylation processes (mice and humans
with a 'fast acetylator' phenotype), a smaller percentage of an administered dose of aniline is
metabolized by the other two pathways, which are known to generate reactive intermediates
(Weisburger and Hudson, 2001). The most toxicologically significant pathway is N-
hydroxylation of aniline by cytochrome P450 (CYP450) mixed function oxidases to form
phenylhydroxylamine. In erythrocytes, the metabolism of phenylhydroxylamine to
nitrosobenzene results in the conversion of hemoglobin to methemoglobin (U.S. EPA, 1992).
CYP450 enzymes also catalyze ring hydroxylation of aniline to aminophenols, which may be
subsequently undergo phase II conjugation reactions to form glucuronide or sulfate metabolites
typically excreted in urine. The metabolism of N-methylaniline is expected to be similar to
aniline following an N-demethylation reaction, which is a function of heme-containing enzymes
such as CYP450 and peroxidases (Hover and Kalkami, 2000). N-demethylation of
N-methylaniline has been demonstrated in erythrocytes (Stecca et al., 1992). N-glucuronidation
has been reported in primary cultures of rat hepatocytes treated with N-methylaniline (Sherrat
and Damani, 1989). In the acidic conditions of the stomach, N-methylaniline reacts with nitrite
to form N-nitroso derivatives (Greenblatt et al., 1971; Sander, 1973).
The acute oral toxicity of N-methylaniline in rats is similar to that of aniline. The acute
oral LD50 values for N-methylaniline were 782 and 716 mg/kg in males and females,
respectively (GINC, 2003; Biosafety Research Center, undated a). Acute oral LD50 values in
several tests for aniline ranged between 440 and 1072 mg/kg for the free base and 840 to 1070
for the hydrochloride salt (U.S. EPA, 1992; Health Canada, 1994).
A characteristic effect of the biotransformation of aromatic amines such as
N-methylaniline and aniline is the formation of methemoglobin, which can occur independent of
the route of exposure (Weisburger and Hudson, 2001). N-methylaniline and aniline have been
shown to induce methemoglobin formation in erythrocytes of exposed animals, resulting in
4

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hemolytic anemia and secondary splenic effects (hemosiderosis, hematopoiesis, splenomegaly)
(Markosyan, 1969; U.S. EPA, 2003a). Evidence for methemoglobinemia has also been provided
in some studies in which methemoglobin was not measured. In a preliminary test for the
pulmonary adenoma assay described above, rats receiving a dietary concentration of 7.8 g/kg per
kg (dose calculated as approximately 1457 mg/kg-day) developed cyanosis, which is a typical
effect of methemoglobinemia (Greenblatt et al., 1971). Erythrocyte effects (reduced hematocrit,
hemoglobin and erythrocyte counts) and splenic effects (increased organ weight, splenomegaly,
pigmentation and hematopoiesis) were observed in rats exposed to N-methylaniline by gavage
for 28 days (GINC, 2003; Biosafety Research Center, undated b).
In evaluating common features of carcinogenicity caused by aromatic amines, Goodman
et al. (1984) concluded that both genotoxic and non-genotoxic mechanisms were possible; both
mechanisms involve biotransformation by CYP450. The genotoxic mechanism relates to the
generation of reactive intermediates during metabolism and is presumably the cause of
carcinogenicity in the liver and other tissues. A non-genotoxic mechanism was proposed for rare
tumors of the spleen, in which accumulation of erythrocytes damaged from methemoglobin
formation would lead to splenic fibrosis, and subsequently, to splenic tumors. However, this
mechanism cannot rule out the possibility that reactive metabolites are transferred from the
damaged erythrocytes to the spleen, resulting in a secondary genotoxic effect.
Less genotoxicity information is available for N-methylaniline than for aniline, but the
results for the available comparable studies were similar for the two compounds.
N-Methylaniline was not mutagenic to Salmonella typhimurium strains TA97, TA98, TA100,
TA 1535 and TA 1537 or to Escherichia coli strain WP2uvrA with or without metabolic
activation (Zeiger et al., 1988; GINC, 2003; Hatano Research Institute, undated a). Similarly,
aniline was not mutagenic in bacteria under standard test conditions (U.S. EPA, 1992, 2003a;
Health Canada, 1994). Both compounds yielded negative results for DNA repair in primary
cultured rat hepatocytes when tested at 10"6 to 10"3 M (Yoshimi et al., 1988).
N-Methylaniline induced chromosomal aberrations in cultured Chinese hamster lung
(CHL/IU) cells after 6 hours of exposure with metabolic activation or 24 hours of exposure
without metabolic activation (GINC, 2003; Hatano Research Institute, undated b). Aniline did
not induce chromosomal aberrations in a different strain of Chinese hamster lung (Don) cells
(U.S. EPA, 1992). In other studies, aniline tested positive for genotoxic effects, increasing the
frequency of sister chromatid exchanges in human and hamster cells, increasing DNA damage in
cultured mouse lymphoma cells, and transforming mouse Balb/3T3 cells (U.S. EPA, 1992). In in
vivo studies, aniline increased the frequencies of micronucleus formation in bone marrow of rats
and mice, and sister chromatid exchanges in bone marrow of mice, but not rats (U.S. EPA,
1992). Because of these uncertainties in mode of action and lack of adequate toxicokinetic
information, a non-genotoxic mode of action may not be appropriate for N-methylaniline.
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PROVISIONAL WEIGHT-OF-EVIDENCE CLASSIFICATION
No human data are available for the carcinogenicity of N-methylaniline and the animal
data are not adequate because the study designs do not meet current standards (Sander, 1971;
Greenblatt et al, 1971): group sizes were too small, exposure durations were insufficient, too few
dose levels were administered, and the range of tissues evaluated for histopathology was limited.
Under the current guidelines (U.S. EPA, 2005b), there is inadequate information to assess the
carcinogenic potential of N-methylaniline.
QUANTITATIVE ESTIMATES OF CARCINOGENIC RISK
Since no chronic inhalation data are available for calculating a quantitative estimate of
carcinogenic risk from inhalation exposure to aniline, no provisional estimate of carcinogenic
risk can be derived for inhalation exposure to N-methylaniline. Lack of oral carcinogenic data
and pharmacokinetic/pharmacodynamic information also precludes derivation of a provisional
oral slope factor for N-methylaniline.
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