*>EPA
EPA/690/R-22/007F | September 2022 | FINAL
United States
Environmental Protection
Agency
Provisional Peer-Reviewed Toxicity Values for
The Aliphatic Low Carbon Range Total Petroleum
Hydrocarbon (TPH) Fraction
(various CASRNs)
U.S. EPA Office of Research and Development
Center for Public Health and Environmental Assessment
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A United Stiles
MKHu Environmental Protection
IbbI # % Agency
EPA/690/R-22/007F
September 2022
https://www.epa.gov/pprtv
Provisional Peer-Reviewed Toxicity Values for
The Aliphatic Low Carbon Range Total Petroleum
Hydrocarbon (TPH) Fraction
(various CASRNs)
Center for Public Health and 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
Elizabeth O. Owens, PhD
Center for Public Health and Environmental Assessment, Cincinnati, OH
CONTRIBUTOR
Glenn E. Rice, ScD
Center for Public Health and Environmental Assessment, Cincinnati, OH
Allison L. Phillips, PhD
Center for Public Health and Environmental Assessment, Cincinnati, OH
Jeff Swartout, MS (deceased)
Center for Public Health and Environmental Assessment, Cincinnati, OH
Jacqueline Weinberger, Student Services Contractor
Oak Ridge Associated Universities
DRAFT DOCUMENT PREPARED BY
SRC, Inc.
7502 Round Pond Road
North Syracuse, NY 13212
PRIMARY INTERNAL REVIEWERS
Jeffry L. Dean II, PhD
Center for Public Health and Environmental Assessment, Cincinnati, OH
M. Margaret Pratt, PhD
Center for Public Health and Environmental Assessment, Washington, DC
PRIMARY EXTERNAL REVIEWERS
Eastern Research Group, Inc.
110 Hartwell Avenue
Lexington, MA 02421-3136
PPRTV PROGRAM MANAGEMENT
Teresa L. Shannon
Center for Public Health and Environmental Assessment, Cincinnati, OH
J. Phillip Kaiser, PhD, DABT
Center for Public Health and Environmental Assessment, Cincinnati, OH
Questions regarding the content of this PPRTV assessment should be directed to the U.S. EPA
Office of Research and Development (ORD) Center for Public Health and Environmental
Assessment (CPHEA) website at https://www.epa.gov/pprtv/forms/contact-us-about-pprtvs.
in
Aliphatic low carbon range TPH fraction
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TABLE OF CONTENTS
COMMONLY USED ABBREVIATIONS AND ACRONYMS v
BACKGROUND 1
QUALITY ASSURANCE 1
DISCLAIMERS 2
QUESTIONS REGARDING PPRTVs 2
1. INTRODUCTION 3
1.1. DEFINITION OF THE ALIPHATIC LOW CARBON RANGE FRACTION 3
1.2. OVERVIEW OF PHYSICOCHEMICAL PROPERTIES AND ENVIRONMENTAL
FATE 3
1.3. OVERVIEW OF MIXTURE ASSESSMENT METHODS 7
1.3.1. Indicator Chemical Approach 8
1.3.2. Hazard Index Approach 8
2. SUMMARY OF TOXICITY AND DOSE-RESPONSE ASSESSMENT APPROACH 10
2.1. IDENTIFICATION OF RELEVANT MIXTURES AND COMPOUNDS WITH
TOXICITY VALUES 12
2.2. IDENTIFICATION OF OTHER RELEVANT TOXICITY DATA 14
2.3. METHODS FOR INDICATOR CHEMICAL SELECTION 15
2.4. DEVELOPMENT OF EXPOSURE-RESPONSE ARRAYS 15
3. REVIEW OF POTENTIALLY RELEVANT DATA 17
3.1. NONCANCEREVIDENCE 17
3.2. CANCER EVIDENCE 20
3.2.1. Human Studies 20
3.2.2. Animal Studies—Oral 20
3.2.3. Animal Studies—Inhalation 21
3.2.4. Cancer Evidence Summary 21
4. TOXICOKINETIC CONSIDERATIONS 22
5. MECHANISTIC CONSIDERATIONS AND GENOTOXICITY 26
6. DERIVATION OF PROVISIONAL VALUES 27
6.1. DERIVATION OF ORAL REFERENCE DOSES 27
6.1.1. Oral Noncancer Assessment Using the Indicator Chemical Method for the
Aliphatic Low Carbon Range Fraction 30
6.1.2. Alternative Oral Noncancer Assessment Using the Hazard Index Method for
the Aliphatic Low Carbon Range Fraction 32
6.2. DERIVATION OF INHALATION REFERENCE CONCENTRATIONS 32
6.2.1. Inhalation Noncancer Assessment Using the Indicator Chemical Method for
the Aliphatic Low Carbon Range Fraction 35
6.2.2. Alternative Inhalation Noncancer Assessment Using the Hazard Index
Method for the Aliphatic Low Carbon Range Fraction 37
6.3. SUMMARY OF NONCANCER PROVISIONAL REFERENCE VALUES 38
6.4. CANCER WEIGHT-OF-EVIDENCE DESCRIPTOR 38
6.5. DERIVATION OF PROVISIONAL CANCER RISK ESTIMATES 39
APPENDIX A. LITERATURE SEARCH AND SCREENING 41
APPENDIX B. COMPOSITION OF MIXTURES RELEVANT TO THE ALIPHATIC
LOW CARBON RANGE FRACTION 43
APPENDIX C. POTENTIALLY RELEVANT NONCANCER EVIDENCE 45
APPENDIX D. REFERENCES 67
iv Aliphatic low carbon range TPH fraction
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COMMONLY USED ABBREVIATIONS AND ACRONYMS
a2u-g
alpha 2u-globulin
IVF
in vitro fertilization
ACGIH
American Conference of Governmental
LC50
median lethal concentration
Industrial Hygienists
LD50
median lethal dose
AIC
Akaike's information criterion
LOAEL
lowest-observed-adverse-effect level
ALD
approximate lethal dosage
MN
micronuclei
ALT
alanine aminotransferase
MNPCE
micronucleated polychromatic
AR
androgen receptor
erythrocyte
AST
aspartate aminotransferase
MOA
mode of action
atm
atmosphere
MTD
maximum tolerated dose
ATSDR
Agency for Toxic Substances and
NAG
7V-acetyl-P-D-glucosaminidase
Disease Registry
NCI
National Cancer Institute
BMC
benchmark concentration
NOAEL
no-observed-adverse-effect level
BMCL
benchmark concentration lower
NTP
National Toxicology Program
confidence limit
NZW
New Zealand White (rabbit breed)
BMD
benchmark dose
OCT
ornithine carbamoyl transferase
BMDL
benchmark dose lower confidence limit
ORD
Office of Research and Development
BMDS
Benchmark Dose Software
PBPK
physiologically based pharmacokinetic
BMR
benchmark response
PCNA
proliferating cell nuclear antigen
BUN
blood urea nitrogen
PND
postnatal day
BW
body weight
POD
point of departure
CA
chromosomal aberration
PODadj
duration-adjusted POD
CAS
Chemical Abstracts Service
QSAR
quantitative structure-activity
CASRN
Chemical Abstracts Service registry
relationship
number
RBC
red blood cell
CBI
covalent binding index
RDS
replicative DNA synthesis
CHO
Chinese hamster ovary (cell line cells)
RfC
inhalation reference concentration
CL
confidence limit
RfD
oral reference dose
CNS
central nervous system
RGDR
regional gas dose ratio
CPHEA
Center for Public Health and
RNA
ribonucleic acid
Environmental Assessment
SAR
structure-activity relationship
CPN
chronic progressive nephropathy
SCE
sister chromatid exchange
CYP450
cytochrome P450
SD
standard deviation
DAF
dosimetric adjustment factor
SDH
sorbitol dehydrogenase
DEN
diethylnitrosamine
SE
standard error
DMSO
dimethylsulfoxide
SGOT
serum glutamic oxaloacetic
DNA
deoxyribonucleic acid
transaminase, also known as AST
EPA
Environmental Protection Agency
SGPT
serum glutamic pyruvic transaminase,
ER
estrogen receptor
also known as ALT
FDA
Food and Drug Administration
SSD
systemic scleroderma
FEVi
forced expiratory volume of 1 second
TCA
trichloroacetic acid
GD
gestation day
TCE
trichloroethylene
GDH
glutamate dehydrogenase
TWA
time-weighted average
GGT
y-glutamyl transferase
UF
uncertainty factor
GSH
glutathione
UFa
interspecies uncertainty factor
GST
glutathione -.S-1 ni n s fc ra sc
UFC
composite uncertainty factor
Hb/g-A
animal blood-gas partition coefficient
UFd
database uncertainty factor
Hb/g-H
human blood-gas partition coefficient
UFh
intraspecies uncertainty factor
HEC
human equivalent concentration
UFl
LOAEL-to-NOAEL uncertainty factor
HED
human equivalent dose
UFS
subchronic-to-chronic uncertainty factor
i.p.
intraperitoneal
U.S.
United States of America
IRIS
Integrated Risk Information System
WBC
white blood cell
Abbreviations and acronyms not listed on this page are defined upon first use in the
PPRTV document.
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PROVISIONAL PEER-REVIEWED TOXICITY VALUES FOR
THE ALIPHATIC LOW CARBON RANGE TOTAL PETROLEUM HYDROCARBON
(TPH) FRACTION
BACKGROUND
A Provisional Peer-Reviewed Toxicity Value (PPRTV) is defined as a toxicity value
derived for use in the Superfund program. PPRTVs are derived after a review of the relevant
scientific literature using established U.S. Environmental Protection Agency (U.S. EPA)
guidance on human health toxicity value derivations.
The purpose of this document is to provide support for the hazard and dose-response
assessment pertaining to chronic and subchronic exposures to substances of concern, to present
the major conclusions reached in the hazard identification and derivation of the PPRTVs, and to
characterize the overall confidence in these conclusions and toxicity values. It is not intended to
be a comprehensive treatise on the chemical or toxicological nature of this substance.
Currently available PPRTV assessments can be accessed on the U.S. EPA's PPRTV
website at https://www.epa.gov/pprtv. PPRTV assessments are eligible to be updated on a 5-year
cycle and revised as appropriate to incorporate new data or methodologies that might impact the
toxicity values or affect the characterization of the chemical's potential for causing adverse
human-health effects. Questions regarding nomination of chemicals for update can be sent to the
appropriate U.S. EPA's eComments Chemical Safety web page
(https://ecomments.epa.gov/chemicalsafetv/).
QUALITY ASSURANCE
This work was conducted under the U.S. EPA Quality Assurance (QA) program to ensure
data are of known and acceptable quality to support their intended use. Surveillance of the work
by the assessment managers and programmatic scientific leads ensured adherence to QA
processes and criteria, as well as quick and effective resolution of any problems. The QA
manager, assessment managers, and programmatic scientific leads have determined under the
QA program that this work meets all U.S. EPA quality requirements. This PPRTV assessment
was written with guidance from the CPHEA Program Quality Assurance Project Plan (PQAPP),
the QAPP titled Program Quality Assurance Project Plan (PQAPP) for the Provisional
Peer-Reviewed Toxicity Values (PPRTVs) and Related Assessments/Documents
(L-CPAD-0032718-QP), and the PPRTV development contractor QAPP titled Quality Assurance
Project Plan—Preparation of Provisional Toxicity Value (PTV) Documents
(L-CPAD-0031971-QP). As part of the QA system, a quality product review is done prior to
management clearance. A Technical Systems Audit may be performed at the discretion of the
QA staff.
All PPRTV assessments receive internal peer review by at least two CPHEA scientists
and an independent external peer review by at least three scientific experts. The reviews focus on
whether all studies have been correctly selected, interpreted, and adequately described for the
purposes of deriving a provisional reference value. The reviews also cover quantitative and
qualitative aspects of the provisional value development and address whether uncertainties
associated with the assessment have been adequately characterized.
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1 DISCLAIMERS
2 The PPRTV document provides toxicity values and information about the adverse effects
3 of the chemical and the evidence on which the value is based, including the strengths and
4 limitations of the data. All users are advised to review the information provided in this document
5 to ensure that the PPRTV used is appropriate for the types of exposures and circumstances at the
6 site in question and the risk management decision that would be supported by the risk
7 assessment.
8 Other U.S. EPA programs or external parties who may choose to use PPRTVs are
9 advised that Superfund resources will not generally be used to respond to challenges, if any, of
10 PPRTVs used in a context outside of the Superfund program.
11 This document has been reviewed in accordance with U.S. EPA policy and approved for
12 publication. Mention of trade names or commercial products does not constitute endorsement or
13 recommendation for use.
14 QUESTIONS REGARDING PPRTVS
15 Questions regarding the content of this PPRTV assessment should be directed to the
16 U.S. EPA ORD CPHEA web site at https://ecomments.epa.gov/pprtv.
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1. INTRODUCTION
This Provisional Peer-Reviewed Toxicity Value (PPRTV) assessment supports a
fraction-based approach to risk assessment for mixtures of petroleum hydrocarbons (U.S. EPA.
2022. 2009c). In this approach, total petroleum hydrocarbon (TPH) fractions are defined based
on expected transport in the environment and analytical methods used to quantify environmental
contamination by TPH mixtures. TPH components were first classified into aliphatics and
aromatics, and each of these two major fractions were further separated into low, medium, and
high carbon range fractions. This PPRTV assessment describes the derivation of toxicity values
for the aliphatic low carbon range fraction of TPH. The toxicity values described herein are used
in the assessment of Complex Mixtures of Petroleum Hydrocarbons that is intended to replace
current approaches used at TPH-contaminated sites (U.S. EPA. 2022. 2009c).
1.1. DEFINITION OF I II I ALIPHATIC LOW CARBON RANGE FRACTION
The aliphatic low carbon range fraction includes aliphatic hydrocarbons with a carbon
(C) range of C5-C8 (contains between 5 and 8 carbons, inclusive) and an equivalent carbon (EC)
number1 index range of EC5-EC82 that occur in, or co-occur with, petroleum contamination.
The EC index is equivalent to the retention time of the compound on a boiling-point gas
chromatography (GC) column (nonpolar capillary column), normalized to the //-alkanes (NJ
DEP. 2010). EC numbers are the physical characteristic that underpin analytical separation of
petroleum components. EC numbers are useful because they are more closely related to
environmental mobility than carbon number. For instance, two chemicals with similar carbon
numbers but different structures (e.g., aliphatic vs. aromatic) could partition differently into
environmental media and, ultimately, have different environmental fates. Grouping based on EC
numbers provides a consistent basis for logically placing petroleum hydrocarbon compounds into
fractions because EC measures correlate with physicochemical properties such as water
solubility, vapor pressure, Henry's law constant, and soil absorption coefficient (log Koc). For
example, cyclohexane, a C6 aliphatic compound, has an EC of 6.59 because its boiling point and
GC retention time are approximately halfway between those of //-hexane (C6 [EC6]) and
//-heptane (C7 [EC7]). Individual compounds in this fraction may include linear and branched
alkanes, alkenes, and alicyclic compounds. The selection of relevant compounds and mixture is
described in Section 2 and Appendix A.
1.2. OVERVIEW OF PHYSICOCHEMICAL PROPERTIES AND ENVIRONMENTAL
FATE
The physicochemical properties for members of the aliphatic low carbon range fraction
that have toxicity values are provided in Table 1. Section 2 details how the fraction members
with toxicity values were identified. As Table 1 shows, the seven chemicals with toxicity values
include representatives from the entire carbon range (C5-C8), and include compounds with
linear, branched, cyclic, and unsaturated structures. All seven compounds are liquids at room
temperature, with moderate water solubility and high vapor pressure. Some members of this
fraction are expected to have high mobility in soil, indicating the potential for some members of
this fraction to leach to groundwater. Measured biodegradation data for several members of the
1 Based on an empirical relationship, the EC value can be estimated from a compound's boiling point (BP; °C) using
the following equation: EC = 4.12 + 0.02 (BP) + 6.5 x 10~5 (BP)2; see Gustafson et at (1997).
2This range reflects EC values rounded to the nearest whole number. For instance, cyclohexene (EC = 6.74) is
included in this fraction because its EC value rounds to 7.
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aliphatic low carbon range fraction have been reported. In Japanese Ministry of International
Trade and Industry (MITI) ready biodegradation tests, //-pentane, //-hexane, and //-heptane
biodegraded an estimated 96, 100, and -100%, respectively, within 4 weeks (J-CUHCK. 2010a.
b, c). However, limited biodegradation of methylcyclopentane occurred under aerobic or
anaerobic conditions in pure culture studies, and slow biodegradation was reported for
2,4,4-trimethylpentene, cyclohexane, and cyclohexene under aerobic conditions. Volatilization is
expected to be the predominant fate process for the fraction members in the environment, based
on available Henry's law constant values. The aliphatic low carbon range hydrocarbons do not
contain hydrolysable functional groups; therefore, the rate of hydrolysis is expected to be
negligible for all members. In the atmosphere, photochemical degradation is expected to be slow
for the saturated category members. The three unsaturated category members (cyclohexene and
the two isomers of 2,4,4-trimethylpentene) are expected to have a moderate rate of
photochemical degradation (NI.M, 2021).
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Table 1. Physicochemical Properties of Aliphatic Low Carbon Range Hydrocarbons with Toxicity Values"
Chemical
rt-Pcntanc
w-Hexane
Methyl-
cyclopentane
Cyclohexane
Cyclohexene
rt-Heptane
2,4,4-Trimethyl-
pentene
Structure
HjC ^ ^
CHj
/x.
H3C 3
CH,
O
O
H3C H3Cx/'H3
h2c
h3c ch^ch3
/^/^rH
H3C
CASRN
109-66-0
110-54-3
96-37-7
110-82-7
110-83-8
142-82-5
25167-70-8
(mixture of two
isomers, 107-39-1 and
107-40-4)
Molecular formula
C5H12
C6H14
G5H12
C
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Table 1. Physicochemical Properties of Aliphatic Low Carbon Range Hydrocarbons with Toxicity Values"
Chemical
M-Pentane
w-Hexane
Methyl-
cyclopentane
Cyclohexane
Cyclohexene
rt-Heptane
2,4,4-Trimethyl-
pentene
Log Koc
455*
1.29 x 103*
467*
531*
196*
5.69 x 103*
2.751
'Data were gathered from the U.S. EPA CompTox Chemicals Dashboard unless otherwise specified; https://comptox.crei.gov/dashboard.
bEC number was developed by the TPHCWG and is proportional to the BP of a chemical. EC number is analogous to an n-paraffin retention time index and can be
estimated using EC = 4.12 + 0.02 (BP) + 6.5 x 10~5 (BP)2 (NIST. 2020; Edwards et al.. 1997; Gustafson et at. 1997).
°OECD (2002).
dU.S. EPA (2012a); HLC calculated based on measured VPAVS with user-entered inputs for WS = 9.5 mg/L and VP = 153 mm Hg.
"U.S. EPA (2012a); HLC calculated based on measured VP/WS with user-entered inputs for WS = 42 mg/L and VP = 138 mm Hg.
fU.S. EPA (2012a); HLC calculated based on measured VP/WS with user-entered inputs for WS = 3.4 mg/L and VP = 46 mm Hg.
gU.S. EPA (2012a); EPI Suite™ estimate with no user-entered inputs (Bond method); representative SMILES C(=CC(C)(C)C)(C)C and C(=C)(CC(C)(C)C)C.
•"Calculated from listed values for log Kow and HLC.
'OECD (2008).
*Predicted value.
BP = boiling point; C = carbon; EC = equivalent carbon; EPI Suite™ = Estimation Programs Interface Suite; HLC = Henry's law constant; Kow = octanol-water partition
coefficient; Koa = octanol-air partition coefficient; Koc = soil adsorption coefficient; SMILES = simplified molecular input line entry system; TPHCWG = Total
Petroleum Hydrocarbon Criteria Working Group; U.S. EPA = U.S. Environmental Protection Agency; VP = vapor pressure; WS = water solubility.
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1.3. OVERVIEW OF MIXTURE ASSESSMENT METHODS
A number of different approaches have been developed and used to estimate risks and
hazards posed by exposures to chemical mixtures encountered in the environment. Among the
simplest of these approaches to implement is the indicator chemical approach (ATSDR. 2018).
The indicator chemical approach estimates the risk or hazards of a mixture by evaluating the
dose-response assessment developed for a component of the mixture to the exposure rate of the
entire mixture. While it has greater uncertainty than the hazard index (HI) approach, the other
approach that will be addressed in this PPRTV assessment, the indicator chemical approach, is
used when there are only measures of the concentrations of this fraction (i.e., no information is
available on the concentrations of individual chemicals in this fraction).
The U.S. Environmental Protection Agency (U.S. EPA) Supplementary Guidance for
Conducting Health Risk Assessment of Chemical Mixtures (U.S. EPA. 2000. 1986) describes the
following two broad categories of approaches for assessing human health risks and health
hazards associated with environmental exposures to chemical mixtures: component methods and
whole mixture methods. Component-based approaches, which involve analyzing the toxicity of a
mixture's individual components, have more uncertainty and are recommended when appropriate
toxicity data on a complex mixture of concern, or on a sufficiently similar mixture (discussed
below), are unavailable (U.S. EPA. 2000. 1986). In this PPRTV assessment, a component
approach, the HI approach, is described for assessing noncancer hazards posed by exposures to
the aliphatic low carbon range fraction.
Chemical mixture assessments are conducted most appropriately with quantitative
dose-response information resulting from comparable exposures to the mixture of concern. If the
dose-response data are insufficient to develop a health reference value for the specific mixture of
concern in the environment, the second option that the U.S. EPA Supplementary Guidance for
Conducting Health Risk Assessment of Chemical Mixtures (U.S. EPA. 2000. 1986)
recommended is a "sufficient similarity" approach that uses a health reference value from a
characterized surrogate mixture to estimate the hazard or risk associated with exposures to the
mixture of concern. This method requires chemistry and toxicity data on both the potential
surrogate mixture and the mixture of concern (e.g., a key event that is related to the apical
endpoint observed in an epidemiological study or whole animal study), and a health reference
value (e.g., from an in vivo study) on the surrogate mixture. If the chemistry and toxicity data
indicate that the mixtures are "sufficiently similar" to one another, then the health reference
value for the surrogate mixture can be used as a proxy for the mixture of concern. No data were
identified that were suitable to implement a whole mixture approach.
The choice of a chemical mixtures risk assessment method is driven by the available data.
Starting with the method requiring the least information and then discussing the method
requiring more information, the following subsections summarize the indicator chemical
approach and the HI approach. Figure 1 summarizes the two approaches and the preference for
using each approach.
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Approaches
Available Exposure Data
Fraction Measure
Aliphatic low carbon fraction
Individual
Component Measures
Oral: n-hexane,
methylcyclopentane, cyclohexene,
n-heptane, 2,4,4-trimethylpentene
Inhalation: n-pentane, n-hexane,
cyclohexane, commercial hexane,
cyclohexene, n-heptane
Approach
Indicator Chemical Approach
Oral: cyclohexene
Inhalation: n-hexane (subchronic);
n-heptane (chronic)
Cancer (inhalation): commercial hexane
Hazard Index Approach
Component HQs; cyclohexene (oral), n-
hexane (inhalation, subchronic), or n-
heptane (inhalation, chronic) are used as a
surrogatefor the remainder of the fraction
mass HQ
3
Oq
1
2
a
Two approaches are available to estimate the noncancer hazards associated with exposure to the aliphatic low
range fraction. Approach selection should be driven by the available exposure data. Increased analytical
characterization of fraction components allows for more refined risk estimates with less inherent uncertainty.
Approach preference is inversely correlated with approach uncertainty.
HQ = hazard quotient.
Figure 1. Provisional Peer-Reviewed Toxicity Approaches for the Aliphatic Low Carbon
Range TPH Fraction Assessment
1.3.1. Indicator Chemical Approach
When the chemical composition of a mixture or a mixture fraction is not known, or
toxicity measures are only available for a few individual chemicals in a mixture, the toxicity of
an individual chemical can be used as an indicator for the toxicity of a mixture or a mixture
fraction (ATSDR. 2018). ATSDR (2018) describes an indicator chemical as "a chemical . . .
selected to represent the toxicity of a mixture because it is characteristic of other components in
the mixture and has adequate dose-response data." Indicator chemical approaches are typically
implemented to assess health risks in a health-protective manner; the chemical chosen as an
indicator is among the best characterized toxicologically and likely among the most potent
components of the mixture. The indicator chemical needs to have adequate dose-response data to
indicate hazard potential or a dose-response relationship for noncancer outcomes, depending on
the purpose of the assessment. The health risk value of the indicator chemical is integrated with
exposure estimates for the mixture or mixture fraction to estimate health hazards associated with
the fraction (i.e., calculate fraction-specific HI for a specific exposure pathway or a fraction-
specific cancer risk estimate for a specific exposure pathway). This approach does not scale for
potency of individual constituents; instead, it assumes that toxicity of all measured members of
the fraction can be adequately estimated, given the purpose of the risk assessment, by the
indicator chemical.
1.3.2. Hazard Index Approach
The HI approach combines estimated population exposures with toxicity information to
characterize the potential for toxicological effects. The HI is not a risk estimate, in that it is not
expressed as a probability, nor is it an estimate of a toxicity measure (e.g., percentage decrement
in enzyme activity). Instead, the HI is an indicator of potential hazard. In the HI approach, a
hazard quotient (HQ) is calculated as the ratio of human exposure (E) to a health hazard
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1 reference value (RfV) for each mixture component chemical (/) (U.S. EPA. 1986). These HQs
2 are summed to yield the HI for the mixture. In health risk assessments, the U.S. EPA's preferred
3 RfVs are the reference dose (RfD) for the oral exposure route, and the reference concentration
4 (RfC) for the inhalation exposure route.
n n
i=l i=l
6 The HI is based on dose addition (U.S. EPA. 2000; Svendsgaard and Hertzberg. 1994);
7 the hazard is evaluated as the potency-weighted sum of the component exposures. The HI is
8 dimensionless, so E and the RfV must be in the same units.
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2. SUMMARY OF TOXICITY AND DOSE-RESPONSE ASSESSMENT
APPROACH
Toxicity and dose-response assessment for the aliphatic low carbon range fraction
depends upon selection of an indicator chemical from among the component chemicals and
mixtures with existing toxicity values and entailed the four basic steps outlined here and
described in more detail below. Mixtures and compounds that met structural criteria (see
definition of the fraction, above) and had available toxicity values from designated sources were
identified.
In Step 1 and Step 2 of the assessment, literature searches were performed for the
mixtures and compounds with toxicity values and for other mixtures and compounds that are
relevant to the fraction. These literature searches were conducted in February 2018 and updated
most recently in August 2021, and were date-limited to identify assessments published after
2009. The searches were designed for two purposes: first, to determine whether new information
suggested that toxicity values for mixtures or compounds relevant to the fraction should be
updated from those identified in the U.S. EPA (2009c) PPRTV assessment for complex mixtures
of aliphatic and aromatic hydrocarbons; and second, to determine whether new noncancer
toxicity values or data on other mixtures or compounds meeting the structural criteria of the
fraction might alter the overall understanding of the toxicity of the fraction. The third step in the
assessment involved searching PubMed for new noncancer toxicity data on compounds and
mixtures lacking either Integrated Risk Information System (IRIS) oral or inhalation toxicity
values. These literature searches were conducted in February 2018 and were date-limited to
studies published from 2007 forward, in order to capture studies that were published since the
searches performed in U.S. EPA (2009c). The fourth step in the assessment involved searching
of recent comprehensive reviews on the toxicity of petroleum components or classes of
compounds relevant to the fraction, as well as Organisation for Economic Co-operation and
Development (OECD) Screening Information Data Set (SIDS) assessments3 and the Petroleum
High Production Volume (HPV) Testing Group website, to identify other mixtures or
compounds within this carbon range with existing toxicity data that may inform hazard
identification for the fraction. Toxicity data criteria included human studies of any duration by
oral, inhalation, and dermal exposure, and animal studies of oral or inhalation exposure lasting at
least 28 days (or any duration of gestational exposure). Mixture toxicity data were considered
relevant only if the mixture composition under study was quantitatively defined to enable
assessment of relevance to the fraction. Figure 2 shows a schematic depiction of the process, and
further detail is provided below.
'The OECD Existing Chemicals Database (https://hpvchemicals.oecd.org') was reviewed for relevant categories, and
dossiers for the following categories were screened: alpha-olefins, higher olefins, C5 aliphatic hydrocarbon solvents,
C7-C9 aliphatic hydrocarbon solvents, and methyl- and ethylcyclohexane. A category of C6 aliphatic hydrocarbon
solvents is under assessment, but dossiers and hazard characterization for this category were not available at the time
of the search (October 2018).
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Step 4 Step 2
1
Identify health outcomes associated with available noncancer toxicity
values; for these health outcomes, compare effects and potencies across
compounds and mixtures that have toxicity data
Compound Structural Criteria:
5 < C<8 and
5 < EC< 8 aliphatic
Mixture Structural Criteria:
>99% of the mixture consists of aliphatic compounds (<1% aromatic) and
>90% of mixture consists of compounds meeting structural criteria (left)
Toxicity Data Criteria:
Human: any duration, oral, inhalation, or dermal exposure
Animal: oral or inhalation exposure for at least 28 days or any duration during gestation
Mixture toxicity data considered relevant only if mixture composition quantitatively
defined to enable assessment of relevance to the fraction
ATSDR = Agency for Toxic Substances and Disease Registry; C = carbon; EC = equivalent carbon; HPV = High Production Volume: IRIS = Integrated
Risk Information System; OECD = Organisation for Economic Co-operation and Development; PPRTV = Provisional Peer-Reviewed Toxicity Value;
RfC = reference concentration; RfD = reference dose; SIDS = Screening Information Data Set.
Figure 2. Selection of Compounds and Mixtures for Aliphatic Low Carbon Range Fraction Hazard Identification and
Dose-Response Assessment
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2.1. IDENTIFICATION OF RELEVANT MIXTURES AND COMPOUNDS WITH
TOXICITY VALUES
The first step (see Figure 2) in assessment of the toxicity for the aliphatic low carbon
range fraction was to identify constituents of the fraction that have existing toxicity values from
any of the sources considered for the U.S. EPA (2009c) PPRTV assessment for complex
mixtures of aliphatic and aromatic hydrocarbons (these included IRIS, PPRTVs, Agency for
Toxic Substances and Disease Registry [ATSDR] Minimal Risk Levels [MRLs], Massachusetts
Department of Environmental Protection [MassDEP], Total Petroleum Hydrocarbon Criteria
Working Group [TPHCWG], and Health Effects Assessment Summary Tables [HEAST]). Of
these sources, only IRIS, PPRTVs, and ATSDR MRLs have been updated since 2009, so only
these sources were consulted for toxicity values. Based on the U.S. EPA's previous assessments
and assessment activities as well as those relevant chemicals reviewed by the MassDEP
(MassDEP, 2003) or TPHCWG (Edwards et al., 1997), the U.S. EPA compiled an initial list of
26 chemicals and 1 mixture (commercial hexane) considered relevant to the fraction [see full list
in Appendix A and description of approach and results in Wang et al. (2012)1. Published toxicity
values were identified from the IRIS, PPRTV, and ATSDR MRL databases. At least one
subchronic or chronic oral or inhalation reference value or cancer toxicity value was available for
six chemicals or mixtures: //-pentane, //-hexane, methylcyclopentane, cyclohexane, commercial
hexane, and //-heptane. Comprehensive toxicity assessments for 2,2,4-trimethylpentane (U.S.
HP A. 2007) and methylcyclohexane (U.S. HP A. 2013) were available, but did not result in the
derivation of noncancer or cancer toxicity values due to inadequate data.
In the second step (see Figure 2), all existing chemicals in the IRIS, PPRTV, and ATSDR
MRL databases were searched to determine whether any other compounds or mixtures (not on
the initial list) meeting the structural criteria for inclusion (C5-C8 and EC5-EC8 aliphatics)
were available. Searches of the IRIS and ATSDR databases did not identify any additional
compounds, but review of the PPRTV database identified two additional compounds that had
toxicity values and met structural criteria for inclusion: 2,4,4-trimethylpentene and cyclohexene.
To evaluate whether these compounds occur in, or co-occur with, petroleum contamination, the
compounds were compared against the list of petroleum mixture constituents in the TPHCWG's
(1998) Selection of Representative TPH Fractions Based on Fate and Transport Considerations
(Volume 3). In that compendium, cyclohexene was identified as a constituent of gasoline
(Gustafson et al.. 1997). In contrast, 2,4,4-trimethylpentene was not identified as a constituent of
petroleum mixtures (Gustafson et al.. 1997). However, other information indicates that
2,4,4-trimethylpentene may be added to gasoline as a fuel additive, antioxidant, or octane booster
(Rankovic et al.. 2015: EU. 2008: Calamur et al.. 2003: Gomez and Basil 1998). Thus, while not
a natural component of petroleum, 2,4,4-trimethylpentene may co-occur with petroleum
contaminants and was therefore considered relevant to the fraction. Including cyclohexene and
2,4,4-trimethylpentene brought the number of compounds or mixtures with toxicity values to
eight (seven chemicals and the commercial hexane mixture). Table 2 shows the toxicity values
available for these compounds.
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Table 2. Summary of Available Toxicity Values for Mixtures and Constituents of Aliphatic Low Carbon Range
Fraction (C5-C8, EC5-EC8)a
CASRN
Name
C
EC
Oral Reference Dose
(mg/kg-d)
Inhalation Reference Concentration
(mg/m3)
Inhalation Unit
Risk (mg/m3)"1
Oral Slope Factor
(mg/kg-d)1
Subchronic
Chronic
Subchronic
Chronic
109-66-0
//-Pcntanc
5
5
-
-
10
1
-
-
110-54-3
«-Hexane
6
6
0.3
-
2
0.7 (IRIS)
-
-
96-37-7
Methylcyclopentane
6
6.27
0.4
-
-
-
-
-
110-82-7
Cyclohexane
6
6.59
-
-
18
6 (IRIS)
-
-
Various
Commercial hexane
6
NA
-
-
27
0.6
0.0002
-
110-83-8
Cyclohexene
6
6.74
0.05
0.005
-
1
-
-
142-82-5
//-Heptane
7
7
0.003
0.0003
4
0.4
-
-
25167-70-8
2,4,4 -T rimethy lpentene
8
6.8
0.1
0.01
-
-
-
-
aExcept where indicated, all toxicity values are from PPRTVs. Where more than one source reported a toxicity value, the values were selected based on the following
hierarchy: IRIS > PPRTV > ATSDR > HEAST > MassDEP > TPHCWG.
bValues in italics are screening provisional values obtained from an existing PPRTV assessment. Screening values are not assigned confidence statements; however,
confidence in these values is presumed to be low. Screening provisional values are derived when the available data do not meet the requirements for deriving a
provisional toxicity value.
ATSDR = Agency for Toxic Substances and Disease Registry; C = carbon; EC = equivalent carbon; HEAST = Health Effects Assessment Summary Tables;
IRIS = Integrated Risk Information System; MassDEP = Massachusetts Department of Environmental Protection; NA = not applicable; PPRTV = Provisional Peer-
Reviewed Toxicity Value; TPHCWG = Total Petroleum Hydrocarbon Criteria Working Group.
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2.2. IDENTIFICATION OF OTHER RELEVANT TOXICITY DATA
Among the 28 compounds and 1 mixture identified (26 chemicals and 1 mixture on the
initial list determined relevant, plus 2,4,4-trimethylpentene and cyclohexene identified through
additional searches), there were 7 compounds and 1 mixture with toxicity values. Of the
29 fraction members, 2 (//-heptane and 2,4,4-trimethylpentene) had toxicity assessments
published within the last 5 years (2016 and 2015, respectively). In Step 3 (see Figure 2),
literature searches were conducted in PubMed to identify any new studies that could fill data
gaps for the remaining 27 fraction members. The literature searches were conducted in
February 2018, were updated in August 2021, and were date-limited to studies published from
2007 forward, in order to capture studies that were published since the searches performed for
the 2009 PPRTV assessment for complex TPH mixtures. A summary of the literature search
strategy is provided in Appendix A. As detailed in the appendix, studies considered relevant to
hazard identification included animal studies using inhalation or oral exposure routes, in which
exposures continued for at least 28 days (or any duration of gestational exposure), at least one
health outcome was assessed, and an untreated or vehicle control group was included. Human
studies of any duration in which exposure was known or presumed to be through oral, inhalation,
or dermal routes and at least one health outcome was assessed were also considered relevant.
Results of the updated literature search are as follows. Ten human studies of occupational
exposure to //-hexane were identified (Jimenez-Garza et al.. 2018; Beckman et al.. 2016; Hassani
et al.. 2014; Jia et al.. 2014; Wang et al.. 2014; Neghab et al.. 2012; Kutlu et al.. 2009; Elci et al..
2007; Prieto-Castello et al.. 2007; Puri et al. 2007). Acute human studies evaluated effects of
cyclohexane following inhalation exposure (Lammers et al.. 2009) or //-octane after dermal
exposure (Schliemann et al .. 2013) in volunteers. Animal studies of oral exposure include
8-week (Wang et al .. 2017) and 24-week (Yin et al.. 2014) studies of //-hexane in rats. Animal
studies of inhalation exposure included a 5-week study of //-hexane in mice (Liu et al.. 2012). a
30-day study of cyclohexane in mice (Campos-Ordonez et al .. 2015). a 4-week study of
3-methylpentane in rats (Chung et al.. 2016). 13-week studies of //-pentane (Kim et al.. 2012)
and //-octane (Sung et al .. 2010) in rats, and two developmental studies of //-hexane in rats (Li et
al.. 2015; I.i et al.. 2014).
In Step 4 (see Figure 2), to determine whether additional relevant compounds or mixtures
had been tested for repeat-dose and/or reproductive/developmental toxicity since 2007, recent
reviews of petroleum toxicity (Mckee et al .. 2015; Baxter. 2012; Carreon and Herri ck. 2012;
Saavedra et al.. 2007). OECD S1DS dossiers (OECD. 2010. 2004. 2000). and the Petroleum High
Production Volume (HPV) Testing Group website were searched. Mixtures considered relevant
to the fraction met the following criteria:
1. at least 90% of the mixture consisted of identified compounds within the C5-C8
and/or EC5-EC8 ranges.
2. 99% of the mixture consisted of aliphatic compounds (<1% aromatic).
3. the mixture has been tested in animals in at least one repeat-dose (>28 days) or
reproductive/developmental toxicity study using inhalation or oral exposure routes
and included an untreated or vehicle control group.
4. human mixture studies of any duration by oral, inhalation, and dermal exposure, and
animal studies of oral or inhalation exposure lasting at least 28 days (or any duration
of gestational exposure).
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None of the mixtures described on the Petroleum HPV Testing Group website met these
criteria. In addition to commercial hexane (already included), Mckee et al. (2015) described two
other mixtures that met these criteria: a C6 mixture without //-hexane, tested in an inhalation
study by Egan et al. (1980); and practical-grade hexane (<40% //-hexane and not included in the
PPRTV assessment for commercial hexane), tested in an oral study by Krasavage et al. (1980).
In addition, OECD (2004) described studies of a C5-C7 alkene mixture that met these criteria.
Thus, toxicity data for four mixtures were considered potentially relevant to the assessment of
the aliphatic low carbon range fraction. Available information on the compositions of these
mixtures is provided in Appendix B.
In addition to the two compounds with IRIS or PPRTV assessments that did not yield
toxicity value derivations (2,2,4-trimethylpentane and methylcyclohexane), searches of the
reviews and OECD assessments identified toxicity data for 10 additional aliphatic low carbon
range compounds.4 Human and animal studies that met criteria outlined above were reviewed to
support selection of surrogates for the aliphatic low carbon range fraction toxicity values.
2.3. METHODS FOR INDICATOR CHEMICAL SELECTION
Only compounds or mixtures with at least one U.S. EPA (IRIS or PPRTV) or ATSDR
toxicity value (see Table 2) were considered for use as potential indicator chemicals (or indicator
mixtures) for derivation of the fraction-specific toxicity values, although toxicity data for other
compounds were used for hazard identification and to assess consistency in effects and potency
across the components of the fraction. The method for selecting indicator chemicals was adapted
from the 2009 complex TPH mixtures document (U.S. EPA. 2009c). First, mixtures consisting of
fraction component chemicals were preferred over individual compounds, provided that the
mixture study was adequate and the mixture exhibited in vivo toxic effects similar to those
exhibited by the individual fraction components. If suitable mixture data were lacking, a
representative compound exhibiting in vivo toxic effects and potency similar to those exhibited
by other compounds in the fraction was chosen. In the event that components of the fraction
varied widely in toxic effects or potency, the toxicity value for the most potent component
(i.e., component with lowest toxicity value) was selected as an indicator chemical for the
fraction. Finally, if toxicity values were available for many or most of the individual compounds
in a fraction, and these compounds are typically monitored at sites of hydrocarbon
contamination, then a component approach would be considered.
2.4. DEVELOPMENT OF EXPOSURE-RESPONSE ARRAYS
In order to assess consistency in effects and potency across the components of the
fraction, experimental data from compound-specific IRIS and PPRTV documents and primary
data sources (identified from literature searches) were used to create exposure-response arrays
provided in Appendix C. Data were extracted only from reliable studies (e.g., studies that
provided dose-response data enabling the identification of no-observed-adverse-effect levels
[NOAELs] and lowest-observed-adverse-effect levels [LOAELs]). Target-organ-specific
NOAELs and LOAELs were determined using the following methodology.
4The 10 additional aliphatic low carbon range compounds identified in searches of the reviews and OECD
assessments are cyclopentane, 2,3-dimethylbutane, 2-methylpentane, 3-methylpentane, 1-hexene,
2-methyl-2-pentene, 2-methylhexane, 2,3-dimethylpentane, ethylcyclohexane, and 1-octene.
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5. Whenever possible, NOAELs and LOAELs were identified from existing IRIS or
PPRTV assessments. For chemicals in which both types of assessments were
available, preference was given to IRIS (in accordance with U.S. EPA Office of
Superfund Remediation and Technology Innovation [OSRTI] hierarchy of human
health toxicity values for Superfund assessments). In general, these assessments
explicitly identified NOAEL and LOAEL values only for the most sensitive target of
toxicity, so characterization of additional adverse effect levels allowed for a
comprehensive comparison of toxic effects across additional endpoints and tissues.
6. All other target-organ-specific effect levels (i.e., for targets other than the most
sensitive target identified in IRIS or PPRTV assessments, and all targets evaluated in
newly identified studies) were determined using professional judgment, taking into
consideration factors such as statistical significance (at ap-value < 0.05), biological
significance (e.g., a greater than or equal to 10% increase in liver weight), magnitude
and direction of change, and study quality. In the case of chemicals with existing IRIS
or PPRTV assessments, NOAELs and LOAELs could often be identified from
existing study summaries.
Dose-response data were presented in exposure-response arrays by health outcome and
exposure route (see Appendix C). From left to right, compounds exhibiting an effect are shown
before those not exhibiting an effect, to facilitate identification of patterns. Within the group
exhibiting an effect, compounds are ordered from lowest LOAEL to highest. For compounds that
do not exhibit an effect, NOAELs in the arrays are ordered by EC number (low to high from left
to right), with mixtures shown last. Both administered doses and exposure concentrations
reported in the arrays and in the text reflect time-weighted average (TWA) exposures to facilitate
comparisons across studies and compounds. Consistency across the fraction was evaluated by
assessing if comparable outcomes were observed for members of the fraction, and if these effects
were observed at similar dose levels.
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3. REVIEW OF POTENTIALLY RELEVANT DATA
3.1. NONCANCER EVIDENCE
Compound-specific IRIS and PPRTV documents, supplemented by the literature search
findings and recent reviews of petroleum toxicity and OECD SIDS dossiers (described above),
were reviewed to evaluate the available noncancer data for the aliphatic low carbon range
fraction compounds. Critical effects identified with existing toxicity values include peripheral
neuropathy, decreased hearing sensitivity, hepatic toxicity, decreased body weight, nasal lesions,
and developmental toxicity (decreased pup weights). Appendix C summarizes the evidence
provided by human and experimental animal studies of noncancer health outcomes. Table 3
presents an overview of the human and animal data available to evaluate these primary
toxicological endpoints for the fraction (neurological, hepatic, body weight, gastrointestinal [GI],
respiratory, and developmental). As Table 3 shows, both oral and inhalation data available to
assess consistency in effects across members of the fraction are discrepant across endpoints.
Body weight was the only endpoint consistently evaluated across most components and
mixtures. Another important data limitation not captured in Table 3 is the lack of chronic
systemic toxicity information for all but three members of the fraction. Only cyclohexene,
methylcyclohexane, and commercial hexane have been tested in comprehensive systemic
toxicity studies in animals exposed for at least 1 year, all by the inhalation route. Furthermore,
most of the oral toxicity studies observed in this database are <13 weeks in duration, and few
examined comprehensive endpoints, as most were focused on selected neurotoxicity or alpha
2u-globulin (a2u-g)-mediated renal effects in male rats. The latter effects, which if established as
acting through this mechanism, are not considered to be relevant to humans (U.S. HP A. 1991).
and are not discussed further in this assessment. In addition, few compounds have been tested for
systemic toxicity in animals exposed orally or after chronic exposure by inhalation.
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Table 3. Overview of Noncancer Human and Animal Data Availability"'b
CASRN
Name
C
EC
Neurological
Hepatic
Body Weight
Gastrointestinal
Respiratory
Developmental
109-66-0
zz-Pentane
5
5
H, I
I
0,1
0,1
I
0,1
287-92-3
Cyclopentane
5
5.66
I
I
I
I
I
79-29-8
2,3 -Dimethylbutane
6
5.68
0
0
107-83-5
2-Methylpentane
6
5.72
0,1
0,1
0
96-14-0
3-Methylpentane
6
5.85
0,1
I
0,1
I
I
592-41-6
1-Hexene
6
5.9
0,1
0,1
0,1
0
I
0
110-54-3
zz-Hexane
6
6
H, 0,1
I
0,1
0,1
I
0,1
625-27-4
2-Methyl-2-pentene
6
6.07
0
0
96-37-7
Methylcyclopentane
6
6.27
0,1
I
0,1
0,1
I
110-82-7
Cyclohexane
6
6.59
H, I
H, I
I
I
I
591-76-4
2-Methylhexane
7
6.68
0
0
565-59-3
2,3 -Dimethylpentane
7
6.69
0
0
110-83-8
Cyclohexene
6
6.74
0,1
0,1
0
25167-70-8
2,4,4-Trimethylpentene
8
6.8
O
0
0
0
0
0
540-84-1
2,2,4-Trimethylpentane
8
6.98
I
0,1
0
142-82-5
//-Heptane
7
7
H, I
H
I
108-87-2
Methylcyclohexane
7
7.22
H, 0
I
0
111-66-0
1-Octene
8
7.89
0
1678-91-7
Ethylcyclohexane
8
7.89
0
111-65-9
//-Octane
8
8
I
I
I
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Table 3. Overview of Noncancer Human and Animal Data Availability"'b
CASRN
Name
C
EC
Neurological
Hepatic
Body Weight
Gastrointestinal
Respiratory
Developmental
NA
Practical-grade hexane,
40% //-hexane
5-6
NA
0
NA
C6 Alkane mixture
without //-hexane
6
NA
I
I
NA
Commercial hexane
6
NA
I
I
I
I
I
I
68526-52-3
C5-C7 Alkene mixture
6-7
NA
0
0
0
0
0
0
includes human and animal studies meeting inclusion criteria. Bolded compounds and mixtures have at least one oral or inhalation toxicity value available (see Table 2).
bCompounds are arranged by increasing EC number.
C = carbon; EC = equivalent carbon; H = human data; I = animal inhalation studies; NA = not applicable; O = animal oral studies.
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Based on the review of the available data, there is evidence that oral or inhalation
exposures to C6 alkanes and //-heptane can induce neurological effects; most of the other
compounds in the fraction have not been explicitly tested for sensitive measures of peripheral
neuropathy or hearing. Thus, consistency in effects and potency across members of the fraction
cannot be adequately assessed for neurological endpoints. Information among a wider range of
compounds suggests that aliphatic low carbon range fraction compounds and mixtures can
induce hepatic effects in the form of increased liver weight, and that potencies are generally
comparable in subchronic inhalation studies (LOAELs range from 2,763.3 to 6,294 mg/m3 in rats
and mice), but not in subchronic oral studies (LOAELs range from 50 to 1,000 mg/kg-day in
rats). However, the small number of compounds with information on liver toxicity after oral
exposure, lack of chronic oral studies, and availability of chronic inhalation studies for only two
fraction members limit conclusions that can be drawn for hepatic effects. Data on body-weight
effects after oral and inhalation exposure to a variety of aliphatic low carbon range fraction
compounds and mixtures indicate that members of the fraction can be expected to induce
body-weight reductions at doses >400 mg/kg-day or duration-adjusted concentrations
>1,000 mg/m3.
The available data are not considered adequate to evaluate consistency in effects or
potencies across fraction members for GI endpoints. Respiratory effects have also not been
consistently shown to be associated with oral or inhalation exposure to members of the aliphatic
low carbon range fraction. Finally, too few members of the fraction have received rigorous
testing for developmental effects to assess consistency in effects or potencies for these endpoints.
In summary, there is evidence to suggest consistency in body-weight changes and hepatic
effects of some aliphatic low carbon range fraction members. However, there is not enough
information to assess consistency across the entire fraction. Data limitations (most notably, a
lack of testing for sensitive measures of peripheral neuropathy or hearing) preclude an
assessment of consistency in neurological effects and potencies for fraction members. There is
little evidence to indicate respiratory tract effects for compounds other than commercial hexane
and //-hexane. The available data are not adequate to provide confidence in an assessment of the
consistency in effects for GI tract and developmental toxicity endpoints. Finally, new studies
suggest that //-hexane may elicit adverse effects on the developing female reproductive tract, but
no other information is available to support this finding or to assess this endpoint for other
compounds.
3.2. CANCER EVIDENCE
3.2.1. Human Studies
No relationship was found between exposure to //-hexane and the occurrence of
intracranial tumors in petrochemical plant workers (U.S. EPA. 2005). No other studies of
carcinogenicity in humans exposed to aliphatic low carbon range compounds have been
identified.
3.2.2. Animal Studies—Oral
No carcinogenicity studies of animals exposed orally to compounds or mixtures in the
aliphatic low carbon range fraction have been identified.
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3.2.3. Animal Studies—Inhalation
Statistically significant increases in the incidences of liver tumors (adenomas and
carcinomas) and pituitary tumors (adenomas and adenocarcinomas) were observed in female
mice exposed to commercial hexane at duration-adjusted concentrations >366 mg/m3 (U.S. EPA.
2009b). There were no increases in tumor incidences among male mice or rats of either sex. The
findings in female mice were the basis for characterizing the weight of evidence (WOE) as
"Suggestive Evidence of Carcinogenic PotentiaF for commercial hexane (U.S. EPA. 2009b). A
screening provisional inhalation unit risk (p-IUR) of 2 x 10 4 per mg/m3 was derived based on
benchmark dose (BMD) modeling of the combined pituitary adenomas and adenocarcinomas
(U.S. EPA. 2009b).
In 2-year carcinogenicity studies of rats and mice exposed to cyclohexene by inhalation,
there was a statistically significant dose-related trend for increased incidence of combined
hepatocellular adenomas and carcinomas at the highest dose in male rats, but not in female rats
or in mice of either sex (U.S. EPA. 2012b). However, these data were not considered adequate to
assess the carcinogenic potential of cyclohexene given the small increase in incidence and lack
of dose-response relationship (U.S. EPA. 2012b).
In rats exposed to methylcyclohexane via inhalation (268 or 1,339 mg/m3) for 1 year, a
statistically significant increase in testicular tumors was observed at the low exposure level
(5/10 compared with 0/11 in controls) but not at the high exposure level (2/11) (U.S. EPA.
2013). No information on tumor histology was reported. Given the lack of dose-response
relationship, small group sizes, and abbreviated duration of exposure, U.S. EPA (2013) did not
consider these data adequate for assessment of carcinogenic potential for methylcyclohexane.
In a study examining the potential for 2,2,4-trimethylpentane to promote renal cell tumor
formation, rats were exposed to 234 mg/m3 by inhalation for up to 61 weeks (U.S. EPA. 2007).
Study groups included an initiation-only group (pre-exposed to A-ethyl -A-hydroxyethyl -
nitrosamine in drinking water for 2 weeks), a promoter-only group (2,2,4-trimethylpentane only,
6 hours/day and 5 days/week), and an initiation-promotion group. No renal cell tumors were
observed in rats exposed only to 2,2,4-trimethylpentane, and the incidence in the
initiation-promotion group was not significantly different from the incidence in the
initiation-only group (U.S. EPA. 2007). These data were not considered adequate for the
assessment of 2,2,4-trimethylpentane carcinogenicity (U.S. EPA. 2007).
3.2.4. Cancer Evidence Summary
Few data with which to assess the carcinogenic potential of compounds and mixtures in
the aliphatic low carbon range fraction are available. No human or animal studies examining
carcinogenicity were located for any compound or mixture other than commercial hexane,
//-hexane, cyclohexene, methylcyclohexane, and 2,2,4-trimethylpentane. In addition, only the
inhalation data for commercial hexane were considered adequate to assess carcinogenic
potential, resulting in a WOE descriptor of "Suggestive Evidence of Carcinogenic Potential."
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Aliphatic low carbon range TPH fraction
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2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
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20
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4. TOXICOKINETIC CONSIDERATIONS
The available toxicokinetic information on compounds and mixtures in the aliphatic low
carbon range fraction has been reviewed extensively (Mckee et al.. 2015; Baxter. 2012; Carreon
and Herrick. 2012). In general, these compounds and mixtures are absorbed by both inhalation
and oral routes and are distributed widely in the body with some preference for adipose tissue
and kidney. Metabolism of alkane compounds is predominantly via hydroxylation to alcohols,
which are further hydroxylated or dehydrogenated to hydroxy and/or ketone derivatives. Alkenes
are metabolized via epoxide intermediates to glycols. Elimination of aliphatic low carbon range
fractions occurs via exhaled air (as carbon dioxide [CO2]) and urine.
Oral absorption of compounds in the aliphatic low carbon range fraction is high.
Estimates of the absorbed fraction of orally-administered doses are 86% for
2,2,4-trimethylpentane (U.S. EPA. 2007) and 90% for cyclohexane (Mckee et al.. 2015). Oral
absorption of aliphatic hydrocarbons was inversely proportional to molecular weight and
independent of structure (linear, branched, or alicyclic) in a rat study examining a wide range of
aliphatic compounds [reviewed by Mckee et al. (2015)1. Based on conclusions from Mckee et al.
(2015). oral absorption of the remaining compounds in the aliphatic low carbon range fraction is
expected to be in the range of 80-90%.
Absorption of inhaled aliphatic low carbon range hydrocarbons is high and increases with
molecular weight and boiling point (Mckee et al.. 2015). as suggested by existing blood-gas
partition coefficients. For example, relatively little //-pentane is absorbed into the bloodstream
after inhalation exposure, because it partitions preferentially into the gas phase (Perbellini et al..
1985). Blood-gas partition coefficients reported in comprehensive toxicity assessments for
fraction members, or in publications cited by these assessments (Gargas et al.. 1989; Perbellini et
al.. 1985) are shown in Table 4. As the table indicates, partition coefficients in humans are higher
for compounds with higher EC (which is linearly correlated to boiling point) and molecular
weight.
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Aliphatic low carbon range TPH fraction
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3
4
5
6
7
8
9
10
11
12
13
14
15
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Table 4. Blood-Gas Partition Coefficients for Aliphatic Low Carbon
Compounds
Compound
C
EC
Molecular Weight (g/mol)
Human
Rat
//-Pentane
5
5
72.15
0.38a
1.48b
2,2 -Dimethy lbutane
6
5.68
86.18
0.26a
-
2-Methylpentane
6
5.72
86.18
0.41a
-
3-Methylpentane
6
5.85
86.18
0.43a
-
«-Hexane
6
6
86.18
0.80a
2.29°
Methylcyclopentane
6
6.27
84.16
0.86a
-
Cyclohexane
6
6.58
84.16
1.4°
1.39°
3-Methylhexane
7
6.76
100.21
1.3a
-
2,2,4-Trimethy lpentane
8
6.98
114.23
1.60°
1.77°
//-Heptane
7
7
100.21
2.85°
4.75°
aPerbellini et at (1985).
bMeulenberg and Viiverberg (2000) as cited in U.S. EPA (2009e).
°Gargas et at (1989).
C = carbon; EC = equivalent carbon.
Compounds in the aliphatic low carbon range are widely distributed in the body after
inhalation or oral exposure. In rats exposed by inhalation, //-pentane was distributed primarily to
liver, kidney, and small intestine (Mckee et al.. 2015). The highest deposition of cyclohexane in
rats exposed orally was in adipose tissue (Mckee et al.. 2015). After oral exposure, radioactivity
from labeled 2,2,4-trimethylpentane was primarily distributed to kidneys in male rats, with
significantly higher levels in the kidneys of male rats compared with female rats (U.S. HP A.
2007). Other deposition sites (primarily peritoneal fat and liver) contained lower amounts of
radioactivity with little difference between the sexes (U.S. HP A. 2007). Alpha-olefins (those
having a double bond at the first carbon) in the C2-C10 range are primarily distributed to the
brain, liver, kidneys, and peritoneal fat (OECD. 2004). In vitro air-tissue partitioning studies
show that many aliphatic low carbon range compounds partition into adipose tissue (coefficients
range from 39.6 to 443) and to a lesser extent into liver, brain, and kidney (coefficients <18.8)
(Gargas et al.. 1989; Perbcllini et al.. 1985).
Metabolism of aliphatic low carbon range compounds is largely dependent on structure
(linear, branched, or cyclic; alkane or alkene). Available information indicates that alkanes are
oxidatively metabolized in the liver to alcohols, ketones, carboxylic acids, dihydrodiols, and
diketones, and are subsequently conjugated to glucuronide or sulfate (Mckee et al.. 2015;
AT SDR. 1999). OECD (2004) reported that short-chain //-alkenes are predominantly
metabolized to epoxide intermediates that are subsequently converted to glycols or conjugated
with glutathione and excreted as mercapturic acids. Table 5 shows the urinary metabolites
identified after in vivo exposure to members of the fraction. Few in vivo data on metabolism of
alkenes were identified. An in vitro study using rat and human liver microsomes exposed to
1 -hexene identified two metabolites: 1 -hexen-3-ol and hexen-3-one (Carreon and Herrick. 2012).
Little is known about the dose dependence of aliphatic low carbon range compound metabolism;
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1 uptake and metabolism of cyclopentane was concentration-dependent, with greater amounts
2 exhaled (and less absorbed or metabolized) at higher concentrations (20% exhaled as
3 unmetabolized parent compound at 100 ppm, but 88% at 1,000 ppm) (Galvin and Marashi.
4 1999).
Table 5. Urinary Metabolites Identified for Aliphatic Low Carbon
Compounds
Compound
Route
Species
Urinary Metabolites
Reference
n- Pentane
(C5 [EC5])
Inhalation
(5% in air for 1 h)
Mouse
2-Pentanol, 3-pentanol, 2-pentanone
U.S. EPA
(2009e)
2-Methylpentane
(C6 [EC5.72])
Inhalation
1,500 ppm for 14 wk)
Rat
2-Methyl-2-pentanol
Frontali et
al. (1981)
3-Methylpentane
(C6 [EC5.85])
Inhalation
(1,500 ppm for 14 wk)
Rat
3 -Methy 1-3 -pentanol, 3 -methyl-2-pentanol
Frontali et
al. (1981)
«-Hexane
(C6 [EC6])
Inhalation
(1,000 ppm for 8 h)
Rat
2-Hexanol, 2,5-hexanedione, 3-hexanol,
1-hexanol, 2-hexanone
U.S. EPA
(2005)
Cyclohexane
(C6 [EC6.58])
Oral
(0.3-400 mg/kg once)
Rabbit
Cyclohexanol, trans-1,2-cyclohexane-diol
Mckee et
al. (2015)
Cyclohexene
(C6 [EC6.74])
Oral
(3 mmol/kg once)
Rat
3-Hydroxycyclohexyl mercapturic acid,
2-hydroxycyclohexylmercapturic acid
U.S. EPA
(2012b)
//-Heptane
(C7 [EC7])
Inhalation
(1,800 ppm for 6 h)
Rat
2-Heptanol, 3-heptanol,
gamma-valerolactone, 2-heptanone,
3-heptanone, 4-heptanone, 2,5-heptanedione
U.S. EPA
(2016)
Methylcyclohexane
(C7 [EC7.22])
Oral
(2-2.5 mmol/kg once)
Rabbit
/ra«s-4-Methylcyclohexane
Mckee et
al. (2015)
Oral
(800 mg/kg on
alternate days for 2 wk
Rat
2-/ra«5-Hydroxy-4-c/5-methylcyclohexanol,
2-c/5-hydroxy-4-/ra«5-methylcyclohexanol,
trans-3 -methy Icy clohexanol,
2-67.Y-hydro\y-4-67.Y-mcthylcyclohc\anol.
/ra«s-4-methylcyclohexanol,
cyclohexylmethanol
Carreoti
and
Merrick
(2012)
//-Octane
(C8 [EC8])
Oral
(1,400 mg/kg every
other day for 14 d)
Rat
2-Octanol, 3-octanol, 5-oxohexanoic acid,
6-oxohexanoic acid
Mckee et
al. (2015)
2-Methylheptane
(C8 [EC7.71])
Oral
Rat
2-Methy 1-2,5 -heptanediol,
2-methyl-5-heptanoloactone,
2-methylheptanoic acid,
2-methyl-1,2-heptanediol
Mckee et
al. (2015)
2,2,4-Trimethy lpentane
(C8 [EC6.98])
Oral
(4.4 mmol/kg once)
Rat
2,4,4-Trimethyl-2-pentanol,
2,4,4-trimethyl-l-pentanol,
2,4,4-trimethylpentanoic acid,
2,4,4-trimethyl-5-hydroxypentanoic acid,
2,2,4-trimethyl-l-pentanol,
2,2,4-trimethylpentanoic acid,
2,2,4-trimethyl-5-hydroxypentanoic acid
U.S. EPA
(2007)
C = carbon; EC = equivalent carbon.
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5
6
7
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Excretion of aliphatic low carbon compounds is predominantly via exhaled air (either as
parent or as CO2) and urine, with little excreted in feces. In rats exposed orally to cyclohexane,
60-80% (depending on dose) of the administered compound was eliminated in exhaled air
(parent and metabolite compositions were not reported) and the rest was excreted via urine
(Mckee et al.. 2015). After oral exposure to radiolabeled 2,2,4-trimethylpentane, excretion of
radioactivity occurred primarily via urine (50-67%) and exhaled air (43-49%); after inhalation
exposure, urinary excretion accounted for 60-70% of the absorbed compound (U.S. HP A. 2007).
Elimination of the aliphatic low carbon compounds is generally rapid; elimination half-lives of
0.13 hours for //-pentane and 14-18 hours for cyclohexane have been reported in rats and
humans exposed by inhalation (Mckee et al.. 2015). After inhalation exposure to //-octane, 50%
of the absorbed dose was eliminated as exhaled CO: within 10 hours after exposure (Mckee et
al.. 2015).
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6
7
8
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5. MECHANISTIC CONSIDERATIONS AND GENOTOXICITY
Of the health effects induced by aliphatic low carbon range compounds, mechanistic
information is available to inform mode of action only for peripheral nervous system effects.
Peripheral neuropathy after exposure to //-hexane has been previously established to result from
production of a y-diketone metabolite, 2,5-hexanedione (U.S. EPA. 2005). Metabolism of
//-hexane yields relatively high levels of the di ketone (U.S. HP A. 2005). Available metabolic
data (see Table 5) show only two compounds (//-hexane and //-heptane) for which y-diketone
formation has been demonstrated; however, few data are available to assess whether other
compounds in the fraction may be metabolized to y-diketone intermediates. Compared to
//-hexane, metabolism of //-heptane yields much smaller amounts of y-diketone (U.S. HP A.
2016).
Among the compounds and mixtures with any genotoxicity data summarized in
comprehensive U.S. EPA toxicity assessments (commercial hexane, //-pentane,
methylcyclopentane, cyclohexane, cyclohexene, //-hexane, //-heptane, 2,2,4-trimethylpentane,
and 2,4,4-trimethylpentene), genotoxicity data were largely negative. Positive findings were
reported for //-hexane (minimal mutagenic activity in Saccharomyces cerevisiae and slightly
increased incidences of chromosomal aberrations [CAs] in rat bone marrow after in vivo
exposure) (U.S. HP A. 2005).
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6. DERIVATION OF PROVISIONAL VALUES
1 6.1. DERIVATION OF ORAL REFERENCE DOSES
2 Subchronic provisional RfDs (p-RfDs) are available for five constituents of the fraction.
3 The critical effects for these subchronic p-RfDs are peripheral nervous system effects
4 (//-hexane), body-weight changes (methylcyclopentane), hepatic changes (2,4,4-trimethyl-
5 pentene, cyclohexene), and forestomach lesions (//-heptane based on read-across analogue
6 analysis). There are three available chronic RfDs for constituent compounds (cyclohexene,
7 //-heptane, and 2,4,4-trimethylpentene); all of these are based on the same studies and points of
8 departure (PODs) as the corresponding subchronic RfDs. Table 6 summarizes the subchronic and
9 chronic RfDs for constituent compounds and mixtures, with PODs, uncertainty factors, critical
10 effects, and associated confidence descriptors.
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Table 6. Available RfD Values for Aliphatic Low Carbon Range Fraction (C5-C8 [EC5-EC8])a
Indicator Chemical
or Components
POD
(mg/kg-d)
POD
Type
UFc
UF
Components
RfD or
p-RfD
(mg/kg-d)
Confidence
in RfD or
p-RfD
Critical Effect(s)
Species, Mode,
and Duration
Reference
Subchronic
«-Hexane
(C6 [EC6])
785
LOAEL
3,000
UFa, UFd,
UFh, UFl
0.3
Low
Reductions in motor
nerve conduction
velocity (nervous)
Rat, gavage,
8 wk
U.S. EPA (2009a): Ono et
al. (1981)
Methylcyclopentane
(C6 [EC6.27])
357
NOAEL
1,000
UFa, UFd,
UFh
0.4
Low
Reduced body weight
(body weight)
Rat, gavage,
5 d/wk for 4 wk
U.S. EPA (2009d): Haider
et al. (1985)
Cyclohexene
(C6 [EC6.74])
4.81
BMDLisd
(HED)
100
UFa, UFd,
UFh
0.05
Low
Increased total serum
bilirubin (hepatic)
Rat, gavage,
one-generation
MHLW (2001) as cited
in U.S. EPA (2012b)
//-Heptane
(C7 [EC7])
3.13
BMDLio
1,000
UFa, UFd,
UFh
0.003h
Low
Based on n-nonane as
analogue; forestomach
histopathology (GI)
Mouse, gavage,
13 wk
Dodd et al. (2003) as cited
in U.S. EPA (2016)
2,4,4 -T rimethy lpentene
(C8 [EC6.8])
41.5
BMDLio
(HED)
300
UFa, UFd,
UFh
0.1h
Low
Increased relative liver
weight (hepatic)
Rat, gavage,
one-generation
Huntingdon Life Sciences
(1997a) as cited in U.S.
EPA (2015)
28 Aliphatic low carbon range TPH fraction
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Table 6. Available RfD Values for Aliphatic Low Carbon Range Fraction (C5-C8 [EC5-EC8])a
Indicator Chemical
or Components
POD
(mg/kg-d)
POD
Type
UFc
UF
Components
RfD or
p-RfD
(mg/kg-d)
Confidence
in RfD or
p-RfD
Critical Effect(s)
Species, Mode,
and Duration
Reference
Chronic
Cyclohexene
(C6 [EC6.74])
4.81
BMDLisd
(HED)
1,000
UFa, UFd,
UFh, UFs
0.005
Low
Increased total serum
bilirubin (hepatic)
Rat, gavage,
one-generation
MHLW (2001) as cited
in U.S. EPA (2012b)
//-Heptane
(C7 [EC7])
3.13
BMDLio
10,000
UFa, UFd,
UFh, UFs
0.0003h
Low
Based on n-nonane as
analogue; forestomach
histopathology (GI)
Mouse, gavage,
13 wk
Dodd et al. (2003) as cited
in U.S. EPA (2016)
2,4,4 -T rimethy lpentene
(C8 [EC6.8])
41.5
BMDLio
(HED)
3,000
UFa, UFd,
UFh, UFs
0.0 lh
Low
Increased relative liver
weight (hepatic)
Rat, gavage,
one-generation
Huntingdon Life Sciences
(1997a) as cited in U.S.
EPA (2015)
aBolded row shows the compound and toxicity value selected as the indicator chemical for the fraction if analytical chemistry data do not identify concentrations of
individual chemicals composing this fraction.
bToxicity values are provisional values obtained from an existing PPRTV assessment. Values in italics are screening provisional values obtained from an existing
PPRTV assessment. Screening values are not assigned confidence statements; however, confidence in these values is presumed to be low. Screening provisional values
are derived when the available data do not meet the requirements for deriving a provisional toxicity value.
BMDL = benchmark dose lower confidence limit; BMDLio = 10% benchmark dose lower confidence limit; C = carbon; EC = equivalent carbon; GI = gastrointestinal;
HED = human equivalent dose; LOAEL = lowest-observed-adverse-effect level; NOAEL = no-observed-adverse-effect level; POD = point of departure;
PPRTV = Provisional Peer-Reviewed Toxicity Value; p-RfD = provisional reference dose; RfD = oral reference dose; SD = standard deviation; UF = uncertainty factor;
UFa = interspecies uncertainty factor; UFC = composite uncertainty factor; UFD = database uncertainty factor; UFH = intraspecies uncertainty factor;
UFS = subchronic-to-chronic uncertainty factor.
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Aliphatic low carbon range TPH fraction
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2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
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As suggested by the disparity in critical effects and values of RfDs for fraction members
and discussed in Appendix C, the available oral toxicity data for aliphatic low carbon range
compounds do not demonstrate significant consistency across fraction members in terms of
toxicological effects or potencies. Thus, there is no basis to identify an indicator chemical or
mixture that is representative of the effects and potency of the fraction as a whole. Therefore, the
most potent component compounds and mixtures were considered as the basis for indicator
chemical selection, as outlined in the methodology (see Section 2.3).
6.1.1. Oral Noncancer Assessment Using the Indicator Chemical Method for the Aliphatic
Low Carbon Range Fraction
If available analytical chemistry data do not identify concentrations of individual
chemicals composing this fraction, the subchronic and chronic p-RfDs (0.05 and
0.005 mg/kg-day, respectively) for cyclohexene are recommended as the indicator chemical for
the aliphatic low carbon range fraction (U.S. EPA. 2012b). The p-RfDs for cyclohexene are
based on hepatic toxicity, and available data generally support the liver as a target of aliphatic
low carbon compounds. Although the RfDs for cyclohexene are not the lowest available, the
subchronic and chronic p-RfD values for //-heptane (0.003 and 0.0003 mg/kg-day, respectively)
are not recommended, for the following three reasons. First, the //-heptane p-RfDs are screening
values based on a read-across analysis and therefore carry additional uncertainty associated with
the analogue approach. Second, the analogue upon which the values are based (//-nonane) is
outside (C9 [EC9]) the carbon range of the fraction. Third, the chronic p-RfD for //-heptane is
highly uncertain, derived with a composite uncertainty factor (UFc) of 10,000. Evaluation of
available data as discussed in Appendix C suggests that use of the cyclohexene p-RfD values is
reasonably anticipated to be protective for effects associated with exposures to other constituents
of the fraction. Users of the indicator chemical method should understand that there could be
more uncertainty associated with the application of this toxicity value to the aliphatic low carbon
range fraction than for its derivation in U.S. EPA (2012b).
The cyclohexene PPRTV assessment cited Ministry of Health, Labour, and Welfare
(MHLW. 2001a. b as cited in U.S. HP A. 2012b) as the principal studies for the subchronic and
chronic p-RfDs:
MHLW (2001a) conducted a subchronic oral toxicity study that also
examined reproductive and developmental effects that will be discussed
separately (MHLW, 2001b). This study appears to be proprietary (may have been
part of a Japanese toxicity assessment conducted by MHLW) and is in Japanese.
OECD SIDS (2002) peer-reviewed and summarized the study (cited as MHLW,
2002) and EPA subsequently had the document translated. The internal and
external peer reviewers of this PPRTV document also concurred that the MHLW
(2001a) study was suitable for deriving a provisional toxicity value. This study
was conducted as a combined repeated dose toxicity study with reproduction/
developmental toxicity screening according to OECD test guideline 422 and was
stated by OECD to be GLP compliant (no GLP statement was provided in the
study report).
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Aliphatic low carbon range TPH fraction
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3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
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Crj:CD(SD)IGS rats (12 animals/sex/treatment group) were administered
0, 50, 150, or 500 mg/kg-day of cyclohexene (98.6% pure) in corn oil via gavage.
Dose formulations were testedfor concentration and stability. Males were dosed
for 48 days andfemales for 43-53 days beginning 14 days before mating,
throughout the mating and gestational period, to Day 4 of lactation. Animals were
observed for clinical signs of toxicity daily. Body weight and food consumption
were measured weekly and at necropsy. Urinalysis was conducted on 5 males/
treatment group at 43-48 days of treatment. At sacrifice (on Day 49 for males
and 5 days after delivery for females), blood was collectedfor hematology and
clinical chemistry in all animals. The brain, liver, kidney, spleen, adrenal glands,
thymus, testis, and epididymis were weighed. Tissues and organs were examined
histologically in at least the control and high-dose group. Statistical analyses
performed included Bartlett 's test for homogeneity of variance, Dunnett's
multiple comparison test (if equal variance), and Steel's test for unequal
variances. The */2 and Fisher's exact probability tests were also used where
appropriate.
Salivation was observed at 150 (for about 5 minutes in 3/12 males and
2/12 females) and 500 mg/kg-day (all animals for 30 60 minutes in males and
6 hours in females). Lacrimation was observed in 2/12 males at 500 mg/kg-day
andfemales at >150 mg/kg-day (1/12 for each dose group). There were some
small—but statistically significant—hematological changes at 500 mg/kg-day.
Increased were the number of reticulocytes and bilirubin in males and
prothrombin time and total bile acids in females. Decreased was the level of
APTT in males. There were no treatment-related significant changes in body
weight, or food consumption, in either sex or in the urinalysis findings for males
(females not measured). There was a dose-dependent decrease in triglyceride in
males (see Table B.l). Even though triglyceride in the 500 mg/kg-day group
males was 43% lower than in the controls, the results were not statistically
significant nor was this effect noted in the females. There was an increase in total
bilirubin in both sexes; reanalysis of the data indicates that there are statistically
significant increases at all doses in males and in high-dose females. Total bile
acid was increased by >10% in all dose groups. However, the results were highly
variable and not dose dependent. Only the 150-mg/kg-day males and the 50- and
500-mg/kg-day females showed statistically significant changes above the
controls. High-dose males had a statistically significant increase in relative
kidney weight that was not accompanied by any histopathological changes and
did not reach 1SD (standard deviation) above the control (see Table B.2). OECD
SIDS (2002) reported a NOAEL of 50 mg/kg-day for the repeated dose toxicity
portion of the test based on transient salivation observed in both sexes at
150 mg/kg-day. Transient salivation is not considered sufficiently adverse to
identify as a critical effect. Although the bile acid increase was not dose
dependent and was variable, the data taken together may indicate bile duct
blockage. Bile duct blockage is also consistent with the statistically significant
increase in alkaline phosphatase in rats noted by Laham (1976) following
inhalation exposure. Based on the statistically significant increase in total bile
acid in females and total bilirubin in males at the lowest dose, no NOAEL can be
determined, and the LOAEL is 50 mg/kg-day.
31
Aliphatic low carbon range TPH fraction
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
EPA/690/R-22/007F
The selected critical effect of total bilirubin in male rats was BMD modeled. The
resultant benchmark dose lower confidence limit with one standard deviation (BMDLisd) of
19.71 mg/kg-day was subsequently converted to a human equivalent dose (HED) of
4.81 mg/kg-day (see Table 6). As reported in U.S. EPA (2012b). confidence in the principal
study was medium. Although the study was described as being conducted according to OECD
Test Guideline (TG) 422 and was subsequently translated by U.S. EPA, the OECD (2002) S1DS
report is a secondary data source. As reported in U.S. EPA (2012b). confidence in the database
was low, because only one oral repeated-dose study was available. Therefore, confidence in the
subchronic and chronic p-RfDs was also low.
6.1.2. Alternative Oral Noncancer Assessment Using the Hazard Index Method for the
Aliphatic Low Carbon Range Fraction
If the available analytical chemistry data quantify the concentrations of //-hexane,
methylcyclopentane, cyclohexene, //-heptane, or 2,4,4-trimethylpentene separately from the
remainder of the low carbon fraction, it is recommended that HQs for the individual chemicals
with analytical data be calculated and an HI for the mixture be developed using the calculated
HQs.
For subchronic oral exposures, the following subchronic p-RfDs can be used as the
denominator in the HQ equations: //-hexane (0.3 mg/kg-day), methylcyclopentane
(0.4 mg/kg-day), cyclohexene (0.05 mg/kg-day), w-heptane (0.003 mg/kg-day), and
2,4,4-trimethylpentene (0.1 mg/kg-day). In this alternative approach, the subchronic p-RfD
(0.05 mg/kg-day) for cyclohexene is recommended for use with the remainder of the fraction,
including any other fraction members analyzed individually (see Table 6).
For chronic oral exposures, the following chronic p-RfDs can be used in the denominator
of the HQ equations: cyclohexene (0.005 mg/kg-day), //-heptane (0.0003 mg/kg-day), and
2,4,4-trimethylpentene (0.01 mg/kg-day). In this alternative approach, the chronic p-RfD
(0.005 mg/kg-day) for cyclohexene is recommended for use with the remainder of the fraction,
including any other fraction members analyzed individually (see Table 6).
6.2. DERIVATION OF INHALATION REFERENCE CONCENTRATIONS
The available subchronic and chronic RfC values, with PODs, uncertainty factors, critical
effects, and confidence descriptors are presented in Table 7. As shown in the table, there are
subchronic and chronic RfCs or provisional RfCs (p-RfCs) for one mixture (commercial hexane)
and four individual compounds (//-pentane, //-hexane, cyclohexane, and //-heptane) relevant to
the aliphatic low carbon range fraction. In addition, there is a chronic p-RfC for cyclohexene.
The critical effects for the subchronic RfCs include peripheral nervous system injury (//-hexane),
diminished hearing sensitivity (//-heptane), decreased body weight and nervous system effects
(commercial hexane), and developmental toxicity (decreased pup weight; cyclohexane). The
critical effects for the chronic RfCs include peripheral nervous system injury (//-hexane),
diminished hearing sensitivity (//-heptane), liver pathology (spongiosis hepatis; cyclohexene),
nasal lesions (hyperplasia; commercial hexane), and developmental toxicity (decreased pup
weight; cyclohexane).
32
Aliphatic low carbon range TPH fraction
-------
EPA/690/R-22/007F
Table 7. Available RfC Values for Aliphatic Low Carbon Range Fraction (C5-C8 [EC5-EC8])a
Indicator
Chemical or
Components
POD
(mg/m3)
POD Type
(all are
HECs)
UFc
UF
Components
RfC or
p-RfC
(mg/m3)
Confidence
in RfC or
p-RfC
Critical Effect(s)
Species, Mode,
and Duration
Reference
Subchronic
//-Pcntanc
(C5 [EC5])
3,658
NOAEL
300
UFa, UFd,
UFh
10
Low
No treatment-related
effects
Rat, 6 h/d, 5 d/wk
for 13 wk
McK.ee and Frank (1998) as
cited in U.S. EPA (2009e)
Commercial
hexane
(C6)
804
NOAEL
30
UFa, UFh
27
Medium
Abnormal gait; decreased
body weight; mild atrophy
of sciatic and/or tibial
nerve and skeletal muscle
(nervous and body weight)
Rat, 22 h/d, 7 d/wk
for 6 mo
IRDC (1992) as cited in U.S.
EPA (2009b)
//-Hexane
(C6 [EC6])
215
BMCLisd
100
UFa, UFd,
UFh
2
Low
Peripheral neuropathy
(nervous)
Rat, 12 h/d,
7 d/wk for 16 wk
Huang (1989) as cited in
U.S. EPA (2009a)
Cyclohexane
(C6 [EC6.58])
1,822
BMCLisd
100
UFa, UFd,
UFh
18
Moderate
Reduced pup weight
(developmental)
Rat, 6 h/d, 5 d/wk,
two-generation
Kreckmann (2000) and
Duootit HLR (1997a). both as
cited in U.S. EPA (2010)
//-Heptane
(C7 [EC7])
1,170
BMCLisd
300
UFa, UFd,
UFh
4
Low
Loss of hearing sensitivity
(nervous)
Rat, 6 h/d, 7 d/wk
for 28 d
Simonsen and Lund (1995) as
cited in U.S. EPA (2016)
33
Aliphatic low carbon range TPH fraction
-------
EPA/690/R-22/007F
Table 7. Available RfC Values for Aliphatic Low Carbon Range Fraction (C5-C8 [EC5-EC8])a
Indicator
Chemical or
Components
POD
(mg/m3)
POD Type
(all are
HECs)
UFc
UF
Components
RfC or
p-RfC
(mg/m3)
Confidence
in RfC or
p-RfC
Critical Effect(s)
Species, Mode,
and Duration
Reference
Chronic
//-Pcntanc
(C5 [EC5])
3,658
NOAEL
3,000
UFa, UFd,
UFh, UFs
1
Low
No treatment-related
effects
Rat, 6 h/d, 5 d/wk
for 13 wk
McK.ee et al. (1998) as cited
in U.S. EPA (2009e)
Commercial
hexane (C6)
17.59
BMCLio
30
UFa, UFh
0.6
Medium
Nasal epithelial cell
hyperplasia (respiratory)
Rat, 6 h/d, 5 d/wk
for 2 yr
Dauehtrev et al. (1999) and
Biodvnamics (1993). both as
cited in U.S. EPA (2009b)
«-Hexane
(C6 [EC6])
215
BMCLisd
300
UFa, UFd,
UFh, UFs
0.7
Medium
Peripheral neuropathy
(nervous)
Rat, 12 h/d, 7 d/wk
for 16 wk
Huang et al. (1989) as cited
in U.S. EPA (2005)
Cyclohexane
(C6 [EC6.58])
1,822
BMCLisd
300
UFa, UFd,
UFh
6
Low-moderate
Reduced pup weight
(developmental)
Rat, 6 h/d, 5 d/wk,
two-generation
Kreckmann et al. (2000) and
DuPont HLR (1997a) as cited
in U.S. EPA (2010)
Cyclohexene
(C6 [EC6.74])
360
NOAEL
300
UFa, UFd,
UFh
lh
Low
Spongiosis hepatis
(hepatic)
Rat, 6 h/d, 5 d/wk
for 104 wk
MHLW (2003) as cited in
U.S. EPA (2012b)
//-Heptane
(C7 [EC7])
1,170
BMCLisd
3,000
UFa, UFd,
UFh, UFs
0.4
Low
Loss of hearing sensitivity
(nervous)
Rat, 6 h/d, 7 d/wk
for 28 d
Simonsen and Lund (1995)
as cited in U.S. EPA (2016)
aBolded row shows the compounds and toxicity value selected as the indicator chemical for the fraction if analytical chemistry data do not identify concentrations of
individual chemicals composing this fraction.
bToxicity values are provisional values obtained from an existing PPRTV assessment. Values in italics are screening provisional values obtained from an existing
PPRTV assessment. Screening provisional values are not assigned confidence statements; however, confidence in these values is presumed to be low. Screening
provisional values are derived when the available data do not meet the requirements for deriving a provisional toxicity value.
BMCL = benchmark concentration lower confidence limit; BMCLio = 10% benchmark concentration lower confidence limit; C = carbon; EC = equivalent carbon;
HEC = human equivalent concentration; NOAEL = no-observed-adverse-effect level; POD = point of departure; PPRTV = Provisional Peer-Reviewed Toxicity Value;
p-RfC = provisional reference concentration; RfC = inhalation reference concentration; SD = standard deviation; UF = uncertainty factor; UFA = interspecies uncertainty
factor; UFC = composite uncertainty factor; UFD = database uncertainty factor; UFH = intraspecies uncertainty factor; UFS = subchronic-to-chronic uncertainty factor.
34
Aliphatic low carbon range TPH fraction
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
EPA/690/R-22/007F
As suggested by the disparity in critical effects and values of RfCs for fraction members
and discussed in Appendix C, the available inhalation toxicity data for aliphatic low carbon
range compounds do not demonstrate significant consistency across fraction members in terms of
toxicological effects or potencies. There is no basis to identify an indicator chemical or mixture
that is representative of the effects and potency of the fraction as a whole. Therefore, the most
potent component compounds and mixtures were considered as the basis for indicator chemical
selection, as outlined in the methodology (see Section 2.3).
6.2.1. Inhalation Noncancer Assessment Using the Indicator Chemical Method for the
Aliphatic Low Carbon Range Fraction
If available analytical chemistry data do not identify concentrations of individual
chemicals composing this fraction, the lowest subchronic and chronic p-RfCs among the
compounds in this fraction, for //-hexane and //-heptane, respectively lYU.S. EPA. 2016. 2009a);
see Table 7] are recommended as indicator chemicals for the aliphatic low carbon range fraction.
Use of these values is anticipated to be protective for exposure to other constituents based on
available information (see Appendix C). However, users of the indicator chemical method should
understand that there could be more uncertainty associated with the application of this toxicity
value to the aliphatic low carbon range fraction than for its derivation in (U.S. EPA. 2016.
2009a).
The U.S. EPA (2009a) //-hexane PPRTV assessment cited Huang et al. (1989) Huang et
al. (1989) as cited in U.S. EPA (2009a) as the principal study for the subchronic p-RfC:
Male Wistar rats (eight/group) were exposed to 0, 500, 1200, or 3000ppm
(0, 1762, 4230, 10,574 mg/m3) n-hexane (>99%pure) for 12 hours/day,
7 days/week for 16 weeks (Huang et al., 1989). The authors measured motor
nerve conduction velocity (MCV) in the tail nerve along with body weight before
exposure and after 4, 8, 12, and 16 weeks of exposure to n-hexane. One animal
from each group was sacrificed at 16 weeks exposure for histopathological
evaluation of the nerve fibers in the tail. In addition, Huang et al. (1989)
measured the levels of neuron-specific enolase and beta-S-100. These nervous
system-specific proteins are a family of calcium binding proteins that are involved
in processes such as cell-to-cell communication, cell growth, intracellular signal
transduction, and development and maintenance of the central nervous system. A
dose-dependent, statistically significant reduction in body weight gain was
observed in the mid- (at 12 weeks) and high-dose (at 8 weeks) rats. Additionally,
there were some neurological deficits in mid- and high-dose rats, including a
reduction in grip strength and a comparative slowness of motion from week 12 of
exposure. However, no hindlimb paralysis was observed by the termination of the
experiment. Rats exposed to the mid and high doses of n-hexane showed a
reduction in MCV. This reduction was statistically significant during weeks 8-16
of the exposure period compared with controls. Increased incidence of paranodal
swellings, along with some evidence of demyelination and remyelination, was
present in the peripheral nerves at both mid and high doses. However, these
histopathological findings were more severe in the high dose group. Among
biochemical changes, there were dose-dependent reductions in nervous system
specific proteins, particularly the beta-S-100 proteins from tail nerve fibers,
which were significantly reduced by approximately 75% at all dose levels. The
35
Aliphatic low carbon range TPH fraction
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
EPA/690/R-22/007F
neurophysiological deficits and histopathological effects that were evident in mid-
and high-dose rats indicate a NOAEL of500 ppm.
The Huang et al. (1989) study as cited in U.S. EPA (2009a) provided adequate
dose-response data for BMD modeling with an estimated POD (benchmark concentration lower
confidence limit [BMCL] human equivalent concentration [HEC]) of 215 mg/m3 (see Table 7).
As reported in U.S. EPA (2009a). confidence in the principal study was medium. The study used
a low, but acceptable, number of animals per group (8/sex); data enabled identification of
NOAEL and LOAEL values for neurological effects. As reported in U.S. EPA (2009a).
confidence in the database was low due to the lack of a multigenerational developmental and
reproductive toxicity study. Therefore, confidence in the subchronic p-RfC was also low.
The U.S. EPA (2009a) //-heptane PPRTV assessment cited Sim on sen and Lund (1995) as
the principal study for the chronic p-RfC:
In this neurotoxicity study, groups of male Long-Evans rats (9- 10/group)
were placed in whole-body chambers and exposed to n-heptane (99.5% pure)
vapors at reported mean concentrations of 0, 801 ± 79, or 4,006 ± 242 ppm,
6 hours/day for 28 days. The study was aimed at evaluating potential effects of
n-heptane on the central auditory system, given that exposure to organic solvents
has been associated with hearing loss in rats and humans (Simonsen and Lund,
1995). Feed and water were available ad libitum except during exposure periods.
Six weeks prior to exposure, screw electrodes were mounted in the skull of the
rats for measurement of auditory brainstem responses. The amplitudes and
latencies of Components la and IV of the auditory brainstem responses elicited at
frequencies 4, 8, 16, or 32 kHz and intensities 25-95 dB were measured in
anaesthetized rats 2 months after cessation of exposure using both implanted
electrodes and needle electrodes. Body weight was monitored throughout the
study. No other systemic endpoints were assessed.
Body-weight gain during the first 2 weeks postexposure was significantly
decreased by 53% in the 4,006-ppm group. However, body weights were similar
in all three exposure groups during the course of treatment. The peak amplitudes
of the la and IV components of the auditory brainstem responses were reduced in
rats exposed to 4,006ppm at all frequencies and intensities, compared with
control (0-ppm treatment group), but not at 801 ppm. Statistically significant
reductions were reported for Component IV, most prominently at higher
frequencies and intensities (see Table B-4). Decreases in amplitude of Component
la displayed a similar pattern to IV; however statistical analyses for this
component were not provided. No exposure-related changes were observed in the
latencies or interpeak latencies of the la and IV components. The reduction in the
peak amplitudes corresponded to an approximate 10-dB increase in the auditory
threshold. The difference in auditory threshold between the control and the
4,006-ppm group was observed at all frequencies, although statistical
significance was only reached at 8 and 16 kHz (see Table B-5; data have been
digitally extracted using Grablt! Software).
36
Aliphatic low carbon range TPH fraction
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
EPA/690/R-22/007F
A NOAEL of 801 ppm and a LOAEL of4,006ppm is identified for
abnormal auditory brainstem responses and increased auditory threshold that
suggest a loss of hearing sensitivity in rats. Concentrations of 801 and 4,006ppm
are converted to human equivalent concentrations (HECs) of 821 and
4,105 mg/m3 for extrarespiratory effects by treating n-heptane as a
Category 3 gas (generally water insoluble and unreactive in the extrathoracic or
tracheobronchial regions) and using the following equation (U.S. EPA, 1994a):
HECEXRESP = (ppm x molecular weight [MW] : 24.45) x (hoursper day
exposed '¦ 24) x (days per week exposed 7) x ratio of blood-gas partition
coefficient (animal: human). For n-heptane, the blood-air partition coefficient for
rats is greater than that for humans (Gargas et al., 1989); thus, a default ratio of
1 is applied (U.S. EPA, 1994a).
BMD analyses were performed to model central auditory effects (all frequencies) in rats
exposed to //-heptane. Only data sets at frequencies of 16 and 32 Hz provided an adequate fit.
The lowest benchmark concentration lower confidence limit with one standard deviation
(BMCLisd) (HEC) of 1,170 mg/m3 was selected as the POD (see Table 7). As reported in U.S.
EPA (20161 confidence in the study was medium. Although the study was peer-reviewed, used
adequate methodology, and provided identification of NOAEL and LOAEL values for auditory
effects, it was a short-term (28 days) study in male rats only, and a limited number of endpoints
were evaluated. As reported in U.S. EPA (2016). confidence in the database was low, because no
developmental or multigeneration studies were available; the chronic study did not provide
organ-weight data or perform thorough histopathological examinations. Therefore, confidence in
the chronic p-RfC was also low.
6.2.2. Alternative Inhalation Noncancer Assessment Using the Hazard Index Method for
the Aliphatic Low Carbon Range Fraction
If the available analytical chemistry data quantify the concentrations of //-pentane,
//-hexane, cyclohexane, cyclohexene, or //-heptane separately from the remainder of the low
carbon fraction, it is recommended that HQs for the individual chemicals with analytical data be
calculated and an HI for the mixture be developed using the calculated HQs.
For subchronic inhalation exposures, the following subchronic p-RfCs can be used as the
denominator in the HQ equations: //-pentane (10 mg/m3), //-hexane (2 mg/m3), cyclohexane
(18 mg/m3), and //-heptane (4 mg/m3). In this alternative approach, the subchronic p-RfC for
//-hexane (2 mg/m3) is recommended for use with the remainder of the fraction, including any
other fraction members analyzed individually.
For chronic inhalation exposures, the following chronic p-RfCs can be used as the
denominator in the HQ equations: //-pentane (1 mg/m3), //-hexane (0.7 mg/m3), cyclohexane
(6 mg/m3), cyclohexene (1 mg/m3), and //-heptane (0.4 mg/m3). In this alternative approach, the
chronic p-RfC for //-heptane (0.4 mg/m3) is recommended for use with the remainder of the
fraction, including any other fraction members analyzed individually.
37
Aliphatic low carbon range TPH fraction
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
EPA/690/R-22/007F
6.3. SUMMARY OF NONCANCER PROVISIONAL REFERENCE VALUES
Table 8 summarizes the noncancer health references values for indicator chemicals used
when available analytical data and exposure estimates are limited to either air concentrations of,
or oral exposure rates associated with, the whole fraction. When analytical results, air
concentrations, or exposure rate measures for individual compounds with reference values are
available, then the hazards associated with these compounds can be assessed separately, using
the HI approach and reference values reported in Tables 6 and 7.
Table 8. Summary of Noncancer Reference Estimates for Indicator
Chemicals for Aliphatic Low Carbon Range (C5-C8 [EC5-EC8]) Fraction
of Total Petroleum Hydrocarbons
Toxicity Type
(units);
Indicator
Chemical
Species/
Sex
Critical
Effect
p-Reference
Value
POD
Method
POD
(HED/HEC)
UFc
Reference
Subchronic
p-RfD (mg/kg-d);
cyclohexene
Rat/M
Hepatotoxicity
(increased total
serum bilirubin)
5 x 10-2
BMDLisd
4.81
100
MHLW (2001) as
cited in U.S. EPA
(2012b)
Chronic p-RfD
(mg/kg-d);
cyclohexene
Rat/M
Hepatotoxicity
(increased total
serum bilirubin)
5 x 1(T3
BMDLisd
4.81
1,000
MHLW (2001) as
cited in U.S. EPA
(2012b)
Subchronic
p-RfC (mg/m3);
//-hexane
Rat/M
Neurotoxicity
(peripheral
neuropathy)
2 x 10°
BMCLisd
215
100
Huang et al. (1989)
as cited in U.S.
EPA (2009a)
Chronic p-RfC
(mg/m3);
//-heptane
Rat/M
Neurotoxicity
(loss of hearing
sensitivity)
4 x KT1
BMCLisd
1,170
3,000
Simonsen and Lund
(1995) as cited in
U.S. EPA (2016)
BMDL = benchmark dose lower confidence limit; C = carbon; EC = equivalent carbon; HEC = human equivalent
concentration; HED = human equivalent dose; M = male; POD = point of departure; p-RfC = provisional reference
concentration; p-RfD = provisional reference dose; SD = standard deviation; UFC = composite uncertainty factor.
6.4. CANCER WEIGHT-OF-EVIDENCE DESCRIPTOR
The inhalation cancer assessment outcomes for mixtures and individual components of
the aliphatic low carbon range fraction that have existing assessments are shown in Table 9. The
only component of the fraction for which there is information available to adequately assess
carcinogenic potential is commercial hexane. The PPRTV assessment for commercial hexane
(U.S. EPA. 2009b) describes the WOE as follows:
Under the 2005 Guidelines for Carcinogen Risk Assessment (U.S. EPA,
2005b), there is "Suggestive Evidence for [the] Carcinogenic Potential" of
commercial hexane in humans. There are no data on carcinogenicity of
commercial hexane in humans. A 2-year carcinogenicity bioassay in mice and
rats exposed to commercial hexane showed an increased incidence of liver tumors
(combined hepatocellular adenomas and carcinomas) in female mice (Daughtrey
etal., 1999; Biodynamics, 1993a, b). No increase in liver tumor incidence was
observed in treated male mice or in either sex of F344 rats exposed to commercial
38
Aliphatic low carbon range TPH fraction
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
EPA/690/R-22/007F
hexane under the same conditions. The study authors also identified a statistically
significant increase in the incidence ofpituitary tumors in female mice. Available
data on the genotoxicity of commercial hexane are limited; no gene reversion or
chromosomal aberrations in mammalian cells and no chromosomal aberrations
in the bone marrow of rats exposed in vivo were observed in the only tests
conducted.
Table 9. Available Cancer WOE Descriptors for Aliphatic Low Carbon
Range Fraction (C5-C8 [EC5-EC8])
Compound or Mixture
Cancer WOE Descriptor
Source
//-Pcntanc (C5 [EC5])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2009e)
Commercial hexane (C6
[EC NA])
"Suggestive Evidence of Carcinogenic Potential"
U.S. EPA (2009b)
«-Hexane (C6 [EC6])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2005)
Methylcyclopentane (C6
[EC6.27])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2009d)
Cyclohexane (C6 [EC6.59])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2010)
Cyclohexene (C6 [EC6.74])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2012b)
2,2,4-Trimethy lpentane
(C8 [EC6.98])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2007)
«-Heptane (C7 [EC7])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2016)
Methylcyclohexane (C7
[EC7.22])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2013)
2,4,4 -T rimethy lpentene
(C8 [EC6.8])
"Inadequate Information to Assess Carcinogenic Potential"
U.S. EPA (2015)
C = carbon; EC = equivalent carbon; NA = not applicable; WOE = weight of evidence.
While data on genotoxicity testing of compounds and mixtures in the aliphatic low
carbon range fraction are limited, available information suggests little to no genotoxic potential
(see Section 5).
6.5. DERIVATION OF PROVISIONAL CANCER RISK ESTIMATES
None of the mixtures or constituents in this fraction had an oral slope factor (OSF) from
IRIS, PPRTVs, HEAST, MassDEP, or TPHCWG. Thus, a provisional OSF (p-OSF) is not
derived for the fraction. The only available inhalation unit risk (IUR) value for members of the
aliphatic low carbon range fraction is a screening p-HJR for commercial hexane (U.S. EPA.
2009b). In the absence of data to support a clear 'best' surrogate for the mixture, the most
health-protective value will be adopted to protect against the carcinogenicity of components of
the mixture. The provisional IUR (p-IUR) of 2 x 10 4 (per mg/m3) for combined pituitary
adenomas and adenocarcinomas in female mice exposed to commercial hexane is selected to
assess inhalation carcinogenicity for this fraction (see Table 10).
39
Aliphatic low carbon range TPH fraction
-------
EPA/690/R-22/007F
Table 10. Summary of Cancer Risk Estimates for Aliphatic Low Carbon
Range (C5-C8 [EC5-EC8]) Fraction of Total Petroleum Hydrocarbons
Toxicity Type (units);
Indicator Chemical
Species/
Sex
Tumor Type
Cancer Risk
Estimate
Reference
p-OSF (mg/kg-d) 1
NDr
p-IUR (lng/in3) 1;
commercial hexane
Mouse/F
Pituitary adenomas or
adenocarcinomas
2 x 1(T4
Dauehtrev et al. (1989) and
Biodvnamics (1993). both as
cited in U.S. EPA (2009b)
C = carbon; EC = equivalent carbon; F = female; NDr = not determined; p-IUR = provisional inhalation unit risk;
p-OSF = provisional oral slope factor.
40
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APPENDIX A. LITERATURE SEARCH AND SCREENING
Literature searches were conducted in February 2018 and updated in August 2021 for
studies relevant to the derivation of provisional toxicity values the aliphatic low carbon range
fraction of total petroleum hydrocarbons (TPHs). The following 27 constituents (CASRNs) were
included for the aliphatic low carbon range fraction: cyclohexane (110-82-7), cyclohexene
(110-83-8), cyclopentane (287-92-3), 2,2-dimethylbutane (75-83-2), 2,3-dimethylbutane
(79-29-8), 2,3-dimethylpentane (565-59-3), 2,4-dimethylpentane (108-08-7), 3-ethylpentane
(617-78-7), commercial hexane (no CASRN), «-hexane (110-54-3), 2-methyl-2-butene
(513-35-9), 2-methyl-2-pentene (625-27-4), methylcyclohexane (108-87-2), methylcyclopentane
(96-37-7), 2-methylheptane (592-27-8), 3-methylheptane (589-81-1), 2-methylhexane
(591-76-4), 3-methylhexane (589-34-4), 2-methylpentane (107-83-5), 3-methylpentane
(96-14-0), «-octane (111-65-9), «-pentane (109-66-0), 2,2,3,3-tetramethylbutane (594-82-1),
2,2,3-trimethylbutane (464-06-2), 2,2,4-trimethylpentane (540-84-1), 2,3,3-trimethylpentane
(560-21-4), and 2,3,4-trimethylpentane (565-75-3). Initial searches were date limited from 2007
to 2018 and were conducted using the U.S. Environmental Protection Agency (U.S. EPA) Health
and Environmental Research Online (HERO) database of scientific literature. The PubMed
database was searched using the HERO interface. The updated search was conducted similarly
using the same search strings in PubMed and Web of Science from February 2018 through
August 2021. There was an additional search of Agency for Toxic Substances and Disease
Registry (ATSDR) and U.S. EPA documents for health risk values for fraction members.
The results of the PubMed searches (title and abstract) were screened for relevance using
the Population, Exposure, Comparison, and Outcome (PECO) criteria outline in Table A-l
below. Full-text screening for relevance to hazard identification was performed using the refined
PECO criteria shown in Table A-2.
Table A-l. PECO Criteria for Title and Abstract Screening of Total
Petroleum Hydrocarbon Constituent Literature Search Results
PECO Element
Inclusion Criteria
Population
Humans (any population) or laboratory mammals (any life stage).
Exposure
Human: Exposure to the subject material alone or as the primary component of a mixture, known
or presumed to occur by oral, inhalation, and/or dermal routes.
Animal: In vivo, exposure to the subject material alone, by oral or inhalation (including
instillation) routes, for all durations of exposures (durations <28 d will be captured as supporting
information), including any duration during gestation. Other routes of exposure will be captured
as supporting information.
Comparison
Human: Includes any comparison/referent group (no exposure, lower exposure).
Animal: Includes concurrent negative (untreated, sham-treated, or vehicle) control.
Outcomes
Assesses any cancer or noncancer endpoint in any tissue, organ, or physiological system.
PECO = Population, Exposure, Comparison, Outcomes.
41
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Table A-2. PECO Criteria for Full Text Screening for Relevance to Hazard
Identification
PECO Element
Inclusion Criteria
Population
Humans (any population) or laboratory mammals (any life stage).
Exposure
Human: Exposure to the subject material alone or as the primary component of a mixture,
known or presumed to occur by oral or inhalation routes.
Animal: In vivo, exposure to the subject material alone, by oral or inhalation routes, for
durations >28 d or any duration during gestation.
Comparison
Human: Includes any comparison/referent group (no exposure, lower exposure).
Animal: Includes concurrent negative (untreated, sham-treated, or vehicle) control.
Outcomes
Assesses any cancer or noncancer health outcome in any tissue, organ, or physiological system.
PECO = Population, Exposure, Comparison, Outcomes.
42 Aliphatic low carbon range TPH fraction
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EPA/690/R-22/007F
APPENDIX B. COMPOSITION OF MIXTURES RELEVANT TO THE ALIPHATIC
LOW CARBON RANGE FRACTION
1 Information on the composition of the C5-7 alkene mixture used in the Springborn Labs
2 study (Springborn Labs. 2003 as cited in OHCD. 2004) is provided in Table B-l. Tables B-2,
3 B-3, and B-4 list the compositions of practical-grade hexane, commercial hexane, and C6
4 mixture without //-hexane mixture in the Krasavage et al. (19801 U.S. EPA (2009b 1 and Egan et
5 al. (1980) studies, respectively.
Table B-l. Composition of C5-C7 Alkene Mixture"
Category
C
Example Compounds
Percentage in Mix
C5 //-olefins
5
//-Pcntcnc
0.5%
C5 /w-olcfms
5
3 -Methyl-1 -butene/isopentene
1.3%
C5 //-paraffins
5
//-Pcntanc
3.3%
C5 /'so-paraffins
5
2-Methylbutane/isopentane
9.3%
C6 //-olefins
6
//-Hcxcnc
10.4%
C6 /.Yo-olcfins
6
4-Methy 1-1 -pentene/isohexene
55.6%
C6 /'so-paraffins
6
2-Methylpentane/isohexane
17.8%
C7 /.Yo-olcfins
7
Isoheptene
1.0%
Total contribution from members of fraction
>99.2%
aSpringbom Labs (2003) as cited in OECD (2004).
C = carbon.
Table B-2. Composition of Practical-Grade Hexane"
CASRN
Name
EC
C
Percentage in Mix
287-92-3
Cyclopentane
5.66
5
9%
79-29-8
2,3 -Dimethylbutane
5.68
6
24%
107-83-5
2-Methylpentane/isohexane
5.72
6
1.8%
96-14-0
3-Methylpentane
5.85
6
24%
110-54-3
//-Hexane
6
6
40%
110-82-7
Cyclohexane
6.59
6
2.5%
Total contribution from members of fraction
100%
aKrasavage et al. (1980).
C = carbon; EC = equivalent carbon.
43
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Table B-3. Composition of Commercial Hexane"
CASRN
Name
EC
C
Percentage in Mix
107-83-5
2-Methylpentane/isohexane
5.72
6
13%
96-14-0
3-Methylpentane
5.85
6
16%
110-54-3
«-Hexane
6
6
52%
96-37-7
Methylcyclopentane
6.27
6
16%
110-82-7
Cyclohexane
6.59
6
<3%
108-08-7
2,4-Dimethylpentane
6.31
7
<3%
Total contribution from members of fraction
100%
aU.S. EPA (2009b).
C = carbon; EC = equivalent carbon.
Table B-4. Composition of C6 Mixture without «-Hexanea
CASRN
Name
EC
C
Percentage in Mix
79-29-8
2,3 -Dimethylbutane
5.68
6
3.4%
107-83-5
2-Methylpentane/isohexane
5.72
6
35.3%
96-14-0
3-Methylpentane
5.85
6
30.0%
110-54-3
«-Hexane
6
6
0.3%
96-37-7
Methylcyclopentane
6.27
6
24.6%
110-82-7
Cyclohexane
6.59
6
6%
Total contribution from members of fraction
>99.6%
aEganetal. (1980).
C = carbon; EC = equivalent carbon.
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APPENDIX C. POTENTIALLY RELEVANT NONCANCER EVIDENCE
DEVELOPMENT OF EXPOSURE-RESPONSE ARRAYS
As described in the main document, dose-response data were presented in
exposure-response arrays by health outcome and exposure route. The following sections
summarize the evidence provided by human and experimental animal studies of noncancer health
outcomes. In order to assess consistency in effects and potency across the components of the
fraction, experimental data from compound-specific Integrated Risk Information System (IRIS)
and Provisional Peer-Reviewed Toxicity Value (PPRTV) documents and primary data sources
(identified from literature searches) were used to create exposure-response arrays.
Exposure-response arrays present dose-response data by health outcome and exposure route.
From left to right, compounds exhibiting an effect are shown before those not exhibiting an
effect, to enable identification of patterns. Within the group exhibiting an effect, compounds are
ordered from lowest lowest-observed-adverse-effect levels (LOAELs) to highest. For compounds
that do not exhibit an effect, no-observed-adverse-effect levels (NOAELs) in the arrays are
ordered by equivalent carbon (EC) number index (low to high from left to right), with mixtures
shown last. Both administered doses and exposure concentrations reported in the arrays and in
text reflect time-weighted average (TWA) exposures, to facilitate comparisons across studies and
compounds. Consistency across the fraction was evaluated by assessing if comparable outcomes
were observed for members of the fraction, and if these effects were observed at similar dose
levels. Unless otherwise specified, the term "significant," used throughout this appendix, refers
to statistical significance at ap-walue < 0.05.
NEUROLOGICAL EFFECTS
Peripheral nervous system effects are the critical effect for the subchronic and chronic
reference concentrations (RfCs) and subchronic provisional reference dose (p-RfD) for //-hexane
(U.S. EPA. 2009a). and a cocritical effect (with decreased body weight) for the subchronic
provisional RfC (p-RfC) for commercial hexane (U.S. EPA. 2009b). A neurological endpoint
(decreased hearing sensitivity) is also the critical effect for the subchronic and chronic p-RfCs
for //-heptane (U.S. EPA. 2016). Neurological effects in humans have been studied for several
additional fraction members, but the majority of the data pertain to peripheral neuropathy
associated with //-hexane. Animal studies examining neurological effects are available for about
half of the compounds or mixtures with toxicity data; however, the studies varied widely with
respect to the spectrum of the neurological effects evaluated.
Human Studies
Neurotoxicity has been observed in humans exposed to aliphatic compounds in the low
carbon range fraction. //-Hexane is the most intensely-studied compound in this fraction, with
studies of occupational exposure resulting in peripheral neuropathy characterized by loss of
distal motor and sensory function (Wang et at.. 2014; Kutlu et at.. 2009; Puri et at.. 2007; U.S.
EPA. 2005). Clinical symptoms of neurotoxicity include weakness, motor impairment,
paresthesia (burning or tingling sensation in limbs), hypoesthesia (partial loss of sensation and/or
diminished sensibility), and changes in tendon reflexes and muscle tone. These symptoms were
usually confined to distal portions of the limbs, and the degree of intensity depended on the
extent of exposure (Wang et at.. 2014; Kutlu et at.. 2009; Puri et at.. 2007; U.S. EPA. 2005).
Electrophysiology measurements in exposed workers revealed decreased maximum conduction
velocity (MCV) and reduced amplitude of the sensory nerve action potential (SNAP) (Wang et
45
Aliphatic low carbon range TPH fraction
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9
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11
12
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al.. 2014; Kutlu et al.. 2009; Puri et al.. 2007; U.S. EPA. 2005). Reduced SNAP amplitude was
also observed in asymptomatic workers exposed to //-hexane, and the magnitude of the effect
was correlated with urinary concentrations of 2,5-hexanedione (Neghab et al.. 2012).
Examination of sural nerve biopsy samples showed axonal swelling, demyelination, and a
selective decrease in long myelinated neurons in workers exposed to //-hexane (Puri et al.. 2007;
U.S. EPA. 2005).
Some human studies have suggested central nervous system (CNS) toxicity resulting
from //-hexane exposure, including clinical signs of Parkinsonism (i.e., tremor, bradykinesia, and
rigidity), memory loss, and impaired visual motor response to neurological assessment (U.S.
EPA. 2005). Pathology and magnetic image resonance findings in these patients indicated loss of
dopaminergic neurons, gliosis in the substantia nigra, and cerebral cortical atrophy. //-Hexane
also affects vision in exposed workers, demonstrated by decreased visual evoked potentials,
color discrimination deficits, and maculopathy, characterized by damage to blood vessels, fluid
leakage into the retina, and pigment dispersion (Bcckman et al.. 2016; Kutlu et al.. 2009; U.S.
EPA. 2005).
No clinical signs of peripheral neuropathy were reported in 18 workers exposed to a
solvent containing >90% //-heptane for 1-9 years (U.S. EPA. 2016). However, electrophy siology
testing of 12 workers revealed a decrease in motor nerve conduction velocity (NCV) correlated
with increased exposure duration and an increase in amplitude desynchronization of the evoked
muscle action potential (U.S. EPA. 2016).
Neurological symptoms (i.e., fatigue, headache, dizziness) were reported in workers
exposed to glue containing at least 75% cyclohexane (U.S. EPA. 2010). El ectrophy si ol ogi cal
abnormalities were also noted (i.e., shorter motor distal latency); however, workers were
previously exposed to //-hexane. Other study limitations included small group sizes (n = 15-18)
and poorly matched controls. No neurological symptoms were reported in a different study of
workers exposed to glue containing at least 75% cyclohexane; however, the findings of this
study were limited by small cohort size, discrepancies in reporting of analytical air
concentrations, and absence of details related to the measured health outcomes (U.S. EPA.
2010). Print shop workers exposed to methylcyclohexane and other solvents for an average of
15 years experienced sleep apnea, mood disturbances, and decreased hand-eye coordination
(U.S. EPA. 2013). Volunteers exposed to 4,000, 8,000, 14,000, or 20,000 mg/m3 //-heptane for
up to 15 minutes reported vertigo, with severity increasing with exposure concentration (U.S.
EPA. 2016). Additional effects observed at the highest concentration included hilarity,
incoordination, and inability to walk straight.
Neurological effects were not observed in volunteers exposed to 15,000 mg/m3 //-pentane
for 10 minutes (Mckee et al.. 2015). or cyclohexane at 86 or 860 mg/m3 for 4 hours in two test
sessions (U.S. EPA. 2010).
46
Aliphatic low carbon range TPH fraction
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2
3
4
5
6
7
8
9
10
11
12
13
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16
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Animal Studies
Animals exposed orally to alkane compounds containing six carbons have exhibited
peripheral nervous system effects; few data on the neurotoxicity of other members of the fraction
were located. Studies for which neurotoxicity effect levels could be determined are shown in an
exposure-response array (see Figure C-l). Decreases in NCV occurred after oral exposure to
several C6 alkanes at doses between 785 and 1,168 mg/kg-day in a comparative toxicity study by
Ono et al. (1981). The relative potency of effects on NCV, based on severity of changes, was
//-hexane > methylcyclopentane >2-methylpentane > 3-methylpentane. In a 24-week
neurotoxicity study of //-hexane that was published after development of the PPRTV and IRIS
documents for that compound (Yin et al.. 2014). a LOAEL of 500 mg/kg-day was identified for
gait abnormalities; this value is comparable to the LOAEL of 785 mg/kg-day identified by Ono
et al. (1981) and was used as the basis for the sub chronic oral p-RfD for that compound. An
8-week study focused on evaluating whether diallyl sulfide mitigates neurotoxic effects of
//-hexane reported gait abnormalities and decreased grip strength in rats exposed to
3,000 mg/kg-day //-hexane (the only dose tested) (Wang et al.. 2017). Krasavage et al. (1980)
reported no hindlimb paralysis in a group of five male rats exposed to 4,000 mg/kg-day
(5 days/week for 13 weeks) practical-grade hexane containing 40% //-hexane, but one of the five
rats exhibited histologic evidence of neuropathy (giant axonal neuropathy) while no control rats
exhibited this effect; the small number of animals tested and the lack of statistically significant
change preclude determination of effect levels for this mixture.
Limited data in rats exposed orally to alkenes do not show evidence of neurotoxicity.
Exposure to 1-hexene did not induce sciatic nerve histopathology at doses up to 1,000 mg/kg-day
for 6-7 weeks (Gingell et al.. 2000 as cited in OECD. 2004) and there was no change in rotarod
performance at doses up to 3,365 mg/kg-day for 4 weeks (Dotti et al.. 1994 as cited in OECD.
2004). Exposure of rats to 2,4,4-trimethylpentene (up to 1,000 mg/kg-day for 4 weeks) did not
result in treatment-related effects on functional observational battery (FOB), sensory reactivity,
grip strength, motor activity, or histopathology in the brain, spinal cord, or sciatic nerve (U.S.
EPA. 2015). Similarly, oral exposure of rats to the C5-C7 alkene mixture at doses up to
1,000 mg/kg-day for 4 weeks did not alter FOB or histopathology of brain, spinal cord, or optic
or peripheral nerve (Springborn Laboratories. 2003 as cited in OECD. 2004).
Neurological effects seen after inhalation exposure to aliphatic low carbon range
compounds include peripheral neuropathy and related signs (abnormal gait and peripheral nerve
atrophy), decreased hearing sensitivity, and mild narcosis or sedation (see Figure C-2). Studies
examining CNS effects, including hearing sensitivity, are displayed in Figure C-3.
47
Aliphatic low carbon range TPH fraction
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48
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and Mixtures
49
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Figure C-3. Hearing Sensitivity and Other Central Nervous System Effects in Animals after Inhalation Exposure to Aliphatic Low
Carbon Range Compounds and Mixtures
50
Aliphatic low carbon range TPH fraction
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5
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9
10
11
12
13
14
15
16
17
18
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34
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39
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44
EPA/690/R-22/007F
As Figure C-2 indicates, eight compounds, commercial hexane, and the C6 alkane
mixture without //-hexane have been tested for different measures of peripheral neuropathy
(e.g., NCV, hindlimb spread, rotarod performance, and tibial or sciatic nerve histopathology) in
studies of at least 13 weeks in duration. Of the compounds tested for any peripheral nervous
system effect, only //-hexane and commercial hexane exhibited evidence of peripheral
neuropathy. Of note, exposure to 2- and 3-methylpentane and methylcyclopentane by inhalation
did not result in significant effects on hindlimb spread or tibial or sciatic nerve histology, despite
the fact that these compounds induced effects on NCV after oral exposure (Ono et al.. 1981). The
study authors of the inhalation study for methylcyclopentane (Yang et al.. 2014) noted that their
study was likely not adequate to evaluate potential neurological effects, as specialized
histopathology preparations (teased nerve fiber preparations or Epon-embedded specimens) may
be necessary to detect axonal changes. Many of the other available studies suffer from similar
limitations; thus, the data from these studies should not be interpreted as providing
incontrovertible evidence for a lack of peripheral nerve damage.
Decreased hearing sensitivity was observed in rats following inhalation exposure to
//-heptane and //-hexane (see Figure C-3), but little information is available for this endpoint. A
single study of brainstem evoked potentials in rats exposed to 1,000 ppm //-hexane for
18 hours/day for 9 weeks suggested slight loss of auditory sensitivity; no effect on auditory
sensitivity was seen after only 4 weeks of exposure to //-hexane (U.S. HP A. 2005). For
//-heptane, decreased hearing sensitivity was the critical effect in the 4-week study used to derive
the p-RfC (U.S. HP A. 2016). No other fraction members were specifically tested for auditory
sensitivity. In mice and rats exposed to cyclohexane, transient decreases in the sensitivity of the
animals to auditory stimuli were reported, but these effects were attributed to sedation (U.S.
HP A. 2010).
Volatile hydrocarbons are well-known to induce narcotic effects after acute exposure to
high concentrations (Mckee et al .. 2015). In longer-term studies of cyclohexane and //-hexane at
lower exposure levels, some evidence of narcosis was observed. Transient sedative effects in the
absence of histopathology changes were observed in 13-week studies of rats and mice exposed to
6,886 mg/m3 cyclohexane (U.S. HP A. 2010); the effects were transient and generally occurred
during the exposure period (U.S. HP A. 2010). Decreased activity was reported in female mice
exposed to //-hexane at a concentration of 8,340 mg/m3; one mouse died at this exposure level
(Liu et al.. 2012). Narcotic effects were not reported in other studies reviewed.
In studies examining primarily other CNS endpoints (including FOB, motor activity, and
histopathology of brain), no effects were seen in rats exposed by inhalation to //-pentane (U.S.
HP A. 2009e). cyclopentane (Toxicity Testing Consortium. 1997 as cited in Galvin and Marashi.
1999; Kim merle and Thvssen. 1975). 3-methylpentane (Chung et al.. 2016). or commercial
hexane (U.S. EPA. 2009b).
Summary of Potentially Relevant Neurological Evidence
Available data indicate that neurological effects associated with oral or inhalation
exposure to saturated members of the aliphatic low carbon range fraction include peripheral
neuropathy, decreased hearing sensitivity, visual deficits, and CNS effects. The lowest LOAELs
(by compound or mixture) for neurological endpoints (excluding transient effects for
cyclohexane) ranged from 1,230 to 8,340 mg/m3 in inhalation studies in rats and mice
(see Figures C-2 and C-3) and from 500 to 1,168 mg/kg-day in subchronic oral studies in rats
51
Aliphatic low carbon range TPH fraction
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45
EPA/690/R-22/007F
(see Figure C-l). In contrast, the limited available data on unsaturated fraction members and
mixtures (1-hexene, 2,4,4-trimethylpentene, and the C5-C7 alkene mixture) do not indicate
neurological effects. There are data demonstrating a causal relationship between //-hexane
exposure and peripheral neuropathy in both humans and animals. Available oral and inhalation
studies of other fraction members suggest that other six carbon alkanes (including 2- and
3-methylpentane and methylcyclopentane) and commercial hexane (a mixture of primarily C6
alkanes) may also induce peripheral neuropathy. While other studies may be limited by lack of
specialized histopathological evaluation for peripheral nerve damage, the remaining studies do
show that compounds other than //-hexane do not induce severe peripheral neuropathy that would
be observed as clinical signs (e.g., gait abnormalities). Exposure to //-hexane and //-heptane via
inhalation have been shown to reduce auditory sensitivity in rats. Supporting data in humans are
lacking, and no other studies evaluating this endpoint in animals exposed to other compounds in
the fraction were located in the sources reviewed. Humans exposed to //-hexane have shown
visual deficits, but data in animals, or in humans after exposure to other members of the fraction,
were not identified. Other CNS effects have been reported to occur in humans (dizziness,
headache, signs of Parkinsonism, memory loss) and animals (sedation) exposed by inhalation to
several aliphatic low carbon range fraction members (//-hexane, cyclohexane,
methylcyclohexane, and //-heptane).
Taken together, the available data indicate that C6 alkanes and //-heptane can induce
neurological effects. However, because most of the other compounds in the fraction have not
been explicitly tested for sensitive measures of peripheral neuropathy or hearing, it is not
possible to evaluate the consistency in these endpoints and their potencies across members of the
fraction.
HEPATIC EFFECTS
Hepatic effects are the critical effects for the subchronic and chronic p-RfDs and chronic
p-RfC for cyclohexene (U.S. EPA. 2012b). and for the subchronic and chronic p-RfD for
2,4,4-trimethylpentene (U.S. EPA. 2015). Critical hepatic effects of cyclohexene exposure
included increased serum bilirubin and spongiosis hepatis, while the critical effect of
2,4,4-trimethylpentene was increased liver weight. Few human data pertaining to the
hepatotoxicity of aliphatic low carbon range fraction members are available, and those data are
limited to clinical chemistry measurements in workers exposed to mixtures. As shown in
Table 3, data on hepatic effects in animals were located for 14 members of the fraction. In
general, the hepatic endpoints evaluated in the animal studies were liver weight and histology,
with a few studies measuring clinical chemistry.
Human Studies
Few data are available to evaluate potential hepatic effects of aliphatic low carbon range
fraction exposures in humans. Workers exposed to methylcyclohexane and //-heptane (in
addition to toluene and xylene) exhibited statistically significant elevations of urinary bile acids,
urinary 6P-hydroxyCortisol, and ratio of 6P-hydroxycortisol to urinary free Cortisol (considered
by the study authors to be sensitive measures of hepatotoxicity) compared with the control
workers with normal liver function (U.S. EPA. 2013). No differences were seen between these
two groups in serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline
phosphatase, y-glutamyl transferase (GGT), bilirubin, or urinary D-glucaric acid levels. No
changes to clinical chemistry parameters were reported in a study of workers exposed to glue
containing at least 75% cyclohexane; however, the findings of this study were limited by the
52
Aliphatic low carbon range TPH fraction
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1
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3
4
5
6
7
8
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38
39
40
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42
43
EPA/690/R-22/007F
small cohort size (n = 38), discrepancies in reporting of analytical air concentrations, and
absence of details related to the clinical chemistry parameters that were evaluated (U.S. HP A.
2013).
Animal Studies
As shown in Figure C-4, data on hepatic effects of oral exposure to aliphatic low carbon
range compounds are limited to 4-8-week rat studies. Exposure to either >300 mg/kg-day
2,4,4-trimethylpentene (U.S. HP A. 2015) or 1,000 mg/kg-day C5-C7 alkene mixture
(Springborn Laboratories. 2003 as cited in OHCD. 2004) induced increases in absolute and/or
relative liver weight, without concomitant histopathology changes. Rats exposed to cyclohexene
for 7 weeks exhibited increased serum total bilirubin and bile acids at doses >50 mg/kg-day
(U.S. HP A. 2012b). No changes in liver weight or histology were observed in rats exposed to
1 -hexene (up to 1,000 mg/kg-day) for 6-8 weeks (Gingett et al.. 2000 as cited in Carreon and
Herrick. 2012; OHCD. 2004).
Figure C-5 displays the exposure-response array for hepatic effects of inhalation
exposures up to 26 weeks in duration. Only two fraction members were tested in longer-term
(1-2-year) studies (cyclohexene and commercial hexane); these data were not arrayed as they
were not considered to be comparable to the shorter-duration studies. Hepatic effects, primarily
consisting of liver weight changes without effects on hepatic histopathology, were reported in
rats exposed for up to 26 weeks to 3-methylpentane, commercial hexane, methylcyclopentane,
cyclohexane, and //-hexane. Histologic changes in the liver were seen only with chronic exposure
to cyclohexene and subchronic exposure to commercial hexane and cyclohexane. Chronic
(2-year) exposure to cyclohexene resulted in an increased incidence of spongiosis hepatis at
concentrations >720 mg/m3 (U.S. HP A. 2012b). Slight hemorrhage and inflammation in the
livers were noted in a few male rats exposed to 5,639 mg/m3 commercial hexane for 13 weeks
(U.S. HP A. 2009b). However, chronic (2-year) exposure to commercial hexane at concentrations
up to 5,639 mg/m3 did not result in any histopathology changes in the livers of rats or mice (U.S.
HP A. 2009b). Increased liver weights and an increase in the incidence of centrilobular
hepatocellular hypertrophy were observed in rats after exposure to 24,101 mg/m3 cyclohexane
for 13 weeks (U.S. HP A. 2010). In a companion experiment in mice, liver weights were
increased without clinical chemistry or histology changes (U.S. HP A. 2010).
Exposure of rats to 2,648 mg/m3 3-methylpentane for 4 weeks (Chung et al .. 2016) or
3,608 mg/m3 methylcyclopentane for 13 weeks (Yang et al.. 2014) resulted in increased relative
liver weights (in the absence of body-weight changes), but no effects on histopathology.
Increased relative liver weights without histopathology changes were observed in mice exposed
to 6,294 mg/m3 //-hexane for 13 weeks, but body-weight decreases also occurred in this group
(U.S. HP A. 2005). Increases in total serum cholesterol and serum albumin were observed in
male, but not female, rats exposed for 13 weeks to 167 mg/m3 //-octane, but there were no other
clinical chemistry changes or effects on liver weight or histopathology; these effects were not
considered to be adverse (Sung et al.. 2010). No hepatic effects were noted in rats after
subchronic exposure to //-pentane (Kim et al.. 2012). cyclopentane (Toxicity Testing
Consortium. 1997 as cited in Galvin and Marashi. 1999; Kim merle and Thvssen. 1975).
1 -hexene (Gingell. 1999 as cited in OHCD. 2004). or 2,2,4-trimethylpentane (IUCLID. 2000 as
cited in Johnson et al.. 2012).
53
Aliphatic low carbon range TPH fraction
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Figure C-5. Hepatic Effects in Animals after Subchronic Inhalation Exposure to Aliphatic Low Carbon Range Compounds and
Mixtures
55
Aliphatic low carbon range TPH fraction
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1
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3
4
5
6
7
8
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33
34
35
36
37
38
39
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41
EPA/690/R-22/007F
Summary of Potentially Relevant Hepatic Evidence
Oral studies examining liver effects were limited to five compounds and one mixture
(C5-C7 alkenes) in studies of 4-7 weeks in duration, and most showed increases in liver weight.
Hepatic effects, primarily consisting of increased relative liver weights in the absence of
body-weight changes, were also seen in inhalation studies in laboratory animals exposed to at
least one compound with six, seven, and eight carbons, and with linear, branched, cyclic, and
unsaturated structures. Histopathological changes in the livers of animals exposed to aliphatic
low carbon range fraction members varied, consisting of hepatocellular hypertrophy in
subchronic oral and inhalation studies and spongiosis hepatis in a chronic inhalation study.
Lowest LOAELs (by compound or mixture) for hepatic endpoints ranged between 2,763.3 and
6,294 mg/m3 in subchronic inhalation studies in rats and mice (see Figure C-5) and varied from
50 to 1,000 mg/kg-day in subchronic oral studies in rats (see Figure C-4). Too few chronic
studies were available to compare effects and potencies after longer exposures. In aggregate, the
data suggest that many aliphatic low carbon range fraction compounds and mixtures can produce
increases in rodent liver weight, occasionally in tandem with histological (hepatocellular
hypertrophy) or serum chemistry (increases in bilirubin, ALT, or GGT) changes, and that
potencies are generally comparable in inhalation studies, but more variable in oral studies.
BODY-WEIGHT EFFECTS
Decreased body weight was a cocritical effect in the study used to derive the subchronic
p-RfC for commercial hexane (U.S. EPA. 2009b). No human studies examining body-weight
effects of aliphatic low carbon range compounds were identified in the sources reviewed.
As Table 3 shows, animal studies that examined body weight as an endpoint are available
for nearly all of the compounds and mixtures with toxicity data; exceptions are ethylcyclohexane
and practical-grade hexane. In this section, body-weight changes of at least 10% relative to
controls in adult animals are considered LOAELs, and smaller changes are not. For studies that
reported body-weight gain but did not report absolute body weights, and for studies of maternal
weight gain during gestation, statistically significant changes from control are described.
Animal Studies
Figure C-6 shows the effects of orally-administered aliphatic low carbon range
compounds and mixtures on body weight; data are available for 14 compounds and one mixture,
including compounds with carbon numbers across the entire range (C5-C8). Body-weight
decreases were seen with several C5-C6 compounds: //-pentane, 2,3-dimethylbutane, //-hexane,
2-methyl-2-pentene, and methylcyclopentane. No effects on body weight were seen in studies of
compounds of higher (EC > 6.68) equivalent carbon number (U.S. EPA. 2015. 2012b; Til et at..
1986 as cited in OECD. 2004; Haider et al.. 1985).
Body-weight effects in animals exposed by inhalation for subchronic (up to 16 weeks) or
chronic durations (26 weeks to 2 years) are shown in Figures C-7 and C-8. In inhalation studies,
reductions in body weight were reported to occur in rats and/or mice exposed for up to 16 weeks
to //-hexane, 2-methylpentane, 1-hexene, and 2,2,4-trimethylpentene at concentrations
>1,000 mg/m3; and in rats, mice, or hamsters exposed for >26 weeks to //-pentane, //-hexane,
methylcyclohexane, and commercial hexane at concentrations ranging from 268 to 5,639 mg/m3.
56
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57
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Range Compounds and Mixtures
58
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EPA/690/R-22/007F
Body-weight changes associated with gestational exposure are not shown in the figures.
Maternal body-weight reductions were reported in pregnant rats and mice exposed during
gestation to //-hexane at a concentration of 14,686 mg/m3 (U.S. EPA. 2005) and in pregnant rats
exposed to commercial hexane (>2,632 mg/m3) (U.S. HP A. 2009b) or cyclohexane
(>1,722 mg/m3) (U.S. HP A. 2010). No effect on maternal body weight was noted in pregnant rats
exposed to //-pentane concentrations up to 7,377 mg/m3 on gestation days (GDs) 6-15 (U.S.
HP A. 2009e).
No body-weight changes were observed in studies of adult rats, mice, or rabbits exposed
by inhalation or oral administration for at least 4 weeks to cyclopentane, 3-methylpentane,
//-heptane, //-octane, or the C6 alkane mixture without //-hexane.
Summary of Potentially Relevant Body Weight Evidence
Compounds and mixtures in the aliphatic low carbon range fraction have been shown to
reduce body weights of rats, mice, and hamsters after oral and inhalation exposure. Individual
compounds that induced body-weight changes after inhalation exposure include compounds
across the entire carbon range (C5-C8) and compounds representing linear, branched, cyclic,
and unsaturated structures. Lowest LOAELs ranged between 1,414 and 5,357 mg/m3 in rats and
between 6,294 and 8,340 mg/m3 in mice in subchronic inhalation studies (see Figure C-7). In
chronic inhalation studies, a LOAEL of 268 mg/m3 was identified in hamsters; LOAELs ranged
between 472 and 5,639 mg/m3 in rats and mice (see Figure C-8). In oral studies, body-weight
decreases were seen with several C5-C6 compounds, but compounds with higher equivalent
carbon numbers (EC > 6.68) did not induce body-weight changes. Lowest LOAELs (by
compound or mixture) for body-weight endpoints ranged between 357 and 1,500 mg/kg-day in
subchronic oral studies in rats (see Figure C-6). Taken together, the inhalation and oral animal
data indicate that compounds in the aliphatic low carbon range fraction can be expected to
induce body-weight reductions at sufficiently high doses (generally >1,000 mg/kg-day for most
compounds or duration-adjusted concentrations >1,000 mg/m3 after less-than-chronic
exposures).
GASTROINTESTINAL EFFECTS
The //-heptane screening subchronic and chronic p-RfDs are based on analogue
read-across analysis using //-nonane as the analogue; forestomach lesions were the critical effect
in the study of //-nonane (U.S. HP A. 2016). No human studies examining gastrointestinal (GI)
effects of aliphatic low carbon range compounds were identified in the sources reviewed. Data
on GI effects of aliphatic low carbon range compounds in animals exposed by oral and inhalation
routes were limited, so exposure-response arrays are not developed for this endpoint.
Animal Studies
The subchronic and chronic oral p-RfDs for //-heptane are based on an analogue,
//-nonane (C9 [EC9]), and forestomach histopathology (hyperplasia and hyperkeratosis at doses
>100 mg/kg-day administered by gavage as neat compound 7 days/week) (U.S. HP A. 2016).
Irritation of the gastric mucosa was noted at both gross and microscopic examination of rats
exposed by gavage to 1-hexene (as neat compound) at doses >1,010 mg/kg-day for 4 weeks
(Dotti et al.. 1994 as cited in OECD. 2004). No histopathology findings were observed in the
stomach or large or small intestines of rats exposed to 2,4,4-trimethylpentene in maize oil at
doses up to 1,000 mg/kg-day for 4 weeks (U.S. HP A. 2015) or the C5-C7 alkene mixture in corn
oil at doses up to 1,000 mg/kg-day for 4-6 weeks (Springbom Laboratories. Inc.. 2003 as cited
60
Aliphatic low carbon range TPH fraction
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1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
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22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
EPA/690/R-22/007F
in OHCD. 2004). None of the remaining oral studies of compounds within the C5-C8 range
evaluated GI tract histopathology, and the only related data available were gross necropsy
findings in the stomach. In the unpublished version of the Haider et al. (1985) gavage study, API
(1985) reported grossly observed stomach changes including ulcers, edema, and reddened areas;
these effects were seen in 80-100% of the animals treated with each of the tested compounds in
the C5-C8 range (affected dose groups were not reported; duration-adjusted doses tested in the
study were 357 and 1,429 mg/kg-day). All of the compounds (including //-pentane,
2,3-dimethylbutane, 2-methylpentane, //-hexane, 2-methyl-2-pentene, methylcyclopentane,
2-methylhexane, 2,3-dimethylpentane, and 2,2,4-trimethylpentane) were administered neat
(without solvent) in that study.
No inhalation studies of aliphatic low carbon range compounds or mixtures have
identified GI effects. Studies that examined the GI tract for histopathology changes reported no
effects in rats after exposure for 4-13 weeks to //-pentane (U.S. HP A. 2009e). cyclopentane
(Kimmerle and Thvssen. 1975), 3-methylpentane (Chung et al.. 2016). methylcyclopentane
(Yang et al .. 2014). or //-octane (Sung et al.. 2010). or in mice exposed to //-hexane for 13 weeks
(U.S. EPA. 2005). Chronic (2-year) studies of commercial hexane in rats and mice exposed by
inhalation to duration-adjusted concentrations up to 5,639 mg/m3 also showed no
treatment-related histopathology in the GI tract (U.S. EPA. 2009b).
Summary of Potentially Relevant Gastrointestinal Evidence
Irritant responses in the GI tract were observed macroscopically in rats exposed by
gavage to neat alkanes in the C5-C8 range (Haider et al.. 1985). and forestomach histopathology
was seen in rats exposed by gavage to neat //-nonane (the analogue for //-heptane). API (1985)
and Haider et al. (1985) reported gross changes in the stomach collectively for the tested C5-C8
compounds as a group; thus, effect levels could not be determined. Histopathology changes were
not seen after inhalation exposure to compounds in the fraction, and histopathology evaluations
of the GI tract were lacking for most of the available oral studies. It appears from these
observations that oral exposure to undiluted members of the fraction may result in direct effects
on the GI tract. However, available data are not considered sufficient to evaluate the consistency
in GI effects and potencies across fraction members.
RESPIRATORY EFFECTS
Nasal and laryngeal lesions represent the critical effect for the chronic RfC for
commercial hexane (U.S. EPA. 2009b). No information on respiratory effects in humans exposed
to aliphatic low carbon range compounds or mixtures was identified in the sources reviewed.
Animal studies examining respiratory tract endpoints are available for nine compounds and two
mixtures (see Table 3); the preponderance of the animal data is from subchronic inhalation
studies.
Animal Studies
Only two of the available oral studies of compounds or mixtures relevant to the aliphatic
low carbon range fraction examined respiratory tract effects in animals, and no oral studies
examined nasal pathology. No histopathology changes were observed in the lungs of rats given
2,4,4-trimethylpentene at doses up to 1,000 mg/kg-day for 4 weeks (U.S. EPA. 2015) or in the
lungs or tracheas of rats given the C5-C7 alkene mixture at doses up to 1,000 mg/kg-day for
4-6 weeks (Springborn Laboratories. Inc.. 2003 as cited in OHCD. 2004). Due to the limited data
61
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and lack of effects, an exposure-response array is not presented for respiratory effects after oral
exposure.
Figure C-9 shows the exposure-response data for respiratory effects from studies of
animals exposed by inhalation. Animal studies in which the nasal cavity, nasal turbinates, and/or
larynx were examined after inhalation exposure include a 4-week rat study of 3-methylpentane;
subchronic rat and mouse studies of //-pentane, 1-hexene, //-hexane, methylcyclopentane, and
//-octane; and chronic studies of commercial hexane in rats and mice. In mice exposed to
>629 mg/m3 //-hexane for 13 weeks, nasal histopathology changes included inflammation,
erosion, regeneration, and metaplasia in the olfactory and/or respiratory epithelium (U.S. EPA.
2005). Nasal and laryngeal histopathology changes (hyperplasia of epithelial and goblet cells,
chronic inflammation, and increased incidence of intracytoplasmic eosinophilic material in nasal
turbinates; squamous metaplasia/hyperplasia of the columnar epithelium in the larynges) were
observed in rats after 2 years of exposure to commercial hexane concentrations >564 mg/m3
(U.S. EPA. 2009b). No histopathology changes in the nasal cavity, nasal turbinates, and/or
larynx were observed in rats exposed to //-pentane by inhalation for 13 weeks (Kim et at.. 2012).
3-methylpentane for 4 weeks (Chung et at.. 2016). 1 -hexene for 13 weeks (Gingett. 1999 as cited
in OECD. 2004). methylcyclopentane for 13 weeks (Yang et at.. 2014). or //-octane for 13 weeks
(Sung et at.. 2010). generally at concentrations exceeding 1,000 mg/m3.
Few reports of lower respiratory tract effects were located in the sources reviewed.
Enlargement of the air spaces in respiratory bronchioles and alveolar ducts and pulmonary
fibrosis, along with papillary tumors of nonciliated bronchial epithelial cells were observed in
rabbits exposed to //-hexane for 24 weeks at a concentration of 2,610 mg/m3 (U.S. EPA. 2005).
Gestational exposure studies of commercial hexane in rats and mice reported gross observations
of pulmonary color change in dams at 7,894 mg/m3 (U.S. EPA. 2009b). Other studies in rats or
mice reported no treatment-related effects on the lung or lower respiratory tract histopathology
after exposure to //-pentane, cyclopentane, 3-methylpentane, 1-hexene, methylcyclopentane,
cyclohexane, or //-octane for 4-13 weeks (see Figure C-9).
Summary of Potentially Relevant Respiratory Evidence
Respiratory effects consisting of nasal and/or laryngeal lesions were reported in animals
exposed to //-hexane and commercial hexane by inhalation, and limited evidence for bronchiolar
and pulmonary changes after exposure to these materials has been reported. LOAELs ranged
from 629 mg/m3 in mice to 2,517 mg/m3 in rabbits (see Figure C-9). Studies of other compounds
did not show effects on the upper and/or lower respiratory tract. Thus, respiratory effects have
not been consistently shown to be associated with oral or inhalation exposure to members of the
aliphatic low carbon range fraction.
62
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Mixtures
63
Aliphatic low carbon range TPH fraction
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EPA/690/R-22/007F
DEVELOPMENTAL EFFECTS
Developmental toxicity, manifested as reduced offspring weights, is the critical effect for
the sub chronic and chronic RfCs for cyclohexane (U.S. EPA. 2010. 2003). No human studies
were available to address the potential for developmental toxicity of the aliphatic low carbon
range total petroleum hydrocarbon (TPH) fraction. Animal studies of developmental toxicity are
available for seven compounds and two mixtures; most of the data are from inhalation studies.
Animal Studies
Developmental studies of aliphatic low carbon range compounds and mixtures in animals
exposed orally include teratogenicity studies of //-pentane and //-hexane, as well as combined
repeated-dose and reproductive/developmental screening studies in rats exposed to 1-hexene,
cyclohexene, 2,4,4-trimethylpentene, methylcyclohexane, ethylcyclohexane, or the C5-C7
alkene mixture. In mice exposed to //-hexane on GDs 6-15, fetal birth weights were decreased at
doses >7,920 mg/kg-day, but maternal mortalities also occurred at these doses (U.S. EPA. 2005).
No developmental effects were seen in rats exposed to //-pentane at doses up to 1,000 mg/kg-day
during gestation (U.S. EPA. 2009e). The screening reproductive and developmental toxicity
studies showed no developmental effects at doses up to 500 mg/kg-day (cyclohexene) (U.S.
EPA. 2012b) or 1,000 mg/kg-day (1 -hexene, 2,4,4-trimethylpentene, methylcyclohexane,
ethylcyclohexane, and the C5-C7 alkene mixture) (U.S. EPA. 2015; OECD. 2014; Gingett et at..
2000 and Springbom Laboratories. Inc.. 2003 as cited in OECD. 2004); however, these studies
included only limited developmental toxicity evaluations (some were limited to pup weight and
viability) and did not assess teratogenicity. Due to the limited data and absence of effects, an
exposure-response array is not presented for developmental effects after oral exposure.
Data on developmental toxicity in animals exposed by inhalation are available for
//-pentane, //-hexane, cyclohexane, and commercial hexane. //-Pentane has been studied only in a
screening-level developmental toxicity assay, while more complete developmental toxicity data
in two species are available for the remaining compounds, and two-generation reproductive
toxicity studies are available for cyclohexane and commercial hexane. Figure C-10 displays the
exposure-response information from these studies. In the screening-level study of //-pentane, no
effects on number of implantations, viable fetuses, or incidences of external malformations were
observed in rats exposed to concentrations up to 7,380 mg/m3 on GDs 6-15 (U.S. EPA. 2009e).
Decreased pup growth was observed in rats exposed to //-hexane during gestation to
concentrations >881 mg/m3 (duration-adjusted) and in mice exposed to 14,686 mg/m3 (U.S.
EPA. 2005). Increased incidences of skeletal variations were also reported in rats exposed to
14,686 mg/m3 //-hexane (U.S. EPA. 2005); this finding may have been influenced by decreased
fetal body weights at this exposure level. In mice exposed to //-hexane during gestation,
decreases in the number of live fetuses per litter were reported at concentrations >7,500 mg/m3
(Li et at.. 2015; Li et at.. 2014; U.S. EPA. 2005); a decrease in percent live implants and an
increase in the incidence of late resorptions were also seen at 14,686 mg/m3 (U.S. EPA. 2005).
64
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65
Aliphatic low carbon range TPH fraction
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Cyclohexane induced decreases in Fi and F2 pup weights (during lactation) at a
duration-adjusted concentration of 4,304 mg/m3 in a two-generation rat reproductive toxicity
study, while no effects on fetal weights or other measures of developmental toxicity were seen in
rats and rabbits exposed to cyclohexane at 6,025 mg/m3 during gestation [GDs 6-15 in rats or
GDs 6-18 in rabbits (U.S. EPA. 2010)1. A two-generation reproductive toxicity study of
commercial hexane also reported decreased Fi and F2 offspring weights (postnatal days
[PNDs] 14 and 7, respectively) in rats at a duration-adjusted concentration of 5,639 mg/m3 (U.S.
EPA. 2009b). Exposure to 7,985 mg/m3 commercial hexane had no effect on GD 21 fetal
weights or developmental toxicity endpoints in rats when exposure was limited to GDs 6-15
(U.S. EPA. 2009b). In mice exposed to commercial hexane at 7,895 mg/m3 during gestation, an
increase in the incidence of skeletal variations was seen in the absence of pup weight changes
(U.S. EPA. 2009b).
Summary of Potentially Relevant Developmental Evidence
Limited developmental toxicity data, which lack teratogenicity assessments, are available
for «-pentane, 1-hexene, cyclohexene, 2,2,4-trimethylpentene, and the C5-C7 alkene mixture.
More robust developmental toxicity data are available for //-hexane, cyclohexane, and
commercial hexane. The available oral and inhalation data suggest that //-hexane, cyclohexane,
and commercial hexane reduced body weights in rat offspring, while 1-hexene, cyclohexene,
2,4,4-trimethylpentene, methylcyclohexane, ethylcyclohexane, and the C5-C7 alkene mixture
did not. Exposure to //-hexane and commercial hexane via inhalation increased the incidences of
skeletal variations in rats and mice, respectively, when exposed during gestation, but
cyclohexane and //-pentane did not; data on skeletal variations and malformations were not
available for the remaining compounds. Only //-hexane exposure (by inhalation) has been shown
to affect embryonic or fetal viability. In summary, too few compounds have received rigorous
testing for developmental effects, so the available developmental toxicity data are not adequate
to assess consistency in effects or potencies of the compounds and mixtures in the fraction.
OTHER EFFECTS
New studies identified in the PubMed searches for //-hexane identified effects on ovarian
function in female mice exposed by inhalation. Liu et al. (2012) reported reduced egg production
and serum progesterone levels at duration-adjusted //-hexane exposure concentrations
>330 mg/m3 and decreases in diestrus duration and number of ovarian follicles after 5 weeks of
exposure (4 hours/day, 7 days/week). Alterations in the proportions of secondary and atretic
ovarian follicles, estrous cycle disruptions, and changes in the secretion of progesterone and
estradiol by cultured ovarian granulosa cells from exposed offspring were also reported in female
offspring of mice exposed to //-hexane during gestation (Li et al .. 2015; Li et al .. 2014). No other
studies of ovarian function in humans or animals exposed to aliphatic low carbon range
compounds or mixtures were located in the sources reviewed. Takcuchi et al. (1980)
66
Aliphatic low carbon range TPH fraction
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APPENDIX D. REFERENCES
API (American Petroleum Institute). (1985). Four-week oral nephrotoxicity screening study in
male F344 rats. (EPA Document No. FYI-AX-1085-0460). Washington, DC: U.S.
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ATSDR (Agency for Toxic Substances and Disease Registry). (1999). Toxicological profile for
Total Petroleum Hydrocarbons (TPH). In Govt Reports Announcements and Index (GRA
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ATSDR (Agency for Toxic Substances and Disease Registry). (2018). Framework for assessing
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Baxter. CS. (2012). Alicvclic hydrocarbons. In E Bingham; B Cohrssen (Eds.), Patty's
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automotive mechanics. Am J Epidemiol 183: 969-976.
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Calamur. N; Carrera. ME; Wilsak. RA. (2003). Butvlenes. In Kirk-Othmer Encyclopedia of
Chemical Technology. John Wiley & Sons, Inc.
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Campos-Ordonez. T; Zarate-Lopez. D; Galvez-Contreras. AY; Mov-Lopez. N; Guzman-Muniz.
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astrogliosis and microglial reactivity in the adult hippocampus mouse brain. Cell Mol
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Carreon. T; Hcrrick. RL. (2012). Aliphatic hydrocarbons. In Patty's toxicology: Volume 2 (6th
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Chung. YH; Shin. SH; Han. JH; Lee. YH. (2016). Subacute inhalation toxicity of 3-
methylpentane. Toxicological Research 32: 245-250.
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series: Volume 4: Development of fraction specific reference doses (RFDs) and reference
concentrations (RFCs) for total petroleum hydrocarbons (TPH). Amherst, MA: Amherst
Scientific Publishers.
Egan. G; Spencer. P; Schaumburg. H; Murray. KJ; Bischoff M; Scala. R. (1980). n-Hexane-
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EU (European Union). (2008). 2,4,4-Trimethylpentene risk assessment. CAS No: 25167-70-8.
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43
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the European Communities. European Chemicals Agency.
https://echa.europa.eu/documents/10162/10656235-2b4a-45dd-alc6-a00312c69bd5
Frontali. N; Amantini. MC; Spagnoio. A; Guard ni. AM; Saltari. MC; Brum one. F; Perbellini. L.
(1981). Experimental neurotoxicity and urinary metabolites of the C5-C7 aliphatic
hydrocarbons used as glue solvents in shoe manufacture. Clin Toxicol 18: 1357-1367.
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Galvin. IB; Marashi. F. (1999). Cvclopentane. CAS#287-92-3 [Review], J Toxicol Environ
Health A 58: 57-74.
Gargas. ML; Burgess. RJ; Voisard. DE; Cason. GH; Andersen. ME. (1989). Partition
coefficients of low-molecular-weight volatile chemicals in various liquids and tissues.
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Gomez. J; Basil. T. Chan. N. (1998). An overview of the use oxygenates in gasoline. California
Environmental Protection Agency. California Air Resources Board. Stationary Source
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Gustafson. JB; Griffith Tell. J; Orem. D. (1997). Total petroleum hydrocarbon criteria working
group series: Volume 3: Selection of representative TPH fractions based on fate and
transport considerations. Amherst, MA: Amherst Scientific Publishers.
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de=shopping&prd kev=146b7eaf-c92d-4e4b-aflf-092c5735al0b
Haider. CA; Holdsworth. CE; Cockrett. BY; Piccirillo. VJ. (1985). Hydrocarbon nephropathy in
male rats: Identification of the nephrotoxic components of unleaded gasoline. Toxicol Ind
Health 1: 67-87. http://dx.doi.org/10.1177/0748233785001003Q5
Hassani. S; Namvar. M; Ghoreishvandi. M; Attarchi. M; Golabadi. M; Sevedmehdi. SM;
Khodarahmian. M. (2014). Menstrual disturbances and hormonal changes in women
workers exposed to a mixture of organic solvents in a pharmaceutical company. 28: 156.
J-CHECK (Japanese CHEmical Collaborative Knowledge database). (2010a). J-CHECK
substance data: n-Heptane (CASRN 142-82-5). Available online at
https://www.nite. go. ip/chem/i check/template. action?ano=4682&mno=2-7&cno=142-82-
5&request locale en
J-CHECK (Japanese CHEmical Collaborative Knowledge database). (2010b). J-CHECK
substance data: n-Hexane (CASRN 110-54-3). Available online at
https://www.nite. go. ip/chem/icheck/template.action?ano=3294&mno=2-0006&cno=l 10-
54-3&request locale en
J-CHECK (Japanese CHEmical Collaborative Knowledge database). (2010c). J-CHECK
substance data: n-Pentane (CASRN 109-66-0). Available online at
https://www.nite.go.ip/chem/i check/template.action?ano=3193&mno=2-0005&cno=109-
66-0&request locale en
Jia. X; Liu. Q; Zhang. Y; Dai. Y; Duan. H; Bin. P; Niu. Y; Liu. J; Zhong. L; Guo. J; Liu. X;
Zheng. Y. (2014). Myelin protein zero and its antibody in serum as biomarkers of n-
hexane-induced peripheral neuropathy and neurotoxicity effects. Chin Med J 127: 1536-
1540. http://dx.doi.Org/10.3760/cma.i.issn.0366-6999.20140202
Jimenez-Garza. O; Guo. L; Bvun. HM; Carried. M; Bartolucci. GB; Barron-Vivanco- BS;
Baccaretti. AA. (2018). Aberrant promoter methylation in genes related to hematopoietic
malignancy in workers exposed to a VOC mixture. Toxicol Appl Pharmacol 339: 65-72.
http://dx.doi.Org/10.1016/i.taap.2017.12.002
Johnson. W. Jr; Bergfeld. WF; Belsito. DV; Hill. RA; Klaassen. CD; Liebler. D; Marks. JG. Jr;
Shank. RC; Slaga. TJ; Snyder. PW; Andersen. FA. (2012). Safety assessment of
68 Aliphatic low carbon range TPH fraction
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