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EPA
TOXICOLOGICAL REVIEW
OF
Cerium Oxide and Cerium
Compounds
(CAS No. 1306-38-3)
In Support of Summary Information on the
Integrated Risk Information System (IRIS)
September 2009
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.
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CONTENTS—TOXICOLOGICAL REVIEW OF CERIUM OXIDE
AND CERIUM COMPOUNDS (CAS NO. 1306-38-3)
LIST OF TABLES v
LIST OF FIGURES vi
LIST OF ABBREVIATIONS AND ACRONYMS vii
FOREWORD ix
AUTHORS, CONTRIBUTORS, AND REVIEWERS x
1. INTRODUCTION 1
2. CHEMICAL AND PHYSICAL INFORMATION 3
3. TOXICOKINETICS 8
3.1. ABSORPTION 8
3.1.1. Oral Exposure 8
3.1.2. Inhalation Exposure 11
3.2. DISTRIBUTION 12
3.2.1. Oral Exposure 12
3.2.2. Inhalation Exposure 13
3.3. METABOLISM 14
3.4. ELIMINATION 15
3.5. PHYSIOLOGICALLY BASED TOXICOKINETIC MODELS 16
4. HAZARD IDENTIFICATION 17
4.1. STUDIES IN HUMANS—EPIDEMIOLOGY, CASE REPORTS, CLINICAL
CONTROLS 17
4.1.1. Oral Exposure 17
4.1.2. Inhalation Exposure 18
4.2. SUBCHRONIC AND CHRONIC STUDIES AND CANCER BIO ASSAYS IN
ANIMALS—ORAL AND INHALATION 20
4.2.1. Oral Exposure 20
4.2.2. Inhalation Exposure 22
4.3. REPRODUCTIVE/DEVELOPMENTAL STUDIES—ORAL AND INHALATION .. 30
4.4. OTHER DURATION-OR ENDPOINT-SPECIFIC STUDIES 30
4.4.1. Acute Toxicity Studies (Oral and Inhalation) 30
4.4.2. Acute Studies (Injection) 31
4.4.2.1. Neurobehavioral andNeurodevelopmentalEffects 31
4.4.2.2. Neurological Effects 32
4.4.2.3. Hepatic Effects 33
4.4.2.4. Cardiovascular Effects 35
4.4.2.5. HematologicalEffects 36
4.4.2.6. Renal Effects 37
4.4.3. Genotoxicity 37
4.5. MECHANISTIC DATA AND OTHER STUDIES 38
4.5.1. In Vitro Studies 38
iii
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4.5.2. Ex Vivo Studies 40
4.5.3. Nanoparticle Studies 40
4.6. SYNTHESIS OF MAJOR NONCANCER EFFECTS 45
4.6.1. Oral 45
4.6.2. Inhalation 45
4.6.3. Mode-of-Action Information 46
4.6.3.1. Respiratory Tissues 46
4.6.3.2. Other Tissues 50
4.7. EVALUATION OF CARCINOGENICITY 52
4.7.1. Summary of Overall Weight of Evidence 52
4.7.2. Synthesis of Human, Animal, and Other Supporting Evidence 52
4.8. SUSCEPTIBLE POPULATIONS AND LIFE STAGES 52
4.8.1. Possible Childhood Susceptibility 52
4.8.2. Possible Gender Differences 52
4.8.3. Possible Susceptible Populations 53
5. DOSE-RESPONSE ASSESSMENTS 54
5.1. ORAL REFERENCE DOSE (RfD) 54
5.1.1. Choice of Principal Study and Critical Effect—with Rationale and Justification 54
5.1.2. Previous RfD Assessment 55
5.2. INHALATION REFERENCE CONCENTRATION (RfC) 55
5.2.1. Choice of Principal Study and Critical Effect—with Rationale and Justification 5 5
5.2.2. Methods of Analysis—Including Models (PBTK, HMD, etc.) 57
5.2.3. RfC Derivation—Including Application of Uncertainty Factors (UFs) 59
5.2.4. RfC Comparison Information 61
5.2.5. Previous RfC Assessment 64
5.3. UNCERTAINTIES IN THE ORAL REFERENCE DOSE (RfD) AND INHALATION
REFERENCE CONCENTRATION (RfC) 64
5.4. CANCER ASSESSMENT 67
6. MAJOR CONCLUSIONS IN THE CHARACTERIZATION OF HAZARD AND DOSE
RESPONSE 68
6.1. HUMAN HAZARD POTENTIAL 68
6.2. DOSE RESPONSE 70
6.2.1. Noncancer/Oral 70
6.2.2. Noncancer/Inhalation 71
6.2.3. Cancer/Oral and Inhalation 73
7. REFERENCES 74
APPENDIX A: SUMMARY OF EXTERNAL PEER REVIEW AND PUBLIC COMMENTS
AND DISPOSITION 83
APPENDIX B: BENCHMARK DOSE MODELING RESULTS FOR THE DERIVATION OF
THE RFC 99
IV
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LIST OF TABLES
Table 2-1. Physical properties of cerium and cerium compounds 4
Table 2-2. Major uses of selected cerium compounds 7
Table 4-1. Hematological changes in male and female Sprague-Dawley rats following inhalation
of cerium oxide aerosol 6 hours/day, 5 days/week for 13 weeks 24
Table 4-2. Absolute lung weight in rats exposed to cerium oxide aerosol 6 hours/day, 5
days/week for 13 weeks 25
Table 4-3. Relative lung weight in rats exposed to cerium oxide aerosol 6 hours/day, 5
days/week for 13 weeks 25
Table 4-5. Relative spleen weight in rats exposed to cerium oxide aerosol 6 hours/day, 5
days/week for 13 weeks 25
Table 4-5. Results of gross pathological examination of lungs of rats exposed to cerium oxide 6
hours/day, 5 days/week for 13 weeks 26
Table 4-6. Results of gross pathological examination of bronchial, medistinal, and pancreatic
lymph nodes of rats exposed to cerium oxide 6 hours/day, 5 days/week for 13 weeks 26
Table 4-7. Incidences of selected histopathological lesions in rats exposed to cerium oxide
aerosol 6 hours/day, 5 days/week for 13 weeks 28
Table 4-8. NOAEL and LOAEL values for effects observed in BRL (1994) 29
Table 5-1. Selected models and BMCs and BMCLs for the increased incidence of alveolar
epithelial hyperplasia in the lungs of male and female rats 58
Table 5-2. Output from RDDR v.2.3 used in the analysis in Section 5.2.2 59
Table B-l. Benchmark Dose modeling used in the derivation of the RfC 100
v
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LIST OF FIGURES
Figure 5-1. Exposure-response array of selected toxicity effects from the BRL (1994) study.... 62
Figure 5-2. Points of Departure for selected endpoints from Figure 5-1 with corresponding
applied uncertainty factors and derived chronic inhalation RfVs 63
VI
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LIST OF ABBREVIATIONS AND ACRONYMS
ALT alanine aminotransferase
AST aspartate aminotransferase
BMC benchmark concentration
BMD benchmark dose
CASRN Chemical Abstract Service Registry Number
CFU colony-forming unit
CI confidence interval
COH coumarin 7-hydroxylase
CT computer tomography
CYP cytochrome P450 (in connection with isozyme abbreviations)
CYP450 cytochrome P450
DAF dosimetric adjustment factor
DCF dichlorodihydrofluorescein diacetate
DPO dendriform pulmonary ossification
EDTA ethylenediamine tetraacetic acid
EGTA ethylene glycol bis(2-aminoethylether)tetraacetic acid
EPA Environmental Protection Agency
GD gestation day
GI gastrointestinal
GSD geometric standard deviation
GSH glutathione
HEC human equivalent concentration
HEI Health Effects Institute
i.v. intravenous
IRIS Integrated Risk Information System
LCso median lethal concnetration
LDX dose that kills X% of animals
LDH lactate dehydrogenase
LOAEL lowest-observed-adverse-effect level
MCP monocyte chemoattractant protein
MDA malodialdehyde
MMAD mass median aerodynamic diameter
MT metallothionein
NOAEL no-observed-adverse-effect level
NTP National Toxicology Program
OCT ornithine-carbamyl transferase
PAM pulmonary alveolar macrophage
PBTK physiologically based toxicokinetic
PCNA proliferating cell nuclear antigen
PND postnatal day
RDDR regional deposited dose ratio
RfC reference concentration
RfD reference dose
vn
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SDH sorbitol dehydrogenase
SOD superoxide dismutase
UF uncertainty factor
Vlll
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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 cerium
oxide and cerium compounds. It is not intended to be a comprehensive treatise on the chemical
or toxicological nature of cerium oxide and cerium compounds.
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 the 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
hotline.iris@epa.gov (email address).
IX
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AUTHORS, CONTRIBUTORS, AND REVIEWERS
CHEMICAL MANAGER
Martin Gehlhaus
National Center for Environmental Assessment
U.S. Environmental Protection Agency
Washington, DC
AUTHORS
Martin Gehlhaus
National Center for Environmental Assessment
U.S. Environmental Protection Agency
Washington, DC
Mark Osier, Ph.D.
Environmental Science Center
Syracuse Research Corporation
Syracuse, NY
Fernando Llados, Ph.D.
Environmental Science Center
Syracuse Research Corporation
Syracuse, NY
Daniel Plewak
Environmental Science Center
Syracuse Research Corporation
Syracuse, NY
Michael Lumpkin, Ph.D.
Environmental Science Center
Syracuse Research Corporation
Syracuse, NY
Marc Odin, Ph.D.
Environmental Science Center
Syracuse Research Corporation
Syracuse, NY
Andrew Rooney, Ph.D.
National Center for Environmental Assessment
U.S. Environmental Protection Agency
RTF, NC
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REVIEWERS
This document has been 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.
INTERNAL EPA REVIEWERS
Ted Berner
Office of Research and Development
National Center for Environmental Assessment
Kevin Dreyer, M.S, Ph.D.
Office of Research and Development
National Health and Environmental Effects Research Laboratory
Experimental Toxicology Division
Pulmonary Toxicology Branch
Lynn Flowers, Ph.D, DABT
Office of Research and Development
National Center for Environmental Assessment
Mary Jane Belgrade, Ph.D.
Office of Research and Development
National Health and Environmental Effects Research Laboratory
Immunotoxicology Branch
Jamie Strong, Ph.D.
Office of Research and Development
National Center for Environmental Assessment
EXTERNAL PEER REVIEWERS
Mitchell D. Cohen, Ph.D.
New York University School of Medicine
Joe L. Mauderly, D. V.M.
Lovelace Respiratory Research Institute
Giinter Oberdorster, Ph.D., D. V.M.
University of Rochester School of Medicine and Dentistry
XI
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Katherine S. Squibb, Ph.D.
University of Maryland School of Medicine
John M. Veranth, Ph.D.
University of Utah
Department of Pharmacology and Toxicology
xn
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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 cerium
oxide and cerium compounds. IRIS Summaries may include oral reference dose (RfD) and
inhalation reference concentration (RfC) values for chronic and less-than-lifetime exposure
durations, and a carcinogenicity assessment.
The RfD and RfC 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 may also be derived for acute
(<24 hours), short-term (>24 hours up to 30 days), and subchronic (30 days up to 10% of
average lifetime) exposure durations, all of which are derived based on an assumption of
continuous exposure throughout the duration specified.
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. 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 are derived from the application of a low-dose
extrapolation procedure. 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 ug/m3 air breathed.
Development of these hazard identification and dose-response assessments for cerium
oxide and cerium compounds has followed the general guidelines for risk assessment as set forth
by the National Research Council (1983). EPA guidelines and Risk Assessment Forum
Technical Panel Reports that were used in the development of this assessment include the
following: Guidelines for Mutagenicity Risk Assessment (U.S. EPA, 1986b), Guidelines for
Developmental Toxicity Risk Assessment (U.S. EPA, 1991), Guidelines for Reproductive Toxicity
Risk Assessment (U.S. EPA, 1996b), Guidelines for Neurotoxicity Risk Assessment (U.S. EPA,
1998a), Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005a), Supplemental Guidance
for Assessing Susceptibility from Early-Life Exposure to Carcinogens (U.S. EPA, 2005b),
1
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Recommendations for and Documentation of Biological Values for Use in Risk Assessment (U.S.
EPA, 1988), (proposed) Interim Policy for Particle Size and Limit Concentration Issues in
Inhalation Toxicity (U.S. EPA, 1994a), Methods for Derivation of Inhalation Reference
Concentrations and Application of Inhalation Dosimetry (U.S. EPA, 1994b), Use of the
Benchmark Dose Approach in Health Risk Assessment (U.S. EPA, 1995), Science Policy Council
Handbook: Peer Review (U.S. EPA, 1998b, 2000a, 2005c), Science Policy Council Handbook:
Risk Characterization (U.S. EPA, 2000b), Benchmark Dose Technical Guidance Document
(U.S. EPA, 2000c), and A Review of the Reference Dose and Reference Concentration Processes
(U.S. EPA, 2002).
The literature search strategy employed for this compound was based on the Chemical
Abstract 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 2009.
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2. CHEMICAL AND PHYSICAL INFORMATION
Cerium is a member of the lanthanide series of metals and is the most abundant of the
rare-earth elements in the earth's crust (average concentration of 50 ppm) (Hedrick, 2004).
Elemental cerium is an iron-gray, ductile, malleable metal (O'Neil, 2001). Cerium metal is very
reactive and is a strong oxidizing agent that is stabilized when associated with an oxygen ligand
(Kilbourn, 2003). When present in compounds, cerium exists in both the trivalent state (Ce3+,
cerous) and the tetravalent state (Ce4+, eerie) (Kilbourn, 2003; Reinhardt and Winkler, 2002).
Chemical structures and selected chemical and physical properties of cerium and cerium
compounds are listed in Table 2-1.
Cerium is found in nature along with other lanthanide elements in the minerals alanite,
bastanite, monazite, cerite, and samarskite; however, only bastanite and monazite are important
sources commercially (Lide, 2005; Kilbourn, 2003). Because of its unique stability in the
tetravalent state (other lanthanides are stable in only the trivalent state), cerium can be separated
out from the other rare-earth elements through oxidation (forming CeC^) followed by variable
solubility filtration (Reinhardt and Winkler, 2002). Cerium salts can be prepared by liquid-
liquid extraction from rare-earth cerium-containing solutions. Cerium metal is prepared by
reacting CeFs with an excess of calcium at approximately 900°C (Kilbourn, 2003). It can also be
obtained by the fused-salt electrolysis of a mixture of cerium chlorides and fluorides (Reinhardt
and Winkler, 2002).
Cerium is most heavily used in the form of mischmetal for metallurgical purposes
(Kilbourn, 2003; Reinhardt and Winkler, 1996). Cerium is the major component of mischmetal
(50-75% by weight for the most common grades), a commercial mixture of metallic light
lanthanides prepared by the electrolysis of mixed lanthanide chlorides and fluorides obtained
from bastanite or monazite (Kilbourn, 2003; Reinhardt and Winkler, 2002). Mischmetal reacts
with the impurities found in metals to form solid compounds, thereby reducing the effect of these
impurities on the properties of the metal (Reinhardt and Winkler, 2002). Mischmetal has been
used in the manufacture of steel to improve shape control, reduce hot shortness, and increase
heat and oxidation resistance. It can be added to cast iron to improve ductility, toughness, and
microstructure. Mischmetal is also used in the manufacture of cerium-iron alloy lighter flints
(Kilbourn, 2003; Reinhardt and Winkler, 2002).
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Table 2-1. Physical properties of cerium and selected cerium compounds
Name
CASRN
Synonyms
Structure
Molecular weight
Molecular formula
Form
Melting point
Density
Water solubility
Other solubility
Cerium
7440-45-1
140.116
Ce
Iron-gray, ductile,
malleable metal
798°C; boiling point =
3443°C
6.770 g/cm3
Decomposes slowly
with cold water and
rapidly with hot water
Soluble in dilute
mineral acids
Cerium oxide
1306-38-3
Cerium dioxide; ceria;
eerie oxide; cerium(IV)
oxide
°^ ^°
^Ce
172.11
CeO2
Pale-yellow, heavy powder
(white when pure);
commercial product is
brown
2400°C
7.65 g/cm3
Insoluble in water
Insoluble in dilute acid
Hydrated cerium oxide
12014-56-1 (hydroxide)
23322-64-7 (hydrate)
Hydrated eerie oxide; cerium
hydrate; eerie hydroxide;
cerium(IV) hydroxide; cerium
perhydroxide; cerium
tetrahydroxide
?H
HO— Ce-OH
OH
or
>'[OH2]
0 L -12
208.148
CeO2-2H2O
The hydroxide precipitate is
amorphous and on drying,
converts to hydrated eerie oxide;
whitish powder when pure.
Not available
Not available
Insoluble in water
Soluble in concentrated mineral
acid
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Table 2-1 [continued]. Physical properties of cerium and selected cerium
compounds
Name
CASRN
Synonyms
Structure
Molecular weight
Molecular formula
Form
Melting point
Density
Water solubility
Other solubility
Cerium nitrate
13093-17-9
Cerium tetranitrate;
nitric acid,
cerium(4+) salt
O
,N+ Ce
O*NXO'
L J4
388.1369
Ce(N03)4
Not available
Not available
Not available
Not available
Not available
Cerous chloride
7790-86-5
Cerium(III) chloride;
cerium trichloride
246.48
CeCl3
White crystals; fine
powder
817°C
3.97g/cm3
Soluble in water
Soluble in ethanol
Cerous fluoride
7758-88-5
Cerium(III) fluoride;
cerium trifluoride
197.11
CeF3
Hexagonal crystals or
powder
1430°C
6.157g/cm3
Insoluble in water
Soluble in acids
(monohydrate)
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Table 2-1 [continued]. Physical properties of cerium and selected cerium
compounds
Name
CASRN
Synonyms
Structure
Molecular weight
Molecular formula
Form
Melting point
Density
Water solubility
Other solubility
Cerous acetate
537-00-8
Cerium(III) acetate; cerium
triacetate; acetic acid;
cerium(3+) salt
r o i
_A Ce3+
[O CH3J3
317.251
Ce(C2H302)3
Not available
Not available
Not available
Not available
Not available
Cerous citrate
512-24-3
Cerium citrate;
cerium(III) citrate;
cerium(3+)
2-hydroxypropane-
1,2,3 -tricarboxy late
/? ^°H
o — \ | o
i Ce3+
cA-°
329.219
C6H807.Ce
Not available
Not available
Not available
Not available
Not available
Sources: ChemlDplus (2006); Lide (2005); Lewis (2001); O'Neil (2001).
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Exposure to commercially used cerium compounds is most likely through exposure to
cerium (eerie) oxide (CeO2). It is used either in the pure form or in a concentrate as a polishing
agent for glass mirrors, plate glass, television tubes, ophthalmic lenses, and precision optics
(Kilbourn, 2003; Reinhardt and Winkler, 2002). Cerium oxide is used as a glass constituent to
prevent solarization and discoloration (especially in the faceplates of television screens)
(Reinhardt and Winkler, 2002). Cerium oxide is also used in emission control systems in
gasoline engines and as a diesel fuel-born catalyst to reduce particulate matter emissions (Health
Effects Institute [HEI], 2001; Reinhardt and Winkler, 1996). Cerium nitrate has been used as a
topical treatment for burn wounds (Monafo et al., 1976). Major uses for selected cerium
compounds are listed in Table 2-2.
Table 2-2. Major uses of selected cerium compounds
Name
Cerium oxide
Hydrated cerium oxide
Cerous chloride
Cerous fluoride
CASRN
1306-38-3
23322-64-7
7790-86-5
7758-88-5
Use
Polishing and decolorizing glass; opacifier in vitreous enamels
and photochromic glasses; heat-resistant alloy coatings; as a
cracking catalyst; as a catalyst for automobile emission control; in
ceramic coatings; in phosphors; in cathodes; in capacitors; in
semiconductors; in refractory oxides; gemstone polishing
Production of cerium salts and cerium oxide
In the manufacturing of cerium metal and cerium salts; catalyst for
the polymerization of olefins
In the preparation of cerium metal; in arc carbons to increase
brilliance
Sources: Kilbourn (2003); HEI (2001); Lewis (2001); O'Neil (2001); Wells and Wells (2001).
Cerium is not expected to exist in elemental form in the environment since it is a reactive
metal (Lewis, 2001). Cerium compounds are not expected to volatilize and will exist in the
particulate form if released into the air. For cerium compounds that are soluble in water, Ce3+
would likely have a pKa close to La3+ (8.5) (Wulfsberg, 2000), which indicates that the hydrated
Ce3+ ion ([Ce(H2O)n]3+) will remain in solution at environmental pHs (4-9). The hydrated Ce4+
ion ([Ce(H2O)n]4+) is expected to hydrolyze and polymerize at environmental pH (Cotton et al.,
1999) and may precipitate out of solution. In general, metal cations in solution are attracted to
the surfaces of soil particles, and the extent of adsorption to soils will depend on the soil
characteristics (e.g., pH, mineral content, organic content) (Evans, 1989).
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3. TOXICOKINETICS
Many of the studies of the toxicokinetics of cerium were conducted using radioactive
cerium. Stable and radioactive cerium are expected to behave in a similar toxicokinetic manner
and possess the same chemical properties. Radioactive cerium is a beta-emitter. As such, data
from studies using either stable or radioactive cerium are presented below. Data characterizing
the toxicokinetics of cerium compounds, such as cerium oxide, are discussed in order to inform
the overall database.
3.1. ABSORPTION
3.1.1. Oral Exposure
Studies evaluating the absorption of cerium compounds following oral exposure in
humans are not available.
In adult animals, cerium compounds are poorly absorbed following oral exposure, while
suckling animals exhibit higher absorption and retention of cerium in the gastrointestinal (GI)
tissues. Observed absorption of radioactive cerium salts from the GI tract of adult rats ranged
from 0.05% to less than 0.1% of the administered dose (Kostial et al., 1989b; Inaba and
Lengemann, 1972; Shiraishi and Ichikawa, 1972). Suckling rats, however, absorbed 40-98% of
administered dose, with the youngest rats retaining the largest percentage of the dose (Kostial et
al., 1989a, b; Inaba and Lengemann, 1972).
Four litters of Sprague-Dawley rats (n = 7-9), age 0, 7, 14, or 26 days, were given a
single dose of [141Ce]-ceric nitrate of unreported concentration by intragastric dosing (Inaba and
Lengemann, 1972). Subsequent, periodic whole-body radioactivity measurements were taken
immediately after dosing and periodically thereafter. In another experiment in this study, two
litters of 1-day-old newborn rats were given single doses of [141Ce]-ceric nitrate, and one rat
from each litter was sacrificed at 1, 3, 5, 7, and 10 days after dosing, after which radioactivity in
the GI tract and whole body was measured and macro-autoradiographs of the GI tract were
produced.
It was observed that on or about day 16 of life, rats began consuming a solid, grain-based
diet and were completely weaned by day 26 (Inaba and Lengemann, 1972). In weanling rats
(rats dosed at 26 days of age), only 0.04% of the administered radioactivity remained in the body
by 3 days after dosing. However, radioactivity in newborns diminished more slowly, dropping
from 98% of administered dose on day 1 to 29% on day 16. The GI tract accounted for nearly
100% of whole-body retention on day 1 and 93% on day 16. After the onset of weaning in the
suckling rats dosed as newborns, whole-body radioactivity fell to 3% of administered dose by
day 24, of which only 17% was measured in the GI tract. Autoradiographs of the GI tract from
the two litters dosed as 1-day-old newborns suggest rapid transit of radioactivity to the lower
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small intestine 1 day after exposure. Autoradiography of rats 5 days after dosing (6 days old)
suggests that the intestinal radioactivity is restricted to the upper two-thirds of the epithelial villi,
which the study authors associated with cerium concentration in the vacuoles.
Kostial et al. (1989b) administered single oral doses of an unreported concentration of
[141Ce]-cerous chloride by intragastric dosing to 6-day-old and 6-8-week-old rats (strain
unreported) to investigate whether distribution of cerium (and other metals) in the GI tract
differed in suckling versus mature rats. Six-day-old rats were measured for whole-body and gut
radioactivity 2, 4, 6, and 12 days after dosing, while adult rats were measured 2 days after
dosing. Orally administered cerium chloride was more readily retained in the whole body, gut,
and carcass of suckling rats than in older rats. The ileum was the main site for cerium chloride
accumulation in the suckling rats following oral administration, while the stomach and large
intestine were the main sites for cerium chloride accumulation in the 6-8-week-old rats. Whole-
body radioactivity 6 days postexposure in 2-week-old suckling rats orally dosed with [141Ce] was
40% of the administered dose, of which 95% was found in the gut. However, whole-body
radioactivity 6 days postexposure in adult (6-8-week-old) rats was only 0.05%, of which 66%
was found in the gut, mostly in the stomach and cecum.
Shiraishi and Ichikawa (1972) administered single oral doses of an unreported
concentration of [144Ce]-cerous chloride by intragastric dosing to 0-, 7-, 14-, and 21-day-old
juvenile and 100-day-old adult Wistar rats. Cortisone acetate, which alters the morphology of
the absorptive epithelium of the small intestine, was injected to a group of 7-day-old rats 4 days
prior to [144Ce]-cerous chloride exposure to observe its effect on whole-body retention.
Periodically (up to 70 days after dosing), small groups of rats from all age groups were sacrificed
and measured for radioactivity in the whole-body, gut, and various excised tissues. The decrease
in retention of whole-body radioactivity among suckling rats dosed at 0, 7, and 14 days after
birth was approximately 11, 6.5, and 1.5%, respectively, of the administered dose. Weanling and
adult rats exhibited a rapid decrease in whole-body radioactivity through 10 days following
dosing. At 2 weeks after dosing, the 21- and 100-day-old adults had whole-body radioactivity of
0.08 and 0.018%, respectively, of the administered dose. The intestinal content accounted for
most of the retained radioactivity in neonates until the age of weaning. Cortisone acetate
treatment resulted in a more rapid loss of the oral dose from young rats, although whether this is
due to lower uptake by intestinal cells or a more rapid release to feces is not presently clear.
This investigation demonstrated that the whole-body retention of cerium chloride by suckling
rats was greater than the retention by weanling and adult rats, and this increased retention by
suckling rats may be due to increased pinocytic activity in the absorptive epithelium of sucklings
(Shiraishi and Ichikawa, 1972).
Yorkshire piglets treated with [144Ce]-cerous chloride by gavage on the first or fourth day
after birth and sacrificed 4 or 18 or 3 or 21 days, respectively, after dosing absorbed 2.5-8% of
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the administered dose (Mraz and Eisele, 1977). Absorption was threefold greater in piglets
treated at 1 day of age versus those treated at 4 days of age. Body content of cerium chloride did
not differ significantly between piglets sacrificed at the earlier dates and those sacrificed later,
indicating that absorption was almost complete within 3-4 days.
Eisele et al. (1980) gave single gavage doses of either [144Ce]-cerous chloride or
[144Ce]-cerous citrate (concentration unreported) to 0-6- or 6-24-hour-old CsH mice and
Sprague-Dawley rats and to 6-24-hour-old Yorkshire piglets. Radioactivity levels were
measured in the GI tract and other tissues, including the remaining carcass, at days 1, 5, 7, 9, 12,
15, 17, 19, and 21. In the 0-6-hour-old mice, a high of 31% of the administered dose of cerium
chloride was retained in the body 9 days following exposure. At 21 days post administration, the
amount of pooled cerium citrate and chloride retained was approximately 25%. The mice dosed
0-6 hours after birth retained more cerium in the GI tract throughout the 21-day observation
period than the mice dosed 6-24 hours after birth. The 0-6- and 6-24-hour-old rats exhibited
absorption of 9-10% of administered cerium 21 days after exposure. In the Yorkshire piglets,
the absorbed dose did not differ significantly over the 21-day observation period.
Two studies in adult rats also reported low absorption of soluble cerium following oral
exposures. Durbin et al. (1956) administered single [144Ce]-ceric nitrate (unreported
concentrations) intramuscular and intragastric doses to adult female Sprague-Dawley rats. The
rats dosed intramuscularly were sacrificed at post-administration days 1, 4, 64, and 256, while
the intragastric-dosed rats were sacrificed 4 days after administration. Less than 0.1% of the
intragastric-administered dose was absorbed from the GI tract.
Stineman et al. (1978) administered single intragastric [141Ce]-cerous chloride doses of
1000 mg/kg (lethal to 5% of the animals [LD5]) and 1163 (LD25) mg/kg to 6-8-week-old male
Swiss ICR mice and sacrificed them at 4 hours or at 1, 3, or 7 days later. No measurements were
made of whole-body radioactivity; however, 97-99% of radioactivity in the 12 sampled tissues
was found in the gut (stomach + duodenum).
In young (suckling) animals, soluble cerium appears to be retained in intestinal cells,
particularly in the ileum, possibly resulting in a much greater absorption than in adult animals
(Kostial et al., 1989a, b; Inaba and Lengemann, 1972). However, cerium retained in intestinal
cells has been demonstrated to be unavailable systemically (Inaba and Lengemann, 1972). The
high gut retention of cerium in young animals may be associated with high pinocytotic activity
of the newborn intestinal cells (Kostial et al., 1989b). Glucocorticoids, such as
methylprednisolone, stimulate production of endogenous corticosteroids, which cause precocious
gut closure (decreased pinocytosis) and maturation of the metal absorptive process (Kargacin
and Landeka, 1990). Administration of methylprednisolone in conjunction with artificial
feeding of [141Ce]-cerous chloride in cows' milk to 4-day-old suckling rats (strain not reported)
resulted in a nearly 40-fold reduction in the amount of cerium chloride detected in gut tissue
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(Kargacin and Landeka, 1990). This finding suggests that the pinocytotic activity of the
intestinal cells contributed to the differences. Similarly, injection with cortisone acetate resulted
in a more rapid loss of an oral dose to feces of young rats (Shiraishi and Ichikawa, 1972),
although whether this is due to lower uptake by intestinal cells or a more rapid release is not
presently clear.
3.1.2. Inhalation Exposure
Studies evaluating the deposition or absorption of cerium compounds following
inhalation exposure in humans are not available. However, cerium has been detected in the lung
tissue and alveolar macrophages of subjects believed to have been exposed to cerium
occupationally.
Case reports and a retrospective occupational investigation provide support for the
limited absorption of cerium deposited in the lung following inhalation exposure.
Transbronchial biopsies in a 60-year-old movie projectionist showed cerium concentrations of
11 ug/g wet weight after 12 years of exposure (Porru et al., 2001). McDonald et al. (1995)
demonstrated particulate material (diameter range from <1 um to 5-10 um) localized within lung
biopsy cells by using a scanning electron microscope. Analysis of bronchioalveolar lavage fluid
from a 58-year-old patient exposed to rare earth dusts and asbestos revealed cerium and
phosphorus in the alveolar macrophages (Pairon et al., 1995). The cerium particles accounted
for 70% of the particles observed in the lung tissue and were also identified in the interstitial
macrophages.
Microscopic examination of the tracheobronchial lymph nodes of a movie projectionist of
25 years revealed grey granules in large macrophages, which were characterized as calcium and
rare earth elements (cerium, lanthanum, and neodymium) by energy-dispersive analysis of
X-rays (Waring and Watling, 1990). Energy-dispersive X-ray also characterized dark particles
(diameter of 1 to 6 um) as cerium, unidentifiable by optical microscopy, in the bronchioalveolar
lavage of a 13-year photoengraver.
Lung tissues from a photoengraver exposed to smoke from cored carbon arc lamps for
46 years (Pietra et al., 1985) were found to have cerium concentrations 2,800-207,000 times
higher than those of urine, blood, or nails, suggesting that cerium particles in the lung are poorly
mobilized. The concentration of cerium in the lung and lymph nodes of a subject exposed to
cerium for 46 years as a photoengraver was 167 and 5 ug/g wet tissue weight, respectively, and
2,400- and 53-times higher, respectively, than the concentration in unexposed control subjects
(Vocaturo et al., 1983; Sabbioni et al., 1982).
Pairon et al. (1994) performed a retrospective evaluation of retention of cerium-
containing particles in the lungs of workers previously exposed to mineral dusts.
Bronchioalveolar lavage and lung tissue samples from mineral dust exposed workers and
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controls were examined for cerium content. In the seven cases that were judged to have high
cerium particle retention (as defined by having lavage fluid or tissue cerium concentrations at
least 5 times higher than those of controls), time since last exposure ranged from present to 29
years, with one patient's exposure time not available.
Limited animal data are available regarding total deposition of cerium aerosols within the
respiratory tract. Thomas et al. (1972) exposed 4-month-old, mixed sex Holtzman rats to two
concentrations (unreported) of aerosolized 1.4 um median aerodynamic diameter (GSD of 2.0)
[144Ce]-ceric hydroxide for 10 minutes. Whole-body radioactivity measurements were used to
identify a 28% deposition rate of inhaled cerium aerosol. Boecker and Cuddihy (1974) reported
an average deposition of 71% in beagles exposed to [144Ce]-cerous chloride for 4-10 minutes. In
both studies, cerium was deposited in the lungs with evidence of absorption from the lung
provided by the subsequent detection of cerium in the skeleton, liver, and kidneys.
3.2. DISTRIBUTION
3.2.1. Oral Exposure
Although cerium appears to be poorly absorbed from the GI tract, the bone and liver were
the organs with the highest cerium levels in rats following oral gavage of cerium chloride
(Shiraishi and Ichikawa, 1972). The concentration of cerium in the kidney, liver, lung, and
spleen of male ICR mice was significantly elevated relative to controls following 6 and 12 weeks
of oral exposure to 20 or 200 ppm cerium chloride (Kawagoe et al., 2005). The lung and spleen
contained the highest cerium concentrations in male ICR mice.
Manoubi et al. (1998) gave a single intragastric dose of stable cerium nitrate (20 mg/mL)
to Wistar rats. Three hours after dosing, cerium was found in the lysosomes of the duodenal
villosity but not in the liver or spleen. In 1-day-old Sprague-Dawley rats given a single
intragastric dose of [141Ce]-ceric nitrate of unreported concentration, Inaba and Lengermann
(1972) found cerium to be localized centrally, likely in the vacuoles, within epithelial cells of the
small intestine.
Cerium is capable of crossing the placenta and entering the fetal circulation in mice, but
the amounts found in the uterus and placenta were generally less than 5% of the maternal body
burden and decreased rapidly with increased time after exposure (Naharin et al., 1969). Fetal
body burdens in rodents were generally less than 1% of the initial maternal body burden after
either injected or oral administration (Levack et al., 2002; Inaba et al., 1992; Naharin et al.,
1969). Small amounts of injected cerium were also found in the maternal milk of mice (Naharin
et al., 1974), although at a small proportion (<0.01%) of the maternal body burden.
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3.2.2. Inhalation Exposure
As poorly soluble particles, cerium oxide particles behave like other airborne particles,
depositing within the respiratory tract based on aerodynamic character (Schulz et al., 2000).
Hamsters inhaling [144Ce]-cerium oxide aerosols with particle activity median diameters of 0.11
and 0.06 um exhibited lung burdens of 3.6% at 5 hours and 50% at 3 hours after exposure,
respectively, of initial body burden (Kanapilly and Luna, 1975).
Once deposited in the lung, insoluble cerium compounds may dissolve slowly, as
evidenced by the low percentage of cerium found in other tissues. In an investigation of the
toxicity of insoluble cerium, Hahn et al. (2001) exposed beagles to aerosolized [144Ce]-fused
aluminosilicate particles for 2-48 minutes and collected a variety of tissues (tissues studied not
reported) at death (up to 6,205 days following exposure). Cerium was dissolved from the lung
into the systemic circulation and observed in the liver, skeleton, and tracheobronchial lymph
nodes. Hahn et al. (2001) found between 1.0 and 10% of the initial lung burden of [144Ce]-fused
aluminosilicate particle aerosol in the liver and skeleton of beagles observed for 800 days
following 2-48-minute inhalation exposures. Hahn et al. (1999) also observed [144Ce]-fused
aluminosilicate particle translocation to the tracheobronchial lymph nodes following acute
inhalation of the particles. Lundgren et al. (1992) exposed adult F344/Crl rats to [144Ce]-cerium
oxide for 5-50 minutes or to bimonthly exposures of 25 minutes for 1 year; rats were sacrificed
at 1 hour and 3, 7, 14, 28, 56, 112, 224, 448, 560, and 672 days after exposure. The lungs, heart,
liver, spleen, kidney, and skeleton (remaining carcass) were measured for cerium. Cerium was
detected in the liver and skeleton in increasing percentages of body burden with respect to time,
while cerium was not detected in the spleen and kidneys.
More soluble forms of cerium (e.g., cerium citrate) may be systemically absorbed more
easily from the lung due to the increased solubility of the compound. Morgan et al. (1970)
exposed Swiss mice to aerosols of [144Ce] in the form of chloride, citrate, or fused clay, with
activity median diameters from 1.3 to 2.75 um, in unreported concentrations or durations. While
the initial body burden of all forms decreased rapidly during the first 2 weeks, likely due to
mucociliary elimination to the GI tract, the remaining lung burden for the relatively insoluble
fused clay remained higher than the chloride or citrate for the duration of the study (130 days).
Conversely, the liver burdens of the citrate and chloride forms remained higher than the fused
clay by about an order of magnitude. Further, as lung burdens of the chloride and citrate forms
decreased, the bone burdens of these forms increased. The bone burden for the fused clay form,
like the liver, was about an order of magnitude lower than the citrate or chloride forms.
Sturbaum et al. (1970) exposed 40 Chinese hamsters via the nose to [144Ce]-cerous
chloride aerosol, activity median diameter of 0.83 um and a GSD of 1.7, for 20 minutes and
sacrificed small groups (n = 4) at 2, 8, 16, 28, 64, 128, and 256 days after exposure. Whole-body
and tissue (types unreported) measurements of cerium radioactivity were made. The liver and
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skeleton exhibited between 1 and 10% of the initial body burden throughout the post-
administration measurements, while the lung portion of the initial body burden diminished from
approximately 20 at two hours to <1% by study's end.
Cerium has been observed to be localized in the cell, particularly in the lysosomes, where
it is concentrated and precipitated in an insoluble form in association with phosphorus. Wistar
rats were exposed to stable cerium chloride aerosol, mean diameter of 0.1 um, 5 hours/day for
either 5 days or 4 days/week for 4 weeks (Galle et al., 1992; Berry et al., 1989, 1988). Several
hours after exposure, cerium was deposited in the lysosomes of alveolar macrophages. The
cerium deposits appeared to be in the form of aggregates of fine granules or fine needles that
varied in length from 30 to 60 nm, with the longer needles resulting from the 4-week exposure.
Cerium was found in the lysosomal fraction of liver centrifugate collected from rats (strain
unreported) given an intravenous (i.v.) injection of 1.3 mg/kg [141Ce]-cerous chloride (Wiener-
Schmuck et al., 1990). Cerium was also found in the lysosomes of the duodenal villosity, but not
in the liver or spleen of Wistar rats following intragastric dosing. Cerium was also localized
centrally, likely in the vacuoles, within epithelial cells of the small intestine of Sprague-Dawley
rats.
3.3. METABOLISM
As an element, cerium is neither created nor destroyed within the body. The particular
cerium compound (e.g., cerium chloride, cerium oxide) may be altered as a result of various
chemical reactions within the body, particularly dissolution, but data have not demonstrated a
change in the oxidation state of the cerium cation. Exposure to cerium has been shown to
change hepatic levels of some cytochrome (CYP) P450 isozymes in a species- and strain-
sensitive manner for mice. Salonpaa et al. (1992) gave i.v. cerous chloride injections of 2 mg/kg
to adult DBA/2 and C57BL/6 mice and observed increases in expression of CYP2A4 and
CYP2A5 in the livers (2 and 3 days after dosing) and in the kidneys (6 hours and 1 day after
dosing) of D2 mice but not in B6 mice. Arvela et al. (1991) gave i.v. cerous chloride injections
of 0.5, 1, and 2 mg/kg to adult male DBA/2 and C57BL/6 mice and found a greater sensitivity to
increased CYP450 expression (isoform not reported) in DBA/2 and C57BL/6 mice 24 hours and
3 days after exposure, respectively. Conversely, Arvela and Karki (1971) observed a 50%
reduction, compared to controls, in CYP450 activity in adult Sprague-Dawley rats 3 days after a
single i.v. injection of 2 mg/kg cerous chloride. The effect of changes in CYP450 levels on the
toxicokinetics or toxicity of cerium, if any, is not known. In addition, the relatively high
intravenous bolus doses used in the available studies may not be relevant to oral or inhaled
exposure to cerium oxide.
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3.4. ELIMINATION
Following inhalation exposure, the initial rapid elimination of insoluble cerium from the
body is due primarily to transport up the respiratory tract by the mucociliary escalator and
eventual swallowing of the material, as with other poorly soluble particles (Boecker and
Cuddihy, 1974). Initial short-term clearance rates range from 35 to 95% of initial cerium body
burden, depending on the species tested and length of clearance time investigated. Lundgren et
al. (1992) exposed adult F344/Crl rats to [144Ce]-cerium oxide aerosol for 5-50 minutes, with
clearance of approximately 90% of the initial body burden by 7 days. Kanapilly and Luna
(1975) exposed hamsters to [144Ce]-cerium oxide aerosols with particle activity median
aerodynamic diameters of 0.11 and 0.06 um and observed decreases in initial body burden of 95
and 60%, respectively, 4 days after exposure. Differences in clearance rates may have been
dependent on particle size differences, with the smaller particles taking more time for
elimination; however, the authors also stated that the difference may have resulted from a leak in
the inhalation chamber used for the first dose group. Boecker and Cuddihy (1974) observed an
early clearance of initial body burden from 35-80% for individual dogs 4 days after exposure.
Thomas et al. (1972) exposed Holtzman rats to two concentrations (unreported) of aerosolized
[144Ce]-ceric hydroxide for 10 minutes and observed approximately 75-95% clearance of initial
body burden within 2 weeks of exposure (Thomas et al, 1972). Sturbaum et al. (1970) reported
clearance of 80% of initial cerium body burden by 7 days in Chinese hamsters exposed to
[144Ce]-cerous chloride aerosol for 20 minutes. After the initial clearance of cerium particles
from the upper respiratory tract, pulmonary clearance is slower, with reported slow-phase
clearance half-times ranging from 100 to 190 days in rodents (Lundgren et al., 1974; Thomas et
al., 1972; Morgan et al., 1970; Sturbaum et al., 1970). The slow-phase clearance was slightly
faster in beagles, with an estimated half-time of 63 days (Boecker and Cuddihy, 1974). Slow-
phase clearance from the lung is a combination of cerium dissolution and absorption (Morgan et
al., 1970) and mechanical clearance from the respiratory tract (Sturbaum et al., 1970).
Elimination of orally administered soluble cerium has been shown to be age dependent in
animals, with suckling animals absorbing cerium into the GI tissues (Inaba and Lengemann,
1972). This cerium may remain in the intestinal cells, may not be available systemically, and
may eventually be eliminated in the feces.
Although quantitative estimates of cerium elimination are rare, it appears that the primary
route of elimination for cerium, whether inhaled, ingested, or injected, is through the feces, with
small (generally <10%) amounts eliminated in the urine (Lustgarten et al., 1976; Durbin et al.,
1956). It has been suggested that the fecal excretion of systemically absorbed cerium is due to
elimination in the bile (Lustgarten et al., 1976), since hepatic clearance was due primarily to
biliary function.
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3.5. PHYSIOLOGICALLY BASED TOXICOKINETIC MODELS
No physiologically based toxicokinetic (PBTK) models for cerium oxide or other cerium
compounds were located in the evaluated literature.
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4. HAZARD IDENTIFICATION
4.1. STUDIES IN HUMANS—EPIDEMIOLOGY, CASE REPORTS, CLINICAL
CONTROLS
4.1.1. Oral Exposure
An association between exposure to rare earth elements, cerium in particular, in food and
the development of endomyocardial fibrosis has been suggested (Eapen, 1998; Kutty et al., 1996;
Valiathan et al., 1989). Cerium levels were elevated in the endomyocardial tissue samples from
patients who died of endomyocardial fibrosis compared with the tissues of controls who died of
accidents or congenital heart disease (Valiathan et al., 1989). Although causality has not been
conclusively demonstrated, a higher incidence of endomyocardial fibrosis has been reported
among a population consuming tubers grown in a region of India with high soil cerium
concentrations, compared to subjects consuming tubers grown in a soil with low cerium
concentrations. Analysis of the geographic distribution of endemic endomyocardial fibrosis in
India suggested a link to high cerium soil concentrations and possibly to magnesium deficiency
during childhood (Kutty et al., 1996).
A case-control study was conducted by Gomez-Aracena et al. (2006) to investigate the
role of chronic cerium exposure in coronary heart disease. Chronic cerium exposure was
represented by toenail cerium concentrations, and the occurrence of first myocardial infarction
was the characterization used for coronary heart disease. Odds ratios were calculated by
comparing the four exposure groups of 111, 142, 171, and 257 ug/kg toenail cerium
concentration to the control group. Gomez-Aracena et al. (2006) found an association between
increased toenail cerium concentrations and the risk of first myocardial infarction, when
controlling for confounding factors, such as smoking, body mass index history of hypertension,
diabetes, family history of coronary heart disease, p-carotene, lycopene, a-tocopherol, selenium,
mercury, and scandium. The odds ratio of first myocardial infarction in smokers with a toenail
cerium concentration of 257 ug/kg and without adjusting for additional risk factors of
myocardial infarction was 1.18 (95% confidence interval [CI]: 0.83-1.66), with ap value for
trend of 0.020. In nonsmokers, the odds ratio of first myocardial infarction with toenail cerium
concentrations of 169 and 227 ug/kg were 2.09 (95% CI: 1.05-4.16) and 2.81 (95% CI: 1.21-
6.52), respectively, with ap value for trend of 0.011, when controlling for the confounding
factors listed above. The results of Gomez-Aracena et al. (2006) suggest a relationship between
chronic cerium exposure and increased risk of acute myocardial infarction, with the strongest
association observed in nonsmokers.
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4.1.2. Inhalation Exposure
Reports have been published describing numerous cases of workers who developed
adverse lung effects, such as interstitial lung disease or pneumoconiosis, associated with
accumulation of cerium in the lungs after prolonged occupational exposure to cerium fumes or
dust (Yoon et al., 2005; Porru et al., 2001; McDonald et al., 1995; Pairon et al., 1995, 1994;
Sulotto et al., 1986; Vogt et al., 1986; Pietra et al., 1985; Vocaturo et al., 1983; Sabbioni et al.,
1982; Husain et al., 1980; Kappenberger and Buhlmann, 1975; Heuck and Hoschek, 1968).
The workers in the above reports had been exposed to cerium for periods of 10-46 years, with
exposures most commonly due to fumes from carbon arc lamps. These lamps, widely used in
the past in the fields of cinematography and photoengraving, have a central core consisting of
approximately 46% cerium oxide and smaller amounts of other rare earth oxides, including
lanthanum, neodymium, praseodymium, and samarium (Waring and Watling, 1990). As the
core burns, it emits oxide, and, to a lesser extent, fluoride dusts of cerium and the other rare
earth elements. Cases of cerium pneumoconiosis not associated with carbon arc lamps all
involved exposure to cerium oxide, either during processing or peripheral to its use as an
abrasive to grind and polish lenses (McDonald et al., 1995). Exposure concentrations of
cerium were not quantified in any of these studies.
Dendriform pulmonary ossification (DPO), a rare condition characterized by branching,
bony spicules often containing marrow, which are found in the lung parenchyma and are
associatied with pulmonary fibrosis, was observed in a 38-year-old man who worked as a
polisher in a crystal factory for 3 years (Yoon et al., 2005). Diffuse reticulonodular infiltrates in
the lung were observed in a chest radiograph, and a computer tomography (CT) scan showed
diffuse, tiny, circular, or beak-like densities with branching structures in the interlobular septum.
A CT scan with bone-setting showed branching, twig-like ossified masses in the right lower
lobe and a few dot-like ossifications in both lower lobes. The lung surface appeared irregular,
emphysematous, and mottled with anthracotic pigmentation during an open lung biopsy, with
several thorn-like hard materials in the lung parenchyma. Microscopic examination revealed
interstitial fibrosis, peripheral emphysema, multiple particles, and pneumonia. The particles
(0.1-0.3 um) were determined to be cerium oxide and phosphates of cerium and lanthanum by
energy dispersive X-ray analysis, with particles of quartz, feldspar, mica, kaolinite, halloysite,
talc, and TiC>2 infrequently detected. Yoon et al. (2005) characterized this case study as the first
to present a case of DPO associated with pneumoconiosis caused by the inhalation of rare earth
metals.
A 60-year-old male who worked as a movie projectionist and was exposed to rare earth
dusts for 12 years presented with diffuse interstitial lung fibrosis, emphysema, and severe
obstructive impairment (Porru et al., 2001). An increase in the lung concentration of rare earth
elements was evident in the subject with the highest concentration for cerium, compared with the
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five unexposed controls. Interstitial fibrosis accompanied by vascular thickening, reactive
alveolar macrophages, abundant macrophages in the air space, and moderate chronic interstitial
inflammation, along with small interstitial clumps of macrophages bearing scant deposits of
grayish-black pigment, was observed in a 68-year-old man who was employed as an optical lens
grinder for 35 years and smoked for 20 years (McDonald et al., 1995). Pairon et al. (1995)
identified particles containing cerium, lanthanum, and phosphorus in the alveolar macrophages
from a 58-year-old smoker with dyspnea who had been exposed to asbestos and rare earth dusts
as a crystal manufacturer, polisher, and movie projectionist from 1951-1967. Diffuse fibrosis of
interalveolar septa and perivascular hyalinized fibrosis was observed from the histologic analysis
(Pairon et al., 1995). Mild interstitial fibrosis, peribronchiolar fibrosis, and diffuse interstitial
fibrosis with emphysema were seen in a photoengraver or glass polisher, nonsmoking foundry
worker, and a movie projectionist and glass polisher, respectively (Pairon et al., 1994).
Deposits of carbon arc lamp fumes were evident in the marcrophages in the
tracheobronchial lymph nodes of a 66-year-old smoking movie projectionist of 25 years,
although pneumoconiosis was not considered because respiratory symptoms and radiographic
and histologic changes were not apparent (Waring and Watling, 1990). Dark particles, identified
as cerium by energy dispersive X-ray analysis, were observed in lung tissue from a 48-year-old
smoker employed as a photoengraver for 13 years (Sulotto et al., 1986). A chest X-ray showed a
micronodular pattern extending to all lung fields although lung examinations were normal and
the patient did not experience respiratory impairment. Vogt et al. (1986) observed five
reproduction photographers, exposed to carbon arc fumes for more than a decade, with slowly
progressive respiratory function restriction, as well as interstitial lung fibrosis and the
accumulation of fine, granular dusts, characterized as rare earth minerals and primarily cerium,
in the lung tissue.
Pulmonary hypertension with increased vascular resistance was observed in a 58-year-old
man who worked in the photoengraving industry for 46 years (Vocaturo et al., 1983; Sabbioni et
al., 1982). Rare earth elements, primarily cerium, were observed by neutron activation analysis
in the lung and lymph node biopsies, and the concentrations of the elements in these tissues were
greater than those in control subjects (Pietra et al., 1985).
Profuse discrete nodular shadowing was present in the chest X-ray of a 34-year-old
employee at a glass rubbing polish plant, although pulmonary function appeared normal (Husain
et al., 1980). The subject refused histologic examination, and an analysis of the occupational
dust concentrations revealed high levels of cerium oxide (50%) and other rare earth oxides.
A 65-year-old man working in the photographic department of a printing plant
demonstrated inactive right apical infiltrates in a chest roentgenogram in 1948; in 1951 diffuse
spotty infiltrates were noted; in 1953 and 1959 the infiltrates were more pronounced in the
middle and lower fields; and in 1965 slight fibrosis of the surrounding tissue was evident, along
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with a perifocal emphysema (Heuck and Hoschek, 1968). Heuck and Hoschek (1968) also
documented fibrosis and small infiltrates in the lung of a 53-year-old male exposed to carbon arc
lamp smoke in printing industries and infiltrates in a 67-year-old man, with chemotherapy-
treated tuberculosis in both upper lobes of the lung, who worked with carbon arc lamps for 26
years.
Collectively, the available studies show that the defining characteristic of cerium
pneumoconiosis is accumulation of cerium particles, as well as other rare earth particles, in the
lungs and lymphoreticular system. In most cases, the initial indication of disease was the
presence of diffuse interstitial or reticulonodular opacities in chest X-rays. Pulmonary function
in the affected workers varied from normal to severe restrictive impairment. In several cases,
thorium, a common impurity in rare earth minerals, and the naturally occurring radioisotopes of
the rareearth elements were quantified, but, in each case, they were found to be present in
quantities too small to produce any effect due to radiation. Exposure to silica, which is
fibrogenic, may have contributed to the effects observed in the cerium oxide and glass workers
but is not a factor for the workers exposed to carbon arc lamp dust, which includes most of the
workers found to have fibrosis. Two reviews of the available case studies concluded that there is
convincing evidence that accumulation of cerium and other rare earth metals in the lungs is
causally related to the development of pulmonary interstitial fibrosis in workers (McDonald et
al., 1995; Waring and Watling, 1990). The human data were inadequate to identify potentially
sensitive subgroups.
4.2. SUBCHRONIC AND CHRONIC STUDIES AND CANCER BIOASSAYS IN
ANIMALS—ORAL AND INHALATION
4.2.1. Oral Exposure
Kawagoe et al. (2005) studied the possible association between cerium exposure and
oxidative stress in the mouse liver. Groups of male ICR mice (four per dose group) were
administered a diet containing 0, 20, or 200 ppm cerium chloride (CeCls) for 6 or 12 weeks.
This corresponds to doses of approximately 2.6 or 26 mg Ce/kg-day, assuming a reference food
consumption of 0.005 kg/day and reference body weight of 0.022 kg (U.S. EPA, 1988).
Evaluations conducted at termination included cerium levels in organs (liver, spleen, kidney, and
lung), levels of glutathione (GSH) and metallothionein (MT) in liver, kidney, and lung, levels of
lipoperoxide in liver and plasma, superoxide dismutase (SOD) activity in blood and liver, as well
as cholesterol levels, triglyceride levels, and aspartate aminotransferase (AST) and alanine
aminotransferase (ALT) activity in serum. Treatment with cerium did not affect food
consumption or body weight but statistically significantly decreased lipoperoxide levels in
hepatic tissues (33% at 20 and 38% at 200 ppm, 6 weeks; 29% at 20 ppm, 12 weeks), increased
liver GSH levels (200 ppm, 12 weeks) and liver MT activity (20 ppm, 12 weeks; 200 ppm,
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6 weeks), and decreased plasma SOD activity (20 and 200 ppm, 6 weeks). Cerium in the kidney,
liver, lung, and spleen was statistically significantly elevated relative to controls in the organs of
mice in the 200 ppm group at 6 and 12 weeks, with the lung and spleen containing higher cerium
concentrations. Pathological alterations were not detected in the kidney, liver, lung, or spleen by
microscopic observation. According to Kawagoe et al. (2005), the increases in hepatic GSH and
MT activity represent a response to cerium-induced oxidative stress. It is unknown whether the
decrease in hepatic lipoperoxide is a consequence of the increase in GSH and MT. The study
authors suggest that the endpoints showing changes as a result of cerium exposure in this study
are indicators of reactive oxygen species generation in the liver.
A study (Cheng et al., 2000) in male Wistar rats (six per dose group) dosed orally (further
details not reported) with 0, 0.2, 2.0, or 20 mg cerium chloride/kg-day (0.1, 1.1, or 11.4 mg
Ce/kg-day) for up to 105 days investigated the effects of CeCb on the structure and oxygen
affinity of hemoglobin in vivo. The highest dose, 20 mg/kg-day, produced a slight increase of
hemoglobin content in the erythrocytes after 40 days of treatment, with an even greater increase
in hemoglobin content after 80 days. The effect on the oxygen affinity of the hemoglobin was
demonstrated by oxygen saturation curves for the dosed rats and control rats. Hemoglobin in
cerium-treated rats also exhibited altered oxygen affinity up to 80 days of exposure,
demonstrated by increased affinity up to 10 mm Hg and a double sigmoidal curve for rats treated
for 40 days and increased affinity above 20 mm Hg for rats treated for 80 days. A significant
change was not observed at 0.2 mg/kg-day for 105 days of exposure. These results suggest that
the oxygen affinity of hemoglobin increases following long-term oral exposure to CeCls. When
the 80-day cerium feeding period was followed by 15 days of cerium-free exposure, the oxygen
affinity did not recover to normal levels. The investigators attributed the altered oxygen affinity
up to 80 days of exposure to conformational changes of hemoglobin, hydrolysis of hemoglobin-
bound diphosphoglyceric acid, and the partial oxidation of heme-Fe(II) to heme-Fe(III).
Karma et al. (1998) examined the effects of cerium chloride, administered in the drinking
water with other rare earth chlorides, on the heart of New Zealand white rabbits (10/group) fed
diets with normal or restricted magnesium for 6 months. The rabbits were randomly distributed
into 4 groups with a male to female ratio of 1:1 within each group. Group 1 was exposed to a
magnesium-sufficient diet only, group 2 was exposed to a magnesium-sufficient diet and cerium,
group 3 was exposed to a magnesium-restricted diet, and group 4 was exposed to a magnesium-
restricted diet and cerium. The drinking water for the rare earth chloride-exposed rabbits was
adulterated with 1 g/L of rare earth chloride of which 56.6% was cerium (lanthanum 11.5%;
praseodymium and neodymium 14.5%; samarium 2.6%). Histologic evaluation of the heart at
the end of the study showed no cardiac lesions in the groups fed the normal magnesium diet,
regardless of whether they consumed water with or without cerium. Cardiac lesions were
evident in 6/10 rabbits from group 3, the magnesium-restricted diet, and in 9/10 rabbits from
21
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group 4, the magnesium-restricted diet and cerium chloride-exposed group. Rabbits fed the
magnesium-restricted diets, treated with or without cerium, showed endocardial, subendocardial,
interstitial, and perivascular fibrosis. The lesions were more severe in those with cerium added
to the drinking water (group 4). The results suggest that cerium may intensify the effect of
magnesium deficiency on heart tissue. A no-observed-adverse-effect level (NOAEL) or lowest-
observed-adverse-effect level (LOAEL) for cerium cannot be established for cerium chloride in
this study because the cardiac lesions were observed in rabbits that were fed a magnesium-
restricted diet.
The only chronic oral exposure study of stable cerium was reported by Kumar et al.
(1996), who exposed mixed-sex (male to female ratio of 1:1) groups (n = 5-9) of Sprague-
Dawley rats to cerium chloride and a magnesium-sufficient or magnesium-deficient diet for
13 months. The rats were randomly distributed into four groups, with groups 1 and 2 fed a
magnesium-sufficient diet and groups 3 and 4 fed a magnesium-deficient diet. The rats in group
1 and 3 were exposed to 0 ppm cerium chloride, while the rats in groups 2 and 4 were exposed to
35 ppm cerium chloride in the drinking water. This cerium chloride exposure corresponds to a
concentration of 19 mg/L, which in turn, assuming a reference water consumption of 0.046 and
0.038 L/day and reference body weight of 0.523 and 0.338 kg (U.S. EPA, 1988) in males and
females, respectively, corresponds to an average daily dose in males of 1.7 mg/kg-day and in
females of 2.1 mg/kg-day. At 13 months, the animals were sacrificed and cardiac tissue was
collected for elemental analysis and histology. No statistically significant changes in serum or
cardiac levels of magnesium or calcium were reported. Cerium levels in cardiac tissue were
statistically significantly elevated in group 4. Cerium-treated animals had a significantly greater
level of collagen in cardiac tissue, relative to group 1, with an enhanced effect in animals fed a
magnesium-deficient diet (groups 3 and 4). No other endpoints were evaluated. This study was
not used to identify a NOAEL or LOAEL due to the limited number of evaluated endpoints and
the investigation of a single dose group.
4.2.2. Inhalation Exposure
No chronic inhalation studies on cerium toxicity are available. However, the National
Toxicology Program (NTP) is considering an evaluation of the chronic inhalation toxicity of
cerium oxide.
A subchronic inhalation study using cerium oxide (CeO2; eerie oxide) was conducted in
7-week-old Sprague-Dawley rats (BRL, 1994). Cerium oxide is the form of cerium typically
encountered in industrial exposures (Reinhardt and Winkler, 1986). This study is an
unpublished study; accordingly, it was externally peer reviewed by EPA in August 2006
(external peer review report available at www.epa.gov/iris).
Groups of 15 male and 15 female Sprague-Dawley CD rats were given nose-only
22
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exposure to a dry powder aerosol (mass median aerodynamic diameter [MMAD] = 1.8-2.2 um,
geometric standard deviation [GSD] = 1.8-1.9) of cerium oxide at concentrations of 0, 0.005,
0.0505, or 0.5075 mg/L (0, 5, 50.5, or 507.5 mg/m3) 6 hours/day, 5 days/week for 13 weeks.
The cerium oxide test material was 99% rare earth oxide with a maximum of 75 ppm Fe2C>3. Of
the 99% rare earth oxide, 99.95% was cerium oxide with a maximum of 25 ppm of both PreOn
and Nd2Os. Praseodymium and neodymium are also rare earth metals.
A functional observational battery was performed on all rats, as well as activity level
testing, hematology, clinical biochemistry, urinalysis, ophthalmological examination, and a gross
pathological examination of selected tissues weighed and retained for histopathologic
examination. No deaths or clinical signs related to cerium oxide were noted. Food consumption
and body weight gain were marginally, but statistically not significantly reduced in males at
507.5 mg/m3 and were considered the result of cerium oxide exposure.
A functional observational battery detected a statistically significant (p < 0.05) 17%
decrease in forelimb grip strength at week 13 in females exposed to 507.5 mg/m3. No other
changes were found in the functional observational battery. Motor activity, measured by
photocells in a figure-8 enclosure, was unaffected by cerium oxide exposure. The
ophthalmology examination was normal.
Hematological analysis revealed a statistically significant (p < 0.05) increase in absolute
neutrophil counts of 105% in 6-week males at 507.5 mg/m3, 130% in 6-week females at
50.5 mg/m3, 85% in 13-week males at 50.5 mg/m3, 75% in 13-week males at 507.5 mg/m3,
210% in 13-week females at 5 mg/m3, 177% in 13-week females at 50.5 mg/m3, and 233% in 13-
week females at 507.5 mg/m3. Differential white blood cell counts were largely restricted to
altered neutrophil counts (as shown in Table 4-1), with the exception being increased absolute
lymphocyte and eosinophil counts in 13-week males at 50.5 mg/m3 by 36% and 187%,
respectively. Differential white blood cell counts revealed changes in relative percentages of
neutrophils and lymphocytes. The relative percentage of neutrophils and lymphocytes were
significantly increased by 102 and 80% in 6-week and 13-week males, respectively, at
507.5 mg/m3. In 13-week males, there was a corresponding 19% decrease in the relative
percentage of lymphocytes. The 118% relative increase in neutrophils was accompanied by a
12% decrease in lymphocytes in 6-week females at 50.5 mg/m3. In 13-week females, the 5, 50.5,
and 507.5 mg/m3 doses were associated with, respectively, 130, 130, and 164% relative increase
in the percentage of neutrophils and 12, 10, and 14% decreases in lymphocytes. Clinical
chemistry and urinalysis were normal.
23
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Table 4-1. Hematological changes in male and female Sprague-Dawley rats
following inhalation of cerium oxide aerosol 6 hours/day, 5 days/week for
13 weeks
Dose
(mg/m3)
Absolute
neutrophils
Mean
SD
Absolute
lymphocytes
Mean
SD
Absolute
eosinophils
Mean
SD
Relative
neutrophils
Mean
SD
Relative
lymphocytes
Mean
SD
Males, week 6
0
5
50.5
507.5
1,509
1,440
2,560
3,088a
1,697
720
882
1,162
10,057
9,891
11,478
10,296
2,455
2,602
1,944
3,670
122
102
83
110
299
136
125
175
11.3
12.5
17.7
22.8a
7.5
5.9
5.6
6.8
85.4
84.6
79
73.7a
8.3
6.5
5.7
6.9
Females, week 6
0
5
50.5
507.5
724
1,230
l,680b
1,328
494
765
651
562
8,791
7,537
8,083
8,916
2,355
2,068
1,737
3,678
75
67
92
87
119
57
88
104
7.8
13.0
17a
13.3
5.7
6.3
6.4
4.3
90.8
84.5
80. lb
84.9
6.0
6.7
7.5
4.5
Males, week 13
0
5
50.5
507.5
1,541
1,638
2,844a
2,698a
909
622
1,038
1,101
6,243
6,254
8,481b
5,238
1,575
1,521
3,009
1,910
62
85
175b
73
94
83
173
98
18.4
20.1
25.4
33.2a
6.4
6.9
10.9
9.5
77.5
76.1
70.7
62.7a
8.7
7.9
11.0
11.0
Females, week 13
0
5
50.5
507.5
325
1,006C
899C
1,081C
162
797
414
399
3,624
3,794
3,802
3,722
1,365
1,200
1,228
1,137
47
61
36
32
58
55
44
42
8.3
19.1a
19.1a
21. 9a
4.2
10.0
7.8
6.3
85.7
75.3a
76.9b
73.9a
5.5
10.7
8.5
7.1
""Statistically significantly different from control (Dunnett's test; p < 0.01).
bStatistically significantly different from control (Dunnett's test; p < 0.05).
Statistically significantly different from control (Dunn's test; p < 0.01).
Source: BRL (1994).
At necropsy, there were treatment-related increases in the weight of the lungs and spleen
that correlated with gross and microscopic findings. Absolute and relative lung weights were
statistically significantly (p < 0.001) increased in both males and females at 50.5 and
507.5 mg/m3 (Tables 4-2 and 4-3). Lung weights, relative to brain weights, were also
statistically significantly increased in male and female rats at 50.5 and 507.5 mg/m3. Relative
spleen weight was statistically significantly (p < 0.05) increased in males at 507.5 mg/m3 (Table
4-4). A statistically significant increase in absolute (29%) and relative (28%) thymus weight in
male mice at 50.5 mg/m3 was not considered by the study authors to be related to cerium oxide
treatment. The increase in absolute and relative thymus weight was observed in only the mid-
dose male rats and a dose-response relationship was not observed in male or female rats. Thus,
the increase in thymus weight in male rats was determined to not be a biologically significant
effect of cerium oxide exposure.
24
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Table 4-2. Absolute lung weight in rats exposed to cerium oxide aerosol
6 hours/day, 5 days/week for 13 weeks
Dose
(mg/m3)
0
5
50.5
507.5
Males
Mean weight (g)
1.611
1.760
2.574a
4.662a
SD
0.1088
0.1398
0.3499
0.5161
%
Change
-
9
60
189
Females
Mean weight (g)
1.145
1.311
1.6513
3.1733
SD
0.0871
0.1490
0.2900
0.3235
%
Change
-
14
44
177
""Statistically significantly different from control (Dunn's test; p < 0.01).
Source: BRL (1994).
Table 4-3. Relative lung weight in rats exposed to cerium oxide aerosol
6 hours/day, 5 days/week for 13 weeks
Dose
(mg/m3)
0
5
50.5
507.5
Males
Mean weight (g%)
0.334
0.371
0.5283
1.0243
SD
0.0256
0.0270
0.0775
0.1621
%
Change
—
11
58
207
Females
Mean weight (g%)
0.477
0.544
0.6973
1.3583
SD
0.0443
0.0557
0.1124
0.1432
%
Change
—
14
46
185
"Statistically significantly different from control (Dunn's test; p < 0.01).
Source: BRL (1994).
Table 4-4. Relative spleen weight in rats exposed to cerium oxide aerosol
6 hours/day, 5 days/week for 13 weeks
Dose
(mg/m3)
0
5
50.5
507.5
Males
Mean weight (g%)
0.162
0.169
0.178
0.1883
SD
0.0190
0.0286
0.0266
0.0298
%
Change
.
4
10
16
Females
Mean weight (g%)
0.216
0.242
0.226
0.222
SD
0.0281
0.0259
0.0445
0.0252
%
Change
.
12
5
3
""Statistically significantly different from control (Dunnett's test; p < 0.05)
Source: BRL (1994).
Gross examination found discoloration or pale areas, pale foci, and uncollapsed
parenchyma in the lungs of male and female rats (Table 4-5). Pale areas and discoloration in the
lung were evident at 50.5 and 507.5 mg/m3 in male and female rats, respectively, with
uncollapsed parenchyma evident at 50.5 and 507.5 mg/m3 in male rats and 507.5 mg/m3 in
female rats. Pale foci in the lungs were only seen in female rats exposed to 5 mg/m3. The
incidence of enlargement or pale discoloration of the mandibular, bronchial, mediastinal, and
pancreatic lymph nodes is shown in Table 4-6. Enlargement or pale discoloration of both lymph
nodes that drain the lungs (the bronchial and mediastinal lymph nodes) was evident in both
25
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males and females at >5 mg/m3 cerium oxide. The mandibular and pancreatic lymph nodes did
not display a dose-response trend of enlargement or discoloration. The study authors (BRL,
1994) judged the mandibular lymph node enlargement, which was observed in control and
cerium-exposed rats, not to be an effect of the cerium oxide exposure, while the pancreatic,
bronchial, and mediastinal lymph node enlargement and/or discoloration were considered to be
related to the cerium oxide treatment.
Table 4-5. Results of gross pathological examination of lungs of rats
exposed to cerium oxide 6 hours/day, 5 days/week for 13 weeks
Lung
Pale foci
Pale areas or discoloration
Uncollapsed parenchyma
Dose (mg/m3)
Male
0
0/15
0/15
0/15
5
0/15
0/15
0/15
50.5
0/15
15/15
2/15
507.5
0/15
15/15
15/15
Female
0
0/15
0/15
0/15
5
4/15
0/15
0/15
50.5
0/15
15/15
0/15
507.5
0/15
15/15
15/15
Source: BRL (1994).
Table 4-6. Results of gross pathological examination of bronchial,
mediastinal, and pancreatic lymph nodes of rats exposed to cerium oxide
6 hours/day, 5 days/week for 13 weeks
Lymph nodes
Mandibular
enlargement
discoloration
Bronchial
enlargement
discoloration
Mediastinal
enlargement
discoloration
Pancreatic
enlargement
discoloration
Dose (mg/m3)
Male
0
5
50.5
507.5
4/15
-
2/15
-
5/15
-
4/15
-
0/15
0/15
4/15
13/15
15/15
15/15
15/15
15/15
0/15
0/15
2/15
2/15
9/15
9/15
9/15
10/15
1/15
-
0/15
-
0/15
-
0/15
-
Female
0
5
50.5
507.5
2/15
-
4/15
-
3/15
-
1/15
-
0/15
0/15
1/15
15/15
14/15
15/15
15/15
15/15
0/15
1/15
1/15
10/15
8/15
9/15
8/15
10/15
2/15
0/15
0/15
0/15
1/15
0/15
0/15
1/15
Source: BRL (1994).
Histologic examination revealed dose-related alveolar epithelial and lymphoid
hyperplasia and pigment accumulation in the lungs, lymph nodes, and larynx of male and female
rats at >5 mg/m3. The incidence data are presented in Table 4-7. The pigment, which was also
found in other parts of the respiratory tract, including the nasal cavities and the trachea, and the
liver and spleen, was considered by the study authors to be the test compound or a product
thereof. The lymphoid hyperplasia in the lymph nodes following cerium oxide exposure was
characterized by the study pathologist as an increase in the number of lymphocytes, with lymph
26
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node paracortices and cortices expansion. The authors reported that the severity of the
hyperplasia in a given tissue, lung or lymph node, was correlated with the amount of pigment
accumulated in the tissue but did not present any supporting data. BRL (1994) considered these
findings to be consistent with antigenic stimulation by cerium oxide; however, they did not
discuss the possibility of non-antigenic stimulation. The metaplasia evident in the larynx was
interpreted by the study pathologist as adaptive and reversible. Lesions were not observed in the
testes or ovaries of the high-dose group.
The NOAEL and LOAEL values for the toxicological effects observed are included in
Table 4-8. This study identified a LOAEL of 5 mg/m3 in rats, based on the increased incidence
of lymphoid hyperplasia in the bronchial lymph nodes of male and female rats. A NOAEL was
not identified.
27
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Table 4-7. Incidences of histopathologic effects in rats exposed to cerium
oxide aerosol 6 hours/day, 5 days/week for 13 weeks
Exposure (mg/m3)
Control
5
50.5
507.5
Males
Larynx
metaplasia
pigment accumulation
Lung
lymphoid hyperplasia
alveolar epithelial hyperplasia
pigment accumulation
Bronchial lymph node
lymphoid hyperplasia
pigment accumulation
Mediastinal lymph node
lymphoid hyperplasia
pigment accumulation
Mandibular lymph node
lymphoid hyperplasia
pigment accumulation
Pancreatic lymph node
lymphoid hyperplasia
pigment accumulation
Spleen
pigment accumulation
0/15
0/15
0/15
0/15
0/15
0/15
0/15
0/0
0/0
0/15
0/15
-
0/15
3/15
6/15a
0/15
1/15
15/153
11/133
13/133
2/2
2/2
0/3
0/3
-
0/15
9/15a
9/15a
0/15
11/153
15/153
15/153
15/153
9/10
8/10
0/5
0/5
-
0/15
13/153
12/153
12/153
14/153
15/153
15/153
15/153
9/9
9/9
2/15
6/15a
-
6/15a
Females
Larynx
metaplasia
pigment accumulation
Lung
lymphoid hyperplasia
alveolar epithelial hyperplasia
pigment accumulation
Bronchial lymph node
lymphoid hyperplasia
pigment accumulation
Mediastinal lymph node
lymphoid hyperplasia
pigment accumulation
Mandibular lymph node
lymphoid hyperplasia
pigment accumulation
Pancreatic lymph node
lymphoid hyperplasia
pigment accumulation
Spleen
pigment accumulation
0/15
0/15
0/15
0/15
0/15
0/15
0/15
0/1
0/1
0/15
0/15
0/2
0/2
0/15
3/15
0/15
0/15
0/15
15/153
13/153
14/153
10/10
10/10
0/5
0/5
0/0
0/0
0/0
6/15a
7/15a
1/15
5/15a
15/153
15/153
15/153
9/9
9/9
0/3
0/3
1/1
1/1
0/0
9/15a
9/15a
7/15a
15/15a
15/15a
15/15a
15/15a
9/10
9/10
0/15
6/15a
0/1
1/1
3/15
"Significantly different from vehicle control group (p < 0.05 by Fisher's exact test)
Source: BRL, 1994.
28
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Table 4-8. NOAEL and LOAEL values for effects observed in BRL (1994)
Toxicological effect
Sex
NOAEL
(mg/m3)
LOAEL
(mg/m3)
Organ weight changes
Absolute lung weight
Relative lung weight
Absolute spleen weight
Relative spleen weight
M, F
M, F
M, F
M
F
5
5
507.5
50.5
507.5
50.5
50.5
-
507.5
-
Gross pathological lesions in lung
Pale areas
Discoloration
Uncollapsed parenchyma
M, F
M, F
M
F
5
50.5
5
50.5
50.5
507.5
50.5
507.5
Gross pathological lesions in the lymph nodes
Mandibular enlargement
Bronchial enlargement
Bronchial discoloration
Mediastinal enlargement
Mediastinal discoloration
M, F
M, F
M, F
M, F
M, F
-
-
-
-
-
5
5
5
5
5
Histopathologic lesions
Metaplasia, larynx
Pigment accumulation, larynx
Lymphoid hyperplasia, lung
Alveolar epithelial hyperplasia, lung
Pigment accumulation, lung
Lymphoid hyperplasia, bronchial lymph node
Pigment accumulation, bronchial lymph node
Pigment accumulation, spleen
Pigment accumulation, mandibular lymph node
Lymphoid hyperplasia, mediastinal lymph node
Pigment accumulation, mediastinal lymph node
M, F
M
F
M
F
M, F
M, F
M, F
M, F
M
M
M, F
M, F
-
-
5
50.5
5
5
-
-
-
50.5
50.5
-
5
5
50.5
507.5
50.5
50.5
5
5
5
507.5
507.5
5
5
Intratracheal instillation of 50 mg of a dust suspension of cerium oxide produced mild
changes in the lung and spleen of white rats examined 8 months later (Mogilevskaya and
Raikhlin, 1967). Conglomerates of dust particles deposited intracellularly and small particles
were observed in the cytoplasm of alveolar macrophages. There was a moderate proliferative
response in the lungs and bronchi, with occasional increased lymphocyte numbers and slight
development of connective tissue and collagen fibers, but diffuse or nodular fibrotic changes
were lacking. In the spleen, macrophages and large multinucleated cells accumulated. No
histologic changes were found in any of the other major organs.
Lundgren et al. (1996) conducted an investigation into the pulmonary carcinogenicity of
beta-particle radiation from inhaled 144CeO2 in F344/N rats, with stable cerium oxide serving as
29
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the control group (n = 1,049). The radiation doses ranged from 3.6 to 37 Gy. The control rats
received stable cerium oxide in a single inhalation dose of comparable mass concentration to the
dose groups, although the concentration was not specified by the study authors. The control rats
were held for life-span observation and were evaluated histologically. The histologic evaluation
showed nonneoplastic lesions, such as inflammation (5.1%), fibrosis (5.6%), alveolar-epithelial
hyperplasia (4.5%), and alveolar macrophage hyperplasia (7.1%), in the lungs of control rats.
Seven primary lung neoplasms were also apparent in the control rats and included four alveolar
or papillary adenomas; one alveolar, papillary, or tubular adenocarcinoma; one squamous cell
carcinoma; and one fibro- or osteosarcoma. Control rats, unexposed to cerium oxide, in a
separate study of inhaled 144CeC>2 in F344/N rats demonstrated an incidence of lung tumors of
6/110 (Lundgren et al., 1992). The observed tumors included two papillary adenomas, two
papillary adenocarcinomas, one adenosquamous carcinoma, and one mesothelioma.
4.3. REPRODUCTIVE/DEVELOPMENTAL STUDIES—ORAL AND INHALATION
Kawagoe et al. (2008) administered 200 mg/kg cerium chloride by gavage to pregnant
ICR mice on day 11.5 of gestation and the embryos were examined on gestation day 14.5.
Histopathology was performed on the liver, spleen, and lungs of the embryos. Pulmonary and
hepatic vascular congestion were observed in the fetal mice. However, evidence of altered tissue
morphology was not observed. In addition, Kawagoe et al. (2008) administered a single dose of
200 mg/kg cerium chloride by gavage to dams immediately after parturition to observe the
effects of cerium chloride on neonatal mice through lactation. Neonatal mice were smaller
compared to controls through lactation. There were no differences in survival of neonatal mice
and controls. As early as 1 day post-administration, pulmonary hemorrhage in the peripheral
alveoli, vascular congestion, and neutrophil infiltration were observed in the lungs of adult mice.
In neonatal mice exposed lactationally, pulmonary hemorrhage, hepatic vascular congestion,
and trachea hemorrhage were also observed.
4.4. OTHER DURATION- OR ENDPOINT-SPECIFIC STUDIES
As indicated in Section 3.1.1, cerium compounds are poorly absorbed from the GI tract of
animals; thus, parenteral administration of cerium compounds has been the preferred route of
exposure to study the systemic effects of these compounds.
4.4.1. Acute Toxicity Studies (Oral and Inhalation)
Single dose and/or acute oral data are limited but include a report of splenic lesions,
including hypertrophy, reticuloendothelial hyperplasia, and hyperactive lymphoid follicles, and
GI irritation, characterized by gastritis and enteritis with focal hemorrhage and necrosis of the
mucosa in the stomach and duodenum, in mice given single gavage doses of 1,000 or
30
-------
1,163 mg/kg (1:3 cerium chloride-sodium citrate complex) (Stineman et al., 1978). Open field
behavior was not affected and the 7-day LD50 was 1,291 mg/kg cerium (95% CI: 1,198-1,440).
Bruce et al. (1963) identified an oral LD50 of 4,200 mg/kg for cerium nitrate in female Sprague-
Dawley rats (n=30).
There is a report of piloerection, hunched posture, unsteady gait, and dark coloring of
eyes in male and female rats, along with lethargy, abnormal respiration, and prostration in
females, given single gavage doses of 5,000 mg/kg cerium as cerium sulfide (Rhodia Inc., 1998).
Ji and Cui (1988) list an oral LD50 in female mice of 1,178 mg/kg (95% CI: 1,043-1,331) for
cerium nitrate (Ce[NO3]3) and 622 mg/kg (95% CI: 550-702) for cerium oxides.
Cerium chloride was administered by gavage to male and female adult ICR mice as a
single dose of 200 or 500 mg/kg (Kawagoe et al., 2008). The mice were sacrificed and
necropsied 1, 3, or 7 days post-administration. Mortality in the mice that received 500 mg/kg
cerium chloride was 60% within 48 hours of gavage, while the mice in the 200 mg/kg group
survived at least 8 weeks after dosing. The mice receiving cerium chloride demonstrated
pulmonary hemorrhage, vascular congestion, and thickened alveolar septae in the lungs; focal
hepatocellular necrosis and neutrophil infiltrations in the midzone of periportal areas in the liver;
congestion of hepatic central and portal veins and sinusoids; kidney vascular congestion,
particularly of the renal tubules; and pancreatic vascular congestion (Kawagoe et al., 2008).
Data on the toxicity of single exposures to airborne cerium in animals are limited. A
review by the HEI (2001) reported that the LCso for cerium oxide in rats was >50 mg/m3 in a
study by Rhone-Poulenc (1983), but the study was unavailable for examination and no further
details were reported.
4.4.2. Acute Studies (Injection)
4.4.2.1. Neurobehavioral and Neurodevelopmental Effects
D'Agostino et al. (1982, 1978a, b) studied the neurodevelopmental effects of cerium
following subcutaneous injection in mice. The three separate publications present the same
experimental design. Pregnant mice were administered a single dose of 80 mg/kg cerium
(sodium/citrate complex) or citrate (control) on gestation day (GD) 7 or 12 or postnatal day
(PND) 2. In order to differentiate the gestational effects from lactational effects or changes in
maternal behavior, a full cross-foster study design was employed. A cross-fostering design
distributed the offspring (3 males and 3 females per dam) of mothers receiving cerium or citrate
during gestation to lactating dams exposed to either cerium or citrate. Body weight and gross
activity of the neonates were assessed on PNDs 7 or 12 (D'Agostino et al., 1978a, b) or PNDs 8
or 13 (D'Agostino et al., 1982), whereas open-field behavior, accelerating rotarod performance,
and passive avoidance learning were assessed on PNDs 60-65. Maternal offspring retrieval
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latency was measured on PND 3. The growth and behavioral data were analyzed via a
multivariate analysis.
Pups exposed to cerium in utero on GDs 7 and 12 demonstrated a statistically significant
decrease in body weight on PNDs 7 and 12 (D'Agostino et al., 1978a, b), whereas the body
weight of offspring of cerium-exposed dams on GD 7 did not differ significantly from that of
control offspring (D'Agostino et al., 1982). Offspring of dams exposed to cerium on GD 12
showed significantly decreased body weights on PNDs 8 and 13 (D'Agostino et al., 1982). Pups
exposed to cerium in utero were retrieved by dams and replaced in the nest significantly faster
than controls. When the dams were injected on PND 2, neonatal weights on PNDs 7 and 12 and
8 and 13 were statistically significantly reduced (D'Agostino et al., 1982, 1978a, b). The study
authors suggested that these effects may be due to altered maternal behavior (e.g., ineffective
suckling or lack of grooming) rather than cerium being transmitted to offspring in the milk. In
addition, offspring of rats exposed to cerium on GD 7 showed a higher frequency of rearings
than controls when evaluated on PNDs 60-70 (D'Agostino et al., 1982, 1978a, b). Rats exposed
to cerium on GD 7 demonstrated a decrease in activity, measured by the circular runway, on
PND 12 (D'Agostino et al., 1978a, b). Other behavioral differences were not reported. No
possible mechanism of action was discussed to explain the neurodevelopmental effects of cerium
administration. The lack of information on maternal health status following administration of
cerium limits the usefulness of these studies.
4.4.2.2. Neurological Effects
Morganti et al. (1978) examined the open-field and exploratory behavior of 6- to 8-week-
old male Swiss ICR mice following subcutaneous administration of cerium. Three days after
receiving a dose of 20 mg/kg cerium (from a 1:3 cerium chloride-cerium citrate complex),
10 mice were observed in the open-field and exploratory apparatuses and then sacrificed. This
procedure, including injection, was repeated every 3 days until the last group of mice had
received a total of 200 mg/kg cerium. Control mice were injected with sodium citrate. The
results showed that cerium exposure statistically significantly (p < 0.05) depressed ambulations,
marginally depressed explorations, but did not affect rearings. Behavioral measures were not
correlated with cerium levels in tissues. According to Morganti et al. (1978), the lack of
correlation between the behavioral measures and the cerium levels in tissue indicated that the
effect of cerium is not a one-step process depending directly on the level of cerium in a single
tissue but instead involves a biological chain of events.
In a different study by the same group of investigators (Stineman et al., 1978) in which
male Swiss mice received single, subcutaneous injections of cerium (136 or 173 mg/kg), there
was an inverse relationship between open field behavior (ambulations and rearings) and levels of
cerium in blood, brain, lung, stomach, intestine, and kidney. The levels of cerium in the brain
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had the strongest association with decreased open field behavior, followed by the lung.
However, splenic levels of cerium were positively correlated with behavior, as mice with higher
levels of cerium in the spleen exhibited less altered behavior. Stineman et al. (1978) suggested
that the spleen may protect against the cerium-induced effects by sequestering cerium from the
circulation after the removal of damaged erythrocytes and leukocytes.
In a subsequent study (Morganti et al., 1980), 6- to 8-week-old adult male Swiss ICR
mice administered single, subcutaneous injections of 136 or 173 mg/kg cerium showed
depression of general activity, as measured in an activity wheel study and a passive avoidance
study. However, the subcutaneous injection of cerium did not significantly affect two-way
active avoidance learning or social behavior, although gross activity was depressed in the social
behavior study (Morganti et al., 1980). The study authors' interpretation of the study data was
that cerium exposure did not affect simple and complex learning, as measured in the above tests.
4.4.2.3. Hepatic Effects
Numerous studies have examined the effects of cerium on liver parameters following
parenteral dosing. The i.v. administration of cerium oxide (0.5, 1.0, 2.0, 3.5, or 4.5 mg/kg to
females and 0, 3.5, 7.0, or 14.0 mg/kg to males) to Carworth Farms Nelson rats resulted in an
increased level of liver lipids compared to controls, but a dose response trend among the dose
groups was not apparent (Snyder et al., 1959). In a second part of the study, i.v. administration
of 3.5 mg/kg cerium oxide to male and female Sprague-Dawley, Charles River, Carworth Farms
Nelson and Wistar rats resulted in an increase in liver lipids, with a more pronounced increase in
liver lipids in female rats (Snyder et al., 1959).
The i.v administration of a dose of 3 mg/kg cerium (specific form not specified) to rats
resulted in increased serum levels of ornithine-carbamyl transferase (OCT), which reached a
peak 1-2 days after dosing (Magnusson, 1962). Blood glucose levels were reduced markedly in
females 2-3 days after dosing but recovered by day 4. No macroscopic changes were seen in the
liver of males, but fatty liver and fatty degeneration were evident in females 1 day after
treatment. Males showed hydropic changes and isolated necrotic cells 1-4 days after dosing. In
females, changes to the ultrastructure of the liver cells were seen 12 hours after administration of
cerium and appeared to reach a maximum 2-3 days later, with only slight ultrastructural changes
observed in males.
Several additional reports have confirmed and expanded on the Magnusson (1962)
findings. Marciniak and Baltrukiewitcz (1981, 1977) showed that the increase in serum OCT
activity after cerium injection was linear in the range 1.5-4.5 mg/kg cerium. Lombardi and
Recknagel (1962) observed that i.v. administration of 5 mg/kg cerium (form not specified) to
female Sprague-Dawley rats produced a fourfold increase in liver triglyceride levels 24 hours
after treatment. Salas et al. (1976) showed that a single i.v. dose of 10 mg/kg cerium chloride
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administered to female Sprague-Dawley rats decreased the total hepatic adenosine triphosphate
level by 12 hours postexposure and depleted liver glycogen levels and increased liver
triglyceride levels within 48 hours. Ultrastructurally, the rough endoplasmic reticulum was the
main target of cerium toxicity, with marked dilation and degranulation, as well as the appearance
of free ribosomes in the cytoplasm, 24 hours postexposure. The hepatic changes returned to
normal between the 5th and 8th day postexposure.
Arvela and Karki (1971) observed fatty degeneration on the first day following i.v.
administration of 2 mg/kg cerium chloride in Sprague-Dawley rats. On the third day after i.v.
administration, there was a 53% reduction in the level of CYP450 in the liver of male Sprague-
Dawley rats. Arvela et al. (1991) showed that C57BL/6N male mice were more resistant to the
hepatotoxic effects, including necrosis, cell membrane disintegration, and microsteatosis, of
cerium than were DBA/2 mice. However, the concentration of cerium in the livers of C57BL/6N
mice was about 50% higher at 72 hours than in the livers of DBA/2 mice. The authors also
showed that in C57BL/6N mice the slight to moderate liver injury 72 hours post administration
was associated with an increase in coumarin 7-hydroxylase (COH) activity; however, at doses
that caused severe damage in DBA/2 mice, COH activity 72 hours post administration was
drastically reduced. It was also reported that the total amount of CYP450 in the liver was
significantly increased in both strains after a dose of 1 mg/kg cerium, but higher doses tended to
decrease CYP450 content, producing a biphasic relationship. A follow-up study found that
cerium increased the amount of hepatic CYP2A5 mRNA only in DBA/2 mice (Salonpaa et al.,
1992), which have been shown to be more sensitive to cerium exposure than the C57BL/6N
strain. Since the CYP2A5 gene encodes P450 isozymes catalyzing COH activity in mice, the
study authors suggested that some association exists between the development of liver damage
and COH induction.
Strubelt et al. (1980) reported dose-dependent increases in AST (reported using the older
name of glutamic-oxaloacetic transaminase) and ALT (reported using the older name of
glutamic-pyruvic transaminase) and sorbitol dehydrogenase (SDH) in male Wistar rats after i.v.
administration of 3, 5, 7, and 10 mg/kg cerium nitrate.
Sex differences in sensitivity were also reported by Wiener-Schmuck et al. (1990).
Intravenous injection of 1.3 mg/kg of cerium chloride to rats caused lipid deposition, damage to
mitochondria, and invaginations of the nuclear membrane in hepatocytes 48 hours after dosing.
These changes, which were accompanied by increased activities of serum transaminases, were
reversible and occurred only in females. However, when isolated hepatocytes of female rats
were incubated in medium with cerium chloride for 20 hours, there was no sign of cell damage.
The lack of toxicity of cerium to isolated hepatocytes in vitro suggests that hepatocellular lesions
and related changes observed following injection of cerium result from an indirect cause.
Wiener-Schmuck et al. (1990) suggested that deposition of injected rare earths in Kupffer cells
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leads to blockage of the reticuloendothelial system in the liver, inducing damage to
macrophages, which release mediators that in turn damage the hepatocytes.
A more recent paper showed that administration of the metal chelator ethylene glycol
bis(2-aminoethylether)tetraacetic acid (EGTA) to mice after dosing with cerous sulfate for 7
days decreased the severity of the histologic effects of cerium on the liver (Shrivastava and
Mathur, 2004). The histologic effects that resulted from this 7-day subcutaneous exposure of 0.5
mL cerous sulfate included lymphocytic infiltration, hepatocyte hypertrophy, cytoplasmic
vacuolation, and stumpy Kupffer cells. In addition to the histologic effects, the activity of ALT
and AST were statistically significantly increased, with maximum increases of 350 and 98%,
respectively, and hepatic acid, alkaline phosphatases, and succinic dehydrogenase were
statistically significantly decreased, 42, 69, and 46%, respectively. The decrease in severity of
the histologic effects would indicate that the presence of circulating cerium is necessary for
hepatotoxicity but does not provide information on a possible mechanism of hepatotoxicity.
In summary, administration of cerium to rodents caused lipid deposition, mitochondrial
damage, and invaginations of the nuclear membrane in hepatocytes, as well as morphological
changes in the liver, characterized by fatty liver, fatty degeneration, and necrosis. In addition,
histologic effects included lymphocytic infiltration, hepatocyte hypertrophy, cytoplasmic
vacuolation, and stumpy Kupffer cells. Arvela and Karki (1971) suggested that liver toxicity
may result indirectly from induction of CYP2A5 and COH.
4.4.2.4. Cardiovascular Effects
Information regarding effects of cerium on the human heart derives mainly from a series
of studies (see Section 4.1.1) in which naturally high levels of cerium in the soil in certain
geographical regions appear to be correlated with higher levels of cerium in serum and cardiac
tissue of individuals with endomyocardial fibrosis (Eapen, 1998; Kutty et al., 1996; Valiathan et
al., 1989).
The i.v. administration of a single dose of 1.3 mg/kg cerium as cerium chloride to female
Sprague-Dawley rats resulted in a statistically significant twofold increase in protein synthesis
(p < 0.001) and transcription (p < 0.01) in cardiac muscle relative to controls 24 hours after
injection (Kumar et al., 1995). This was consistent with findings by the same group of
investigators who reported that incubation of cardiac fibroblasts in vitro with 100 nM cerium
increased RNA synthesis approximately 64%, but the rate of DNA synthesis was unaffected
(Shivakumar et al., 1992). However, it should be noted that higher concentrations of cerium in
the medium were inhibitory. This was taken as evidence suggesting that cerium at low levels
may act at the level of transcription to stimulate collagen and non-collagen protein synthesis.
This, in turn, may contribute to the accumulation of collagen in endocardial fibrosis. In a
follow-up study, Kumar and Shivakumar (1998) reported that a single i.v. dose of 1.3 mg/kg
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cerium increased lipid peroxidation by 30% in cardiac tissue of Sprague-Dawley rats and
increased proliferation of cardiac fibroblasts by 23%. Treatment with cerium also statistically
significantly decreased collagen degradation by 7% and increased the rate of deposition of newly
synthesized collagen by 27% in cardiac tissue 48 hours after cerium administration (Kumar and
Shivakumar, 1998).
In summary, the limited data from oral exposure experiments in rats and rabbits (see
Section 4.2.1) and i.v. administration studies in rats suggest that cerium increases collagen
accumulation in cardiac muscle by increasing synthesis and decreasing degradation by unknown
mechanisms.
4.4.2.5. Hematological Effects
The i.v. administration of a single dose of 10 mg/kg cerium as the chloride, citrate
complex, or ethylenediamine tetraacetic acid (EDTA) complex to anesthetized dogs every
10 minutes for 10 total doses, significantly increased prothrombin levels and coagulation time
within minutes of the injection (Graca et al., 1964). The magnitude of the increased prothrombin
levels and coagulation time was greatest for cerium chloride, followed by the citrate complex,
and finally the EDTA complex. No other hematological endpoint was significantly affected by
cerium exposure. Talbot et al. (1965) implanted a pellet of cerium metal under the skin of
C57BL mice and collected blood samples from five male and five female mice every 6 months
for hematological determinations. They stated that there were no significant differences between
coagulation times of cerium-implanted mice and controls; however, a table shows that after
6 months, coagulation time in implanted male mice was approximately twice that in controls. In
this study, the only statistically significant difference (p < 0.05) between cerium implanted and
nonimplanted mice was a decrease in total leukocyte counts in males and females at 6, 12, and
18 months relative to controls. However, analysis of differential counts showed no significant
differences between treated and control mice. In another study, Shrivastava and Mathur (2004)
injected subcutaneous doses of cerous sulfate (Ce2[SC>4]3) daily for 7 days into male mice and
reported 18 and 37% decreases in hemoglobin and red blood cell counts, respectively, and 93
and 39% increases in sedimentation rate and hematocrit, respectively, all of which appeared to
reach a maximum on day 14 (7 days after the last injection) and appeared to return to control
levels approximately 60 days posttreatment.
The information available from these few studies is insufficient to determine a possible
mechanism of action by which cerium might be causing hematological alterations. Alterations in
prothrombin and coagulation times could be secondary to liver dysfunction, but more
information is necessary to confirm this hypothesis.
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4.4.2.6. Renal Effects
The i.v. administration of a single dose of 2 mg/kg of cerium chloride to adult male
DBA/2 and C57BL/6 mice resulted in a more than fourfold increase in COH activity in the
kidneys of DBA/2 mice, but less than a twofold increase in C57BL/6 mice (Salonpaa et al.,
1992). The effect was maximized in DBA/2 mice 4 hours after dosing, and the enzyme activity
returned to predosing levels 6 hours after treatment. Cerium also increased CYP2A5 mRNA in
the kidneys of DBA/2 mice sevenfold 6 hours after dosing and sixfold 1 day after the injection,
but no such increase occurred in C57BL/6 mice.
Injection of subcutaneous doses of 0.5 ml (1 mM) cerous sulfate to male mice for 7 days
produced statistically significant (p < 0.05) decreases up to 62 and 42% in the activities of renal
acid and alkaline phosphatases, respectively, and up to 31% in succinic dehydrogenase activity
(Shrivastava and Mathur, 2004). Cerium also induced histologic damage to the kidneys,
consisting of hypertrophy in the epithelial cells, deformed Bowman's capsules, exfoliated nuclei
in tubular lumen, and leukocyte infiltration. Maximum injury was observed on day 21 (14 days
after the last injection). As with the findings regarding liver toxicity, mice administered EGTA
after cerium exhibited less severe necrotic effects in the kidney than mice not receiving EGTA.
The mechanism of kidney toxicity of cerium is unknown.
In summary, cerium exposure initiated an increase in COH activity and CYP2A5
expression and decreased acid and alkaline phosphatases and succinic dehydrogenase activities,
as well as kidney epithelial hypertrophy, exfoliated nuclei in the tubular lumen, and leukocyte
infiltration.
4.4.3. Genotoxicity
No information was located regarding genotoxic effects of cerium or cerium compounds
in humans and only three studies were identified with pertinent information in mammals and in
lower organisms (Sharma and Taluker, 1987; Shimizu et al., 1985; Nishioka, 1975). The
available information is insufficient to ascertain the genotoxicity of cerium.
Shimizu et al. (1985) examined the potential mutagenicity of cerium oxide in five strains
of Salmonella typhimurium (TA98, TA100, TA1535, TA1537, and TA1538) and mEscherichia
coli WP2uvrA. Tests were conducted with and without metabolic activation (S9 fraction from
male Sprague-Dawley rats). Eight different concentrations were tested, ranging from 1 to
5,000 jig/plate. No increase in the number of revertant colonies per plate was observed at any
dose level; in some cases the highest concentration produced growth inhibition. In a short
communication, Nishioka (1975) reported that cerium chloride did not induce DNA damage in
two strains of Bacillus subtilis using the rec-assay. Cerium nitrate was reported to induce
chromosomal breaks and reduce the mitotic index in rat bone marrow in vivo (Sharma and
Talukder, 1987).
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4.5. MECHANISTIC DATA AND OTHER STUDIES
4.5.1. In Vitro Studies
The cytotoxicity of soluble cerium chloride and insoluble cerium oxide was assayed in
Sprague-Dawley rat pulmonary alveolar macrophages (PAMs) and compared to the cytotoxic
and fibrogenic cadmium chloride and oxide (Palmer et al., 1987). Cell viability, effect on
lysosomal enzyme release, and cell morphology were investigated. Cerium chloride was
cytotoxic to rat PAMs with an LCso (concentration inducing 50% cell death) of 29 uM, while
cerium oxide was less toxic, with an LC50 of approximately 4,700 uM; the LC50 values in this
assay for cadmium chloride and oxide were 28 and 15 uM, respectively. Cerium chloride and
oxide did not affect lysosomal enzyme release, although the sensitivity of this assay was
questioned by the study authors. Cerium oxide induced an increase in cells with a featureless
surface and a decrease in cells consistent with the control population. Cells typified by blebs on
the cell surface and/or surface structure were absent. The induction of cells with a featureless
surface was considered by the study authors to be minimal. Cerium chloride was not evaluated
for effect on cell morphology due to an experimental error during cell culture preparation.
Cerium chloride was more cytotoxic than cerium oxide and of similar cytotoxicity to cadmium
chloride, which had an LCso of 28 uM.
Shivakumar and Nair (1991) conducted an in vitro study to examine the effect of cerium
on protein synthesis in cultured rat heart cells and human lung fibroblasts exposed to normal and
reduced levels of magnesium in the growth medium. Cerium exposure resulted in the inhibition
of protein synthesis in rat heart cells and human lung fibroblasts, evident by the decreased
amount of [3H]-tyrosine incorporated into heart cell and fibroblast proteins, 73 and 76%,
respectively; the cell cultures grown on low-magnesium medium displayed a more pronounced
decrease in protein synthesis, with [3H]-tyrosine incorporation decreased 51 and 40% in heart
cell and fibroblast proteins, respectively. However, the mechanism of protein synthesis
inhibition is unknown.
Another in vitro study was conducted to investigate the toxicity of lanthanides, which
includes cerium, towards cultured rat alveolar macrophages (Lizon and Fritsch, 1999). Rat
alveolar macrophages were acquired by pulmonary lavage of 2-month-old male Sprague-Dawley
rats and were cultured for 1 day on culture medium and then for 3 days on medium containing
soluble cerium concentrations of 1 x 10~4 and 5 x 10~6 M. The fraction of apoptotic cells
increased with concentration and as a function of time, with observations at 24, 48, and 78 hours.
As cerium concentration increased from 10~6 to 5 x 10~5 M, the fraction of normal cells
decreased and the fraction of post-apototic and unstained cells increased. The LCso for cerium
was approximately 5 x io~5 M. Cerium exposure to cultured rat alveolar macrophages resulted
in significant alveolar macrophage death.
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Studies with cardiac fibroblasts from neonatal rats in vitro showed that the stimulation of
fibroblast proliferation was accompanied by an increase in the generation of free radicals. Preeta
and Nair (1999) isolated cells from the hearts of 3-4-day-old neonatal Wistar rats. Fibroblasts
were selected from the cultured isolated heart cells via selective adhesion and reincubated in
fresh medium. To assess fibroblast growth, the selected fibroblasts were exposed to cerium
concentrations of 0.1, 0.5, 1, 10, and 100 uM and were harvested 96 hours postexposure.
Growth dynamics were evaluated at 24, 48, and 96 hours postexposure. Immunohistochemical
labeling for proliferating cell nuclear antigen (PCNA) was also conducted to determine if an
increase in cell number was due to cell proliferation. Intracellular free-radical generation was
determined by spectrophotometric assay with reduced nitroblue tetrazolium, with the cardiac
fibroblasts exposed to the same cerium concentrations as in the growth assessment. In addition,
the role of free radicals in cell proliferation was investigated using cardiac fibroblasts exposed to
0.5 uM cerium, SOD (100 U/mL), and catalase (120 U/mL) with cell counts measured after
96 hours.
The results of this investigation showed that cardiac fibroblast proliferation followed a
concentration-dependent response to cerium exposure, with increased proliferation at low
concentrations and decreased proliferation at high concentrations (Preeta and Nair, 1999).
Increased proliferation was evident from 0.1 to 1 uM, with a statistically significant (p < 0.01)
peak at 0.5 uM, while decreased proliferation was evident from 10-100 uM. A statistically
significant increase in the proliferation of PCNA immunoreactive cells was evident at low
cerium concentrations (0.1-1.0 uM), while a decrease was evident at higher concentrations. The
addition of SOD inhibited the increased PCNA expression. The reduction of nitroblue
tetrazolium to formazan, which peaked at 0.5 uM, showed the increase in free radicals from the
fibroblasts exposed to cerium. An increase in free radical production resulted from the fibroblast
exposure to cerium at 0.5 uM, and the cerium-induced stimulatory response was inhibited by the
addition of SOD. This in vitro study demonstrates that low concentrations of cerium stimulate
cardiac fibroblast proliferation in association with an increase in intracellular generation of free
radicals.
A comparative study of cardiac and pulmonary fibroblasts in vitro by Nair et al. (2003)
was conducted to investigate the mitogenic effect of cerium. Fibroblasts were isolated from
heart and lung tissue of 3- to 4-day-old neonatal Wistar rats through selective adhesion. The
cardiac and pulmonary fibroblasts were exposed to fetal bovine serum, a known nonspecific
mitogen, which resulted in cell proliferation in association with intracellular free radical
generation in both types of fibroblasts. Exposure to SOD resulted in significant reductions in
intracellular superoxide anion content and cell density for both types of fibroblasts. The cardiac
fibroblast proliferation was stimulated by exposure to cerium at 0.5 uM for 96 hours, whereas
lung fibroblast proliferation was not stimulated by 0.5 uM cerium exposure for 96 hours. These
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data indicate that cardiac fibroblasts may be more sensitive to cerium exposure than lung
fibroblasts.
Du et al. (2001) exposed packed human erythrocytes to cerium chloride at concentrations
ranging from 4.9 x 10~5 to 3.9 x 10~3 M Ce3+ to investigate the aggregation of membrane
proteins after exposure to cerium. At the highest concentration tested, 3.9 x 10~3 M, aggregation
of membrane proteins was clearly evident, with aggregation increasing gradually with increasing
concentration from 4.9 x 10~5 to 3.9 x 10~3 M. Further analysis, using SDS-PAGE of the
membrane proteins and light scattering measurements, showed that the membrane protein
aggregation was mainly due to non-covalent cross-linking and to a lesser extent oxidative cross-
linking through disulfide bond formation.
4.5.2. Ex Vivo Studies
Manju et al. (2003) utilized isolated papillary muscle in an effort to assess the mechanical
response of the myocardium to varying levels of cerium. Cerium chloride concentrations of 0,
0.1, 0.5, 1, 5, 10, 20, 50, and 100 uM were applied to papillary muscle isolated from Sprague-
Dawley rats and the force of contraction was recorded using a force transducer. The role of SOD
was also investigated by superfusing the muscle with SOD prior to cerium chloride exposure and
recording of contractile force. A statistically significant (p < 0.01) reduction in contractile force
was evident at cerium chloride exposures as low as 0.1 uM, with the lowest dose reducing the
force of contraction approximately 15% (Manju et al., 2003). Complete recovery of contractile
force was apparent at or below 0.5 uM when cerium exposure was removed. The addition of
SOD completely inhibited the effect of cerium exposure, up to 5 uM, on the contractile force of
the isolated papillary muscle. These results provide further evidence for the involvement of
reactive oxygen species in the effects of cerium on the heart.
4.5.3. Nanoparticle Studies
In addition to the in vitro studies, data on nano-sized cerium particles provide
information on absorption and cytotoxicity of cerium. Limbach et al. (2005) measured the
uptake of cerium oxide nanoparticles at four different sizes, 20-50, 40-80, 80-150, and 250-500
nm, into cultured human lung fibroblasts. The cultured fibroblasts absorbed the nanoparticle
cerium linearly with exposure time, with absorption occurring at concentrations as low as 100
ng/g. Particles were not found outside of the vesicles or flowing freely in the cytoplasm and
were present exclusively in the form of agglomerates. The size of the cerium nanoparticle
greatly affected the amount of cerium incorporated into the cell, with better absorption of larger
nanoparticles. Particle size was a more important factor in absorption than particle number and
total surface area (Limbach et al., 2005).
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Brunner et al. (2006) evaluated the cytotoxicity of nanoparticle CeC>2 to human
mesothelioma and rodent fibroblast cell lines by measuring metabolic activity and cell
proliferation. Cerium oxide was tested at exposures of 0, 3.75, 7.5, and 15 ppm for 6 days and 0,
7.5, 15, and 30 ppm for 3 days. The cerium oxide particles had a specific surface area derived
particle size of 6 nm and a hydrodynamic particle size of 19 nm, a specific surface area of
124 ± 3% m2/g, and a GSD of 1.49. Metabolic activity was spectroscopically measured as the
total mitochondrial activity through the conversion of the leuko form of a formazan-type dye to
the active dye, and cell proliferation was measured by the total DNA content of the cells
measured spectroscopically after converting DNA with an intercalating dye into a highly
fluorescent complex (Brunner et al., 2006). The human mesothelioma cells were more sensitive
to CeC>2 than the rodent fibroblast cells, with cell activity and DNA content decreased
approximately 50% after 3 days. The mesothelioma and fibroblasts were, however, not
completely killed at 30 ppm. After 6 days, the cell activity was not significantly altered and
DNA content was increased slightly in the mesothelioma cells; in the rat fibroblasts, the DNA
content increased slightly and the cell activity was not significantly affected.
The impact of a model water dispersion of 7 nm CeC>2 nanoparticles (specific surface
area of 400 m2/g) on E. coli and cytotoxicity, assessed by counting colony-forming units (CPUs),
was investigated at concentrations ranging from 0 to 730 mg/L by Thill et al. (2006). Cerium
was almost completely adsorbed to the bacteria cell surface at a concentration of approximately
30 mg/L, and above this concentration an increasing amount of cerium was found in the
supernatant with a maximum adsorbed concentration of approximately 48 mg/L. The adsorption
appeared to be due to electrostatic attraction between the cerium oxide nanoparticles and the cell
membrane. The percentage of CPUs was strongly affected as the CeO2 nanoparticle
concentration increased, with a 50% survival rate at around 5 mg/L and no survival above
230 mg/L. Thill et al. (2006) also showed that the speciation of the adsorbed cerium was
modified and the nanoparticles reduced. The study authors concluded that direct contact
between the E. coli and the CeC>2 nanoparticles needs to be established for CeC>2 cytotoxicity to
occur and that the reduction of the nanoparticles occurs at or close to the surface of the bacteria
and may be associated with cytotoxicity.
The toxicity of cerium nanoparticles was also investigated using the human
bronchioalveolar carcinoma-derived cell line A549 (Lin et al., 2006). The A549 cell line was
exposed to 20 ± 3 nm cerium oxide nanoparticles at 0, 3.5, 10.5, or 23.3 ug/mL for 24, 48, or
72 hours. The cells were evaluated for cytotoxicity, as well as for intracellular reactive oxygen
species generation, lactate dehydrogenase (LDH) activity, dichlorodihydrofluorescein diacetate
(DCF) fluorescence, GSH levels, a-tocopherol levels, and malondialdehyde (MDA) levels. Cell
viability was decreased at all three dose levels and exposure durations and followed a dose- and
time-dependent decrease, with viability at 3.5, 10.5, and 23.3 ug/mL at 72 hours decreased 12,
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22, and 46%, respectively. LDH levels, indicative of cell membrane damage, increased 15, 32,
and 71% at 24, 48, and 72 hours, respectively, and were greatest at 23.3 ug/mL. DCF
fluorescence, indicative of oxidative stress, increased 70, 139, and 181% after exposure to 3.5,
10.5, and 23.3 ug/mL cerium oxide, respectively. Antioxidant levels were decreased, with
cellular GSH levels reaching a maximal decrease of approximately 70% at 48 hours, with a
possible recovery of GSH levels at 72 hours, and a-tocopherol levels decreased 38, 76, and 88%
at 3.5, 10.5, and 23.3 ug/mL, respectively. MDA levels, indicative of lipid peroxidation, were
significantly increased in a dose- and time-dependent manner in the 3.5, 10.5, and 23.3 ug/mL
dose groups at 48 and 72 hours. Lin et al. (2006) demonstrated the induction of significant
oxidative stress at levels of 3.5, 10.5, and 23.3 ug/mL of 20 nm cerium oxide particles. The
elevated reactive oxygen species levels, increased lipid peroxidation, increased membrane
damage, and reduced antioxidant levels are evidence of the increased oxidative stress from
cerium oxide nanoparticle exposure.
An investigation by Park et al. (2007) described an in vitro screening strategy to examine
the potential for human health effects following exposure to cerium oxide nanoparticles (9 nm
mean particle size). The in vitro assays applied in this strategy were used to investigate local
toxicological effects (skin irritation), cytological effects (structural, functional, or pathological
effects on cells), and genotoxic effects (Ames test). An in vivo immobilization study was also
conducted in Daphnia magna. In the Epiderm™ Skin Irritation Test, nano-cerium oxide and
non-nano-cerium oxide (320 nm mean particle size) particles were not cytotoxic in the MTT
assay (Park et al., 2007). The MTT cytotoxicity assay measures the amount of MTT converted
by the mitochondria of the cell and a reduction of activity is indicative of cell death (Fletcher et
al., 2001). Cytotoxicity was not demonstrated in the BS EN ISO 10993-5 cytotoxicity assay for
both nano-cerium oxide and non-nano-cerium oxide. A cytotoxicity grade of 0 for both nano-
and non-nano-cerium oxide was the same as the cytotoxicity grade for the negative control
(polypropylene pipette tips) and less than the cytotoxicity grade of 3 for the positive control (tin-
impregnated PVC strips) (Park et al., 2007). An increase in the frequency of revertant colonies
in the OECD 471 Ames assay were not observed for any of the tested Salmonella strains
(TA100, TA1535, TA102, TA98, TA1537) at any dose level of nano- or non-nano-cerium oxide
with or without metabolic activation (Park et al., 2007). There were no toxic effects observed at
any time period in the immobilization study in Daphnia magna. The study authors concluded
that the results support a lack of any additional toxicological effects for nano-cerium oxide when
compared to non-nano-cerium oxide (Park et al., 2007).
Fall et al. (2007) exposed lung slices from female Wistar rats to a continuous flow of
nano-sized cerium oxide particles for 3 hours at 14, 76, or 157 mg/m3 to evaluate the potential
biological impact of cerium oxide on lung slices. The cerium oxide particles were approximately
10 nm in diameter, with a mean mobility diameter of approximately 140 nm. The following
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biological endpoints were investigated: cell viability (ATP levels and total glutathione), tumor
necrosis factor (TNF)-a levels, and analysis of a range of antioxidant enzyme levels including
selenium-dependent glutathione peroxidase, catalase, and superoxide dismutase. The viability of
the lung tissue slices, glutathione-dependent metabolism, superoxide dismutase activity, TNF-a,
and proinflammatory factors were not affected by the 3-hour exposure to nanoparticle cerium
oxide. Catalase levels were statistically significantly increased over basal levels at all three
exposure levels, but did not lead to a reduction in cell viability.
A study by Park et al. (2008a) was conducted to examine a range of in vitro cell and cell-
free endponts to assess the toxicity of cerium oxide nanoparticles. The assays included in this
assessment included; general toxicity assays, studies using organotypic cultures of lung slices,
studies on oxidative potential and cytotoxicity, and interleukin (IL)-8 release by A549 cells. The
general toxicity assays conducted by Park et al. (2007) were reported again by Park et al.
(2008a). The studies using organotypic cultures of lung slices used a continuous flow of cerium
oxide aerosol over a 3-hour period were conducted and reported by Fall et al. (2007). The
studies on oxidative potential and cytotoxicity included three experiments: 1) ascorbate depletion
by metal oxide nanoparticles in simple, single antioxidant model, 2) respiratory tract lining fluid
(RTFL) model studies with metal oxide nanoparticles, and 3) cytotoxicity - in vitro cell viability
using A549 cells.
Nano-cerium oxide did not significantly deplete ascorbate in the single antioxidant
model, but did decrease ascorbate in a dose-dependent manner in the RTLF model (Park et al.,
2008a). Nano-cerium oxide at concentrations up to 1000 jiM had no effect on A549 cells
viability, while copper oxide, zinc oxide, and titanium dioxide decreased the viability of A549
cells with LD50 of 300, 375, and 1600 jiM, respectively (Park et al., 2008a). Nano-cerium oxide,
at concentrations of 6.25-100 |ig/ml, did not cause any detectable cytotoxicity to epithelial cells
based on LDH leakage and did not have a significant stimulating effect on IL-8 (Park et al.,
2008a).
Park et al. (2008b) evaluated the cytotoxic effect of cerium oxide nanoparticles using
BEAS-2B cells and compared the results to the cytotoxicity in the T89G and H9C2 cell lines.
The BEAS-2B cell line is derived from human bronchial epithelial cells, where as the T98G cell
line is from human brain fibroblasts and the H9C2 cell line is from rat cardiomyocytes. BEAS-
2B, T89G, and H9C2 cell viability was tested following treatment with 10, 20, and 40 |ig/ml of
30 nm-sized particles for 24, 48, 72, and 96 hours. BEAS-2B, T89G, and H9C2 cells were also
treated with 15, 25, 30, or 45 nm-particles to compare cytotoxicities of different size
nanoparticles. To measure reactive oxygen species (ROS) generation, BEAS-2B cells were
pretreated with 5, 10, 20, 40 |ig/ml of 30 nm-sized particles for 24 hours and then incubated with
40 jiM 2,7-dichlorofluoroscein diacetate for 30 minutes. The relationship between the increased
ROS and the level of antioxidant materials in the cells was investigated by treating cells with 30
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nm-particles at 5, 10, 20, or 40 |ig/ml for 24 hours. Caspase-3 activity was determined by
incubating cells with 5, 10, 20, or 40 |ig/ml 30 nm-particles and lysing the cells and testing for
enzyme activity. BEAS-2B cells were also treated with 40 |ig/ml of 30 nm-particles for 0.5, 1, 2,
4, and 8 hours for inclusion in gene expression analysis.
BEAS-2B cells viability was decreased by treatment with nanoparticles (15-45 nm) in a
time- and dose-dependent manner, and differences in cerium oxide nanoparticle size had no
effect on cell viability (Park et al., 2008b). Cell viability was not reduced in T98G or H9C2 cells
(Park et al., 2008b). ROS were generated in a dose-dependent manner by 5, 10, 20, or 40 |ig/ml
30 nm cerium oxide particles in BEAS-2B cells, and the levels of GSH were decreased in the
nanoparticle-treated groups (Park et al., 2008b). Caspase-3, which plays a key role in the
apoptotic pathway, was increased following treatment with 5, 10, 20 or 40 |ig/ml 30 nm cerium
oxide particles (Park et al., 2008b). In addition, oxidative stress-related genes, including
catalase, glutathione-S-transferase, heme oxygenase-1, and thioredoxin reductase, were induced
by 40 |ig/ml of 30 nm-particles in the BEAS-2B cells (Park et al., 2008b). The cytotoxicity in
BEAS-2B cells appears to be caused by oxidative stress, as cerium oxide nanoparticles increased
the level of ROS with the induction of ROS-related gene expression and decreased the level of
intracellular GSH (Park et al., 2008b). The cerium oxide nanoparticles were also absorbed by
the cells and were observed around the peri-region of the nuclear membrane using a phase-
contrast microscope (Park et al., 2008b).
In addition to the evidence of cytotoxicity, there is evidence of neuroprotective and
cardioprotective effects of nanoparticle cerium exposure. Schubert et al. (2006) exposed the
HT22 hippocampal nerve cell line, derived from the rodent nervous system, to CeC>2
nanoparticles and monitored the intracellular generation of reactive oxygen species with a
nonfluorescent compound that fluoresces when in contact with reactive oxygen species. The
cerium oxide nanoparticles were characterized as single, monodisperse crystals. The CeC>2
nanoparticles were, for the most part, nontoxic to the HT22 cell line, with the exception of the
1 um CeC>2 nanoparticles at concentrations >20 ug/mL. The 6 and 12 nm, as well as 1 um,
cerium particles were protective of oxidative stress to HT22 cells, and a difference in level of
protection offered by the 6 nm, 12 nm, and 1 um sized particles could not be produced. It was
also shown that 12 nm CeC>2 particles at 20 and 200 ug/mL were able to rapidly reduce pools of
reactive oxygen species formed by the cells after 8 hours of exposure to glutamate. Cerium
nitrate and cerium chloride, when tested at the same concentrations as cerium oxide, displayed
no protective effects. Schubert et al. (2006) were able to provide evidence that cerium oxide
nanoparticles have antioxidant properties that promote nerve cell survival under oxidative stress
conditions.
The cardioprotective effects of nano-sized CeCh were evident in cultured cells from MCP
mice administered 15 nmol of 7 nm CeC>2 intravenously twice a week for two weeks (Niu et al.,
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2007). The cardiac-specific expression of monocyte chemoattractant protein (MCP-1) in mice
causes ischemic cardiomyopathy. The treatment of MCP mice with nanoparticle CeC>2 inhibited
monocyte/macrophage infiltration into the myocardial interstitial space, suppressed
proinflammatory cytokine production in the myocardium, and limited myocardial oxidative
stress.
4.6. SYNTHESIS OF MAJOR NONCANCER EFFECTS
4.6.1. Oral
No studies evaluating the oral toxicity of cerium in humans were located, but an
association between exposure to cerium in food and the development of endomyocardial fibrosis
has been suggested (Eapen et al., 1998; Kutty et al., 1996; Valiathan et al., 1989). Long-term
studies in animals are limited to a 13-month drinking water study in rats, which investigated the
effects of a single dose group (Kumar et al., 1996), a 6-month drinking water study in rabbits in
which the administered dose consisted of a mixture of rare earth chlorides (Kartha et al., 1998), a
12-week dietary study in mice (Kawagoe et al., 2005), and a 105-day gavage study in rats
(Cheng et al., 2000). The study of Kumar et al. (1998) suggested that cerium may increase the
levels of collagen in the heart, while the findings of Kartha et al. (1998) suggested that cerium
may intensify the cardiac effects of magnesium deficiency. Kawagoe et al. (2005) suggested that
cerium may increase oxidative stress in tissues of mice, and Cheng et al. (2000) reported that
cerium increased the oxygen affinity of hemoglobin in rats.
A single dose study in mice observed splenic lesions and GI irritation in treated mice
(Stineman et al., 1978). Other acute oral data consist of a report of piloerection, hunched
posture, unsteady gait, and dark coloring of eyes in male and female rats (along with lethargy,
abnormal respiration, and prostration in females only) given single gavage doses of 5,000 mg/kg
cerium as cerium sulfide (Rhodia Inc., 1998); a 7-day LD50 in mice of 1,291 mg/kg cerium as the
chloride/citrate complex (Stineman et al., 1978); and an LD50 of 1,178 mg/kg for cerium nitrate
and 622 mg/kg for cerium oxide, also in mice (Ji and Cui, 1988).
The few long-term studies available in animals identify cardiac tissue and hemoglobin
oxygen affinity as possible health effects, but the animal studies are of limited scope and
insufficient duration and experimental design. Toxicokinetics data have shown that orally
administered cerium compounds are poorly absorbed, and this may be reflected in the results of
the studies conducted with these compounds by the oral route of exposure.
4.6.2. Inhalation
Inhalation data in humans consist of reports describing numerous cases of workers who
developed pneumoconiosis associated with accumulation of cerium in the lungs after prolonged
occupational exposure to cerium fumes or dust (Yoon et al., 2005; Porru et al., 2001; McDonald
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et al., 1995; Pairon et al., 1995, 1994; Sulotto et al., 1986; Vogt et al., 1986; Pietra et al., 1985;
Vocaturo et al., 1983; Sabbioni et al., 1982; Husain et al., 1980; Kappenberger and Biihlmann,
1975; Heuck and Hoschek, 1968). In these cases, the exposure was to cerium oxide, and cerium-
induced pneumoconiosis was characterized by accumulation of cerium particles (and other rare
earth particles) in the lungs and lymphoreticular system. Exposure was not quantified in any of
these cases. Potentially sensitive subgroups were not identified in the available human studies.
Information regarding long-term inhalation exposure in animals is derived from a single
subchronic study in rats (BRL, 1994). Sprague-Dawley rats were exposed nose only to cerium
oxide aerosol 6 hours/day, 5 days/week for 13 weeks. Endpoints evaluated included a functional
observational battery, hematology and clinical chemistry, urinalysis, and gross and microscopic
morphology of tissues. The results revealed statistically significant increases in absolute and
differential neutrophil counts in the blood, treatment-related increases in the absolute and
relative weight of the lungs in both males and females dosed at 50.5 and 507.5 mg/m3 and in the
relative spleen weight of male rats at 507.5 mg/m3, discoloration or pale areas and uncollapsed
parenchyma in the lungs of male and female rats at >50 mg/m3 and pale foci in female rats at
5 mg/m3, and dose-related alveolar epithelial and lymphoid hyperplasia and pigment
accumulation in the lungs, lymph nodes, and larynx of males and females at >5 mg/m3. The
lowest exposure level, 5 mg/m3, was a LOAEL for lymphoid hyperplasia of the lymph nodes. A
NOAEL was not identified.
Acute inhalation data are limited to the determination of an LCso greater than 50 mg/m3
for cerium oxide in rats (Rhone-Poulenc, 1983) and a report of mild histologic alterations,
including slight connective tissue and collagen fiber development and increased lymphocyte
numbers, in the lung, as well as the accumulation of macrophages and large, multinucleated cells
in the spleen, of rats that received an intratracheal instillation of cerium oxide 8 months prior to
examination (Mogilevskaya and Raikhlin, 1967).
4.6.3. Mode-of-Action Information
4.6.3.1. Respiratory Tissues
The mode of action for pulmonary toxicity in humans following chronic inhalation of
cerium oxide has not been demonstrated. However, pathological data from human case reports
demonstrate accumulation of cerium in the lungs following occupational exposure to cerium
fumes or dust. Animal data provide evidence that supports a hypothesized mode of action for
effects observed in the lungs. The occurrence of alveolar epithelial and lymphoid hyperplasia in
the lungs of test animals accompanied by pigmentation in the lungs due to the accumulation of
cerium oxide suggests that the mode of action for cerium oxide inhalation toxicity may be
mediated by cytokine and fibrogenic effects resulting from pulmonary macrophage activation
and immobilization. Taken together, the accumulation of insoluble cerium particles in the
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respiratory tract of humans and animals following chronic and subchronic inhalation exposures,
respectively, suggests that at high exposure concentrations impaired clearance may influence
pulmonary toxicity for both species. Pathology observed in the lung may be due to an immune
response to inhaled cerium dust in which clearance by pulmonary macrophages becomes
overloaded.
One question related to the impairment of clearance in the lung is whether the toxicity
results from the chemical characteristics of cerium oxide versus the physical presence of the
particle. The concept of dust overloading of the lungs refers to inhalation exposures that are
sufficiently intense as to overwhelm the pulmonary clearance mechanisms, specifically,
macrophagic phagocytosis in the alveolar spaces with mucociliary escalation in the bronchial
airways (Morrow, 1988). The concept of particle overload applies to particles of relatively low
cytotoxicity that secondarily induce pulmonary toxicity via chronic activation of immune-
responsive cells (Oberdorster, 1995). In addition to immune cellular activation, the uncleared
particles may traverse the pulmonary epithelial boundary to the interstitial spaces, where
fibrogenesis may occur (Oberdorster, 1995).
Haley (1991) proposed that rare earth metals, including cerium, may exert toxicity both
from innate chemical characteristics as well as pulmonary dust burden. A suggested mechanism
explaining both inflammatory and fibrogenic responses involves the activation of PAMs
following particle overload. Tissue-destructive release products of activated PAMs include acid
hydrolases, elastases, collagenases, and several reactive oxidant species (Haley, 1991).
Additionally, the release of fibroblast growth factor and fibronectin may stimulate the
proliferation of fibroblasts, and PAMs may release neutrophil attractant factors (Haley, 1991).
The subsequently enlisted neutrophils can release several oxidants and proteases that are known
to result in connective tissue damage, possibly stimulating fibrogenesis (Hunninghake et al.,
1984). Although BRL (1994) did not collect data on macrophage activation or the release of
chemokines, the significant increase in mature neutrophils in rats exposed to high levels of
cerium oxide observed in this study is consistent with stimulation of neutrophils by PAMs. The
exposure-related increase in pigment accumulation and lymphoid hyperplasia in lymph nodes
draining the lungs (i.e., bronchial and mediastinal lymph nodes) observed in the BRL (1994)
study and the absence of significant effects in pancreatic and mandibular lymph nodes further
supports the role of pulmonary macrophages.
The immobilization of PAMs by excessive cerium dust loads, as hypothesized by
Morrow (1992, 1988), results in macrophages carrying a heavy cerium oxide load to lose the
ability to move either toward the mucociliary escalator system, for subsequent clearance to the
gut, or to the lymphoid vasculature. A dense tissue population of activated, yet immobile,
macrophages may serve to induce significant cell damage by effectively increasing the
concentration of inflammatory cytokines and fibrogenic growth factors within the pulmonary
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epithelium. Reduction in the ability to clear insoluble cerium oxide from the lung spaces due to
macrophage immobilization is consistent with reports of extracellular presence of pigment in
mid- and high-dose rats relative to low-dose animals (BRL, 1994). The presence of
inflammatory cytokines and growth factors was not examined.
The ability of immobilized macrophages to induce toxicity via concentration of immune
signaling requires minimal cytotoxicity of the macrophages themselves. In vitro experiments
have shown that cytotoxicity of rat pulmonary macrophages is high for soluble cerium (cerium
chloride) (Lizon and Fritsch, 1999; Palmer et al., 1987) but quite low for the insoluble salt
(cerium oxide) (Palmer et al., 1987). This suggests that the pulmonary responses to inhaled
cerium reported in human cases and animal bioassays (which predominantly involved insoluble
cerium oxide) may not have caused significant macrophage cell death. This finding is consistent
with pathological findings in human case reports, animal studies, and in vivo assays of PAMs in
which cerium-related granule pigmentation was visible in the lung and lymphoid tissues,
suggesting that the cerium sequestration and concentration occurred in viable PAMs.
The precipitation and concentration of both soluble cerium chloride and insoluble cerium
oxide within cytosolic lysosomes has been demonstrated (Berry et al., 1997; Berry, 1996). The
high phosphate concentration and extensive enzymatic hydrolysis activity (acid phosphatase)
within lysosomes was shown to effectively precipitate cerium as cerium phosphate. Rats
exposed to 5 ug insoluble cerium oxide via intratracheal dosage for 30 days displayed rat
alveolar macrophages that contained very fine needles or granules in the lysosomes (Berry et al.,
1997). The needle or granule inclusions contained both phosphorus and cerium. After 2 days of
intratracheal dosage, the rat alveolar macrophage lysosomes contained cerium particles
approximately 1 um in length and displayed a crystalline structure. Rats exposed to soluble
cerous chloride displayed fine granules and needles in the lysosomes (Berry et al., 1997). Berry
(1996) showed that cerium precipitated with phosphorus in the lysosomes of hepatocytes and of
bone marrow and splenic macrophages of rats after intraperitoneal injections of cerium.
Granular or needle-like precipitate deposits were observed in the lysosomes of alveolar
macrophages of rats exposed to an aerosol solution of cerium chloride (Galle et al., 1992). The
authors (Berry et al., 1997; Galle et al., 1992) suggest that the precipitation and concentration of
cerium within the lysosomes may serve to inhibit diffusion of the metal to other tissues.
The presence of insoluble cerium oxide particles in the alveolar macrophages of rats
exposed via the intratracheal route (Mogilevskaya and Raikhlin, 1967) is consistent with the
appearance of cerium-related particles within the alveolar macrophages of a worker in an
occupation associated with cerium oxide exposure (McDonald et al., 1995). The particles in the
human alveolar macrophages ranged from small, pinpoint particles to oblong or needle-shaped.
Pairon et al. (1995) revealed particles of phosphorus, calcium, lanthanum, and cerium in human
alveolar macrophages following occupational exposure and identified interstitial macrophages
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that contained both phosphorus and rare earth elements. Cerium was essentially always
associated with phosphorus in the biological samples studied by Pairon et al (1995).
The presence of the insoluble cerium oxide particles in the rat alveolar macrophages
(Berry et al., 1997) and cerium in the lysosomes of hepatocytes and splenic macrophages (Berry,
1996) is also consistent with the involvement of macrophages in the appearance of cerium-
related pigmentation in the rat lung, liver, and spleen following inhalation exposures (BRL,
1994).
Shivakumar and Nair (1991) have shown that cerium exposure resulted in the inhibition
of protein synthesis in human lung fibroblasts, while lung fibroblast proliferation was not
stimulated by 0.5 uM cerium exposure (Nair et al., 2003). However, cell types other than
fibroblasts may be involved in the inflammatory response associated with cytokine and growth
factor release that leads to a fibrotic response (Nair et al., 2003).
Inhalation data on other poorly soluble metal oxides provides support for the
hypothesized mode of action for cerium oxide-induced lung effects. Cullen et al. (2000)
demonstrated overwhelmed pulmonary clearance in male Wistar rats following inhalation
exposure to the poorly soluble metal oxide, titanium dioxide (TiO2), in which the particles were
of similar size and concentration to that used for cerium oxide in the BRL (1994) study. Cullen
et al. (2000) investigated pulmonary effects in male Wistar rats following the 4- and 7-month
inhalation exposure of TiO2 particles with a MMAD of 2.1 |j,m administered at 25 mg/m3 for 209
calendar days and 50 mg/m3 for 118 calendar days. To compare the pulmonary clearance and
effects of TiCMo another poorly soluble particle, barium sulfate particles, with a MMAD of 4.3
|j,m, were administered to male Wistar rats at 37.5 mg/m3 for 203 days and 75 mg/m3 for 119
days. The lymph node burdens, measured in mg/day, for TiC>2 demonstrated the initiation of
overload retardation after approximately 50 days at 50 mg/m3 and 150 days at 25 mg/m3. In
addition, inhalation of TiO2 led to an accumulation of pulmonary macrophages around dust
deposition sites, and most of the macrophages contained phagocytized dust particles. Cullen et
al. (2000) did not include exposure concentrations as low as the BRL cerium oxide study (i.e., 5
mg/m3); however at the lowest concentration of TiC>2 examined, overload was also observed,
although time to overload was longer at the lower concentration (140 days at 25 mg/m3).
Overwhelmed pulmonary clearance, as marked by increased translocation of dust to lymph
nodes, was not observed for barium sulfate (BaSC^) (Cullen et al., 2000). The demonstration of
pulmonary overload by Cullen et al. (2000) in an exposure time frame and at a concentration of
TiC>2 particles with a similar MMAD to the cerium oxide particles in the BRL (1994) study
supports the proposed mode of action of overwhelmed pulmonary clearance for the relatively
insoluble cerium oxide. Although pulmonary overload is a mode of action that may apply to
other relatively insoluble particles of low cytotoxicity, it is clear from the Cullen et al. (2000)
study that there are strong differences between exposures to the dust of different chemicals, such
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that exposure to TiO2 leads to overloading of the lung, whereas overloading was not observed for
BaSC>4. Tran et al. (2000) hypothesizes that the differences between the TiO2 and BaSC>4 in the
level of inflammation and translocation to the lymph nodes may be explained when the lung
burden is expressed as total particle surface area. Barium sulfate, with a low specific surface
area, may be cleared effectively from the lung, as the volumetric capacity of the alveolar
macrophage was not exceeded and overload was not initiated (Tran et al., 2000).
An alternative mode of action may involve the presentation or delivery of cerium oxide
particles to the bronchial lymph nodes from the tracheobronchial region of the lung by dendritic
cells. An important requirement in the induction of adaptive immunity is the transport of antigen
to the T-cell area of the draining lymph nodes from the initial exposure region (Lambrecht et al.,
2001). The lymphoid hyperplasia in the bronchial lymph nodes may represent the presentation
of cerium oxide particles to the lung-draining lymph nodes by dendritic cells. While this mode
of action provides a biologically plausible basis for cerium oxide-induced pulmonary toxicity,
data are lacking that directly evaluate adaptive immunity and cerium oxide exposure.
The proposed mode of action for the pulmonary effects observed following cerium oxide
exposure is the overloading of the PAMs by cerium oxide particles at high doses, leading to
immobility of the PAMs in the alveoli, resulting in the sustained release of inflammatory
cytokines and fibrogenic growth factors and subsequent cell damage. However, additional data
demonstrating pulmonary overload, such as type II cell proliferation, altered alveolar epithelial
integrity, and/or chronic acute inflammation with an influx of PMNs are unavailable following
cerium oxide exposure.
4.6.3.2. Other Tissues
No mode of action for the effects of cerium compounds in non-respiratory tissues was
identified from the available studies. The studies utilized primarily parenteral routes of exposure
and suggest that the production of free radicals may be involved in cerium-associated toxicity, as
well as actions at the RNA transcription level. However, details of potential mode(s) of action in
non-respiratory tissues are lacking.
Cerium, as a cerium chloride-cerium citrate complex, administered subcutaneously was
found to have minor effects on some tests of spontaneous motor behavior in young adult mice
(Morganti et al., 1980, 1978; Stineman et al., 1978) and in mice exposed in utero (D'Agostino et
al., 1982, 1978a, b); however, a mode of action was not discussed or apparent in these studies.
Interestingly, behavioral measures were not correlated with cerium levels in tissues, which
suggested an unknown, indirect mode of action involving a biological chain of events (Morganti
etal., 1978).
Liver effects have been reported in several studies in rats and mice following i.v.
administration of cerium compounds. The effect is characterized by fatty liver, fatty
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degeneration, and necrosis (Magnusson, 1962; Lombard! and Recknagel, 1962). Female rats
appeared more sensitive than male rats (Wiener-Schmuck et al., 1990; Magnusson, 1962).
Incubation of hepatocytes from female rats with cerium did not result in damage to the cells
(Wiener-Schmuck et al., 1990), suggesting that the damage is not a result of direct contact of
cerium with the hepatocytes but that cerium exposure may trigger a series of events that
ultimately result in liver damage. This seems consistent with the results of a study in two strains
of mice having different sensitivities to cerium toxicity that showed that cerium levels in the
liver of the more resistant strain were 50% higher than those in the more sensitive strain (Arvela
et al., 1991). The latter study and an additional study by Salonpaa et al. (1992) presented
evidence suggesting that some association exists between the development of liver damage and
COH induction. Enzyme induction was accompanied by increased expression of CYP2A5
mRNA, but the mechanism by which cerium increases CYP2A5 expression is unknown.
Cerium chloride exposure has been found to increase the levels of collagen in the heart of
animals in an oral study (Kumar et al., 1996) and following i.v. dosing (Kumar and Shivakumar,
1998). Cerium chloride exposure decreased collagen degradation and increased the rate of
deposition of newly synthesized collagen in cardiac tissue of rats (Kumar and Shivakumar,
1998). The increase in collagen synthesis is consistent with a marked increase in mRNA in
cardiac fibroblasts incubated with cerium chloride (Shivakumar et al., 1992), pointing to an
action at the level of transcription. Cerium-induced production of free radicals was also
associated with stimulation of fibroblast proliferation in studies in vitro (Nair et al., 2003; Preeta
and Nair, 1999). While cerium-induced collagen proliferation can explain the occurrence of
cardiac fibrosis, the mechanism by which cerium triggers this event at the molecular level
remains unknown.
In vitro studies demonstrated that low cardiac fibroblast proliferation is stimulated by
exposure to cerium (0.5 uM) (Nair et al., 2003), that low concentrations of cerium stimulate
cardiac fibroblast proliferation in association with an increase in intracellular generation of free
radicals (Preeta and Nair, 1999), and that cerium exposure results in the inhibition of protein
synthesis in rat heart cells (Shivakumar and Nair, 1991). Kuruvilla and Kartha (2006)
demonstrated that cerium reduced the incorporation of [H3]-thymidine into DNA of cardiac
fibroblasts grown in endocardial endothelial cells conditioned medium. These results suggest
that the cardiac lesions in endomyocardial fibrosis may result from the direct stimulation, and
not through toxic effects on the endocardial endothelium, of subendomyocardial fibroblasts by
cerium (Kuruvilla and Kartha, 2006).
Cerium inhibited contraction of isolated rat ventricular papillary muscle, an effect that
could be partially prevented by the free radical scavenger SOD (Manju et al., 2003), which adds
support for the involvement of free radicals in the cerium-induced cardiac effects.
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4.7. EVALUATION OF CARCINOGENICITY
4.7.1. Summary of Overall Weight of Evidence
Data were unavailable regarding the carcinogen!city of cerium compounds in humans or
experimental animals. In accordance with U.S. EPA (2005a) Guidelines for Carcinogen Risk
Assessment, there is "inadequate information to assess the carcinogenic potential" of cerium in
humans.
4.7.2. Synthesis of Human, Animal, and Other Supporting Evidence
No relevant human or animal data are available. In addition, the available information is
insufficient to ascertain the mutagenicity of cerium compounds. The NTP has recently begun an
evaluation of the chronic inhalation toxicity of cerium oxide, including a cancer bioassay. The
date of completion and public availability of the results is unknown but may be expected in
2009.
A study in various strains of S. typhimurium demonstrated negative evidence of
mutagenicity under the conditions of the assay (Shimizu et al., 1985). Cerium chloride did not
induce DNA damage in two strains of B. subtilis by using the rec-assay (Nishioka, 1975), but
cerium nitrate was reported to induce chromosomal breaks and reduce the mitotic index in rat
bone marrow in vivo, and cerium sulfate was reported to cause differential destaining of
chromosomal segments in plants (Sharma and Talukder, 1987).
4.8. SUSCEPTIBLE POPULATIONS AND LIFE STAGES
4.8.1. Possible Childhood Susceptibility
No studies were located regarding possible childhood susceptibility to cerium
compounds. Kutty et al. (1996) stated that high cerium soil concentrations and magnesium
deficiency during childhood may lead to endomyocardial fibrosis, although causation has not
been established. Increased GI uptake of cerium in preweaning, suckling animals compared with
adults is apparent, due primarily to pinocytosis in intestinal cells, but it is unclear if this has any
toxic consequences, since the cerium remains in the intestinal cells, may be minimally available
systemically, and is eventually eliminated in the feces as the intestinal cells die and are replaced
(Kargacin and Landeka, 1990; Kostial et al., 1989a, b; Inaba and Lengemann, 1972).
4.8.2. Possible Gender Differences
No information was located regarding gender differences in humans in response to
exposure to cerium compounds. Information addressing gender differences in animals is from a
study by Magnusson (1962) in which i.v. administration of cerium to rats resulted in noticeably
more severe liver effects in females than in males. This was confirmed in another study in rats
also treated intravenously with cerium (Wiener-Schmuck et al., 1990). There is no explanation
52
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for this gender-related difference in susceptibility, and it is unknown whether it also applies to
endpoints other than the liver. Additionally, it appears that female rats may be more susceptible
to the hematological changes (Table 4-1) observed as the result of cerium oxide inhalation (BRL,
1994).
4.8.3. Possible Susceptible Populations
An association between exposure to rare earth elements, cerium in particular, in food and
the development of endomyocardial fibrosis has been suggested (Eapen, 1998; Kutty et al., 1996;
Valiathan et al., 1989). Cerium levels were elevated in the endomyocardial tissue samples from
patients who died of endomyocardial fibrosis compared with the tissues of controls who died in
accidents or from congenital heart disease (Valiathan et al., 1989). A higher incidence of
endomyocardial fibrosis has been reported among a population consuming tubers grown in a
region of India with high cerium soil concentrations, compared to subjects consuming tubers
grown in a soil with low cerium concentrations. Analysis of the geographic distribution of
endemic endomyocardial fibrosis in India suggested a link to high cerium soil concentrations and
possibly to magnesium deficiency during childhood (Kutty et al., 1996). Populations exposed to
both cerium in food and a diet deficient in magnesium may be susceptible to cardiac effects,
although causality has not been conclusively demonstrated.
A variety of health effects could result from the accumulation of a sufficient amount of
persistently retained particles in the lung (Morrow, 1988). Smoking has suppressive effects on
pulmonary clearance in humans (Morrow et al., 1992), and the co-exposure of cerium with
cigarette smoke, or particulates that accumulate in the lungs, may potentially lead to more severe
pulmonary effects in exposed populations. Chen et al. (2006) found that pulmonary
inflammation, in this case from lipopolysaccharide treatment, may play an integral role in
enhancing the extrapulmonary translocation of particles.
Particle size is another factor that influences overload and toxicity that may be
particularly relevant for human exposure if human exposure is associated with fumes from
carbon arc lamps, as fumes include smaller particles from the low nm range to <1 |im
aggregates. Impaired lung clearance and lung effects in rats exposed to ultrafine particles (<100
nm) occur at lower mass concentrations than in rats exposed to fine particles (<10 jim) (Baan et
al., 2006). Translocation of particles to the insterstitium is a function of number of particles, and
appears to be dependent on the dose and particle size (Ferin et al., 1992). Excessive
translocation into the insterstitium may cause damage to epithelial cells, pulmonary edema, and
eventual fibrosis (Ferin 1994, Oberdorster et al., 1990). Populations exposed to ultra-fine cerium
oxide may have a higher than expected toxicity when compared to cerium oxide particles of a
larger size, as ultrafine particles have higher than expected toxicity when compared to similar
particles of a larger size (Ferin, 1994).
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5. DOSE-RESPONSE ASSESSMENTS
5.1. ORAL REFERENCE DOSE (RfD)
5.1.1. Choice of Principal Study and Critical Effect—with Rationale and Justification
The available human and animal studies demonstrate that cerium may have an effect on
cardiac tissue and hemoglobin oxygen affinity. However, an RfD for cerium was not derived
because the available studies were not suitable for quantitation of effects for various reasons,
including unknown exposure concentrations in the available human studies, lack of a dose-
response, uncertain toxicological significance (e.g., changes in measures of oxidative stress), and
study design (e.g., effects noted only under conditions of a restricted diet).
An association between exposure to cerium in food and the development of
endomyocardial fibrosis has been suggested (Eapen, 1998; Kutty et al., 1996; Valiathan et al.,
1989). In addition, Gomez-Aracena et al. (2006) suggest a relationship between chronic cerium
exposures, characterized by cerium toenail concentrations, and increased risk of acute
myocardial infarction.
The data set for long-term animal studies consists of a 13-month drinking water study in
rats (Kumar et al., 1996), a 6-month drinking water study in rabbits (Kartha et al., 1998), a 12-
week dietary study in mice (Kawagoe et al., 2005), and a 105-day gavage study in rats (Cheng et
al., 2000).
Kumar et al. (1996) demonstrated increased, highly variable cerium levels in cardiac
tissue. Cerium chloride-treated rats, relative to untreated controls, had an increased level of
collagen in the cardiac tissue, with an enhanced effect in rats fed a magnesium-deficient diet.
This study suggested that cerium might increase the levels of collagen in the heart. This study
was conducted on a small number of rats at one dose level (control and dosed rats) and evaluated
few endpoints, and the observed effects were highly variable and not statistically significant.
Kartha et al. (1998) suggested that cerium chloride may intensify the cardiac effects of
magnesium deficiency. Cardiac lesions were apparent in 6/10 rabbits fed a magnesium-deficient
diet with no cerium exposure and 9/10 rabbits fed a magnesium-deficient diet with cerium
exposure. Rabbits fed magnesium-restricted diets, treated with or without cerium, showed
endocardial, subendocardial, interstitial, and perivascular fibrosis and the lesions were more
severe in those with cerium added to the drinking water. Cardiac lesions were absent from the
groups fed the normal magnesium diet regardless of whether they consumed water with or
without cerium. However, this study used only one dose group. The authors reported that
cerium may intensify the cardiotoxicity associated with a magnesium-deficient or restricted diet
but did not elicit a cardiac effect when tested under conditions of a normal magnesium diet.
Cheng et al. (2000) reported that cerium chloride exposure in rats produced a slight
increase of hemoglobin content in erythrocytes after 40 days of treatment with an even greater
54
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increase in hemoglobin content after 80 days of exposure. The effect on the oxygen affinity of
hemoglobin was demonstrated by altered oxygen saturation curves for the dosed rats compared
to control rats. Hemoglobin in cerium-treated rats exhibited altered oxygen affinity up to 80
days of exposure, demonstrated by increased affinity up to 10 mm Hg and a double sigmoidal
curve for 40-day rats and increased affinity above 20 mm Hg for 80-day rats.
Kawagoe et al. (2005) demonstrated that cerium chloride statistically significantly
decreased liver lipoperoxide levels, increased liver GSH levels and liver MT activity, and
decreased plasma SOD activity in mice. Cerium concentrations in the kidney, liver, lung, and
spleen were statistically significantly elevated relative to controls, with the lung and spleen
containing the highest levels. The study authors suggested that the endpoints showing changes
as a result of cerium exposure in this study are indicators of reactive oxygen species generation
and resultant oxidative stress but indicated that their toxicological significance was uncertain.
The authors did not report any other effects in the liver.
5.1.2. Previous RfD Assessment
This is the first IRIS assessment for cerium oxide and cerium compounds; thus, no oral
RfD was previously available on IRIS.
5.2. INHALATION REFERENCE CONCENTRATION (RfC)
5.2.1. Choice of Principal Study and Critical Effect—with Rationale and Justification
Exposure to cerium compounds in the environment is most likely through cerium (eerie)
oxide. There are numerous case reports of workers who developed pneumoconiosis or
interstitial lung disease associated with the accumulation of cerium particles in the lungs after
prolonged occupational exposure to cerium fumes or dust (Yoon et al., 2005; Porru et al., 2001;
McDonald et al., 1995; Pairon et al., 1995, 1994; Sulotto et al., 1986; Vogt et al., 1986; Pietra et
al., 1985; Vocaturo et al., 1983; Sabbioni et al., 1982; Husain et al., 1980; Kappenberger and
Biihlmann, 1975; Heuck and Hoschek, 1968). In these cases, the inhalation exposure was
primarily to cerium oxide, and cerium-induced pneumoconiosis was characterized by
accumulation of cerium particles (and other rare earth particles) in the lungs and lymphoreticular
system. Cerium exposure in humans was associated with interstitial, peribronchial, and
perivascular fibrosis, a restriction of respiratory function and/or pulmonary hypertension (Porru
et al., 2001; Pairon et al., 1995, 1994; Vogt et al., 1986; Vocaturo et al., 1983; Sabbioni et al.,
1982). The available human studies were not selected for the derivation of an inhalation RfC
because the cerium exposures were not available in any of the case reports.
Information regarding long-term inhalation exposure in animals is derived from a single
subchronic study in rats (BRL, 1994). The BRL (1994) study is an unpublished study;
accordingly, it was externally peer reviewed by EPA in August 2006 to evaluate the accuracy of
55
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experimental procedures, results, and interpretation and discussion of the findings presented.
Sprague-Dawley rats (n = 30) were exposed nose only to cerium oxide aerosol 6 hours/day,
5 days/week for 13 weeks. Endpoints evaluated included a functional observational battery,
hematology and clinical chemistry, urinalysis, and gross and microscopic morphology of tissues,
as discussed in Section 4.2.2. As in humans, BRL (1994) demonstrated that the lung and
lymphoreticular system appeared to be the targets of toxicity in rats following inhalation
exposure to cerium oxide. Low-dose effects observed included a statistically significant increase
in absolute and relative lung weight with discoloration or pale areas and uncollapsed
parenchyma in the lungs in both male and female rats at >50.5 mg/m3. Female rats also showed
pale foci in the lungs at >5 mg/m3. In addition, the study authors reported an increased incidence
of pigment accumulation and lymphoid hyperplasia in the bronchial lymph nodes at >5 mg/m3 in
both males and females. In the lungs, an increased incidence of alveolar epithelial hyperplasia
and pigment accumulation was observed at >50 and 5 mg/m3, respectively, in both males and
females. The authors reported that the severity of the hyperplasia in a given tissue, lung or
lymph nodes, was correlated with the amount of pigment accumulated in the tissue (data not
reported). BRL (1994) considered these findings to be consistent with antigenic stimulation by
cerium oxide; however, they did not discuss the possibility of non-antigenic stimulation. Other
effects observed included an increased incidence of metaplasia and pigment accumulation in the
larynx in males at >50.5 mg/m3 and at 50.5 and 5 mg/m3, respectively, in females. The
metaplasia observed in the larynx was interpreted by the study pathologist to be adaptive and
reversible. Hematological changes reported were statistically significant increased numbers of
absolute neutrophils in males at >50 mg/m3, absolute and relative neutrophils in females at > 5
mg/m3, and relative lymphocytes in females at >5 mg/m3 at 13 weeks. The biological
significance of the observed hematological changes is not known.
The mode of action for cerium oxide-induced lung and lymphoreticular toxicity is not
known. However, the available mode of action information from humans and animals (Section
4.6.3.1) suggests that cerium toxicity may be the result of nonspecific stimulation of PAMs that
are activated and immobilized by the accumulation of insoluble cerium particles. The
subchronic BRL (1994) study in rats indicated increased lung weight; discoloration or pale areas,
pale foci, and uncollapsed parenchyma in the lungs; enlargement or pale discoloration of the
bronchial, mediastinal, and pancreatic lymph nodes; and dose-related alveolar epithelial and
lymphoid hyperplasia and pigment accumulation in the lungs and lymph nodes. These effects
are similar to the pneumoconiosis described in the human case reports, which were characterized
by the accumulation of cerium particles in the lungs and lymphoreticular system and histologic
effects throughout the lung. In addition, Hahn et al. (2001, 1999) demonstrated the retention of
cerium-alumniosilicate particles in the tracheobronchial lymph nodes of dogs for several years
following a single inhalation exposure. Clearance of foreign substances from the lung by
56
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macrophages involves removal to the stomach and GI tract, the lymph nodes, and the pulmonary
vasculature (Witschi and Last, 2001). Following inhalation exposure to cerium oxide, it is
possible that macrophages that have phagocytized cerium particles and have retained their
mobility, would be removed from the lung and may accumulate in the lymph nodes. However, if
this process is overwhelmed, PAMs that have been activated and immobilized by the absorption
of cerium oxide particles may be observed.
The only available study reporting low-dose effects of subchronic inhalation exposure to
cerium oxide in animals is the BRL (1994) study. This study was well designed with three dose
groups of 30 animals per group per sex. Numerous tissues and endpoints were assessed, and
methods and observed effects were thoroughly reported. This study identified statistically
significant dose-dependent effects on the lungs and lymphoreticular system in both male and
female rats. Thus, EPA has selected the BRL (1994) study to serve as the principal study.
Alveolar epithelial hyperplasia in the lungs was selected as the critical effect in the derivation of
the inhalation RfC. EPA chose alveolar epithelial hyperplasia in the lungs as the basis for the
determination of the point of departure because this effect represents a sensitive endpoint
occurring at low doses that is considered to be indicative of cerium oxide-induced toxicity.
While it is recognized that lymphoid hyperplasia in the bronchial lymph nodes occurs at a lower
concentration, EPA concluded that this effect may not represent a specific response to cerium
oxide-induced toxicity. Nevertheless, lymphoid hyperplasia in the bronchial lymph nodes may
represent a generalized response to cerium oxide exposure at low concentrations..
5.2.2. Methods of Analysis—Including Models (PBTK, BMD, etc.)
Benchmark dose (BMD) modeling was conducted using the EPA's Benchmark Dose
Software (BMDS, version 2) to analyze the increased incidence of alveolar epithelial hyperplasia
in the lungs of male and female rats (see Appendix B for details). The software was used to
calculate potential points of departure for deriving the chronic RfC by estimating the effective
dose at a specified level of response (BMCX) and its 95% lower bound (BMCLX). A benchmark
response (BMR) of 10% was selected as the level of response because this effect level is
considered to represent a minimal biologically significant change and the BMCLio remained
near the range of the observed data.
Table 5-1 presents BMCs and their corresponding lower 95% confidence limits (BMCLs)
for the increased incidence of alveolar epithelial hyperplasia in the lungs of male and female rats.
The candidate BMC for the critical effect in each sex was identified by comparing the BMDS
outputs from the fitted models. Adequacy of each model fit was determined by assessing the
chi-square goodness-of-fit statistic using a significance level of a = 0.1. The best-fitting models
were selected from those models exhibiting adequate fit by considering the Akaike Information
57
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Criterion (AIC) value of each model (i.e., the AIC value is a measure of the deviance of the
model fit that allows for comparison across models for a particular endpoint) (see Appendix B).
Table 5-1. Selected models and BMCs and BMCLs for the increased incidence
of alveolar epithelial hyperplasia in the lungs of male and female rats.
Sex
male
female
Model
Log-logistic
Multistage
Quantal-linear
Goodness of
fit p-value
0.5278
0.9962
0.8329
X2 p-value
1.28
0.06
0.87
AIC
37.2
21.2
22.7
BMD10
(mg/m3)
4.9
25.9
12.5
BMDL10
(mg/m3)
1.7
8.7
7.2
The log-logistic model fit the alveolar epithelial hyperplasia data in male rats with a
BMCio of 4.9 mg/m3 and BMCLio of 1.7 mg/m3, and both the multistage and the quantal-linear
models fit the increased incidence of alveolar epithelial hyperplasia in female rats with a BMCio
of 25.9 and 12.5 mg/m3 and BMCLio of 8.7 and 7.2 mg/m3, respectively. The BMCLio of 8.7
mg/m3 from the multistage analysis of the increased incidence of alveolar epithelial hyperplasia
in female rats is the selected point of departure for the derivation of the RfC because the
multistage model provided the best fit as characterized by the lowest AIC.
The human equivalent concentration (HEC) was calculated from the point of departure
by adjusting to a continuous exposure (24 hours a day, 7 days a week) and multiplying by a
dosimetric adjustment factor (DAF), which, in this case, was the regional deposited dose ratio
(RDDR) for the pulmonary region of the lung. Adjustment to a continuous exposure was
calculated as follows:
= BMCLio x (6 hours)/(24 hours) x (5 days)/(7 days)
= 9 mg/m3 x 0.25 x 0.71
= 1.6 mg/m3
The RDDR was calculated using the RDDR v.2.3 program (Table 5-2), as described in
Methods for Derivation of Inhalation Reference Concentrations and Application of Inhalation
Dosimetry (U.S. EPA, 1994b). The pulmonary region of the lung was selected as the deposition
site because the critical effect and the mode of action data indicate that the pulmonary region of
the lung is the initial location of cerium oxide toxicity.
The human parameters used were body weight, 70 kg; minute volume, 13.8 L;
extrathoracic surface area, 200 cm2; tracheobronchial surface area, 3,200 cm2; and pulmonary
surface area, 54.0 m2. The parameters used for the rat were body weight, 345 g (calculated by
averaging the mean body weights for each dose group for the course of the experiment); minute
volume, 234.18 mL; extrathoracic surface area, 15 cm2; tracheobronchial surface area,
22.50 cm2; and pulmonary surface area, 0.34 m2. The GSD for the cerium oxide particles in the
58
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BRL (1994) study ranged from 1.8 to 1.9 and the MMAD for the particles was approximately
2.0 um. The RDDRs calculated from the above model parameters vary depending upon the
deposition site in the lung (Table 5-2). The RDDR for pulmonary deposition is 0.536. The
calculation is as follows:
BMCLHEc = BMCLADj x RDDR
= 1.6mg/m3 x 0.536
= 0.86 mg/m3
Table 5-2. Output from RDDR v.2.3 used in the analysis in Section 5.2.2
Species
Rat
Human
Rat
Human
Ratio
Body
weight (g)
345
70000
0.005
VE (ml)
234.2
13800.0
0.017
RDDR
Ratio
RDDR
Extrathoracic
SA (cm2)
15.000
200.000
0.075
dep
0.542
0.352
1.540
0.348
Thoracic
SA (cm2)
0.342
54.320
0.006
dep
0.091
0.125
0.724
0.738
Tracheobronchial
SA (cm2)
22.500
3200.000
0.007
dep
0.042
0.085
0.490
1.183
Total RT
SA (cm2)
0.344
54.340
0.006
dep
0.632
0.682
0.927
2.487
Pulmonary
SA (m2)
0.340
54.000
0.006
Dep
0.049
0.245
0.199
0.536
Extrarespiratory
SA (m2)
345
70000
0.005
Dep
0.632
0.682
0.927
3.191
MMAD = 2.00; Sigma g = 1.85.
5.2.3. RfC Derivation—Including Application of Uncertainty Factors (UFs)
Based on the selection of alveolar epithelial hyperplasia in the lungs of male and female
CD rats (BRL, 1994) as the critical effect, the use of the BMCLHEc value of 0.86 mg/m3 as the
point of departure for the derivation of the RfC for cerium oxide is presented below. A total UF
of 1,000 was applied to this point of departure: 3 for extrapolation from animals to humans
(UFA: animal to human), 10 for consideration of interindividual variability (UFH: human
variability), 10 for extrapolation from a subchronic study (UFS), and 3 for database deficiencies
(UFD). The rationale for the application of the UFs is described below.
A factor of 3 was selected to account for uncertainties in extrapolating from rats to
humans (UFA). This value is adopted by convention where an adjustment from an animal-
specific LOAELADJ to a LOAELnEc has been incorporated. Application of a full UF of 10 would
depend on two areas of uncertainty (i.e., toxicokinetic and toxicodynamic uncertainties). In this
assessment, the toxicokinetic component is mostly addressed by the determination of a HEC as
described in the RfC methodology (U.S. EPA, 1994b). The toxicodynamic uncertainty is also
accounted for to a certain degree by the use of the applied dosimetry method.
59
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A factor of 10 was used to account for variation in susceptibility among members of the
human population (UFH). Insufficient information is available to predict potential variability in
susceptibility among the population to inhaled cerium oxide and cerium compounds.
A factor of 10 was used to account for uncertainty in extrapolating from a subchronic to
chronic (UFs) exposure duration, since the BRL (1994) study, which was selected as the
principal study, is a subchronic study. The critical effect, increased incidence of alveolar
epithelial hyperplasia, may be more pronounced at longer exposure durations.
An UF for LOAEL-to-NOAEL extrapolation was not applied in this case, because the
current approach is to address this factor as one of the considerations in selecting a BMR for
benchmark dose modeling. The data for alveolar epithelial hyperplasia in the lungs were
modeled using BMD modeling and a BMR of a 10% was selected under an assumption that it
represents a minimal biologically significant change.
A factor of 3 was used to account for deficiencies in the cerium oxide database. The
database includes multiple case reports of inhalation exposure to workers and a single 13-week
subchronic inhalation study in rats. The effect from the subchronic rat inhalation study that was
used for the derivation for the RfC (i.e., alveolar epithelial hyperplasia) may be an early indicator
of the more overt toxicity that is found in humans (i.e., interstitial lung disease) exposed to
cerium oxide in the workplace. The database does not include an exposure and recovery study
that could demonstrate the persistence or, conversely, the adaptive nature of either the alveolar
epithelial hyperplasia or lymphoid hyperplasia in the bronchial lymph nodes.
Toxicity via the inhalation route is expected to be a portal-of-entry effect. Cerium oxide
is a relatively insoluble metal oxide and absorption or translocation from the lung to the
circulation is expected to be minimal at low doses. The pulmonary effects observed in the
human case reports and in the BRL (1994) study are likely due to the physical deposition of
cerium oxide particles in the lung and the immunological reaction to the particles, and are not
likely to be due to a chemical reaction of cerium oxide with lung tissues. The observed
immunological response is a portal-of-entry effect and systemic circulation and effects are not
expected because of the insoluble nature of the cerium oxide particles.
In considering the impact of database deficiencies on the derivation of the RfC,
substantial weight was given to the available data demonstrating similarities between the effects
observed in humans following prolonged exposure to cerium oxide and the effects observed in
rats in the subchronic principal study, along with the data on deposition and absorption of cerium
oxide in the lung. Thus, these data support the assumption that the respiratory system may be
the most sensitive target of toxicity following inhalation exposure to cerium and that inhalation
exposure to cerium may primarily involve portal-of-entry effects.
The database for cerium oxide lacks both a two-generation reproductive toxicity bioassay
and a developmental toxicity bioassay. Systemic effects following the inhalation of cerium
60
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oxide, with an MMAD of approximately 2.0 um and a GSD from 1.8 to 1.9, are not likely to be
observed. While it is recognized that the investigation of systemic effects following cerium oxide
exposure has not been the focus of existing studies, there is no reason to expect that
reproductive, developmental, or other systemic effects would occur, and a UF of 3 is considered
to be sufficient for the absence of data on these effects.
The RfC for cerium oxide based on alveolar hyperplasia in the lung was calculated as
follows:
RfC = BMCLHEc-UF
= 0.86mg/m3-1000
= 9 x 1CT4 mg/m3 (rounded to one significant figure)
Note that the RfC was quantified for cerium oxide particles with an MMAD of
approximately 2.0 um and a GSD from 1.8 to 1.9, and may not characterize the potential toxicity
from exposures to cerium oxide particles with smaller MMADs and GSDs, including nano-sized
cerium particles. The use of the RfC for cerium compounds other than cerium oxide is not
recommended as the similarity between this form of cerium and other cerium compounds is
unknown.
5.2.4. RfC Comparison Information
Figure 5-1 is an exposure-response array, which presents NOAELs, LOAELs, and the
dose range tested corresponding to selected health effects from the BRL (1994) study, some of
which were considered candidates for chronic RfC derivation. Figure 5-2 presents the point of
departure, calculated as a human equivalent dose, applied uncertainty factors, and derived
sample chronic inhalation reference values for selected endpoints from Figure 5-1. This
comparison is intended to provide information for additional endpoints associated with cerium
oxide inhalation.
61
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1000
100
O)
10
^ V
*• .<^
f <5S>
v
• NOAEL ^ *f $f / # ^ ^ /•
• I OAF I ^ ^ ^
l_ vx r\ I I— ,J2» ^^ />O
The vertical lines represent the .^ $ &
range of doses tested in a given ^° ^
study. ^ ^
Figure 5-1. Exposure-response array of selected toxicity effects from the BRL (1994) study
Source: BRL, 1994.
^
62
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Figure 5-2. Points of departure for selected endpoints (with critical effect circled) from Figure 5-1 with corresponding
applied uncertainty factors and derived chronic inhalation RfVs.
10
I
0.1
0.01
0.001
0.0001
ssssss,
ssssss,
ssssss,
ssssss.
ssssss.
Lymphoid
hyperplasia
- bronchial
lymph
node (&$)
Pigment
accumulation
- bronchial
lymphoid
Pigment
accumulation
-lung(c?,$)
POD
RfV
Increased
absolute lung
Lymphoid
hyperplasia
-lung (?)
Alveolar
epithelial
hyperplasia
Lymphoid
hyperplasia
- lung
UFH
UFsc
UFL
UFD
63
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5.2.5. Previous RfC Assessment
This is the first IRIS assessment for cerium oxide and cerium compounds; thus, no
inhalation RfC was previously available on IRIS.
5.3. UNCERTAINTIES IN THE ORAL REFERENCE DOSE (RfD) AND INHALATION
REFERENCE CONCENTRATION (RfC)
Risk assessments need to portray associated uncertainty. The following discussion
identifies uncertainties associated with the RfC for cerium oxide. As presented earlier in this
chapter (Sections 5.2.2 and 5.2.3), uncertainty factors were applied to the point of departure for
the RfC. Factors accounting for uncertainties associated with a number of steps in the analyses
were adopted to account for extrapolating from an animal bioassay to human exposure, a diverse
population of varying susceptibilities, extrapolating from a subchronic to chronic exposure
duration, extrapolating from a LOAEL to a NOAEL, and database deficiencies.
A limited range of animal toxicology data is available for the hazard assessment of
cerium oxide, as described throughout the previous sections (see Sections 4 and 5). For the oral
route, human case reports showing an association between exposure to cerium in food and the
development of endomyocardial fibrosis are available. Long-term oral studies in animals are
limited to a 13-month drinking water study in rats, which investigated the effects of a single dose
group; a 6-month drinking water study in rabbits in which the administered dose consisted of a
mixture of rare earth chlorides; a 12-week dietary study in mice; and a 105-day gavage study in
rats. An RfD for cerium oxide and cerium compounds was not derived since the available
studies were not suitable for quantitation of effects for various reasons, including unknown
exposure concentrations in the available human studies, lack of a dose response, uncertain
biological significance (e.g., changes in measures of oxidative stress), and study design (e.g.,
effects noted only under conditions of a restricted diet).
The inhalation database includes numerous case reports of workers who developed
pneumoconiosis associated with accumulation of cerium particles in the lungs after prolonged
occupational exposure to cerium fumes or dust and a single, 13-week subchronic inhalation
bioassay of cerium oxide exposure in rats (BRL, 1994). The exposure concentrations utilized in
the BRL (1994) study were higher than expected human environmental exposures to cerium
oxide. Thus, the selection of the BRL (1994) as the principal study may overestimate the risk to
cerium oxide particles. In addition, due to significant uncertainty related to toxicokinetic and
toxicodynamic differences between soluble and insoluble cerium compounds, the RfC for cerium
oxide should not be applied to other cerium compounds.
Consideration of the available dose-response data to determine an estimate of inhalation
exposure that is likely to be without an appreciable risk of adverse health effects over a lifetime
led to the selection of the 13-week subchronic inhalation bioassay in Sprague-Dawley rats (BRL,
64
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1994) and increased incidence of alveolar epithelial hyperplasia in the lungs of male and female
rats as the principal study and critical effect for deriving the RfC for cerium oxide.
The mode of action for the observed pulmonary effects, along with the lymphoreticular
effects is not known. The hypothesized mode of action for the pulmonary effects observed
following cerium oxide exposure is the overloading of the PAMs by cerium oxide particles,
leading to immobility of the PAMs in the alveoli, resulting in the sustained release of
inflammatory cytokines and fibrogenic growth factors and subsequent cell damage. This
proposed mode of action is based on the results from mechanistic studies, toxicity reviews of
cerium oxide or similar compounds, in vitro cerium oxide and soluble cerium studies,
intratracheal cerium oxide instillation studies, and human case reports. Additional data
demonstrating particle overload following cerium oxide exposure, such as type II cell
proliferation, altered alveolar epithelial integrity, and/or chronic acute inflammation with an
influx of PMNs, are unavailable. The data demonstrating particle overload following exposure
to cerium oxide were at relatively high doses. Thus, the mode of action for toxicity induced by
chronic exposure to lower doses of cerium oxide may not be related to overloaded pulmonary
clearance mechanisms, and the effects observed following low dose chronic exposures may also
be different.
Based on the available evidence the effects observed in the lungs and lymph nodes of
male and female rats following inhalation exposure to cerium oxide in the BRL (1994) study
were considered to be indicative of overwhelmed pulmonary clearance and potential key events
in the proposed mode of action. However, as described in Section 4.6.3.1, evidence for
overwhelmed pulmonary clearance as a mode of action for pulmonary and lymphoreticular
system effects observed with inhalation exposure to cerium oxide relies on data from other
relatively insoluble particles (such as titanium dioxide) due to the limited mechanistic data on
cerium oxide. In the absence of evidence indicating otherwise, these effects are assumed to
represent cerium oxide-induced toxicity.
Benchmark dose (BMD) modeling was conducted using the EPA's Benchmark Dose
Software (BMDS, version 2) to analyze the increased incidence of alveolar epithelial hyperplasia
in the lungs of male and female rats (see Appendix B for details). The software was used to
calculate potential points of departure for deriving the chronic RfC by estimating the effective
dose at a specified level of response (BMCX) and its 95% lower bound (BMCLX). A benchmark
response (BMR) of 10% was selected for the modeling of the increased incidence of alveolar
epithelial hyperplasia in male and female rats. The lung and lymphoreticular system effects
observed by BRL (1994) are comparable to the effects observed in humans that were
characterized by the accumulation of cerium particles in the lungs and lymphoreticular system
and histologic effects throughout the lung. Thus, EPA has selected the BRL (1994) study to
serve as the principal study. Alveolar epithelial hyperplasia in the lungs was selected as the
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critical effect in the derivation of the inhalation RfC. While it is recognized that lymphoid
hyperplasia in the bronchial lymph nodes occurs at a lower concentration, EPA concluded that
this effect may not represent a specific response to cerium oxide-induced toxicity. Nevertheless,
it should be noted that lymphoid hyperplasia in the bronchial lymph nodes may represent a
generalized response to cerium oxide exposure at low concentrations.
Extrapolating from animals to humans embodies further issues and uncertainties. The
effect and its magnitude associated with the concentration at the point of departure in rodents are
extrapolated to human response. Pharmacokinetic models are useful to examine species
differences in pharmacokinetic processing; however, dosimetric adjustment using
pharmacokinetic modeling was not possible for the effects observed following inhalation
exposure to cerium oxide. For the RfC, a HEC was calculated from the point of departure by
multiplying the BMCLADJ by a DAF. The calculated DAF in this assessment is an RDDR. The
RDDR was calculated using the RDDR v.2.3 program, as described in Methods for Derivation of
Inhalation Reference Concentrations and Application of Inhalation Dosimetry (U.S. EPA,
1994b).
Heterogeneity among humans is another uncertainty associated with extrapolating doses
from animals to humans. Uncertainty related to human variation needs consideration, also, in
extrapolating doses from a subset or smaller sized population, say of one sex or a narrow range
of life stages typical of occupational epidemiologic studies, to a larger, more diverse population.
Human variation may be larger or smaller; however, cerium-specific data to examine the
potential magnitude of over- or underestimation is unavailable.
Critical data gaps have been identified with uncertainties associated with regards to
chronic toxicity, especially evidence demonstrating a persistence of alveolar epithelial
hyperplasia, and reproductive and developmental toxicity associated with inhalation exposure to
cerium oxide. The available oral cerium exposure information in both humans and rats identifies
cardiac tissue and hemoglobin oxygen affinity as possible health effects, but the animal studies
are of insufficient duration and experimental design and the human studies do not provide
exposure characterization. The lack of a study to derive an RfD given the possible effects
demonstrated represents a critical data gap in the oral database. In addition, it is unclear whether
the cerium oxide particles in the lung or lymph nodes can disassociate and release cerium
systemically, possibly causing systemic effects, such as cardiotoxicity.
A chronic, cerium oxide inhalation exposure study in animals is unavailable; however,
the workers in the human case reports were exposed to cerium oxide for periods of 10 to
46 years, and, while the case reports do not contain the necessary exposure analysis for dose
response assessment, they do provide evidence of respiratory effects following long-term
exposure to cerium oxide. There are limited data available addressing possible reproductive or
neurodevelopmental toxicity following exposure to cerium. The accumulation of insoluble
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cerium particles in the respiratory tract of humans and animals following chronic and subchronic
inhalation exposures, respectively, suggests that impaired clearance may influence pulmonary
toxicity in both rats and humans and limit systemic availability; therefore, possible reproductive
or developmental toxicity may be expected to occur, if at all, at doses higher than those at which
portal-of-entry effects occurred.
The RfC derived in this assessment is for cerium oxide particles with an MMAD of
approximately 2.0 um and a GSD of 1.8 to 1.9, and the application of the RfC to cerium oxide
particles with smaller MMADs and GSDs, including nano-sized cerium particles is not
recommended. The lack of data to support the derivation of an RfC for nano-cerium oxide is a
significant area of uncertainty.
5.4. CANCER ASSESSMENT
Studies addressing the carcinogenic effects of cerium or cerium compounds upon which
to base a cancer assessment are unavailable. Lundgren et al. (1996) observed lung neoplasms
(7/1,049) in cerium oxide exposed control F344/N rats in an investigation of the carcinogenicity
of the beta-particle emitter, 144Ce. The incidence of lung neoplasms in the stable cerium oxide
exposed rats was not compared to a treatment-free group or to historical background levels in
F344/N rats.
In accordance with U.S. EPA (2005a) guidelines for carcinogen risk assessment, there is
"inadequate information to assess the carcinogenic potential" of cerium oxide and cerium
compounds. Lack of carcinogenicity data precludes derivation of an oral slope factor or
inhalation unit risk. Genotoxicity evidence was insufficient to assess the genotoxic potential.
This overall lack of information represents a data gap and does not allow for a quantitative
assessment of the carcinogenicity of cerium oxide and cerium compounds. A previous cancer
assessment was not available for cerium oxide and cerium compounds.
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6. MAJOR CONCLUSIONS IN THE CHARACTERIZATION OF
HAZARD AND DOSE RESPONSE
6.1. HUMAN HAZARD POTENTIAL
Cerium oxide (CeO2; eerie oxide) is used either in the pure form or in a concentrate as a
polishing agent for glass mirrors, plate glass, television tubes, ophthalmic lenses, and precision
optics (Kilbourn, 2003; Reinhardt and Winkler, 2002). Cerium oxide is used as a glass
constituent to prevent solarization and discoloration (especially in the faceplates of television
screens) (Reinhardt and Winkler, 2002). Cerium oxide is also used as a diesel fuel-born catalyst
to reduce paniculate matter emissions (HEI, 2001). Cerium is not expected to exist in elemental
form in the environment since it is a reactive metal (Lewis, 2001). Cerium compounds are not
expected to volatilize and will exist in the particulate form if released into the air.
Toxicokinetic studies in rodents have examined the absorption, distribution, metabolism,
and elimination of cerium. In adult rats, cerium compounds are poorly absorbed following oral
exposure, while suckling animals exhibit higher absorption and retention of cerium in the GI
tissues (Kostial et al., 1989a, b; Inaba and Lengemann, 1972). Following inhalation exposure,
cerium, a poorly-soluble particle, behaves like other airborne particles, depositing within the
respiratory tract based on aerodynamic character (for review, see Schulz et al., 2000). Cerium
has been detected in lung tissues and in alveolar macrophages of subjects believed to have been
exposed occupationally (Vocaturo et al., 1983; Sabbioni et al., 1982), with cerium concentrations
in the lung tissues 2,800-207,000 times higher than those found in the urine, blood, or nails
(Pietra et al., 1985). The early clearance of the radioactive cerium administered dose suggests
that the majority of cerium aerosol is deposited in the airways, where it is subject to removal via
the mucociliary escalator, swallowing, and elimination in the feces.
Once absorbed into the body, cerium tends to accumulate primarily in the bone, liver,
heart, and lung. Cerium has been observed to be localized in the cell, particularly in the
lysosomes, where it is concentrated and precipitated in an insoluble form in association with
phosphorus. As an element, cerium is neither created nor destroyed within the body. The
particular cerium compound (e.g., cerium chloride, cerium oxide) may be altered as a result of
various chemical reactions within the body, particularly dissolution, but data have not
demonstrated a change in the oxidation state of the cerium molecule (Berry et al., 1989, 1988).
Following inhalation exposure, the initial rapid elimination of cerium from the body is
due primarily to transport up the respiratory tract by the mucociliary escalator and eventual
swallowing of the material, as with other poorly soluble particles (Boecker and Cuddihy, 1974).
After the initial clearance of cerium particles from the upper respiratory tract, pulmonary
clearance is slower, with reported slow-phase clearance half-times ranging from 100 to 190 days
in rodents (Lundgren et al., 1974; Thomas et al., 1972; Morgan et al., 1970; Sturbaum et al.,
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1970). Elimination of orally administered cerium has been shown to be age dependent in
animals, with suckling animals absorbing cerium into the GI tissues (Inaba and Lengemann,
1972).
An epidemiological study reports a higher incidence of endomyocardial fibrosis among a
population consuming tubers grown in a high cerium soil in India (Eapen, 1998; Kutty et al.,
1996; Valiathan et al., 1989), and a case control study found an association between increased
toenail cerium concentrations and the risk of first myocardial infarction (Gomez-Aracena et al.,
2006). The few long-term animal studies available included dietary, gavage, and drinking water
administrations, although clear indicators of health effects were not evident.
Numerous case reports have been published describing cases of workers who developed
adverse lung effects, such as interstitial lung disease or pneumoconiosis, associated with
accumulation of cerium in the lungs after prolonged occupational exposure to cerium fumes or
dust (Yoon et al., 2005; Porru et al., 2001; McDonald et al., 1995; Pairon et al., 1995, 1994;
Sulotto et al., 1986; Vogt et al., 1986; Pietra et al., 1985; Vocaturo et al., 1983; Sabbioni et al.,
1982; Husain et al., 1980; Kappenberger and Buhlmann, 1975; Heuck and Hoschek, 1968). The
human cases of cerium exposure demonstrate the accumulation of cerium particles in the lungs
and lymphoreticular system, with pulmonary function varying from normal to severe restriction
and interstitial fibrosis in one case and granulomas in another. Interstitial fibrosis accompanied
by vascular thickening, reactive alveolar macrophages, abundant macrophages in the airspace,
and moderate chronic interstitial inflammation, along with small interstitial clumps of
macrophages bearing scant deposits of grayish-black pigment, was observed in a 68-year-old
man who was employed as an optical lens grinder for 35 years (McDonald et al., 1995).
Additionally, particles characterized as cerium were identified in alveolar macrophages,
macrophages in the tracheobronchial lymph nodes, and lung and lymph node tissue (Porru et al.,
2001; Pairon et al., 1995; Waring and Watling, 1990; Sulotto et al., 1986; Vogt et al., 1986;
Pietra et al., 1985). Cerium exposure was associated with interstitial, peribronchial, and
perivascular fibrosis, a restriction of respiratory function, and/or pulmonary hypertension (Porru
et al., 2001; Pairon et al., 1995, 1994; Vogt et al., 1986; Vocaturo et al., 1983; Sabbioni et al.,
1982).
Data from a subchronic toxicity test in Sprague-Dawley rats (BRL, 1994) identified an
increased incidence of alveolar epithelial hyperplasia as the critical effect for noncancer effects.
Histologic examination revealed dose-related alveolar epithelial and lymphoid hyperplasia and
pigment accumulation in the lungs, lymph nodes, and larynx of male and female rats at >5
mg/m3.
The mode of action for cerium-oxide induced lung and lymphoreticular system effects
observed in rats is not known. However, evidence suggests that the overloading of PAMs by
cerium oxide particles at high doses, leading to immobility of PAMs in the alveoli, resulting in
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the sustained release of inflammatory cytokines and fibrogenic growth factors and subsequent
cell damage may be a plausible mode of action for these effects. The accumulation of insoluble
cerium oxide particles in the respiratory tract of humans and rodents following chronic and
subchronic exposure, respectively, suggests that impaired clearance may influence pulmonary
toxicity for rats and humans. A population of immobilized, activated macrophages may serve to
induce significant cell damage by effectively increasing the concentration of inflammatory
cytokines and fibrogenic growth factors within the pulmonary epithelium.
6.2. DOSE RESPONSE
6.2.1. Noncancer/Oral
The database for oral exposure to cerium is limited to geographical distribution studies
evaluating the possible association between exposure to cerium in food and the development of
endomyocardial fibrosis (Eapen et al., 1998; Kutty et al., 1996; Valiathan et al., 1989), and long-
term animal studies, including a 13-month drinking water study in rats (Kumar et al., 1996), a
6-month drinking water study in rabbits (Kartha et al., 1998), a 12-week dietary study in mice
(Kawagoe et al., 2005), and a 105-day gavage study in rats (Cheng et al., 2000).
The geographical distribution studies suggest that there is an association between
exposure to cerium in food and the development of endomyocardial fibrosis but are unsuitable
for RfD derivation. The above animal studies are unsuitable for derivation of an RfD for various
reasons. The 13-month study by Kumar et al. (1996) was conducted on a small number of rats at
only one dose level (control and dosed rats) and evaluated few endpoints, and the health effects
were highly variable and not statistically significant. Kartha et al. (1998) conducted a 6-month
drinking water study in rabbits that utilized a rare earth chloride mixture in the drinking water,
which suggests that cerium may enhance the effect of magnesium deficiency in heart tissue. The
Cheng et al. (2000) study was limited in scope, since it investigated the effects of cerium
chloride exposure on the structure and affinity of hemoglobin in rats. The evaluations conducted
in Kawagoe et al. (2005), showing statistically significant changes as a result of cerium exposure
(e.g., decreased lipoperoxide levels), increased GSH levels and MT activity, and decreased SOD
activity, are of unknown biological significance and may represent oxidative stress in response to
cerium exposure.
The available oral cerium exposure information in both humans and rats identifies
cardiac abnormalities and changes in hemoglobin oxygen affinity as possible effects, but the
animal studies are of insufficient duration and experimental design and the human studies do not
provide adequate exposure characterization. The lack of a study to derive an RfD given the
effects demonstrated represents a critical data gap in the oral database.
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6.2.2. Noncancer/Inhalation
There are numerous case reports of workers who developed pneumoconiosis, associated
with accumulation of cerium particles in the lungs, after prolonged occupational exposure to
cerium fumes or dust (Yoon et al., 2005; Porru et al., 2001; McDonald et al., 1995; Pairon et al.,
1995, 1994; Sulotto et al., 1986; Vogt et al., 1986; Pietra et al., 1985; Vocaturo et al., 1983;
Sabbioni et al., 1982; Husain et al., 1980; Kappenberger and Buhlmann, 1975; Heuck and
Hoschek, 1968). Information regarding long-term inhalation exposure in animals is derived
from a single subchronic study in rats (BRL, 1994). Sprague-Dawley rats were exposed nose
only to cerium oxide aerosol 6 hours/day, 5 days/week for 13 weeks. Endpoints evaluated
included a functional observational battery, hematology and clinical chemistry, urinalysis, and
gross and microscopic morphology of tissues.
Consideration of the available dose-response data to determine an estimate of inhalation
exposure that is likely to be without an appreciable risk of adverse health effects over a lifetime
led to the selection of the 13-week subchronic inhalation bioassay in Sprague-Dawley rats (BRL,
1994) and increased incidence of alveolar epithelial hyperplasia of male and female rats as the
principal study and critical effect for deriving the RfC for cerium oxide. Based on the available
evidence the effects observed in the lungs and lymph nodes of male and female rats were
considered to be indicative of overwhelmed pulmonary clearance and potential key events in the
proposed mode of action. However, as described in Section 4.6.3.1, evidence for overwhelmed
pulmonary clearance as a mode of action for pulmonary and lymphoreticular system effects
observed with inhalation exposure to cerium oxide relies on data from other relatively insoluble
particles (such as titanium dioxide) due to the limited mechanistic data on cerium oxide. In the
absence of evidence indicating otherwise, these effects are assumed to represent cerium oxide-
induced toxicity.
EPA selected the BRL (1994) study to serve as the principal study. Alveolar epithelial
hyperplasia in the lungs was selected as the critical effect in the derivation of the inhalation RfC.
EPA chose alveolar epithelial hyperplasia in the lungs as the basis for the determination of the
point of departure because this effect represents a sensitive endpoint occurring at low doses that
is considered to be indicative of cerium oxide-induced toxicity. While it is recognized that
lymphoid hyperplasia in the bronchial lymph nodes occurs at a lower concentration, EPA
concluded that this effect may not represent a specific response to cerium oxide-induced toxicity.
Benchmark dose (BMD) modeling of the increased incidence of alveolar epithelial
hyperplasia in rats was conducted using the EPA's Benchmark Dose Software (BMDS, version
2) to determine the point of departure. The BMCLio of 8.7 mg/m3 from the multistage analysis
of the increased incidence of alveolar epithelial hyperplasia in female rats is the selected point of
departure. The human equivalent concentration (HEC) of 0.86 mg/m3 was calculated from the
point of departure by adjusting to a continuous exposure (24 hours a day, 7 days a week) and
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multiplying by a DAF (0.536), which, in this case, was the regional deposited dose ratio (RDDR)
for the pulmonary region of the lung. The RDDR was calculated using the RDDR v.2.3
program, as described in Methods for Derivation of Inhalation Reference Concentrations and
Application of Inhalation Dosimetry (U.S. EPA, 1994b).
The RfC of 9 x l(T4 mg/m3 was calculated from a BMCLHEc value of 0.86 mg/m3 for
increased incidence of alveolar epithelial hyperplasia in the lungs of male and female rats
following subchronic cerium oxide inhalation exposure (BRL, 1994). A total UF of 1,000 was
used: 3 for interspecies extrapolation, 10 for intraspecies variability, 10 for subchronic to chronic
extrapolation, and 3 for database deficiencies.
A factor of 3 was selected to account for uncertainties in extrapolating from rats to
humans, which is adopted by convention where an adjustment from an animal-specific BMCLADJ
to a BMCLHEc has been incorporated. Insufficient information is available to predict potential
variability in susceptibility among the population, thus a human variability UF of 10 was
applied. A 10-fold UF was used to account for uncertainty in extrapolating from a subchronic to
chronic exposure duration. Data gaps have been identified with uncertainties associated with
database deficiencies with regards to reproductive and developmental toxicity associated with
cerium inhalation exposure. A database UF of 3 was applied with special consideration of the
information pertaining to the deposition and absorption of cerium oxide, the effects observed in
humans following prolonged exposure, the mode of action data, and the effects observed in
animals in the BRL (1994) study, in addition to the lack of reproductive and developmental
studies.
The overall confidence in this RfC assessment is low. Confidence in the principal study
(BRL, 1994) is medium. EPA conducted an external peer review to evaluate the accuracy of the
experimental procedures, results, and interpretation and discussion of the results presented in this
study. The peer reviewers considered the BRL (1994) study conclusions to be supported by the
data. In addition, the results observed in the BRL (1994) study were consistent with the
observed effects in the human case reports (Yoon et al., 2005; Porru et al., 2001; McDonald et
al., 1995; Pairon et al., 1995, 1994; Sulotto et al., 1986; Vogt et al., 1986; Pietra et al., 1985;
Vocaturo et al., 1983; Sabbioni et al., 1982; Husain et al., 1980; Kappenberger and Buhlmann,
1975; Heuck and Hoschek, 1968). Confidence in the database is low. The database lacks
chronic exposure information on cerium via any route of exposure and multigenerational
developmental and reproductive toxicity studies. However, there is evidence of cerium
pneumoconiosis in humans exposed to cerium compounds, and the anticipated critical effects
observed are point-of-entry effects that would be expected in humans. Reflecting medium
confidence in the principal study and low confidence in the database, confidence in the RfC is
low.
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6.2.3. Cancer/Oral and Inhalation
Under the Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005a), the database
for cerium oxide and cerium compounds is inadequate to assess the human carcinogenic
potential and to calculate quantitative cancer risk estimates.
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APPENDIX A: SUMMARY OF EXTERNAL PEER REVIEW AND
PUBLIC COMMENTS AND DISPOSITION
The Toxicological Review of cerium oxide and cerium compounds has undergone a
formal external peer review performed by scientists in accordance with EPA guidance on peer
review (U.S. EPA, 2006a, 2000a). The external peer reviewers were tasked with providing
written answers to general questions on the overall assessment and on chemical-specific
questions in areas of scientific controversy or uncertainty. A summary of significant comments
made by the external reviewers and EPA's responses to these comments arranged by charge
question follow. In many cases the comments of the individual reviewers have been synthesized
and paraphrased in development of Appendix A. EPA also received scientific comments from
the public. These comments and EPA's responses are included in a separate section of this
appendix.
On April 10, 2008, EPA introduced revisions to the IRIS process for developing chemical
assessments. As part of the revised process, the disposition of peer reviewer and public
comments, as found in this Appendix, and the revised IRIS Toxicological Review was provided
to the external peer review panel members for an additional opportunity to comment on October
15, 2008. Information on the additional peer review is included at the end of this Appendix.
EXTERNAL PEER REVIEW PANEL COMMENTS
The reviewers made several editorial suggestions to clarify specific portions of the text.
These changes were incorporated in the document as appropriate and are not discussed further.
In addition, the external peer reviewers commented on decisions and analyses in the
Toxicological Review under multiple charge questions, and these comments were organized
under the most appropriate charge question.
A. General Comments
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: The reviewers concluded that the Toxicological Review was logical, clear,
and concise, and that EPA has accurately, clearly, and objectively represented and
synthesized the scientific information. The Toxicological review was characterized as
'repetitive' by some of the reviewers. One reviewer stated that the synthesis of the
scientific evidence for noncancer hazard appears to be more subjectively represented and
is in part more speculative than proven by the data.
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Response: EPA has reviewed and modified the document to address the reviewer
concerns about the repetitious nature of the document. The evidence provided by the
human case reports and the BRL (1994) study is sufficient to formulate a hazard
identification. The only available study reporting low-dose effects of subchronic
inhalation exposure to cerium oxide in animals, BRL (1994), was chosen as the principal
study. This study was well designed with three dose groups of 30 animals per group per
sex. Numerous tissues and endpoints were assessed, and methods and observed effects
were thoroughly reported. This study identified statistically significant dose-dependent
effects on the lungs and lymphoreticular system in both male and female rats. The
observed effects included increased lung weight; discoloration or pale areas, pale foci,
and uncollapsed parenchyma in the lungs; enlargement or pale discoloration of the
bronchial, mediastinal, and pancreatic lymph nodes; and dose-related alveolar epithelial
and lymphoid hyperplasia and pigment accumulation in the lungs and lymph nodes. The
lung and lymphoreticular system effects observed by BRL (1994) are consistent with
effects observed in humans, that were characterized by the accumulation of cerium
particles in the lungs and lymphoreticular system and histologic effects throughout the
lung. Thus, EPA selected this study as the principal study for the derivation of the RfC
and chose alveolar epithelial hyperplasia as the critical effect because it was determined
that this effect represents the most sensitive endpoint occurring at the lowest dose that
was indicative of toxicity.
2. Please identify any additional studies that should be considered in the assessment of the
noncancer and cancer health effects of cerium oxide and cerium compounds.
Comment: The following references demonstrating the hepatic toxicity of soluble CeCb
injected intravenously were recommended by a reviewer:
Hirano, S. and K.T. Suzuki. Exposure, metabolism, and toxicity of rare earths
and related compounds. Env. Hlth. Perspect. 104 Suppl. 1: 85-95, 1996
Snyder, F. Cress, E.A., Kyker, G.C. Liver lipid response to intravenous rare
earths in rats. J. Lipid Res. 1: 125- 1131, 1959
Response: The references cited in Hirano and Suzuki (1996) that address cerium-induced
liver toxicity are referenced in Section 4.4.2.3, Hepatic Effects, of the draft Toxicological
Review. Hirano and Suzuki (1996) was not added to the document because it is a review
and the references presented in the article are already included in the document. Snyder
et al. (1959) was added to Section 4.4.2.3, Hepatic Effects.
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Comment: The following references were recommended by a reviewer; the first
reference provides in vitro data for nano-sized CeC>2 and the second reference is a new
study of the ingestion of CeCb by mice.
EJ Park, J Choi, YK Park, K Park, "Oxidative stress induced by cerium oxide
nanoparticles in BEAS-2B cells." Toxicology 245 p 90-100, 2008
Kawagoe et al. J Trace Elem Med Bio (2008)
Response: Both references were added to the document. The Park et al. (2008) reference
was added to Section 4.5.3, In Vitro and Ex Vivo Nanotoxicology Studies. The Kawagoe
et al. (2008) reference was added to Section 4.3, Reproductive/Developmental Studies -
Oral and Inhalation and Section 4.4.1, Acute Toxicity Studies (Oral and Inhalation).
3. Please discuss research that you think would be likely to increase confidence in the database
for future assessments of cerium oxide.
Comment: Reviewers identified the following areas of research that would likely increase
the confidence in the database:
• Study with a longer timeframe than 13-weeks to provide clearer support for the
current choice for the "critical effect"
• Study using an exposure level in the range of 0-5 mg CeCVm3 to permit a
narrower point of departure to be generated
• A new investigation should include an analysis of pulmonary immunologic
endpoints and of cerium burdens in the various lymph nodes
• Improved animal study with concentrations beginning at the lower end of the
range of the BRL (1994) study and extend downward toward a plausible
environmental range
• Studies related to the biokinetics of cerium compounds, in particular with specific
and different particle sizes, and determine alveolar macrophage clearance
function in an effort to determine the cause of particle overload
• Animal bioassay of soluble forms of cerium to determine systemic absorption and
target organ effects, with evaluations of cardiotoxicity, hepatotoxicity, and renal
toxicity
• More comprehensive animal exposure study that uses ultrafine cerium oxide and
includes more sensitive toxicological endpoints
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Response: EPA agrees that the above research recommendations would improve future
hazard identifications of cerium compounds.
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: The relevance of the BRL (1994) study to environmental exposures is an area
of uncertainty that a reviewer highlighted as being inadequate.
Response: Text has been added to Section 5.3, Uncertainties in the Oral Reference Dose
(RfD) and the Inhalation Reference Concentration (RfC), addressing uncertainty
associated with the high exposure concentrations of the BRL study and the potential for
overestimating risk.
Comment: Another area of uncertainty that was inadequately discussed was the
importance of particle size-related differences in the biokinetics and potential effects,
especially for nano-cerium oxide.
Response: Text has been added to Section 5.3, Uncertainties in the Oral Reference Dose
(RfD) and the Inhalation Reference Concentration (RfC)., discussing the uncertainly of
the RfC and it's application to nano-sized cerium oxide. However, data are insufficient
to derive an RfC for nano-cerium oxide, and the RfC derived in the draft document is for
cerium oxide particles with an MMAD of approximately 2.0 um and a GSD of 1.8 to 1.9.
Comment: More discussion is also needed highlighting the differences between effects
elicited in the alveolar region of the lung and the conducting airways (tracheobronchial
region).
Response: Effects were observed in the alveolar region of the lung and in the bronchial
lymph nodes of rats exposed to cerium oxide aerosol for 13-weeks. The bronchial lymph
nodes, in addition to the mediastinal lymph nodes, are part of the lymphatic system that
collects particles from the lung as a whole. The exposure-related increase in pigment
accumulation and lymphoid hyperplasia in the lymph nodes draining the lung supports
the role of pulmonary macrophages in the clearance of particles deposited in the lung.
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The deposition of cerium oxide particles in the pulmonary region of the lung may lead to
pulmonary alveolar macrophage (PAM) activation and migration of PAMs containing
cerium oxide particles to the tracheobronchial lymph nodes.
Comment: One reviewer recommended that the discussion of the mode of action should
be limited to situations where exposure to insoluble cerium oxide is at high doses. In
addition, this reviewer suggested that the uncertainty for the derived RfC increases when
it is applied generally to cerium compounds.
Response: Text has been added to Section 5.3, Uncertainties in the Oral Reference Dose
and Reference Inhalation Concentration, stating that the proposed mode of action for
toxicity induced by lower dose chronic exposures to cerium oxide may not be related to
overloaded pulmonary clearance mechanisms, and the effects observed following low
dose chronic exposures may also be different.
The derived RfC, 9 x 1CT4 mg/m3, is not recommended for cerium compounds
other than cerium oxide. Text has been added to Section 5.3, Uncertainties in the Oral
Reference Dose (RfD) and the Inhalation Reference Concentration (RfC), addressing
uncertainty associated with the application of the derived RfC to cerium compounds
other than cerium oxide.
B. Oral Reference Dose (RfD) for cerium oxide and cerium compounds
1. A chronic RfD for cerium compounds has not been derived. Has the scientific justification
for not deriving an RfD been transparently and objectively described? Please identify and
provide the rationale for any studies that should be selected as the principal study. Please
identify and provide the rationale for any endpoints that should be considered in the selection of
the critical effect.
Comment: The reviewers commented that the rationale for not deriving a RfD provided
in the document was appropriate.
Response: No response.
Comment: The reviewers did not identify any studies for consideration as the primary
study or endpoints for selection as the critical effect. One reviewer commented that the
available studies indicate potential target organs (cardiac and hepatic) for toxicity
following oral exposure to soluble cerium compounds.
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Response: While there may be potential for cardiac effects following oral exposure, the
cardiac effects were observed in conjunction with a magnesium-restricted diet. An
association between exposure to rare earth elements, cerium in particular, in food and the
development of endomyocardial fibrosis in humans has been suggested (Eapen, 1998;
Kutty et al., 1996; Valiathan et al., 1989); although, analysis of the geographic
distribution of endemic endomyocardial fibrosis in India suggested a link to high cerium
soil concentrations and possibly to magnesium deficiency during childhood (Kutty et al.,
1996).
Karma et al. (1998) suggested that cerium chloride may intensify the adverse
cardiac effects of magnesium deficiency; however, the authors reported that cerium may
intensify the cardiotoxicity associated with a magnesium-deficient or restricted diet but
did not elicit a cardiac effect when tested under conditions of a normal magnesium diet.
Parenteral administration of cerium to rodents caused lipid deposition,
mitochondrial damage, and invaginations of the nuclear membrane in hepatocytes, as
well as morphological changes in the liver, characterized by fatty liver, fatty
degeneration, and necrosis. Unfortunately, these effects were not observed following
oral administration of cerium chloride and do not inform the oral hazard identification
analysis.
C. Inhalation Reference Concentration (RfC) for cerium oxide and cerium compounds
1. A chronic RfC for cerium oxide has been derived from the 13 week inhalation study (BRL,
1994) in rats. 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? Are the criteria and rationale for the selection of this study 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.
Comment: One reviewer commented that the study was not adequately justified because
the lowest exposure concentration used in the study was not environmentally relevant,
and is a concentration that would predictably cause nonspecific effects due to particle
overload. In addition, the reviewer stated that existing knowledge would not support the
notion that 2 jim diameter CeC>2 as relevant to the most likely target form of cerium
oxide.
Response: Text was added to Section 5.3, Uncertainties in the Oral Reference Dose
-------
(RfD) cmd the Inhalation Reference Concentration (RfC), of the document addressing the
utilization of high exposure concentrations in the BRL (1994) study and the possibility
that the utilization of the BRL study may overestimate the risk to cerium oxide particles.
It should be noted that while the low dose in the BRL (1994) study was
considered high in comparison to environmental concentrations, it should not be assumed
to predictably cause nonspecific effects due to pulmonary overload. For example,
BaSO4, at higher exposure levels than those of the BRL (1994) study, did not elicit a
nonspecific overload response, whereas, pulmonary overload was demonstrated by TiCh
at similar exposures but higher particle surface area. It was hypothesized that the particle
surface area of BaSC>4 (BaSC>4 has a low specific surface area) influences the level of
inflammation (Tran et al., 2000). The data for TiO2 and BaSC>4 suggests that a high
particle exposure level does not necessarily lead to a pulmonary overload response. In
the case of cerium oxide, the exposure was such that pulmonary clearance was
overloaded.
The derived RfC is relevant to 2 jim diameter CeO2 particles and this is specified
in Section 5.2.3, RfC Derivation-Including Application of Uncertainty Factors. Data are
currently unavailable to better evaluate the hazard identification and dose-response of
nano-sized cerium oxide.
2. Increased incidence of lymphoid hyperplasia in the bronchial lymph nodes of male rats was
selected as the critical toxicological effect. The selection of increased incidence of lymphoid
hyperplasia in the bronchial lymph nodes as the critical effect for cerium oxide is because it is
considered by EPA to be a precursor to an adverse effect. Please comment on whether the
selection of this critical effect has been scientifically justified. Are the criteria and rationale for
this selection transparently and objectively described in the document? Please provide a detailed
explanation. Please comment on whether EPA's rationale about the adversity of the critical
effect has been adequately and transparently described and is supported by the available data.
Please identify and provide the rationale for any other endpoints that should be used instead of
lymphoid hyperplasia to develop the RfC.
Comment: Four of the five reviewers disagreed with the selection of lymphoid
hyperplasia in the bronchial lymph nodes as the critical effect and recommended using
the alveolar hyperplasia as the critical effect. One reviewer stated that it was only weakly
sufficient to claim that the lymphoid hyperplasia in the bronchial lymph nodes as the
"critical" effect, while another questioned whether the data supported the conclusion that
lymphoid hyperplasia in bronchial lymph nodes was a precursor effect. Several
reviewers questioned the proposed mode of action for the critical effect, stating that
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lymphoid hyperplasia is not known to be on the pathogenic pathway to more clearly
adverse effects and may not be adverse itself. The reviewers commented that the alveolar
epithelial hyperplasia in the lungs may be relevant to progressive adverse effects.
Response: Based on the recommendations of the external peer review panel, EPA has
reconsidered the BRL (1994) study data and has selected alveolar hyperplasia in the
lungs as the critical effect to determine the point of departure for the derivation of the
RfC. Specifically, EPA selected alveolar epithelial hyperplasia in the lungs as the basis
for the determination of the point of departure because this effect likely represents a
sensitive endpoint occurring at low doses that is indicative of cerium oxide-induced
toxicity. While it is recognized that lymphoid hyperplasia in the bronchial lymph nodes
occurs at a lower concentration, EPA concluded that this effect may not represent a
specific response to cerium oxide-induced toxicity.
3. Some mode of action evidence exists suggesting that lymphoid hyperplasia in the bronchial
lymph nodes represents a sensitive endpoint that occurs early in a series of critical events leading
to more severe effects in the lung. Specifically, the data suggest that lymphoid hyperplasia in the
bronchial lymph nodes may represent the point at which normal clearance of particles from the
lung by alveolar macrophages becomes overwhelmed and particles are no longer cleared
effectively. This delayed clearance leads to increased accumulation of cerium oxide particles in
the respiratory tract, an inflammatory response, and subsequent cell proliferation. Please
comment on whether the available mode of action data supports this proposed mode of action for
cerium oxide-induced bronchial lymphoid hyperplasia. Is this proposed mode of action
scientifically justified and transparently and objectively described?
Comment: One reviewer commented that the mode of action is not justified based on the
sole evidence provided in the BRL (1994) study. Specifically, the hyperplasia may be
suggestive of problems with the clearance of the particles out of the lymph nodes and not
necessarily the airways. Another reviewer commented that the BRL study does not give
any evidence of impaired clearance nor does it provide evidence of other key results
related to dust overload, such as type II cell proliferation, altered alveolar epithelial
integrity, and/or chronic acute inflammation with an influx of PMNs, and recommended
the addition of an alternative mode of action to the discussion. The alternative mode of
action proposed included an immune response due to the cerium oxide particles delivered
by dendritic cells from the tracheobronchial region to the bronchial lymph nodes. A third
reviewer stated that it is not certain that hyperplasia in lung-associated lymph nodes is a
direct result of overloading exposures. A reviewer commented that chronic exposure to
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lower concentrations could cause effects via a different mode of action. Finally, a
reviewer also expressed concern that the lymphoid hyperplasia may be a non-specific
response to particles and not strictly an effect of cerium oxide.
Response: The proposed mode of action analysis, while including a discussion of the
results from the BRL (1994) study, also incorporated pulmonary overload and particle
clearance studies, toxicity reviews, in vitro studies, intratracheal instillation studies, and
the pulmonary effects observed in human studies to propose a potential mode of action
for cerium oxide toxicity. The mode of action analysis included a discussion of the
concept of dust overload in the lung following inhalation exposure, an example of
overwhelmed pulmonary clearance following exposure to titanium dioxide, the
comparison of symptoms of pulmonary overload with effects observed in the BRL (1994)
study, and an analysis of cellular responses to insoluble particles and the relationship to
pulmonary overload. EPA recognizes that the proposed mode of action is based, in part,
on data for other insoluble particles (titanium dioxide for example), and that there is
uncertainty for the mode of action due to the lack of comprehensive data from cerium
oxide on some aspects of the mode of action. Text was added to Section 5.3,
Uncertainties in the Oral Reference Dose and Inhalation Reference Concentration.,
discussing the absence of key pulmonary overload data.
Chronic exposure to lower doses of cerium oxide may cause effects via a different
mode of action; however, data and/or information on an alternative mode of action at
lower doses are unavailable. Text was added to Section 5.3, Uncertainties in the Oral
Reference Dose and Inhalation Reference Concentration, stating that lower doses of
cerium oxide may act through a different mode of action.
The delivery of cerium oxide particles to bronchial lymph nodes from the
tracheobronchial region of the lung by dendritic cells is a proposed alternative mode of
action. Text has been added to Section 4.6.3.1 Respiratory Tissues addressing this
potential alternative mode of action.
However, based on the recommendations of the external peer review panel, EPA
has selected alveolar hyperplasia as the critical effect and the text in Section 5.2.1,
Choice of Principal Study and Critical Effect—with Rationale and Justification has been
changed accordingly. .
The proposed mode of action for cerium oxide may, in fact, be a non-specific
response to cerium oxide in the lung, i.e. the physical presence of cerium oxide in the
lung leading to pulmonary toxicity. The available data support the proposed mode of
action of overloading of PAMs by cerium oxide particles, leading to the release of
inflammatory cytokines and fibrogenic growth factors and subsequent cell damage,
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observed as alveolar epithelial hyperplasia in the lung. However, the mode of action for
pulmonary toxicity in humans following chronic inhalation of cerium oxide is not known.
4. The chronic RfC has been derived utilizing the NOAEL/LOAEL approach to define the point
of departure. Please provide comments with regards to whether this is the best approach for
determining the point of departure. Please identify and provide rationale for any alternative
approaches for the determination of the point of departure, and if such approaches are preferred
to EPA's approach.
Comment: Four of the reviewers stated that the use of the NOAEL/LOAEL approach to
define the point of departure was a reasonable selection and that this decision was
adequately justified. Several reviewers stated that data for alveolar epithelial hyperplasia
should be selected as the critical effect for determination of the point of departure using
benchmark dose modeling.
Response: EPA has selected alveolar hyperplasia as the critical effect as indicated above.
Benchmark dose modeling was used to derive the point of departure.
5. 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: One reviewer commented that the database uncertainty factor (UFD) should
consider and address the issue of nano-cerium oxide, which may give rise to systemic
effects that could not be observed with the larger particle size used in the BRL (1994)
study.
Response: The database uncertainty factor is applied to the point of departure for cerium
oxide particles with an MMAD of approximately 2.0 um and a GSD of 1.8 to 1.9. It
would not be appropriate to apply a database UF to this point of departure for lack of data
for another particle size. Text was added to Section 5.3 Uncertainties in the oral
reference dose (RfD) and inhalation reference concentration (RfC) was added to address
the lack of nano-particle cerium oxide data as an uncertainty in the assessment.
Comment: One reviewer commented that the uncertainty factors are adjustments for an
outcome that is probably not relevant to environmental exposures. Additionally, a
reviewer commented that the Toxicological Review has taken a response to a high
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exposure level, applied a large combined uncertainty factor, and ended up with a
reference concentration comparable to the values for better studied materials with well-
demonstrated toxicity.
Response: Please see first comment and response under A.4. for a discussion of the
hazard identification and relevancy to environmental exposures.
In this draft document, uncertainty factors have been applied to a point of
departure for the critical effect from the principal study to derive a reference
concentration for cerium oxide.
Comment: A reviewer recommended including a discussion of magnesium deficiency
and susceptibility to cardiotoxic effects of cerium. The document would be strengthened
by referring to these studies as a potential mechanism causing increased risk within
human subpopulations.
Response: The studies suggesting magnesium deficiency might increase susceptibility to
cardiotoxic effects of cerium chloride were not included in the discussion of the
intraspecies uncertainty factor for cerium oxide. The studies that demonstrated that
magnesium deficiency might increase the susceptibility to cardiotoxic effects of soluble
cerium chloride exposure are oral studies, and the qualitative extrapolation of these
results to the derivation of the inhalation RfC is not supported. The cerium compound in
the oral studies is cerium chloride, a soluble cerium compound, while the RfC was
derived for cerium oxide, an insoluble cerium compound. The difference in solubility
may have a significant effect on the toxicokinetics and toxicodynamics of the cerium
compound, and qualitatively extrapolating the potential for cardiac effects from the oral
studies of cerium chloride to the portal-of-entry effects following cerium oxide exposure
via inhalation is not appropriate. Text has also been added to Section 4.8.3, Possible
Susceptible Populations, highlighting magnesium deficiency and susceptibility to cardiac
effects.
6. Please comment on the transparency, scientific rationale and justification for the LOAEL-to-
NOAEL uncertainty factor of 3. Are the criteria and rationale for this selection transparently and
objectively described in the document? The point of departure for this analysis was based on the
critical effect of lymphoid hyperplasia in the bronchial lymph nodes. This effect is described as a
sensitive effect occurring early in the series of critical events leading to more severe effects in
the lung, and hence a default 10-fold uncertainty factor was not applied. The mode of action for
the critical effect is thought to be related to pulmonary clearance overload, in which normal
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clearance of particles from the lung by alveolar macrophages becomes overwhelmed and
particles are no longer cleared effectively, leading to an increasing accumulation of particles in
the lung and airways, an inflammatory response, and subsequent cell proliferation. Please
comment on whether the justification for selection of the LOAEL-to-NOAEL uncertainty factor
based on these data is scientifically justified and transparently described.
Comment: One reviewer commented that the underlying premise for the choice of the
point of departure, and a UFL of 3, was insufficiently justified based on the critical effect
selected and mode of action proposed. One reviewer commented that because the
lymphoid hyperplasia in the bronchial lymph nodes does not show a dose response in the
BRL (1994) study, a UFL of 10 should also be considered.
Response: Note that the critical effect has been changed to alveolar hyperplasia as
indicated above. The BRL (1994) study and human case reports support the conclusion
that alveolar epithelial hyperplasia likely represents the most sensitive effect that may
occur early in a series of critical events leading to more severe effects in the lung. The
critical effect along with other effects observed in the lungs and lymph nodes may be
representive of disruption of the normal clearance of particles from the lung by alveolar
macrophages. Specifically, alveolar macrophages become overwhelmed and particles are
no longer cleared effectively. Delayed clearance could possibly lead to increased
accumulation of cerium oxide particles in the respiratory tract, an inflammatory response,
and subsequent cell proliferation. A factor of 3 was used to account for uncertainty in
extrapolating from a LOAEL to a NOAEL (UFL) because the point of departure was a
LOAEL. The critical effect selected to determine the point of departure for this analysis,
was considered to be minimally biologically significant; therefore, an uncertainty factor
of 3 was applied.
Comment: One reviewer was not sure that the lymphoid hyperplasia represents the point
at which normal clearance of particles from the lung by macrophages has become
overwhelmed. The reviewer proposed an alternative mode of action in which an immune
response is initiated by cerium-oxide particles translocated to the lymph node by
dendtritic cells. The LOAEL-to-NOAEL uncertainty factor can be avoided by using
alveolar epithelial hyperplasia as the critical effect and benchmark dose modeling.
Response: Please see responses above regarding the choice of the critical effect.
7. Please comment on the transparency, scientific rationale and justification for the selection of
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the database uncertainty factor. Please comment on whether the application of the database
uncertainty factor adequately addresses the lack toxicity data for cerium oxide. Specifically,
please comment on whether studies addressing additional endpoints of concern (e.g.
reproductive and developmental toxicity studies) would likely result in a lower point of
departure. Are the criteria and rationale for this selection transparently and objectively described
in the document? An uncertainty factor of 3 was applied with special consideration of the
information pertaining to the deposition and absorption of cerium oxide, the effects observed in
humans following prolonged exposure, the mode of action data, and the similar effects observed
in animals in the principal study.
Comment: One reviewer commented that a lower point of departure might be developed
if studies assessing immunologic endpoints in the lung were available.
Response: While it is possible that additional immunologic studies may lead to a lower
point of departure, there are no data to inform this assumption. No changes have been
made in response to the comment.
Comment: One reviewer suggested EPA consider different particle sizes of cerium oxide
rather than just focusing on what size happened to be used in the BRL (1994) study. For
example, the reviewer recommended modeling the deposition of inhaled cerium oxide
particles at different sizes by using the MPPD deposition model and assuming normal
clearance in rats and humans.
Response: While toxicity resulting from particle overload in the lungs may be expected
with other micro-sized cerium oxide particles, the toxicity and mode of action of nano-
sized particles is unknown. Extrapolating the toxicity resulting from exposure to micro-
sized cerium oxide to nano-sized cerium oxide exposures would likely impart significant
uncertainty. It was not undertaken as a part of the assessment.
Comment: One reviewer commented that the lack of a lifetime exposure/recovery study
in a second animal species with a longer lifetime is a major shortcoming since 1)
progressive effects (such as fibrosis and lung cancer) cannot be observed in from short
term studies, and 2) it's unclear whether the particles in the lung and/or lymph nodes
slowly release cerium systemically, possibly causing cardiotoxicity over time.
Response: The lack of a lifetime exposure/recovery study is a data gap in this
assessment. In considering this data gap, an uncertainty factor of 10 was used to account
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for the extrapolation from a subchronic to chronic exposure duration. The lack of a
chronic study is also addressed in Section 5.3, Uncertainties in the Oral Reference Dose
(RfD) and the Inhalation Reference Concentration (RfC).
Cerium as an element is a strong oxidizing agent and is stabilized when
associated with an oxygen ligand (Kilbourn, 2003). Text was added to Section 5.3,
Uncertainties in the Oral Reference Dose (RfD) and the Inhalation Reference
Concentration (RfC) presenting the possibility for systemic effects, including
cardiotoxicity, following inhalation absorption.
8. The RfC has been derived using data from inhalation exposure to cerium oxide (BRL, 1994).
Is the statement to not use the RfC for cerium compounds other than cerium oxide scientifically
justified? Is there enough information on and discussion of cerium compounds to warrant the
title "cerium oxide and cerium compounds?"
Comment: One reviewer commented that the end of the document needs additional
discussion of cerium oxide as the most critical compound of exposure (given the
solubility of cerium salts) because of the long retention as a particle, provided low
inhaled concentrations are present.
Response: Text was added in Section 6.1, Human Hazard Potential., to highlight cerium
oxide exposures as the most relevant of the cerium compounds.
Comment: One reviewer commented that statements in the text drawing conclusions or
making generalizations should specifically state the cerium compound being discussed or
reference the solubility of the cerium.
Response: The document has been revised, where feasible, in order to specifically state
the cerium compound being discussed by each reference article.
D. Carcinogenicity of cerium oxide and cerium compounds
1. Under the EPA's 2005 Guidelines for carcinogen risk assessment (www.epa.gov/iris/backgr-
d.htm), there is "inadequate information to assess the carcinogenic potential" of cerium
compounds. Please comment on the scientific justification for the cancer weight of the evidence
characterization. Has the scientific justification for the weight of evidence characterization been
sufficiently, transparently, and objectively described? Has the scientific justification for not
deriving a quantitative risk estimate been transparently and objectively described?
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Comment: The reviewers agreed with the conclusion that data were inadequate to assess
carcinogenic potential of cerium oxide.
PUBLIC COMMENTS
A. Inhalation Reference Concentration (RfC) for cerium oxide and cerium compounds
Comment: A public submission provided three references addressing nanoparticle cerium
oxide:
• Park et al., Particle and Fibre Toxicology 4:12, 1 (2007)
• Park et al., Inhalation Toxicology 20:6, 547 (2008)
• Fall et al., Nanotoxicology 1(3):227 (2007)
Response: The Park et al. (2007) and (2008) and Fall et al. (2007) references were added to
Section 4.5.3, Nanotoxicology Studies of the document.
Comment: A public submission stated that it would be more logical to determine hazard and
dose response specific to cerium oxide separately from that of other cerium compounds. The
inclusion of all cerium compounds in the review is misleading.
Response: The last paragraph in Section 5.2.3, RfC Derivation—Including Application of
Uncertainty Factors (UFs), includes language specifying that the RfC was quantified for
cerium oxide particles, and the use of the RfC for cerium compounds other than cerium oxide
is not recommended as the similarity between this form of cerium and other cerium
compounds is unknown. The data on additional cerium compounds are presented as
additional information for the reader. However, these data are sparse and are not suitable for
the derivation of toxicity values for other specific cerium compounds.
Comment: A public submission commented that the results presented in Section 4.1.2 are
from high exposures and likely have little relevance for low environmental exposures. In
addition, the submission commented that the doses used in the BRL (1994) study cannot be
scientifically defended as of being or relevance to human exposure.
Response: The human case reports demonstrated the development of pneumoconiosis or
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interstitial lung disease associated with the accumulation of cerium particles in the lungs
after prolonged occupational exposure to cerium fumes or dust (Yoon et al., 2005; Porru et
al., 2001; McDonald et al., 1995; Pairon et al., 1995, 1994; Sulotto et al., 1986; Vogt et al.,
1986; Pietra et al., 1985; Vocaturo et al., 1983; Sabbioni et al., 1982; Husain et al., 1980;
Kappenberger and Biihlmann, 1975; Heuck and Hoschek, 1968). However, the available
human studies were not selected for the derivation of an inhalation RfC because the cerium
exposures were not available in any of the case reports.
Text has been added to Section 5.3, Uncertainties in the Oral Reference Dose (RfD)
and the Inhalation Reference Concentration (RfC), addressing uncertainty associated with the
high exposure concentrations of the BRL study and the potential for overestimating risk.
ADDITIONAL EXTERNAL PEER REVIEW PANEL COMMENTS IN RESPONSE TO
REVISIONS
Based on the comments received from the panel,, the revised assessment was
provided to the external peer review panel for additional opportunity to comment. The
comments focused on the selection of alveolar epithelial hyperplasia in the lungs as the
critical effect for the derivation of the RfC as opposed to lymphoid hyperplasia. The
responses are subsumed into the comments above to indicate that alveolar hyperplasia was
chosen as the critical effect.
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APPENDIX B: BENCHMARK DOSE MODELING RESULTS FOR THE DERIVATION
OF THE RFC
Alveolar hyperplasia in the lung was selected as the critical effect and benchmark dose
(BMD) modeling was conducted using the EPA's Benchmark Dose Software (BMDS, version
2.. 13) to identify the point of departure for the derivation of an RfC for cerium oxide. The
modeling was conducted in accordance with the draft EPA guidelines (U.S. EPA, 2000b) using
Benchmark Dose Software Version 2.13. The BMD modeling results for the derivation of the
chronic RfC are summarized in Table B-l. The model output results for all of the models per
endpoint in male and female rats are presented in the corresponding tables. A brief discussion of
the modeling results is presented below.
The increased incidence of alveolar epithelial hyperplasia in male and female rats was
modeled, with a benchmark response (BMR) of 10%. Adequacy of each model fit was
determined by assessing the chi-square goodness-of-fit statistic using a significance level of a =
0.1. The best-fitting models were selected from those exhibiting adequate fit by considering the
Akaike Information Criterion (AIC) value of each model.
In male rats, the log-logistic model, with a BMCLio of 1.7 mg/m3, adequately fit the
alveolar epithelial hyperplasia incidence data. In female rats, the multistage model, with a
BMCLio of 8.7 mg/m3, and the Quantal-linear, with aBMCLio of 7.2 mg/m3, model adequately
fit the incidence data. For the computation of the point of departure, the multistage model
analysis of the incidence of alveolar epithelial hyperplasia in female rats was selected. The
multistage model and resulting BMCLio were selected because the multistage model provided
the lowest AIC.
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Table B-l. Benchmark Dose modeling used in the derivation of the RfC
Sex
male
female
Model
Gamma
Logistic
Log-logistic
Log-Probit
Multistage
Probit
Weibull
Quantal-linear
Gamma
Logistic
Log-logistic
Log-Probit
Multistage
Probit
Weibull
Quantal-linear
Goodness of
fit p-value
0.0000
0.0001
0.5278
0.0001
0.0000
0.0000
0.0000
0.0000
1.0000
1.0000
1.0000
1.0000
0.9962
1.0000
1.0000
0.8329
X2 p-value
29.95
19.67
1.28
18.19
29.95
20.13
29.95
29.95
0.00
0.00
0.00
0.00
0.06
0.00
0.00
0.87
AIC
43.8
58.2
37.2
41.2
43.8
58.5
43.8
43.8
23.1
23.1
23.1
23.1
21.2
23.1
23.1
22.7
BMD10
(mg/m3)
8.9
42.7
4.9
9.5
8.9
48.1
8.9
8.9
37.7
46.7
42.3
36.6
25.9
43.1
38.2
12.5
BMDL10
(mg/m3)
5.8
23.6
1.7
5.8
5.8
32.4
5.8
5.8
8.9
24.6
13.4
13.4
8.7
22.1
8.9
7.2
Logistic Model. (Version: 2.12; Date: 05/16/2008)
Input Data File: C:\USEPA\BMDS2\Temp\tmpl79.(d)
Gnuplot Plotting File: C:\USEPA\BMDS2\Temp\tmpl79.plt
Tue Aug 12 14:33:37 2008
BMDS Model Run
The form of the probability function is:
P [response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose)
Dependent variable = alv_epi_hyper
Independent variable = DOSE
Slope parameter is restricted as slope >= 1
Total number of observations = 4
Total number of records with missing values = 0
Maximum number of iterations = 250
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = -4.14211
slope = 1.14259
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by the user,
and do not appear in the correlation matrix )
intercept
slope
100
-------
intercept 1 -0.94
slope -0.94 1
Parameter Estimates
95.0% Wald Confidence Interval
Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit
background 0 * * *
intercept -4.12525 * * *
slope 1.2208 * * *
* - Indicates that this value is not calculated.
Analysis of Deviance Table
Model Log(likelihood) # Param's Deviance Test d.f. P-value
Full model -16.0466 4
Fitted model -16.6185 2 1.14373 2 0.5645
Reduced model -41.0539 1 50.0146 3 <.0001
AIC: 37.237
Goodness of Fit
Dose
0 .0000
5 .0000
50.5000
507.5000
Est. Prob.
0. 0000
0. 1034
0.6599
0.9701
Expected
0 .000
1 .550
9.898
14 .552
Observed
0 .000
1 .000
11.000
14 .000
Size
15
15
15
15
Scaled
Residual
0. 000
-0.467
0.601
-0.836
Chi*2 = 1.28 d.f. = 2 P-value = 0.5278
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 4.8516
BMDL = 1.73808
101
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Log-Logistic Model with 0.95 Confidence Level
T3
-------
Multistage Model. (Version: 3.0; Date: 05/16/2008)
Input Data File: C:\USEPA\BMDS2\Temp\tmpl97.(d)
Gnuplot Plotting File: C:\USEPA\BMDS2\Temp\tmpl97.plt
Tue Aug 12 14:55:57 2008
BMDS Model Run
The form of the probability function is:
P [response] = background + (1-background)*[1-EXP(
-betal*dose*l-beta2*dose*2)]
The parameter betas are restricted to be positive
Dependent variable = alv_epi_hyper
Independent variable = DOSE
Total number of observations = 4
Total number of records with missing values = 0
Total number of parameters in model = 3
Total number of specified parameters = 0
Degree of polynomial = 2
Maximum number of iterations = 250
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
Default Initial Parameter Values
Background = 0
Beta(l) = 0
Beta(2) = 3.89529e+014
Beta (2)
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -Background -Beta(l)
have been estimated at a boundary point, or have been specified by the user,
and do not appear in the correlation matrix )
Beta(2)
1
Variable
Background
Beta(l)
Beta(2)
Estimate
0
0
0.000157092
Parameter Estimates
Std. Err.
95.0% Wald Confidence Interval
Lower Conf. Limit Upper Conf. Limit
* - Indicates that this value is not calculated.
Model
Full model
Fitted model
Reduced model
Analysis of Deviance Table
Log(likelihood)
-9.54771
-9.60698
-38.1909
# Param's
4
1
1
Deviance Test d.f.
0.118528
57.2863
P-value
0.9895
<.0001
AIC:
21.214
Goodness of Fit
Dose
0 .0000
5.0000
50.5000
Est. Prob.
0. 0000
0.0039
0.3301
Expected
0 .000
0.059
4 .951
Observed
0 .000
0.000
5.000
Size
15
15
15
Scaled
Residual
0. 000
-0.243
0.027
103
-------
507.5000 1.0000 15.000 15.000 15
Chi*2 = 0.06 d.f. = 3 P-value = 0.9962
0.000
Benchmark Dose Computation
Specified effect =
Risk Type
Confidence level =
BMD =
BMDL =
BMDU =
0 .1
Extra risk
0. 95
25.8977
8.71717
39.45
Taken together, (8.71717, 39.45 ) is a 90
interval for the BMD
% two-sided confidence
Multistage Model with 0.95 Confidence Level
I
c
o
'
0.8
0.6
0.4
0.2
Multistage
BMD Lower Bound
100
200 300
dose
400
500
14:55 08/12 2008
104
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Quantal Linear Model using Weibull Model (Version: 2.12; Date: 05/16/2008)
Input Data File: C:\USEPA\BMDS2\Temp\tmpl9A.(d)
Gnuplot Plotting File: C:\USEPA\BMDS2\Temp\tmpl9A.plt
Tue Aug 12 15:00:02 2008
BMDS Model Run
The form of the probability function is:
P [response] = background + (1-background)*[1-EXP(-slope*dose)]
Dependent variable = alv_epi_hyper
Independent variable = DOSE
Total number of observations = 4
Total number of records with missing values = 0
Maximum number of iterations = 250
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
Default Initial (and Specified) Parameter Values
Background = 0.03125
Slope = 0.00676648
Power = 1 Specified
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -Background -Power
have been estimated at a boundary point, or have been specified by the user,
and do not appear in the correlation matrix )
Slope
Slope 1
Parameter Estimates
Variable
Background
Slope
Estimate
0
0.008453
Std. Err.
NA
0.0028472
95.0% Wald Confidence Interval
Lower Conf. Limit Upper Conf. Limit
0.00287259
0.0140334
NA - Indicates that this parameter has hit a bound
implied by some inequality constraint and thus
has no standard error.
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-9.54771
-10.3953
-38.1909
# Param's
4
1
1
Deviance Test d.f.
P-value
1.69527
57.2863
0.638
<.0001
AIC:
22 . 7907
Goodness of Fit
Dose
0 .0000
5 .0000
50.5000
507.5000
Est. Prob.
0. 0000
0. 0414
0.3475
0.9863
Expected
0 .000
0 .621
5.212
14 .794
Observed
0 .000
0 .000
5.000
15.000
Size
15
15
15
15
Scaled
Residual
0. 000
-0. 805
-0.115
0.457
Chi*2 = 0.87
d.f. = 3
P-value = 0.8329
Benchmark Dose Computation
105
-------
Specified effect
Risk Type
Confidence level
BMD
BMDL
I
C
o
=8
ro
0.8
0.6
0.4
0.2
0.1
Extra risk
0.95
12.4643
7.19255
Quantal Linear Model with 0.95 Confidence Level
Quantal Linear
BMD Lower Bound
liMDLlJB
MD
100
200 300
dose
400
500
15:00 08/122008
106
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