f/EPA
EPA/635/R-08/003F
www.epa.gov/iris
TOXICOLOGICAL REVIEW
OF
PROPIONALDEHYDE
(CAS No. 123-38-6)
In Support of Summary Information on the
Integrated Risk Information System (IRIS)
September 2008
U.S. Environmental Protection Agency
Washington, DC
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DISCLAIMER
This document has been reviewed in accordance with U.S. Environmental Protection Agency
policy and approved for publication. Mention of trade names or commercial products does not
constitute endorsement or recommendation for use.
11
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CONTENTS—TOXICOLOGICAL REVIEW OF PROPIONALDEHYDE
(CAS No. 123-38-6)
LIST OF TABLES v
LIST OF FIGURES v
LIST OF ACRONYMS vi
FOREWORD vii
AUTHORS, CONTRIBUTORS, AND REVIEWERS viii
1. INTRODUCTION 1
2. CHEMICAL AND PHYSICAL INFORMATION 3
3. TOXICOKINETICS 5
3.1. ABSORPTION 5
3.1.1. Oral 5
3.1.2. Inhalation 5
3.2. DISTRIBUTION 6
3.3. METABOLISM 6
3.4. ELIMINATION 7
3.5. PHYSIOLOGICALLY BASED TOXICOKINETIC MODELS 7
4. HAZARD IDENTIFICATION 8
4.1. STUDIES IN HUMANS—EPIDEMIOLOGY, CASE REPORTS, CLINICAL
CONTROLS 8
4.2. SUBCHRONIC AND CHRONIC STUDIES AND CANCER BIO AS SAYS IN
ANIMALS—ORAL AND INHALATION 8
4.2.1. Oral Studies 8
4.2.2. Inhalation Studies 8
4.3. REPRODUCTIVE/DEVELOPMENTAL STUDIES-INHALATION 9
4.4. OTHER STUDIES 12
4.4.1. Genotoxicity 12
4.4.1.1. Bacteria 12
4.4.1.2. Mammalian Cells In Vitro 14
4.4. \.2.l.Mutagenicity 14
4.4.1.2.2. Chromosomal aberrations 17
4.4.1.2.3. DNA damage 18
4A.I.2A. Non-DNA adductformation 21
4.4.1.3. Genotoxicity Summary 21
4.4.2. Cardiovascular Effects 22
4.4.3. Immunotoxicity 24
4.4.4. Cytotoxicity 25
4.4.5. Comparative Toxicity of Related Aldehydes 25
4.5. SYNTHESIS AND EVALUATION OF MAJOR NONCANCER EFFECTS 29
4.5.1. Oral 29
4.5.2. Inhalation 30
iii
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4.6. WEIGHT-OF-EVIDENCE EVALUATION AND CANCER CHARACTERIZATION 32
4.6.1. Summary of Overall Weight of Evidence 32
4.7. SUSCEPTIBLE POPULATIONS AND LIFE STAGES 34
4.7.1. Possible Childhood Susceptibility 34
4.7.2. Possible Gender Differences 34
4.7.3. Possible Genetic Differences 34
4.7.4. Possible Sensitive Subgroups - Asthmatics 34
5. DOSE-RESPONSE ASSESSMENTS 36
5.1. ORAL REFERENCE DOSE (RfD) 36
5.2. INHALATION REFERENCE CONCENTRATION (RFC) 36
5.2.1. Choice of Principal Study and Critical Effect 36
5.2.2. Methods of Analysis 39
5.2.3. RfC Derivation—Including Application of Uncertainty Factors (UFs) 39
5.3. CANCER ASSESSMENT 42
5.4. GENERAL UNCERTAINTY IN THE PROPIONALDEHYDE NONCANCER AND
CANCER ASSESSMENT 42
6. MAJOR CONCLUSIONS IN THE CHARACTERIZATION OF HAZARD AND DOSE
RESPONSE 45
6.1. HUMAN HAZARD POTENTIAL 45
6.2. DOSE RESPONSE 47
7. REFERENCES 49
APPENDIX A. SUMMARY OF EXTERNAL PEER REVIEW AND PUBLIC COMMENTS
AND DISPOSITION A-l
APPENDIX B. BENCHMARK CONCENTRATION MODELING RESULTS B-1
IV
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LIST OF TABLES
Table 2-1. Chemical and physical properties of propionaldehyde 3
Table 4-1. Summary of nasal lesion incidence data in female and male rats exposed to various
concentrations of propionaldehyde 11
Table 4-2. Mutagenicity of various aldehydes in Salmonella typhimurium 13
Table 4-3. Mutagenicity of various aldehydes in mammalian cells 16
Table 4-4. Aldehyde-induced chromosome damage in mammalian cells in vitro 18
Table 4-5. Aldehyde-induced DNA damage in vitro 19
Table 4-6. Effects of inhalation of propionaldehyde on blood pressure and heart 23
Table 4-7. RD50 values for propionaldehyde and selected, related aldehydes measured in B6C3Fi
and Swiss-Webster mice 27
Table 4-8. Concentration [M] of selected aldehydes required to produce a 50% change from
control in each cytotoxic endpoint 29
Table 5-1. Propionaldehyde References for Exposure-Response Array 37
Table 5-2. Summary of general uncertainty in the propionaldehyde noncancer and cancer risk
assessments 42
Table B-l. Olfactory atrophy incidence data in male rats exposed to various concentrations of
propionaldehyde B-l
Table B-2. BMC model outputs for olfactory atrophy B-2
LIST OF FIGURES
Figure 5-1. Exposure-Response Array for Propionaldehyde 36
Figure B-l. BMCio Weibull model for olfactory atrophy (Union Carbide, 1993) B-2
Figure B-2. BMC05 Weibull model for olfactory atrophy (Union Carbide, 1993) B-3
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LIST OF ACRONYMS
ADJ
AIC
ALDH
ATPase
BMC
BMCL
BMD
BMDS
BMR
CASRN
CHED
CHO
DPX
EC50
EPA
FDA
GD
HEC
HGPRT/HPRT
IC50
IPCS
IRIS
i.v.
KC1
LD50
LOAEL
M, mM, jiM
MR
NLM
NOAEL
NOEL
ppb, ppm
PND
POD
RD50
RfC
RfD
RGDR
S9
SA
SCE
SDS
UDS
UF
VE
WHO/JECFA
dosimetrically adjusted
Akaike Information Criterion
aldehyde dehydrogenase
adenosine triphosphatase
benchmark concentration
95% lower confidence limit of the benchmark concentration
benchmark dose
benchmark dose software
benchmark response
Chemical Abstracts Service Registry Number
Chinese hamster embryonic diploid (cells)
Chinese hamster ovary (cells)
DNA protein cross-link
median effective concentration
U.S. Environmental Protection Agency
U.S. Food and Drug Administration
gestation day
human equivalent concentration
hypoxanthine-guanine phosporibosyltransferase
median inhibitory concentration
International Programme on Chemical Safety
Integrated Risk Information System
intravenous
potassium chloride
median lethal dose
lowest-observed-adverse-effect level
Molar (M), milli molar (10"3M), micro molar (10"6M)
molecular reactivity
National Library of Medicine, Hazardous Substances Database
no-observed-adverse-effect level
no-observed-effect level
parts per billion, parts per million
postnatal day
point of departure
concentration required to elicit a 50% decrease in respiratory rate.
reference concentration
reference dose
regional gas dose ratio
post mitochondrial microsomal liver fraction
surface area
sister chromosome exchanges
sodium-dodecyl sulfate
unscheduled DNA synthesis
uncertainty factor
ventilation rate
World Health Organization/Joint Expert Committee on Food Additives
VI
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FOREWORD
The purpose of this Toxicological Review is to provide scientific support and rationale
for the hazard and dose-response assessment in IRIS pertaining to chronic exposure to
propionaldehyde. It is not intended to be a comprehensive treatise on the chemical or
toxicological nature of propionaldehyde.
The intent of Section 6, Major Conclusions in the Characterization of Hazard and Dose
Response, is to present the major conclusions reached in the derivation of the reference dose,
reference concentration and cancer assessment, where applicable, and to characterize the overall
confidence in the quantitative and qualitative aspects of hazard and dose response by addressing
the quality of data and related uncertainties. The discussion is intended to convey the limitations
of the assessment and to aid and guide the risk assessor in the ensuing steps of the risk
assessment process.
For other general information about this assessment or other questions relating to IRIS,
the reader is referred to EPA's IRIS Hotline at (202) 566-1676 (phone), (202) 566-1749 (fax), or
(email address).
vn
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AUTHORS, CONTRIBUTORS, AND REVIEWERS
CHEMICAL MANAGER
John Stanek, Ph.D.
National Center for Environmental Assessment
U.S. Environmental Protection Agency
Research Triangle Park, NC
AUTHORS
John Stanek, Ph.D.
National Center for Environmental Assessment
U.S. Environmental Protection Agency
Research Triangle Park, NC
Ms. Susan Goldhaber
Mr. Frank Stack
Errol Zeiger, Ph.D.
Alpha-Gamma Technologies, Inc.
Raleigh, NC
EXECUTIVE DIRECTION
(U.S. Environmental Protection Agency; Office of Research and Development;
National Center for Environmental Assessment):
Ila Cote, Ph.D. (Acting RTF Division Director)
Debra Walsh, M.S. (Deputy Division Director)
Fred Dimmick, M.S.E. (Acting Branch Chief)
Lynn Flowers, Ph.D., DABT (Assistant Center Director for Risk Analysis)
John Vandenberg, Ph.D. (Associate Director for Health)
REVIEWERS
This document has been peer reviewed by EPA scientists, interagency reviewers from
other federal agencies, and the public, and peer reviewed by independent scientists external to
EPA. A summary and EPA's disposition of the comments received from the independent
external peer reviewers and from the public is included in Appendix A of the Toxicological
Review of Propionaldehyde.
INTERNAL EPA REVIEWERS
Ila Cote, Ph.D.
National Center for Environmental Assessment
Office of Research and Development
Lynn Flowers, Ph.D.
National Center for Environmental Assessment
Office of Research and Development
viii
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Gary Foureman, Ph.D.
National Center for Environmental Assessment
Office of Research and Development
Marion Hoyer, Ph.D.
Environmental Scientist
Office of Transportation and Air Quality
Nagu Keshava, Ph.D.
National Center for Environmental Assessment
Office of Research and Development
Connie Meacham, M.S.
National Center for Environmental Assessment
Office of Research and Development
D. Charles Thompson, Ph.D.
National Center for Environmental Assessment
Office of Research and Development
John Vandenberg, Ph.D.
National Center for Environmental Assessment
Office of Research and Development
Debra Walsh, M.S.
National Center for Environmental Assessment
Office of Research and Development
John Whalan, Ph.D.
National Center for Environmental Assessment
Office of Research and Development
EXTERNAL PEER REVIEWERS
John Morris, Ph.D. (Chair)
University of Connecticut School of Pharmacy
Andrew Salmon, Ph.D.
CA Office of Environmental Health Hazard Assessment
Jeffry Schroeter, Ph.D.
The Hamner Institutes for Health Sciences
Richard Schlesinger, Ph.D., Fellow ATS
Dept. of Biology and Health Sciences, Pace University
IX
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1. INTRODUCTION
This document presents background information and justification for the Integrated Risk
Information System (IRIS) Summary of the hazard and dose-response assessment of
propionaldehyde. IRIS Summaries may include oral reference dose (RfD) and inhalation
reference concentration (RfC) values for chronic and other exposure durations, and a
carcinogenicity assessment.
The RfD and RfC, if derived, provide quantitative information for use in risk assessments
for health effects known or assumed to be produced through a nonlinear (presumed threshold)
mode of action. The RfD (expressed in units of mg/kg-day) is defined as an estimate (with
uncertainty spanning perhaps an order of magnitude) of a daily exposure to the human
population (including sensitive subgroups) that is likely to be without an appreciable risk of
deleterious effects during a lifetime. The inhalation RfC (expressed in units of mg/m3) is
analogous to the oral RfD, but provides a continuous inhalation exposure estimate. The
inhalation RfC considers toxic effects for both the respiratory system (portal-of-entry) and for
effects peripheral to the respiratory system (extrarespiratory or systemic effects). Reference
values are generally derived for chronic exposures (up to a lifetime), but may also be derived for
acute (<24 hours), short-term (>24 hours up to 30 days), and subchronic (>30 days up to 10% of
lifetime) exposure durations, all of which are derived based on an assumption of continuous
exposure throughout the duration specified. Unless specified otherwise, the RfD and RfC are
derived for chronic exposure duration.
The carcinogenicity assessment provides information on the carcinogenic hazard
potential of the substance in question and quantitative estimates of risk from oral and inhalation
exposure may be derived. The information includes a weight-of-evidence judgment of the
likelihood that the agent is a human carcinogen and the conditions under which the carcinogenic
effects may be expressed. Quantitative risk estimates may be derived from the application of a
low-dose extrapolation procedure. If derived, the oral slope factor is a plausible upper bound on
the estimate of risk per mg/kg-day of oral exposure. Similarly, an inhalation unit risk is a
plausible upper bound on the estimate of risk per |ig/m3 air breathed.
Development of these hazard identification and dose-response assessments for
propionaldehyde has followed the general guidelines for risk assessment as set forth by the
National Research Council (NRC) (1983). EPA Guidelines and Risk Assessment Forum
Technical Panel Reports that may have been used in the development of this assessment include
the following: Guidelines for Mutagenicity Risk Assessment (EPA, 1986b), Recommendations for
and Documentation of Biological Values for Use in Risk Assessment (EPA, 1988), Guidelines for
Developmental Toxicity Risk Assessment (EPA, 1991), Methods for Derivation of Inhalation
Reference Concentrations and Application of Inhalation Dosimetry (EPA, 1994), Use of the
Benchmark Dose Approach in Health Risk Assessment (EPA, 1995), Guidelines for Reproductive
1
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Toxicity Risk Assessment (EPA, 1996), Guidelines for Neurotoxicity Risk Assessment (EPA,
1998), Science Policy Council Handbook: Risk Characterization (EPA, 2000b), Benchmark
Dose Technical Guidance Document (EPA, 2000c), A Review of the Reference Dose and
Reference Concentration Processes (EPA, 2002), Guidelines for Carcinogen Risk Assessment
(EPA, 2005a), Supplemental Guidance for Assessing Susceptibility from Early-Life Exposure to
Carcinogens (EPA, 2005b), Science Policy Council Handbook: Peer Review (EPA, 2006b), and
A Framework for Assessing Health Risks of Environmental Exposures to Children (EPA, 2006a).
The literature search strategy employed for this compound was based on the Chemical
Abstracts Service Registry Number (CASRN) and at least one common name. Any pertinent
scientific information submitted by the public to the IRIS Submission Desk was also considered
in the development of this document. The relevant literature was reviewed through June 2008.
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2. CHEMICAL AND PHYSICAL INFORMATION
Propionaldehyde is an aldehyde also known as propanal, propionic aldehyde,
methylacetaldehyde, propyl aldehyde, propaldehyde, and propylic aldehyde. Some relevant
chemical and physical properties are listed in Table 2-1.
Table 2-1. Chemical and physical properties of propionaldehyde
Propionaldehyde
CAS registry number
Empirical formula
Molecular weight
Vapor pressure
Vapor density
Boiling point
Melting point
Density /specific gravity
Solubilities
Viscosity
Octanol/water partition coefficient (as log P)
Auto ignition temperature
Conversion factors (in air)
o
^H
123-38-6
C3H60
58.08
3 17 mm Hg (at 25°C) (-400,000 ppm)
1.8(atlOO°F = 37.8°C)
49°C
-81°C
0.8657 (at 25°C)
Water = 3.06 x 105 mg/L at 25°C; soluble in
chloroform; miscible with alcohol and ether
0.3167cP(at26.7°C)
0.59
207°C
1 ppm = 2.38 mg/m3; 1 mg/m3 = 0.42 ppm
Sources: National Library of Medicine (NLM) (2004); International Programme on Chemical Safety (IPCS) (1993).
Propionaldehyde is a colorless liquid with a suffocating, fruity odor. It is used in the
manufacturing of propionic acid and polyvinyl and other plastics, in the synthesis of rubber
chemicals, and as a disinfectant and preservative. It is prepared by treating propyl alcohol with a
bichromate oxidizing mixture or by passing propyl alcohol vapor over copper at a high
temperature (NLM, 2004).
Propionaldehyde can form explosive peroxides and may polymerize with the addition of
acids, bases, amines, and oxidants, resulting in a fire or explosion hazard. It decomposes on
burning, producing toxic gases and irritating fumes (International Programme on Chemical
Safety [IPCS], 1993).
The chemical is released to the environment primarily through the combustion of wood,
gasoline, diesel fuel, and polyethylene (NLM, 2004). Propionaldehyde is also a component of
both mainstream and sidestream cigarette smoke (Counts et al., 2005). Municipal waste
incinerators can also release propionaldehyde to ambient air. In air, propionaldehyde is expected
to exist solely as a vapor; it may be degraded in the atmosphere by reaction with
photochemically produced hydroxyl radicals with a half-life of 19.6 hours for this reaction in air.
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Studies have indicated that propionaldehyde is readily biodegradable in wastewater, and its
potential for bioconcentration in aquatic organisms appears to be low (NLM, 2004).
Propionaldehyde has been detected in ambient and indoor air in several studies. Baez et
al. (2003) measured the concentrations of propionaldehyde in indoor and outdoor air in Mexico
to be 0.0002-0.018 mg/m3 and 0.0002-0.016 mg/m3, respectively. A North Carolina roadside
study of 23 hydrocarbons and 10 aldehydes reported that propionaldehyde accounted for
approximately 4% of the total aldehydes measured (Zweidinger et al., 1988). Propionaldehyde
was detected at concentrations <14 parts per billion (ppb) (0.014 parts per million [ppm] or
0.033 mg/m3) in Los Angeles air when measured during severe photochemical pollution episodes
(Grosjean, 1982) and at concentrations ranging from 0.007-0.025 ppm (0.017-0.06 mg/m3) in
the exhaust from a jet airplane, measured at 50 meters behind the engine at an idle power setting
(Miyamoto, 1986).
Propionaldehyde has also been approved by both the U.S. Food and Drug Administration
(FDA) and World Health Organization/Joint Expert Committee on Food Additives
(WHO/JECFA) as a synthetic flavoring ingredient for direct addition to food; the alcohol
(propanol) and acid (propionic acid) are similarly approved (FDA, 2003; WHO, 1999; IPCS,
1998). Propionaldehyde was determined to pose no safety concern since its expected oral intake
(140 jig/day) is below the threshold for human intake (1800 |ig/day, as defined by WHO) and it
is oxidized to propionic acid, which is metabolized via the citric acid cycle (WHO, 1999; IPCS,
1998).
Limited information is available on the occurrence of propionaldehyde in water. In the
National Organics Reconnaissance Survey conducted in the 1970s, propionaldehyde was found
to be one of the 18 organic chemicals detected most frequently in the drinking water of the
10 cities surveyed (Bedding et al., 1982).
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3. TOXICOKINETICS
There are a limited number of published studies on the toxicokinetics of
propionaldehyde. The absorption of propionaldehyde in the respiratory tract of dogs has been
measured after inhalation exposure. The metabolism of propionaldehyde via aldehyde
dehydrogenase (ALDH) (NADP- and NAD-dependent) has been investigated in rodent hepatoma
cell lines. The distribution and localization of ALDH in rat respiratory tract tissues, and presence
in human tissues, have also been examined. The urinary elimination of propionaldehyde formed
via lipid peroxidation has been examined in rats.
3.1. ABSORPTION
3.1.1. ORAL
There are no studies available examining the absorption or the bioavailability of
propionaldehyde via the oral route of exposure.
3.1.2. INHALATION
Egle (1972a) reported the regional retention levels (percent of amount inhaled) in the
respiratory tract of mongrel dogs of both sexes after exposure to concentrations ranging from
0.4-0.6 |ig/mL (403-604 mg/m3 or 168-252 ppm) propionaldehyde via nasal inhalation through
a fitted mask. Retention levels of propionaldehyde were measured for the total respiratory tract
as well as for the surgically isolated upper and lower respiratory tracts. Ventilatory rates were
varied, ranging from 6 to 20/minute. Neither detailed information on the inspiratory flow rates
nor the time period of exposure were reported. Average retention levels were reported from 6-20
experiments, with at least four dogs per experiment exposed to propionaldehyde. The retention
of propionaldehyde by the total respiratory tract was between 70 and 80%, and there was a
significant inverse relationship between retention and ventilation rate (p < 0.01). Retention of
propionaldehyde in the isolated upper respiratory tract under cyclic breathing conditions also
averaged 70-80% with a significant effect of ventilation rate (p < 0.01). However, under
unidirectional breathing conditions, retention in the isolated upper respiratory tract averaged
approximately 63% over the range of ventilation rates. In the lower respiratory tract,
propionaldehyde retention averaged between 65 and 75% with a significant inverse relationship
between retention and ventilation rate (p < 0.01). No effect of exposure concentration on total
respiratory tract retention was noted in animals exposed over a concentration range of 0.4-
1.2 |ig/mL (403-1,200 mg/m3 or 168-500 ppm) propionaldehyde. Variation in tidal volume over
a range of 100-200 ml was also without affect on propionaldehyde retention.
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3.2. DISTRIBUTION
Based on its physical-chemical properties, propionaldehyde likely crosses biological
membranes and thus could distribute throughout various bodily fluids. However, no specific
studies are available that describe the distribution of propionaldehyde.
3.3. METABOLISM
Propionaldehyde is oxidized to its corresponding carboxylic acid (i.e., propionic acid) via
ALDH (NADP- and NAD-dependent) (Bassi et al., 1997). The metabolisms of propionaldehyde
and three other aldehydes (acetaldehyde, benzaldehyde, and valeraldehyde) were examined in
two metabolically competent rodent hepatoma cell lines. Propionaldehyde, as well as the other
aldehydes tested, was efficiently metabolized in the rat hepatoma cell line. In the mouse
hepatoma cell line, low enzyme activities were observed. The authors concluded that the
differences in the metabolic activities between these two cell lines could be attributed to greater
oxidative activity in the rat cell line and greater reductive than oxidative activity in the mouse
cell line.
Respiratory tract tissues of both rats and humans contain ALDH (Zhang et al., 2005;
Stanek and Morris, 1999; Bogdanffy et al., 1998,1986; Morris, 1997; Casanova-Schmitz et al.,
1984). Acetaldehyde metabolism rates have been measured directly in rat nasal tissue
homogenates (Stanek and Morris, 1999; Morris, 1997; Casanova-Schmitz et al., 1984). These
studies identified a low-affinity, high-capacity isozyme, as well as a high-affinity, low-capacity
isozyme. In the rat, the distribution and localization of ALDH in the respiratory tract has been
examined (Bogdanffy et al., 1986). ALDH activity examined qualitatively was detected
principally in the nasal respiratory epithelium, while low activity was observed in the olfactory
epithelium. Epithelial cells of the trachea also demonstrated little enzyme activity; however, the
Clara cells of the bronchioles showed high enzyme activity. The authors noted that the pattern of
lower enzyme activity and localization correlated with the pattern of lesion distribution observed
after exposure to acetaldehyde, which is most notable in the olfactory epithelium. Bogdanffy et
al. (1998) also compared the enzyme activities of ALDH and carboxyl esterase in rat and human
nasal tissues for vinyl acetate. Enzyme activities were measured indirectly via the disappearance
of vinyl acetate or acetaldehyde form the headspace of incubation vials containing nasal tissue.
Rat respiratory epithelium ALDH activity was approximately twofold higher than that of humans
but was equivalent in the olfactory epithelium. Km values did not differ between species. In
addition, the presence of ALDH in fetal and adult human nasal tissues has been confirmed by
using gene expression analysis (Zhang et al., 2005).
Additionally, the Krebs (citric acid or tricarboxylic acid) cycle is thought to play a role in
the metabolism of aldehydes after oxidation to their corresponding carboxylic acids. After oral
intake, the Krebs cycle is expected to efficiently metabolize a number of aldehydes used as food
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additive flavoring agents (WHO, 1999). For propionaldehyde, its metabolite, propionic acid, is
also the end product of the metabolism of odd chain fatty acids via the p-oxidation pathway.
Propionic acid reacts with coenzyme A to form propionyl-CoA, which enters the Krebs cycle
after conversion to succinyl-CoA via methylmalonyl-CoA (Stipanuk, 2000; Voet and Voet,
1990). Succinyl-CoA is an intermediate in the Krebs cycle. In comparison, acetic acid, the
metabolite of acetaldehyde, condenses with coenzyme A. This complex undergoes P-oxidation to
form acetyl-CoA. Acetyl-CoA can enter the Krebs cycle directly or be used anabolically in fatty
acid and cholesterol synthesis (Voet and Voet, 1990). The fate of formic acid, formed by the
oxidation of formaldehyde via formaldehyde dehydrogenase, includes binding to tetrahydrofolic
acid, which is used in transmethylation reactions and as a source of single carbon additions
(Stipanuk, 2000; Voet and Voet, 1990).
Wang et al. (2002) performed a genotype analysis of the ALDH2 gene in the livers of
human volunteers in order to investigate the metabolism of a variety of aldehydes. Of a total of
39 subjects, 8 were heterozygotes of the wild-type (ALDH2*1) and mutant (ALDH2*2) alleles,
and the others were homozygotes of the wild-type allele. The ability of mitochondria to
metabolize propionaldehyde was significantly (p < 0.05) lower (80% for propionaldehyde) in the
heterozygotes (ALDH2*l/*2) compared to the homozygotes (ALDH2*1/*1), showing
differences in metabolism between the two genotypes.
Oyama et al. (2007) evaluated the inhalation toxicity of acetaldehyde in ALDH2
knockout (KO) mice. Male C57BL/6 wild-type and KO mice were exposed to 0, 125, or 500
ppm acetaldehyde 24 hours/day for 14 days via whole body-inhalation. Although the average
blood acetaldehyde concentration was greater in the KO mice compared to the wild-type, no
differences in liver and lung effects between the two groups were noted. However, the incidence
of erosion of the respiratory epithelium and subepithelial hemorrhage in the nasal cavity, and
degeneration of the respiratory epithelium in the larynx, pharynx, and trachea were greater in the
KO mice compared to the wild-type. These results indicate that the ALDH2 KO mice are more
sensitive to acetaldehyde-induced effects.
3.4. ELIMINATION
No information specific to the elimination of administered propionaldehyde is available.
De Tata et al. (2001) reported age-related effects in the urinary excretion of aldehydes formed
via lipid peroxidation in male Sprague-Dawley rats fed either a normal ad libitum diet or kept on
a restricted diet (every other day feeding, or 40% caloric restriction). The results showed that the
urinary excretion of propionaldehyde increased with age between 6 and 27 months and was
higher in animals on a restricted diet compared with animals fed ad libitum.
3.5. PHYSIOLOGICALLY BASED TOXICOKINETIC MODELS
No physiologically based toxicokinetic models were identified for propionaldehyde.
7
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4. HAZARD IDENTIFICATION
4.1. STUDIES IN HUMANS—EPIDEMIOLOGY, CASE REPORTS, CLINICAL
CONTROLS
No studies in humans were identified for propionaldehyde.
4.2. SUBCHRONIC AND CHRONIC STUDIES AND CANCER BIOASSAYS IN
ANIMALS—ORAL AND INHALATION
4.2.1. ORAL STUDIES
No subchronic or chronic oral studies were identified for propionaldehyde. However, the
structurally-related aldehydes, formaldehyde and acetaldehyde, appear to be less hazardous by
the oral route compared to the inhalation route (Morris et al., 1996).
4.2.2. INHALATION STUDIES
No subchronic or chronic inhalation studies were identified for propionaldehyde. In a
short-term study, Gage (1970) exposed four male and four female Alderley-Park rats to
1300 ppm (3,094 mg/m3) propionaldehyde for 6 hours/day for 6 days via whole-body inhalation.
Urine was collected after the last exposure for analysis of pH, bilirubin, and protein. Blood was
collected during sacrifice for analysis of hemoglobin concentration, blood differential counts,
platelets, clotting function, and the concentration of urea, sodium, and potassium. The lungs,
liver, kidneys, spleen, and adrenals; and occasionally, the heart, intestines, and thymus were
collected for microscopic examination. No changes in body weight were noted. At autopsy,
histological examination of all principal organs and tissues revealed liver cell vacuolation. No
other findings were noted. Four male and four female rats were also exposed to 90 ppm
(214 mg/m3) for 6 hours/day for 20 days. All organs were reported to be normal at autopsy, and
no clinical signs of toxicity were noted. Thus, a no-observed-effect level (NOEL) of 90 ppm can
be derived from this study.
In a short duration inhalation study, Steinhagen and Barrow (1984) determined the
concentration of propionaldehyde required to elicit a 50% decrease in respiratory rate (RD50) as a
measure of sensory irritation potential of propionaldehyde in B6C3Fi and Swiss-Webster mice.
Groups of three to four mice per strain were exposed via inhalation in a head-only exposure
chamber for 10 minutes to varying concentrations of propionaldehyde. Respiratory rates were
measured by a method in which animals were sealed in airtight plethysmographs and attached to
a head-only exposure chamber, and concentration-response curves were constructed to determine
the RDso. In animals, sensory irritants produce a reflex decrease in respiratory rate characterized
as a pause at the onset of expiration. The RD50 for propionaldehyde was calculated to be 2,078
ppm or 4,946 mg/m3 in B6C3Fi mice and 2,052 ppm or 4,884 mg/m3 in Swiss-Webster mice.
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4.3. REPRODUCTIVE/DEVELOPMENTAL STUDIES-INHALATION
Two short-term rat developmental inhalation studies were conducted by Union Carbide
(1993, 1991).l In a range-finding study, young adult female CD rats (seven per group) were
exposed to 0, 500, 1,000, 1,500, or 2,500 ppm (0, 1,190, 2,380, 3,570, or 5,950 mg/m3)
propionaldehyde for 6 hours/day via whole-body inhalation on gestation days (GDs) 0-20,
following successful mating with naive males (Union Carbide, 1991). Clinical observations were
made daily following the exposure, and maternal body weights were measured on GDs 0, 7, 14,
and 21. Food consumption was measured weekly throughout the study. At sacrifice on GD 21,
the dams were evaluated for liver and uterine weights, number of corpora lutea, and number and
status of implantation sites. Fetuses were dissected from the uterus, weighed, and examined
externally for malformations and variations. The pregnancy rate was equivalent among the
groups. None of the groups displayed any exposure-related clinical signs. Maternal toxicity was
noted as exposure-related differences in body weight gain, which were 82 and 72% (-28.9 and
-43.3 g, respectively,/? < 0.01) of control over the entire gestation period at exposure
concentrations of 1,500 and 2,500 ppm. At 1,000 ppm, body weight gain was depressed only
during the first week of exposure. However, these decreases in body weight gain were
accompanied by statistically significant decreases in food consumption compared those of
controls (p < 0.05) throughout the gestation period at 1,000, 1,500, and 2,500 ppm. The average
food consumption ranged from 82-89% of control at these exposure concentrations. None of
these effects were noted at 500 ppm. In addition, there were no exposure-related differences in
gestational parameters, including total number of implants and the number of viable and
nonviable implants. In the high exposure group, there was a significant reduction in fetal body
weights of approximately 12% (-0.6 g) compared with controls (p < 0.01), but no other evidence
of any treatment-related external malformations or variations was observed. The results of this
study indicate a no-observed-adverse-effect level (NOAEL) for developmental toxicity of 1,500
ppm. Indications of maternal effects (i.e. changes in body weight gain) related to
propionaldehyde exposure were most notable at 2,500 ppm.
In the second study, young adult male and female CD rats (15/sex/group) were exposed
to 0, 150, 750, or 1,500 ppm (0, 357, 1,785, or 3,570 mg/m3) propionaldehyde for 6 hours/day,
7 days/week via whole-body inhalation, during a 2-week premating period and a 14-day
(maximum) mating phase (Union Carbide, 1993). The mated females were exposed daily
through GD 20 only for a minimum of 35 days and a maximum of 48 days depending upon when
they mated (average exposure period -38 days). The females were then allowed to deliver their
litters naturally and raise their offspring until postnatal day (PND) 4 both free of exposure to
1 The Union Carbide studies (1991 and 1993) are unavailable in the peer-reviewed literature. These unpublished
studies were submitted to EPA under the Toxic Substances Control Act. An external peer review was conducted to
evaluate the accuracy of experimental procedures, results, and interpretation and discussion of the findings
presented. See References for more information.
-------
propionaldehyde. The males continued to be exposed for a total of 52 exposures until sacrifice in
week 7. Clinical observations were made daily, following exposure, and body weight and food
consumption were measured at regular intervals throughout the study. Offspring body weight,
viability, and disposition were monitored from birth until PND 4. Following the last exposure,
males were fasted and blood samples were obtained for clinical pathology analyses prior to
necropsy. On PND 4, necropsies were performed on adult females, and a number of organs and
tissues, including at least two sections of the nasal cavity (sectioning details not provided), were
examined histologically. The offspring were examined externally and sacrificed without
pathologic evaluation.
No exposure-related clinical signs were noted in the adult females. During the first week
of exposure to 750 and 1,500 ppm, body weight gains were decreased to approximately 60 and
71% (p < 0.01), respectively, of controls, and food consumption was decreased by approximately
7% (p < 0.05) of controls at both concentrations. No differences were observed during the
second week of exposure. During gestation, body weight (over GDs 0-14) and food consumption
(over GDs 0-21) were decreased in the high exposure group compared with controls, but no
significant differences in body weight gain were observed. At sacrifice, no gross lesions
attributable to propionaldehyde exposure were found. However, microscopic examination of the
nasal cavity revealed propionaldehyde-induced vacuolization of the olfactory epithelium in the
150 and 750 ppm exposure groups and atrophy of the olfactory epithelium in the 750 and
1,500 ppm exposure groups. These effects were noted to be localized to the dorsal anterior two
sections of the nasal cavity. The incidence of atrophy was 0/15, 0/15, 2/15, and 15/15 at 0, 150,
750, and 1,500 ppm, respectively (see Table 4-1). The severity of this nasal lesion increased with
exposure concentration being minimal to mild at 750 ppm and moderate to marked at 1,500 ppm.
No evidence of squamous metaplasia was found in olfactory or respiratory epithelium. Low
incidences of minimal to mild rhinitis involving the respiratory epithelium were also noted at
150, 750, and 1,500 ppm. No significant effects of exposure on any of the reproductive
parameters assessed were found. Litter size and viability were similar among the groups. Pup
body weights on the day of birth and PND 4 were not affected by exposure, although at the high
concentration only body weight gain for that period was significantly depressed (p < 0.05, -0.8
g) compared with controls. The biological significance of this finding is difficult to assess since
changes in absolute body weight were not demonstrated and the time period of observation was
relatively short.
The adult males did not display any overt signs of toxicity at any time during the study.
Body weight, weight gain, clinical observation, and food consumption were similar among all
exposure groups and controls. Hematology and clinical chemistry analyses revealed elevated
erythrocyte counts, with a corresponding increase in hemoglobin and hematocrit values and an
increase in monocytes in the males exposed to 1,500 ppm. These effects were considered to be
consistent with and indicative of dehydration. At necropsy (examination performed as per the
10
-------
adult females), no gross lesions were found that could be attributable to propionaldehyde
exposure. However, similar to effects in the females, microscopic examination revealed
exposure-related effects in the olfactory epithelium of the nasal cavity that consisted of
vacuolization and atrophy in the low, intermediate, and high exposure groups. These effects were
also noted to be localized to the dorsal anterior two sections of the nasal cavity. The incidence of
atrophy was 0/15, 2/15, 10/15, and 15/15 at 0, 150, 750, and 1,500 ppm, respectively (see
Table 4-1).
Table 4-1. Summary of nasal lesion incidence data in female and male rats
exposed to various concentrations of propionaldehyde
Group
Females3
Males3
Nasal lesion
Vacuolization - Olfactory
minimal
mild
moderate
Atrophy - Olfactory
minimal
mild
moderate
marked
Necrosis - Respiratory
moderate
Rhinitis - Respiratory
minimal
mild
Vacuolization - Olfactory
minimal
mild
moderate
marked
Atrophy - Olfactory
minimal
mild
moderate
marked
Squamous metaplasia -
Respiratory
mild
moderate
Rhinitis - Respiratory
minimal
mild
moderate
0
0/15
0
0
0
0/15
0
0
0
0
0/15
0
0/15
0
0
0/15
0
0
0
0
0/15
0
0
0
0
0/15
0
0
0/15
0
0
0
Exposure
150
15/15b
8
7
0
0/15
0
0
0
0
0/15
0
1/15
1
0
12/15b
6
4
2
0
2/15
2
0
0
0
0/15
0
0
0/15
0
0
0
concentration (ppm)
750
15/15b
0
7
8
2/15
1
1
0
0
0/15
0
6/15c
0
6
14/15b
2
o
J
2
7
10/15b
1
6
3
0
1/15
1
0
7/1 5b
1
5
1
1,500
0/15
0
0
0
15/15b
0
0
6
9
1/15
1
1/15
0
1
2/15
0
0
0
2
15/15b
0
1
8
6
2/15
0
2
14/15b
3
7
4
"Females were exposed daily only until GD 20 and sacrificed on PND 4; males were exposed daily until
sacrifice. See Section 4.3 for details
bSignificantly different from control atp < 0.01.
cSignificantly different from control atp < 0.05.
Source: Union Carbide (1993).
11
-------
The severity of this nasal lesion increased with exposure concentration being minimal at
150 ppm, minimal to moderate at 750 ppm, and mild to marked at 1,500 ppm. Squamous
metaplasia of the respiratory epithelium was reported in one male from the 750 ppm group and
two males from the 1,500 ppm group. An increased incidence of minimal to moderate rhinitis
involving the respiratory epithelium was also noted at 750 and 1,500 ppm. The results of this
study indicate a lowest-observed-adverse-effect level (LOAEL) for portal-of-entry toxicity of
150 ppm as a result of olfactory atrophy graded by Union Carbide (1993) as being of minimal
severity by the study authors and supported by the presence of vacuolization.
4.4. OTHER STUDIES
4.4.1. GENOTOXICITY
A number of other structurally related aldehydes, including acetaldehyde, formaldehyde,
butyraldehyde (butanal), and isobutyraldehyde (isobutanal) were evaluated concurrently for their
genotoxic potential. The results of these other aldehydes tested for similar genotoxic endpoints
are included in the evaluation of propionaldehyde for comparative purposes where available.
However, the results and comparisons presented for these aldehydes do not provide an
examination of the aldehyde database as a whole. No in vivo studies examining the genotoxicity
of propionaldehyde are available.
4.4.1.1. Bacteria
The mutagenicity test results for nonmammalian systems are summarized in Table 4-2.
Propionaldehyde was found to be mutagenic in Salmonella typhimurium strain TA1534 at
incubation concentrations >20 mM, but nonmutagenic in strains TA1950 and TA1952 at
concentrations up to 50 mM (Sampson and Bobik, 2008). Propionaldehyde was found to be
nonmutagenic in strains TA98, TA100, TA1535, and TA1537 when tested at concentrations up
to 10 mg/plate in the preincubation procedure with or without rat or hamster liver
postmitochondial microsonal liver fraction S9 (Aeschbacher et al., 1989; Mortelmans et al.,
1986) or when tested in strains TA100, TA102, and TA104 in the presence or absence of rat or
mouse liver S9 (Dillon et al., 1998; Aeschbacher et al., 1989). It was also nonmutagenic in
strains TA100, TA102, and TA104, when tested as a vapor in a desiccator at concentrations up to
3.3% in air with or without rat or mouse liver S9 (Dillon et al., 1998). In a plate test procedure,
propionaldehyde was not mutagenic in strain TA1535 at concentrations up to 2.5 (imol/plate
(equivalent to 145 jig/plate) with or without rat liver S9 (Pool and Wiessler, 1981).
Acetaldehyde was also found to be nonmutagenic in S. typhimurium strains TA98,
TA100, TA1535, and TA1537 when tested at concentrations <10 mg/plate in a preincubation
procedure with or without rat or hamster liver S9 (Mortelmans et al., 1986) or when tested in
strains TA98, TA100, and TA102 at concentrations up to 1.7 mmol/plate with or without rat liver
12
-------
S9 (Aeschbacher et al., 1989). It was nonmutagenic in strains TA100 and TA104 when tested at
concentrations <1 mL/desiccator chamber with or without rat or mouse S9, but an equivocal
response was seen in strain TA102 at 1 mL/desiccator chamber in the presence of rat liver S9
(Dillon et al., 1998). In a plate test procedure, acetaldehyde was not mutagenic in strain TA1535
when tested at concentrations up to 2.5 jimol/plate with or without rat liver S9 (Pool and
Wiessler, 1981).
Table 4-2. Mutagenicity of various aldehydes in Salmonella typhimurium
Aldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Acetaldehyde
Acetaldehyde
Acetaldehyde
Acetaldehyde
Acetaldehyde
Acetaldehyde
Formaldehyde
Formaldehyde
Formaldehyde
Formaldehyde
Strains
TA 1534
TA 1950
TA 1952
TA98, 100,
1535, 1537
TA98, 100,
102
TA100,
102, 104
TA100,
102, 104
TA1535
TA98, 100,
1535, 1537
TA98, 100,
102
TA100,
102, 104
TA100,
104
TA102
TA1535
TA100
TA98,
1535, 1537
TA100,
102, 104
TA1535
Protocol
Preincubation
Preincubation
Preincubation
Preincubation
"Modified"
preincubation
Preincubation
Vapor in
desiccator
Plate test
Preincubation
"Modified"
preincubation
Preincubation
Vapor in
desiccator
Vapor in
desiccator
Plate test
Preincubation
Preincubation
Preincubation
Plate test
S9, species
None
None
None
None, rat,
hamster
None, rat
None, rat,
mouse
None, rat,
mouse
None, rat
None, rat,
hamster
None, rat
None, rat,
mouse
None, rat,
mouse
Rat
None, rat
None, rat,
hamster
None, rat,
hamster
None, rat,
mouse
None, rat
Result3
+
-
-
-
-
-
-
-
-
-
-
-
?
-
+
-
+
-
LED (HTD)b
20mM
50 mM
50 mM
10 mg/plate
0.13 mmol/plate
(7.5 mg/plate)
10 mg/plate
3. 3% in air
2.5 |imol/plate
(145 |ig/plate)
10 mg/plate
1.7 mmol/plate
(75 mg/plate)
N/A (toxic
level)
1.0 mL/
desiccator
1.0 mL/
desiccator
2.5 |imol/plate
(110 |ig/plate)
10 jig/plate
333 jig/plate
15 |ig/plate
2.5 |imol/plate
(75 |lg/plate)
Reference
(Sampson and
Bobik, 2008)
(Sampson and
Bobik, 2008)
(Sampson and
Bobik, 2008)
(Mortelmans et
al., 1986)
(Aeschbacher
etal., 1989)
(Dillon et al.,
1998)
(Dillon et al.,
1998)
(Pool and
Wiessler, 1981)
(Mortelmans et
al., 1986)
(Aeschbacher
etal., 1989)
(Dillon etal.,
1998)
(Dillon etal.,
1998)
(Dillon etal.,
1998)
(Pool and
Wiessler, 1981)
(Haworth et al.,
1983)
(Haworth et al.,
1983)
(Dillon etal.,
1998)
(Pool and
Wiessler, 1981)
13
-------
Aldehyde
Butyraldehyde
Butyraldehyde
Butyraldehyde
Isobutyraldehyde
Isobutyraldehyde
Strains
TA98, 100,
1535, 1537
TA100,
102, 104
TA1535
TA98, 100,
1535, 1537
TA100,
102, 104
Protocol
Preincubation
Preincubation
Plate test
Preincubation
Preincubation
S9, species
None, rat,
hamster
None, rat,
mouse
None, rat
None, rat,
hamster
None, rat,
mouse
Result3
-
-
-
-
-/?
LED (HTD)b
3,333 Jig/plate
1000 Jig/plate
2.5 |imol/plate
(180 ng/plate)
10,000 jig/plate
5,000 jig/plate
Reference
(Mortelmans et
al., 1986)
(Dillon etal.,
1998)
(Pool and
Wiessler, 1981)
(Mortelmans et
al., 1986)
(Dillon etal.,
1998)
aTest results are either positive (+), negative (-), or equivocal (?).
^ED is the lowest effective concentration for positive test results;
negative or inconclusive results. N/A = not applicable.
HTD is the highest tested concentration for
Formaldehyde was mutagenic in S. typhimurium strain TA100 when preincubated with
rat and hamster S9 at concentrations between 10 and 100 jig/plate and weakly mutagenic without
S9 (Haworth et al., 1983). It was also found to be mutagenic in strains TA100, TA102, and
TA104 when tested over a concentration range of 6.25-50 jig/plate with and without rat and
mouse liver S9 (Dunnett's test; no statistical values nor effective concentrations reported) (Dillon
et al., 1998). Formaldehyde was not mutagenic in strains TA98, TA1535, or TA1537 when
tested at concentrations up to 333 jig/plate under the same conditions (Haworth et al., 1983) (no
statistical evaluation). Formaldehyde was not mutagenic in strain TA1535 when tested at
concentrations up to 2.5 jimol/plate (75 jig/plate) by using a plate test procedure with and
without rat liver S9 (Pool and Wiessler, 1981).
Butyraldehyde was nonmutagenic in S. typhimurium strains TA98, TA100, TA1535, and
TA1537 when tested at concentrations up to 3,333 jig/plate with rat and hamster liver S9 in a
preincubation procedure (Mortelmans et al., 1986). Butyraldehyde was also nonmutagenic in
strains TA100, TA102, and TA104 when tested at concentrations <1,000 jig/plate in the presence
and absence of rat or mouse liver S9 (Dillon et al., 1998). It was not mutagenic in TA1535 when
tested up to 2.5 (imol/plate (180 jig/plate) with and without rat liver S9 and using a plate test
procedure (Pool and Wiessler, 1981).
Similarly, isobutyraldehyde was nonmutagenic in S. typhimurium strains TA98, TA100,
TA1535, and TA1537 when tested at concentrations up to 10,000 jig/plate with rat and hamster
liver S9 in a preincubation procedure (Mortelmans et al., 1986). Isobutyraldehyde produced
equivocal responses in strains TA100, TA102, and TA104, but was judged to be nonmutagenic
overall when tested at concentrations between 50-5,000 jig/plate in the presence and absence of
rat or mouse liver S9 (Dillon et al., 1998).
4.4.1.2. Mammalian Cells In Vitro
4.4.1.2.1. Mutagenicity.
Propionaldehyde produced a concentration-related increase in HGPRT and ouabain
14
-------
mutants in V79 hamster cells following a 60-minute exposure over a concentration range of 3-90
mM. The increase in HGPRT mutants was significant (p < 0.01 versus controls) at 30 and 90
mM, and the increase in ouabain mutants was significant at 10, 30, and 90 mM (equivalent to
0.58, 1.7, and 5.2 mg/mL) (Brambilla et al., 1989). However, these increases were associated
with significant decreases in cell viability at > 30 mM in hypoxanthine-guanine phosphoribosyl
transferase (HGPRT) and at 90 mM in ouabain mutants. In a subsequent study, propionaldehyde
was not mutagenic at the HGPRT locus in V79 hamster cells exposed to 1 or 2 jiM (equivalent to
0.058 or 0.12 |ig/mL) for 2 hours; toxicity was seen at 2 |iM (Smith et al., 1990).
Acetaldehyde was found to induce mutations in the HPRT locus in cultured human
lymphocytes (He and Lambert, 1990). Cells treated with 1.2-2.4 mM acetaldehyde for 24 hours
or 0.2-0.6 mM acetaldehyde for 48 hours showed a 3- to 16-fold increase in mutant frequency.
This effect was accompanied by a dose-dependent decrease in cell survival.Formaldehyde did
not induce mutations at the hypoxanthine-guanine phosphoribosyltransferase (HPRT) locus in
V79 hamster cells treated for 4 hours with concentrations ranging from 125-500 uM (Merk and
Speit, 1999). Formaldehyde concentrations >250 uM produced significant decreases in cell
survival. In Chinese hamster ovary cells (CHO Kl), formaldehyde was found to induce a 4.7-
fold increase in HPRT mutations following a 1 hour exposure to 1 mM (Graves et al., 1996).
Butyraldehyde also induced concentration-related increases in the frequencies of HGPRT
and ouabain mutants in V79 hamster cells, following 60-minute exposures (Brambilla et al.,
1989). Significant increases in HGPRT and oubain mutants (p < 0.05-0.01 versus controls) were
observed at 10 and 30 mM. Similarly, the additional aldehydes tested, including pentanal,
hexanal, and nonanal, all induced concentration-related increases in the frequencies of HGPRT
and ouabain mutants in V79 hamster cells, following 60-minute exposures (Brambilla et al.,
1989). Significant increases in HGPRT mutants (p < 0.05-0.01 versus controls) were observed at
10 and 30 mM for pentanal, 30 mM for hexanal, and 0.1 and 0.3 mM for nonanal. Significant
increases in ouabain mutants (p < 0.05-0.01 versus controls) were observed at 10 and 30 mM for
pentanal, 3 and 10 mM for hexanal, and 0.3 mM for nonanal. The majority of these increases
were also associated with decreases in cell viability.
Isobutyraldehyde was found to be strongly mutagenic in the mouse lymphoma assay in
the absence of S9 (NTP, 1999). In this study, L5178Y mouse lymphoma cells were treated with
isobutyraldehyde for 4 hours at concentrations ranging from 62.5 tolOOO ug/mL. Concentrations
>125 ug/mL produced significant increases in mutations; 1000 ug/mL was found to be cytotoxic.
The results of the mutagenicity tests conducted in mammalian systems are compiled in
Table 4-3.
15
-------
Table 4-3. Mutagenicity of various aldehydes in mammalian cells
Aldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Acetaldehyde
Acetaldehyde
Formaldehyde
Formaldehyde
Butyraldehyde
Butyraldehyde
Isobutyraldehyde
Pentanal
Pentanal
Hexanal
Hexanal
Nonanal
Nonanal
Cells
V79
V79
V79
Human
lymphoctyes
Human
lymphoctyes
V79
CHOK1
V79
V79
L5178Y
V79
V79
V79
V79
V79
V79
Endpoint
HGPRT
HGPRT
Ouabain
HPRT
HPRT
HPRT
HPRT
HGPRT
Ouabain
—
HGPRT
Ouabain
HGPRT
Ouabain
HGPRT
Ouabain
Results"
+
-
+
+
+
-
+
+
+
+
+
+
+
+
+
+
LED (HTD)b
30 mM
(1.7 mg/mL)
[30mM]
2|lM
(0.12 |Ig/mL)
[2 JIM]
10 mM
(581 ng/mL)
[90 mM]
1.2 mM
(53 |lg/mL)
[2.4 mM]
0.2 mM
(9 |lg/mL)
[0.4 mM]
500 uM
(15 ng/mL )
[250 uM]
ImM
(30 |ig/mL)
10 mM
(720 Hg/mL)
[30 mM]
10 mM
(720 ng/mL)
125 ng/mL
[1000 ng/mL]
10 mM
(860 Hg/mL)
[30 mM]
10 mM
(860 Hg/mL)
[30 mM]
30 mM
(3.0 mg/mL)
[10 mM]
3mM
(300 ng/mL)
[10 mM]
100 uM
(14 |lg/mL)
[300 |IM]
300 |IM
(43 |lg/mL)
Reference
(Brambillaetal.,
1989)
(Smith etal., 1990)
(Brambillaetal.,
1989)
(He and Lambert,
1990)
(He and Lambert,
1990)
(Merk and Speit,
1999)
(Graves et al., 1996)
(Brambillaetal.,
1989)
(Brambillaetal.,
1989)
(NTP, 1999)
(Brambillaetal.,
1989)
(Brambillaetal.,
1989)
(Brambillaetal.,
1989)
(Brambillaetal.,
1989)
(Brambillaetal.,
1989)
(Brambillaetal.,
1989)
aTest results are either positive (+), negative (-), or equivocal (?).
bLED is the lowest effective concentration for positive test results; HTD is the highest tested concentration
for negative or inconclusive results; [ ] is the test concentration that resulted in notable decreases in cell
viability ortoxicity.
16
-------
4.4.1.2.2. Chromosomal aberrations. The results for chromosome damage in mammalian cells
in vitro are summarized in Table 4-4. Propionaldehyde induced a concentration-related increase
in chromosome aberrations in cultured Chinese hamster embryonic diploid (CHED) cells treated
with concentrations of 5 x 10"4 (0.0005), 1 x 10~3 (0.001), and 2 x 10~3 (0.002) % (equivalent to
4.3, 8.7, and 17 |ig/mL) for 1.5 hours (Furnus et al., 1990). Aneuploidy was induced at all three
concentrations but not in a concentration-related manner. No increase in the proportions of
polyploid cells was observed. An increase in lagging chromosome fragments, which is indicative
of chromosome breaks, was observed in Chinese hamster ovary (CHO) cells treated with 2.5,
5.0, and 7.5 x 10~4% propionaldehyde (equivalent to 2.2, 4.3, and 6.5 |ig/mL) for 8 hours
(Seoane and Dulout, 1994). Only the increase at the highest concentration tested (7.5 x 10^%)
was statistically significant (p < 0.05 versus untreated controls). No other aldehydes were
examined in this study.
Increases in chromosomal aberrations were induced in CHED cells treated with 4 and 6 x
10~3%, but not 2 x 10~3% acetaldehyde for 24 hours, and aneuploidy at all concentrations tested
(Dulout and Furnus, 1988). Concentrated-related increases in SCE were induced in CHO cells
treated with 2.5-15 x io~4% acetaldehyde for 24 hours (Obe and Beek, 1979).
Formaldehyde increased the frequency of sister chromatid exchanges (SCE) in a
concentrated-related manner in V79 hamster cells treated for 4 hours with concentrations up to
125 uM (Merk and Speit, 1999). Concentrated-related increases in SCE were also induced in
CHO cells treated with 1-4 x 10~4% formaldehyde for 24 hours, and in human lymphocytes
treated with 10~4-10"3% formaldehyde for 24 or 48 hours (Obe and Beek, 1979).
Treatment of CHO cells with butyraldehyde at concentrations of 59, 90, and 135 |ig/mL
for up to 26 hours in the absence of S9 did not induce increases in chromosomal aberrations
(Galloway et al., 1987). Butyraldehyde exposure did induce increases in SCE in CHO cells
treated with 9 to 90 |ig/mL for 25-29 hours in the absence of S-9 (Galloway et al., 1987), but
not in human lymphocytes treated with 2 x 10~3% butyraldehyde for 24 and 48 hours (Obe and
Beek, 1979).
Isobutraldehyde was also tested for the induction of chromosomal aberrations and SCE in
CHO cells (NTP, 1999). In cells treated with 16-4000 |ig/mL without S-9 for 12 hours,
isobutyraldehyde induced a concentration-related increase in chromosomal aberrations.
Similarly, isobutyraldehyde treatment induced a concentration-related increase in SCE in cells
treated for 26 hours over a concentration range of 5-500 |ig/mL in the absence of S-9.
17
-------
Table 4-4. Aldehyde-induced chromosome damage in mammalian cells
in vitro
Aldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Acetaldehyde
Acetaldehyde
Acetaldehyde
Formaldehyde
Formaldehyde
Formaldehyde
Butyraldehyde
Butyraldehyde
Butyraldehyde
Isobutyraldehyde
Isobutyraldehyde
Cells
CHED
CHO
CHED
CHED
CHED
CHO
V79
CHO
Human
lymphocytes
CHO
CHO
Human
lymphocytes
CHO
CHO
Endpoint
Aberrations
Fragments
Aneuploidy
Aberrations
Aneuploidy
SCE
SCE
SCE
SCE
Aberrations
SCE
SCE
Aberrations
SCE
Results"
+
+
+
+
+
+
+
+
+
-
+
—
+
+
LED (HTD)b
5 x 10^%
(4.3 |lg/mL)
0.75 x 1(T5%
(0.64 Hg/mL)
5 x 10^%
(4.3 |lg/mL)
4 x 10"3%
2 x lQ-3%
2.5 x 10^%
125 uM
1 x KT* %
1 x 10^%
135 Hg/mL
9 |lg/mL
2 x 10"3%
500 Hg/mL
5 |Ig/mL
Reference
(Furnus et al., 1990)
(Seoane and Dulout, 1994)
(Furnus et al., 1990)
(Dulout and Furnus, 1988)
(Dulout and Furnus, 1988)
(ObeandBeek, 1979)
(Merk and Speit, 1999)
(ObeandBeek, 1979)
(ObeandBeek, 1979)
(Galloway et al., 1987)
(Galloway et al., 1987)
(ObeandBeek, 1979)
(NTP, 1999)
(NTP, 1999)
4.4.1.2.3. DNA damage. The results for DNA damage caused by propionaldehyde and other
aldehydes are summarized in Table 4-5. Propionaldehyde induced a concentration-related
increase in unscheduled DNA synthesis (UDS) in rat hepatocytes at concentrations of 10, 30, and
100 mM (equivalent to 0.58, 1.7, and 5.8 mg/mL) following a 20-hour exposure in vitro
(Martelli, 1997; Martelli et al., 1994). UDS increases of 36-37% repair were statistically
significant at 30 and 100 mM (p < 0.001 compared with controls). A parallel test conducted in
human hepatocytes provided no evidence for UDS. Propionaldehyde concentrations of 300 mM
(equivalent to 17.4 mg/mL) were toxic to both cell lines.
18
-------
Table 4-5. Aldehyde-induced DNA damage in vitro
Aldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Propionaldehyde
Acetaldehyde
Acetaldehyde
Acetaldehyde
Acetaldehyde
Formaldehyde
Formaldehyde
Formaldehyde
Butyraldehyde
Butyraldehyde
Butyraldehyde
Pentanal
Pentanal
Hexanal
Hexanal
Hexanal
Hexanal
Species
Human
Human
Rat
Hamster
Hamster
N/AC
Human
Hamster
Hamster
N/A
Hamster
Hamster
N/A
Human
Rat
N/A
Human
Rat
Human
Rat
Hamster
Hamster
Cells
Hepatocytes
Lymphoma
Hepatocytes
CHO-K1
CHO-K1
Cell-free
plasmid
Lymphoma
CHO-K1
CHO-K1
Cell-free
plasmid
CHO-K1
CHO-K1
Cell-free
plasmid
Hepatocytes
Hepatocytes
Cell-free
plasmid
Hepatocytes
Hepatocytes
Hepatocytes
Hepatocytes
CHO-K1
CHO-K1
Endpoint
UDS
Cross-links
UDS
Strand
breaks
Cross-links
Cross-links
Cross-links
Strand
breaks
Cross-links
Cross-links
Strand
breaks
Cross-links
Cross-links
UDS
UDS
Cross-links
UDS
UDS
UDS
UDS
Strand
breaks
Cross-links
Results"
+
+
+
—
+
+
-
+
+
-
+
+
-
+
+
-
+
-
+
+
-
LED (HTD)b
100 mM
(5.8 mg/mL)
[300 mM]
75mM
(4.4 mg/mL)
30 mM
(1.7 mg/mL)
[300 niM]
4.5 mM
(261|lg/mL)
4.5 mM
(261 Hg/mL)
295 mM (17.1
mg/mL)
17.5 mM
(771 mg/mL)
4.5 mM
(198 |Ig/mL)
4.5 mM
(198 Hg/mL)
115 mM
(5.0 mg/mL)
4.5 mM
(135.1 |Ig/mL)
4.5 mM
(135.1 |Ig/mL)
1.5 HM
(0.045 ng/mL)
100 mM
(7.2 mg/mL)
30 mM
(2.2 mg/mL)
[300 mMl
360 mM
(26.0 mg/mL)
30 mM
(2.6 mg/mL)
3mM
(0.26 mg/mL)
[100 mMl
30 mM
(3.0 mg/mL)
30 mM
(3.0 mg/mL)
[100 mMl
4.5 mM
(0.45 mg/mL)
4.5 mM
(0.45 mg/mL)
Reference
(Martellietal.,
1994)
(Costa et al.,
1997)
(Martellietal.,
1994)
(Marinari et al.,
1984)
(Marinari et al.,
1984)
(Kuykendall and
Bogdanffy, 1992)
(Costa et al.,
1997)
(Marinari et al.,
1984)
(Marinari et al.,
1984)
(Kuykendall and
Bogdanffy, 1992)
(Marinari et al.,
1984)
(Marinari et al.,
1984)
(Kuykendall and
Bogdanffy, 1992)
(Martellietal.,
1994)
(Martellietal.,
1994)
(Kuykendall and
Bogdanffy, 1992)
(Martellietal.,
1994)
(Martellietal.,
1994)
(Martellietal.,
1994)
(Martellietal.,
1994)
(Martellietal.,
1994)
(Martellietal.,
1994)
19
-------
Aldehyde
Nonanal
Nonanal
Species
Human
Rat
Cells
Hepatocytes
Hepatocytes
Endpoint
UDS
UDS
Results"
-
-
LED (HTD)b
30 mM
(4.3 mg/mL)
30 mM
(4.3 mg/mL)
[100 mM]
Reference
(Martellietal.,
1994)
(Martellietal.,
1994)
"Test results are either positive (+), negative (-), or equivocal (?).
YED is the lowest effective concentration for positive test results; HTD is the highest tested concentration for
negative or inconclusive results; [ ] is the test concentration that resulted in notable decreases in cell viability
ortoxicity.
°N/A = not applicable.
The aldehydes butanal, pentanal, and hexanal also induced concentration-related
increases in UDS in rat hepatocytes, following a 20-hour exposure in vitro (Martelli, 1997;
Martelli et al., 1994). Significant increases in UDS (p < 0.001 compared with controls) were
observed at butanal concentrations of 30 and 100 mM (equivalent to 2.16 and 7.21 mg/mL),
pentanal concentrations of 3, 10, and 30 mM (equivalent to 0.258, 0.86, and 2.58 mg/mL), and a
hexanal concentration of 30 mM (equivalent to 3.0 mg/mL). The increases in UDS (20-30%
repair) induced by these aldehydes were comparable in potency to those produced by
propionaldehyde (36-37% repair). Nonanal did not induce UDS at the concentrations tested. No
significant increase in UDS (0-9% repair) was seen in human hepatocytes treated under similar
conditions with butanal, pentanal, hexanal, or nonanal at any of the concentrations tested.
Propionaldehyde produced a weak, concentration-related increase in DNA protein cross-
links (DPXs) in cultured human lymphoma cells, following a 4-hour exposure to concentrations
of 0.75, 3, 15, and 75 mM (equivalent to 0.044, 0.17, 0.87, and 4.4 mg/mL) (Costa et al., 1997).
The increase in DPX formation was significant (p < 0.05 compared with controls) at 75 mM, a
concentration that was toxic at a longer duration of exposure. Similar results were shown for
acetaldehyde. Acetaldehyde produced a weak, concentration-related increase in DPXs in cultured
human lymphoma cells, following a 4-hour exposure to concentrations of 0.035, 0.175, 0.875,
3.5, and 17.5 mM (equivalent to 0.0015, 0.008, 0.039, 0.154, and 0.77 mg/mL). The increase in
DPX formation was significant (p < 0.05 compared with controls) at 17.5 mM, a concentration
that was toxic at longer durations of exposure.
Treatment of CHO-K1 cells with 0.5, 1.5, and 4.5 mM (equivalent to 0.029, 0.087, and
0.26 mg/mL) propionaldehyde or hexanal (equivalent to 0.05, 0.15, and 0.45 mg/mL) for
90 minutes induced DNA single-strand breaks but not cross-links, based on concentration-
dependent decreases in the relative retention of DNA as measured by alkaline elution (Marinari
etal., 1984).
In contrast, treatment of CHO-K1 cells with formaldehyde and acetaldehyde produced
DPXs but not single-strand breaks when tested at concentrations of 0.5, 1.5, and 4.5 mM
(equivalent to 0.015, 0.045, 0.135, and 0.022, 0.066, 0.2 mg/mL, respectively). It was noted that
formaldehyde produced minimal cytotoxicity in this study.
20
-------
A filter-binding assay based on sodium dodecyl sulphate-potassium chloride (SDS-KC1)
precipitation of protein and covalently attached DNA was used to study the kinetics of plasmid-
histone cross-link formation with saturated and unsaturated aldehydes in vitro. In this study, 295
mM (equivalent to 17.1 mg/mL) propionaldehyde produced one cross-link per plasmid molecule
(Kuykendall and Bogdanffy, 1992). In comparison, the other aldehydes tested, acetaldehyde,
acrolein, formaldehyde, and butyraldehyde, produced one cross-link per plasmid molecule at
concentrations of 116 mM, 170 jiM, 1.6 jiM, and 357 mM, respectively.
4.4.1.2.4. Non-DNA adductformation. Propionaldehyde (5 mM ~ 290 jig/mL) has been shown
to form protein adducts with adult human hemoglobin (1 mM) in vitro (Hoberman and San
George, 1988). In another study, propionaldehyde (25 mM ~ 1,450 |ig/mL) did not form protein
adducts with freshly prepared human hemoglobin (~ 150 mg Hb/mL) in the absence of an added
arachidonic acid lipid peroxidation system (Kautiainen, 1992).
Acetaldehyde (5 mM ~ 220 |ig/mL) and butyraldehyde (5 mM ~ 360 |ig/mL) were also
shown to form protein adducts with adult human hemoglobin (1 mM) in vitro. The efficiency of
formation was noted to be inversely proportional to the aldehyde chain length (Hoberman and
San George, 1988). No protein hemoglobin adducts were recovered following treatment of
freshly prepared human hemoglobin (~ 150 mg Hb/mL) with pentanal (25 mM -2,150 |ig/mL)
or hexanal (25 mM ~ 2,500 jig/mL) in the absence of a supplementary oxidizing system
(Kautiainen, 1992). Low levels of adducts were seen when an arachidonic acid lipid peroxidation
system was added.
4.4.1.3. Genotoxicity Summary
In summary, the genotoxicity of propionaldehyde has been studied in bacteria and a
number of mammalian cells in vitro. Propionaldehyde was found to be mutagenic in
S. typhimurium strain TA1534 (Sampson and Bobik, 2008), and nonmutagenic in all other strains
tested (Dillon et al., 1998; Aeschbacher et al., 1989; Mortelmans et al., 1986). The positive
mutagenic response was observed in a mutated strain in which a bacterial microcompartment
thought to mitigate toxicity was inactivated (Sampson and Bobik, 2008). Propionaldehyde
produced concentration-related increases in HGPRT and ouabain mutants in V79 hamster cells
(Brambilla et al., 1989). These effects, however, were associated with decreases in cell viability
in these test systems. Smith et al. (1990) determined that propionaldehyde was not mutagenic at
the HGPRT locus in V79 hamster cells exposed to lower, noncytotoxic concentrations.
Propionaldehyde produced a concentration-related increase in chromosome aberrations in
Chinese hamster embryonic cells (Furnus et al., 1990) and chromosome breaks in CHO cells
(Seoane and Dulout, 1994). In addition, propionaldehyde induced a concentration-related
increase in unscheduled DNA synthesis in rat, but not human, hepatocytes (Martelli, 1997;
Martelli et al., 1994) and a weak, concentration-related increase in DPXs in cultured human
21
-------
lymphoma cells (Costa et al., 1997). Although the information provided in these in vitro studies
suggests that propionaldehyde is DNA reactive, supportive information from in vivo animal
bioassay studies is unavailable.
The relevance of the in vitro test doses and effects in these studies is unclear as a number
of these results were associated with significant cellular toxicity. In addition, the relevance of the
in vitro dose and effect to in vivo dose and effect is also difficult to ascertain. For example,
increases in DPXs have been used as a tissue dose surrogate and serve as an important aspect in
describing the dosimetry and the mode of action for formaldehyde-induced nasal lesions based
on its correlation with cell replication at specific sites in the nose (Conolly et al., 2000; Casanova
et al., 1994). However, studies measuring DPX formation in the nose in response to inspired
acetaldehyde have been negative although DPX formation was detected in nasal tissue
homogenates treated with acetaldehyde (Dorman et al., 2008; Stanek and Morris, 1999).
Consequently, with respect to DPX formation, formaldehyde may represent a special case
because of its highly focal deposition and toxicity in the nose as well as its greater potency to
form cross-links compared to other aldehydes.
In general, this information indicates that the rank order of potency of aldehydes across
similar genotoxic endpoints appears to be as follows: formaldehyde » acetaldehyde ~
propionaldehyde > butyraldehyde and isobutyraldehyde. For DPX, the rank order of potency is:
formaldehyde » acetaldehyde > propionaldehyde > butyraldehyde.
4.4.2. CARDIOVASCULAR EFFECTS
Egle (1972b) investigated the effects of propionaldehyde on arterial blood pressure and
heart rate. Male Wistar rats were exposed to propionaldehyde concentrations ranging from 3.0-
200 |ig/mL (3,000-200,000 mg/m3 or 1260-84,000 ppm) via inhalation for 1-minute intervals.
Propionaldehyde-induced changes in blood pressure and heart rate (expressed as percent change
± SE) were compared with those in control rats (n = 93) exposed to clean air. The results are
summarized in Table 4-6.
22
-------
Table 4-6. Effects of inhalation of propionaldehyde on blood pressure and heart
Exposure concentration,
jig/mL (mg/m3)
Control (air)
3.0 (3000)
10.0 (10,000)
20.0 (20,000)
30.0 (30,000)
50.0 (50,000)
100.0 (100,000)
150.0 (150,000)
200.0 (200,000)
Blood pressure (% change ± SE)a Heart rate (% change ± SE)a
| 0.8 ±0.7
t 3.2 ±1.0
t5.9±1.13b
t 10.6±1.5C
t20.8±2.6c
t20.6±2.1c
t27.1±6.3c
t41.6±4.7c
t47.0±4.9c
| 0.9 ±0.6
| 3.3 ±0.6
t 3.0 ±1.2
t 6.1 ±1.1°
t5.0±1.0c
t 1.6 ±0.7
t 1.7 ±2.2
t 3.4 ±4.2
4 26.0 ±9.1°
Increase (t); decrease (4).
bSignificantly different from control atp < 0.05.
Significantly different from control atp < 0.01.
Source: Egle (1972b).
A slight but nonsignificant rise in blood pressure was seen at 3.0 |ig/mL (3.2 ± 1.0%;
n = 7), while exposure-related significant increases (p < 0.05) in blood pressure were seen at 10
|ig/mL (5.9 ± 1.13%; n = 6), 20 |ig/mL (10.6 ± 1.5%; n = 6), 30 |ig/mL (20.8 ± 2.6%; n = 5), 50
|ig/mL (20.6 ± 2.1%; n = 6), 100 |ig/mL (27.1 ± 6.3%; n = 3), 150 |ig/mL (41.6 ± 4.7%; n = 3),
and 200 |ig/mL (47.0 ± 4.9%; n = 3). The lowest exposure concentration (3.0 jig/mL; n = 7) was
without effect on heart rate, while concentrations of 20 (6.1 ± 1.1%; n = 6) and 30 |ig/mL (5.0 ±
1.0%; n = 5) produced significant increases in heart rate (p < 0.01 versus controls). No change in
heart rate was seen in the 50-150 |ig/mL exposure groups as compared with that in controls.
However, exposure to 200 |ig/mL propionaldehyde resulted in a significant decrease (-26.0 ±
9.1%; n = 3) (p < 0.01) in heart rate. Based on the data, 3,000 mg/m3 (3 jig/mL) appears to be a
NOEL for rat cardiac responses. However, the biological significance of these changes is
uncertain as relatively high concentrations of propionaldehyde were required to produce effects,
and thus limits the usefulness of these data.
In another study, Egle et al. (1973) examined the effects of intravenous (i.v.)
administration of propionaldehyde on blood pressure and heart rate. Male Wistar rats (7-
10/dose/treatment group) were administered propionaldehyde at dosing regimens of 5 mg/kg at
10-minute intervals and 10, 20, and 40 mg/kg at 20-minute intervals. A group of control animals
(n = 9) received saline injections that were found to have no effect on resting blood pressure and
heart rate. Results were expressed as the percent change ± SE from the initial resting blood
pressure or heart rate in each dose/treatment group. Multiple observations were made in each
dose/treatment group, and data were reported as the frequency of each response as a function of
the number of observations (e.g., a dose/treatment group of seven rats may yield a frequency of
response of 18 for 21 [18/21] total observations). After administration of 5 and 10 mg/kg
propionaldehyde, pressor responses predominated as average increases in blood pressure of 10.5
± 1.1% (17/17) and 12.4 ± 1.9% (18/21), respectively, were observed. Although pressor
23
-------
responses were still evident, depressor responses predominated after administration of 20 and 40
mg/kg propionaldehyde as average decreases in blood pressure of 40.0 ± 8.1% (11/20) and 63.9
± 7.2% (13/16), respectively, were observed. The pressor responses induced by propionaldehyde
were partially inhibited by the adrenergic antagonists reserpine (a depletor of monoamine
neurotransmitters) and phentolamine, and the depressor responses were reduced by the
anticholinergic agent atropine as well as by bilateral vagotomy. Administration of 40 mg/kg
propionaldehyde also induced a profound decrease in heart rate of 71 ± 6.1% (n = 16) from
baseline. This response was partially attenuated by phentolamine and atropine and completely
reversed by bilateral vagotomy. Based on the results of this study, the authors concluded that
propionaldehyde exerts two opposing actions on the cardiovascular system at different dose
levels—a sympathomimetic effect that results primarily from release of norepinephrine and
produces vasoconstriction and an increase in blood pressure and a secondary reflex stimulation
of the vagus nerve that results in bradycardia and hypotension.
The effect of propionaldehyde on isolated smooth muscle systems was studied (Beckner
et al., 1974). In the first part of the study, isolated vas deferens from Wistar rats was treated with
propionaldehyde and contractile responses and concentration-response relationships were
examined. The isolated rat vas deferens was first exposed to 14C-norepinephrine for 15 minutes,
and the ability of the aldehydes to produce an increase in loss of radioactivity was then
examined. Propionaldehyde (p < 0.05) significantly reduced 14C-concentration in tissue. The
contractile response produced by propionaldehyde was reversible and blocked by reserpine
pretreatment. In the second part of the study, the effect of propionaldehyde on 45Ca binding in
the aorta isolated from New Zealand white rabbits was examined. Propionaldehyde significantly
(p < 0.05) reduced calcium binding in isolated rabbit aorta at approximately 10~2 M after
30 minutes of exposure. The authors concluded that propionaldehyde can cause the release of
endogenous catecholamines (e.g., norepinephrine) and may interact with tissue norepinephrine
stores by inhibiting Na+, K+-dependent adenosine triphosphatase (ATPase) and affect nonspecific
membrane calcium-binding sites. These results provide further support that the cardiovascular
effects induced in animals after exposure to propionaldehyde appear to be due to their
sympathomimetic activities.
4.4.3. IMMUNOTOXICITY
Poirier et al. (2002) assessed propionaldehyde as a chemical component of tobacco
smoke for its effects on viability and proliferation of mouse lymphocytes in vitro.
Propionaldehyde significantly inhibited T-lymphocyte and B-lymphocyte proliferation, with
median inhibitory concentration (IC50) values of approximately 3 x 10~5 M after 3 hours of
exposure. Other chemical components that also inhibited T-lymphocyte and B-lymphocyte
proliferation were formaldehyde, catechol, acrylonitrile, acrolein, crotonaldehyde, and
hydroquinone with ICso values in the range of 1.19 x 10~5 to 5.86 x 10~4M. Based on their ICso
24
-------
values, propionaldehyde was determined to be more inhibitory than formaldehyde but less than
acrolein and crotonaldehyde. Propionaldehyde did not affect lymphocyte cell viability since the
IC50 for lymphocyte cell viability was in the same range as the control. Acrolein and
crotonaldehyde were the only compounds shown to affect lymphocyte cell viability. These
results suggest that propionaldehyde may have effects on important lymphocyte function.
4.4.4. CYTOTOXICITY
In a cytotoxicity study, Bombick and Doolittle (1995) used the neutral red uptake assay,
which measures cellular membrane damage and cell viability, to investigate the cytotoxic
potential and chemical structure of low molecular weight aldehydes, including propionaldehyde.
CHO cells were treated with propionaldehyde for 24 hours, and the median effective
concentration (ECso) (the chemical concentration required to reduce the absorbance value by
50% after a 24-hour exposure) was determined. The ECso for propionaldehyde was 17.2 mM.
In another cytotoxicity study, Koerker et al. (1976) treated the NBP2 clone of C1300
mouse neuroblastoma cells in culture with propionaldehyde and investigated their effects on the
inhibition of cell growth and viability, changes in the morphologic appearance of the cells, and
increase in the percentage of cells sloughing into the medium. For propionaldehyde, the molar
(M) concentrations producing a 50% change from control in each cytotoxic endpoint after 24
hours of exposure ranged from 2 x 10~4to 1 x 10~2 M.
4.4.5. COMPARATIVE TOXICITY OF RELATED ALDEHYDES
Several studies that provide information on the comparative toxicity of various aldehydes
were identified in the literature. The majority of these studies examined and compared the
relative potencies of aldehydes in a variety of in vivo and in vitro systems. The studies discussed
below are limited primarily to those studies in which a number of aldehydes were examined
together and/or studies in which endpoints relevant to propionaldehyde were evaluated, allowing
for more direct comparisons. The endpoints evaluated include respiratory and cardiac effects,
effect on smooth muscle, and cellular cytotoxicity.
Guth (1996) reviewed and assessed the noncancer effects of propionaldehyde based on
comparative toxicity with other low molecular weight aldehydes, such as formaldehyde, acrolein,
and acetaldehyde. The effects of i.v. administration of acetaldehyde or propionaldehyde on blood
pressure and heart rate in rats were very similar (Egle et al., 1973), and the effects from
inhalation on blood pressure and heart rate showed that acetaldehyde and propionaldehyde also
have similar potencies by this route of exposure (Egle, 1972b). Guth (1996) concluded that these
results, taken together, suggest that acetaldehyde and propionaldehyde are absorbed and
distributed similarly after inhalation exposure, since changes in heart rate and blood pressure are
systemic effects. In a comparative kinetic study conducted in dogs, Egle (1972a) observed
similar magnitudes of respiratory tract deposition after inhalation exposure for acetaldehyde,
25
-------
acrolein, and propionaldehyde, with deposition averaging between 70 and 80%. In addition,
propionaldehyde and acetaldehyde exhibit similar median lethal doses (LD50) after oral exposure
(1930 and 1410 mg/kg, respectively) and subcutaneous dosing (640 and 820 mg/kg). In
comparing the RDso values among various aldehydes, Steinhagen and Barrow (1984) observed
that the unsaturated aldehydes and formaldehyde were approximately 2 orders of magnitude
more potent than the longer-chain saturated aldehydes (e.g., propionaldehyde).
In a study designed to test general and portal-of-entry toxicity, the most sensitive
noncancer effect identified in rats for acetaldehyde was degeneration of the olfactory nasal
epithelium (Woutersen et al., 1986; Appelman et al., 1982;). Appelman et al. (1982) exposed
male and female Wistar rats to 400, 1,000, 2,200, or 5000 ppm acetaldehyde 6 hours/day,
5 days/week for 4 weeks. Small reductions in weight gain were seen at exposure concentrations
of 1000 ppm and greater. Degeneration of the nasal olfactory epithelium was observed at the
lowest exposure concentration tested (400 ppm), and this effect increased in severity with
increasing exposure concentration. Similar results were obtained by Appelman et al. (1986),
when degeneration of the olfactory epithelium was observed in rats exposed to 500 ppm
acetaldehyde 6 hours/day, 5 days/week for 4 weeks. Reductions in weight gain were not noted in
these animals, and no compound-related effects were seen in animals exposed to 150 ppm
acetaldehyde. In a recent study by Dorman et al. (2008), a similar pattern and progression of
nasal olfactory lesions were observed in F344 rats exposed to acetaldehyde concentrations of
150, 500, and 1,500 ppm via whole-body inhalation for 6 hours/day, 5 days/week for up to 65
exposure days. Olfactory epithelial degeneration, noted as the most sensitive endpoint observed,
increased in incidence and severity with both exposure concentration and duration. The presence
of vacuolization was also noted, but the severity was not graded. In animals exposed to 50 ppm,
no lesions were observed. The portal-of-entry effects of the structurally-related aldehyde
isobutyraldehyde was also evaluated in a 2-year chronic toxicology and carcinogenicity study
(NTP, 1999). In this study groups of male and female F344 rats were exposed to 0, 500, 1,000, or
2,000 ppm isobutyraldehyde by inhalation of 6 hours/day, 5 days/week for 105 weeks. No
increases in neoplastic nasal lesions were observed in this study. Nonneoplastic lesions in the
nose consisted of squamous metaplasia of the respiratory epithelium, degeneration of the
olfactory epithelium, and suppurative inflammation. Incidences of minimal to mild squamous
metaplasia in 1,000 and 2,000 ppm males and females and in 500 ppm females were significantly
greater than those in the chamber controls. Another lesion associated with exposure was minimal
to mild degeneration of the olfactory epithelium in 2,000 ppm males and females. The incidences
of suppurative inflammation (rhinitis) in male and female rats exposed to 2,000 ppm were
increased compared to the chamber controls.
Although studies of comparable design examining the effects of propionaldehyde on the
nasal epithelium are unavailable, a similar pattern and progression of nasal olfactory lesions were
reported in adult male and female CD rats in a propionaldehyde inhalation reproductive and
26
-------
developmental study conducted by Union Carbide (1993, 1991) (see Sections 4.3 and 4.5.2).
Increases in olfactory epithelium atrophy were observed at 150, 750, and 1,500 ppm
propionaldehyde. In toto, these comparisons suggest that acetaldehyde and propionaldehyde
produce similar respiratory and cardiac effects, and may produce nasal lesions at similar
exposure concentrations.
Steinhagen and Barrow (1984) compared the RDso values of 14 aldehydes in B6C3Fi and
Swiss-Webster mice as a measure of sensory irritation potential. Groups of three to four mice per
strain were exposed via inhalation in a head-only exposure chamber for 10 minutes to varying
concentrations (usually five) of the test aldehyde. Respiratory rates were measured by a method
in which animals were sealed in airtight plethysmographs and attached to a head-only exposure
chamber, and concentration-response curves were constructed to determine the RD50. In animals,
sensory irritants produce a reflex decrease in respiratory rate characterized as a pause at the onset
of expiration. The RD50 values for propionaldehyde, acetaldehyde, formaldehyde, and acrolein
for each mouse strain are shown in Table 4-7. Other aldehydes tested included crotonaldehyde,
isovaleraldehyde, butyraldehyde, caproaldehyde, valeraldehyde, and isobutyraldehyde.
Comparing the values for the aldehydes tested, the RD50 values spanned approximately 3.5
orders of magnitude. The a,P~unsaturated aliphatic aldehydes (acrolein and crotonaldehyde) and
formaldehyde were approximately two orders of magnitude more potent than the saturated
aliphatic aldehydes (propionaldehyde, isovaleraldehyde, butyraldehyde, caproaldehyde,
valeraldehyde, acetaldehyde, and isobutyraldehyde) in producing a 50% decrease in respiration
rate.
Table 4-7. RDso values for propionaldehyde and selected, related aldehydes
measured in B6C3Fi and Swiss-Webster mice
Aldehyde
Propionaldehyde
Isobutyraldehyde
Acetaldehyde
Formaldehyde
Acrolein
B6C3F!a
2,078 ppm (1,803-2402)
4,946 mg/m3 (4,291-5,717)
30 16 ppm (2568-36 10)
2,932 ppm (2,627-3,364)
4.90 ppm (3. 9-6.4)
1.41 ppm (1.16-1.73)
Swiss- Webster
2,052 ppm (1,625-3,040)
4,884 mg/m3 (3,868-7,235)
4 167 ppm (3258-5671)
2,845 ppm (1,967-3,954)
3. 2 ppm (2. 1-4.7)
1.03 ppm (0.70-1. 52)
aRanges for RD50 values shown in parentheses.
Source: Steinhagen and Barrow (1984).
The effects of propionaldehyde, acetaldehyde, formaldehyde, and acrolein on isolated
smooth muscle systems were studied (Beckner et al., 1974). In the first part of the study, isolated
vas deferens from Wistar rats was treated with the four aldehydes and contractile responses and
concentration-response relationships were examined. The isolated rat vas deferens was first
exposed to 14C-norepinephrine for 15 minutes, and the ability of the aldehydes to produce an
increase in loss of radioactivity was then examined. Propionaldehyde (p < 0.05) and
27
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acetaldehyde (p < 0.01) at 10 2 M and formaldehyde (p < 0.05) and acrolein (p < 0.01) at 10 3 M
significantly reduced 14C-concentration in tissue. The contractile responses produced by
propionaldehyde and acetaldehyde, but not formaldehyde and acrolein, were reversible and
blocked by reserpine pretreatment. In the second part of the study, the effect of these aldehydes
on 45Ca binding in the aorta isolated from New Zealand white rabbits was examined. All four
aldehydes significantly (p < 0.05) reduced calcium binding in isolated rabbit aorta in the same
concentration range (10~2 M) after 30 minutes of exposure. The authors concluded that these
results suggest that propionaldehyde and acetaldehyde can cause the release of endogenous
catecholamines (e.g., norepinephrine), and all four aldehydes may interact with tissue
norepinephrine stores by inhibiting Na+, K+-dependent ATPase and affect nonspecific membrane
calcium-binding sites. These results provide further support that the cardiovascular effects
induced in animals after exposure to propionaldehyde and other aldehydes appear to be due to
their indirect sympathomimetic activities (see Egle et al., [1973] in Section 4.4.2).
Wang et al. (2002) performed a genotype analysis of the ALDH2 gene in the livers of
human volunteers in order to investigate the metabolism of a variety of aldehydes. Of a total of
39 subjects, 8 were heterozygotes of the wild-type (ALDH2*1) and mutant (ALDH2*2) alleles,
and the others were homozygotes of the wild-type allele. The ability of mitochondria to
metabolize propionaldehyde, acetaldehyde, formaldehyde, n-butyraldehyde, capronaldehyde, and
heptaldehyde was significantly lower (p < 0.05) (between 37 and 93%, depending on the
aldehyde; 80% for propionaldehyde) in the heterozygotes (ALDH2*l/*2) compared to the
homozygotes (ALDH2*1/*1), showing differences in metabolism between the two genotypes.
However, the mitochondrial activity was not lower for octylaldehyde, decylaldehyde,
retinaldehyde, benzaldehyde, 3-hydroxybenzaldehyde, 2,5-dihydroxybenzaldehyde,
phenylacetaldehyde, and 3-phenylpropionaldehyde, showing similar metabolism between the
two genotypes. Based on these results, the authors hypothesized that the polymorphisms of the
ALDH2 gene may only alter the metabolism of the short aliphatic chain aldehydes.
In a cytotoxicity study, Bombick and Doolittle (1995) used the neutral red uptake assay,
which measures cellular membrane damage and cell viability, to investigate the relationship
between the cytotoxic potential and chemical structure of low molecular weight aldehydes. CHO
cells were treated with formaldehyde, acetaldehyde, propionaldehyde, acrolein, pyridine, 2-vinyl
pyridine, 4-vinyl pyridine, 4-picoline, butanol, and ammonium hydroxide for 24 hours, and the
chemical concentrations required to reduce the absorbance value by 50% after a 24-hour
exposure (ECso) were determined. The ECso values for the aldehydes were as follows: 0.009 mM
for acrolein, 0.6 mM for formaldehyde, 2.3 mM for acetaldehyde, and 17.2 mM for
propionaldehyde. Thus, formaldehyde was considered more toxic than acetaldehyde, which was
more toxic than propionaldehyde, with the a-, p-unsaturated aldehyde, acrolein, being the most
toxic compound by almost three orders of magnitude. Based on these results, the authors
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concluded that cytotoxicity generally appears to decrease with increasing (saturated) aldehyde
chain length.
In another cytotoxicity study, Koerker et al. (1976) treated the NBP2 clone of C1300
mouse neuroblastoma cells in culture with propionaldehyde, acetaldehyde, and acrolein
formaldehyde, and investigated their effects on the inhibition of cell growth and viability,
changes in the morphologic appearance of the cells, and the increase in the percentage of cells
sloughing into the medium. For each aldehyde, the molar concentrations producing a 50%
change from control in each cytotoxic endpoint after 24 hours of exposure are shown in Table
4-8. Based on these results, the authors noted that toxicity increased with decreasing aldehyde
chain length, perhaps reflecting the ease of cross-linking or the reactivity of the carbonyl group.
For example, acrolein was considerably more toxic than propionaldehyde for each endpoint,
illustrating the increased activity of the carbonyl group caused by the presence of the conjugated
double bond.
Table 4-8. Concentration [M] of selected aldehydes required to produce a
50% change from control in each cytotoxic endpoint
Effect
Sloughed cells
Neurite formation
Viability of sloughed cells
Total cell number
Viability of harvested cells
Propionaldehyde
2.2 x 1(T3
2.1 x KT4
1.0 x 1(T3
1.0 x 10~2
4.8 x 10~3
Acetaldehyde
5.4 x IQ-4
7.9 x IQ-4
6.4 x 10~3
6.4 x 10~3
9.0 x 10~3
Acrolein
1.0 x 10~6
7.6 x 10~6
5.3 x 10~6
5.8 x 10~4
3.0 x 10~5
Formaldehyde
8.3 x 10~6
2.0 x 1Q-6
4.5 x 10~6
2.8 x 1Q-6
2.2 x 10~4
Source: Koerker etal. (1976).
Egyud (1967) investigated the effects of a variety of chemical groups, including the
aldehydes, on cell division in Escherichia coll. The chemicals were added to logarithmically
growing bacteria, and the reaction was followed by measuring the increase in the optical density
on a colorimeter. The concentration of the aliphatic aldehydes tested was 10~3 M. Formaldehyde
and acetaldehyde completely and irreversibly inhibited cell division, while the other aldehydes,
including propionaldehyde, produced a transient inhibitory effect.
The studies summarized above provide some insight in comparing the relative potencies
of various aldehydes for the same endpoint(s) and in the same or similarly conducted studies.
Whether the endpoint be portal-of-entry effects, decrease in respiration, or in vitro cytotoxicity,
the rank order of potency appears to be acrolein > formaldehyde » acetaldehyde ~
propionaldehyde > isobutryaldehyde with potency further decreasing with increasing (saturated)
aldehyde chain length.
4.5. SYNTHESIS AND EVALUATION OF MAJOR NONCANCER EFFECTS
4.5.1. ORAL
No human or animal studies are available on the oral effects of propionaldehyde.
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4.5.2. INHALATION
The most notable propionaldehyde-induced effects reported in animal inhalation
exposure studies are respiratory tract irritation, histopathology, and cardiovascular perturbations.
Two short-term reproductive/developmental inhalation studies were conducted by Union
Carbide, one for 20 days (Union Carbide, 1991) and the second for a duration of 7-8 weeks
(Union Carbide, 1993).
In a range-finding study, young adult female CD rats (seven per group) were exposed to
0, 500, 1,000, 1,500, or 2,500 ppm propionaldehyde for 6 hours/day, on GDs 0 through 20,
following successful mating with naive males (Union Carbide, 1991). Maternal toxicity was
noted as exposure-related decreases in body weight gain; however, these decreases in body
weight gain were accompanied by decreases in food consumption throughout the gestation
period. There were no exposure-related differences in gestational parameters, including total
number of implants and the number of viable and nonviable implants. No other evidence of any
treatment-related external malformations or variations was observed.
In the second study, young adult male and female CD rats (15/sex/group) were exposed
to 0, 150, 750, or 1,500 ppm propionaldehyde for 6 hours/day, 7 days/week, during a 2-week
premating period and a 14-day mating phase (Union Carbide, 1993). The mated females were
exposed daily through GD 20 for a minimum of 35 days and a maximum of 48 days depending
upon when they mated (average exposure period -38 days). The females were then allowed to
deliver their litters naturally and raise their offspring until day 4 of lactation, when they were
sacrificed. The males continued to be exposed until sacrifice in week 7, for a total of 52
exposures.
In the adult females, no exposure-related clinical signs were noted. Body weight gains
and food consumption were slightly decreased during the first week of exposure to 750 and
1,500 ppm. During gestation, body weight and food consumption were decreased in the high
exposure group compared with controls, but no differences in body weight changes were
observed. No significant effects of exposure on any of the reproductive parameters assessed were
found. Litter size and viability were similar among the groups. At sacrifice, no gross lesions
attributable to propionaldehyde exposure were found. However, microscopic examination of the
nasal cavity revealed propionaldehyde-induced vacuolization of the olfactory epithelium in the
150 and 750 ppm exposure groups and atrophy of the olfactory epithelium in the 750 and 1,500
ppm exposure groups. These effects were noted to be localized to the dorsal anterior two sections
of the nasal cavity. The incidence of atrophy was 0/15, 0/15, 2/15, and 15/15 at 0, 150, 750, and
1,500 ppm, respectively (see Table 4-1). The severity of this nasal lesion increased with
exposure concentration being minimal to mild at 750 ppm and moderate to marked at 1,500 ppm.
No evidence of squamous metaplasia was found in olfactory or respiratory epithelium. Low
incidences of minimal to mild rhinitis involving the respiratory epithelium were also noted at
150, 750, and 1,500 ppm.
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In the males, body weights, weight gains, clinical observations, and food consumption
were similar among all exposure groups and controls. At necropsy, no gross lesions were found.
However, similar to effects in the females, microscopic examination revealed exposure-related
effects in the olfactory epithelium of the nasal cavity that consisted of vacuolization in the low
and intermediate exposure groups and atrophy in the intermediate and high exposure groups.
These effects were noted to be localized to the dorsal anterior two sections of the nasal cavity.
The incidence of atrophy was 0/15, 2/15, 10/15, and 15/15 at 0, 150, 750, and 1,500 ppm,
respectively. The severity of this nasal lesion increased with exposure concentration being
minimal at 150 ppm, minimal to moderate at 750 ppm, and mild to marked at 1,500 ppm.
Squamous metaplasia of the respiratory epithelium was reported in one male from the 750 ppm
group and two males from the 1500 ppm group. An increased incidence of minimal to moderate
rhinitis involving the respiratory epithelium was also noted at 750 and 1,500 ppm. The decrease
in incidence and severity of the nasal lesions in females relative to males is likely to be
attributable to the differences in exposure duration and approximate 6-day period between
cessation of exposures after GD 20 and sacrifice on day 4 of lactation. This observation may also
indicate that these effects are reversible and that repair and regeneration of the olfactory
epithelium has been initiated. However, pathological indications (e.g., cell proliferation,
hyperplasia) that these processes have started in the female rats were not noted. Consequently,
although the incidence of olfactory epithelium atrophy was not the most sensitive effect observed
after exposure to propionaldehyde, the U.S. EPA considers this endpoint to be a biologically
significant effect (as discussed in Section 5.2.1).
The respiratory tract effects induced by propionaldehyde are consistent with the portal-
of-entry effects reported for other aldehydes, such as acrolein, acetaldehyde, and formaldehyde,
all of which deposit significantly in the upper respiratory tract. In addition, isobutyraldehyde
produces a similar pattern of portal-of-entry effects; however, respiratory tract uptake
information is lacking. Egle (1972a) demonstrated in dogs that approximately 70-80% of inhaled
propionaldehyde is retained in the upper respiratory tract. In addition, when comparing the
sensory irritation potential (i.e., RDso values) among aldehydes, propionaldehyde was found to
be two orders of magnitude less potent than acrolein and formaldehyde, slightly more potent than
acetaldehyde, and approximately 1.5-fold more potent than isobutyraldehyde (Steinhagen and
Barrow, 1984). This reflex decrease in respiratory rate is mediated via stimulation of nasal
trigeminal nerves and is characterized as a pause at the onset of expiration. In studies examining
the effects of propionaldehyde on blood pressure and heart rate in rats after both i.v. and
inhalation exposure, propionaldehyde was shown to produce dose-related pressor (at low doses)
and depressor (at high doses) responses (Egle et al., 1973; Egle, 1972b). The pressor responses
induced by propionaldehyde were partially inhibited by the adrenergic antagonists reserpine and
phentolamine, and the depressor responses were reduced by the anticholinergic agent atropine as
well as by bilateral vagotomy. Administration of 40 mg/kg propionaldehyde i.v. also induced a
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profound decrease in heart rate from baseline (a response also observed at the high inhalation
exposure concentration). This response was partially attenuated by phentolamine and atropine
and completely reversed by bilateral vagotomy. Based on the results of these studies, it can
reasonably be surmised that propionaldehyde exerts two opposing actions on the cardiovascular
system at different dose levels—a sympathomimetic effect that results primarily from release of
norepinephrine and produces vasoconstriction and an increase in blood pressure and a secondary
reflex stimulation of the vagus nerve that results in bradycardia and hypotension.
Similar results were observed when propionaldehyde, acetaldehyde, formaldehyde, and
acrolein were tested in vitro on isolated smooth muscle systems (Beckner et al., 1974). In the
first part of the study, the contractile responses produced by propionaldehyde and acetaldehyde,
but not formaldehyde and acrolein, were reversible and blocked by reserpine pretreatment. In the
second part of the study, all four aldehydes significantly reduced calcium binding in isolated
rabbit aorta in the same concentration range. The authors concluded that taken together these
results suggest that propionaldehyde and acetaldehyde can cause the release of endogenous
catecholamines (e.g., norepinephrine), and all four aldehydes may interact with tissue
norepinephrine stores by inhibiting Na+, K+-dependent ATPase and affect nonspecific membrane
calcium-binding sites. In addition, these results provide support that the cardiovascular effects
induced in animals after exposure to propionaldehyde and other aldehydes appear to be due to
their sympathomimetic activity.
Gage (1970) exposed four male and four female Alderley-Park rats to 1,300 ppm
propionaldehyde 6 hours/day for 6 days via whole-body inhalation. No changes in body weight
were noted; however, microscopic examination revealed liver cell vacuolation. Four male and
four female rats were also exposed to 90 ppm 6 hours/day for 20 exposures. All organs were
reported to be normal at autopsy and no clinical signs of toxicity were noted.
4.6. WEIGHT-OF-EVIDENCE EVALUATION AND CANCER CHARACTERIZATION
4.6.1. SUMMARY OF OVERALL WEIGHT OF EVIDENCE
In accordance with the Guidelines for Carcinogen Risk Assessment (EPA, 2005a), there is
"inadequate information to assess the carcinogenic potential" for propionaldehyde. No human
health effects data or chronic animal bioassay studies are available that assess the carcinogenic
effects of propionaldehyde.
The genotoxicity of propionaldehyde has been studied in bacteria and a number of
mammalian cells in vitro. Propionaldehyde was found to be mutagenic in S. typhimurium strain
TA1534 (Sampson and Bobik, 2008) and nonmutagenic in all other strains tested (Dillon et al.,
1998; Aeschbacher et al., 1989; Mortelmans et al., 1986), but produced concentration-related
increases in HGPRT and ouabain mutants in V79 hamster cells (Brambilla et al., 1989). These
effects, however, were associated with decreases in cell viability in these test systems. Smith
32
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et al. (1990) determined that propionaldehyde was not mutagenic at the HGPRT locus in V79
hamster cells exposed to lower, noncytotoxic concentrations. Propionaldehyde produced a
concentration-related increase in chromosome aberrations in Chinese hamster embryonic cells
(Furnus et al., 1990) and chromosome breaks in CHO cells (Seoane and Dulout, 1994). In
addition, propionaldehyde induced a concentration-related increase in UDS in rat, but not
human, hepatocytes (Martelli, 1997; Martelli et al., 1994) and a weak, concentration-related
increase in DPXs in cultured human lymphoma cells (Costa et al., 1997). For formaldehyde,
increases in DPXs serve as an important aspect in describing the dosimetry and the carcinogenic
mode of action for nasal tumors (Conolly et al., 2000; Casanova et al., 1994). Although the
information provided in the in vitro studies suggests that propionaldehyde is DNA reactive,
information from in vivo animal bioassay studies is unavailable. This overall lack of information
represents a data gap and does not allow for either a quantitative or a qualitative assessment of
the carcinogenic potential of propionaldehyde or a definitive statement concerning its mutagenic
potential.
It is important to note that inhalation exposure to propionaldehyde produced a low
incidence of respiratory epithelium squamous metaplasia in male rats in the intermediate and
high exposure groups (Union Carbide, 1993). Although this alteration may be viewed as an
adaptive response typical of nasal epithelial tissues in response to continued irritant insult, the
lesion may become part of a progression from nasal tissue injury and toxicity (e.g., epithelial
degeneration and atrophy) to hyperplasia to increased cell proliferation and lastly to nasal
tumorigenesis (Renne et al., 2007; Boorman et al., 1990). Squamous metaplasia is also noted in
studies examining the nasal effects of both acetaldehyde and formaldehyde in which marked to
severe metaplasia and/or hyperplasia and increases in cell proliferation are observed prior to
nasal tumor formation during chronic exposure (Monticello et al., 1996; Zwart et al., 1988;
Woutersen et al., 1984; 1986; Appelman et al., 1982). A similar pattern of nasal lesions were
also noted in a 2-year chronic study evaluating the toxicology and carcinogenesis of
isobutyraldehyde, but no increases in neoplastic lesions were observed (NTP, 1999). In contrast,
the exposure concentrations required to induce similar nasal effects were higher compared to
formaldehyde and acetaldehyde. Thus, the pattern of nasal tissue effects and the carcinogenicity
of related aldehydes raise concern. However, the more specific alterations observed for related
aldehydes, such as squamous metaplasia with atypia and disorganization, concurrent hyperplasia,
changes in cell proliferation, and tumor formation in nasal tissues, were not observed after
exposure to propionaldehyde (Union Carbide, 1993). Therefore, as it relates to the effects
observed after exposure to propionaldehyde, the presence of squamous metaplasia alone is
considered to be a nonneoplastic lesion in nasal tissue and is of limited quantitative use in
assessing cancer risk.
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4.7. SUSCEPTIBLE POPULATIONS AND LIFE STAGES
4.7.1. POSSIBLE CHILDHOOD SUSCEPTIBILITY
No studies are available on possible childhood or other age group susceptibility to
propionaldehyde. However, in general, children may represent a particularly sensitive population
to the effects of airborne pollutants (Bateson and Schwartz, 2008). Their enhanced sensitivity is
most likely a result of their higher ventilation rates and undeveloped lungs relative to adults
which together may increase their exposure to toxicants and susceptibility to respiratory tract
injury.
4.7.2. POSSIBLE GENDER DIFFERENCES
No studies investigating the possible gender differences in susceptibility specific to
propionaldehyde are available.
4.7.3. POSSIBLE GENETIC DIFFERENCES
Wang et al. (2002) performed a genotype analysis of the ALDH2 gene in the livers of
human volunteers in order to investigate the metabolism of a variety of aldehydes. Of a total of
39 subjects, 8 were heterozygotes of the wild-type (ALDH2*1) and mutant (ALDH2*2) alleles,
and the others were homozygotes of the wild-type allele. The ability of mitochondria isolated
from these livers to metabolize propionaldehyde, acetaldehyde, formaldehyde, w-butyraldehyde,
capronaldehyde, and heptaldehyde was significantly (p < 0.05) lower (between 37 and 93%,
depending on the aldehyde; 80% for propionaldehyde) in the heterozygotes (ALDH2*l/*2)
compared to the homozygotes (ALDH2*1/*1), showing differences in metabolism between the
two genotypes. However, the mitochondrial activity was not lower for octylaldehyde,
decylaldehyde, retinaldehyde, benzaldehyde, 3-hydroxybenzaldehyde,
2,5-dihydroxybenzaldehyde, phenylacetaldehyde, and 3-phenylpropionaldehyde, showing
similar metabolism between the two genotypes. Based on these results, the authors hypothesized
that polymorphisms of the ALDH2 gene appear to exist in the human population, which may
alter the metabolism of the short aliphatic chain aldehydes. It is not clear, however, if the
potential increase to parent aldehyde exposure exists in vivo for heterozygotes.
4.7.4. POSSIBLE SENSITIVE SUBGROUPS - ASTHMATICS
Although no studies investigating the possible increased susceptibility of asthmatic
specific to propionaldehyde are available, asthmatics represent another potential sensitive
subgroup to inhaled irritant aldehydes (Singh and Busse, 2006; Leikauf, 2002). Asthmatics may
have an increased susceptibility to lower concentrations of inhaled irritants which can augment
symptoms in persons with asthma. Aggravation of underlying asthma can result from moderate
exposures (Nowak, 2002). One possible mechanism by which inhaled irritants may exert their
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effects is via airway sensory receptor-mediated reflex bronchoconstriction (Nowak, 2002).
Specifically, formaldehyde might exacerbate asthma and induce bronchoconstriction via irritant
mechanisms as well as via deregulation of the endogenous bronchodilator S-nitrosoglutathione
(Thompson and Grafstrom, 2008).
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5. DOSE-RESPONSE ASSESSMENTS
5.1. ORAL REFERENCE DOSE (RfD)
No human or animal oral studies for propionaldehyde were identified on which to base an
oral RfD.
5.2. INHALATION REFERENCE CONCENTRATION (RFC)
5.2.1. CHOICE OF PRINCIPAL STUDY AND CRITICAL EFFECT
No human inhalation studies are available for propionaldehyde. No subchronic or chronic
animal inhalation studies were identified for propionaldehyde. However, one short-term animal
inhalation study (Gage, 1970) and two short-term reproductive/developmental animal inhalation
studies were identified (Union Carbide, 1993, 1991). In addition, two acute animal studies were
identified (Steinhagen and Barrow, 1984; Egle, 1972b). The database for propionaldehyde is
further depicted in the Exposure-Response Array, Figure 5.1, and outlined in Table 5-1. The
exposure-response array shows the low- and high-exposure concentrations, and if identified, the
NOAEL and/or LOAEL (y-axis) for the respective study arranged by exposure duration,
endpoint and species (x-axis). The database consists of 5 animal (rat and mouse) studies.
1-minute 10-minutes
6-h/d
20-days x 20-days x 20-days x 48-days x 48-days x 52-days x
6-h/d 6-h/d 6-h/d 6-h/d 6-h/d 6-h/d
:
Q. '
-------
Table 5-1. Propionaldehyde References for Exposure-Response Array
Array (#)
1
2
o
J
3
4
4
4
5
5
Endpoint
Cardiovascular
RD50
Maternal body weight
Developmental toxicity
Maternal body weight
Developmental toxicity
Nasal pathology
Pathology/Clinical toxicity
Pathology/Clinical toxicity
Species
Rats
Mice
Rats
Rats
Rats
Rats
Rats
Rats
Rats
Reference
Egle (1972b)
Steinhagen and Barrow (1984)
Union Carbide (1991)
Union Carbide (1991)
Union Carbide (1993)
Union Carbide (1993)
Union Carbide (1993)
Gage (1970)
Gage (1970)
The Union Carbide (1993) study was selected as the principal study for derivation of the
RfC. Based on the database available for propionaldehyde, this study provided the most adequate
exposure concentration response and longest duration information for derivation of a reference
value. The study was conducted over a range of exposure concentrations, included a control
group, and demonstrated an exposure concentration-related effect more extensively than each of
the reported liver and cardiac effects described in Section 4.5.2. In addition, the studies
examining cardiac and liver effects were conducted over much shorter durations or required
much higher exposure concentrations to produce observable effects (Egle et al., 1973; Egle,
1972b; Gage, 1970). The critical endpoint chosen for analysis from this study was the incidence
of atrophy (diminished cell size and function) of the olfactory epithelium in male rats. This effect
is considered biologically relevant effect, exhibited a concentration-response relationship, and
was observed at the lowest exposure concentration tested (150 ppm). The atrophy observed at the
lowest exposure concentration was of minimal severity and not noted in females, possibly as a
result of the greater exposure duration of the male rats compared to the female rats in this study.
The atrophy observed at the middle exposure concentration (750 ppm) was characterized as
being of minimal to moderate severity. The induction of nasal lesions by propionaldehyde is
consistent with the irritant properties and the portal-of-entry effects observed in studies
conducted for other aldehydes (e.g., acetaldehyde, isobutyraldehyde and formaldehyde).
Along with an increased incidence of atrophy, an increased incidence of vacuolization of
the olfactory epithelium was also noted in propionaldehyde-exposed rats (Union Carbide, 1993).
Vacuolization (i.e., intracellular autophagy) is a normal cellular functional, homeostatic, and
adaptive response (Robbins and Angell, 1976). It is a characteristic of and often accompanies
cells/tissues undergoing atrophy (Kumar et al., 2004; Robbins and Angell, 1976). The presence
of these effects may also include observable inflammation and hypertrophic/hyperplastic
responses (Boorman et al., 1990). However, the qualitative and quantitative biological
relationship between vacuolization and atrophy is unclear and unknown. In vitro studies
conducted in nutrient depleted cells indicate that severe levels of vacuolization may also result in
37
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cell death via apoptosis or autophagic cell death characterized by an accumulation of autophagic
vacuoles (Gonzalez-Polo et al., 2005). Indications that either process occurred was not noted in
the principal study. High incidence levels of vacuolization were observed at 150 and 750 ppm
propionaldehyde. At 1,500 ppm, it appears that olfactory epithelium atrophy had progressed to
the point where cellular function was sufficiently diminished so that vacuolization was not
observed in this exposure group. In general, atrophied cells and tissues may have diminished
function, but are not considered to be devoid of function. However, atrophy may progress to
more severe cell injury and eventually cell death with continued exposure (Kumar et al., 2004).
Atrophy was chosen from the principal study as the critical endpoint because it is considered to
be an adverse effect and is observed, along with vacuolization, at the lowest exposure
concentration. This does not imply that the vacuolization observed in the same nasal tissue is not
relevant. Vacuolization is also considered to be a compound- related effect not noted in controls,
and typically accompanies atrophy. Vacuolization was observed in male and female animals
exposed at the lowest concentration. If vacuolization was a known precursor to atrophy, this
endpoint would be considered the critical effect. However, as this effect is considered an
autophagocytic response and accompanies the minimal atrophy at the lowest dose, atrophy was
selected as the critical effect and is considered an effect that is on the continuum to severe cell
injury and cell death.
The decrease in incidence and decreased severity of the nasal lesions in females relative
to males is likely to be attributable to the differences in exposure duration and approximate 6-day
period between cessation of exposures after GD 20 and sacrifice on PND 4. This observation
may also indicate that these effects are reversible and that repair and regeneration of the
olfactory epithelium has been initiated. Regeneration and repair of the olfactory epithelium are
dynamic processes characterized initially by disorganized cell proliferation of basal cells, which
may begin within 24 hours, but complete turnover of cells takes approximately 30 days
(Harkema et al., 2006; Hardisty et al., 1999). However, pathological indications (e.g., cell
proliferation, hyperplasia) that these processes were ongoing in the female rats were not noted.
Taken together, the nasal lesion data for propionaldehyde over the range of exposure
concentrations tested show a progression in both severity and incidence from no effects in
controls to manifestations of more definitive cellular injury, diminished cellular function, and
nasal tissue toxicity (i.e., atrophy accompanied by vacuolization, necrosis, and squamous
metaplasia). This progression was observed in both males and females. In addition, this pattern
of nasal lesion progression is similar to that observed with exposure to acetaldehyde in animals
(Woutersen et al., 1986; 1984; Appelman et al., 1982). In these studies, inhalation exposure to
acetaldehyde over a period for up to 28 months produced olfactory degeneration/atrophy with
and without hyperplasia/metaplasia at 4 weeks, followed by progression to focal basal cell
hyperplasia of the olfactory epithelium and squamous metaplasia of the respiratory epithelium at
12-15 months and finally by squamous cell carcinomas and adenocarcinomas at 16-28 months.
38
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The severity and incidence of these nasal effects were dependent on exposure concentration and
duration. A similar pattern and progression of nasal olfactory lesions were observed in rats
exposed to acetaldehyde for up to 65 exposure days (Dorman et al., 2008). Olfactory epithelial
degeneration, noted as the most sensitive endpoint observed, increased in incidence and severity
with both exposure concentration and duration. The presence of vacuolization was also noted,
but the severity was not graded. In this study, olfactory degeneration was observed prior to the
appearance of vacuolization upon interim sacrifice at each exposure concentration tested.
Vacuolization was not observed at exposure concentrations that did not induce degeneration. In
rats exposed chronically to isobutyraldehyde, nonneoplastic lesions in the nose consisted of
squamous metaplasia of the respiratory epithelium at concentrations >500 ppm, degeneration of
the olfactory epithelium at 2,000 ppm, and suppurative inflammation at 2,000 ppm (NTP, 1999).
No increases in neoplastic nasal lesions were observed in this study. Exposure to formaldehyde
for 13 weeks also produced similar effects in the nasal respiratory epithelium, consisting of
epithelial hyperplasia, squamous metaplasia, and increases in cell proliferation at concentrations
as low as 3 ppm (Zwart et al., 1988). Formaldehyde-induced nasal tumors are reported at
concentrations >6 ppm after chronic exposure (Monticello et al., 1996).
5.2.2. METHODS OF ANALYSIS
A benchmark concentration (BMC) analysis was conducted on the incidence of atrophy
of the olfactory epithelium in male rats as observed in the Union Carbide (1993) study. This
nasal lesion in male rats was the most biologically and lexicologically relevant response
identified, and the available concentration-response information supports the use of this
analytical approach. The results from the BMC analysis and the model outputs are discussed in
Section 5.2.3 and shown in Appendix B.
5.2.3. RFC DERIVATION—INCLUDING APPLICATION OF UNCERTAINTY
FACTORS (UFS)
The benchmark dose (BMD) approach provides the BMC and its 95% lower confidence
limit (BMCL) associated with a particular benchmark response (BMR). The BMCL is then used
as the point of departure (POD) in determining the RfC. A BMR of 10% extra risk of olfactory
atrophy was considered appropriate for derivation of the RfC under the assumption that it
represents a minimally biologically significant response level, owing to the minimal degree of
atrophy (i.e., 2/15 (-13%) animals responding with a severity scoring of minimal at the low dose
of 150 ppm) observed at this response level. This response level is also within the range of the
experimental data, minimizing extrapolation uncertainty. The critical effect, olfactory atrophy, is
compound related, biologically significant, consistent with lesion progression at higher exposure
concentrations, and not noted in control groups.
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Overall, the data were best fit by the Weibull model, which calculated a BMCio of
149.8 ppm or 366 mg/m3 and a BMCLio of 53.7 ppm or 128 mg/m3 (for details of this BMD
calculation see Appendix B). A BMR of 5% extra risk was also considered for comparison
purposes, and resulted in a BMCLos of 26. 1 ppm, about twofold lower than the BMCLio (see
Appendix B). The BMCLio was adjusted for duration from the experimental exposure regimen
of 6 hours/day, 7 days/week for 7 weeks (52 total exposures) to a continuous exposure as
follows:
BMCL10ADJ = 128 mg/m3 x 6/24x7/7
= 32 mg/m3
In accordance with the guidance for deriving inhalation RfCs (EPA, 1994), a regional gas
dose ratio (RGDR) for a gas with extrathoracic (i.e., nasal region to larynx) respiratory effects
was then derived by using a calculated ventilation rate (VE) of 0.264 L/minute (based on the
average body weight of the male CD rats reported in the principal study) and a default value of
13.8 L/minute for humans, along with default extrathoracic region surface area (SA) values of
15.0 cm2 for the rat and 200 cm2 for humans. The resulting equation is as follows:
RGDR
VE (human) / SA (human)
0.264/15
13.8/200
= 0.26
Applying the RGDR of 0.26 to the BMCLio/ADj of 32 mg/m3 yields a BMCLio/ADj
dosimetrically adjusted to a human equivalent concentration (HEC) (BMCLio HEC) of 3.4 ppm or
8 mg/m3.
The BMCLio/HEc of 3.4 ppm (8 mg/m3) was used as the POD for calculating the RfC, and
to this a total UF of 1,000 was applied: 3 (101/2) for extrapolation from animals to humans (UFA),
10 for intrahuman variability (UFH), 10 for subchronic to chronic duration (UFS), and 3 for
database deficiency (UFo).
A default UFA of 3 (101/2) was applied to account for interspecies (animal-to-human
extrapolation). This factor incorporates two areas of uncertainty given equal weight:
pharmacokinetics and pharmacodynamics. Because the pharmacokinetic component was
addressed in this assessment by the calculation of the HEC by applying the RGDR in
extrapolating from animals to humans according to the procedures in the RfC methodology
(EPA, 1994), only the pharmacodynamic component of this factor of uncertainty remains.
40
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A default UFn of 10 was applied for intraspecies uncertainty to account for human
variability and sensitive subpopulations as there was very limited information available to
definitively address the variability in the severity or range of response from propionaldehyde
exposure among individuals, and available data suggest there are differences among humans in
metabolism of propionaldehyde. Recent PBPK modeling investigating the impact of ALDH2
polymorphisms on rat and human nasal tissue dosimetry demonstrated a negligible impact on
olfactory tissue dose (Teeguarden et al., 2008). Additionally, application of this UF considers the
potential sensitivity of children and individuals with conditions such as asthma.
A default UFs of 10 was applied to account for adjustment from subchronic to chronic
duration. A subchronic study was used to derive the RfC, as no other supportive studies of
similar or longer durations were available for propionaldehyde.
A UFo of 3 (101/2) was applied to account for database deficiencies. The database for
propionaldehyde consists of several short-term inhalation animal studies, ranging from 6 days to
7 weeks in duration, and two reproductive/developmental toxicity studies. The database is
lacking a multigeneration reproductive toxicity study. Although the principal study used for the
RfC derivation was a reproductive/developmental study (Teeguarden et al., 2008), this study
provided limited reproductive and developmental information, since the pups were sacrificed on
PND 4 and pathology in the pups was not evaluated; only an external examination for the
presence of malformations was performed. Although limited nasal sectioning (i.e. 2-3 sections
compared to typical 4-6) was performed at necropsy, the critical effect identified was atrophy of
the olfactory epithelium in adult male rats (also observed in females), which is concordant with
the portal-of-entry effects attributable to the aldehydes acrolein, formaldehyde, acetaldehyde,
and isobutyraldehyde as well as other irritant gases. In addition, none of these aldehydes appear
to induce direct systemic effects, as measured by clinical chemistry and pathology, at exposure
concentrations that produce initial portal-of-entry effects. Similarly, propionaldehyde would not
be anticipated to have significant systemic distribution based on its deposition, solubility, and
reactivity in the respiratory tract. The uptake of propionaldehyde in the upper respiratory tract
measured in dogs is approximately 70-80% (Egle, 1972a). In the same study, moderate to high
respiratory tract uptake was observed for both acrolein (-80%) and formaldehyde (near 100%).
In the rat, acetaldehyde uptake in the upper respiratory tract averaged from 76 to 26% over a
concentration range of 1-1,000 ppm (Stanek and Morris, 1999; Morris and Blanchard, 1992). In
general, the toxicological information and limited kinetic information available for
propionaldehyde is consistent with other structurally related aldehydes and provides support for
41
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the critical effect chosen. However, the lack of a multigeneration reproductive toxicity study
warrants the application of a UFo of 3.
No LOAEL to NOAEL UF was applied since BMC analysis was used to determine the
POD, and this factor was addressed as one of the considerations in selecting the BMR. Based on
the data, a BMR of 10% change in the incidence of minimal olfactory atrophy was selected
under an assumption that it represents a minimal biologically significant change.
Application of a total UF of 1,000 (101/2 x 10 x 10 x 101/2) to the BMCLio HEC of
8 mg/m3 yields an RfC of 8 x l(T3 mg/m3.
5.3. CANCER ASSESSMENT
No studies are available on the carcinogenic effects of propionaldehyde on which to base
a cancer assessment.
5.4. GENERAL UNCERTAINTY IN THE PROPIONALDEHYDE NONCANCER AND
CANCER ASSESSMENT
The paucity of data for this compound, especially for those effects that could serve as
alternate sources for quantitative evaluation, prevent a further meaningful in-depth quantitative
analysis of uncertainty. It is anticipated, however, that the potential uncertainty of this
assessment could be informed both in qualitative and quantitative terms from the more robust
databases of the structurally related aldehydes, formaldehyde, acetaldehyde and
isobutyraldehyde. The areas of uncertainty for consideration in the assessment for
propionaldehyde are outlined in Table 5-2.
Table 5-2. Summary of general uncertainty in the propionaldehyde
noncancer and cancer risk assessments
Area of
consideration
Choice of study
Choice of noncancer
endpoint
Potential impact"
No RfC.
Use of cardiac
responses vs. olfactory
epithelium atrophy
could t RfC several-
fold.
Decision
Union Carbide
(1993) study chosen.
RfC is based on the
most biologically
relevant endpoint,
atrophy of olfactory
epithelium.
Justification
No alternative choices are available.
Chosen endpoint is consistent with expected
chemical irritation properties of agent and is
reasonably anticipated to be relevant for humans for
the same reasons. Cardiac responses observed in
acute studies conducted at exposure concentrations
at least eightfold higher than those showing nasal
effects.
42
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Area of
consideration
Human relevance of
data (portal-of-entry
vs. site-specific
respiratory tract
effect)
Potential deficiency
in necropsy of target
tissue
Choice of gender
Choice of species
POD derivation
method for
noncancer RfC
Choice of model for
BMCL derivation
Statistical
uncertainty at POD
Use of dosimetry in
calculation of HEC
Potential impact"
Assuming no relevance
of results would
indicate that RfC may
be unnecessarily low or
not applicable.
Limited sectioning per
animal may have
resulted in missed
lesions that could
underestimate actual
incidence per exposure
group, assuming such
lesions would be
observed in all sections
and underestimate risk
such that the RfC could
possibly be |.
RfC could be t or | if
based on another
gender.
RfC could be t or | if
based on another
species.
Little difference as
LOAEL is at 13%
response and thus is
near the BMCLJ o
Other models t (approx.
1.5 -fold) or I (approx.
1.3-fold) RfC.
POD would be -40%
higher if BMC (vs.
BMCL) were used.
Use of dosimetry
increases scientific
robustness of
assessment.
Decision
Assume human
relevancy.
Use Union Carbide
(1993) study (only
available repeated-
concentration study).
RfC is based on
olfactory atrophy in
males. Males are
observed to be more
sensitive possibly as
a result of study
design.
RfC is based on the
most clearly relevant
endpoint in the only
species tested, rat.
BMD method used.
Weibull model
chosen.
BMCL used per U.S.
EPA BMD guidance
(EPA, 2000c).
Apply dosimetry.
Justification
Due to the irritation-type mode of action involving
the general reactivity of the functional group (i.e.,
aldehyde) with tissue constituents regardless of
source or site within the respiratory tract, there is
comparatively little uncertainty concerning
applicability of relevance to humans.
This same reasoning may be used to assume site
concordance (i.e., portal-of-entry) although the
relative effects at different sites in the respiratory
tract may differ between species due to differences
in airflow patterns and regional uptake within the
respiratory tract.
Although sectioning in target tissues (nasal tract)
was limited (two sections vs. typical three to six per
animal), effects, including atrophy, were found at all
concentrations. The pathology findings are
consistent with nasal lesions observed after exposure
to other aldehydes and irritants.
Although progression of nasal effects is seen in both
males and females, there was a clear decrease in
incidence and decreased severity in females (likely
to be attributable to the differences in exposure
duration approximate 6-day period between
cessation of exposures after GD 20 and sacrifice on
PND 4 versus continued exposure in males during
this period). Comparable incidence data from
females not available based on this study design.
Only species tested in the available study.
Comparable effects for propionaldehyde in other
strains or species not known.
Advantages include capacity to account for sample
size that is quantitatively reflected in providing
confidence bounds on dose.
U.S. EPA (2000c) BMD technical guidance used to
choose best fitting model.
Limited size of bioassay results in sampling
variability; lower bound is 95% confidence interval
on administered exposure.
Dosimetry methodology accommodates estimation
of dose at the site of toxicity (nasal tract), thus
providing target-tissue dosimetry.
43
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Area of
consideration
Potential impact3
Decision
Justification
Human population
variability
Risk unknown.
Default 10-fold
uncertainty factor
applied to derive the
RfC value.
10-fold UF is applied principally because of lack of
definitive and quantifiable information on the
variability of response with this mode of action.
Specific subgroups (e.g. asthmatics and children)
may be more sensitive to inhaled irritants and
aldehydes, but definitive information for
propionaldehyde is lacking. The default factor for
intrahuman variability is used to ensure that the risk
to chemicals and stressor are not underestimated.
Potential for cancer
Risk unknown.
Note concern for
carcinogenic
potential.
The presence of the more resilient squamous
metaplasia (without atypia) is an anticipated
response of airway portal-of-entry tissues being
exposed to irritants such as aldehydes. However, the
presence of nasal tumors in conjunction with
squamous metaplasia in lifetime studies of related
aldehydes (formaldehyde and acetaldehyde) raises a
concern that cannot be addressed with the
propionaldehyde since the Union Carbide (1993)
study was only 7 weeks in duration.
= increase; J, = decrease.
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6. MAJOR CONCLUSIONS IN THE CHARACTERIZATION OF HAZARD
AND DOSE RESPONSE
6.1. HUMAN HAZARD POTENTIAL
Propionaldehyde is an aldehyde used primarily to manufacture polyvinyl, other plastics,
and propionic acid. It is released to the environment mainly through wood and gasoline
combustion and from municipal waste incinerators. Propionaldehyde has been detected in
ambient air, indoor air, and drinking water (NLM, 2004). Propionaldehyde is also a component
of both mainstream and sidestream cigarette smoke (Counts et al., 2005). The primary route of
exposure to propionaldehyde is expected to be via inhalation. No studies on the effects of
propionaldehyde administered by the oral route have been performed. Propionaldehyde has also
been approved by both U.S. FDA and WHO/JECFA as a synthetic flavoring ingredient for direct
addition to food; the alcohol (propanol) and acid (propionic acid) are similarly approved (FDA,
2003; WHO, 1999; IPCS, 1998).
Limited data are available on the pharmacokinetics of propionaldehyde. In an inhalation
study conducted in dogs, Egle (1972a) determined that the animals retained approximately 70-
80% of the inspired concentration of propionaldehyde. An in vitro study in a rat hepatoma cell
line showed propionaldehyde to be efficiently metabolized via aldehyde dehydrogenase (Bassi et
al., 1997). Wang et al. (2002) performed a genotype analysis of the ALDH2 gene in human
volunteers and found polymorphisms in the ALDH gene that appeared to alter propionaldehyde
metabolism. It is not clear, however, if this alteration would lead to a significant increase in
parent aldehyde exposure in those individuals with specific polymorphisms of this gene. A rat
study demonstrated increased urinary excretion of propionaldehyde, formed via lipid
peroxidation, with age and for animals on a restricted diet (De Tata et al., 2001).
No studies in humans are available for propionaldehyde. No subchronic or chronic oral
animal studies are available for the chemical. However, three short-term inhalation animal
studies, ranging from 6 days to 7 weeks in duration, are available. Gage (1970) exposed male
and female rats to 90 ppm propionaldehyde 6 hours/day for 20 exposures or to 1,300 ppm
propionaldehyde 6 hours/day for 6 days. No changes in body weight or clinical signs were noted.
Microscopic examination revealed liver cell vacuolation in animals exposed to 1,300 ppm
propionaldehyde. Two short-term rat developmental inhalation studies conducted by Union
Carbide (1993, 1991) are also available. In a range-finding study (Union Carbide, 1991),
maternal toxicity was noted as exposure-related decreases in body weight gain were observed at
exposure concentrations of 1,000 ppm and above. However, these decreases in body weight gain
were accompanied by decreases in food consumption throughout the gestation period. In the high
concentration group, there was a significant reduction in fetal body weights, but no other
evidence of any treatment-related external malformations or variations was observed. In the
second study, young adult male and female rats were exposed to propionaldehyde during a
45
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2-week premating period and a 14-day mating phase (Union Carbide, 1993). The mated females
were exposed daily through GD 20 for a minimum of 35 days and a maximum of 48 days. The
males continued to be exposed until sacrifice in week 7, for a total of 52 exposures. No
significant effects of exposure on any of the reproductive parameters assessed were found. Litter
size and viability were similar among the groups. Absolute pup body weights on PND 0 and 4
were not affected by exposure, although, at the high concentration, body weight gain for that
period was significantly depressed. The biological significance of this finding is difficult to
assess, since changes in absolute body weight were not demonstrated and the period of
observation was relatively short. The most significant exposure-related effects were found in the
nasal cavity. In the adult females, microscopic examination revealed propionaldehyde-induced
vacuolization in the low and intermediate exposure groups and atrophy of the olfactory
epithelium in the low, intermediate, and high exposure groups. The incidence of atrophy
increased with exposure concentration. No evidence of squamous metaplasia was found in
olfactory or respiratory epithelium. In the adult males, as in the females, microscopic
examination revealed exposure-related effects in the olfactory epithelium, consisting of
vacuolization and atrophy in the low, intermediate, and high exposure groups. The incidence of
atrophy increased with exposure concentration and was greater than observed in the females. In
both males and females, the severity of this nasal lesion increased with exposure concentration.
In males only, a low incidence of squamous metaplasia in the respiratory epithelium was
reported in both the intermediate and high exposure groups. A decrease in incidence and
decrease in severity of the nasal lesions in females relative to males was observed and could be
attributable to the differences in exposure duration and approximate 6-day period between
cessation of exposures after GD 20 and sacrifice on PND 4. This observation may also indicate
that these effects are reversible and that repair and regeneration of the olfactory epithelium has
been initiated. However, pathological indications (e.g., cell proliferation, hyperplasia) that these
processes have started in the female rats were not noted.
Squamous metaplasia was noted as a compound-related lesion in the upper airways of
rats exposed to propionaldehyde. Although the occurrence of this lesion, especially in the upper
airways, may occur as a response to repeated irritation whereby a resistant type of epithelium
replaces a more susceptible one, it has also been noted along with nasal tumors in lifetime
studies of related aldehydes, including formaldehyde and acetaldehyde. Thus, this pattern of
nasal tissue effects in this relatively short-term study and nasal carcinogenicity of related
aldehydes raises some concern for the carcinogenic potential of this compound.
The genotoxicity of propionaldehyde has been studied in bacteria and a number of
mammalian cells in vitro. Propionaldehyde was found to be mutagenic in S. typhimurium strain
TA1534 (Sampson and Bobik, 2008) and nonmutagenic in all other strains tested (Dillon et al.,
1998; Aeschbacher et al., 1989; Mortelmans et al., 1986), but produced concentration-related
increases in HGPRT (with notable decreases in cell viability) and ouabain mutants in V79
46
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hamster cells (Brambilla et al., 1989). Propionaldehyde produced a concentration-related
increase in chromosome aberrations in Chinese hamster embryonic cells (Furnus et al., 1990)
and chromosome breaks in CHO cells (Seoane and Dulout, 1994). In addition, propionaldehyde
induced a concentration-related increase in unscheduled DNA synthesis in rat, but not human,
hepatocytes (Martelli, 1997; Martelli et al., 1994) and a weak, concentration-related increase in
DPXs in cultured human lymphoma cells (Costa et al., 1997). Propionaldehyde also formed
protein adducts with hemoglobin in vitro (Hoberman and San George, 1988).
Two studies have shown that propionaldehyde produces concentration/dose-related
changes in blood pressure and heart rate after inhalation or i.v. administration in rats (Egle et al.,
1973; Egle, 1972b). A study on mouse lymphocytes demonstrated significant inhibition of T-
lymphocyte and B-lymphocyte proliferation, with no effects on cell viability (Poirier et al.,
2002). Studies on the toxicity relationships (in terms of cytotoxicity) among propionaldehyde
and other aldehydes showed that acrolein was the most toxic compound, formaldehyde next,
followed by acetaldehyde, and finally propionaldehyde, with the conclusion that cytotoxicity
generally decreased with increasing (saturated) aldehyde chain length (Bombick and Doolittle,
1995; Koerker et al., 1976). Similar relationships among various aldehydes were noted when
comparing RDso values in mice (Steinhagen and Barrow, 1984). The a,P~unsaturated aliphatic
aldehydes (acrolein and crotonaldehyde) and formaldehyde were approximately two orders of
magnitude more potent than the saturated aliphatic aldehydes (e.g., propionaldehyde,
acetaldehyde butyraldehyde, and isobutyraldehyde) in producing a 50% decrease in respiration
rate. In a review by Guth (1996), it was concluded from a comparison of the effects of
propionaldehyde and acetaldehyde for a variety of endpoints that there should not be major
differences in toxicity between acetaldehyde and propionaldehyde. Similarly, analysis and
comparison of the nasal lesion data between acetaldehyde and propionaldehyde supports this
conclusion.
Based on the information provided from animal studies, the most likely adverse human
health effects that would be anticipated from exposure to propionaldehyde would be primarily
respiratory tract irritation and secondarily cardiovascular perturbations. No human health effects
data or chronic animal bioassay studies are available that assess the carcinogenic effects of
propionaldehyde. Therefore, in accordance with the Guidelines for Carcinogen Risk Assessment
(EPA, 2005a), there is "inadequate information to assess the carcinogenic potential" for
propionaldehyde.
6.2. DOSE RESPONSE
Quantitative estimates of cancer risk for propionaldehyde were not developed due to the
lack of data on the potential carcinogenicity of the compound.
Quantitative estimates of noncancer risk from the oral route of exposure were not
developed for propionaldehyde because of the lack of human or animal data.
47
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A quantitative estimate of the noncancer risk for the inhalation route of exposure was
developed from animal data, since no human data are available. An RfC of 8 x icr3 mg/m3 was
derived from the incidence data of olfactory atrophy in adult male rats reported in a 7-week
(52 total exposures) reproductive and developmental study conducted by Union Carbide (1993).
BMC analysis of this data was best fit by the Weibull model, which calculated a BMCLio of
53.7 ppm or 128 mg/m3.
The RfC was derived by duration adjusting the BMCLio of 128 mg/m3 from the
experimental exposure regimen of 6 hours/day, 7 days/week for 7 weeks (52 total exposures) to a
continuous exposure yielding a BMCLio/ADj of 32 mg/m3. Applying the RGDR calculated for a
gas with extrathoracic respiratory effects of 0.26 (EPA, 1994) resulted in an HEC (BMCLio/HEc)
of 8 mg/m3. The BMCLi0/HEc was used as the POD for calculating the RfC. A total UF of 1000
was applied: 3 (101/2) for extrapolation from animals to humans (UFA), 10 for intrahuman
variability (UFH), 10 for subchronic to chronic duration (UFS), and 3 for database deficiency
(UFD). Application of a total UF of 1000 (10'/2 x 10 x 10 x io'/2) to the BMCLi0/HEc of 8 mg/m3
yielded an RfC of 8 x l(T3 mg/m3.
Confidence in the principal study (Union Carbide, 1993) is judged to be low to medium
because few details were provided specific to the study results. In addition, the key study
provided limited developmental information as the pups were sacrificed on PND 4 and pathology
was not evaluated; only an external examination for the presence of malformations was
performed. However, the critical effect identified was atrophy of the olfactory epithelium in
adult male rats (also observed in females), which is concordant with the portal-of-entry effects
attributable to irritant gases and other aldehydes. Thus, this endpoint is supported by the
aldehyde inhalation exposure-effects database as a whole. Confidence in the critical effect
identified in the principal study is medium. Confidence in the overall database specific to
propionaldehyde is low because there are no additional and/or supporting subchronic or chronic
animal studies available to evaluate and support the concentration-response effect of
propionaldehyde on multiple endpoints. Therefore, confidence in the RfC is judged to be low to
medium.
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APPENDIX A. SUMMARY OF EXTERNAL PEER REVIEW AND PUBLIC
COMMENTS AND DISPOSITION
The "Toxicological Review of Propionaldehyde" has undergone a formal external peer
review performed by scientists in accordance with EPA guidance on peer review (EPA, 2006b).
For the external peer review, the reviewers were tasked with providing written answers to
general questions on the overall assessment and on chemical-specific charge questions,
addressing key scientific issues of the assessment. A summary of significant comments made by
the external reviewers and EPA's responses to these comments arranged by charge question
follow. Editorial comments were considered and incorporated into the document as appropriate
and are not discussed further. EPA also received comments from the public. A summary of
public comments and EPA's responses are also included.
EXTERNAL PEER REVIEWER COMMENTS
General Charge Questions:
Charge Question 1: Is the Toxicological Review logical, clear and concise? Has EPA
accurately, clearly and objectively represented and synthesized the scientific evidence for
noncancer and cancer hazard?
Comment:
All four reviewers commented that the Toxicological Review was written in a logical and
generally clear and concise manner, although there is some repetition in various sections. The
document provides a strong review of the available data relative to derivation of an RfC value for
propionaldehyde. It does fall short of the ideal in some key respects resulting from the lack of a
dedicated long-term inhalation toxicity study which would address the question of possible
carcinogen! city. Two of the reviewers agreed that a strength of the document is that it evaluates
the toxicity information on propionaldehyde in the context of the complete database on
structurally similar aldehydes such as formaldehyde and acetaldehyde while recognizing that a
full structure activity relationship analysis is not possible. The use of this comparative structure-
activity approach to address uncertainties and data gaps in regard to propionaldehyde is
particularly useful in presenting a coherent overall picture. In this regard inclusion of information
on isobutyraldehyde toxicity would be highly valuable. Weaknesses in the database are clearly
delineated; the document appropriately identifies the principal study while clearly indicating its
deficiencies. Two of the reviewers also commented that the document correctly identifies the
critical effect and does so in a logical, clearly expressed and transparent fashion. The overall
conclusion that olfactory atrophy represents the critical effect is sound and demonstrates a strong
knowledge of the nasal toxicological issues that represent important considerations in a risk
evaluation. Since the structurally related compound acetaldehyde also produces this lesion the
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text correctly highlights the current information available on this compound. One reviewer
commented that the derivations are consistent with EPA policy but that the scientific basis for
the default nasal dosimetric assumptions is questionable. Overall, the four reviewers agreed that
the review accurately and succinctly synthesizes the available scientific evidence for noncancer
effects in a clear manner. The available studies are accurately represented and the selection of the
principal study is presented in an objective manner.
Response:
For the sake of completeness and clarity, information on the structurally-related aldehydes (i.e.
butyraldehyde and isobutyraldehyde) was added to the genotoxicity section where available. The
results and a discussion of the chronic NTP study (1999) conducted for isobutyraldehyde was
also added to complete the comparison discussion of related aldehyde-induced portal-of-entry
effects. For this assessment, an attempt was made to include and compare only the genotoxicity
and portal-of-entry results from the other aldehydes (formaldehyde, acetaldehyde, butyraldehyde
and isobutyraldehyde) that are relevant to those observed for propionaldehyde. Therefore, the
review is not meant to reflect information from the aldehyde database as a whole. However, a
few genotoxicity results for longer-chain aldehydes are included for comparison although in vivo
toxicity studies are lacking. Comments regarding EPA policy on nasal dosimetry assumptions
will be considered whenever EPA revisits this guidance issue.
Charge Question 2: Please identify any additional studies that should be considered in the
assessment of the noncancer and cancer health effects of propionaldehyde.
Comment:
Two of the four reviewers were unaware of any additional studies that should be considered in
the assessment of propionaldehyde. Two of the four reviewers suggested the additional studies
by Dorman et al. (2008), Teeguarden et al. (2008), Sampson and Bobik (2008), Oyama et al.
(2007), and the NTP (1999) study conducted for isobutyraldehyde should be considered in the
draft assessment. Additional studies for possible inclusion were noted similarly in response to
other charge questions.
Response:
These references, as well as others cited specifically in response to other charge questions, were
reviewed and incorporated into the assessment as appropriate.
Charge Question 3: Please discuss research that you think would be likely to increase the
confidence in the database for propionaldehyde in future assessments.
In response to this question, the reviewers offered the following points:
• The identified areas of uncertainly provide sound direction for future research. For
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example, a well designed and performed chronic inhalation study for propionaldehyde
which includes complete nasal sectioning would enhance the database, as would a
multigenerational study. In addition, since there are no carcinogenicity data, a two year
bioassay would provide useful information. The priority for such a study depends on the
degree of importance one places on the carcinogenic response to acetaldehyde vs.
isobutylaldehyde relative to the potential response to propionaldehyde. Finally, since the
current dosimetric extrapolation procedures of the US EPA are so controversial, precise
information on the inhalation dosimetry of propionaldehyde would strongly aid in the
formation of a scientifically based quantitative inhalation risk assessment of this
compound.
• A major deficiency of the existing database is the lack of a long-term inhalation
carcinogenicity study. Supporting studies of deposition in the respiratory tract, cell
proliferation and formation of DNA-protein crosslinks have provided extensive
additional understanding of the effects of formaldehyde and, to a lesser extent,
acetaldehyde. It would be helpful if at least some of these studies were extended to
include propionaldehyde.
• It was stated that the literature search was conducted before July, 2007. We found no
other studies with propionaldehyde that are relevant to health effects. However, a main
argument in the RfC derivation is that propionaldehyde is lexicologically similar in many
aspects to acetaldehyde. There were recently two publications on acetaldehyde
inhalation: Teeguarden et al., 2008. A PBPK Model for Evaluating the Impact of
Aldehyde Dehydrogenase Polymorphisms on Comparative Rat and Human Nasal Tissue
Acetaldehyde Dosimetry. Inhalation Toxicology 20:375-390, 2008, and Dorman et al.,
2008. Derivation of an Inhalation Reference Concentration Based Upon Olfactory
Neuronal Loss in Male Rats Following Subchronic Acetaldehyde Inhalation. Inhalation
Toxicology 20:245-256, 2008. These publications should be considered in future risk
assessments of propionaldehyde. As far as the current review is concerned, these
publications only strengthen the argument for using olfactory atrophy as the critical
effect, since this was also the endpoint selected for acetaldehyde.
Response:
The assessment has been updated to incorporate the newer information available from the studies
on acetaldehyde published since July 2007 as well as the results from the new study on
propionaldehyde mutagenicity in salmonella.
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Charge Question 4: Please comment on the identification and characterization of sources of
uncertainty in sections 5 and 6 of the assessment document. Please comment on whether the key
sources of uncertainty have been adequately discussed. Have the choices and assumptions made
in the discussion of uncertainty been transparently and objectively described? Has the impact of
the uncertainty on the assessment been transparently and objectively described?
Comment:
All four reviewers commented that the majority of uncertainties were appropriately identified
and characterized, and that Table 5-1 is quite useful in this regard. The impact of the
uncertainties on the assessment has been transparently and objectively described.
One of the four reviewers noted that the potential for reactive irritants to exacerbate conditions
such as asthma should be addressed in the document.
Two of the four reviewers commented that the human relevance of data in the uncertainty
discussion could be enhanced by adding points that the relative effects at different sites in the
respiratory tract may differ between species due to differences in airflow patterns and regional
uptake within the respiratory tract. The major consideration being that animals with more
extensive nasal passages than found in humans may have greater effect in the upper respiratory
tract, while there may be greater penetration into the lower respiratory tract in humans.
Response:
A discussion of the effect irritants may have on sensitive subpopulations such as asthmatics was
added to the document (Section 4.7.4).
Additional discussion points concerning the human relevancy of the animal data in consideration
of potential differences of specific site of effect in the respiratory tract was added to the
document.
Chemical-Specific Charge Questions:
(A) Oral reference dose (RfD) for propionaldehyde
Charge Question 1: No oral RfD has been derived in the current draft assessment based on the
lack of studies available that examine the effects of propionaldehyde administered via the oral
route. Are there available studies missing from the draft document that might be useful for
deriving an oral RfD or that should be considered in this decision?
Comment:
None of the four reviewers were aware of any studies that would inform development of an RfD.
One of the four reviewers suggested that a reference could be made that formaldehyde and
acetaldehyde appear to be less hazardous by the oral route compared to the inhalation route
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(Morris et al., 1996). One of the four reviewers suggested that some insight into tolerable levels
of propionaldehyde taken in orally would be useful.
Response:
The write up in section 2 of the assessment does provide information on the intake levels of
propionaldehyde considered to be tolerable as a food additive (WHO, 1999). The Morris et al.
1996 citation was added to the text.
(B) Inhalation reference concentration (RfC) for propionaldehyde
Charge Question 1: The current draft IRIS assessment for propionaldehyde uses a combined
reproductive/developmental exposure study by Union Carbide (1993) as the principal study for
the derivation of the RfC. Please comment on whether the selection of this study as the principal
study has been scientifically justified. Has this study been transparently and objectively
described in the document? Please identify and provide the rationale for any other studies that
should be selected as the principal study. Is this study appropriate for use in this assessment?
Comment:
Two of four reviewers commented that the selection of this study of the principal study was
scientifically justified. One of four reviewers commented that the selection of this study was
transparently and objectively described. Two of the four reviewers also commented that the
description of the principal study was appropriately or clearly described in the document. Two of
four reviewers also commented that as appropriately noted, this study is far from ideal for the
purpose, is the only study available, but nevertheless is sufficient to provide a basis for the RfC.
One reviewer also commented that the concerns in using this study have been adequately
addressed in this document.
One of the four reviewers also commented that the section on the comparative toxicity of
aldehydes represents a strong component of the overall risk assessment. In addition, given the
structural similarity between acetaldehyde and propionaldehyde and the similarity of nasal
lesions induced by both compounds, the recent study of Dorman et al., (2008) on subchronic
acetaldehyde inhalation toxicity should appropriately be included. Similarly information on
inhalation toxicity of butyraldehyde and isobutyraldehyde including the NTP studies should be
included as well.
Response:
Discussions of the data relevant to propionaldehyde from studies on acetaldehyde,
butyraldehyde, and isobutyraldehyde have been added to the appropriate sections of the
document.
Charge Question 2: Has the most appropriate critical effect (increase incidence of olfactory
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atrophy in male rats,) presented in Sections 4.3 and 4.5.2 of the Toxicological Review been
selected? Is the rationale for this selection transparently and objectively described in the
document? Please comment on whether the selection of this critical effect has been scientifically
justified. Please comment on the choice of olfactory atrophy as the critical effect as opposed to
other endpoints (e.g., vacuolization) and the rationale that this endpoint was chosen because it is
on the continuum leading to overtly adverse effects such as cell death. Has the qualitative
pathological relationship between effects observed been adequately and appropriately
characterized? Please provide a detailed discussion. Please identify and provide the rationale for
any other endpoints that should be used instead of increased incidence of olfactory atrophy in
male rats to develop the RfC.
Comment:
Based on the propionaldehyde database, three of the four reviewers were in general agreement
that the choice of the critical endpoint lesion is scientifically justified, reasonable, appropriate,
and represents the critical response. One reviewer noted, however, due to the paucity of the
database this was the only effect that could be selected for derivation of the RfC. Had other
studies been available, perhaps this effect would not have been the one of greatest biological
significance. One reviewer further commented that the rationale for the selection of the critical
response is scientifically sound, transparently and objectively described, and is consistent with
the critical effects for other reactive aldehydes such as formaldehyde, acetaldehyde and acrolein.
One reviewer also noted that the description of the lesions is adequate to relate propionaldehyde-
induced lesions to the lesions induced by these other nasal toxicants and that olfactory atrophy
represents a toxic lesion. Two of the four reviewers commented that the data clearly indicate a
concentration response continuum for these lesions and use of 150 ppm as the LOAEL, based on
olfactory atrophy and vacuolization is appropriate. The incidence of olfactory atrophy has a well-
defined dose response curve for establishing a POD and extrapolating to human health effects.
Three of the four reviewers commented and were in general agreement on the characterization of
the continuum of the qualitative pathological lesions and their concentration-response
relationships - with vacuolization at the lower end and more severe effects, including various
degrees of metaplasia and even carcinogenesis, at the upper end. However, one reviewer
commented that the justification for placing a specific cut-off level for these effects at the level
of atrophy as opposed to vacuolization is questionable and the discussion could be improved in
that it is inappropriate to argue that the vacuolization is for some reason an "adaptive" or
"compensatory" response which is not relevant, as opposed to the slightly more severe atrophy.
Response:
The choice of atrophy as the critical endpoint instead of vacuolization was not meant to imply
that a level of vacuolization as a response is not relevant. The choice was made for atrophy as the
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critical endpoint because it is the endpoint most closely associated with adversity (a decrease in
cell function). Nothing in the literature could be cited indicating vacuolization is a precursor
effect to atrophy (or any other lesion) in nasal tissue - only that the two effects often are
observed together and are part of a continuum possibly leading to more severe effects with
increasing exposure concentration and duration. Similar to propionaldehyde, the presence of
vacuolization was noted along with olfactory degeneration in rats exposed to acetaldehyde
(Dorman et al., 2008). In this study, the incidence of vacuolization decreased as atrophy
increased with exposure concentration. The severity of vacuolization was not noted in this study
and olfactory degeneration was considered to be the most sensitive endpoint. In addition,
degenerative olfactory lesions were observed prior to the detection of vacuolization upon interim
sacrifice at all exposure concentrations tested. Vacuolization was not detected at exposure
concentrations that did not induce degeneration.
As noted elsewhere, discussion of the Dorman et al. (2008) study was added to the appropriate
sections of the document.
Charge Question 3: BMD methods were applied to incidence data on olfactory atrophy in male
rats to derive the POD for the RfC. Please provide comments with regard to whether BMD
modeling is the best approach for determining the POD. Has the BMD modeling been
appropriately conducted and objectively and transparently described? Has the BMR selected for
use in deriving the POD (i.e., 10% extra risk of olfactory atrophy) been scientifically justified?
Please comment on EPA's decision to treat all cases of olfactory atrophy similarly, without
consideration of the severity of the atrophy seen at different dose levels. Please provide a
detailed discussion and any suggestions for consideration of severity in determining the POD
including identifying and provide rationales for any alternative approaches for the determination
of the POD and discussion of whether such approaches are preferred to EPA's approach
considering the available data.
Comment:
All four reviewers commented that the BMD approach is the preferred method and suitable for
determining the POD for this data.
Two of the four reviewers commented that the choice of a 10% response rate was justifiable and
consistent with the animal data and represents a biologically minimal significant response level.
One of these two reviewers also commented that the value in the severity scores (increasing with
exposure concentration) serves as justification for the use of olfactory atrophy as the critical
effect.
A-7
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Two reviewers commented that in consideration of severity and incidence, there may be some
justification for reduction to the 5% level from the proposed 10% because the data appear more
like a LOAEL than a NOAEL for straightforward quantal responses in animal toxicity studies.
Therefore, it might be more appropriate to choose a BMR of 5%, and the 95% lower confidence
limit on the dose producing this response rate, as the POD for this type of data.
Response:
All endpoints observed were considered for BMC modeling, however, only the atrophy data
were amenable to this approach. Other modeling approaches (e.g. Categorical Regression) were
considered for the data as a means to incorporate severity. However, it was determined that
based on the nature of the responses observed - vacuolization and atrophy - that a degree of
subjectivity in grading the severity among responses would be introduced. For example, would
marked vacuolization be considered more severe than mild atrophy? A logical pathological
progression model would be needed to definitively address such issues. In addition, evaluation of
the individual animal data did not result in any definitive correlation between severity of
vacuolization and atrophy. Therefore, the spectrum and severity of incidence effect data was
used qualitatively to support our choice of endpoint to be modeled using BMD - atrophy.
As noted in response to the comments for Question 4, justification for using a BMR of 10% in
determining the POD is provided in the text and consistent with EPA guidance and the power of
the study (i.e. n = 15).
A-8
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Charge Question 4: Please comment on the selection of the uncertainty factors applied to the
POD for the derivation of the RfC. For instance, are they scientifically justified and transparently
and objectively described in the document?
Comment:
Two of the four reviewers commented that the selection of uncertainty factors is clearly and
transparently described, appear to follow EPA policy, and are consistent with precedent. One
reviewer further commented that the factor of 10 for inter-individual differences is appropriate
because of the known human polymorphisms in aldehyde dehydrogenase and the likely role of
this enzyme in detoxifying propionaldehyde. Presumably this factor of 10 is sufficient to account
for the potential sensitivity of persons with chronic lung disease such as asthma or COPD. This
reviewer commented that these points might be explicitly stated in the text.
One reviewer commented that the basis for the default value of 3 for interspecies extrapolation
was not clear.
One reviewer commented that if the proper POD (i.e. the lower 95% confidence limit on the dose
producing a 5% rather than 10% response) for a quantal analysis had been selected the UF
estimates selected and their justification would be acceptable. This reviewer discussed the use an
alternative pseudo-continuous score-based variable assuming that any statistically
distinguishable deviation constitutes an unacceptable impact (especially if the continuous change
constitutes a demonstrably adverse and undesirable change at higher levels of response, as here).
Response:
Text describing the presumptions and intent of the application of the UF for inter-individual
differences was added to the document.
Other approaches (e.g., Categorical Regression) and PODs for the data were considered in
accordance with EPA guidance. However, each approach was considered to add subjectivity into
analyzing the data specifically in regards to grading severity among the responses. Converting
the incidence data to continuous data as suggested would also increase this subjectivity.
Therefore, the spectrum and severity of incidence effect data was used qualitatively to support
our choice of endpoint to be modeled using BMD - atrophy. The choice of a 10% BMR is
consistent for the type of incidence data, effect level, and number of animals in accordance with
the BMD Technical Guidance. The POD calculated (53 ppm) is also consistent with the
observation of a NOAEL for acetaldehyde-induced nasal effects (vacuolization and
degeneration) of 50 ppm (Dorman et al., 2008). Justification for this approach is provided in the
document.
The use of a factor of 3 (to account for dynamics) for interspecies extrapolation is used when the
RGDR approach (accounting for kinetics) is used to calculate an HEC from animal data.
A-9
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Charge Question 5: Please comment specifically on the database uncertainty factor of 3 applied
in the RfC derivation. Please comment on the body of information regarding reproductive and
developmental toxicity on propionaldehyde, the relevance of toxicity data on other aldehydes,
and the relevance of toxicokinetic data regarding the likelihood of portal-of-entry effects as the
critical effects in the determination of the database uncertainty factor. Please comment on
whether the selection of the database uncertainty factor for the RfC has been scientifically
justified. Has this selection been transparently and objectively described in the document?
Comment:
Three of the four reviewers agreed that a database UF of 3 is appropriate for the data cited,
which is restricted to consideration of possible deficiencies in the animal study database due to
the lack of a multi-generational study. One of these three reviewers noted a limitation of the
database is the absence of chronic toxicity data, however a UF =10 was included for subchronic
to chronic extrapolation. This reviewer also noted that a UF = 3 for this deficiency is consistent
with other EPA documents previously reviewed. One of these three reviewers was concerned
that the UF database does not take into account possible post-natal effects such as asthma. In
addition, one reviewer commented on highlighting the modeling results for acetaldehyde
(Dorman et al., 2008; Teeguarden et al., 2008) to provide insight into the interspecies
extrapolation for propionaldehyde. One of these three reviewers also commented that the lack of
nasal sectioning performed for propionaldehyde compared to typical practices warrants mention
here as well as in Table 5-2.
One of the four reviewers commented that the justification provided in the document does not
support the value of 3 but would support a value of 10 since the database is clearly lacking a long
term study.
Response:
The justification of the application of a UF of 3 for database deficiencies in deriving the RfC was
clarified in the document. The aldehyde database - which includes longer-term studies showing
lack of systemic effects at concentrations producing similar portal-of-entry effects and
information on respiratory tract dosimetry for three structurally-related aldehydes - was used to
support the choice of endpoint as well as the limited propionaldehyde database. Information
specific to propionaldehyde (and definitive studies for other aldehydes) on the exacerbation of
asthma is scant. A database UF of 3 was applied primarily due to the lack of a multigenerational
study and is consistent with EPA practices.
Lack of a chronic study is addressed under a separate UF.
A discussion of the acetaldehyde modeling results in the context of interspecies extrapolation
was added to the uncertainty factor discussion.
A-10
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Consideration of limited nasal sectioning was also added to the text. Although limited sectioning
was performed, had nasal pathology not been evaluated - especially in light of the known nasal
effects induced by other aldehydes - consideration for a database UF of 10 would have been
warranted.
(C) Carcinogenicity of propionaldehyde
Charge Question 1: Under the EPA's 2005 Guidelines for Carcinogen Risk Assessment
(www.epa.gov/iris/backgr-d.htm), the Agency concluded that data are inadequate for an
assessment of the human carcinogenic potential of propionaldehyde Please comment on the
scientific justification for the cancer weight of evidence characterization. In addition, has the
Agency properly characterized the potential for concern for carcinogenicity of propionaldehyde
based on the available data on propionaldehyde and other aldehydes?
Comment:
All four reviewers commented that the document properly concludes that there are insufficient
data to characterize the carcinogenic potential of propionaldehyde. In addition, the reviewers
agreed with the effort to use structure/activity comparisons with other aldehydes for both
carcinogenicity and mutagenicity in order to place the potential concern for carcinogenicity in
context. However, one reviewer commented that the database for comparison should also include
isobutyraldehyde for the sake of completeness and transparency. The most compelling aspect of
the comparison between upper respiratory tract lesions in these three compounds is that all three
show progression with exposure concentration and duration on a similar continuum of lesion
severity. Three of the four reviewers noted that the finding of increased DPX in vitro for
propionaldehyde (and acetaldehyde) compared to formaldehyde should be placed into context as
one reviewer commented that formaldehyde may represent a special case because its deposition
is so focal and because it's ability to form DPX is so much greater (orders of magnitude?) than
the other aldehydes. One reviewer also noted that the recent positive finding of mutagenicity in
Salmonella (Sampson and Bobik, 2008) be put into context as well considering that the majority
of other mutagenicity tests were negative.
Response:
The results for isobutyraldehyde in comparison with the other aldehydes discussed were added
where appropriate. In addition, the DPX results for formaldehyde, acetaldehyde and
propionaldehyde have been added and put into context. The study results on mutagenicity have
also been added to the genotoxicity section as well as the genotoxicity summary.
A-ll
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Other comments from the External Peer Reviewers:
All reviewers provided editorial comments on the document as well as additional text and studies
that could be added to further improve the clarity of the assessment.
Response:
These comments were all addressed and incorporated as appropriate within the scope of the
assessment and in accordance with EPA policy and procedures.
Comments from the Public
Comment:
One reviewer commented on the analysis of the genotoxicity and carcinogenicity in the draft
assessment concerning what they viewed as inappropriate degree of weight given to comparisons
with formaldehyde and acetaldehyde, but not with other structurally-related aldehydes (e.g.
isobutyraldehyde), thus suggesting an incomplete analysis of the data available on other
aldehydes.
Response:
This comment was given serious consideration and aspects of it were incorporated into the
assessment to provide a more complete analysis. It should be noted that this assessment was not
meant to be a complete analysis of the available data on all aldehydes, only those structurally-
related aldehydes for which data were available on similar genotoxic, nonneoplastic, and
carcinogenic endpoints in order to provide a degree of potency and effect comparisons with
propionaldehyde. That being stated, more complete information, where available, is provided in
the text for all of these endpoints, including the addition of comparative information for
isobutyraldehyde for which a 2-year bioassay is available from NTP (1999). Therefore, EPA
believes a more complete and robust analysis is provided.
Comment:
One reviewer provided the link to the Material Safety and Data Sheet (MSDS) for
propionaldehyde: http://www.sciencelab.com/xMSDS-Propionaldehyde-9924730. And provided
the following comment: "Since there is no information available on safety of this product on
humans it should never be used on this earth. It is clear that not enough testing has been done."
Response:
MSDS sheets are readily available for public reading from a variety of sources and thus, are not
part of the assessment document.
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APPENDIX B. BENCHMARK CONCENTRATION MODELING RESULTS
Benchmark concentration modeling was performed to identify potential critical effect
levels for derivation of the RfC for propionaldehyde. The modeling was conducted according to
draft EPA guidelines (EPA, 2000c) by using benchmark dose software (BMDS) Version 1.4.1,
available online from EPA (http://www.epa.gov/ncea/bmds.htm). A brief discussion of the
modeling results is presented below.
The incidence data for atrophy of the olfactory epithelium in male rats from the Union
Carbide (1993) study were chosen as the critical endpoint for benchmark analysis. The incidence
data are depicted in Table B-l, and the various modeling output results at the designated BMR of
10% (BMCio) are summarized in Table B-2. A BMR of 10% change in the incidence of minimal
olfactory atrophy was selected under an assumption that it represents a minimal biologically
significant change (see Section 5.2.3). Graphical representation of the model of choice is shown
in Figure B-l. As shown in Table B-2, several of the models had similar Akaike Information
Criteria (AICs) and overall chi-square values (scaled residuals) and fit for the data at the lowest
exposure concentration, 150 ppm. In accordance with benchmark dose technical guidance (U.S.
EPA, 2000c), the Weibull model was chosen as the model for use in derivation of the RfC
because it was the model with the lowest AIC and it had a lower-scaled residual at the exposure
concentration closest to the BMCio compared to the model with the next lowest AIC (i.e., the
multistage 1). The corresponding BMCLio of 53.7 ppm was used in further derivation of the RfC.
For comparison purposes, the modeling results depicting a BMR of 5% change in the incidence
of minimal olfactory atrophy are shown in Figure B-2. The BMCos was calculated to be 94.6
ppm, and the corresponding BMCL0s to be 26.1 ppm.
Table B-l. Olfactory atrophy incidence data in male rats exposed to various
concentrations of propionaldehyde
Exposure concentration
Oppm
150 ppm
750 ppm
1,500 ppm
Incidence of olfactory
atrophy
0/15
2/15
10/15
15/15
Source: Union Carbide (1993).
B-l
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Table B-2. BMC model outputs for olfactory atrophy
Model
Weibulf
Multistage 1
Gamma
Probit
Logistic
BMCio
(ppm)
149.8
61.2
142.6
145.7
146.9
BMCLio
(ppm)
53.7b
42.6
50.2
79.5
62.9
AIC
35.97
36.33
36.42
37.52
37.86
x2
0.81
2.24
1.07
1.87
2.04
p Value
0.6659
0.5238
0.5852
0.3912
0.3612
X2 residual,
150 ppm
0.4275
-0.871
0.3104
0.3387
0.3737
aModel of choice (see text for details).
b53.7ppm= 128mg/m3.
Source: Union Carbide (1993).
Weibull Model with 0.95 Confidence Level
Weibull
0.8
0.6
o 0.4
o
ctf
0.2
0
BMCL..
0 200 400 600 800 1000 1200 1400
Exposure Concentration (ppm)
Figure B-l. BMCio Weibull model for olfactory atrophy (Union Carbide, 1993).
B-2
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Weibull Model with 0.95 Confidence Level
0 200 400 600 800 1000 1200 1400
Exposure Concentration (ppm)
Figure B-2. BMCos Weibull model for olfactory atrophy (Union Carbide, 1993).
B-3
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