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# TOXICOLOGICAL REVIEW

OF

THALLIUM AND COMPOUNDS

(CAS No. 7440-28-0)



## In Support of Summary Information on the Integrated Risk Inform ation System (IRIS)

September 2009

U.S. Environmental Protection Agency Washington, DCDISCLAIMER

This document has been reviewed in acco rdance with U.S. Environmental Protection Agency policy and approved for publication. Men tion of trade names or commercial products
does not constitute endorsement or recommendation for use.

iiCONTENTS—TOXICOLOGICAL REVIEW OF THALLIUM AND COMPOUNDS (CAS No. 7440-28-0)

LIST OF TABLES .......................................................................................................................... v LIST OF FIGURES ...................................................................................................................... vii LIST OF ABBREVIAT IONS AND ACRONYMS .................................................................... viii FOREWORD

.............................................................................................................................. ... ix AUTHORS, CONTRIBUTORS, AND REVIEWERS .................................................................. x

1. INTRODUCTION ..................................................................................................................... 1

2. CHEMICAL AND PHYSICAL INFORMATION ................................................................... 3

3. TOXICOKINETICS .................................................................................................................. 5 3.1. ABSORPTION .................................................................................................................. 5 3.2. DISTRIBUTION ............................................................................................................... 5 3.3. METABOLISM ................................................................................................................. 6 3.4. ELIMINATION ................................................................................................................. 6 3.5. PHYSIOLOGICALLY BASE D TOXICOKINETIC MODELS ...................................... 8

4. HAZARD IDENTIFICATION .................................................................................................. 9 4.1. STUDIES IN HUMANS—EPIDEMIOLOGY, CASE REPORTS, CLINICAL CONTROLS ...................................................................................................................... 9 4.1.1. Incident/Case Reports ............................................................................................. 9 4.1.2. Population Surveys ............................................................................................... 16 4.1.3. Occupational Exposure ......................................................................................... 17 4.2. LESS-THAN-LIFETIME AND CHRONIC STUDIES AND CANCER BIOASSAYS IN ANIMALS—ORAL AND INHALATION ........................................ 18 4.2.1. Oral Exposure ....................................................................................................... 18 4.2.2. Inhalation Exposure .............................................................................................. 26 4.3. REPRODUCTIVE/DEVELOPMENTAL STUDIES ..................................................... 26 4.3.1. Reproductive Toxicity .......................................................................................... 26 4.3.2. Developmental Toxicity ....................................................................................... 31 4.4. OTHER ENDPOINT-SPECIFIC STUDIES ................................................................... 32 4.4.1. Liver and Kidney Toxicity ................................................................................... 33 4.4.2. Cardiotoxicity ....................................................................................................... 40 4.4.3. Neurotoxicity ........................................................................................................ 40 4.5. MECHANISTIC DATA AND OTHE R STUDIES IN SUPPORT OF THE MODE OF ACTION ....................................................................................................... 45 4.5.1. Interference with Potassium Transport ................................................................. 45 4.5.2. Disturbance of Mitochondrial Function and Energy Generation ......................... 46 4.5.3. Induction of Oxidative Stress ............................................................................... 46 4.5.4. Reaction with Thiol Groups ................................................................................. 47 4.5.5. Other Endpoint-specific Mechanistic Data .......................................................... 47 4.5.6. Genotoxicity ......................................................................................................... 50 4.6. SYNTHESIS OF MAJO R NONCANCER EFFECTS ................................................... 51 4.6.1. Oral ....................................................................................................................... 51 4.6.2. Inhalation .............................................................................................................. 54 4.6.3. Mode-of-Action Information ................................................................................ 54
iii4.7. EVALUATION OF CARCINOGENICITY ................................................................... 55 4.8. SUSCEPTIBLE POPULATIONS AND LIFE STAGES ................................................ 56 4.8.1. Possible Childhood Susceptibility ........................................................................ 56 4.8.2. Possible Gender Differences ................................................................................ 56

5. DOSE-RESPONSE ASSESSMENTS ..................................................................................... 57 5.1. ORAL REFERENCE DOSE (RfD) ................................................................................ 57 5.1.1. Choice of Principal Study and Critical Effect—with Rationale and Justification .......................................................................................................... 57 5.1.2. Methods of Analysis ............................................................................................. 60 5.1.3. RfD Derivation—Inclu ding Application of Uncer tainty Factors (UFs) ............... 63 5.1.4. Candidate RfD Comparison Information ............................................................. 67 5.1.5. Previous RfD Assessment .................................................................................... 75 5.2. INHALATION REFERENCE CONCENTRATION (RfC) ........................................... 75 5.3. CANCER ASSESSMENT .............................................................................................. 75

6. MAJOR CONCLUSIONS IN THE CHARACTERIZATION OF HAZARD AND DOSE RESPONSE ...................................................................................................................... ....... 76 6.1. HUMAN HAZARD POTENTIAL ................................................................................. 76 6.2. DOSE RESPONSE .......................................................................................................... 78

7. REFERENCES ........................................................................................................................ 79

APPENDIX A. Summary of External Peer Review and Public Comments and Disposition ..... A-1

APPENDIX B. Documentation of Benchmark Dose Modeling ................................................. B-1

ivLIST OF TABLES

Table 2-1. Chemical and physical propert ies of thallium and selected thallium compounds ................................................................................................................... 4

Table 3-1. Urine concentrations of thal lium for the U.S. population from NHANES, 1999–2002 ................................................................................................................... 7

Table 4-1. Thallium toxicity in hu mans following oral exposure ............................................... 10

Table 4-2. Selected clinical observations in Sprague-Dawley rats treated with thallium sulfate for 90 days ...................................................................................................... 20

Table 4-3. Incidence of alopecia in rats ..................................................................................... .. 21

Table 4-4. Selected blood chemistry values ................................................................................ 22

Table 4-5. Thallium toxicity in an imals following oral exposure ............................................... 29

Table 4-6. Thallium toxicity in animals via injection ................................................................ 366

Table 5-1. Incidence data and BMD modeling re sults for selected clin ical observations in Sprague-Dawley rats treated with thallium sulfate for 90 days ................................. 61

Table B-1. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of rough coat in male Sprague-Dawley ra ts exposed to thallium sulfate via gavage for 90 days ................................................................................................... B-1

Table B-2. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of piloerection in male Sprague-Dawley ra ts exposed to thallium sulfate via gavage for 90 days ................................................................................................... B-3

Table B-3. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of shedding in male Sprague-Dawley rats e xposed to thallium sulfate via gavage for 90 days ............................................................................................................... B-5

Table B-4. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of alopecia in male Sprague-Dawley rats e xposed to thallium sulfate via gavage for 90 days ............................................................................................................... B-6

Table B-5. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of lacrimation in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 day ..................................................................................................... B-6

Table B-6. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of exophthalmos in male Sprague-Dawley ra ts exposed to thallium sulfate via gavage for 90 days ................................................................................................... B-8

vTable B-7. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of miosis in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days ............................................................................................................. B-10

Table B-8. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of behavioral findings in male Sprague-Dawl ey rats exposed to thallium sulfate via gavage for 90 days ........................................................................................... B-12

Table B-9. A summary of BM DS (version 1.4.1) modeling resu lts based on incidence of rough coat in female Sprague-Dawley ra ts exposed to thallium sulfate via gavage for 90 days ................................................................................................. B-13

Table B-10. A summary of BMDS (version 1.4.1) modeling results based on incidence of piloerection in female Sprague-Daw ley rats exposed to thallium sulfate via gavage for 90 days .......................................................................................... B-17

Table B-11. A summary of BMDS (version 1.4.1) modeling results based on incidence of shedding in female Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days ................................................................................................ B-22

Table B-12. A summary of BMDS (version 1.4.1) modeling results based on incidence of alopecia in female Sprague-Dawley ra ts exposed to thallium sulfate via gavage for 90 days ................................................................................................ B-24

Table B-13. A summary of BMDS (version 1.4.1) modeling results based on incidence of lacrimation in female Sprague-Dawle y rats exposed to thallium sulfate via gavage for 90 days ................................................................................................ B-28

Table B-14. A summary of BMDS (version 1.4.1) modeling results based on incidence of exophthalmos in female Sprague-Dawle y rats exposed to thallium sulfate via gavage for 90 days .......................................................................................... B-29

Table B-15. A summary of BMDS (version 1.4.1) modeling results based on incidence of miosis in female Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days ................................................................................................ B-31

Table B-16. A summary of BMDS (version 1.4.1) modeling results based on incidence of behavioral findings in female Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days ............................................................................... B-31

viLIST OF FIGURES

Figure 5-1. POD (mg/kg-day) with correspondi ng derived candidate reference value that would result if histopathol ogic changes of the skin (hair follicle atrophy) were used as the critical effect. ................................................................................. 70

Figure 5-2. PODs (mg/kg-day) with corres ponding derived candidate reference values that would result if clini cal observations from MRI (1988) were used as the critical effect ........................................................................................................... 71

Figure 5-3. POD (mg/kg-day) with correspondi ng derived candidate reference value that would result if clinical chemistry change s (suggesting the liver or kidney as a target) were used as th e critical effect ...................................................................... 72

Figure 5-4. PODs (mg/kg-day) with corres ponding derived candidate reference values that would result if repro ductive toxicity endpoints we re used as the critical effect ........................................................................................................................ . 73

Figure 5-5. PODs (mg/kg-day) with correspondi ng derived candidate reference values that would result if alternativ e endpoints were used as the critical effect .........................74

viiLIST OF ABBREVIATIONS AND ACRONYMS

AchE acetyl cholinesterase AIC Akaike Information Criterion ALA aminolevulinic acid ALT alanine aminotransferase AST aspartate aminotransferase ATSDR Agency for Toxic Substance and Disease Registry BMD benchmark dose BMD 10 benchmark dose correspondi ng to a 10% extra risk BMDL 10 95% lower bound on the benchmark dose corresponding to a 10% extra risk BMDS benchmark dose software BMR benchmark response BUN blood urea nitrogen CASRN Chemical Abstracts Service Registry Number ChAT choline acetyltransferase CHO Chinese hamster ovary EC 50 effective concentration nece ssary to produce a 50% response EPA Environmental Protection Agency GI gastrointestinal GLP good laboratory practice GSH reduced glutathione 5-HT 5-hydroxytryptamine i.p. intraperitoneal IPCS International Programme on Chemical Safety i.v. intravenous IRIS Integrated Risk Information System LD 50 median lethal dose LDH lactate dehydrogenase LOAEL lowest-observed-adverse-effect level MAO monoamine oxidase MDA malondialdehyde MEPP miniature endplate potential MRI Midwest Research Institute NA nucleus accumbens NHANES National Health and Nutrition Examination Survey NLM National Library of Medicine NOAEL no-observed-adverse-effect level NOEL no-observed-effect level OSHA Occupational Safety a nd Health Administration PAD peripheral arterial disease POD point of departure PBPK physiologically based pharmacokinetic RfC inhalation reference concentration RfD oral reference dose

s.c. subcutaneous SCE sister chromatid exchange Tl thallium UF uncertainty factor

viiiFOREWORD

The purpose of this Toxicological Review is to provide scientific support and rationale for the hazard and dose-response assessment in IR IS pertaining to chronic exposure to thallium and compounds. It is not intended to be a comprehensive treatise on the chemical or toxicological nature of thallium and compounds. The intent of Section 6, Major Conclusions in the Charac terization of Hazard and Dose Response , is to present the major c onclusions reached in the deri vation of the reference dose, reference concentration, and cance r assessment, where applicable, and to characterize the overall confidence in the quantitative a nd qualitative aspects of hazard and dose response by addressing the quality of the data and re lated uncertainties. The discus sion 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 a ssessment or other questi ons 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).

ixAUTHORS, CONTRIBUTORS, AND REVIEWERS

CHEMICAL MANAGER/AUTHOR

Octavia Conerly, M.S.P.H. Office of Water U.S. Environmental Protection Agency Washington, DC

AUTHORS

Susan Rieth, M.P.H. Office of Research and Development U.S. Environmental Protection Agency Washington, DC

CONTRACTOR SUPPORT

Robyn B. Blain, Ph.D. ICF Consulting Fairfax, VA

OFFICE OF RESEARCH AND DEVELOPMENT CO-LEAD

Susan Rieth, M.P.H. Office of Research and Development U.S. Environmental Protection Agency Washington, DC

REVIEWERS

This document has been reviewed by EPA sc ientists, interagency reviewers from other federal agencies and White House offices, and the public, and peer reviewed by independent scientists external to EPA. A summary and EP A’s disposition of the co mments received from independent external peer re viewers and from the public is included in Appendix A.

INTERNAL EPA REVIEWERS

Joyce Donohue, Ph.D. Office of Water

Elizabeth Doyle, Ph.D. Office of Water

Steven Kueberuwa, Ph.D. Office of Water

xxi Mike Hughes, Ph.D. Office of Research and Development

Amal Mahfouz, Ph.D. Office of Water

David Thomas, Ph.D. Office of Research and Development

EXTERNAL PEER REVIEWERS

Ronald Baynes, DVM, Ph.D. North Carolina State University

George Cherian, Ph.D. University of Western Ontario

Lucio G. Costa, Ph.D. (chair) University of Washington

George Daston, Ph.D. The Procter & Gamble Company

Robert Hoffman, M.D. New York University

Deborah Rice, Ph.D. Maine Center for Disease Control1. INTRODUCTION

This document presents background information and justification for the Integrated Risk Information System (IRIS) Summary of the h azard and dose-response assessment of thallium and compounds. IRIS Summaries may includ e oral reference dose (RfD) and inhalation reference concentration (RfC) values for ch ronic and other exposure durations, and a carcinogenicity assessment. The RfD and RfC, if derived, provide quantitat ive information for use in risk assessments for health effects known or assumed to be pr oduced 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 ma gnitude) of a daily exposure to the human population (including sensitive subgroups) that is lik ely to be without an appreciable risk of deleterious effects during a lif etime. The inhalation RfC (expressed in units of mg/m 3 ) is analogous to the oral RfD, but provides a c ontinuous inhalation exposure estimate. The
inhalation RfC considers toxic effects for both the respiratory system (por tal of entry) and for effects peripheral to the respiratory system (ext rarespiratory or systemic effects). Reference values are generally derived for chronic exposures (up to a lifetime), but may also be derived for acute ( ≤ 24 hours), short-term (>24 hours up to 30 days ), and subchronic (>30 days up to 10% of lifetime) exposure durations, all of which are derived based on an assumption of continuous exposure throughout the duration sp ecified. Unless specified othe rwise, the RfD and RfC are derived for chronic exposure duration. The carcinogenicity assessment provide s information on the carcinogenic hazard potential of the substance in que stion and quantitative estimates of risk from oral and inhalation exposure may be derived. The information in cludes a weight-of-evidence judgment of the likelihood that the agent is a human carcinogen and the conditi ons under which the carcinogenic effects may be expressed. Quantitative risk esti mates may be derived from the application of a low-dose extrapolation procedure. If derived, th e oral slope factor is a plausible upper bound on the estimate of risk per mg/kg-day of oral expos ure. Similarly, an inhalation unit risk is a plausible upper bound on the estimate of risk per μg/m 3 air breathed. Development of these hazard identificati on and dose-response assessments for thallium and compounds has followed the general guidelin es for risk assessment as set forth by the National Research Council ( 1983). EPA guidelines and Risk Assessment Forum Technical Panel Reports that may have been used in th e development of this assessment include the following: Guidelines for the Health Risk Assessment of Chemical Mixtures (U.S. EPA, 1986a), Guidelines for Mutagenicity Risk Assessment (U.S. EPA, 1986b), Recommendations for and Documentation of Biological Values for Use in Risk Assessment (U.S. EPA, 1988), Guidelines for Developmental Toxicity Risk Assessment (U.S. EPA, 1991a), Interim Policy for Particle Size

1and Limit Concentration Issu es in Inhalation Toxicity (U.S. EPA, 1994a), Methods for Derivation of Inhalation Refere nce Concentrations and Applica tion of Inhalation Dosimetry (U.S. EPA, 1994b), Use of the Benchmark Dose Approach in Health Risk Assessment (U.S. EPA, 1995), Guidelines for Reproductive Toxicity Risk Assessment (U.S. EPA, 1996), Guidelines for Neurotoxicity Risk Assessment (U.S. EPA, 1998), Science Policy Council Handbook : Risk
Characterization (U.S. EPA, 2000a), Benchmark Dose Technical Guidance Document (U.S. EPA, 2000b), Supplementary Guidance for Conducting He alth Risk Assessment of Chemical
Mixtures (U.S. EPA, 2000c), A Review of the Reference Dose and Reference Concentration
Processes (U.S. EPA, 2002), Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005a), Supplemental Guidance for Assessing Susceptibility from Early-Life Exposure to Carcinogens (U.S. EPA., 2005b), Science Policy Council H andbook: Peer Review (U.S. EPA, 2006a), and A Framework for Assessing Health Risks of Environmental Exposures to Children (U.S. EPA, 2006b). The literature search strategy employed fo r this compound was based on the Chemical Abstracts Service Registry Number (CASRN) a nd 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 re levant literature was reviewed through May 2009.

22. CHEMICAL AND PH YSICAL INFORMATION

Metallic thallium (Tl) is bluish white or grey, very soft, malleable, and insoluble in water. Thallium is a Group IIIA metal, one whose salts do not hydrolyze at pH ≥ 7 to form insoluble hydroxides. According to Mulkey and Oehme (1993) , this is a physical pr operty that contributes to thallium’s marked toxicity. Thallium exists in monovale nt (thallous, thallium (I), Tl +1 ) and trivalent (thallic, thallium (III), Tl +3 ) states. Monovalent thallium is favored in the standard potential of Tl +3 /Tl +1 , coupling with a redox potential of +1.25V (Tl +3 + 2e – becomes Tl +1 ). According to Pearson (1963), monov alent thallium is a Lewis acid (e lectron pair receiver) that prefers to interact with inor ganic and organic sulfur, carbon, phosphorous, and arsenic moieties as the electron pair donor (Lewis base). Monova lent thallium ions also are more stable in aqueous solution, but trivalent thallium (Tl +3 ) can be stabilized by comp lexing agents (Sabbioni et al., 1980a). Trivalent thallium forms more stable organic compounds than monovalent
thallium. Monovalent thallium is similar to potassium (K + ) in ionic radius and electrical charge,
which contribute to its toxic natu re. Many of the thallium salts are soluble in water with the exception of thallium (III) oxide, wh ich is insoluble. Thallium compounds and their chemical and physical properties are listed in Table 2-1. Thallium occurs naturally in the earth’s crus t, with a crustal abundance of approximately 1 mg/kg. In soil, thallium c oncentrations are on the orde r of 0.1 to 1 mg/kg; higher concentrations occur in the vicinity of metallic ore deposits. Measureable concentrations of thallium are also found in marine water, freshwat er, and air. Thallium is taken up by vegetation, with the extent of uptake determined by so il acidity and plant species (Kazantzis, 2007). According to the International Programme on Chemical Safety (IPCS) (1996), thallium is used only in small amounts by industry, and thus worldwide production of pure thallium is low. Sources for the production of thallium are zinc, lead, and sometimes copper or iron smelters and sulfuric acid plants as well as a by-product of cadmium production. In 1981 the production of
thallium in the U.S. was discontinued. Thalliu m is released to the environment through the combustion of fossil fuels (in particular from coal -fired power-generating pl ants), refinement of oil fractions, smelting of ferrous and non-ferrous ores (including lead, copper, and zinc), and by some other industrial processes such as cement production and brickworks (Kazantzis, 2007; IPCS, 1996). Due to its ability to remove hair, thallium (I) sulfate was used in the past as a depilatory agent. Thallium (I) sulfate was once used in me dicine to treat infections, such as venereal diseases, ringworm of the scalp, typh us, tuberculosis, and malaria. It was also used in the past as a pesticide for various rodents a nd insects but has been banned fo r this use in the U.S. since 1972. Currently, thallium is used in the semi conductor industry and the manufacture of optic

3lenses. When thallium is alloyed with mercury, it is used on switches and closures, which can operate at subzero temperatures. Thallium comp ounds are also used to manufacture low-melting glass, low-temperature thermometers, alloys, elec tronic devices, mercury lamps, fireworks, and imitation gems. Thallium radioisotopes are used in medicine for scintigraphy of certain tissues and the diagnosis of melanoma (Ibrahim et al ., 2006; National Library of Medicine [NLM], 1998; IPCS, 1996; Agency for Toxic Substan ce and Disease Regist ry [ATSDR], 1992; U.S. EPA, 1991b).

Table 2-1. Chemical and physical pr operties of thallium and selected thallium compounds

Name CASRN Chemical formula Molecular weight Meltin g point (°C) Boiling point (°C) Solubility in water (g/L)
Metallic thallium 7440-28-0 Tl 204.38 303.5 1,457 Insoluble Thallium (I) acetate 563-68-8 TlC 2 H 3 O 2 263.43 131 No data Very soluble Thallium (I) carbonate 6533-73-9 Tl 2 CO 3 468.78 273 No data 40.3 (15.5°C)

Thallium (I) chloride 7791-1 2-0 TlCl 239.84 430 720 Very soluble (20°C) Thallium (I) nitrate 10102-45-1 TlNO 3 266.39 206 430 95.5 (20°C)

Thallium (I) oxide 1314-12-1 Tl 2 O 424.77 596 No data Soluble (as TlOH) Thallium (III) oxide 1314-32-5 Tl 2 O 3 456.76 717 875 Insoluble Thallium (I) selenite 12039-52-0 Tl 2 SeO 3 535.72 No data No data No data Thallium (I) sulfate 7446-18-6 Tl 2 SO 4 504.82 632 Decomposes 48.7 (20°C)

Sources: IPCS (1996); Downs (1993); ATSDR (1992).

43. TOXICOKINETICS

3.1. ABSORPTION Studies in humans and animals indicate that thallium compounds are readily absorbed through various routes of exposure, but few studies provide quantitative m easures of absorption. Mulkey and Oehme (1993) reported that water sol uble salts are rapidly and completely absorbed from the respiratory tract, gastrointestinal (GI) tr act, or skin but did not provide data or cite references to support this conclusion. Thallium ions have been detected in the urine of exposed humans (Ludolph et al., 1986; Davi s et al., 1981; Schaller et al ., 1980; Cavanagh et al., 1974; Gefel et al., 1970) and animals (Thomas and McK eever, 1993; Waters et al., 1992; Leloux et al., 1987; Talas and Wellhöner, 1983), which implie s absorption from environmental sources. Shaw (1933) determined that 61.6% of an oral dose of thallium (I) sulfate (25 mg/kg Tl) was absorbed by a dog. Lie et al. (1960) determin ed that thallium was completely absorbed via the GI tract, following oral administration of 767 μ g/kg 204 Tl as thallium (I) nitrate. This was
based on observations in male Wistar-der ived rats where the body burden decreased exponentially and extrapolated to 100% absorption. The same results were obtained when thallium (as thallium nitrate) was administered by other routes of exposure (intravenous [i.v.],
38 μ g/kg; intramuscular, 96 μ g/kg; subcutaneous [s.c.], 96 μ g/kg; intratracheal, 123 μ g/kg; and intraperitoneal [i.p.], 146 μ g/kg). Eighty percent of a single dose of 10 nmol of thallium, as thallium (I) sulfate, was absorbed within 1 hour from tied-off jejunal segments in anesthetized rats (Forth and Rummel, 1975; Leopold et al., 1968). No information was found regarding the absorption of thallium salts via inhalation. There are a few case reports (Hirata et al., 1998; Ludolph et al., 1986) in which occupational exposure has been associated with toxicity, but it could not be determined if exposure occurred
via inhalation or another rout e (e.g., oral or dermal). The use of thallium salts in the past as de pilatory agents, treatment for ringworm of the scalp, and treatment for night sweats associated with tuberculosis suggests dermal absorption (Léonard and Gerber, 1997; Reed et al., 1963; Lie et al., 1960).

3.2. DISTRIBUTION Thallium ions are rapidly distributed (as early as 1 hour after exposure) throughout the body in both experimental animals (Careaga-O livares and Gonzalez-Ramirez, 1995; GalvánArzate and Rios, 1994; Aoyama, 19 89; Rios et al., 1989; Talas a nd Wellhöner, 1983; Sabbioni et al., 1980a, b; Lameijer and van Zwieten, 1977; Andre et al., 1960; Downs et al., 1960; Lie et al., 1960; Lund, 1956) and humans (Talas et al., 1983 ; Davis et al., 1981; Cavanagh et al., 1974; Barclay et al., 1953), regardless of the route of exposure, dose, and length of exposure (Sabbioni et al., 1980a, b; Lameijer and van Zwieten, 1977). The highest thallium concentrations have

5typically been found in the kidney and the lowest concentrations in the brain, with none being detected in fat tissue. Thallium also has been demonstrated to cross the placenta in humans (Hoffman, 2000) and experimental animals (Gibson and Becker, 1970).
The distribution of thal lium in newborn Wistar rats differed from that in adult Wistar rats.
Newborns administered an i .p. dose of 16 mg/kg thallium (I) acet ate (12.4 mg/kg Tl) had the highest levels of thallium in the testis, heart, and kidneys , in that order, 24 hours after administration (Galván-Arzate and Rios, 1994). Levels in the liver and brain were approximately three- to fourfold lower. In adu lt rats, the level of thal lium in the kidney 24 hours after an i.p. dose of 16 mg/kg thallium (I) sulfat e was approximately twofold higher than the level present in the testis (Rio s et al., 1989). Galván-Arzate and Rios (1994) also demonstrated age-related differences in the regional distribu tion of thallium in the brain. Twenty-four hours after i.p. injection of 16 mg/kg thallium (I) acetate, the thallium content among all regions of the
brain of newborn rats was homogeneous, whereas the thallium content in the brains of 5- to 20-day-old rats showed a region- dependent distribution, with thal lium levels in the cortex significantly lower than leve ls in the hypothalamus.

3.3. METABOLISM Because thallium is an element, it is not me tabolized. It is not known if thallium is
transformed from one valence state to another in vivo.

3.4. ELIMINATION Thallium salts are eliminated mainly via urine and feces, but the amount excreted via each route varies depending on the species. Thallium also has been found to be excreted in breast milk, sweat, saliva, and tears. Thallium depos ition into hair and nails also is considered an important route of elimination (Kazantzis, 2007; IPCS, 1996). A study of a human cancer patient orally administered thallium (I) sulfate and radiolabeled thallium (I) nitrate ( 204 TlNO 3 ) demonstrated that thallium was mainly excreted in the urine; 15.3% of the thallium salts were recovered in the urine over 5.5 days with 0.4% recovered in the feces over 3 days (Barclay et al., 1953). In a survey of 776 member s of the general population ( ≥ 40 years of age) that participated
in the 1999–2000 National Health and Nutrition Examination Survey (NHANES), the geometric
mean level of thallium in the urine was 0.16 μ g/L, with a maximum of 0.86 μ g/L (Navas-Acien et al., 2005).
The Third National Report on Human Exposure to Environmental Chemicals (Centers for Disease Control and Prevention [CDC], 2005) pr ovides ongoing biomonitoring data for the U.S. population for environmental chemicals over the periods 1999–2000 and 2001–2002 collected from NHANES participants. Selected urinary thallium level data from this survey are provided in Table 3-1. For the U.S. population (ages 6 an d older), the geometric mean urinary thallium

6concentration for survey years 2001–2002 was 0.165 μ g/L, and the 95 th percentile concentration
was 0.440 μ g/L.

Table 3-1. Urine concentrations of thallium for the U.S. population from NHANES, 1999–2002

Survey
years Sample size Urinary concentration of thallium in μ g/L urine [ μ g/g creatinine] a Selected percentiles of the U.S. population
Geometric mean 50th 95th Total, ages 6 and older 99–00 2413 0.176 (0.162–0.192) [0.166 (0.159–0.173)] 0.200 (0.180–0.210) [0.168 (0.162–0.176)] 0.450 (0.420–0.470) [0.366 (0.338–0.387)]
01–02 2653 0.165 (0.154–0.177) [0.156 (0.151–0.162)] 0.180 (0.170–0.200) [0.156 (0.148–0.164)] 0.440 (0.410–0.470) [0.348 (0.337–0.365)] Age group 6–11 years 99–00 336 0.201 (0.167–0.243) [0.221 (0.197–0.248)] 0.200 (0.150–0.260) [0.221 (0.196–0.236)] 0.440 (0.350–0.590) [0.424 (0.356–0.600)]
01–02 362 0.172 (0.147–0.202) [0.211 (0.198–0.226)] 0.200 (0.160–0.220) [0.207 (0.198–0.221)] 0.380 (0.360–0.420) [0.411 (0.389–0.456)] 12–19 years 99–00 697 0.202 (0.181–0.225) [0.153 (0.146–0.160)] 0.210 (0.200–0.240) [0.154 (0.146–0.162)] 0.460 (0.430–0.510) [0.321 (0.265–0.364)]
01–02 746 0.200 (0.182–0.220) [0.143 (0.137–0.150)] 0.210 (0.190–0.240) [0.145 (0.135–0.152)] 0.460 (0.400–0.500) [0.307 (0.299–0.333)] 20 years & older 99–00 1380 0.170 (0.157–0.183) [0.162 (0.153–0.171)] 0.180 (0.170–0.200) [0.167 (0.154–0.176)] 0.450 (0.420–0.470) [0.364 (0.325–0.389)]
01–02 1545 0.159 (0.147–0.173) [0.153 (0.147–0.159)] 0.190 (0.170–0.200) [0.152 (0.144–0.161)] 0.440 (0.400–0.490) [(0.342 (0.313–0.362)]
Gender
Males 99–00 1200 0.197 (0.179–0.217) [0.154 (0.147–0.161)] 0.220 (0.190–0.240) [0.156 (0.149–0.164)] 0.440 (0.420–0.480) [0.338 (0.300–0.364)]
01–02 1313 0.184 (0.173–0.196) [0.146 (0.140–0.153)] 0.200 (0.190–0.220) [0.148 (0.141–0.156)] 0.420 (0.390–0.460) [0.307 (0.291–0.342)]
Females 99–00 1213 0.159 (0.145–0.175) [0.178 (0.167–0.189)] 0.180 (0.150–0.200) [(0.182 (0.169–0.196)] 0.450 (0.410–0.490) [0.380 (0.333–0.462)]
01–02 1340 0.149 (0.137–0.163) [0.167 (0.158–0.176)] 0.150 (0.150–0.170) [0.167 (0.153–0.179)] 0.430 (0.400–0.500) [(0.375 (0.348–0.402)]

a 95 th percentile confidence interval in parentheses.

Source: CDC (2005).

As noted above, thallium elimination is not limited to renal excretion. IPCS (1996)
estimated that in humans renal excretion accounts for approximately 70% of total daily excretion
of thallium. This estimate is based on limited human data. In contrast to humans, thallium is excreted to a greater extent in the feces than in the
urine of rats and rabbits. IPCS (1996) estimated that in rats about 2/3 of the intake of thallium

7was excreted via the GI tract and about 1/3 vi a the kidney. Lund (1956) determined that, after 26 days, 51.4% of an i.p. dose of 10 mg/kg thallium (I) sulfate in the rat wa s eliminated via the feces, while 26.4% was excreted in the urine. Talas and Wellhöner (1983) demonstrated that thallium (I) acetate administered to rabbits via i.v. injection (as a radioactive tracer) was excreted mainly in the feces. Both studies found that, although the feces was the major route of excretion in the rat and rabbit, neither spec ies had high levels in the bile, suggesting that ex cretion via the liver was relatively low. Lund (1956) determined that thallium was mainly excreted in the feces through gastric and intestinal secretions, which is likely associated with potassium excretion. Lund (1956) demonstrated that rabbits excreted thallium through the kidneys by glomerular filtration, but approximately one-half the dose filtered was reabsorbed in the tubuli. In Syrian golden hamsters, thallium (I) sulfat e was mainly excreted in the feces after i.p. administration but was excreted at an equal ra te in the feces and ur ine after an oral dose (Aoyama, 1989). Shaw (1933) demonstrated th at 32 and 61.6% of a single oral dose of 25 mg/kg Tl as thallium (I) sulfate administered to a dog was excreted in the urine at 3 and 36 days after dosing, respectively. Sabbioni et al. (1980b) determ ined that thallium (I) sulfate administered at doses of 0.00004–2,000 μ g/rat was persistent in the kidneys for 8 days (192 hours) after dosing with 2.5% of the dose still present at that time (suggesting a half-life of approximate ly 1.5 days). Lehmann and Favari (1985) and Lie et al. (1960) estimated the biological half-life of thallium in rats to range from 3–8 days. The biological half-life in humans has been estimated to be approximately 10 days, with values up to 30 days reported (IPCS, 1996).

3.5. PHYSIOLOGICALLY BASE D TOXICOKINETIC MODELS There are no physiologically based toxic okinetic models for thallium compounds.

89 4. HAZARD IDENTIFICATION

4.1. STUDIES IN HUMANS—EPIDEMIOLOGY, CASE REPORTS, CLINICAL
CONTROLS Studies of thallium toxicity in humans are comp rised of clinical report s, case studies, and medical surveys. Because case reports larg ely involved accidental ingestion, intentional poisoning, or suicide attempts, they do not provi de useful information on thallium toxicity associated with chronic exposure. Available epidemiology studies involving long-term exposure to thallium are limited by small study populations and insufficient characterization of long-term exposure. Health effects information was based on self-reporting (via questionnaire) or medical histories/physical examinations of uncertain scope. In adults, the average lethal oral dose has been estimated to range from 10 to 15 mg/kg (Gosselin et al., 1984; Schoer, 1984) . Without treatment, death typically follows in about 10– 12 days, but death as soon as 8–10 hours also has been documented (IPCS, 1996).

4.1.1. Incident/Case Reports As indicated by case reports, the acute toxi city of thallium is ch aracterized by alopecia (hair loss), severe pain in th e extremities, lethargy, ataxia, abdominal pain or vomiting, back pain, abnormal reflexes, neuropathy, muscle weakness, coma, convulsion, other neurological symptoms (i.e., mental abnormalities, tremors, abnormal movements, abnormal vision, and headache), and death (Lu et al., 2007; Tsai et al., 2006; Saha et al., 2004 : Sharma et al., 2004; Rusyniak et al., 2002; Atsmon et al., 2000; Hi rata et al., 1998; Feldman and Levisohn, 1993; Yokoyama et al., 1990; Heyl and Barlow, 1989; Roby et al., 1984; Limos et al., 1982; Davis et al., 1981; Cavanagh et al., 1974; Gefel et al., 1970; Reed et al., 1963). Symptoms were
observable within 14 hours after a high dose (i.e., 5–10 g of thallium (I) nitrate), with death occurring 8 days later (Davis et al., 1981). The lowest known singl e dose of thallium associated with adverse effects was reported to be 0.31 g of thallium (I) acetate (3.4 mg/kg Tl, assuming a 70 kg body weight) (Cavanagh et al., 1974). This dose caused paresthesia, pain, weakness, vomiting, and alopecia in a 26-ye ar-old male. Approximately 1 month after the onset of symptoms, complete recovery occurred following treatment. In adults, doses ranging from 6 to 40 mg/kg have been reported to be lethal (IP CS, 1996). Table 4-1 su mmarizes the individual
case reports.Table 4-1. Thallium toxicity in humans following oral exposureReferenceSexAgeDoseSymptomsaFinal outcomeMales—adultGefel et al. (1970)Male41 yearsUnknown butchronic;urine thallium0.15 mg/100 mLHigh blood pressure; lower back pain; vomiting;severe pain in the feet; weakness of the calf muscle;alopecia; slurred speech;atrophic lower limbs;limited visionDeathCavanagh et al. (1974)Male60 years0.93 g thallium (I)acetate in 2 divideddosesDiarrhea; vomiting; dizziness; back pain; paresthesiaof the feet and lower legs;high blood pressure; facialweakness; dysphagia; difficulty breathingDeath within a week ofsymptomsCavanagh et al. (1974)Male56 years0.93 g thallium (I)acetate in 3 divideddosesAbdominal pain; diarrhea; vomiting; paresthesia;photophobia, nystagmus, visual impairment; facialweakness; bilateral ptosisDeath within 3 weeks ofsymptomsCavanagh et al. (1974)Male26 years0.31 g thallium (I)acetateParesthesia in both feet; chest pain; tenderness overthe sternum; vomiting, weakness, pain in the kneesand ankles that inhibited walking; alopeciaRecoveryDavis et al. (1981)Male19 years5–10 g thallium (I)nitrateNausea; vomiting; slurred speech; paresthesia ofhands and feet; respiratory weaknessDeathLimos et al. (1982)Male56 yearsUnknownVisual disturbances; alopecia; elevated AST andALT; high blood glucose and creatine kinase;decreased myelinated fibers; denervated Schwanncell clustersBedridden; could notspeakLimos et al. (1982)Male26 yearsUnknownVisual disturbances; alopecia; elevated AST andALT; high blood glucose and creatine kinase;decreased myelinated fibers; denervated Schwanncell clustersResidual tremors of theextremities and muscleweakness of the lowerlimbsRoby et al. (1984)Male45 yearsUnknown;urine thallium:2,000μg/LBurning pain in feet; inability to walk; alopecia;acute fibrillationContinued neurologicaldysfunction10Table 4-1. Thallium toxicity in humans following oral exposureReferenceSexAgeDoseSymptomsaFinal outcome4/5 recovered; 1/5experienced permanentneurological damageHeyl and Barlow (1989)Male“Five youngmen”UnknownFollicular plugging of the skin (nose, cheeks, andnasolabial folds) by keratinous material; crustedeczematous lesions and acneiform eruptions on theface; dry scaling on palms and soles; and alopecia(scalp, eyelashes, lateral eyebrows, arms, and legs).Skin biopsies (scalp and cheek): disintegrating hairshafts, gross follicular plugging, and eosinophilickeratohyalin granules in the epidermis; necroticsebaceous glands; pustular lesions on the face:folliculitis and necrosis of the follicles; (feet) markedhyperkeratosis and hypergranulosis.Yokoyama et al. (1990)Male31 yearsUnknown;urine thallium:3.5 mg/LNausea, vomiting; leg pain; alopecia; abnormalbehavior; decreased conduction velocity of fastnerve fibersRecoveryHantson et al. (1997)Male48 years200 mg thallium (I)sulfateNo overt symptoms within 24 hours; increase inbinucleated cells with micronuclei 15 days afterexposureRecoveryHirata et al. (1998)Male29 yearsUnknown; hairthallium: 20 ng/g(32 months afterpossible exposure)Alopecia; abdominal pain; diarrhea; tingling inextremities; neuropathyRecoveryAtsmon et al. (2000)Male40 yearsUnknown;urine thallium: 7 mgWeakness of the limbs; vomiting; severeneurological symptoms; alopecia; high bloodpressure; increased ALT and AST; Mees lines;decreased visual acuity; bilateral foot dropRecoverySharma et al. (2004)Male48 yearsUnknown;serum thallium:870μg/100 mLurine thallium:5,000μg/mLPainful peripheral neuropathy, decreasedconsciousnessDeath11Table 4-1. Thallium toxicity in humans following oral exposureReferenceSexAgeDoseSymptomsaFinal outcomeFemales—adultRoby et al. (1984)Female51 yearsUnknown;serum thallium:50μg/100 mL;urine thallium:5,000μg/LNumbness and weakness of the legs and hands;alopecia; fluctuating pulse and blood pressure;bradycardia; hypotensionPersistent ventricularectopy and neurologicaldysfunction,necessitating placementat a nursing homeRoby et al. (1984)Female61 yearsUnknown;serum thallium:740μg/100 mLBurning chest pain; paresthesia; difficulty speakingand swallowing; inability to walk; hypotension;acute respiratory distress syndrome (ARDS)DeathRoby et al. (1984)Female80 yearsUnknown;serum thallium:422μg/100 mL;urine thallium:21,600μg/LARDSDeathHoffman (2000)FemalePregnant; agesnot specified150–1,350 mgthallium (I) sulfateParesthesia; abdominal pain; muscle weakness;lethargy; alopecia; Mees linesNone specifiedSaha et al. (2004)Female26 yearsUnknown;serum thallium:12μg/100 mLHeadache, lethargy, abdominal pain, muscle cramps,joint pain, backache, numbness of fingers, alopecia,erosion of nailsNot specifiedBoth sexes—adultBrockhaus et al. (1981)BothNot reportedUnknownSleep disorders; tiredness; weakness; nervousness;headache; other psychic alterations; neurological andmuscular symptomsNot reportedSchoer (1984); Gosselinet al. (1984)BothAdult10–15 mg/kgthalliumNone specifiedDeath (average lethaldose)Rusyniak et al. (2002)BothVariousUnknown; variouslevels were detectedin urineMyalgia; arthralgia; paresthesia; dysesthesia; jointstiffness; insomnia; alopecia; abdominal painRecovery in 7 adults; 5had ongoing psychiatricproblems12Table 4-1. Thallium toxicity in humans following oral exposureReferenceSexAgeDoseSymptomsaFinal outcomeTsai et al. (2006)Both48-year oldfemale; 52-yearold male1.5–2 .4 gConfusion, disorientation, hallucination, anxiety,depression, memory impairment, peripheralneuropathy, erythematous skin rashes, diarrhea,tachycardia, alopeciaImpairment of memoryand verbal fluencyremained at 6 months;neuropsychologicalimpairment persisted at9 monthsAt 1-year follow-up,persistent paresthesia,dysesthesia, andimpairment of smallsensory nerve fibers inskinLu et al. (2007); Kuo etal. (2005)Both48 and 52 years1.5 and 2.3 g/person(estimated);Serum thallium:950–2,056μg/LUrine thallium:11,325–14,520μg/LNausea, vomiting; general aching muscle pain;numbness of tongue and mouth within a few hours;severe paresthesia and dysesthesia in hands and feet(one day postexposure); erythematous rash; diarrhea;urine retention; hyporeflexia; muscle weakness;hypoesthesia; acneiformeruptions; alopecia (1–3weeks); Mees lines (2–3 months).Skin biopsy: parakeratosis; dilated hair folliclesfilled with keratin and necrotic sebaceous materials;mild epidermal atrophy; vacuolar degeneration of thebasal layer.Cutaneous nerve biopsy: axonal degeneration; lossof epidermal nerves indicating involvement of thesmall sensory nerves (2 months).ChildrenReed et al. (1963)Both1–11 yearsUnknownAlopecia; lethargy; ataxia; abdominal pain;vomiting; abnormal reflexes; neuropathy; muscleweakness; coma; convulsionNeurologicalabnormalities;retardation; psychosis;deathFeldman and Levisohn(1993)Male10 yearsUnknown;serum thallium:296μg/L;urine thallium:322μg/24 hoursAlopecia; leg paresthesia; abdominal pain; seizuresRecoveryHoffman (2000)BothTransplacentalUnknownPremature birth; lowbirth weight; alopeciaNone specified1314Table 4-1. Thallium toxicity in humans following oral exposureReferenceSexAgeDoseSymptomsaFinal outcomeAmmendola et al. (2007)Male16 years1.3 g thalliumsulfate;urine thallium:3,400μg/LAcute stage: GI disturbances, alopecia, and clinicaland electrodiagnostic signs of severepolyneuropathy.3 years post-poisoning:neurologicalsymptoms makingprogress;electrophysiologicalsigns of peripheralneuropathy mainlyconfined to lower limbs.6 years post-poisoning:persistent weakness andsensory disturbances ofdistal lower extremities;neurological andelectrodiagnosticabnormalities affectingmainly the feet.aALT = alanine aminotransferase; AST = aspartate aminotransferase.High blood pressure or fluctuating blood pressure was noted upon hospital admission in several cases (Roby et al., 1984; Cavanagh et al., 1974; Gefe l et al., 1970). Elevated serum aspartate aminotransferase (AST) (formerly referred to as serum glutamic oxaloacetic transferase) and serum alanine aminotransferase (ALT) (formerl y referred to as serum glutamate pyruvate transaminase), high blood glucose, and creatine ki nase values also have been noted in case reports of thallium exposure (Atsmon et al., 2000; Limos et al., 1982). The same symptoms were noted across age and sex groupings. Reta rdation and psychosis were the most common findings in children (1–11 years ol d) after nonlethal thallium expos ure. Several cases were so severe that institutionalization was necessary (Reed et al., 1963). Thallium significantly
decreased the conduction velocities of faster nerve fibers in a 31-year-old male, who ingested a thallium-containing rodenticide, compared with ba seline levels recorded following recovery. In most case-study reports, thallium was detectab le in the urine or tissues. In some cases, thallium could not be definitively associated with the symptoms because other heavy metals were also found in the blood or urine of the subject. Hantson et al. (1997) evaluated cytogeneti c changes in blood from a 48-year-old man who accidentally ingested 200 mg of thallium (I) sulfate intended for rodenticide use. Despite the lack of overt symptoms 24 hours after ingest ing the thallium, the man was admitted to the emergency room and Prussian blue treatments were commenced. Blood samples were obtained on days 1 and 15 for cytogenetic analysis. Slight increases in mean sister chromatid exchange (SCE) numbers on days 1 and 15 were not consider ed related to thallium exposure. A 3.5-fold increase in binucleated cells with micronuclei (3 5% versus 10% in the historical controls) was noted on day 15. The thallium level was determined to be 14.4 μ g/dL in blood at the time of hospital admission, and the con centration in urine was 3,804 μ g/g Tl creatinine (reference value,
<1 μ g/g Tl). Fifty-one case histories of women treated for thallium poisoning following external application of a 3 to 8% thallium (I) acetate oi ntment were reviewed for signs of possible thallium intoxication (Munch, 1934). Neurological and GI symptoms were observed in 29 cases after an unspecified number of applications with 2–24 ounces of the ointment. This was approximately equivalent to a dose of 53–636 mg/ kg Tl per application us ing a 5.5% ointment on a 50 kg woman. Alopecia followed seve ral weeks after beginning treatment. Hoffman (2000) provided case reports and a co mprehensive literature review of thallium poisoning that occurred during preg nancy. Exposures were primarily oral, but some of the cases involved dermal exposure. The majority of the doses were unreported, but those doses that were documented ranged from 150–1,350 mg thallium (I) sulfate. Of the 18 cases that met Hoffman’s criteria (cases were excluded if maternal or fetal outcomes were not provided), 5 women were exposed during the first trimester and 5 during the second trimester; the remaining 8 were exposed during the third trimester. The ages of the women were not reported. While the mothers developed the classic symptoms of thalli um poisoning, including paresthesia, abdominal

15pain, muscle weakness, lethargy, alopecia, and Mees lines (single transverse white bands occurring on the nails), the only consistent fi nding in their offspring was a trend toward prematurity and low birth weight. Several of the children had alopecia, particularly those exposed during the third trimester.

4.1.2. Population Surveys Several published studies have surveyed populations living near a cement plant in Lengerich, a small city in northwest Germany. These populations were studied because of their potential to experience exposure to thallium as a result of its pr esence as an impurity in pyrite and its release during the roasting of pyrite for use in making some types of cement. Thallium was discharged to outdoor air, deposited in so ils, and taken up by lo cal crops and indigenous plants. People who lived near the plant and consumed home-grown foods thus were exposed to thallium through their diets. Prior to 1979, the concentration of thallium in the pyrite was 400 ppm. After 1979, a pyrite with lower le vels of thallium (2 ppm) was used. Brockhaus et al. (1981) conducted an epidemiologic study of a group of 1,200 people living near the cement plant in Lengerich. Urinar y thallium data were also collected from two reference populations without increased tha llium intake—one group consisting of 31 persons living in a small (rural) city in northwest Ge rmany and a second group consisting of 10 persons living in an urban area in Dusseldorf, Germany. The study investigators di d not perform specific tests for toxicity but surveyed for the presence of certain symptoms by using questionnaires. Thallium exposure was assessed by measurements in urine and hair. The thallium body burden of the study population was increa sed over the referen ce populations, as i ndicated by a mean urinary thallium level of 5.2 ± 8.3 μ g/L (range: <0.1–76.5 μ g/L) in the study population compared to the reference population means of 0.4 ± 0.2 μ g/L (rural) and 0.3 ± 0.2 μ g/L (urban) (range: 0.1–1.2 μ g/L). The predominant contributing factor to the thallium burden was consumption of homegrown fruits and vegetables . When the consumption of homegrown foods was restricted, thallium exposure was reduced, as i ndicated by decreased thallium in the urine. No correlation between dermal or GI symptoms and thallium level was observed. There was a negative correlation between ur inary thallium and hair loss ( 13.6% with urine levels <2 μ g/L,
6.6% with urine levels 2–20 μ g/L, and 5.9% with urine levels >20 μ g/L; 10.7% with hair levels <10 ng/g, 9.6% with hair levels 10–50 ng/g, and 2.3% with hair levels >50 ng/g). These data appear to conflict with other reports that indi cate hair loss increases with increasing thallium exposure. A positive association was observed among thallium levels in urine or hair and the following self-reported symptoms: sleep disorders, tiredness, w eakness, nervousness, headache, and psychological alterations as we ll as neurological and muscular symptoms (Brockhaus et al., 1981). Dolgner et al. (1983) examined the potential developmental effects of thallium in this same German population. Of 300 births regist ered in Lengerich between January 1, 1978, and

16August 31, 1979, questionnaires on health status and maternal risk factors were completed by the mothers of 297 infants. One hundred fifty-four urine and 164 hair samples were analyzed for thallium content. All children with suspected congenital malformations or other abnormalities were examined physically, and medical histories of mothers were taken. Eleven out of the 297 births were identified as exhibiting congenita l malformations or abnormalities (confirmed by a pediatrician) with five major malformations note d. Two of the five major malformations in the study population were determined by the authors to likely be due to hereditary factors. The observed rate of congenital malforma tions in the study population (5/297) was compared to the expected rate of 0.8/297 births based on annual statistics from the North RhineWestphalia region of Germany for 1974–1978. C ongenital malformations in the reference population were thought to be underr eported because reporting of bi rth defects is not required on
birth certificates in that area of Germany. The study authors not ed that other investigations reported an incidence of 2–3% of birth defects am ong live births, a value that is consistent with
1.7% incidence of birth defect s in the study population (5/297 for major malformations) and 3.7% (11/297) for all malformations. The study authors concluded that a causal relationship between thallium exposure and congenital malf ormations in this population was unlikely. However, study deficiencies, includ ing lack of information on exposure to thallium at the time of pregnancy, limit the strength of this study. Navas-Acien et al. (2005) examined the a ssociation between urinar y levels of various metals, including thallium, with peripheral arterial disease (PAD) in a cross-sectional analysis of 790 participants in NHANES 1999–2000. Thallium was not associated with PAD in this sample of the U.S. population.

4.1.3. Occupational Exposure Schaller et al. (1980) examined 128 men (ages 16–62 years) who were exposed to thallium for 1–42 years in three cement manufact uring plants in the Franconia region of Germany. Health effects were determined thr ough medical histories and a physical examination for symptoms. Information on the scope of th e physical examinations was not provided. Analyses of roasted pyrites and electro-filter dus t confirmed the presence of thallium in various production areas in the plants. The median con centration of thallium in the urine in exposed
workers was 0.8 μ g/g Tl creatinine w ith a range of <0.3–6.3 μ g/g Tl creatinine. The range in
20 individuals without known o ccupational exposure was <0.3–1.1 μ g/g Tl creatinine (median concentration not reported). Me dical histories and physical ex aminations did not indicate thallium poisoning. The health status of exposed workers, however, was not compared with an unexposed reference population, and a single measur ement of urinary thallium did not provide a measure of past exposures. Thirty-six cement plant workers (presumabl y in Germany) were examined for clinical and electrophysiological parameters (Ludolph et al., 1986). Thallium levels were found to be

17elevated in the blood of 16 workers, urine of 5 work ers, and hair of 5 work ers. It was not noted if these were all separate cases or if elevations in all three parameters occurred in the same individuals. The study determin ed that 28–39% of the individual s had some form of peripheral and central motor and sensory impairment. The neurological impairments could not conclusively be attributed to thallium expos ure because half the patients suffered from concurrent diseases (including pe ptic ulcer, diabetes, disorders of joints and connective tissue, and hypertensive vascular diseas e), which could possibly cause ne uromuscular impairment. No controls were employed, and no correlations were made with the levels of thallium in individuals
and their disease states. In another occupational study, Marcus (1985) examined medical r ecords for 86 workers (sex not reported) occupationally exposed to thallium at a magnesi um seawater battery factory. Exposure was determined by measuring thallium in urine samples. Marcus also examined the records of 79 unexposed workers matched for age, length of employment, shift pattern, and type of work. Exposed workers did not have an incr ease in incidence of be nign neoplasms or any other clinical diagnoses when compared with unexposed workers. This study is limited by lack of exposure quantitation, the size of th e cohort, and unknown length of follow-up. Although there are many case reports of thal lium poisoning in the literature, the doses were largely unknown because ingestion was accide ntal or occurred through criminal poisoning. Given the severity of reported symptoms, most of the exposures were likely to have been relatively large. The few epidemiology studies that looked at populat ions surrounding a cement factory that released thallium only attempted to compare thallium exposure with congenital malformations or surveyed symptoms. None of the studies specifically studied cancer as an endpoint. Overall, the available epidemiology litera ture is considered limited and inconclusive.

4.2. LESS-THAN-LIFETIME AND CHRONI C STUDIES AND CANCER BIOASSAYS
IN ANIMALS—ORAL AND INHALATION 4.2.1. Oral Exposure
4.2.1.1. Acute and Subchronic Studies
4.2.1.1.1. Rats. In a study performed by Midwest Resear ch Institute (MRI) (1988) for EPA’s Office of Solid Waste, male and female Sprague-D awley rats (45 days ol d, 20/sex/group) were administered 0 (untreated and vehicle cont rols), 0.01, 0.05, or 0.25 mg/kg-day of an aqueous solution of thallium (I) sulfate (approximately 0, 0.008, 0.04, or 0.20 mg/kg-day Tl) by gavage for 90 days. The study was conducted in comp liance with EPA good laboratory practice (GLP) mandates. The MRI (1988) study is an unpublished study; accordingly, an external peer review was initiated by EPA in November 2006. Body weight, food consumption, hematologic and clinical chemistry parameters, ophthalmologic examinations, gross pathological observations, and organ weights (liver, kidneys , brain, gonads, spleen, heart, a nd adrenals) were recorded for all animals. Neurotoxicological examinations (three times/week) were performed on six

18rats/sex/group; these examinations were appare ntly observational (further details were not provided in the study report). Tissues from three rats/sex/group were prepared for neuropathologic examination. Complete histopathologic examinations (including neuropathologic examinations) were conducted fo r the vehicle control and 0.2 mg/kg-day Tl groups only; for the other three groups, only the livers, lungs, kidneys and gross lesions were examined histopathologically. Ne uropathologic examinations incl uded the following: dorsal and ventral root fibers of the spinal nerves, dorsal root ganglia, spin al cord at C3–C6 and L1–L4, and
six sections of the brain. There were no statistically significant differences in body weight, food consumption, or absolute and relative organ weights among control groups and groups receiving thallium (I) sulfate. The study authors concluded that the histopathologic examina tion did not reveal any
treatment-related effects. Lacrimation (secretion of t ears), exophthalmos (abnormal pr otrusion of the eyeball), and miosis (contraction of the pupil) were observed at higher incidences in the treated male and female rats compared with both untreated and vehicle controls (Table 4-2). Ophthalmologic examination and gross and hist opathologic examinati on of the eyes, however, revealed no
treatment-related abnormalities. The incidence of clinical obse rvations related to the coats (including rough coat, piloerecti on, shedding, and alopecia) and be havior (including aggression, tension/agitation, hyperactivity, voca lization, and self-mutilation) were also elevated in male and female rats at the higher doses (Table 4-2).

19Table 4-2. Selected clinical observat ions in Sprague-Dawley rats treated with thallium sulfate for 90 days

Observation a Untreated control Vehicle control 0.008 mg/kg-day 0.04 mg/kg-day 0.2 mg/kg-day

Male
Coat/skin

Rough coat 1/20 3/20 11/20 16/20 19/20

Piloerection 0/20 0/20 1/20 4/20 13/20 Shedding 0/20 0/20 4/20 10/20 8/20 Alopecia 2/20 1/20 4/20 9/20 4/20 Eyes

Lacrimation 1/20 6/20 19/20 20/20 20/20 Exophthalmos 1/20 5/20 12/20 20/20 20/20

Miosis 0/20 1/20 5/20 7/20 15/20

Behavior b 3/20 0/20 7/20 6/20 7/20

Female
Coat/skin

Rough coat 1/20 0/20 1/20 5/20 11/20

Piloerection 0/20 0/20 0/20 3/20 8/20 Shedding 0/20 0/20 2/20 3/20 13/20 Alopecia 4/20 1/20 4/20 9/20 12/20 Eyes

Lacrimation 7/20 6/20 20/20 20/20 20/20 Exophthalmos 5/20 6/20 19/20 20/20 20/20

Miosis 2/20 3/20 1/20 11/20 8/20

Behavior b 2/20 2/20 0/20 1/20 7/20

a Listed as number of animals with the sign observed at least once during the 90-day study.
b Animals exhibiting one or more behavioral observations at least once during the 90-day study, including the

following: aggression, tension/agitation, hyperactivity, vocalization, self-mutilation.

Source: MRI (1988).

As noted above, the incidence of alopecia was increased, particularly in female rats (see Table 4-3). Examination of individual animal clin ical observation data for female rats from the MRI (1988) study showed that alopecia was first observed in control and treated groups anywhere from study day 44 to 60. Based on a st atistical analysis performed by the U.S. EPA 1

, the incidence of alopecia (total number of cases in each dose group) was statistically

1 A statistical analysis of the incide nce of alopecia (based on the total num ber of cases of alopecia in each dose group) was performed by EPA by using Fisher’s exact test. Incidence in the treated groups was compared with incidence in the untreated control, vehicle control, and pooled control.
20significantly elevated relative to controls in mid-dose males and mid- and high-dose females. Most instances of alopecia in females were attr ibuted to barbering be havior (where fur was present but cropped short). Of the 12 high-dose females with alopecia, 5 instances were not totally attributed to barbering behavior. Histopathologic examin ation revealed atrophy of the hair follicles in two high-dose female rats. Be cause the skin was examined for histopathologic changes only in the vehicle control and high-dose groups, no information on dermal histopathology was available for the low- and mi d-dose groups. The two high-dose females with atrophy of the hair follicles also had alopecia; whether the hair follicle atrophy and alopecia
occurred at the same location on the rats could not be determined from the study report. The study authors concluded that the alop ecia was attributable to the cyclic pattern of hair growth in rodents. Consequently, the authors did not c onsider these findings to be toxicologically significant.

Table 4-3. Incidence of alopecia in rats

Dose (mg/kg-day Tl) Males Females Alopecia a, b Hair follicle atrophy c Alopecia a, b Hair follicle atrophy c 0 (untreated control) 2/20

-- d 4/20

-- d 0 (vehicle control) 1/20 0/20 1/20 0/20
0.008 4/20

-- d 4/20

-- d

0.04 9/20 e

-- d 9/20 f

-- d

0.2 4/20 0/20 12/20 e 2/20

a Number of animals with alopecia at least once during the 90-day stud y based on clinical observations.
b Of the animals with alopecia, the following are the number s of cases in each dose group that the study authors stated were “not totally attributed to barbering behavior”:

Males: untreated control, 1; vehicle control, 0; 0.008 mg/kg-day, 2; 0.04 mg/kg-day, 4; 0.2 mg/kg-day, 1. Females: untreated control, 0; vehicle control, 0; 0.008 mg/kg-day, 1; 0.04 mg/kg-day, 3; 0.2 mg/kg-day, 5.

c Based on histopathologic observation.
d Skin was not examined for histopathologic lesions.

e Incidence of alopecia (total number of cases ) was statistically significantly elevated ( p < 0.05) relative to incidence in vehicle control, incidence in untreated control, and pooled incidence of vehicle and untreated control, based on Fisher’s exact test performed by EPA.
f Incidence of alopecia (total number of cases ) was statistically significantly elevated ( p < 0.05) relative to incidence in vehicle control and pooled incidence of vehicle and untreated control, based on Fisher’s exact test performed by EPA.

Source: MRI (1988).

Subtle but statistically significant change s were observed in several blood chemistry parameters that the investigators considered probably treatment relate d. Specifically, doserelated increases in AST, lactate dehydrogenase (LDH), and sodium levels and decreases in blood sugar levels were detected in male and female rats after 30 and 90 days of exposure. Reported values for the selected blood chemistry parameters are summarized in Table 4-4. Other

21changes in blood chemistry parameters were le ss consistent across species, dose groups, and exposure durations. At 90 days, the differences in AST, LDH, sodium, and blood sugar levels in dosed male and female rats were no greater than +31, +38, +4, and –21%, respectively, of the vehicle control group values. The investigators observed that the increases in AST and LDH levels could indicate a possible effect of treatment on cardiac function, that increases in LDH coupled with subtle changes in electrolytes could indicate an effect on renal functi on, and that, in rare instances, a decrease in blood s ugar coupled with an increase in sodium occurs as a defense mechanism for maintaining cellula r integrity. The investigators concluded that none of the changes observed in the blood chemistries of ma le or female rats during the study were of sufficient magnitude to significantly affect the health status of the animals. Further, histopathologic evaluation did not confirm a ny cellular damage suggested by the clinical chemistry findings.

Table 4-4. Selected blood chemistry values

Endpoint Study day Untreated control Vehicle control 0.008 mg/kg-day 0.04 mg/kg-day 0.2 mg/kg-day
Males a
AST (I.U.) 30 90 91 ± 26.5 77 ± 19.7 108 ± 18.6 87 ± 17.8 128 ± 24.5 b
99 ± 20.4 134 ± 29.0 b ,c 113 ± 27.0 b,c 152 ± 20.1 b ,c 114 ± 31.1 b,c

LDH (I.U.) 30 90 795 ± 322 587 ± 305 1206 ± 424 b
856 ± 385 1333 ± 340 b
1003 ± 363 b 1396 ± 407 b
1071 ± 507 b 1802 ± 341 b ,c 1119 ± 477 b

Na (meq/L) 30 90 148 ± 1.3 144 ± 1.6 149 ± 2.4 147 ± 2.0 b 152 ± 4.0 b
147 ± 1.9 b 154 ± 2.5 b ,c 149 ± 2.0 b,c 153 ± 2.1 b ,c 151 ± 2.2 b,c Blood sugar (mg/100 mL) 30 90 100 ± 22.1 158 ± 15.6 97 ± 18.1 138 ± 16.8 b 93 ± 10.0 131 ± 17.6 b 90 ± 18.3 121 ± 15.7 b 62 ± 14.8 b ,c 113 ± 22.4 b,c

Females a
AST (I.U.) 30 90 95 ± 22.8 77 ± 19.2 115 ± 30.3 90 ± 19.1 127 ± 27.8 b
93 ± 33.1 149 ± 26.8 b ,c 111 ± 30.7 b 154 ± 18.2 b ,c 112 ± 31.0 b

LDH (I.U.) 30 90 1047 ± 335 745 ± 320 1277 ± 495 881 ± 273 1402 ± 501 823 ± 354 1763 ± 370 b ,c 1044 ± 436 1764 ± 361 b ,c 1219 ± 338 b

Na (meq/L) 30 90 148 ± 1.7 146 ± 1.8 150 ± 1.9 146 ± 1.0 153 ± 4.1 b ,c 148 ± 1.8 b,c 154 ± 2.8 b ,c 150 ± 2.0 b,c 155 ± 2.5 b ,c 152 ± 1.0 b,c Blood sugar (mg/100 mL) 30 90 103 ± 23.9 110 ± 28.7 80 ± 13.3 b
89 ± 15.9 80 ± 9.0 b
103 ± 19.9 67 ± 20.0 b
88 ± 20.4 50 ± 11.8 b ,c 70 ± 18.0 b

a Mean ± standard deviation of 7–10 rats.
b Significantly different ( p < 0.05) from the untreated control group.

c Significantly different ( p < 0.05) from the vehicle control group.

Source: MRI (1988)

.

The authors concluded that the minor dose-rela ted changes in this study did not affect the
health status of the treated animals and ther efore were not toxicolo gically significant and

22identified the highest dose, 0.25 mg/kg-day th allium (I) sulfate (0.20 mg/kg-day Tl), as a noobserved-effect level (NOEL). However, upon further analysis by EPA of the MRI (1988) findings as part of this health assessment, a different determination was reached regarding the
no-observed-adverse-effect leve l (NOAEL) and lowest-observed- adverse-effect level (LOAEL) (see the discussion in Section 5.1.1). Manzo et al. (1983) administered drinking water containing thallium (I) sulfate at a concentration of 10 mg/L Tl (a pproximately equivalent to a do se of 1.4 mg/kg-day Tl based on reported thallium intakes and an assumption that the rats weighed 200 g) to 80 female SpragueDawley rats for 36 weeks. Mortality was 15 and 21% after 40 and 240 days of treatment, respectively. After 4+ weeks (32 days) of trea tment, hair loss appeared and involved about 20%
of the animals thereafter. Functional and histopathologic ch anges were observed in the peripheral nerves, including cha nges in motor and sensory action potentials and histopathologic changes in the sciatic myelin sheath and a xonal destruction charac terized by Wallerian degeneration (degeneration of th e axon and its myelin sheath dist al to a site of injury), mitochondrial degeneration, neurofilamentous cl ustering, and elevated lysosomal activity.
Ten adult male albino rats were ad ministered 0.8 mg/kg (1/20th of the LD 50 ) of thallium (I) sulfate orally (presumably via gavage) fo r 3 months (El-Garawany et al., 1990). Blood samples were obtained initially a nd at monthly intervals. At a ll three monthly intervals, the treated group had statis tically significantly ( p < 0.001) increased levels of blood urea, serum creatinine, serum bilirubin, and serum ALT. The largest increase (<90%) in these parameters occurred in the first month, with smaller incr eases (>15%) occurring for each additional month. Mourelle et al. (1988) examined the effect s of silymarin, an antioxidant, on various biochemical indicators of liver damage in male Wistar rats (200–250 g) induced by oral administration (gavage) of thallium (I) sulfate (10 mg /kg) dissolved in water. The controls were given vehicle only. Ten rats per group were sacrificed at 0, 24, 48, 72, and 96 hours and 5, 10, and 20 days after treatment. Without silymarin administration, thallium administration produced a statistically significant ( p < 0.05) decrease in the content of glycogen and reduced glutathione (GSH) and caused a sta tistically significant ( p < 0.05) increase in malondialdehyde (MDA) production and triglycerides in the liver 48 hours after treatment. (MDA production and GSH
content in the liver served as indicators of lipid peroxidati on.) Levels of serum alkaline phosphatase were increased and liver cell membra ne alkaline phosphatase activity was decreased after 24 hours and remained unchanged for 5 days. Furthermore, Na + /K + -ATPase activity in the liver cell membranes was rapidly reduced within 24 hours of thallium treatment; the decrease persisted through day 5 and bega n to rebound by day 10, with values similar to the control by day 20. Serum and liver cell membrane gamma- glutamyl transpeptidase and serum ALT were significantly ( p < 0.001) elevated by 24 hours and rema ined elevated through day 5. Administration of silymarin (100 mg/kg i.p.) comp letely prevented these biochemical changes. The authors suggested that silymarin acted by stabilizing membranes via some antioxidant

23property. During the 20 days, none of the rats tr eated with thallium al one died, but the rats exhibited signs of toxicity that in cluded hypomotility and piloerection. Downs et al. (1960) fed groups of Wistar-derived albino rats (5/sex/dose) diets containing nominal concentrations of 0, 5, 15, or 50 mg thallium (I) acetate/kg (or ppm) in the diet (corresponding to approximately 0, 0.4, 1.2, or 3.9 mg/kg-day Tl for 100 g rats, assuming food consumption of 10 g/day). Animals were allowed ad libitum access to these diets for 15 weeks. At the 50 ppm dose level, mortality was 100% by w eek 5 in males and by week 13 in females. By week 15, 4/10 control animals died (2/sex), ma king interpretation of surv ival in the remaining dose groups difficult (15 ppm, 3/5 males and 1/5 females died; 5 ppm, 2/6 males and 0/4 females died). An additional treatment group (30 ppm) and control group (corresponding to 0 and 2.4 mg/kg-day Tl) were added 6 weeks after the st udy had been initiated and were maintained on the diet for 9 weeks. At the end of the 9 weeks, 2/5 male and 1/5 female controls were dead and 4/5 males and 3/5 females at 30 ppm were dea d. At termination, the only gross finding was alopecia in the 15 and 30 ppm groups. The alopecia was noted be ginning 2 weeks after commencement of the diet, with the rats nearly fr ee of hair at terminati on. The authors reported a slight increase in kidney weight (doses not specified; data not show n). The authors also reported that histopathologic eval uations did not indicate treatm ent-related pathology, but they did not prepare skin sections. The study findi ngs for alopecia suggest a NOAEL and LOAEL of 0.4 mg/kg-day Tl and 1.2 mg/kg- day Tl, respectively, for this endpoint. Because mortality
occurred in rats in both the c ontrol and treated groups, it is not possible to determine whether the deaths in low-dose (5 ppm) male rats were re lated to thallium exposure. Therefore, a NOAEL and LOAEL cannot be reliably established for this study. Downs et al. (1960) also examined the effects of thallium (III) oxide on weanling Wistarderived albino rats (five rats/sex/treatment). Rats received 0, 20, 35, 50, 100, or 500 mg/kg (or ppm) thallium (III) oxide in the diet for 15 week s. This was equivalent to doses of 0, 1.8, 3.1, 4.5, 9.0, or 44.8 mg/kg-day Tl, respectively. All rats (males and females) treated with 50 ppm and greater in the diet died before 8 weeks. The mortality rates in the remaining groups at 15 weeks were as follows: 1/5 control males, 0/5 males treated with 20 ppm, and 4/5 males treated with 35 ppm; 0/5 control females, 2/ 5 females treated with 20 ppm, and 2/5 females treated with 35 ppm. Thalli um (III) oxide caused a dose-related decrease in body weight at 15 weeks. Body weight reductions relative to the cont rol were 50 and 180 g in males treated with 20 and 35 ppm dietary doses, respectively, and 50 g in female s treated with 35 ppm in the
diet. Males treated with either 20 or 35 ppm in the diet had marked hair loss beginning around 4 weeks, with near complete hair loss after 6 weeks; females were less affected. There was a statistically significant ( p ≤ 0.05) increase in absolute kidney weights in males and females treated with 20 ppm and females treated with 35 ppm and a dose-response trend in kidney to body weight ra tio. Histopathologic examination did not reveal any alterations in the kidney related to thallium treatment. Hi stopathologic evaluation of the skin revealed a

24decrease in the number of hair follicles and hair shafts, atrophy of the remaining follicles, decrease in the size of the sebaceous glands, and hyperkeratinized epidermis. However, the incidence by dose was not pres ented. The lowest level te sted, 1.8 mg/kg-da y Tl (20 ppm thallium (III) oxide in the diet), is considered to be a LOAEL based on findings of alopecia and significant elevations in kidney weights for male and female rats. A NOAEL was not identified for this study. Leloux et al. (1987) investigated the acute toxicity of oral exposure to thallium (I) nitrate in the adult Wistar rat. In the first experime nt, a single dose of 20 mg/k g thallium (I) nitrate was administered via gavage to male and female ra ts (three per sex); all males and females were found dead within 40 and 54 hours post-dosing, respect ively. Increases in absolute kidney (36%, females; 61%, males) and adrenal (47%, fe males; 100%, males) weights were observed following the single exposure. The second experime nt involved administering four daily gavage doses of 1 mg/kg-day thallium (I) ni trate to 20 animals of each sex. Male rats treated with four doses began to lose their hair 96 hours after the first exposure. A ll treated animals had diarrhea. After the fourth gavage dose, 2/20 males and 2/20 females died. Two more females died within 126 hours, and 11 females and 15 males died within 168 hours. Three rats of each sex were sacrificed at 126 hours post-dosing for gross pathological examination and organ weight changes. The remaining two females were sacr ificed at 192 hours post- dosing. Treated animals weighed less than the untreated controls. Th e tissues did not demons trate any macroscopic degenerative changes, but there was an increase in the absolute weight s of the kidneys (33%, females; 48%, males) and eyes (54%, females; 34%, males). Histopath ology was not performed.

4.2.1.1.2. Dogs. Reports of thallium toxicity in dogs ar e limited to a few cases in the literature of accidental exposure. A 9-m onth-old Doberman pinscher accide ntally consumed mole bait containing 1% thallium (Waters et al., 1992). Two days later the dog was lethargic, vomited blood, and had bloody feces. The dog had moderate hypoproteinemia and a slight prolonged activated clotting time. The dog’s condition was improved by the third day following supportive care, including treatment with activated charcoal. Thomas and McKeever (1993) reported a case of a 1-year-old neutered male miniature schnauzer that had ingested an unknown amount of bread soaked in thallium (the level in one piece of bread was 1.6 ppm). Beginning sympto ms were lethargy, followed 2 weeks later by severe, rapidly progressing alopeci a. No abnormalities were f ound in a complete blood count test, serum chemistry profile, urinalysis, or abdominal radiographs. Diphenylthiocarbazone treatments (40 mg/kg three times daily) were star ted upon establishing thallium toxicity. On the second day of veterinary treatment, the dog sh owed signs of respir atory distress and was euthanized due to its poor condition. An autops y revealed severely congested and edematous lungs, congestion of the liver and kidneys, a nd areas of congestion and hemorrhage in the

25pancreas. Histologic evaluations demonstrated abnormalities in the lungs, kidneys, liver, and pancreas. Thallium was detected in the liver ( 11 ppm), kidneys (12 ppm), and spleen (7 ppm). Histopathology of the skin from 13 cases of thallium poisoning in dogs revealed dyskeratotic and necrolytic changes in the skin and hair follicles (Schwartzman and Kirschbaum, 1961). The most prominent features were massive parakeratosis, spongiform abscess formation, and induction of telogen follicles.

4.2.1.2. Chronic Studies and Cancer Bioassays
There are no chronic animal studies or can cer bioassays for thallium reported in the
literature.

4.2.2. Inhalation Exposure
No studies were identified that examined the effects of inhaled thallium in animal
models.

4.3. REPRODUCTIVE/DEVE LOPMENTAL STUDIES 4.3.1. Reproductive Toxicity Effects of thallium on male reproduction have been investigated in rats (Formigli et al., 1986; Gregotti et al., 1985; Zasukhina et al., 198 3) and mice (Wei, 1987). These studies suggest that thallium exposure can produce effects on the testes and sperm. N one of the available reproductive toxicity studies, however, used st andard protocols for evaluating reproductive endpoints. No studies of the potential reproductive toxicity of thallium in female experimental
animals were identified. Male Wistar rats (10/group) were ad ministered drinking wa ter containing 10 ppm thallium (I) sulfate (approximately 0.7 mg/kg- day Tl based on reported daily thallium consumption [270 μ g Tl/rat] and initial body weights [350–380 g]) (Formigli et al., 1986). The compound was administered for 30 and 60 days. Although the study authors stated that the controls were pair-fed, they also stated that food was available ad libitu m and that thallium did not affect food consumption. No abnormalities we re observed after 30 days of treatment. However, after 60 days of treatment, the fo llowing testicular effects were observed: disarrangement of the tubular epithelium, cytoplasmic vacuolation and distention of smooth endoplasmic reticulum of the Sertoli cells, reduced testicular β -glucuronidase activities (an enzyme primarily located in th e Sertoli cell and spermatogonia), high concentrations of thallium in the testes, and reduced sperm motility. Plasma te stosterone levels were within normal limits. β -Glucuronidase activity was also affected after 60 days of trea tment and ultrastructural changes
were observed in the Sertoli cells. From thes e results, a LOAEL of 0.7 mg/kg-day Tl (10 mg/L)
was identified.

26Gregotti et al. (1985) also reported β -glucuronidase activity and ultrastructural changes in the Sertoli cells after 60 days of treatment. Greg otti et al. (1992) further examined this effect
in vitro and demonstrated that thallium (even at the lowest concentration) causes a dose- and time-dependent detachment of germ cells from Se rtoli cells when testicular cells were treated with thallium concentrations corresponding to 1.4, 7, and 35 μ g/g testis, estimated from protein
content of cultures. Zasukhina et al. (1983) performe d a dominant lethal test with male rats that were given daily oral doses of thallium (I) carbonate (0.005, 0.05, and 0.5 μ g/kg-day) for 8 months and subsequently mated with untreated females. The authors concluded that thallium carbonate produced a treatment-related enhancement of embr yonic mortality. The authors’ conclusion was not, however, supported by the data presented in the paper. The number of implantation sites (10.11 ± 0.48 to 11.05 ± 0.49), number of “yellow bodies” in pregnant rats (11.11 ± 0.35 to 11.75 ± 0.33), and number of live embryos (9.77 ± 0.48 to 10.87 ± 0.37) were similar across the control and treated groups. (“Yellow bodies” we re not further defined in the paper but may mean corpora lutea.) Because the paper lacked statistical analys is, it was unclear whether the differences were statistically si gnificant. Only the number of resorptions in the control group (0.87 ± 0.13) was appreciably higher than that in the treated groups (0.22 ± 0.12 to 0.33 ± 0.13). In addition, the authors’ calcula tion of total embryonic deaths could not be reproduced from the data provided. Finally, over all confidence in the reported findings was low because of inadequate reporting (e.g., the number of male rats exposed and the rat stra in were not reported), the use of nonstandard terminology, the relatively small number of pregnant females (16–18 per group), and lack of statistical analysis. Wei (1987) administered 0, 0.001, 0.01, 0.1, 1.0, or 10.0 mg/L thallium (I) carbonate in drinking water for 6 months to groups of ma le Kunming mice (20/group) weighing 15–20 g at study initiation. (Assuming an average body wei ght of 20 g over the exposure period and drinking water ingestion rate of 6 mL/day [D erelanko and Hollinger, 1995], these exposure levels are approximately equivalent to doses of 0.0003, 0.003, 0.03, 0.3, and 3 mg/kg-day Tl. These doses are approximate only because the ag e of animals at study initiation, terminal body weights, and water consumption were not reported. ) At the end of the exposure period, half of the male mice (10/group) were sacrificed for epididymal sperm examination. The remaining males (10/group) were housed with untreated fema les (1:2 ratio) for 1 w eek to evaluate male reproductive function. At the end of the 1-week mating period, these male mice were also sacrificed for epididymal sperm examination. Untreated female mice were sacrificed on gestation day 20 and evaluated for the following measures of reproductive function: number of pregnant female mice, number of live and dead fetuses, number of implantations, and number of early resorptions. Water intake, body weights, behavior, and animal health were reportedly assessed; however, this informa tion was not provided in the st udy report. The author reported that sperm motility (rapid speed, sperm immobility ) was affected at the lowest drinking water

2728 concentration (0.001 mg/L) test ed. Effects were shown to increase with increasing concentration, thus indicating a dose-response relationship. At 0.01 mg/L and higher, the number of dead sperm was statistically significantly increased. Sperm count was statistically significantly reduced and the percent of deformed sperm was increased at concentrations of 0.1 mg/L and higher. The author observed that there was an adverse effect on sperm quality (motility) at low doses, and, as the dose increased , there was an accompanying decrease in sperm count in addition to the motility change. Exam ination of reproductive function in the group of
mice housed for 1 week with untreated females re vealed that the reproduc tive index (number of pregnant female mice/number of mated female mice) and the number of implantations were not statistically different between tr eated and control animals. The mean number of live fetuses was statistically significantly increa sed at concentrations of 0.01 mg/L and above. The percent of dead fetuses was significantly lower than in the control group at c oncentrations of 0.001, 0.01, and 0.1 mg/L but was increased at the two highest concentrations (1 and 10 mg/L). Review of the reported results reveals that a number of male mice were not accounted for at study termination. Of the initial 20 male mice/group, sperm results were provided for only 12 mice in the 0.1 mg/L group, 10 mice in the 10 mg/L group, and 16 mice in the remaining groups. No explanation is provided for the loss of animals over the 6-month study. The author concluded that the lowest dose tested (0.001 mg/L thallium (I) carbonate) was a LOAEL, causing reproductive effects in male mice. Table 4-5 summarizes thallium toxicity in animals following oral exposure.Table 4-5. Thallium toxicity in animals following oral exposureReferenceSpeciesAgeSexRouteDose and durationNOAELLOAELEffectAcute studiesLeloux et al.(1987)Rat3/sexAdultBothOral(gavage)20 mg/kg thallium(I) nitrate; singledoseNIa15 mg/kg TlDifficulty breathing; rough coat; increasedabsolute kidney, adrenal weights; deathLeloux et al.(1987)Rat10/sex/groupAdultBothOral(gavage)0, 1 mg/kg thallium(I) nitrate; once dailyfor 4 daysNI0.77 mg/kg TlAlopecia; diarrhea; increased absolutekidney, eye weights; deathMourelle et al.(1988)Rat10/groupNSbMaleOral(gavage)0, 10 mg/kg thallium(I) sulfate; singledose. Sacrificed at24 hours to 2 daysafter dosingNI8.1 mg/kg TlLiver changes: increased triglycerides andlipid peroxidation; decreased glutathione andglycogen; increased alkaline phosphatase inserum and liver cell membranesSubchronic studiesDowns et al.(1960)Rat/5/sex/groupNSBothOral(feed)0, 5, 15, or 50 ppmthallium (I) acetate(corresponding to 0,0.4, 1.2, or3.9 mg/kg-day Tl);15 weeks0 or 30 ppm(corresponding to 0or 2.4 mg/kg-dayTl); 9 weeks0.4 mg/kgday Tl*1.2 mg/kg-day Tl*Alopecia; increased kidney weight; mortalityin treated and control groups.*The NOAEL and LOAEL are for alopecia.Because of reported mortality in the controland treated groups, a study NOAEL andLOAEL cannot be reliably determined.Downs et al.(1960)Rat5/sex/groupWeanlingBothOral(feed)0, 20, 35, 50, 100,and 500 ppmthallium (III) oxide(corresponding to 0,1.8, 3.1, 4.5, 9.0, and44.8 mg/kg-day Tl);15 weeksNI1.8 mg/kgday Tl (20ppm)Reduced body weight; alopecia; increasedmortality; increasedabsolute and relativekidney weightsEl-Garawanyet al. (1990)Ratn = 10NSMaleOralc0.8 mg/kg thallium(I) sulfate; 90 daysNI0.65 mg/kgday TlIncreased blood urea; serum creatinine;serum bilirubin; serum ALTManzo et al.(1983)Ratn = 80NSFemaleOral(DWd)10 mg/L Tl asthallium (I) sulfate;36 weeksNI1.4 mg/kgday TlNerve histopathology; alopecia; mortality2930Table 4-5. Thallium toxicity in animals following oral exposureReferenceSpeciesAgeSexRouteDose and durationNOAELLOAELEffectMRI (1988)Rat20/sex/group45 daysBothOral(gavage)0, 0.01, 0.05, or 0.25mg thallium (I)sulfate/kg(corresponding to0,0.008, 0.04, or0.20 mg/kg-day Tl);90 daysNI0.008 mg/kgday TleIncreased incidence of alopecia and otherobservations related to coat (rough coat,piloerection, shedding); lacrimation,exophthalmos, and miosis; and variousbehavioral observations; statisticallysignificant increases in AST, LDH, andsodium levels; decreased blood sugar levels.The study authors identified 0.2 mg/kg-dayTl as the NOAEL.Reproductive and developmental toxicityFormigli et al.(1986);Gregotti et al.(1985)Rat10/groupAdultMaleOral(DW)0, 10 ppm thallium(I) sulfate; 30 or 60daysNI0.7 mg/kgday TlTesticular effects: tubular epitheliumdisarrangement; cytoplasmic vacuolation;reduced sperm motility; distention of smoothendoplasmic reticulum of Sertoli cells;reducedβ-glucuronidase activityWei (1987)MouseNSMaleOral(DW)0, 0.001, 0.01, 0.1,1.0, and 10 mg/Lthallium (I)carbonate(corresponding to 0,0.0003, 0.003, 0.03,0.3, and 3 mg/kgday Tl); 6 monthsNI0.0003 mg/kg-day TlDecreased sperm motility and counts;increase in deformed sperm; decrease in livefetuses.Dose estimated from an assumed averagebody weight of 20 g and drinking wateringestion rate of 6 mL/day.Rossi et al.(1988)RatPerinatalBothMother’s,thenpup’s DW(DW)0, 1 mg/dL ofthallium (I) sulfateDay 1 of gestation toweaning then thru 60daysNINIPrenatal and postnatal exposure caused adelay in the development of the pilusapparatus by 50 days; reduction of theα- andβ-adrenergic and muscarinic vasomotorreactivity noted.aNI = not identified.bNS = not specified.cPresumably via gavage.dDW = drinking water.eSee discussion of the NOAEL and LOAEL determination in Section 5.1.1.4.3.2. Developmental Toxicity Developmental toxicity studies in the rat (Barroso-Moguel et al., 1992; Rossi et al., 1988; Gibson and Becker, 1970) and chicken embr yo (Hall, 1985, 1972; Karnofsky et al., 1950) provide evidence that thallium exposure during development can produce abnormalities (including effects on the developi ng vascular autonomic nervous system and bones) and reduced fetal body weight. Of the studies in rats, onl y one involved oral dri nking water exposure to thallium (Rossi et al., 1988); in other developmental rat studies , dams were exposed by i.p. injection. A group of NOS albino male and female rats was administered 1 mg/dL of thallium (I) sulfate from day 1 of gestation to weaning (22 da ys after birth) via the dams’ drinking water then through their own drinking water until 60 days of age (Rossi et al., 1988). These rats were considered prenatally exposed. Another group of NOS albino male and female rats was exposed to 1 mg/dL of thallium (I) sulfate via the dams’ drinking water from birth until weaning (22 days after birth) then through their own drinking water until 60 days of age. These rats were considered postnatally exposed. Both situations (pre- and postnatal exposur e) caused a delay in the development of the pilus appara tus by 50 days. A reduction of the α - and β -adrenergic and muscarinic vasomotor reactivity also was noted. Authors noted that this reduction may be due to one of the following mechanisms: probable reducti on in the number and/or sensitivity of both α and β -adrenergic and muscarinic receptors or a cha nge of cell membrane in relation to a possible modification of potassium cell concentration. Gibson and Becker (1970) administered thalli um (I) sulfate (i.p.) to pregnant Simonsen Sprague-Dawley rats during earl y (2.5 mg/kg on days 8, 9, and 10) or late (2.5 or 10 mg/kg on days 12, 13, and 14) gestation. Fetuses were ex amined for abnormalities. All three thallium
treatments caused a sta tistically significant ( p < 0.05) reduction in fetal body weight. Thallium treatment (2.5 mg/kg) during early ge station caused a slight (not statistically signific ant) increase in the incidence of hydronephrosis (29% in treated ve rsus 16% in control) and missing or nonossified vertebral bodies (36% in treated versus 17% in controls). The 2.5 mg/kg thallium (I) sulfate treatment administered during late gestation caused a statistically significant ( p < 0.05) increase in the incidence of hydr onephrosis (47% in treated vers us 16% in control) and missing or non-ossified vertebral bodies (60% in treated ve rsus 17% in controls). Increasing the dose to 10 mg/kg thallium (I) sulfate during late ge station did not increase the incidence of developmental abnormalities. In fact, 10 mg/kg thallium (I) sulfate administered during late gestation had no effect on the incidence of hydr onephrosis and was comparable to the 2.5 mg/kg dose administered during late gestation in th e induction of missing or non-ossified vertebral bodies (i.e., 60% in treated versus 17% in cont rols). Maternal toxi city (diarrh ea, lethargy, irritability, poor hair luster , and hair loss) was noted. Barroso-Moguel et al. (1992) administered a single i.p. injection of 32 mg/kg aqueous thallium (I) acetate to 20 newborn (24-hour-old) Wist ar rats. Results were compared with those

31of five vehicle controls. Rats (four treated a nd one control per time point ) were sacrificed at 24, 48, and 72 hours and at 7 and 50 days. Cartilaginous and osseous tissue alte rations were noted. Diarrhea was observed through 72 hours postinj ection. Two rats surviving to 50 days postinjection had persisting alopecia (one irreversible and one with discrete recovery). Although skeletal images of 72-hour-old animals did not show any differences when compared with the control, microscopic images of th e distal third of the tibia show ed disorganization and edema of the fibroblasts of the fibrous layer. By day 7, de lays in ossification in the right forelimb were noted. Microscopic examination demonstrated a majority of pyknotic chondrocytes and the lack
of bone trabeculae calcification. Profound skeletal alterations were noticeable 50 days after injection. Many of the cartilaginou s cells were altered or dead, leading to a decrease of the growth cartilage and scanty bone trabeculae with few osteoblasts. The bone marrow also had few myeloblasts and megakaryocytes. Hall (1972) incubated chick embryos in a for ced-draft Humidaire incubator and injected 0.6 mg thallium (I) sulfate/0.5 mL saline into each embryo via the chorioallantoic membrane at 7 days of incubation. This dose caused a minimal le thal effect with surviv al varying from 94 to 100%. Treated embryos were smaller than cont rols from 10 days of incubation onward and by 18 days were 26% smaller than controls. Trea ted embryos failed to commence ossification or had not progressed to similar de velopmental stages observed in the control. The long bones of treated embryos were smaller (the tibia to a greater degree than the femur), contained less organic material, and contained more water (as a percent of dry weight) than the untreated control embryos. An abnormal distribution of the acid mucopolysaccharides and necrotic areas in maturing hypertrophic chondrocytes were detected histologically. In addition, biochemical assays verified the reduced acid mucopolysaccharide activity. Hall (1985) further demonstrated that the critical period of sus ceptibility ended at 8 2/3 days of incubation. Tibial growth was inhibited by thallium sulfate in 8-day-old embryos but not in 9-day-old embryos. Achondroplasia (a birth defect characteri zed by imperfect bone formation) was also induced in embryonic chicks via in vitro cultures w ith injection into the chorioallantoic membrane (Hall, 1985, 1972) or injection into th e yolk sac (Karnofsky et al., 1950). Karnofsky et al. (1950) determined that thallium (I) sulfate was lethal to 2-day embryos at a dose lower than would be necessary to induce ac hondroplasia. Treatment of 4- day-old chick embryos with 0.2, 0.5, 1.0, or 2.0 mg/egg induced 0, 45, 92, and 100% in cidence of achondroplasia, respectively. Although the data were not presented, the study authors reported that thallium (I) nitrate produced achondroplasia at similar doses.

4.4. OTHER ENDPOINT-SPECIFIC STUDIES A number of investigators have specifically examined the effect of thallium compounds administered to experimental animals by injectio n (s.c., i.p., or i.v.) a nd reported effects on the liver, kidneys, heart, and nervous system.

324.4.1. Liver and Kidney Toxicity Liver and kidney were among the organs a ffected when male and female SpragueDawley rats were given s.c. injection of thallium (I) acetate as acute (single dose of 20– 50 mg/kg), subacute (2–3 weekly injections of 10–15 mg/kg), or chronic (10–20 mg/kg, followed by weekly injections of 5 mg/kg or occasio nally 2.5 mg/kg for up to 26 weeks) exposures (Herman and Bensch, 1967). Toxicity was obser ved in all treatment groups. Animals dosed acutely displayed the symptoms earlier than t hose on subacute or chronic dosing schedules. Animals were sacrificed when signs of toxicity became apparent. Acutely exposed animals had the following changes observed via light microscopy: eosinophilic granular casts in 50–75% of the renal proximal and dist al tubules, mild to moderate enteritis, moderate to severe colitis, and dens e infiltration of polymorphonuclear leukocytes and lymphocytes that extended through all layers of the wall of the large intestine. Electron microscopy revealed severe degenerative changes in the mitochondria of the renal tubular cells and hepatocytes. In rats that received sub acute injections of thal lium acetate, eosinophilic granular casts occurred in one-t hird of the proximal and distal tubules. Electron microscopy revealed moderately prominent mitochondrial granules in the kidne y, dense bodies in the cytoplasm of the cells in the loops of Henle, and distal convoluted tubules. Mitochondrial granules of hepatocytes were slightly en larged and lacked electron-lucent cores. In the chronically exposed rats, electr on microscopy revealed increased size of mitochondrial granules in the proximal convolut ed tubules and an in crease in cup-shaped
mitochondria in the distal convoluted tubules. Hepatocytes had increased numbers of large complex residual bodies and lipid droplets, and the mitochondria were swollen with enlargement of mitochondrial granules. Yoshida et al. (1997) administered a si ngle i.p. injection of thallium (I) sulfate (25 mg/kg) to ICR mice (30–35 g). As was observed in Mourelle et al. (1988) after a single oral dose of thallium (I) sulfate (10 mg/kg) to Wistar rats, liver Na + /K + -ATPase was statistically significantly decreased by 12 hours. However, the rebound occurred by 24 hours instead of the 5 days observed in the Mourelle et al. (1988) study. While the Na + /K + -ATPase activity was decreased, the ATP levels were increased and retu rned to control values by 12 hours post-dosing. The effects were slightly differe nt in the kidney. At 6 hours Na + /K + -ATPase activity was statistically significan tly decreased but had rebounded by 12 hours. The ATP levels were significantly ( p < 0.01) increased through 12 hours then de creased to levels significantly ( p < 0.01) lower than controls by 24 hours; AT P levels did not rebound until 240 hours after
thallium administration. Male albino rats (210–260 g) receiving a single i.p. injection of 30, 60, or 120 mg/kg thallium (I) sulfate had st atistically significant ( p < 0.05) increased levels of AST and ALT above controls (0.5 mL of 0.9% saline) 16 hours after treatment, regardless of dose (Leung and Ooi, 2000). There was a dose-dependent incr ease in ALT and AST between 30 and 60 mg/kg,

33but the levels did not increase from 60 to 120 mg/kg. Some 30 mg/kg animals exhibited weakness, sluggishness, loss of hair , ptosis of the eyelids, dia rrhea, and respiratory difficulty; they were sacrificed 4 days post-dosing. The AL T and AST levels in these animals were still elevated over the controls by a factor of 2–2.5. Serum creatinine levels also were elevated by approximately 2.5 times over the control (0.5 mg/dL control versus 1.33 mg/dL treated). Histologic evaluation confirmed damage to both th e kidney and the liver. The damage in the
liver consisted of necrosis and swollen and v acuolated cells, which appeared to reduce the sinusoidal space. Kidney tubules we re atrophied and vacuolated w ith cell outlines less distinct and cells containing many pyknotic nuclei. Amorphous material was apparent in the lumen of the proximal tubules, and the br ush borders were disorganized. Appenroth et al. (1995) examined the effect s of a single i.p. dose of thallium (I) sulfate (5, 10, 15, or 20 mg/kg) on renal function and mo rphology in adult female Wistar rats. Low doses (i.e., 5 and 10 mg/kg) of thallium (I) sulf ate increased the volume of urine (measured on day 2) but did not affect the protein level. Higher doses (i.e., 15 and 20 mg/kg) caused a reduction in urinary volume along with an increas e in the urinary protein concentration. The glomerular filtration rate was statistically significantly ( p ≤ 0.05) reduced at 2 days after treatment with 20 mg/kg but had returned to control levels by day 10. Blood urea nitrogen (BUN) levels were significantly ( p ≤ 0.05) increased 2 days after tr eatment but had returned to normal values by day 10. Histopathology showed a thickening of ascending limb of the loop of Henle notable on day 2 after treatment and resolved by day 10. Na + /K + -ATPase activity was significantly ( p ≤ 0.05) increased in the medulla on day 2 but was significantly ( p ≤ 0.05) reduced by day 5. No changes in Na + /K + -ATPase activity were noted in the cortex. In follow-up studies, Appenroth et al. ( 1996) and Fleck and Appenroth (1996) examined the age-related nephrotoxicity of thallium (I) sulfate. Both studies demonstrated that nephrotoxicity was more severe in the adult rat (Wistar) than in young rats (10 or 20 days old) after a single i.p. injection of 20 mg/kg thallium (I) sulfate. Appe nroth et al. (1996) determined
that there were several biochemical differences in kidney function between 10- and 20-day-old rats (Wistar) administered 20 mg/kg thallium (I) sulfate, but there were no structural changes indicating kidney damage in either group of youn g rats. In comparison, the thick ascending limb of loop of Henle in 55-day-old rats showed damage. Thallium did not affect the glomerular filtration rate in either 10- or 20-day-old rats but caused a significant reduction in the rate in 55-day-old rats (Appenroth et al., 1996; Fleck and Appenroth, 1996). Thallium caused a statistically significant ( p < 0.05) increase in the fractional excr etion of a few amino acids (i.e., ß-alanine, taurine, and 1-methylhis tidine), a statistically significant ( p < 0.05) decrease in the fractional excretion of glycin e in 10-day-old rats, and a statistically significant ( p < 0.05)
increase in the fractional excretion of 13 amino acids in 55-day-old rats. Fleck and Appenroth (1996) determined that thallium affects renal tubular amino acid resorption and causes kidney damage only when mature kidney function is present.

3435 Woods and Fowler (1986) examined the e ffects of a single i.p. dose of thallium (III) chloride (TlCl 3 ) at doses of 50, 100, or 200 mg/kg on liv er structure and function in male Sprague-Dawley rats (CD strain ; 150–200 g) 16 hours after treatment. They determined a doserelated effect on the volume de nsity of mitochondria (increased), rough endoplasmic reticulum (increased), lysosomes (increased), and cytopl asm (decreased). The surface densities of the inner cristae of mitochondria and the rough endop lasmic reticulum also were determined to increase in a dose-dependent manne r. Statistically significant ( p < 0.05) increases occurred in monoamine oxidase (MAO) (100 and 200 mg/kg) and ferrochelatase (50, 100, and 200 mg/kg). Statistically significant ( p < 0.05) decreases occurred in aminole vulinic acid (ALA) synthetase (50, 100, and 200 mg/kg), aminopyrine demethyl ase (200 mg/kg), aniline hydroxylase (50, 100, and 200 mg/kg), and NADPH cytochrome c (P45 0) reductase (50, 100, and 200 mg/kg). In addition, in vitro studies using 50, 100, or 200 μ g/mL thallium (III) chloride demonstrated a significant ( p < 0.05) reduction in ALA synthetase, ferro chelatase, aniline hydroxylase, ALAdehydratase, and NADPH cytochrome c (P450) reductase for all dose concentrations. A doserelated loss of ribosomes from the smooth endoplasmic reticulum and proliferation of the rough endoplasmic reticulum were observed through ultrastructural examination. Also observed were generalized mitochondrial swelling and increa sed numbers of electron-dense autophagic lysosomes. Table 4-6 summarizes toxicity data from anim al studies involving i.p. or i.v. injection.Table 4-6. Thallium toxicity in animals via injectionSpeciesAge orweightSexRoute/exposureperiodDosesNOAEL/LOAELStudy type/effectReferenceAcute studies (single dose)Ali et al. (1990)RatAdultMalei.p./single injection20 mg/kg Tl asthallium (I) acetateLOAEL = 20 mg/kgTlNeurological toxicity:neurochemical changes in thebrain that were resolved within 24hrsAppenroth et al.(1995)RatAdultFemalei.p./single injection5, 10, 15, and20 mg/kg thallium(I) sulfateLOAEL = 12 mg/kgTlKidney toxicity: decreased urinevolume; increased urine protein;thickened ascending limb of theloop of Henle; changes in brainNa+/K+-ATPaseBarroso-Moguelet al. (1990)RatNewbornBothi.p./single injection32 mg/kg thallium(I) acetateLOAEL = 25 mg/kgTlNeurological toxicity: neuronaland vascular damage in the brainBarroso-Moguelet al. (1992)RatNewbornBothi.p./single injection32 mg/kg thallium(I) acetateLOAEL = 25 mg/kgTlDevelopmental toxicity: diarrhea;alopecia; cartilaginous and osseoustissue alterations; profoundskeletal alterationsBarroso-Moguelet al. (1996)RatNewbornBothi.p./single injection16 mg/kg thallium(I) acetateLOAEL = 12 mg/kgTlDevelopmental toxicity: diarrhea;muscle atrophy; small size;alopecia; death; interstitial edemabetween myelin sheaths; thinnermuscle fibers; nerve fiber damageKuperberg et al.(1998)Rat250–300 gMalei.p./single injection25 mg/kg thallium(I) acetateLOAEL = 19 mg/kgTlBladder and neurological toxicity:distended bladder; reduced acetylcholinesterase activity in the brainand bladder; increased cholineacetyltransferase activity in thebrain and bladderLameijer andvan Zwieten(1976)RatYoungadultMalei.v./single injection3–100 mg/kgthallium (I) sulfateLOAEL = 24 mg/kgTl (30 mg/kg thallium(I) sulfate)Cardiotoxicity: hypertension36Table 4-6. Thallium toxicity in animals via injectionSpeciesAge orweightSexRoute/exposureperiodDosesNOAEL/LOAELStudy type/effectReferenceLeung and Ooi(2000)Rat210–260 gMalei.p./single injection30, 60, 120 mg/kgthallium (I) sulfateLOAEL = 24 mg/kgTlGeneral toxicity: increased ASTand ALT; weakness; sluggishness;alopecia; ptosis of the eyelids;diarrhea; respiratory difficulty;liver necrosis; kidney damageOsorio-Rico etal. (1995)Rat200–250 gMalei.p./single injection30 or 50 mg/kgthallium (I) acetateLOAEL = 23 mg/kgTlNeurological toxicity: increase inMAO and 5-hydroxytryptamine inthe brainWoods andFowler (1986)RatYoungadultMalei.p./single injection50, 100, and 200mg/kg thallium(III) chlorideLOAEL = 42 mg/kgTlLiver toxicity: increased volumedensity of mitochondria,lysosomes, and rough endoplasmicreticulum of the liver; decreasedcytoplasm in the liver; increasedMAO and ferrochelatase; changesin several liver enzymesAcute studies (3–10 doses)Brown et al.(1985)Rat250 gMalei.p./6 days4 or 8 mg/kg-daythallium (I) acetateLOAEL = 3.1 mg/kgday TlNeurological toxicity: increasedlipid peroxidation in the brain;increasedβ-galactosidase activityin the brain; behavioral changesGibson andBecker (1970)RatFetus; 8,9, and 10or 12, 13,and 14days ofgestationBothTransplacental viai.p. injection todam/3 days2.5 or 10 mg/kgday thallium (I)sulfateLOAEL = 1.9 mg/kgday TlDevelopmental toxicity: reducedfetal body weight; increase inhydronephrosis; increase inmissing or non-ossified vertebralbodiesGibson andBecker (1970)RatPregnantFemalei.p./3 days2.5 or 10 mg/kgday thallium (I)sulfateLOAEL = 1.9 mg/kgday TlGeneral toxicity: diarrhea;lethargy; irritability; poor hairluster; alopecia37Table 4-6. Thallium toxicity in animals via injectionSpeciesAge orweightSexRoute/exposureperiodDosesNOAEL/LOAELStudy type/effectReferenceHasan et al.(1977)Rat~150 gMalei.p./7 days5 mg/kg thallium(I) acetateLOAEL = 3.9 mg/kgday TlGeneral toxicity: anorexia; poorhair luster; diarrhea; difficultywalking; abnormal head rotation;lethargy; death; changes in Golgicomplexes and smooth cisternae/vesicles of hypothalamic neurons;decreased succinic dehydrogenaseand guanine deaminase activitiesin the brainHasan et al.(1978)Rat~150 gMalei.p./7 days5 mg/kg thallium(I) acetateLOAEL = 3.9 mg/kgday TlNeurological toxicity: decreaseddopamine, norepinephrine, and5-hydroxytryptamine in the brainHasan and Ali(1981)Rat~150 gMalei.p./7 days5 mg/kg thallium(I) acetateLOAEL = 3.9 mg/kgday TlGeneral toxicity: anorexia; poorhair luster; diarrhea; difficultywalking; abnormal head rotation;lethargy; increased lipidperoxidation; aggregation oflipofuscin granules in theperikarya of cerebellar neuronsHasan andHaider (1989)Rat~150 gMalei.p./6 days5 mg/kg thallium(I) acetateLOAEL = 3.9 mg/kgday TlNeurological toxicity: GSHKuperberg et al.(1998)Rat250–300 gMalei.p./5 days0, 0.1, 1.0, or5.0 mg/kg thallium(I) acetateLOAEL =0.08 mg/kg-day TlNeurological toxicity: difficultywalking and maintaining pressureon the hind paws; loss ofcoordination in motor activity;lethargy; reduced foodconsumption; distended bladder;decreased acetyl cholinesteraseactivity in the bladderSubchronic studiesGalván-Arzateet al. (2000)Rat200–250 gMalei.p./30 days0.8 or 1.6 mg/kgday thallium (I)acetateLOAEL = 0.6 mg/kgday TlNeurological toxicity: increasedlipid peroxidation in the brain3839SpeciesAge orweightSexRoute/exposureperiodDosesNOAEL/LOAELStudy type/effectTable 4-6. Thallium toxicity in animals via injectionReferenceKennedy andCavanagh(1977)CatNotspecifiedFemales.c./4–26 weeks2.3–4.5 mg/kg perweek thallousacetate(unclear if dosewas reported asthallous acetate oras thallium)0.32 mg/kg-day(unclear if dose wasreported as thallousacetate or as thallium)Dying back of central andperipheral sensory neurons; nomotor nerve fiber degeneration4.4.2. Cardiotoxicity Male Wistar rats (180 to 220 g) injected i.v. with 3 to 100 mg/kg thallium (I) sulfate, while under pentobarbital anesthesia, rapidly developed hypotension with the lowest blood pressures reached within 3 to 5 minutes (Lameijer and van Zwieten, 1976). Blood pressures dropped in a dose-dependent manner with dos es of 30–100 mg/kg causing a drop of 20–40%
from initial values and a maximum effect achieve d in the 50 to 100 mg/kg range. Thallium had a greater effect on the diastolic pressure. Afte r 10 minutes, animals injected with doses ranging from 3–40 mg/kg had blood pressures resembling those prior to thallium injection. The higher doses had a more permanent effect on blood pres sure. In addition to lower blood pressure, the rats had a dose-dependent decrease in heart rate with no maximum achieved. Rats treated with 100 mg/kg had heart rates that were one-third their pre- injection rates. The same effects were
observed in anesthetized cats in jected i.v. with monovalent thal lium but not when the thallium
was infused into the left vertebral ar tery (Lameijer and van Zwieten, 1976).

4.4.3. Neurotoxicity No studies of thallium neurotoxicity following exposures by the inhalation or dermal routes of exposure were iden tified. Manzo et al. (1983) demonstrated functional and histopathologic changes in periphe ral nerves in rats that received thallium sulfate in drinking water at a dose equivalent to approximate ly 1.4 mg/kg-day Tl (see Section 4.2.1.1). Study findings reported below involved administrati on of thallium compounds by injection (i.p., s.c., i.v.). A single i.p. injection of 20 mg/kg thallium (I) acetate in 8–12 (exact number per group not specified) adult male Sprague-Dawley rats (~300 g) resulted in a statistically significant ( p < 0.02) decrease in aspartate a nd taurine in the hippocampus 6 hours after treatment that was resolved by 24 hours (Ali et al., 1990). Glutam ine and taurine levels were statistically significantly increased ( p < 0.05) in the frontal cortex at 6 hours. While the glutamine returned to control levels by 24 hours, the taurine was still significantly elevat ed. A dose-dependent decrease in dopamine and muscarinic cholinergic receptor binding in caudate nucleus was not reported in these treated rats but was observed in caudate nucl eus incubated in vitro with thallium. The study report indicated that the e ffect was not observed 24 hours after the last subacute dose (5 mg/kg daily for 10 days, i.p.) of thallium (I) acetate in a separate study, but the results were not presented. The subacute st udy demonstrated a statistically significant ( p < 0.05) increase in dopamine, 3,4-dihydroxyphenyl acetic acid, and 5-hydroxytryptamine (5-HT [serotonin]) levels in the amygdala nucleus, as well as an increase in 5-HT in the hypothalamus. Dopamine, 3,4-dihydroxyphenylacetic ac id, and 5-HT remained simila r to control concentrations in the caudate nucleus, frontal cortex, and hippocampus.

40Kennedy and Cavanagh (1977) examined the neur otoxic properties of thallous acetate in 16 female cats. Cats were injected subcutaneously with thallous acetate in distilled water once per week (or in some cases every other week or every few weeks) at mean weekly doses of 2.3– 4.5 mg/kg per week for 4–26 weeks. (It was unclear if doses were reported as thallous acetate or thallium). Six cats died duri ng the study without opportunity fo r neurological assessment. Remaining animals were observed daily for clinic al signs. At interval s from 4–12 weeks after exposure was initiated, nine animal s were sacrificed and nervous sy stem tissues were subject to perfusion fixation; one cat was exposed week ly for 26 weeks. Ataxia and hypotonia were observed in nearly all the cats. Histopathologic examination revealed a distal degeneration confined to the central and peri pheral axons of primary sensory neurons. No motor nerve fiber degeneration was found. The inve stigators proposed that muscle tissue, which has the largest
reservoir of K + ions in the body, may serve as a sink for thallium ions such that thallium ions
never reach a concentration in the extracellu lar space sufficient to damage nerve fibers. Experimental evidence for this hypoth esis, however, was not available. A set of companion studies in newborn Wistar rats (Barroso-Moguel et al., 1996, 1990) demonstrated severe and progres sive lesions in nerve fibers following i.p. administration of thallium (I) acetate. In the first study (B arroso-Moguel et al., 1990), 32 mg/kg thallium (I) acetate was given to 15 newborn Wi star rats. Equal numbers were sacrificed at 24, 48, and 72 hours and on days 7 and 51 (three rats/time point) . Alterations in the capillary vessel walls of the brain, observed within the first hours after thallium inj ection, progressed to irregular thickened walls and fibrotic sclerosis, which obstructed the lumen by 51 days postexposure. Other changes in the brain began in a diffuse manne r with all sections affected. Cortical neurons developed the first and most intense lesions; by 51 days, the cortical neurons had mostly disappeared and lesions we re apparent in the cen tral grey nuclei. In a follow-up study (Barroso-Moguel et al., 1996), 16 mg/kg thallium (I) acetate was administered via i.p. injection to 20 newborn Wistar rats (10 each sa crificed at 8 and 50 days of age). General toxicity was manifested through diarrhea, progressive muscular atrophy, small body size, and persistent alopecia. Eight of the 20 thallium-treated rats died during the study. By 8 days of age, thallium-treated rats had interstitial edema between the myelin sheaths (causing separation of the nerve fibers), edema around some axons and within the myelin, and, in some cases, initial damage and degeneration of th e myelin sheaths. In addition, muscle fibers were thinner and showed signs of beginning progressive muscular atrophy. Hemorrhage, necrosis, and destruction of the st riation were present in some area s of the muscle. By 50 days of age, the rats developed nerve damage with pr ogressive disappearance of nerve fibers and granular, filiform, and amorphous inclusions in abnormal axons and collapsing myelin sheaths. At this time, the muscle fibers lost their transv erse striation. Other musc le fibers were observed

41to be atrophic, fragmented, and exhibiting hyaline degeneration and ini tial fibroblast reaction; further, some were infiltrated with phagocytic macrophages. Osorio-Rico et al. (1995) m easured MAO activity and 5-HT turnover rates in different regions of the brain in 127 male Wistar rats (200–250 g) 24 hours after an i.p. administration of 30 or 50 mg/kg aqueous thallium (I) acetate. Results demonstrated MAO was significantly increased ( p < 0.05) at 30 mg/kg in the midbrain ( 27.7% over controls) and pons (37% over controls) sections. MAO increases also were observed at 50 mg/kg in the midbrain (48% over controls) and pons (47%). 5-HT turnover was significantly increased in the pons (172% over controls; p < 0.001) after 30 mg/kg treatment and in the pons (166.7% over controls; p < 0.001) and midbrain (56% over controls; p < 0.01) after 50 mg/kg treatment. No significant changes were observed in the dopamine turnover rate. Subcutaneous injection of thallium (I) ac etate as acute (single dose of 20–50 mg/kg), subacute (2–3 weekly injections of 10–15 mg/ kg), or chronic doses (10–20 mg/kg, followed by weekly injections of 5 mg/kg or occasionally 2.5 mg/kg for up to 26 weeks) to male and female Sprague-Dawley rats (250–500 g) caused toxicity in all treatment groups (Herman and Bensch, 1967). Symptoms reported in all groups included diarrhea, marked weight loss, anorexia, and lethargy. Animals dosed acutely displayed the symptoms earlier than those on subacute or chronic dosing schedules. Chronically exposed animals also had hair loss (maximal at 2– 4 weeks after initial injection), irritability, and dragging of the hind limbs. Animals were sacrificed when signs of toxicity became apparent. In acutely exposed animals, the mitochondria of the brain were fre quently filled with an
overabundance of stacked mitochondria l cristae. Three of four an imals administered thallium (I) acetate subacutely had changes in the brain, in cluding occasional foci of perivascular cuffing with lymphocytes and hemosiderin-filled macropha ge, acute necrosis, and swollen histiocytelike cells. Peripheral nerves had occasional de nse bodies in an unmyelinated nerve plexus. Brain neurons had numerous lipofuscin bodies as did the neurons of the chronically exposed
animals. Twenty-four hours after a single dose of 25 mg/kg thallium (I) acetate, acetyl cholinesterase (AchE) activity was reduced in the hypothalamus and nucleus accumbens (NA) regions of the brain and the activity of c holine acetyltransferase (ChAT) activity was significantly ( p ≤ 0.05) increased (Kuperberg et al., 1998) . At 48 hours, the AchE in the hypothalamus was still significantly ( p ≤ 0.05) reduced, and the AchE in the NA region of the brain was back to control levels. AchE activity also was reduced in the duodenum and the sphincter-trigon region of the bladder following th is single high dose, while ChAT activity was significantly ( p ≤ 0.05) increased in the ileum, duodenum, and both regions of the bladder. After 48 hours, the AchE levels in th e duodenum and the sphincter-tri gon and detrusor regions of the bladder were still significantly ( p ≤ 0.05) reduced.

42Adult male Sprague-Dawley rats (2 50–300 g) administered 0.1, 1.0, or 5.0 mg/kg thallium (I) acetate (i.p.) daily for 5 days exhibite d difficulty walking and maintaining pressure on their hind paws, loss of coordination in motor activity, leth argy, and reduced food consumption (Kuperberg et al., 1998). Most of the rats (6/8) treated at the high dose died by 48 hours posttreatment. The only significant cha nges observed 24 hours after the last dose were
a reduction of AchE levels in th e NA region of the brain with the 1.0 mg/kg dose and an increase
in the AchE levels in the striatum and midbrai n of rats treated with 5 mg/kg. There was a decrease in AchE activity in the sphincter-trigon region of the bladder at 24 hours in all of the repeat-dose groups (i.e., 0.1, 1.0, and 5.0), which returned to control values by 48 hours.
Decreased bladder AchE levels were also observe d in the detrusor region of the bladder 24 hours after 1.0 or 5.0 mg/kg doses and 48 hours after the 0.1 mg/kg dose. The bladders of these rats
were distended and contained twice the am ount of urine seen in the controls. Brown et al. (1985) examined lipid peroxidatio n in the brain after thallium exposure. In this study, groups of male Sprague-Dawley rats (250 g) were administered da ily i.p. injections of 4 or 8 mg/kg thallium (I) acetate in water for 6 da ys. Controls received saline. Twenty-four hours after the final injection be havioral analysis was performe d and the following morning, rats were sacrificed. A dose-dependent increase in lipid peroxidation was observed in the cerebellum, brain stem, and striatum but not in the midbrain and hippocampus. The 8 mg/kg dose also caused a statistically significant ( p < 0.05) increase in the lip id peroxidation of the cortex. Beta-galactosidase activity followed a do se-dependent increase in the cerebellum, brain stem, and cortex. The 8 mg/kg dose al so caused a statistically significant ( p < 0.05) increase in beta-galactosidase activity in the midbrain a nd hippocampus. The beta-g alactosidase activity in the striatum was not statistically significantly changed by either dose. In general, thallium decreased the frequency of groo ming behavior while the frequenc y of exploratory and attention behaviors was increased. However, the changes were not dose dependent. Hasan et al. (1977) administ ered 5 mg/kg thallium (I) acetate i.p. for 7 days to albino male rats (weighing approximately 150 g). Contro ls received sodium acetate solution in equal
volumes with the same molar concentration. Cl inical symptoms included anorexia, failure to gain weight, irritability, tende rness during handling, poor hair lu ster, lethargy, diarrhea, dragging hind limbs, and fits of abnormal rotation of head and neck, curving the body. Eight of 55 treated rats died by day 7. Electron microscopy showed an increased incidence of well-developed Golgi complexes and curved conformation of smooth ci sternae and vesicles of the neurons in the hypothalamus. More significantly, there was a peculiar isolation of a xonal endings of the anterior hypothalamus by membranous circumferentia l lamellae, which appear ed to have arisen from the neighboring astrocytic processes. In addition, the succinic dehydrogenase and guanine deaminase activities in the cerebrum were signifi cantly decreased in thallium-treated rats. Protein levels, MAO, adenosine triphosphate, and protease levels in the cerebrum were

43unaffected. Mitochondrial succinic dehydrogenase also was decreased in the cerebrum of
thallium-treated rats. In a study by the same group of investigator s (Hasan and Ali, 1981), male Charles Foster rats (approximately 150 g) injected with 5 mg/k g thallium (I) acetate daily for 7 days showed clinical symptoms similar to those reported by Hasa n et al. (1977). All rats were lethargic after 4–5 days of treatment. Rats were sacrificed the day af ter the final dose (day 8), and their brains were removed and separated into sec tions. A statistically significant ( p < 0.001) increase in lipid peroxidation was reported in the cerebral he misphere, cerebellum, and brain stem by 49, 142, and 116%, respectively. Electron microscopy demons trated prominent aggregation of lipofuscin granules in the perikarya cerebellar neurons (c ell body of neurons in th e brain) in thalliumtreated rats that were hardly discernible in control rats. Co mparisons made with nickel- and
cobalt-treated rats demonstrated differences in the areas of increased lip id peroxidation. Nickel and cobalt both had the greatest impact on the br ain stem, whereas thalliu m had a greater effect on the cerebellum. Although Hasan and Ali (1981) could not relate this to the differences in behavioral observations, they di d note that nickel- and cobalt-tr eated rats were irritable and restless; these symptoms were not obser ved in the thallium-treated rats. Hasan and colleagues also examined the effects of thallium (I) acetate on
neurotransmitter levels and sulf hydryl groups in the brain. Charle s Foster rats (approximately 150 g) were administered 5 mg/kg thallium (I) ac etate i.p. for 6 days (Hasan and Haider, 1989) or 7 days (Hasan et al., 1978). Dopamine, norep inephrine, and 5-HT were reduced in the four sections of the brain examined (hypothalamus a nd limbic area, corpus striatum, cerebellum, and brain stem), but not all reductions were statisti cally significant. Dopamine was significantly reduced in the hypothalamus and limbic area (50%; p < 0.05) and corpus striatum (64%;
p < 0.01). Norepinephrine was reduced by 9–33%, de pending on the brain region, but none of these reductions were statistically significant. 5-HT was significantly reduced in the corpus striatum (53%; p < 0.001), cerebellum (36%; p < 0.05), and brain stem (66%; p < 0.001) (Hasan et al., 1978). Glutathi one was significantly ( p < 0.001) reduced in the cerebrum (56%), cerebellum (62%), and brain stem (74%), and sulfhydryl radicals were significantly ( p < 0.05) reduced in the cerebellum ( 25%) and brain stem (32%) (H asan and Haider, 1989). Galván-Arzate et al. (2000) administered 0.8 mg/kg (considered 1/40 of the median lethal dose [LD 50 ]) or 1.6 mg/kg (considered 1/20 of the LD 50 ) thallium (I) acetate in deionized water via i.p. injection for 30 days to male Wistar ra ts (200–250 g). Three days after treatments ended,
rats were sacrificed and their brains were dissected into five different regions (hypothalamus, cerebellum, frontal cortex, hippocampus, and cor pus striatum). In each region, with the exception of the cerebellum, significant ( p < 0.01) increases in thallium content were observed after administration of 1.6 mg /kg compared to administratio n of 0.8 mg/kg. There were no statistically significant differences in the depositi on of thallium within each region of the brain

44for each dose. The rate of lipid peroxidation, a marker of oxidative stress, was increased significantly ( p < 0.01) in the corpus striatum (182%) and cerebellum (130%) after treatment with 0.8 mg/kg. At 1.6 mg/kg, all five regions exhibi ted statistically signifi cant increases in lipid peroxidation over controls (cor pus striatum, 161% increase, p < 0.05; hippocampus, 114% increase, p < 0.01; hypothalamus, 100% increase, p < 0.01; cerebellum, 81% increase, p < 0.01; and frontal cortex, 80% increase, p < 0.05). The two regions affected at 0.8 mg/kg (i.e., corpus striatum and cerebellum) were not affected to a greater extent at the higher dose. Lipid peroxidation was measured to dete rmine if oxidative stress plays a role in thallium’s toxicity. The study authors concluded that additional studies need to be performed to establish the precise mechanism of neurotoxicity.

4.5. MECHANISTIC DATA AND OTHER STUDI ES IN SUPPORT OF THE MODE OF
ACTION Several, possibly related, mechanisms ha ve been postulated for the toxic action of thallium; however, the exact mechanism or mechanisms of to xicity are unknown.

4.5.1. Interference with Potassium Transport Monovalent thallium is similar to potassium (K + ) in ionic radius and electrical charge, which may contribute to its toxic properties.
Monovalent thallium has been demonstrated to have a 10-fold higher affinity for rabbit kidney Na + /K + -ATPase than does potassium (Britten and Blank, 1968). Barrera and GómezPuyou (1975) reported that monovalent thallium also inhibits the influx and efflux of potassium
in rat liver mitochondria at concentrations (10–15 nmol bound monovalent thallium per mg of mitochondrial protein) that do not affect oxidati ve phosphorylation. This inhibitory effect of thallium seemed to be specific for potassium si nce it did not affect the movement of sodium

. Monovalent thallium was completely equilibr ated by human red blood cells in 30 minutes in a high (140.5 mM)-sodium (Na + ) medium (Cavieres and Ello ry, 1974) but was equilibrated
even faster when the medium cont ained a low concentration (5 mM Na + ). In the high-Na + medium containing 1 mM external potassium, monovalent thallium caused a dose-dependent decrease in the ouabain-sensitive potassium infl ux. Monovalent thallium had a different effect in a medium containing 0.17 mM potassium; low (0.2 mM or less) monovalent thallium ion
concentrations stimulated the ouabain-sensitiv e potassium influx but inhibited it at higher
concentrations. Monovalent thal lium also had an inhibitory effect on the ouabain-sensitive sodium efflux. It was suggested that the e ffects on ouabain-sensitive sodium efflux and potassium influx are related to thallium’s high-affi nity substitution of potassium at the external potassium sites of the sodium pu mp, which is actively transpor ting monovalent thallium ions in while pumping sodium ions out.

45Tao et al. (2008) reported that thallium (I) could functionally replace potassium ion when applied in an in vitro system and that glutamat e transporters could inte ract with thallium (I).

4.5.2. Disturbance of Mitochondrial Function and Energy Generation Thallium may exert toxicity by disturbing mitochondrial function. Thallium (I) acetate caused an uncoupling of oxidative phosphorylati on and swelling of isolated mitochondria and induced an increase in oxygen consumption and l actic acid production in ascites tumor cells in vitro (IPCS, 1996).
There is evidence that thallium reduces th e available energy within peripheral nerves (resulting in a “dying back” type of neuronal degeneration) and other metabolically highly demanding cells (Kazantzis, 2007; Cavanagh, 1991). Cavanagh (1991) proposed that thallium in tissues produces a tissue deficiency of available riboflavin, leading to dist urbances of metabolic reactions dependent on flavoproteins. Among these reactions would be steps in the passage of a number of important intermediate metabolites in the electron transport chain. This metabolic disruption could lead to substa ntial impairment of energy pro duction in the cells. Cavanagh (1991) noted that support for this hypothesis includes the following: (1) riboflavin deficiency in the rat will lead to peripheral neuropathy and in the primate to loss of hair with circumoral skin lesions, and thallium toxicity in the primate produces a similar clinical picture to riboflavin deficiency; (2) the testis, which is peculiarly dependent on glucose as a substrate for energy metabolism, is sensitive to thallium toxicity; and (3) heart muscle, also known for high energy utilization, is also damaged as a re sult of chronic thiamine deficiency.

4.5.3. Induction of Oxidative Stress Other research suggests that thallium may trigger toxicity through induction of oxidative stress. Hanzel et al. (2005) investigated effects of thal lium (III) hydroxide on metabolism of GSH, which plays a key role in th e regulation of cell redox state, in an in v itro system using rat brain cytosolic fractions. Thallium hydroxide decreased the content of GSH and inhibited glutathione peroxidase and glutathione reductase activity, suggesting that thallium impairs the glutathione-dependent antioxida nt defense system. Using rat pheochromocytoma (P12) cells in vitro incubated with both thallium (I) nitrate and thallium (III) nitrate, Hanzel and Verstraeten (2006) found a concentration- and time-depende nt decrease in cell viability, decreased mitochondrial membrane potential, increased st eady-state levels of mitochondrial hydrogen peroxide (H 2 O 2 , a product of partial reduction of molecular oxygen whose generation is enhanced when electron transport is impaired), and GSH content. These investigators postulated that both ionic species of thallium enhance reactive oxygen species production in the cell, decreasing mitochondrial functional ity and cell viability. In a fo llow-up study with P12 cells in vitro, Hanzel and Verstraeten (2009) observed th at thallium (I) and (III) do not cause cell

46necrosis, but significantly increased the number of cells with apoptotic f eatures. The oxidatation state (i.e., I vs III) appeared to influence the apoptotic pathways are involved. Thallium (I)- mediated cell apoptosis was mainly associated with mitochondrial damage, whereas thallium (III) showed a mixed effect triggering both the intrinsic and extrinsic pathways of apoptosis. Galván-Arzate et al. (2005) i nvestigated the effect s of a single dose (8 or 16 mg/kg, i.p.) of thallium acetate on lipid peroxidation in different brain regions of Wistar rats (as an indicator of oxidative damage) and alterati ons in endogenous antioxidant systems. Lipid peroxidation was increased in three of five brain regions from animals that received 16 mg/kg thallium acetate at day 7 postexposure (but not at days 1 and 3); antioxidants GSH and superoxide dismutase showed only a modest depletion in only one or two brain regions.

4.5.4. Reaction with Thiol Groups The capacity of thallium to react with thiol groups, thereby interfering with a variety of processes, is postulated as another mechanis m of toxicity, although interference with the metabolism of sulfur-containing amino acids does not seem to be directly involved in toxicity (IPCS, 1996). Thallium (I) chloride formed co mplexes with a number of sulfur-containing amino acids (i.e., L-cysteine, DL-penicilla mine, N-acetyl-L-cysteine, and N-acetyl-DLpenicillamine) in aqueous solution (Bugarin et al., 1989). Because the thallium (I) complexes formed were weaker than those formed using dimethylthallium (III), the study authors concluded that this was unlikely to be the major mechanism of toxicity.

4.5.5. Other Endpoint-specific Mechanistic Data
4.5.5.1. Cardiotoxicity
Monovalent thallium ions caused a dose-d ependent decrease in the heart rate and contractile force of spontaneously beating atria of guinea pigs (Lameijer and van Zwieten, 1976).
At a concentration of 10 –3 M, monovalent thallium reduced the heart rate by approximately 60%.
When the isolated atria were el ectrically driven, monovalent th allium ions at doses up to 10 –3 M were not able to significantly decrease the amplitude of contra ction. Neither potassium (ranging from 2.4 to 9.4 mM) nor cocaine (10 –6 or 10 –5 M) were able to influence the reduction in heart rate or contractile force of spontaneously bea ting guinea pig atria cause d by monovalent thallium ions (0.5 mM) (Lameijer and van Zwieten, 1976). Isolated rat heart (from albino rats of both sexes) perfused with a nitrate-Krebs solution containing monovalent thallium ions (as thallium (I) nitrate) in place of potassium had a rapid decrease in beat frequency (Hughe s et al., 1978). The heart stopped completely in an average of 7 minutes. Placing the hearts in normal or nitrat e-Krebs saline allowed for some recovery in approximately 10 minutes. The mon ovalent thallium ions were st ill present in the heart tissue (9 mmol Tl + /kg wet tissue) after 30 minutes in thallium -free solution; this residual thallium

47caused the heartbeat frequency and amplitude to remain low. When only a portion of the potassium was replaced with monov alent thallium ions, there was a concentration ratio and time dependency on the reduction of th e heart rate. In a separate experiment, it was noted that injecting thallium (I) nitrate into the perfusion stream close to the heart caused an initial acceleration of the heartbeat followed by a reduction in amplitude until at certain concentrations the heart stopped. The heartbeat could recove r somewhat following nitr ate-Krebs perfusion, but the heartbeat frequency and amplitude did not return to initial values and were instead comparable to those values noted prior to the heart stopping. There was a dose-dependent effect on the length of the cardiac paralysis (5–25 μ mol monovalent thallium ions injected). The use of potassium nitrate instead of thallium (I) nitrate also caused a dose-dependent increase in the length of cardiac paralysis, but the length of pa ralysis was shorter than that with thallium (I) nitrate. Furthermore, potassium nitrate-treated hearts recovered completely when washed with nitrate-Krebs solution (Hughes et al., 1978).

4.5.5.2. Neurotoxicity Wiegand et al. (1984) recorded the freque ncies (reflects presynaptic processes) and amplitudes (reflects postsynaptic processes) of miniature endplate potentials (MEPPs) from neuromuscular junctions of ra t (strain not specified) phren ic nerve (of the diaphragm) preparations. Investigators repo rted a gradual increase in the fr equency of MEPPs by a factor of
10 within 30 and 180 minutes at concentrations of 1 × 10 –3 and 5 × 10 –4 M thallium (I) acetate, respectively, which was reversible. The amplit ude was unchanged. Therefore, it was concluded that thallium interfered presyna ptically with spontaneous transmitter release. A follow-up experiment using triangularis st erni muscles of adult mice (strai n not specified) demonstrated that, although thallium disturbed the presynaptic transmission in a manner similar to divalent metal cations, thallium (monovalent; compound used not specified) acted via a different
mechanism than either divalent cobalt or cad mium (Wiegand et al., 1990). This study also
demonstrated that thallium did not influence th e presynaptic potassium or calcium channels. Windeback (1986) used an in vitro model system to compare the inhibitory effects of four metals on neurite outgrowth. Metals were adde d to cultures of E15 rat embryo dorsal root ganglion neurons; neurite outgrow th was measured during the lin ear phase of growth after 24 and 40–80 hours. Compared to mercury and arse nic, which inhibited neurite outgrowth at very low concentrations (50% inhibition at 3.9 and 9.6 × 10 –6 M, respectively), neurons were relatively resistant to thallium (50% inhibition at 1.3 × 10 –4 M), suggesting that populations of neurons have different suscepti bilities to various metals. The basis for these different
sensitivities was not e xplored in this study. Hippocampal slices from adult guinea pigs or rats (strain not specified) were used to
examine the effects of thallium on central ne uronal activity (Lohmann et al., 1989). The study

48authors did not note any differen ces between the guinea pig and ra t results and appear to have combined the results. Light microscopy did not show any morphologic changes in thalliumtreated hippocampal slices even with a high concen tration (1–1.2 mM) for 6 hours. Thallium was determined to reversibly reduce the amp litudes of the compound action potential of CA1 pyramidal cells in a dose- and time-related manner. Thallium did not alter intracellular response parameters, indicating that membrane potential and input resistance we re not affected, but postsynaptic potentials were inhibited. The study authors concluded that in the hippocampal slices thallium reacts mainly with postsynaptic target sites and exerts an unknown influence on
intracellular metabolism of CA1 pyramidal cells (Lohmann and Wiegand, 1996; Lohmann et al., 1989). Diaphragms from male and female albino rats perfused with a nitrate-Krebs solution containing monovalent thallium (as thalliu m (I) nitrate) in place of potassium (K + ) had an initial (1–2 minute) increase in the contraction amplitude, followed by a steady decline in response (Hughes et al., 1978). Four experiments were performed with th e sequence taking 20– 40 minutes to block indirect (n erve) stimulation and 70–100 minutes to block direct (muscle) stimulation. After returning the diaphragms to normal nitrate-sa line, the block in response was reversed, but contraction amplitudes were 30% of the original response for both nerve and muscle after a 75-minute perfusion. The pres ence of monovalent thallium in the solution, whether as a replacement or an addition to pot assium, caused a dose dependency to the response. Diaz and Monreal (1994) examined the effects of thallium compounds (thallium (III) chloride, thallium (III) nitrate, and thallium (I) acetate) on proton and chloride permeabilities through myelin lipid bilayers using an in vitro system of liposomes prepared with lipids from brain myelin. Trivalent thallium, but not monovalent thallium, mediated a rapid chloride/hydroxyl ion exchange th rough the lipid bilayers. Trival ent thallium in the presence of reducing agents did not have the same reaction. The ion excha nge was faster with trivalent thallium than with mercury (Hg +2 ). In addition, the reaction o ccurred with a 10-fold lower concentration of trivalent thallium than of mercury.

4.5.5.3. Dermal Toxicity Arbiser et al. (1997) examined the effects of thallium acetate on three types of skin cells, human keratinocytes, primary endothelial cells , and melanoma cells, to determine whether thallium affected cell growth and differentiation in vitro. Inhibition of proliferation of all three cell types was observed. In melanoma cells, thallium caused dose-dependent decreases in cell dendricity and shape but not cellular motility. In normal human keratinocytes, thallium appeared to interfere with the normal progr am of cutaneous keratinization. In an in vivo study by the same investigators using piebald LPJ mice with both mela nin-rich and -poor areas in the same animal, one week administration of thallium acetate (5 mg /kg daily) by i.p. injection produced evidence

49of lipid peroxidation in skin in a perifollicul ar distribution (Arbiser et al., 1997). (The investigators noted that lipid peroxidation in vivo results in oxidation of lipid membranes, resulting in increased concentr ations of aldehydes, which can react with the Schiff reagent, thereby producing a colored product. ) It was suggested that lipi d peroxidation may result in cell death due to membrane damage and may pa rtly account for thallium-induced alopecia.

4.5.6. Genotoxicity Positive results were obtained for thallium (I) nitrate (1 mM) in the recombination-repair (Rec) assay using Bacillus subtilis strains H17 and M45; whether or not hepatic homogenates were used was not specified (Kada et al, 1980; Kanematsu et al., 1980). These positive results were obtained using “cold incubation,” which in creases the sensitivit y of the assay by 20– 50 times for many drugs. In this test, plates cont aining the bacteria with a 10 mm filter paper disk containing the metal solution (0.05 mL) were incubated at 4°C for 24 hours prior to being incubated at 37°C overnight. The differences in the inhibition of growth between the Rec + strain
and the Rec

– strain were measured. Thallium (I) nitrate was not mutagenic in reverse mutation assays using Salmonella typhimurium strains TA98, TA100, TA1535, TA1537, and TA1538 (histidine reversions) and Escherichia coli strains B/r WP2 tr – and WP2 her

–

tr – ; whether or not hepatic homogenates were used wa s not specified (Kanematsu et al., 1980). Negative results were obtained in a screening assay for the induc tion of mitogenic gene conversion and reverse mutation in the yeast Saccharomyces cerevisiae at a 0.1 M concentrati on of thallium (I) nitrate (Singh, 1983). Thallium (I) nitrate did not affect cell division in S. cerevisiae (isolated from baker’s yeast) and E. coli (strain B) but proved toxic to the aerobic growth processes of S. cerevisiae (Loveless et al., 1954) . A dose of 250 μ g/mL thallium (I) nitrate caused a 50% reduction in aerobic growth processes. The report did not iden tify specific doses of th allium (I) nitrate tested. The organisms were treated under conditi ons of logarithmic phase growth. S. cerevisiae was incubated for 4 hours with thallium (I) nitrate, and E. coli was incubated for 1.5 hours. The study authors noted that several of the other compounds tested (e.g., iodoacetamide) that were specific inhibitors of sulfhydryl groups also reduced the growth processes with no effect on
cellular division. A concentration of 1,000 μ g/mL thallium (I) acetate reduced viability of Chinese hamster ovary (CHO) cells in culture to 20% with a conc omitant decrease in DNA synthesis (i.e., 1% of control values) (Garrett and Lewtas, 1983). The EC 50 (effective concentration necessary to produce a 50% response) values were 307 μ g/mL for viability and 18 μ g/mL for DNA synthesis. Thallium acetate also depressed ATP and protein synthesis in culture. Single-strand DNA breaks occurred in cell cultures of C57BL/6 mouse and rat embryo fibroblasts exposed to thallium (I) carbonate at bo th concentrations tested in mouse fibroblasts

50(i.e., 10 –5 and 10 –4 M) and all three concentrations te sted in rat fibroblasts (i.e., 10 –6 , 10 –5 , and
10 –4 M) (Zasukhina et al., 1983). However, tha llium (I) carbonate did not induce single-strand
DNA breaks in CBA mouse fibroblas ts after treatment with 10 –4 –10 –6 M concentrations. Zasukhina et al. (1983) performe d a dominant lethal test on ma le white rats that received daily oral doses of thallium (I) carbonate (0.005–0.5 μ g/kg-day) for 8 months and were subsequently mated with untreated females. Female rats were sacrificed on day 20, and mutagenic potential was evaluated based on ev idence of embryotoxicity. The investigators reported an increase in embryonic death, suggestive of a dominant lethal effect. As reported previously (Section 4.3), however, confidence in this study is low. The number of resorptions was highest in the control group. In addition, methods were not adequately reported and results were not analyzed statistically.

4.6. SYNTHESIS OF MAJOR NONCANCER EFFECTS
4.6.1. Oral Thallium is readily absorbed through the GI tract and distributed throughout the organs and tissues of the body. Although thal lium is not metabolized, it occurs in two valence states. If or how the body modifies the valence state of thallium is unknown, but orally administered monovalent thallium and trivalent thallium app ear to be distributed in a similar manner throughout the body (Sabbioni et al., 1980a, b). Once thallium is di stributed, elimination occurs mainly in the urine and feces with the amounts in each varying by species. Most of the available human case reports ar e the results of poisonings, suicide attempts, or accidental ingestion of rode nticides. The lowest known dose to cause symptoms is a single dose of 0.31 g; the patient recovered after treatment (Cavanagh et al., 1974). The only studies of repeated oral exposure to thallium were two su rveys of populations exposed to thallium through contaminated homegrown foods (D olgner et al., 1983; Brockhaus et al., 1981). Limitations in these epidemiology studies included the lack of objective tests for t oxicity, reliance on the incidence of symptoms obtained from questionn aires, and characteriza tion of chronic thallium exposure by measuring the levels in urine and hair at a single point in time. Three studies of occupationally exposed populations (Ludolph et al., 1986; Marcus, 1985; Schaller et al., 1980) did not conclusively establish an association between thallium exposure and impaired health status; however, all three studies were limited in terms of size of th e study population and study design. Symptoms of thallium toxicity are diverse in both humans and animals. The triad of gastroenteritis, polyneuropathy, a nd alopecia has been regarded as the classic syndrome of thallium poisoning (IPCS, 1996), although not all th ree of these effects are observed in all poisoning cases, and other sympto ms develop in varying sequen ce depending on the magnitude and duration of thallium exposure.

51The nervous system as a target organ of thallium is supported by observations from human case reports and animal st udies. Relatively high doses of thallium cause neurological symptoms in humans (e.g., paresthesia of the hands and feet, weakness, tremors, coma, and convulsions) (see Table 4-1). Some of these neurological symptoms (e.g., paresthesia and weakness) were reversible, althoug h recovery was slow. Other eff ects, including mental and/or psychological problems, were more persistent . Neurological sympto ms have also been associated with chronic exposure to thallium in humans. These symptoms include sleep disorders, tiredness, weakness, nervousness, headache, psychological alterations, and neurological and muscular problems. Although paresthesia of the hands and feet ar e trademark symptoms of thallium toxicity, it is generally alopecia that leads to a diagnos is of thallium poisoning in humans. Alopecia occurs about 2 weeks after exposur e and is reversible af ter exposure to thallium is discontinued. Alopecia has also been repeatedly observed in experimental animals exposed to thallium compounds. Thallium exposure in humans has been associated with respiratory effects and GI effects, including diarrhea and vomiting. Other toxic effects associated with or al thallium exposure in humans and animals are changes in blood pressure (high, low, and fluctuating values have all been noted) and liver and kidney damage (kidney damage is age dependent and occurs only in mature kidneys), all of which appear to be reve rsible with the removal of thallium exposure.
Doses that do not affect survival have been shown to affect clinical chemistry parameters such as ALT, AST, BUN, blood glucose, and blood sodium levels, indicating liver and kidney damage with subchronic exposures (Leung and Ooi, 2000; Appenroth et al., 1996, 1995; Fleck and Appenroth, 1996; El-Garawany et al., 1990; Mourelle et al., 1988). In experimental animal studies, thallium e xposure has been associated with biochemical changes, lipid peroxidation, and histopatholog ic changes in the br ain and functional and histopathologic changes in periphe ral nerves (see Section 4.4.3). Th e areas affected in the brain differ with the age of the treated animal; neve rtheless, all measured endpoints (symptoms, biochemical measurements, and histopathology) indi cate that high doses (c lose to lethal doses) of thallium induce significant degradation of the nervous system. Results from in vitro studies
further confirm these observations. Despite the fact that the ne rvous system is a known target of thallium toxicity, studies using standard measures of neurobe havioral toxicity have not been performed. In a 90-day oral gavage study, MRI (1988) observed ra ts daily for clinical signs. Rats exposed to thallium sulfate showed consistently increased incidences of clin ical observations relate d to the coat (rough coat, piloerection, shedding, and alopecia), eyes (lacrimation, exophthalmos, and miosis), and behavioral signs compared with untreated or ve hicle controls. The underl ying mode of action for
these clinical observations is not known. Collectively, however, these observations suggest a

52treatment-related effect of thallium on the rat and possibly an indirect measure of stress or other effects on the nervous system. For example, it has been suggeste d that barbering (or overgrooming) in rodents may represent a stress -evoked behavioral respon se or other nervous system dysfunction (Welch et al ., 2007; Kalueff et al., 2006; Kalu eff and Tuohimaa, 2005; Greer and Capecchi, 2002). Thallium salts have been shown to affect reproductive function. A dose as low as 0.7 mg/kg-day Tl (10 ppm of thallium (I) sulfate) resulted in testicular damage and reduced sperm motility in male Wistar rats within 60 da ys (Formigli et al., 1986). Wei (1987) reported that doses as low as 0.001 mg/L in the drinking water for 6 months in Kunming mice reduced sperm motility (rapid speed only), and 0.01 mg/L reduced overall sperm motility and sperm counts and caused a reduction of live offspring. However, confidence in this study is low due to the non-reporting of several obs ervations (water intake, body we ights, behavior, and animal health). In addition, the nu mbers of animals examined for sperm and reproductive endpoints were ambiguously reported. Half of the animal s were sacrificed at the end of the exposure period, while the other half were allowed to ma te for 1 week and then sacrificed. The study results for male mice used for analysis of sperm and reproductive endpoints were reported together. That is, results for sperm count, sperm mobility, and all other epididymal sperm analysis for males sacrificed at the end of the exposure period were combined with the results for
males that were allowed to mate and thus permitted a one-week rec overy period prior to sacrifice. In addition, at the initiation of the study each group c onsisted of 20 animals. In the reporting of the results, however, sperm data we re missing for 4 mice from each of the control, 0.001, 0.01, and 1 mg/L groups, for 8 mice from th e 0.1 mg/L group, and for 10 mice from the 10 mg/L group. No explanation for the loss of male mice (ranging from 20 to 50%) is provided. Limited data in humans and experimental animals suggest that thallium may produce developmental toxicity. A revi ew of case studies of women e xposed orally to high levels (approximately 120–1,100 mg) of thallium during pr egnancy suggested a trend toward premature and low-birth-weight infants, es pecially if exposure took place in the last trimester; no other developmental abnormalities were identified (Hof fman, 2000). Dolgner et al. (1983) examined birth defects in a German population living near a cement plant emitting thallium dusts during the mid-1970s and found a higher incidence of c ongenital malformations than the incidence documented in the government birth records from the area. The association between the number of birth defects and thallium exposure was w eak, however, because two of the malformations were considered hereditary and the incidence of birth defects, alt hough greater than that determined from civil records, was consistent with that reported in the lit erature. Confidence in this study was limited by lack of exposure data during pregnancy and po ssible underreporting in
controls. In vivo data in rats support an association between i.p. thallium exposure and low birth

53weight, although such an associatio n has not been reported with or ally administered thallium.
In vitro data demonstrated an increase in bone malformations in both rat and chick embryos.

4.6.2. Inhalation There are currently no studies that examine the effects of inhaled thallium. A few case reports (Hirata et al., 1998; Ludolph et al., 1986 ) suggest an associati on between occupational exposure and toxicity (including al opecia, GI symptoms, and neuropa thy), but the route or routes of exposure in these workplace setting could not be established. A study of a population living near a cement factory emitting thallium (Dolgner et al., 1983) determined that thallium exposure occurred via consumption of plants grown in thal lium-contaminated soil to a greater extent than
via inhalation.

4.6.3. Mode-of-Action Information The underlying mode of action for clinical ly observed effects of thallium has been studied. Verstraeten (20 06) determined that Tl 1+ can oxidize membrane fatty acids, causing an increased fluidity in the membrane and an increa sed concentration of cellu lar oxidants associated with thallium toxicity. Ensuing axonal degenera tion characterized by a decrease in density of large myelinated fibers and loss of epidermal ne rves, indicating small se nsory nerve involvement in thallium toxicity, has been reported (Kuo et al ., 2005). The skin experiences parakeratosis and vacuolar degeneration of the basa l layer, resulting in a loss of epidermal nerves and persistent damage to sensory nerve endings (Lu et al., 200 7). Collectively, these observations suggest a
treatment-related effect of th allium on the rats and direct effects on the nervous system. Both potassium and thallium are monovale nt cations with similar atomic radii (Tl + : 1.50 Å; K + : 1.38 Å) (Ibrahim et al., 2006). Thallium has been shown to replace potassium in the

reaction of Na + /K + -ATPase (Barrera and Gómez-Puyou, 1975; Britten and Blank, 1968) and to mimic the biological actions of potassium. Mono valent thallium has been shown to have a 10-fold higher affinity than potassium for Na + /K + -ATPase and thus replaces potassium as a substrate for this enzyme (Barrera and Góme z-Puyou, 1975; Britten and Blank, 1968). In other
studies it caused a decrease in Na + /K + -ATPase activity in the liver and kidney (Yoshida et al., 1997; Mourelle et al., 1988). After a single oral or i.p. dose of thallium (10 mg/kg orally or 25 mg/kg i.p.), the disruption of Na + /K + -ATPase activity was found to be reversible.
In addition to effects related to interferen ce with potassium transport and function, there is evidence that thallium uncouples oxidativ e phosphorylation, adversely affects protein synthesis, reduces the available energy within peripheral nerve a xons, and inhibits a number of enzymes, including alkaline phosphatase and succinic dehydrogenase (Kazantzis, 2007). Thallium’s activity as a Lewis acid with an affinity for organosulfur compounds (Lewis bases) may account for its adverse effect on ha ir production. Keratin is the primary protein

54found in hair. It is rich in the amino acid cystei ne and its low solubility is, in large part, the product of the formation of inter-polypept ide cysteine-cysteine cross-links during posttranslational modification of th e nascent polypeptides. Thalli um prevents keratinization of hair proteins by binding with cy steine and preventing the form ation of the cross-linking bonds (Mulkey and Oehme, 1993), a property that may be related to the alopecia observed in humans and animals following thallium exposure. Binding to cysteine may also account for inhibition of enzymes with active site cysteine residues and increases oxidative stress as a result of GSH modification (Mulkey and Oehme, 1993).

4.7. EVALUATION OF CARCINOGENICITY
Under EPA’s Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005a), there is “inadequate information to assess the carci nogenic potential” for thallium and thallium compounds. There are presently no st udies that evaluate the carcinog enic potential of thallium in animals and no adequate studies of huma ns chronically exposed to thallium.
Two studies of chronic hea lth effects in workers exposed to thallium are available (Marcus, 1985; Schaller et al ., 1980), but these studi es are inadequate for the assessment of carcinogenicity. The study by Marcus (1985) is limited by the examination of medical records only, lack of exposure quantitation, small cohor t size, and the unknown length of observation. Schaller et al. (1980) identified health effects in a worker po pulation at a single time point through medical histories and physical examina tions for unspecified symptoms. Worker exposures to thallium were limited to a single measure of urinary thallium, which would not provide an adequate measure of past exposure. This health evalua tion was not adequate to detect any carcinogenic response. Relatively few studies have examined th e genotoxicity of thallium compounds; these studies provide inconsistent evidence for genot oxicity. Positive results were obtained at 0.001M for thallium (I) nitrate in the Rec assay using B. subtilis strains H17 and M45 (Kanematsu et al., 1980). However, negative results were obtaine d in reverse mutation assays using several S. typhimurium and E. coli strains and mitogenic gene convers ion and reverse mutation tests in yeast. Cytotoxic levels (1,000 μ g/mL) of thallium (I) acetate caused depressed DNA synthesis in CHO cells (Garrett and Lewtas , 1983). Single-strand DNA breaks occurred in C57Bl/6 mouse and rat embryo fibroblasts exposed to thalliu m carbonate but not in similarly exposed CBA mouse fibroblasts (Zasukhina et al., 1983). A dose of 200 mg th allium sulfate caused a slight increase in SCEs in peripheral blood lymphocyt es taken from a 48-year-old man on day 1 and day 15 postexposure and caused a 3.5-fold increase in binucleat ed cells with micronuclei (Hantson et al., 1997).

554.8. SUSCEPTIBLE POPULA TIONS AND LIFE STAGES
There are little or no data available to establish a particular subpopulation as being particularly susceptible to the to xic effects of thallium compounds.

4.8.1. Possible Childhood Susceptibility Exposures in human case reports generally are poorly characterized. Therefore, no comparison can be made between children and adults with regard to susceptibility. In rats, doses of thallium that caused maternal toxicity have be en demonstrated to affect the developing fetus (Gibson and Becker, 1970). The only studies that examined the toxic effects of thallium at different ages were those of Appenroth et al . (1996) and Fleck and Appenroth (1996), which focused on the age-related effects on nephrotoxic ity. In these studies, mature rats were determined to be more susceptible to kidney damage than young (10- or 20-day-old) rats, because mature kidney function appeared necessary for thallium to adversely affect the kidney.

4.8.2. Possible Gender Differences Leloux et al. (1987), MRI (1988), and Downs et al. (1960) are the only available toxicity studies of thallium compounds that used both ma le and female rats. Leloux et al. (1987) administered only a single lethal dose of thalli um (I) nitrate to rats and thus did not provide findings useful for discerning pos sible gender differences. Downs et al. (1960) reported slight differences in thallium (III) oxide toxicity betwee n the sexes, with male s dying earlier, exhibiting greater and more severe alope cia, and having more profound decreases in body weight than females. No sex-related differences in respons e were noted in rats tr eated with thallium (I) acetate (Downs et al., 1960). No marked differen ces in response were noted in male and female rats after exposure to thallium (I) sulfate (MRI , 1988); however, high-dose female rats exhibited a higher incidence of alopecia than males, a nd hair follicle atrophy was observed in females only. Overall, the limited data do not identif y any consistent pattern of gender-related differences in response to thallium exposure.

565. DOSE-RESPONSE ASSESSMENTS

5.1. ORAL REFERENCE DOSE (RfD) 5.1.1. Choice of Principal Study and Critical Effect—with Rationale and Justification As discussed in Section 4.6.1., most informa tion on thallium toxicity in humans comes from poisonings, suicide attempts, or accidental e xposures. Epidemiologic studies of either the general population or occupatio nally exposed groups are limited by inadequate study design and/or insufficient exposure char acterization. Thus, available hum an studies do not provide data useful for dose-response analysis. There are only four repeat-dos e oral toxicity studies of thal lium that used more than one dose level: MRI (1988), Wei (1987), Zasukhina et al. (1983), and Do wns et al. (1960). Of these studies, the subchronic (90-day) toxicity study of thallium (I) sulfate in Sprague-Dawley rats by MRI (1988) is the most comprehensive study of th allium toxicity. In the Downs et al. (1960) study of thallium (I) acetate, mortality in two control groups was 30–40%, complicating interpretation of the findings in treated rats (mortality and al opecia). A study of thallium (III) oxide by the same investigators (Downs et al., 1 960) failed to identify a NOAEL. Further, death
in 2/5 female rats at the lowe st dose tested in this study wa s observed and may have been treatment related. The repeat-dose dominant le thal study of Zasukhina et al. (1983) suffered from critical reporting deficiencies and internal inconsistencies (see Section 4.3). Wei (1987) reported effects on sperm count, motility, and viab ility in Kunming mice exposed to drinking water concentrations of thallium (I) carbonate as low as 0.001 mg/L; however, when the treated males were mated with untreated females, the reproductive index and number of implantations were not affected at drinking water concentrations up to 10 mg/L (10,000-fold higher than the concentration identified by the st udy investigators as the LOAEL fo r sperm effects). The percent of dead fetuses was significantly lower than in the control group at c oncentrations of 0.001, 0.01, and 0.1 mg/L but was increased at the two highest concentrations (1 and 10 mg/L). Although the effects on sperm appeared to be dose related, some uncertainty is associated with the reporting of study findings (see Sections 4.3 and 4.6.1). Further, the unexplained absen ce of reported sperm
results for 20–50% of the male mice in each of th e treatment groups lowers confidence in these findings. Supporting literature on th e reproductive toxicity of thallium is limited. Formigli et al. (1986) reported adverse effects on the testes and sperm of rats exposed to thallium sulfate in drinking water but at a drinking water con centration 10,000-fold higher than the LOAEL reported by Wei (1987). In the ab sence of confirmatory findings of sperm effects at the low drinking water concentrations used in the Wei (1987) st udy, the study findings were not considered sufficiently reliable to serve as the basis for RfD derivation.

57Other repeat-dose studie s of thallium oral toxicity used study designs that included only a single-dose group and thus did not provide data useful for dose-re sponse analysis (El-Garawany et al., 1990; Rossi et al., 1988; Fo rmigli et al., 1986; Gregotti et al., 1985; Manzo et al., 1983). Manzo et al. (1983) reported mo rtality at the only dose tested , making this study unsuitable for RfD derivation. Finally, doses in mg/kg-day coul d not be estimated from the study information provided in Rossi et al. (1988). Based on the above considerations, the 90- day MRI (1988) study was selected as the principal study for derivation of the RfD. In the MRI (1988) study, rats (20/sex/group) were treated by gavage daily for 90 consecutive da ys with 0, 0.01, 0.05, or 0.25 mg/kg-day of an aqueous solution of thallium (I) sulfate (approximately 0, 0.008, 0.04, or 0.20 mg/kg-day Tl). No differences in body weight, body weight gains, food consump tion, or absolute and relative organ weights were observed am ong control groups and groups receiving thallium sulfate. Lacrimation, exophthalmos, and miosis were observ ed at higher incidences in the treated male and female rats at all doses compared with bot h untreated and vehicle controls (see Table 4-2); however, ophthalmologic examination and gross a nd histopathologic examin ation of the eyes
revealed no treatment-related abnor malities. The incidence of clinic al observations related to the coat and skin (including rough coat, piloerect ion, shedding, and alopecia) and behavioral changes (including combined incidences of a ggression, tense/agitated behaviors, hyperactivity, vocalization, and self-mutilation) were also elevated in treated male and female rats at all doses (see Table 4-2). The investigators attributed most , but not all, instances of alopecia to barbering behavior. For example, of the 12 high-dose fema les with alopecia, 5 instances were not totally attributed to barbering. Review of individual animal data reveal ed no discernable differences in either the severity or distri bution pattern of alopecia acro ss control and treated groups. Histopathologic examination did not reveal any stat istically significant treatment-related effects, although it was noted that atrophy of the hair follicles occurred in two high-dose female rats. Tissue samples of the skin from low- and mi d-dose groups, however, were not examined for histopathologic changes. Subtle , but statistically significant, ch anges were observed in several blood chemistry parameters (AST, LDH, sodium and blood sugar levels) (see Table 4-4). These changes were not confirmed by any histopathologic findings a nd were not considered by the investigators to be toxicologically significant. Although the study authors id entified the highest dose ( 0.20 mg/kg-day Tl) as a NOAEL based on lack of biological significance of the observed effects, EPA reached the determination that histopathologic findings in the skin of fema le rats and dose-related clinical findings, when considered collectively, may reflect an a dverse effect on the health of the rats. As noted above, histologic examination of skin samples from two high-dose females showed atrophy of hair follicles. These two animals also exhibited alopecia. Hair loss (alopecia) is characteristic of thallium poisoning in humans and experimental animal s (Ibrahim et al., 2006;

58Galván-Arzate and Santamaría, 1998), and typically occurs in humans within two weeks of exposure. It is hypothesized that thallium’s affi nity for sulfhydryl groups may be responsible for alopecia; thallium prevents keratinization of ha ir proteins by binding with cysteine. Skin biopsies have been taken from a limited number of patients with alopecia and other symptoms of thallium poisoning; these biopsies have revealed atrophic and necrotic changes of the skin (Lu et al., 2007; Heyl and Barlow, 1989; Saddique and Peterson, 1983). For example, skin biopsy findings from two patients who ingested water th at contained thallium included parakeratosis,
dilated hair follicles filled with keratin and necrotic sebaceous materials, mild epidermal atrophy, and vacuolar degeneration of the basal layer (Lu et al., 2007). Thus, the finding of two cases of atrophy of the hair follicles in high-dose female ra ts with alopecia is consis tent with the atrophic changes observed in cases of hu man thallium poisoning, and suggest s that alopecia at the highdose (0.2 mg/kg-day Tl) may be related to thalliu m exposure. This endpoint was considered a
candidate critical effect fo r derivation of the RfD. The clinical observations from the MRI (1988) study were also considered as candidate
critical effects for derivation of the RfD. As discussed in Section 4.6.1, dos e-related increases in the incidence of lacrimation, exophthalomos, and miosis, coat-related fi ndings, and behavioral observations suggest a possible effect on the he alth of the exposed rats from exposure to thallium, although the underlying basis for these observations is unknown. Increased incidences of barbering and behavioral cha nges are consistent with possible effects on the nervous system, a known target of thallium. Clinical chemistry changes were not consider ed as the basis for determining the point of departure (POD) for the RfD. At 90 days, AST, LDH, sodium, and sugar levels in the blood of high-dose male and female rats differed by +31, +38, +4, and -21%, respectively, from vehicle control group values. Even with these changes, these clinical chemistry parameters are all well
within two standard deviations of the mean for control Sprague-Dawley rats as reported by Petterino and Argentino-Storino (2006) and Matsuzawa et al. (1993). In the low-dose group, none of the blood chemistry parameters was statis tically significantly different from the vehicle control group with the exception of the sodium level in female rats, which was increased over control values by only 1.4%. In light of these modest changes in blood chemistry parameters and the lack of other findings in exposed animals to confirm the biological significance of these changes, clinical chemistry parameter data were not selected for dose-response modeling. In summary, two endpoints were considered as potential critical effects for determination of the POD for the RfD: (1) hair follicle atr ophy in female rats that also had alopecia, and (2) clinical observations, incl uding those related to animal coat (rough coat, piloerection, shedding, and alopecia), eyes (including lacrimat ion, exophthalmos, and miosis), and behavior.

595.1.2. Methods of Analysis Hair Follicle Atrophy The NOAEL-LOAEL approach was used for dos e-response analysis of hair follicle atrophy in rats with alopecia. A benchmark dose (BMD) analysis was not conducted because the incidence of histopathologically-d etermined hair follicle atrophy was not considered amenable to BMD methods. Histopathological examination of the skin was performed for two groups only— the high-dose group and vehicle c ontrol. Two of 20 female rats in the high-dose group (10%) had hair follicle atrophy a nd alopecia that is consistent with th allium toxicity in both animals and humans and was thus characterized by EPA as possibly treatment-related. The high dose (0.2 mg/kg-day Tl) was therefore ch aracterized as a LOAEL. Because skin tissue from rats in the low- and mid-dose groups was not examined for histopathologic changes, the NOAEL for this endpoint cannot be determin ed with certainty. Given the low incidence of hair follicle atrophy in females in the high-dose group and absence of cases of hair follicle atrophy in male rats, the mid-dose can reasonably be assumed to approximate a NOAEL for skin histopathology. Thus, an estimated NOAEL of 0.04 mg/kg-day Tl was used as the POD for hair follicle atrophy from the MRI (1988) study.

Clinical Observations BMD modeling (U.S. EPA, 2000b) was used to analyze the clinical observation data from MRI (1988). Incidence data for the selected clinical observations in male and female rats
are summarized in Table 5-1. All of the ava ilable dichotomous models in U.S. EPA’s BMDS (version 1.4.1) (U.S. EPA, 2007) were fit to these incidence data.

60Table 5-1. Incidence data and BMD modeling resul ts for selected clinical observations in Sprague-Dawley rats treated with thallium sulfate for 90 days a

Dose (mg/kg-day)

BMD 10 b (mg/kg-day) BMDL 10 b (mg/kg-day) Untreated

control Vehicle control Observation 0.008 0.04 0.2

Male

Coat/skin Rough coat 1/20 3/20 11/20 16/20 19/20 BMD and BMDL well below experimental
range. d

Piloerection 0/20 0/20 1/20 4/20 13/20 0.020 0.013

Shedding 0/20 0/20 4/20 10/20 8/20 All dichotomous models in BMDS exhibited statistically significant lack of fit. e Alopecia 2/20 1/20 4/20 9/20 4/20 All dichotomous models in BMDS exhibited statistically significant lack of fit. e
Eyes

Lacrimation 1/20 6/20 19/20 20/20 20/20 BMD and BMDL well below experimental
range. d
Exophthalmos 1/20 5/20 12/20 20/20 20/20 BMD and BMDL well below experimental
range. d

Miosis 0/20 1/20 5/20 7/20 15/20 0.0069 0.0039

Behavior c 3/20 0/20 7/20 6/20 7/20 All dichotomous models in BMDS exhibited statistically significant lack of fit. e

Female

Coat/skin Rough coat 1/20 0/20 1/20 5/20 11/20 0.021 f 0.013 f

Piloerection 0/20 0/20 0/20 3/20 8/20 0.043 f 0.026 f
Shedding 0/20 0/20 2/20 3/20 13/20 0.020 0.013 Alopecia 4/20 1/20 4/20 9/20 12/20 0.018 f 0.010 f
Eyes

Lacrimation 7/20 6/20 20/20 20/20 20/20 All dichotomous models in BMDS exhibited statistically significant lack of fit. e Exophthalmos 5/20 6/20 19/20 20/20 20/20 BMD and BMDL well below experimental
range. d

Miosis 2/20 3/20 1/20 11/20 8/20 All dichotomous models in BMDS exhibited statistically significant lack of fit. e

Behavior c 2/20 2/20 0/20 1/20 7/20 0.14 f 0.054 f

61Table 5-1. Incidence data and BMD modeling resul ts for selected clinical observations in Sprague-Dawley rats treated with thallium sulfate for 90 days a

Dose (mg/kg-day)

Observation BMD 10 b (mg/kg-day) BMDL 10 b (mg/kg-day) Untreated

control Vehicle control 0.008 0.04 0.2

a Listed as number of animals with the sign observed at least once during the 90-day study.
b BMD 10 = Benchmark dose correspond ing to a 10% extra risk; BMDL 10 = 95% lower bound on the dose corresponding to a 10% extra risk.

c Animals exhibiting one or more behavioral observations at least once during the 90-day study, including the following: aggressive, tense/agitated, hyperactive, vocalization, self-mutilation.
d Data set not amenable to modeling because of steep slope of the dose-response curve in low-dose region; BMD and BMDL well below experimental range.

e Some data sets shown in this table exhibited a statistically significant lack of fit with the models available in BMDS. Where lack of fit is due to characteristics of the dose-response data for high doses, one option sometime used is to adjust the data set by eliminating the high-dose group(s) (U.S. EPA, 2000b). In the absence of a mechanistic understanding of the biological response or other observed toxicity in the tha llium-exposed animals that could explain the dose response at the higher doses, dose-response assessment with adjusted data sets (i.e., removing the high-dose group(s)) was not performed.
f Reported BMD 10 and BMDL 10 represent averages of results from several similar model fits. See Appendix B for more details.

Source: MRI (1988).

Consistent with EPA (2000b) BMD technical guidance, consideration was given to identifying biologically relevant response levels for developing the RfD. A BMR of 10% is generally used to facilitate a consistent basis of comparison ac ross assessments and was used for these endpoints in the absence of information regarding the level of change considered to be biologically significant. Doses (i.e., BMD 10 [BMD corresponding to a 10% extra risk] and
BMDL 10 [95% lower bound on the BMD corresponding to a 10% extra risk]) associated with a
BMR of 10% extra risk are presented in Table 5-1. Details of the BMD modeling conducted fo r each endpoint presented in Table 5-1 are provided in Appendix B. In gene ral, model fit was assessed by a chi-square goodness-of-fit test (i.e., models with p < 0.1 failed to meet goodness-of-fit crit erion) and visual inspection of the respective plots of observed versus predicte d values from the various models. BMDL 10 estimates from these models that were within a factor of three of each other suggested no appreciable model dependence. Fitted models exhibiting adequate fit (i.e., p ≥ 0.1) with Akaike Information Criterion (AIC) values within two units of the lowest AIC were considered indistinguishable from one another (B urnham and Anderson, 2002), and thus BMD 10 and
BMDL 10 values from these models were averaged. Model fits that yielded the same mathematical model were counted as a single mo del for averaging purposes (see Appendix B for details). As Table 5-1 shows, clinical observation data sets for fema le rats were generally more amenable to BMD modeling than the male rat data sets. The BMDL 10 values for endpoints

62related to the coat/skin in female rats ra nged from 0.010 to 0.026 mg/kg-day, a range of only
2.6-fold. The lowest BMDL 10 value for female rats (0.010 mg/kg-day) comes from averaging
the BMDL 10 values from several similar model fits of incidence data for alopecia. As discussed in Section 5.1.1, alopecia is a hallmark of thallium poisoning. To the extent that instances of alopecia resulted from barbering, the occurrence of alopecia could also be consistent with possible effects of thallium on th e nervous system, although no direct evidence for neurotoxicity was provided. In male rats, the smallest BMDL 10 (0.0039 mg/kg-day) is base d on data for miosis. While elevated in thallium-treated animals, the bi ological significance of miosis is less clear than
the collection of effects related to the rats coat an d skin. Review of individual animal data from the MRI (1988) study revealed that at the high dose, miosis was observed between days 6 to 10 only, and in the majority of animals (approxima tely 70%) miosis was observed on only one or two days.

5.1.3. RfD Derivation—Including Applic ation of Uncertainty Factors (UFs)
5.1.3.1. Soluble Thallium Salts: Acetate, Carbon ate, Chloride, Nitrate, and Sulfate The principal study considered for Rf D derivation – MRI (1988) – involved administration of thallium (I) sulfate. There ar e no studies of thallium (I) acetate, thallium (I) carbonate, thallium (I) chloride, or thallium (I) nitrat e that are appropriate to use as the basis for an RfD. For the following reasons, it was cons idered appropriate to treat these monovalent thallium salts as toxicologically equivalent to thallium (I) sulfate when expressed in terms of thallium. It is likely that the mechanism of toxicity is the same for these salts due to the fact that they all contain monovalent thallium ions and are water soluble. Only small differences in the toxicity of various water-soluble thallium (I) salt s exist in mice, rats, rabbits, and dogs. In general, for most laboratory species at an obs ervation period of approximately 1–2 weeks, the

LD 50 or minimum effective dose values range between 10 and 30 mg/kg body weight for thallium (I) salts, independent of the exposure route (IPCS, 1996). Therefore, the use of thallium (I) sulfate as a surrogate for the other th allium salts is considered appropriate.
Candidate RfD values for thallium were de rived using PODs for two endpoints from the MRI (1988) study – a NOAEL of 0.04 mg/kg-day Tl for hair follicle atrophy and an average

BMDL 10 of 0.01 mg/kg-day Tl for alop ecia (as representative of clinical observations more generally). Some degree of uncertainty is associated with both PODs. Hair follicle atrophy was reported in the MRI (1988) study on ly in high-dose female rats. Because skin tissue from the low- and mid-dose groups was not examined histopathologically, the NOAEL for this endpoint could only be estimated. The biol ogical significance of the dose-re lated clinical observations in rats in the MRI (1988) study is unclear. In general, the collection of clinical observations data is

63relatively subjective and less rigor ously measured than endpoints su ch as clinical chemistry or histopathology. While c linical observations repo rted in the MRI (1988) study could represent an
indirect measure of stress or possible effects on the nervous syst em, no direct support for these etiologies is available. The increased incidence of alopecia in female ra ts was dose-related (less clearly so in male rats); howev er, the background inci dence of alopecia in control animals and attribution of some cases of alopecia by the st udy authors to barberi ng behavior introduce uncertainty in dose-response analysis of this e ndpoint. Furthermore, ex amination of individual animal data from the MRI (1988) study revealed no discernable difference in the severity or distribution of alopecia across cont rol and treated groups. Effects on the eyes based on clinical observations (in particular lacrimation and exophthalmos) were not supported by ophthalmic examination or by gross or histopat hological examination of the eyes. A total uncertainty factor (UF) of 3,000 ( 10 for interspecies extrapolation, 10 for intraspecies extrapolation, 3 fo r extrapolation from a subchroni c to a chronic study, and 10 for database deficiencies) was applied to both PODs to estimate candidate oral RfD values for
soluble thallium salts.

• A default interspecies UF of 10 was applie d for extrapolation from laboratory animals
to humans. No information was availabl e to characterize th e toxicokinetic or toxicodynamic differences between e xperimental animals and humans.

• A default intraspecies UF of 10 was app lied to account for variation in human susceptibility in the absence of information on the variabil ity of response to thallium in the human population.

B ecause no chronic toxicity studies for thalliu m are available, an UF of 3 was applied to account for extrapolation from subchroni c to chronic exposure duration. Oral toxicity data for thallium s uggest that an UF of 10 w ould overestimate the difference in response following subchroni c and chronic oral exposures . Effects on the coat/skin
as well as other clinical observations occur within weeks of exposure to thallium (i.e., these sensitive effects do not require chr onic exposure in order to manifest).

• An UF for LOAEL to NOAEL extrapolation was not needed for ha ir follicle atrophy in rats with alopecia because a NOAEL was estimated from data provided in the principal study.

S imilarly, a UF to account for extrapolat ion from a LOAEL to NOAEL was not used for alopecia (as representative of clinical observations) because BMD methods were

64used for dose-response analysis of these endpoints. The current approach is to address this extrapolation as one of the considerations in selecting a BMR for BMD modeling. In this case, a BMR of 10% in crease in the incidence of alopecia was selected under the assumption that it represents a mini mally biologically significant change.

• The thallium database includes several subchr onic oral toxicity studies in rats. Studies of reproductive and developmental t oxicity of thallium co mpounds in rats and mice are available; however, these studies used nontraditional study designs that did not provide adequate testi ng of reproductive or developmental endpoints. In reproductive toxicity studies by Gregotti et al . (1985) and Formigli et al. (1986), male rats only were exposed for periods up to 60 days and evaluation of reproductive toxicity was limited to examination of ma le reproductive organs. In two other reproductive toxicity studies, male rats (Zasukhina et al., 1983) or mice (Wei, 1987) only were exposed to thallium compounds for 6 to 8 months and mated with untreated
females. Confidence in these latter two st udies was low (see Sections 4.3 and 4.6.1). No studies of reproductive toxicity of thallium in females or multigeneration reproductive toxicity studies were identified. Developmen tal toxicity by the oral route was limited to Rossi et al. (1988) in which rats were exposed to thallium sulfate from gestation day 1 through 60 days of age or from birth through 60 days of age. No investigation of developmental endpoints at the end of the gestation period was performed. Despite the limitations in th e available reproductive and developmental toxicity studies, these studies provide suggestive evidence that thallium compounds can adversely affect male reproductive orga ns and the developing fetus and highlight
the deficiencies in the current thalli um database. Limited neuropathologic
examinations were included in the subc hronic toxicity studie s by MRI (1988) and Manzo et al. (1983), but no standard neurobehavioral studies were identified. Because the nervous system is a sensitiv e target of thallium toxicity, the limited investigation of thallium neurotoxicity represents a data deficiency. Thus, a database UF of 10 was applied to account for a lack of adequate developmental toxicity studies and a two-generation reproductive toxici ty study, and additional uncertainty
associated with the limited data available on neurotoxicity in light of the potential for neurotoxicity to represent a sensitive endpoint for thallium exposure.

T hus, the candidate RfDs for thallium (I) are calculated as the POD (NOAEL or BMDL) divided by the composite UF, as follows:

65Derivation of candidate RfD based on hair follicle atrophy in animals with alopecia: 0.04 mg/kg-day Tl ÷ 3000 = 1 x 10 -5 mg/kg-day Tl

Derivation of candidate RfD based on clinical observations/alopecia: 0.01 mg/kg-day Tl ÷ 3000 = 3 x 10 -6 mg/kg-day Tl

The available toxicity databa se for thallium contains stud ies that are generally of poor quality. The MRI (1988) study that was selected as a candidate principal study suffers from certain critical limitations (e.g., high background incidence of alopeci a, lack of histopathological
examination of skin tissue in low- and mid-dose groups, and inad equate examination of objective measures of neurotoxicity), and there are particular difficulties in the selection of appropriate endpoints. Therefore, even though an RfD would generally be derived with a combined uncertainty factor of 3000, an RfD for soluble tha llium salts is not derived in this specific case.

5.1.3.2. Insoluble Thallium Salts: Thallium (III) Oxide No oral studies of thallium (III) oxide were found that are adequate to support derivation of an RfD. Downs et al. (1960) administered thallium (III) oxide to Wist ar-derived albino rats
via the diet for 15 weeks at concentrati ons of 0, 20, 35, 50, 100, and 500 ppm. The study reported only body weights, mortality, kidney wei ghts, and limited histopathology. Because all relevant endpoints were not evaluated, treatment group sizes were small (five per group per sex), and few animals survived to termination, this st udy could not be used to derive an RfD. Because of differences in the physical-che mical properties of thallium (I) sulfate and thallium (III) oxide, toxicity info rmation for thallium (I) sulfate cannot be used to inform the toxicity of thallium (III) oxide. Unlike thalliu m (I) sulfate, thallium (III) oxide is insoluble in water. Thallium (I) sulfate c ontains monovalent thallium (Tl +1 ), while thallium (III) oxide contains trivalent thallium (Tl +3 ). Although the gastric environment may increase the solubility of thallium (III) oxide, no studies are available th at examine the effects of the gastric pH and gastric environment on the valence state of thallium (III) oxide. Limited evidence from the toxicology lit erature suggests that the distribution of thallium (I) and thallium (III) compounds and the lethality of these two compounds may be comparable. Sabbioni et al. (1980a) determined that, following oral administration of either inorganic monovalent thallium (Tl +1 ) as thallium (I) sulfate or trivalent thallium (Tl +3 ) as thallium (III) chloride, a similar distribution of th allium in the tissues was seen (valence state could not be determined) at 16 hours and 8 days after administration, indi cating that the valence
state of thallium did not affect tissue distribution (and presumably uptake). Downs et al. (1960) demonstrated similar oral 7-day LD 50 values for thallium (I) acetate (32 mg/kg Tl) and thallium (III) oxide (39 mg/kg Tl) in female rats, indicatin g that lethality may be independent of valence

66state. Whether other endpoints would respond similarl y to different valence states of thallium is unknown. Monovalent thallium compounds have been determined to behave similarly to potassium (K + ), thus disrupting Na +/ K + -ATPase and the systems depe ndent on this transporter (e.g., liver and kidney). Trivalent thallium has no t been demonstrated to behave in the same manner. A single in vitro study of mono- and tr ivalent thallium compou nds (Diaz and Monreal, 1994) suggested that biological responses to thallium in the (I) and (III) valence states may differ. Using an in vitro system with liposomes prepared with lipid from brain myelin, these investigators reported that trivalent thallium, but not monovalent thallium, mediated a rapid chloride/hydroxyl ion exchange through the lipid bilayers. Overall, the available toxicity informati on for thallium (III) oxide specifically and thallium compounds more generally is insufficient to support derivation of an RfD for thallium (III) oxide.

5.1.3.3. Thallium (I) Selenite No toxicity studies of thallium (I) selenite are available. Thallium (I) selenite contains
monovalent thallium as does tha llium (I) sulfate. No information could be found in the literature, however, on the water solubility of th allium (I) selenite. In th e absence of solubility information, toxicity information for thallium (I) sulf ate cannot be used to inform the toxicity of thallium (I) selenite. Accordingl y, the available data do not support derivation of an RfD for thallium (I) selenite.

5.1.4. Candidate RfD Comparison Information
PODs and candidate RfD values based on se lected studies included in Table 4-5 are arrayed in Figures 5-1 to 5-5 and provide pers pective on the magnitude of PODs and candidate
RfD values associated with differe nt toxicity endpoints. These fi gures should be interpreted with
caution because the PODs across studies are not nece ssarily comparable, nor is the confidence in
the data sets from which the PODs were derived the same. PODs in these figures may be based on a LOAEL or a BMDL, and the nature, severit y, and incidence of effects occurring at a LOAEL are likely to vary. To some extent, the confidence associated with the data sets is reflected in the magnitude of the total UF applie d to the POD (i.e., the si ze of the bar); however,
the text of Sections 5.1.1 and 5.1.2 should be cons ulted for a more complete understanding of the
issues associated w ith each data set. Thallium causes toxicity in a wide range of target organs, including the nervous system, kidney, cardiovascular system, liver, skin, re productive system, and possibly the developing fetus. Figure 5-1 provides a graphical display of dose-response information for skin histopathology (hair follicle atrophy in female rats with alopecia) based on the 90-day MRI (1988) study. Uncertainties in this da ta set are discussed in Section 5.1.3.1.

67Figure 5-2 provides a graphical display of dose-response information for clinical observations from the 90-day study conducted by MRI (1988). Collectiv ely, the dose-related increased incidences of effects on the coat, ey es, and behavior suggest a potential treatmentrelated effect of thallium, po ssibly including some measure of stress or other effect on the nervous system; however, the underlying basis for these clinical observa tions is unknown. It should be noted that Figure 5-2 includes PODs derived using BMDL 10 values as the POD for
those data sets for which a model fit could be obtained from the models in BMDS, as well as a reference value derived using the LOAEL as the POD (since some data sets were not fit by the models in BMDS). Uncertainties in thes e data sets are disc ussed in Section 5.1.3.1. In some studies, doses of thallium that do not affect survival have been shown to affect clinical chemistry parameters (e.g., BUN, serum creatinine, b ilirubin, and ALT) that may be indicative of effects on the kidney and liver. Cha nges in clinical chemistry parameters observed in El-Garawany et al. (1990) are pl otted in Figure 5-3. It should be noted that application of a composite UF of 10,000, reflecting five areas of uncertainty, indicates that derivation of a reference value using such a data set is highl y uncertain. In such cases, U.S. EPA (2002)
recommends that an RfD not be derived. Evidence from experimental animal studies su ggests that thallium may produce effects on reproductive function. Figure 5-4 in cludes plots of LOAELs and a ssociated UFs for data sets from Formigli et al. (1986) and Wei (1987). Form igli et al. (1986) reported testicular effects in rats exposed to thallium sulfate for 60 days but used only a single dose level of thallium. Therefore, this study has limited utility for dose-response analysis. Wei (1987) found effects on sperm in mice exposed to thallium carbonate at drinking water concentrations as low as 0.001 mg/L. As discussed in Sections 4.6.1 and 5.1.1, confidence in this study is low because of uncertainties associated with estimates of expos ure, reporting inconsistencies, and unexplained absence of sperm data for male mice in all dos e groups (ranging from 20 to 50%). Composite UFs of 100,000 and 10,000 were applie d to the LOAELs from the Fo rmigli et al. (1986) and Wei (1987) data sets. As with the data from El -Garawany et al. (1990), the magnitude of the uncertainty associated with these data sets i ndicates that they are insufficient to support
derivation of a reference value. In humans, numerous case reports of neur opathy following thallium poisoning have been reported. In experimental animals, effects on the nervous system have been most clearly observed following oral e xposure at levels also associated wi th increased mortality (e.g., Manzo et al. [1983]) or following injection exposure (see Section 4.4.3). Therefore, although the nervous system is known to be a target of thal lium toxicity, the availabl e studies do not provide adequate dose-response information on standard measures of nervous system toxicity at nonlethal doses that are appropriate for defining a POD. Figure 5-5 displays PODs for the major targ ets of toxicity (from Figures 5-1 to 5-4) associated with oral exposure to thallium, including skin histopathology, alopecia and other

6869 general clinical observations, changes in clinical chemistry possibly indicative of the kidney and liver as targets of thallium toxi city, and the reproductive system , along with the UFs that would be applied for candidate RfD derivation.0.0000010.000010.00010.0010.010.1MRI (1988); 90-d rat study (gavage); hair follicle atrophy; NOAEL

Dose (mg/kg-day Tl)zPODUF--animalto humanUF--humanvariationUF--LOAEL toNOAELUF--subchronicto chronicUF--databasedeficienciesDerivedcandidate RfDFigure 5-1. POD (mg/kg-day) with corresponding derived candidate reference valuethat would result if histopathologicchanges of the skin (hairfollicle atrophy) were usedas the critical effect.701.0E-071.0E-061.0E-051.0E-041.0E-031.0E-021.0E-01 MRI (1988); 90-d rat study (gavage); coat/skinpiloerection/male; BMDL MRI (1988); 90-d rat study (gavage); eyes-miosis/male; BMDL MRI (1988); 90-d rat study (gavage); coat/skin-rough coat/female; BMDL MRI (1988); 90-d rat study (gavage); coat/skinpiloerection/female; BMDL MRI (1988); 90-d rat study (gavage); coat/skinshedding/female; BMDL MRI (1988); 90-d rat study (gavage); coat/skinalopecia/female; BMDL MRI (1988); 90-d rat study (gavage); behavior/ female; BMDL MRI (1988); 90-day rat study (gavage); other clinical endpoint data sets not fit by BMDS; LOAEL

Dose (mg/kg-day Tl)UF--animalto humanUF--humanvariationUF--LOAEL toNOAELUF--subchronicto chronicUF--databasedeficienciesDerivedcandidate RfDzPODFigure 5-2. PODs (mg/kg-day) with corresponding derived candidate reference values thatwould result if clinical observationsfrom MRI (1988) were usedas the critical effect.711.0E-051.0E-041.0E-031.0E-021.0E-011.0E+00El-Garawany et al. (1990); 90-d rat study (gavage); clinical chem; LOAEL

Dose (mg/kg-day Tl)UF--animalto humanUF--humanvariationUF--LOAEL toNOAELUF--subchronicto chronicUF--databasedeficienciesDerivedcandidate RfDzPODFigure 5-3. POD (mg/kg-day) with corresponding derived candidatereference value that wouldresult if clinical chemistrychanges (suggesting the liver or kidney as a target) were used as the critical effect.721.0E-081.0E-071.0E-061.0E-051.0E-041.0E-031.0E-021.0E-011.0E+00Formigli et al. (1986); 60-d rat study (DW);testicular effects; LOAELWei (1987); 6-month mouse study (DW);sperm effects; LOAEL

Dose (mg/kg-day Tl)zPODUF--animalto humanUF--humanvariationUF--LOAEL toNOAELUF--subchronicto chronicUF--databasedeficienciesDerivedcandidate RfDFigure 5-4. PODs (mg/kg-day) with corresponding derived candidate reference values that would result if reproductivetoxicity endpoints were usedas the critical effect.73741.0E-081.0E-071.0E-061.0E-051.0E-041.0E-031.0E-021.0E-011.0E+00Skin histopathology Behavior/stress/generalhealthReproduction Clinical chemistry (liver,kidney)

Dose (mg/kg-day Tl)Figure 5-5. PODs (mg/kg-day) with corresponding derived candidate reference values that would result if alternativeendpoints were used asthe critical effect.UF--animalto humanUF--humanvariationUF--LOAEL toNOAELUF--subchronicto chronicUF--databasedeficienciesDerivedcandidate RfDzPOD5.1.5. Previous RfD Assessment The previous IRIS RfD values for thallium (I) acetate, thallium (I) carbonate, thallium (I) chloride, thallium (I) nitrate, and thallium (I) sulf ate were posted to the IRIS database in September 1988. These assessments were based on the same principal study considered for the principal study in the current assessment (MRI, 1 988). (The principal study was previously cited as U.S. EPA [1986c]. MRI subsequently issued a revised final report in 1988, which is the basis for the candidate RfD values developed in the current assessment. There are no substantive differences in the findings and conclusions between the 1986 and 1988 versions of the MRI report.) Previous RfD values (adjusted for di fferences in molecular weight) ranged from 8 × 10 –5

to 9 × 10 –5 mg/kg-day. These RfD va lues were based on a NOAEL of 0.25 mg/kg-day thallium sulfate, the highest dose tested by MRI (1988), and application of a composite UF of 3,000 (10 to extrapolate from subchronic to chronic data, 10 for intraspecies extrapolation, 10 to account for interspecies variability, and 3 to account for lack of reproductive and chronic toxicity data). Although both the previous and current assessments for soluble thallium salts (acetate, carbonate, chloride, nitrate, and sulfide) identified the same study as principal, the assessments differed in terms of the interpretation of the MRI (1988) findings and method for dose-response analysis. Most significantly, the current assessment doe s not recommend a value for the RfD.

5.2. INHALATION REFERENCE CONCENTRATION (RfC) Information on the inhalation toxicity of thallium is insufficient to derive an inhalation RfC. No studies of inhaled thallium in experi mental animals were id entified, and occupational epidemiology studies involving possible inhala tion exposures to thallium were limited and
inconclusive.

5.3. CANCER ASSESSMENT
There are no human or animal studies av ailable that are adequate to assess the carcinogenic potential of thallium salts.

756. MAJOR CONCLUSIONS IN THE CHARACTERIZATION OF HAZARD
AND DOSE RESPONSE

6.1. HUMAN HAZARD POTENTIAL Thallium is well absorbed from the GI tract, skin, and respiratory tract and is distributed throughout the body. Thallium, as an element, is not metabolized, but the extent to which it may
be converted from one valen ce state to another (i.e., Tl +1 and Tl +3 ) in the body is not known. Excretion of thallium occurs mainly through th e urine and feces, but the amounts are species dependent. Humans mainly excret e thallium through the urine, but it also has been detected in the hair, sweat, tears, and breast milk. Thallium salts cause a wide spectrum of adverse effects in humans and animals. Alopecia is an effect that is characteristic of thallium expos ure. Alopecia generally occurs within 2 weeks of exposure and is reversible when thallium e xposure is removed. Only one epidemiologic study reported a ne gative correlation between thalli um exposure and hair loss (Brockhaus et al., 1981); however , this study lacked measures of chronic exposure to thallium and was limited by reliance on questionnaires to determine symptomology in thallium-exposed
individuals. Some study observations suggest that the ne rvous system may be the primary target organ for thallium salt toxicity. Neurotoxicity has been observed afte r a single dose of 0.31 g thallium acetate in an adult male or 1 mg/kg th allium (I) nitrate for 4 days in adult rats. A variety of neurological effects have been repo rted in humans, includ ing lethargy, back pain, paresthesia of the hands and feet, weakness (including facial weakness), inability to walk, and prolonged mental defects. Some of the effects are reversible depending on the severity, while others are irreversible and may require long-term care. Animal studies support these findings in
humans. Kidney damage in humans has been noted by increases in BUN levels and serum
creatinine and is reversib le with treatment and/or discontinue d exposure to thallium. Data from animal studies support those in humans with rega rd to kidney damage; thallium-related effects include increased or decreased urine output (d epending on dose), protein in the urine, and changes in electrolyte balance. Histopathol ogic examination of thallium-exposed animals revealed changes in kidney morphology, includ ing atrophied and vac uolated kidney tubules, amorphous material in the lume n of the proximal tubules, di sorganized brush borders, and thickening ascending limb of the loop of Henle. Na + /K + -ATPase activity in the medulla also was significantly reduced. None of these changes we re dose related and were generally seen with large doses of thallium. Animal studies also suggest that kidney toxicity requires mature kidney function. The subchronic (90-day) toxicity study in rats used to derive the RfD showed moderate increases in AST and LDH, which are genera l indicators of tissue damage (MRI, 1988).

76However, a specific relation to possible kidney da mage was not indicated due to the lack of changes in other clinical chemistry parameters (i.e., BUN and creatinin e) and histopathologic changes in the kidney. Cardiotoxicity findings in thallium-exposed humans are variable. Many case reports indicate hypertension, while a fe w reported hypotension. Animal studies indicate that thallium
affects heart rate and causes a decrease in blood pressure, which also can be related to kidney effects and is supported by in vitro results. Many of the case reports in humans repor ted increased ALT and/or AST levels, indicating liver toxicity. These effects returned to normal after patients received medical care for thallium exposure. Various indicators of liver damage, including increases in ALT, AST, serum bilirubin, lipid peroxidation, tr iglycerides, and serum alkalin e phosphatase and decreases in glycogen, glutathione, and liver Na + /K + -ATPase, have been reported in animal studies. In addition, histopathology revealed sw ollen and vacuolated cells a nd swollen mitochondria. Many
of these effects were reversible in animals after a single dose of thallium. Statistically significant
increases in AST and LDH were observed in the 90-day MRI ( 1988) toxicity study in rats; however, the increases remained within the range reported in control rats and were not accompanied by histopathologic changes of the live r or changes in relate d clinical chemistry parameters (i.e., ALT). Low birth weight is a likely adverse effect of thallium exposure in females (humans and animals) exposed during pregnancy. Male mice e xposed to thallium were reported to have low sperm counts, low sperm motility, and increases in the number of deformed sperm. Testicular effects observed in animals included disarrange ment of the tubular epithelium, cytoplasmic vacuolation and distention of smooth endoplasmic reticulum of the Sert oli cells, and reduced beta-glucuronidase activities. Mating exposed male mice to unexposed female mice appeared to
cause a decrease in the number of live fetuses and an increase in the overall rate of dead fetuses; however, critical information concerning the doses of thallium admi nistered makes it difficult to
assess these results. There are no human studies relating thallium exposure to developmental toxicity. A literature review of pregnant women exposed to thallium during various stages of pregnancy reported an association with low birth weight. A survey of ch ildren born near a cement plant emitting thallium found an increase in congenita l malformations over those reported to the government; however, the study author s did not consider these malfor mations to be related to thallium exposure because two of the cases were cons idered attributable to hereditary factors and the incidence was similar to that reported in th e literature. Because of various study limitations, the findings are considered inconclusive. In ra ts exposed transplacentally and/or via mother’s milk, then via the drinking water until 60 days of age, thallium (1 mg/dL in the drinking water of dams’ then offspring) affected bone developm ent and vasomotor reactivity. Chick embryos

77exposed to thallium developed achondroplasia; these data furthe r support the pot ential role of thallium in the disruption of bone development. There are no studies available to determine the carcinogeni c potential of thallium in animals and no adequately conducted studies in hu mans. The limited number of studies of the genotoxicity of thallium compounds provides inconsistent evidence of genotoxic potential.
Under EPA’s Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005a), the Agency
concluded that there is “ina dequate information to assess the carcinogenic potential.”

6.2. DOSE RESPONSE The 90-day oral toxicity study of thallium (I) sulfate in Sprague-Dawley rats (MRI, 1988) was selected as a potential pr incipal study. PODs for two alte rnative endpoints from the MRI (1988) study were selected for dose-response analysis – (1) hair follicle atrophy in female rats with alopecia, and (2) clinical observations, including those rela ted to animal coat (rough coat, piloerection, shedding, and alopecia), eyes (inc luding lacrimation, exophthalmos, and miosis), and behavior. Using hair follicle atrophy, the hi gh dose (0.2 mg/kg-day Tl) was identified as a LOAEL; the mid dose (0.04 mg/kg-day Tl) wa s considered a NOAEL and was used as a
candidate POD. For clinical observations, an average BMDL 10 of 0.01 mg/kg-day Tl was derived using BMD modeling methods. A total uncertainty factor of 3,000 (10 for in terspecies extrapolati on, 10 for intraspecies extrapolation, 3 for extrapolation from a subc hronic to chronic study, and 10 for database deficiencies) was applied to the PODs to yield candidate RfD values for thallium in the form of
soluble thallium salts of 1 × 10 -5 mg/kg-day Tl (for hair follicle atrophy) or 3 × 10 -6 mg/kg-day Tl (for clinical observations). The available toxi city database for thalliu m contains studies that are generally of poor quality. The MRI (1988) study that was sele cted as a candidate principal study suffers from certain critical limitations (e.g., high background incidence of alopecia, lack of histopathological examination of skin tissue in low- and mid-dose groups, and inadequate examination of objective measures of neurotoxicity ), and there are particular difficulties in the selection of appropriate endpoint s. Therefore, even though an RfD would generally be derived with a combined uncertainty factor of 3000, an Rf D for soluble thallium sa lts is not derived in this specific case. The available data are not adequate to deri ve an RfD for thallium (III) oxide or thallium (I) selenite. Inhalation toxicity studies are not available to support derivation of RfCs for any thallium compounds.

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Summary of External Peer Review a nd Public Comments and Disposition

The Toxicological Review of Thallium and Compounds (U.S. EPA, 2009) 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 th e 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 re sponses 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.

EXTERNAL PEER REVIEW PANEL COMMENTS
EPA revised the Toxicological Review to present dose-response analyses for two data sets from MRI (1988), hair follicle atrophy and clin ical observations, and derived candidate RfDs taking the peer reviewers recomm endations into account. However, as pointed out by the peer reviewers, the MRI (1988) study that was selected as a principal study suffers from certain critical limitations (e.g., high background incidence of alopecia, lack of histopathological
examination of skin tissue in low- and mid-dose groups, and inad equate examination of objective measures of neurotoxicity), and there are particular difficulties in the selection of appropriate endpoints. For these reasons, an RfD for soluble thallium salts was not derived in this specific case. Text explaining the basis for EPA’s deci sion was added to Section 5.1.3.1. This decision is considered consistent with the peer review ers observations that the toxicity database for thallium is weak. EPA has provided responses to the reviewers’ comments in detail below,
because the derivations of the candidate RfDs were retained in the assessment. 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.

I. General Comments
1. Is the Toxicological Review logical, clear, and concise? Has EPA accurately, clearly, and objectively represented and synthesized the sc ientific evidence for noncancer and cancer
hazard?

Comments : Four reviewers considered the Toxicological Review to be clearly wr itten and/or to present a critical evaluation of th e published literature. One of th ese reviewers stated that despite

A-1considerable shortcomings in the overall databa se for thallium (which are acknowledged in the report), the authors of the revi ew did a good job at synthesizing the available information and presenting it in a logical fashion. One reviewer observed that the synthesis was somewhat less than ideal in that it was interjected with editorial comments (some from original authors) and that it might have been more desirable to present the facts and leave the interpretation to the reader.
This reviewer also believed that the Toxicological Review did not provide a sufficient review of the foreign language material or of environmental surveys. One reviewer stated that the reasons
for EPA’s reevaluation of the data used as th e basis for the RfD currently on IRIS and the different conclusion regardi ng the NOAEL and LOAEL from th e MRI (1988) study were not fully explained. This reviewer did not consid er the comparison of the LOAEL in the MRI study
with LOAELs from other studies th at were three- to ninefold hi gher to be a “logical argument.”
One reviewer noted that EPA’s case for thallium-induced alopecia could have been strengthened if the Toxicological Review had included a discussion of the hair growth cycle in
rats and how a clinical conditi on such as alopecia can be dis tinguishable from hair loss associated with normal growth cycle in rats. One reviewer stated that the MRI (1988) study
could have been better described and ev aluated. Two reviewers criticized the Toxicological
Review for rejection of studies (inc luding Zasukhina et al. [1983] and Wei [1987]) that may have provided useful information for reasons that were inadequate. One reviewer noted that the title Toxicological Review of Thallium and Compounds was a bit unclear and confusing and suggested as potential alternatives: Toxicological Review of Thallium and Its Salts or Toxicological Review of Thallium and Thallium Compounds

. Other comments were provided in response to this general charge question that were repeated in response to other more specific charge questions. In these cases, the peer reviewer comments (and EPA’s responses) were summarized under the more specific charge question.

Response : The general structure of a toxicological review is to present a factual summary of toxicity studies in Section 4 and critical interpretation/synthesis in Section 5. The study authors’ conclusions were presented in Sections 4 and 5 only where necessary (e.g., where it was necessary to document that EPA’s interpreta tion of the study findings differed from the authors’.) The comment regarding insufficient cons ideration of the foreign language literature is addressed under genera l charge question 2. The health effects of thallium exposure were reevaluated in light of the availability of new data and the need for an up-to-date asse ssment for regulatory purposes. The comment
related to determination of the NOAEL and LO AEL from MRI (1988) is considered further under charge question A3. Comparison to LOAELs from other studies was removed. Alopecia was one of a number of potential cr itical effects considered for dose-response analysis. The POD derived from alopecia usi ng BMD modeling was consistent with the PODs
derived from other related clinical effects. EPA did not consider a discussion of the hair growth

A-2cycle in rats and how alopecia can be distingu ished from normal cyclical hair loss to be

necessary. The Toxicological Review was revised to include an e xpanded discussion of the findings from MRI (1988) (see Section 4.2.1.1). The studies by Zasukhina et al . (1983) and Wei (1987) were critically reevaluated. EPA again reached the conclusion that the Zasukhi na et al. (1983) study suffered from critical
deficiencies and did not provide a scientifically defensible ba sis for the oral RfD. More thorough justification for this determination was added to Section 4.3. The discussion of the Wei (1987) study was revised (Section 4.3 and entry in Table 4-5) . In light of the uncertainties associated with incomplete reporting and re porting inconsistencies and lack of confirmation of these lowexposure findings on sperm, the Wei (1987) st udy was not used as the principal study (see discussions in Sections 4.6.1 and 5.1.1). Nevert heless, the study was included in Section 5.1.5, Candidate RfD Comparison Information. See also the response to charge question A.1 for additional response to this comment rela ted to consideration of other studies. The title, Toxicological Review of Thallium and Compounds , was retained for consistency with other similar a ssessments on the IRIS database.

2. Please identify any additional stud ies that should be considered in the assessment of the
noncancer and cancer health effects of thallium and thallium compounds.

Comments : Three reviewers did not identify any a dditional studies that should be included in
the assessment. One of these reviewers noted th at a rapid Medline search identified several articles dealing with thallium toxi city that were not cited in the Toxicological Review ; however,
this reviewer did not review the studies to determine whethe r they provided important new
additional information. Other re viewers identified the following thallium studies that were not
cited in the Toxicological Review :

• C avanagh, JB. (1988) Book chapter in Recent Adva nces in Nervous System Toxicology, Vol. 100, pp. 177–202 Galli, GL; Manzo, L; Spencer , PS, eds. New York: Plenum Press.

• Kazantzis, G. (2007) Th allium. In: Nordberg, GF; Fowler, BA; Nor dberg, M; et al.; ed s. Handbook on the toxicology of metals. New York, NY: Elsevier; pp. 827–837.

• Granero, S; Domingo, JL. (2002) Levels of metals in soils of Alcalá de Henares, Spain: human health risks. Environ Int 28(3):159–164.

• Kennedy, P; Cavanagh, JB. (1977) Sensory neuropathy pr oduced in the cat with thallous acetate. Acta Neuropathol 39:81–88.

• Windebank, AJ. (1986) Specific inhibition of myelination of lead in vitro; comparison with arsenic, thallium and mercury. Exp Neurol 94:203–212.

O ne of the reviewers noted that the foreign literature may be very significant. This reviewer identified the following papers and suggested these and other foreign language
literature be translated and included in the review:

• Kamil'dzhanov, AKh. (1993) [Experimental substantiation of maximum permissible concentration of thallium carbonate in environmental air]. Gig Sanit 5:8–10.

A-3• Gerasimova, IL. (1991) [Establishment of MPEL for thallium iodide activated cesium iodide in the working zone air]. Gig Tr Prof Zabol 1:31.

• Viereck, L; Kramer, M; Eikmann, T; et al. (1990) [Determining gui delines for metals in children’s playgrounds in North Rhine-Westphalia]. Öffentl Gesundheitswes 53(1):7–15.

• Krasovski ĭ , GN; Kenesariev, UI. (1984) [Methodological ou tline for the experimental substantiation of a system of indices of the adverse effect of metals on the health status of the population (the example of thallium)]. Gig Sanit 2:22–25.

• Zasukhina, GD; Vasil'eva, IM; Sdirkova, NI.(1980) [Approach to the determination of the mutagenic potential of environmental pollutants with the example of detecting the mutagenic action of thallium carbonate]. Dokl Akad Na uk SSSR 250( 3):766–768.

O ne reviewer noted that “ove r-grooming” in any species may be a sign of stress, pain, or other changes in the central or peripheral ner vous systems and provided the following references
that address this issue:

• Greer, JM; Capecchi, MR. (2002) Ho xb8 is required for normal groom ing behavior in mice. Neuron 33:23–34.

• Kalueff, AV; Tuohimaa, P. (2005) Contrasting grooming phenotypes in three mouse strains markedly different in anxiety and activity (129S1, BALB/c and NMRI). Behav Brain Res 160:1–10.

• Kalueff, AV; Minasyan, A; Keisala, T; et al. (2006) Hair barbering in mice: implications for neurobehavioral research. Behav Proc 71:8–15.

• Welch, JM; Lu, J; Rodriguiz, RM; et al. (2007) Cortico-striatal synaptic defects and OCD-like behaviours in Sapap3-mutant mice. Nature 448:894–900.

I n addition, one reviewer noted that some of the cited references that include experimental studies using more than the LD 50 values have little significance to the toxicity of thallium because of the high dose and sugge sted deleting most of them from the Toxicological
Review

.

Response : Information from the papers on thal lium toxicity by Cavanagh and colleagues, Windebank, and Kazantzis was added to relevant sections of the Toxicological Review . A discussion of the evidence that over-grooming may be indicative of effects on the nervous system was also added, including citati ons to the papers identified above. The paper by Granero and Domingo (2002) pr esented a risk assessment for 12 metals in soil in a region of Spain. For thallium, the EPA RfD previously on IRIS was used in the risk
assessment. No further discussion of health effects information for thallium was presented. Three of the studies from the foreign liter ature identified by one reviewer appear to address the derivation of regulatory limits (K amil'dzhanov, 1993; Gerasimova, 1991; Viereck et al., 1990); limits developed by other regulatory ag encies are generally not included in IRIS toxicological reviews. Therefor e, these papers from the foreig n language literature were not considered useful additions to the thallium toxicological review. The paper (in Russian) by Zasukhina et al. (1980) appears to be the same as the Zasukhina et al. (1983) paper published in English. In general, a toxicological review is not intended to be a comprehensive treatise (see
Foreword to the thallium Toxicological Review ). EPA acknowledges that additional literature on

A-4thallium may be available; however, EPA consider s all literature that has a bearing on the derivation of toxicity reference valu es to have been included in the Toxicological Review

.

3. Please discuss research that you think would be likely to incr ease confidence in the database for future assessments of thallium and compounds.

Comments : Several reviewers noted that the toxic ity database for thallium is very limited. Research suggested by the reviewers to increase the confidence in the thallium database included the following study types and suggestions fo r investigation of specific endpoints:

• A cute dose-range finding studies and, if needed , a chronic inhalation study to identify the
hazard of inhaled thallium.

• A chronic bioassay that would provide in formation on chronic noncancer effects and cancer endpoints. (One re viewer noted that such st udies are expensive and timeconsuming, and sources of funding may be difficult to find.)

• Studies of general reproductive toxicity (in particular to im prove the ability to interpret
the results in Kunmi ng mice observed by Wei).

• Studies of developmental toxicity and developmental neurotoxicity.

• A modern in vivo genotoxicity evaluation. Depending on the outcome of a genotoxicity assessment, the value of conducting a lifet ime chronic bioassay could be weighed.

• A much larger human investigation of an expos ed population. (The reviewer noted that a human investigation would be very costly and that an animal model might be more practical.)

• Endpoints that the various reviewers suggested would be better characterized in studies of
subchronic or chronic duration included al opecia and its sequelae and neurological, reproductive, endocrine, and cardiovascular endpoints.

• Additional clinical chemistry, functional, a nd histopathologic asse ssments to determine the source and intensity of the clinical chemistry change s observed in the MRI (1988) subchronic study.

• Studies that more accurately define dose -response relationship for males and females with appropriate dose ranges that capture the alopecia e ndpoint, including supportive histopathologic examination of alopecia skin a nd apparently unaffected skin areas of all treated and untreated animals and suppor tive in vivo and in vitro experiments demonstrating thallium interaction with hair follicles at various stages of the hair cycle in
both males and females.

• More information on the interaction of thallium with trace elements such as selenium and potassium.

A-5• More studies on the potential mechanisms of action of thallium, including alopecia and neurotoxicity (as perhaps the most economical way of addressing unc ertainties about the target organs for thallium and the plausibility of some of the res ponses that have been observed).

• Studies to establish whether absorption, distribution, and elimination of thallium are linear following oral and dermal absorption of thallium salts.

• Studies on metabolism of thallium that addr ess the question as to whether the body has the capability to convert thal lium from one valence state to the other to determine the
extent to which studies on monovalent thallium are relevant to the toxicity of trivalent thallium. (The reviewer noted that if there is little or no conversion, then it will be important to characterize the toxicity of trivalent thallium separately.)

• Pharmacokinetic studies that would support the development of a physiologically based pharmacokinetic model for thallium (to better understand thallium dosimetry) as well as relative toxicity and pharmacokineti cs of the various thallium salts.

R esponse : No response required.

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 discusse d. Have the choices and assumptions made in the discussion of uncertainty been transparen tly and objectively descri bed? Has the impact of the uncertainty on the assessment been tr ansparently and objectively described?

Comments : Three reviewers observed that the sources of uncertainty were adequately addressed
and that the discussion of uncerta inty was comprehensive. One re viewer considered alopecia to be an acceptable marker of thallium toxicity until better markers become available, at least for female rats, but further noted that it may be difficult to use alop ecia as a reliable marker in male
rats in the absence of a statis tically significant dose response e ffect for alopecia in males.
On e reviewer observed that the very lim ited data set for thallium can be further appreciated by comparing the proposed RfD of 1 × 10 –5 mg/kg-day (0.01 μg/kg-day) to background exposure to environmental levels of thal lium. This RfD suggests that a 70 kg adult can consume 0.7 μg/day. The 90 th percentile adult urinary thal lium elimination from NHANES is 0.380–0.390 μg/L, which is about 0.760–0.780 μg/day in the urine alone. Because thallium has substantial fecal elimination, this suggests th at, if the RfD is adopted , greater than 10% of Americans, and in reality probably close to 50% of Americans, woul d ingest more than the RfD. Because there is no evidence that thallium in the current U.S. diet poses any threat and because there is no possible remediation, this reviewer noted that adop tion of this RfD would produce unnecessary concern if the above ca lculation were correct. This reviewer further noted that a

A-6more thorough use of the MRI (1988) study woul d probably further lower the RfD, suggesting possibly that the entire U.S. population is exposed above threshold. This reviewer considered such an analysis to be an example of how poor the existing data are and further questioned the validity of the analysis. Five reviewers agreed that the Toxicological Review correctly characterized confidence in
the RfD for thallium as low and/or that the thal lium database is weak. In light of this low confidence, one of these reviews offered the opinion that it may be better to suggest a range of RfD values for thallium by using various UFs because of uncertain endpoint, inadequate scientific data, and high UFs. A second reviewer stated that, in lig ht of the weakness of the data set, it is important to openly discuss in the document the pros and cons of calculating an IRIS RfD for thallium. A third reviewer noted that the available data are so poor that it was questionable whether such an analysis is even valid. A fourth review er considered the low
confidence to be a reflection of the limited databa se, including the lack of studies addressing the known toxic effects of thallium, particularly neurotoxicity, devel opmental toxicity, and endocrine effects, failure of the MRI (1988) st udy to identify a NOAEL if all relevant endpoints are considered, and consideration of other studies rejected by EPA that suggest that effects may occur at lower doses than in the MRI study. Other comments offered on specific UFs are summarized under charge question A5 along with EPA’s response.

Response : In response to the reviewer who suggested that different UFs be used for males and females, EPA observes that the overall thallium database does not suggest appreciable gender differences in response to thallium (see Secti on 4.8.2) and that the development of genderspecific reference values is not supported. EPA disagrees with the sugge stion that a range of RfD va lues be developed by using various UFs because of uncertain endpoint, inade quate scientific data, and high UFs. Comments from the peer reviewers in response to charge questions A5 and A6 provide little agreement on
alternative UFs. EPA considers the limitations in the database for thallium not to support an UF smaller than 3,000, and therefore a range of RfD values based on di fferent composite UFs is not scientifically justified. A summary of NHANE S biomonitoring data was added to Section 3.4. Because an assessment of potential exposure is generally outside the scop e of an IRIS assessment, a discussion of estimated intakes from background sources of thallium was not included in the Toxicological Review

.

A-7II. Chemical-Specific Charge Questions A. Oral Reference Dose (RfD) for Thallium

A1. The 90-day oral gavage study by MRI (1988) was selected as the basis for the RfD. Please
comment on whether the selection of this study as the principal study has been scientifically justified. Has this study been transparently and objectively described in the document? Please identify and provide the ratio nale for any other studies that should be selected as the principal study.

Comments : Four reviewers generally agreed with the selection of the MRI (1988) study as the most appropriate basis for the RfD. One review er noted that this study may be an acceptable basis for the RfD, although this was not clear without further analysis of other studies. One other reviewer raised significant concer ns about the study, incl uding the fact that th e choice of animal model, the doses given, and the duration of inve stigation were not well substantiated; sample size, statistical analysis, and calcu lations were not presented; and there was lack of publication in a peer-reviewed journal. This reviewer did not identify an alternative st udy to use as the basis
for the RfD.
Several reviewers recommended that the summ ary of the MRI (1988) study be expanded to include additional study details. One reviewer suggested that the Toxicological Review provide a better time line for the appearance of alop ecia as well as hair loss associated with the normal hair cycle in the rat and a better description of the distribution pattern of alopecia. Four
reviewers indicated that othe r toxicological endpoints (inc luding LDH activity, exophthalmos, lacrimation, and miosis) needed to be better presented/explained.
One reviewer noted that the Toxicological Review considered one or two other studies for use as the principal study and th at a good rationale was provided as to why they were not as good a choice as the MRI study. A second reviewer st ated that he was not aware of any other study that could have been used as a primary study to derive the POD and the RfD but, given the paucity of data available, suggested that othe r studies could be analyzed more in depth and information compared to that obtained from the MRI study. Three reviewers indicated that EPA’s justification for excluding other in vivo studies as the principal studies were not
sufficient. One of these reviewers recommended that RfDs be derived from the other endpoints identified in the MRI study and from the Wei ( 1987) and Zasukhina et al . (1983) studies and the results compared to determine where the MRI study lies with respect to sensitivity. One reviewer observed that epidemiologic studies w ould be ideal, but all available epidemiologic studies provided limited evidence of cause and ef fect associated with exposure in food or
inhalation.

A-8Response : Consistent with the feedback from the majority of reviewers, the MRI (1988) study was retained as a potenti al principal study. This study was subj ect to an independent peer review (as noted in Section 4.2.1.1). None of these independent reviewer s considered the study to be inadequate as the basis for the RfD. The summary of the MRI (1988) findings, including LDH activity, exophthalmos, lacrimation, and miosis, was expanded in the Toxicological Review , Section 4.2.1.1. Information on the time that alopecia was first observed was al so added to this section based on examination of individual animal observational findings in th e MRI (1988) study. Across control and treated groups, alopecia was first observe d somewhere between study days 44 and 60. Severity scores across control and treated groups ranged from 1 to 5. There were no disc ernable differences in either the severity or dist ribution pattern of alopecia ac ross control and treated groups. Further consideration was give n to the use of other in vivo studies as the basis for the RfD (see Sections 5.1.1 and 5.1.5). Where certain in vivo studies were not considered by EPA to be sufficiently reliable as the basis for the POD, more thorough justification was provided (see Section 5.1.1). Expanded discussi on of the limitations of the Za sukhina et al. (1983) study was added to Section 4.3. As noted in response to ge neral charge question 1, the discussion of the Wei (1987) study was revised (Sec tion 4.3), and the study findings were further considered as the potential basis for the POD in Sections 5.1.1 and 5.1.5. For the reasons discussed in Sections 4.6.1 and 5.1.1, the Wei (1987) study was not considered sufficiently reliable as the basis for the
RfD.

A2. Alopecia (hair loss) was selected as the most appropriate critical effect for the RfD. EPA characterizes alopecia as being an adverse e ffect. Please comment on whether the science and mode-of-action information supports alopeci a as an adverse effect. EPA has stated: “Whether alopecia is itself an adverse effect merits consideration. In humans, alopecia is generally reversible upon cessation of thallium exposure. Alopecia, however, appears to be a part of a continuum of dermal changes observed following thallium exposure, as well as one of a spectrum of effects on target organs that include the nervous and gastrointestinal systems. For these reasons, alopecia supporte d by two cases of hair follicle atrophy is
considered an adverse effect.” Please comm ent on whether the selection of this critical effect has been scientifically justified. Is EPA’s choice transparently and objectively described in the document? Please provide a detailed explanation. Please identify and provide the rationale for any other endpoints that should be used instead of alopecia to develop the RfD.

Comments : Three reviewers agreed that alopecia should be considered an adverse effect in and
of itself. One reviewer consid ered alopecia to be undesirable. Another reviewer considered alopecia with histopathology of the hair follicle to be adverse. A final review er stated that use of

A-9alopecia “as a biomarker” is justified until a better marker for adverse effect such as neurotoxic
effects is identified. Several reviewers observed that the fact that alopecia may be reversible does not influence its adversity, whereas one review stated that a drawback to the use of alopecia as an endpoint for RfD is that it is a reversible effect. Reviewers pointed to other limitations in the use of alopecia as the critical effect. One re viewer considered limitati ons to the alopecia data
set to be the facts that hi stopathology was only noted in th e assigned LOAEL dose group and was not examined in the assigned NOAEL and lower dose groups and that there is no evidence
that the skin sections examined were from th e alopecia areas of the sk in. A second reviewer considered a drawback to be that the MRI ( 1988) study did not show a statistically significant dose-response effect for alopecia in male rats. One reviewer suggested that the Toxicological
Review could provide a more detailed discussion of mechanisms of alopecia, particularly with regard to thallium. Two reviewers disagreed with the use of alopecia as the critical effect for RfD
determination for thallium. One reviewer based th is opinion on the fact that it was not clear to this reviewer from either the description of the study or the peer review of the study as to whether this effect was really treatment relate d. This reviewer noted that there was a high background incidence of alopecia in the study population and that the observation in the study of “barbering behavior” in the animals may have account ed for much of the alopecia. It was this reviewer’s experience with lab animals that ther e can be any number of reasons for hair loss, including changes in caging or husbandry, and th e fact that there was such a high background incidence of alopecia suggested that the effect could have been unr elated to treatment. A second reviewer disagreed with the choice of alopecia in the MRI (1988) study because several findings (changes in biochemical parameters, exophthalmos, miosis, and changes in coat) appear to show better dose-response eff ects than alopecia. In response to this and charge question A3, four reviewers su ggested that other effects be considered as the basis for the RfD, including cl inical chemistry, exophthalmos, miosis, changes in coat, lacrimation, and behavioral effects that appear to show dose-response effects and could be more defensible than alopecia as the critical effect.
One reviewer stated that data from several limited subchronic studies (Wei, 1987; Formigli et al., 1986) and in vitro studies (Grego tti et al., 1992) suggest ed that reproductive (especially male) endpoints are worthy of further i nvestigation. This reviewer noted that these studies were poorly reported by the authors, a nd a dose-response relati onship was difficult toascertain, but that this was “however a poor excuse for not attempting to determine a doseresponse relationship and providi ng NOAELs and BMDLs.” This re viewer pointed to the study by Galván-Arzate et al. (2005) that describes a dose-related increase in thallium deposition in the brain and lipid peroxidation in male Wistar rats after 30 day exposure to 0.8 mg/kg or 1.6 mg/kg doses. This reviewer also suggest ed that EPA provide a more critic al review of the 36-week oral

A-10study by Manzo et al. (1983) that described in some detail functional and histopathologic changes in the peripheral nervous system.

Response : Consistent with the position of three review ers who agreed that alopecia is an adverse effect, alopecia was considered al ong with other clini cal observations that showed an increased incidence with dose (see Table 5-1) as a candidate critical effect for establishing the POD for the RfD and as part of a spectrum of clinical obse rvations in thallium-exposed rats. The limitations in the skin histopathology data were recognized (see Sections 4.2.1.1 and 5.1.1). EPA acknowledges that the incidence of alopecia in male rats does not increase monotonically with dose, although the incidence is high er in all groups of treated male rats compared with untreated and vehicle controls. Further, the incidences of other clinical observations were increased in treated male rats over the control groups, consistent with a treatment-related effect in male rats. Available information on the mechanism by whic h thallium induces alopecia is presented in
Sections 4.5.5 and 4.6.3. Consistent with the input from the reviewer s, EPA expanded the analysis of the critical
effect to include other clini cal observations that showed a dose-response effect, including observations related to coat, la crimation, exophthalmos, miosis, and behavior. For the reasons presented in Section 5.1.1, changes in clinical chemistry parameters were not considered as the basis for the POD. EPA acknowledges that ot her factors such as caging and husbandry can cause alopecia in laboratory rodent s; however, the incidence was cl early elevated in both male and female rats over controls. Further, to the ex tent that alopecia was due to barbering, research has shown that barbering in rode nts can reflect a stress-evoked be havioral response (see Section 4.6.1). Accordingly, EPA retained alopecia as one of a set of clinical ob servations that were indicative of a potential treatment-re lated effect in rats. Limitations in clinical observation data
were further discussed in Section 5.1.3.1. EPA reconsidered the findings from other st udies, including Galván-A rzate et al. (2000), Gregotti et al. (1992, 1985), Wei (1987), Formigli et al. (1986), and Manzo et al. (1983), as the basis for the critical effect. As discussed in Section 5.1.1, Formigli et al . (1986), Gregotti et al. (1985), and Manzo et al. ( 1983) were all single-dose studies and thus did not provi de data useful for dose-response analysis. In addition, these studi es administered doses higher than those used in the MRI (1988) study and theref ore provided less information on lo w-dose toxicity. In vitro studies such as Gregotti et al . (1992) and single administration st udies such as Galván-Arzate et al. (2005) do not provide informa tion appropriate for characteriza tion of long-term oral exposure to thallium. The Wei (1987) study was considered further as the basis for the POD for the oral RfD (see Section 5.1.5). In res ponse to one reviewer comment, EPA reexamined the discussion of the Manzo et al. (1983) study in Section 4.2.1.1 and determined it to be adequate. Given the increased mortality in thallium-exposed rats at 40 and 240 days after treatment and the fact that

A-11only a single dose level was us ed in this study, Manzo et al . (1983) was not considered appropriate as a principal study for RfD derivation.

A3. At the high dose in the MR I (1988) study, two female rats e xhibited moderate to severe alopecia that could not be attributed to se lf-barbering or normal cyclic hair growth. Histologic examination of skin samples from these high-dose females showed atrophy of hair follicles. EPA considered these findings to be adverse and thus the high dose in this study (0.25 mg/kg-day thallium su lfate) to be the lowest-obs erved-adverse-effect level (LOAEL). The mid-dose group ( 0.05 mg/kg-day thallium sulfat e) was identified as the noobserved-adverse-effect level (NOAEL). Is EPA’s interpretation of the study findings scientifically justified? Has this interpretation of the findings been transparently and objectively described in the document?

As part of the evaluation of alopecia as a critical effect for the RfD, EPA performed a
series of Fisher’s exact tests to determine if the incidence of alopecia in any of the three dose groups was statistically si gnificantly elevated above controls by using all cases of alopecia reported by MRI (1988). Please comm ent on whether EPA chose the appropriate data set and the appropriate stat istical test for this analysis.

The study investigators reached a different interpretation of the study findings than did EPA. The investigators consider ed alopecia to be attributable to the cyclic pattern of hair growth in rodents and, consequently, did not consider these findings to be biologically significant. The high dose (0.25 mg/kg-day th allium sulfate) was id entified in the MRI (1988) study as the NOAEL. Is the study au thors’ conclusion that the high dose (0.25 mg/kg-day thallium sulfate) represents a NOAEL justified and suppor ted by the study data?

Comments : One reviewer offered the opinion that EPA correctly identified an adverse effect that cannot be “attributable to the cyclic pattern of hair growth in rodent s.” This reviewer stated that EPA needs to clearly define what a NOAEL is ve rsus a LOAEL as it pertains to alopecia; the Toxicological Review needs to make clear whether a dose of 0.04 mg/kg that caused significant incidence of alopecia with no evidence of hist opathologic change in sk in/hair follicle is a NOAEL. A second reviewer could not fully agree with lowering the NOAEL value to 0.05 mg/kg because it was based on a change in inte rpretation of the data rather than any new
scientific evidence and because male rats did not show a statistically significant dose response for alopecia. One reviewer considered the high dose of thallium (0.2 mg/kg- day), and possibly the middle dose (0.04 mg/kg), to be a LOAEL rather th an a NOAEL. This reviewer also considered data on the incidence of alopecia not attributable to barbering to be amenable to BMD modeling.

A-12One reviewer found it difficult to conclude de finitively that the al opecia in two animals with hair follicle atrophy was in fact treatment re lated. This reviewer noted that hair follicles go through a natural cycle of activity and inactivity; it was not clear fr om the descriptions of atrophy that what was being observed was anything more than an observation of a normal condition in these animals; there was no systematic evaluation of dermal tissue from other areas in the same animals; and there was no evaluation of hair follicl e status in all dose groups. Because this is not a standard assessment in subchronic studies, it was not possible to know whether what was
observed is within th e range of normal. One reviewer found the use of alopecia findings to be questionable, noting that the middose group also had alopecia and, although it may have been a result of barbering, barbering is not a normal behavior and suggests that the animals were under stress, possibly in pain; thus, this
dose level could not be a NOAEL. This reviewer also stated that it was not valid to discount a sex difference because sex may affect sensitivity to a given toxin. Finally, this reviewer noted that other parameters that show a dose-res ponse relationship might be amenable to BMD analysis.
One reviewer observed that the incidences of alopecia in middle- and high-dose groups were both significantly differentfrom controls, whereas EPA considered the middle dose to represent a NOAEL with no explanation. It appeared to this reviewer th at EPA considers hair follicle atrophy to be the adverse effect. Because histopathologically observed atrophy was examined only at the high dose, this reviewer be lieved that the high dose should be considered a LOAEL for hair follicle atrophy and a UF of 10 applied. More importantly, this reviewer believed that barbering represents a change in behavior (e.g., caused by pain, an increased arousal level, or stress) and should be considered an adverse effect. Th is reviewer noted that barbering increased in a dose- dependent manner. This revi ewer considered many of the endpoints in the rat study to be consistent with neurological ef fects in humans and that they
should not have been dismissed. This reviewerconsidered the lowest dose to be a LOAEL. One reviewer suggested that EPA recognize in their review that there are possible estrogen receptor pathways within the dermal pa pilla that regulate the telogen-anagen follicle transition and that diffusible factors associated with the anagen follicle influence cell proliferation in the epidermis (Oh and Smart, 1996). This may explain male versus female differences observed in the MRI study.

Response : In response to peer reviewer comments , EPA broadened the evaluation of the MRI (1988) study findings beyond alopeci a in the presence of hair follicle atrophy to include all
clinical observations in male and female rats and presented the possibil ity that the dose-related increase in the incidence of clinical effects indicated a possible treatment-related effect. EPA identified two possible critical effects from the MRI (1988) study – hair follicle atrophy and
clinical observations – reflecting different interpretations of MRI study findings. With respect to

A-13hair follicle atrophy, EPA in Section 5.1.2 clarif ied that skin tissue was not examined for histopathological changes in th e low- and mid-dose groups. Th e mid-dose group was assumed to approximate a NOAEL for this endpoi nt in light of the low inciden ce of hair follicle atrophy in high-dose female rats and absence in male rats at all dose levels . EPA acknowledges this to be a change in the interpretation of the data that wa s made in light of all relevant literature on thallium toxicity and considered this to be an appropriate basis for reaching a different
determination of the NOAEL and LOAEL fr om the authors of the MRI (1988) study. In the revised discussion of alopecia and ot her clinical observations, EPA considered the possibility that barbering may i ndicate that animals were under st ress or that barbering (and other clinical observations) may refl ect some other change in be havior (e.g., see Section 4.6.1). As suggested by several reviewers, BMD me thodology was applied to the various clinical
observation data sets and was used to derive the potential PODs, including the POD for alopecia. EPA appreciated one reviewer’s suggestion of a possible estrogen receptor pathway as an underlying basis for alopecia; however, there is no ev idence in the thallium toxicity literature for
such a mode of action. Because of the lack of da ta to support this hypothesis, it was not added to
the Toxicological Review

. In Section 4.8.1, EPA concluded that the available thallium toxi city literature as a whole showed no consistent pattern of gend er-related difference in toxicity.

Comments : Two reviewers considered the nonparametric statistical analysis using Fisher’s exact test to be appropriate. One of the reviewers, however, noted th at the choice to include all animals that had alopecia is problematic, especi ally given that the st udy investigators did not attribute all instances of alopecia to thallium treatment. A third re viewer stated that a Fisher’s exact test is a standard procedur e for pair-wise comparisons but fails to take advantage of the fact that there are three dose groups. This reviewer suggested that a more appropriate analysis would be a trend analysis. A fourth reviewer stated th at, whereas a Fisher’s exact test is specifically designed for a simple comparison, a more comple x analysis is generally performed in a doseresponse relationship and it is quit e likely that there would be no significance had that approach
been taken.

Response : As noted previously, EPA revised the dose- response analysis to include consideration of a range of clinical observ ations and used a BMD appro ach in place of a NOAEL/LOAEL approach for identifying a POD. Therefore, less importance was placed on pair-wise comparison as the basis for identifying a NOAEL a nd LOAEL from the MRI (1988) study.

Comments : Two reviewers specifically commented on the study authors’ identification of the
NOAEL. One reviewer stated that the conclusion as to whether th e observation of alopecia is an adverse one is a matter of judgment and that th e investigators who wrote the original study report

A-14had a different interpretation of the alopecia; howev er, it was less clear to this reviewer why they
concluded that the clinical chem istry observations or exophthalmos were not adverse. A second reviewer noted that the presence of follicle changes sugge sts that this was truly a toxic effect and that barbering represented stress and possibly pain. This review er further noted that the study investigators’ interpretation of these data adde d considerable doubt on thei r ability to interpret the other findings in their study and further calls into question the choice of this study as the critical data for determining the RfD.

Response : EPA disagrees with the reviewer who called into question th e choice of the MRI (1988) study as a potential principal study because of doubt about the ability of the study authors to interpret study findings. EPA notes that this study was subject to an in dependent peer review. The peer reviewers of the MRI (1988) study ge nerally found the study to have been conducted according to guideline protocols in place at the tim e. Further, EPA reached its own conclusions based on an independent review of study findi ngs and did not rely only on the authors’ interpretation of the data. Accordingly, EPA c onsiders the selection of the MRI (1988) study as a principal study to be supported, although it suffers from some limitations. Limitations in study design and uncertainties in data sets from MRI (1988) were addre ssed in Section 5.1.3.1. Additional comments in response to this char ge question, related to the consideration of other endpoints as critical effects, are summarized under charge question A2.

A4. The traditional NOAEL-LOAEL ap proach was used to define the point of departure (POD) for the RfD. A benchmark dose (BMD) anal ysis was considered but was not conducted because of the nature of the data set for alope cia. Please provide comments with regards to whether a NOAEL-LOAEL approach is the best approach for determining the POD. Has the approach been scientifically justified? Is it transparently and objectively described? Please identify and provide a rationale for any alternative approaches for the determination of the POD and if such approaches are preferred to EPA’s approach.

Comments : Five reviewers agreed that the NOAEL- LOAEL approach is the most appropriate method to define the POD if it is derived using data for alopecia. One reviewer stated that, if the LOAEL-NOAEL approach from the MRI (1988) st udy is used, the lowest dose should be the
POD as a LOAEL.
Five reviewers noted that there are data sets for other endpoints from the MRI (1988) study (e.g., exophthalmos, lacrimation, blood chem istry) for which BMD modeling should be
considered. One of these reviewers also noted th at the BMD approach should be tried for each of the endpoints that exhibits a dose-response relationship in the Wei (1987) and Zasukhina et al. (1983) studies. In this case, the POD used to derive the RfD may be th e lowest BMDL, the one
that EPA thinks is the most reliable or most or derly under analysis, or so me kind of average.

A-15One reviewer suggested deriving a range of Rf D values rather than a single RfD value.

Response : BMD modeling of all appropriate data sets from the MRI (1988) study was conducted and presented in Appendix B of the Toxicological Review . BMD modeling was not performed for the Zasukhina et al. (1983) study b ecause, as discussed in the response to charge question A1 comments, the study results were not considered scientifically reliable. The Wei (1987) study was included in Section 5.1.5, Candi date RfD Comparison Information, and the LOAEL was considered as a potential POD. Give n the uncertainties associated with incomplete reporting and the lack of confirmatory findings at dose levels close to those used in the Wei (1987) study, data sets from this study were not analyzed using BMD methods. As noted in response to general charge que stion 4 comments, it is EPA’s general practice in developing IRIS reference values to derive a single value and not a range.

A5. Please comment on the selection of the unc ertainty factors applied to the POD for the
derivation of the RfD. For inst ance, are they scientifically justified and transparently and objectively described in the document? If chan ges to the selected uncertainty factors are proposed, please identify and provide a rationale(s).

Comments : None of the reviewers disagreed with th e UF applied for interspecies or LOAEL to NOAEL extrapolation. Two reviewers questioned the need for application of an intraspecies UF. (One of these comments was offered in response to general charge ques tion 4 but is summarized here.) In one case, this opinion was based on th e fact that metals such as thallium are not
metabolized and that differences in the sensi tivity to thallium in humans will therefore be insignificant. In the second case, the reviewer identified limited dose-response data for human dermal exposure (resulting from the routine practi ce of dermal administration of thallium salts to children with ringworm) and, in particular, 1930s papers from Munch (1934) that demonstrated the distinction between epilation and toxicity and suggested a po ssible range for which animal
effects can be correlated with human effects. Th is reviewer noted that it was unclear whether the addition of these data would improve uncertainty. Only one reviewer questioned the subchronic to chronic UF of 3. It was this reviewer’s opinion that there is little or no uncertainty when alopecia occurs following oral exposure to thallium. The effect occurs in less than a subchronic time frame, and therefore the UF for subchronic to chronic extrapolati on may not be necessary. This re viewer also stated that EPA needs to counter this argument if they are c oncerned about toxic effects other than alopecia following chronic exposure. Input on the database UF is discu ssed under charge question A6 below.

A-16Response : Consistent with the inpu t from the peer reviewers, the UFs for interspecies and LOAEL to NOAEL extrapolation were retained. The paper by Munch (1934) was reviewed for information that would support reducing the uncertainty associated with intraspecies va riability. This paper, which generally reported symptoms of overt toxicity, including death, was determined not to be useful for characterizing variation in intraspecies sensitivity to envir onmental exposures to thallium. Accordingly, the intraspecies UF of 10 was retained. Consistent with the majority of reviewers, a subchronic to chronic UF of 3 was retained. One reviewer suggested that a subchronic to ch ronic UF might not be necessary; however, the UF of 3 was retained because, in the absence of any chronic toxicity information, it is unknown whether effects other than alopecia and other clin ical observations could manifest at low doses.

A6. Please comment specifically on the database uncertainty factor of 10 applied in the RfD
derivation. Please comment on the use of the da tabase uncertainty f actor specifically for the lack of adequate developmental toxici ty studies and a two-generation reproductive toxicity study and additional uncertainty asso ciated with the limited data available on neurotoxicity in light of the potential for neur otoxicity to represent a sensitive endpoint for thallium exposure. Please comment on whether the selection of the database uncertainty
factor for the RfD has been scie ntifically justified. Has this selection been transparently and objectively describe d in the document?

Comments : Three reviewers considered EPA’s database UF to be appropriate. One of these reviewers stated that the absence of specific dose-response studies a ddressing the issues of neurotoxicity, reproductive toxicit y, and developmental toxicity of thallium suggests that a UF to account for an incomplete database is appropr iate. Use of a database UF was supported by evidence that neurotoxicity is se en in humans upon (high) exposure to thallium and in animal studies (e.g., Manzo et al., 1983) a nd that reproductive toxicity is also seen in animal studies (Wei, 1987) at doses below those used in the MRI (1988) study. A second reviewer similarly
noted that neurotoxic effects and reproductive effects were not adequately evaluated in the
available animal studies and that available developmental toxic ity studies, although not state-ofthe-art, appear adequate to c onclude that developmental toxi city would not drive the risk
assessment for thallium. A third reviewer considered the database to be so weak that a UF of 10
must be utilized and that this choi ce was clearly descri bed and appropriate.
One reviewer stated that a UF of 10 for defi ciencies in the thallium toxicity database may
be deemed to be too conservative. Another revi ewer noted that a databa se UF of 10 may not be necessary if all the endpoints in the MRI (1988) study are consid ered, some of which may be indicative of neurotoxicity. This reviewer noted that the lack of a robust database on developmental toxicity is probl ematic, that developmental neur otoxicity and endocrine studies

A-17have apparently not been performed, and that there are some data providing evidence for reproductive toxicity, but only for one sex. This re viewer suggested that the three studies should be used to generate sample RfDs from the endpoi nts that exhibit a dose-e ffect function; in this case, a database UF of 3 may be more appropriate.
One reviewer did not take issue with the database UF but proposed changing the single RfD to a range of RfDs.

Response : Consistent with the opinion of three of th e external peer reviewers that the database UF of 10 is appropriate, the opini on of essentially all of the reviewers that the limited database for thallium toxicity results in an RfD of low confidence, the lack of adequate developmental toxicity studies and a two-gene ration reproductive toxicity study, a nd the uncertainty associated with the limited neurotoxicity data in light of the potential fo r neurotoxicity to represent a sensitive endpoint for thallium exposure, the full default database UF of 10 was retained. One reviewer observed that, based on the available studies, developmental to xicity would not likely
drive the risk assessment for thallium. EPA does not believe that this observation should change
the size of the database UF since the database de ficiencies associated with reproductive toxicity and neurotoxicity also s upport a database UF of 10.

B. Inhalation Reference Concentration (RfC) for Thallium

B1. Has the rationale and justifi cation for not deriving an RfC fo r thallium been transparently
described in the document?

Comments : Three reviewers agreed that the availabl e data on inhalation exposure to thallium were insufficient to support deri vation of an RfC. One review er noted that an Occupational Safety and Health Administration (O SHA) threshold limit value of 0.10 mg/m 3 Tl is available and should be reported in the Toxicological Review

.

Response : IRIS toxicological review s generally do not include a summary of other Agency regulatory guidelines and standard s. Regulatory values included in an IRIS assessment could become irrelevant with time as they are updated. Further, OSHA occupational limits take into consideration technological achievability and economi c costs in addition to toxicity and therefore are not comparable to IRIS reference values. Therefore, the OSHA o ccupational limit was not
included in the thallium Toxicological Review

.

A-18C. Carcinogenicity of Thallium and Compounds

C1. Under the EPA’s (2005a) Guidelines for Carcinogen Risk Assessment (www.epa.gov/iris/backgr-d.htm), the Agency concluded that ther e is “inadequate information to assess the carcinogenic poten tial” of thallium and compounds. Please comment on the scientific justification for the cancer weight-o f-evidence characterization. A quantitative cancer assessment was not derived for thallium. Has the scientific justification for not derivi ng a quantitative cancer assessm ent been transparently and objectively described?

Comments : All six reviewers agreed that the available literature does not support a characterization of the carcinogenicity of tha llium and that the rationale for not performing a quantitative risk assessment for cancer was appropriate.

Response : No response needed.

PUBLIC COMMENTS Two submissions from the public were recei ved. The submissions received during the official public comment period were distributed to the external pe er review panel prior to the public meeting and discussion of the assessment. Submissions offered some editorial comments and suggestions for clarification of specific portions of the text. Changes were incorporated in
the Toxicological Review as appropriate and are not discus sed further in this appendix.

A. General Comments

Comment : One public commenter considered thallium to be a very dangerous substance and urged EPA to retain the IRIS assessmen t for thallium on the IRIS database.

Response : EPA intends to retain an assessment of the health effects of thallium and compounds
on the IRIS database. The draft Toxicological Review provided for public comment and external peer review represented an effort to update th e health assessment for thallium and thallium compounds to reflect the current available sc ience on this compound. However, EPA has not recommended an RfD for thallium compounds in th is specific case. Justification for this decision is provided in Section 5.1.3.1.

A-19B. Oral Reference Dose (RfD) for Thallium

Comment : One public commenter did not consider th e 0.25 mg/kg-day dose to be a biologically significant LOAEL because there is inadequate evidence to link the high dose alopecia with biologically significant eff ects. Specific reasons for failing to establish this link included the following: (1) complete histopathologic examina tions were conducted only in vehicle control and high

‐ dose groups; ther efore, the dose

‐ response relationship of this effect was not established; (2) background occurrence of al opecia in study animals and the potential for misc lassification adds uncertainty regarding the incidence of treatment

‐ related alopecia in study animals; (3) patterns of alopecia and hair follicle atrophy were inconsistent between male and female rats; increases in the incidence of alopecia in male rats were not dose related; and atrophy of the hair follicles in male rats was absent; and (4) there is incomplete information on, and inconsistencies in, anatomical patterns of effects.

Response : As discussed in response to peer revi ewer comments on charge question A2, EPA expanded the analysis of potential critical eff ects to include other clin ical observations that showed a dose-response effect, including obs ervations related to coat, lacrimation, exophthalmos, miosis, and behavior. Collectively, these effects were considered evidence of a possible treatment-related effect and to be biologically significan t. Uncertainties associated with these endpoints were also discussed (Section 5.1.3.1).

Comment : One public commenter stated that EPA failed to adequately support its determination that the study dose selected as the POD constitu tes a biologically significant endpoint. This commenter observed that alopecia sh ould not be considered adverse because it is reversible after exposure ceases and noted that one reviewer of the MRI (1988) study desc ribed alopecia as an “early (low grade) lesion” and a dded that “it is not a lesion that could be considered as an adverse health effect at that stage, an d it certainly is not life threatening.”

Response : External peer reviewers gene rally considered alopecia to be an adverse effect. Several reviewers strongly recommended, however, that EP A consider other effects from the MRI (1988) study in addition to (or in place of) alopecia b ecause these other effects were either more sensitive than alopecia, displayed a better dose response, and/or were more clearly treatment related. A more rigorous dose-resp onse analysis of data sets for these other effects was added to
the Toxicological Review

.

Comment : One public commenter observed that available human data do not provide support for
the choice of the LOAEL or NOAEL establishe d by EPA, noting that, although limited in number and methodology, population-based survey s do not provide evidence for alopecia in
A-20thallium-exposed populations. This commenter also noted that the draft RfD is inconsistent with the World Health Organization eval uation of thallium toxicity (pre sented in IPCS [1996]) that considers urinary concentrations <5 μ g/L (assumed to be associated with a daily intake of 10 μ g thallium) unlikely to cause adve rse health effects based on the Brockhaus et al . (1981) study of residents living near a thallium

‐ emitting cement plant in Germany. The commenter further observed that thallium exposures up to 100 times greater than the draft RfD did not include
effects consistent with the draft RfD.

Response : EPA considered the use of the Brockha us et al. (1981) and other population-based surveys as a basis for the RfD; however, the available studies we re not considered sufficiently rigorous for reference value derivation.

Comment : One commenter considered the draft RfD (0.7 μ g/day Tl for a 70 kg adult) to be of questionable relevance for protection of public h ealth because intake of thallium from background sources (according to ATSD R [1992] as high as 5 μ g/day from food and 2 μ g/day from drinking water) exceeds levels of thallium oral exposure corresponding with the draf t RfD. The commenter noted that, although some of this intake may be in less soluble and less well absorbed forms of thallium than the soluble salts upon which the draf t RfD is based, these forms may better represent the forms of thallium present in soil a nd produce at many contaminated sites.

Response : Because the scope of an IRIS assessment does not generally include an analysis of potential exposure, a discussion of intake of thallium compounds from background sources was
not included in the Toxicological Review . As noted in response to general charge question 4, EPA is not recommending an RfD value for thallium compounds.

Comment : One public commenter observed that the draf t RfD applies only to soluble forms of thallium and strongly urged EPA to restrict application of the draf t RfD for soluble salts to only those cases where these forms of thallium are known to be present.

Response : EPA is not recommending an RfD for soluble thallium salts. Therefore, restrictions on the application of an RfD is not an issue at this time.

Comment : One public commenter observed that, a lthough human biomonitoring studies provide direct evidence of widespread, low-level th allium exposures in the general population, biomonitoring estimates from these studies describe uptake rather than intake, do not account for the unabsorbed fraction of the ingested dose, and need to be coupled with a better understanding of the absorption fraction for dietary thallium and fraction excreted in urine in order to estimate daily intakes more reliably.

A-21A-22 Response : A summary of NHANES biomonitoring data was added to Section 3.4. Because an assessment of potential exposure is generally outside the scope of an IRIS assessment, an analysis of thallium intake based on availabl e biomonitoring data wa s not included in the Toxicological Review

.APPENDIX B

Documentation of Benchmark Dose Modeling

B.1. Summary of BMDS Modeling Results for Clinical Observations in the Male Rat (MRI, 1988)

Table B-1. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of rough coat in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.0000 95.1404 0.003737 0.002460 Logistic 0.0000 104.547 0.009731 0.006365 Log-logistic 0.8762 85.7472 0.001063 0.0005675 Multistage (1°) 0.0000 95.1404 0.003737 0.002460 Probit 0.0000 106.954 0.01361 0.009597 Log-probit 0.0008 92.4736 0.004137 0.002679 Weibull 0.0000 95.1404 0.003737 0.002460

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

Only the log-logistic model had a p ≥ 0.1; however, the data set was not considered amenable to modeling because of the steep sl ope of the dose-response curve in the low-dose region and because the BMD 10 and BMDL 10 were well outside experimental range of the data. BMDS output from the log-logist ic model is provided below.

==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\ROUGH_COAT-MALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\ROUGH_COAT-MALE.plt Fri Jun 13 08:32:57 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background+(1-background)/[1+EXP(-intercept-slope*Log(dose))]

Dependent variable = #RoughCoat Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

B-1Default Initial Parameter Values background = 0.1 intercept = 4.47108 slope = 1

Asymptotic Correlation Matrix of Parameter Estimates

*** The model parameter(s) -slope have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

background intercept

background 1 -0.26

intercept -0.26 1

Parameter Estimates 95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0.101788 * * * intercept 4.64934 * * * slope 1 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -40.7444 4 Fitted model -40.8736 2 0.258303 2 0.8788 Reduced model -69.3147 1 57.1405 3 <.0001

AIC: 85.7472

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.1018 4.072 4 40 -0.037 0.0080 0.5108 10.216 11 20 0.351 0.0400 0.8266 16.532 16 20 -0.314 0.2000 0.9590 19.180 19 20 -0.203

Chi^2 = 0.26 d.f. = 2 P-value = 0.8762

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.00106311

BMDL = 0.000567469

B-20 0.2 0.4 0.6 0.8

1

0 0.05 0.1 0.15 0.2Fraction Affected

dose Log-Logistic Model with 0.95 Confidence Level

08:32 06/13 2008 BMDLBMD Log-Logistic

Table B-2. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of piloerection in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p -value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.9976 55.896 0.01958 0.01339 Logistic 0.0868 63.7889 0.06642 0.04852 Log-logistic 0.9609 57.9327 0.01813 0.008735 Multistage (1°) 0.9976 55.896 0.01958 0.01339 Probit 0.1124 63.1776 0.0596 0.04436 Log-probit 0.3004 60.4202 0.03242 0.02167 Weibull 0.9976 55.896 0.01958 0.01339

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

All models with the exception of the logistic model provided adequate fits of the data as assessed by a chi-square goodness-of-fit test ( p ≥ 0.1) and visual inspect ion of the respective plots of observed versus predicted values from the various models. The gamma, multistage, and Weibull models provided the same fit of the data and were judged to prov ide the best model fit
based on the lowest AIC value. The BMDS out put for the gamma model is provided below.

B-3==================================================================== Gamma Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\PILOERECTION-MALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\PILOERECTION-MALE.plt Fri Jun 13 09:33:49 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response]= background+(1-background)*CumGamma[slope*dose,power], where CumGamma(.) is the cummulative Gamma distribution function

Dependent variable = #Piloerection Independent variable = Dose(mg/kg-d) Power parameter is restricted as power >=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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values Background = 0.0121951 Slope = 6.76558 Power = 1.26625

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

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0 NA Slope 5.38237 1.31322 2.8085 7.95624 Power 1 NA

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 Log(likelihood) # Param's Deviance Test d.f. P-value Full model -26.9273 4 Fitted model -26.948 1 0.0414636 3 0.9978 Reduced model -47.1393 1 40.4241 3 <.0001

AIC: 55.896

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0000 0.000 0 40 0.000 0.0080 0.0421 0.843 1 20 0.175 0.0400 0.1937 3.874 4 20 0.071 0.2000 0.6592 13.184 13 20 -0.087

Chi^2 = 0.04 d.f. = 3 P-value = 0.9976

B-4Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0195751

BMDL = 0.0133909

0 0.2 0.4 0.6 0.8

0 0.05 0.1 0.15 0.2Fraction Affected

dose Gamma Multi-Hit Model with 0.95 Confidence Level

09:33 06/13 2008 BMDL BMD

Gamma Multi-Hit

Table B-3. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of shedding in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.0001 97.3226 0.02650 0.01557 Logistic 0.0000 101.522 0.06984 0.04768 Log-logistic 0.0018 89.706 0.009591 0.005740 Multistage (1°) 0.0001 97.3226 0.02650 0.01557 Probit 0.0001 101.233 0.06524 0.04472 Log-probit 0.0000 103.483 0.06468 0.02975 Weibull 0.0001 97.3226 0.02650 0.01557

a Chi-square p -value = p value from the Chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

This data set was not fit by any of the models in BMDS.

B-5Table B-4. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of alopecia in male Sprague -Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.0006 106.08 201851 0.04072 Logistic 0.0029 103.671 0.2356 0.07835 Log-logistic 0.0032 103.436 0.1419 0.02992 Multistage (1°) 0.0031 103.514 0.1701 0.04426 Probit 0.0030 103.655 0.2283 0.07445 Log-probit 0.0006 106.08 0.8226 0.1092 Weibull 0.0031 103.514 0.1701 0.04426

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

This data set was not fit by any of the models in BMDS.

Table B-5. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of lacrimation in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 day

Model Chi-square
p -value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.9986 51.0387 0.0004624 0.0001676 Logistic 1.0000 49.0387 0.0008749 0.0005251 Log-logistic 0.9996 51.0387 0.004725 6.03 × 10 –6 Multistage (2°) 1.0000 51.0387 0.000374 0.0001676 Probit 1.0000 49.0387 0.000879 0.0005970 Log-probit 0.9990 51.0387 0.002302 0.0001857 Weibull 1.0000 49.0388 0.0003007 0.0001676

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

This data set was not considered amenable to modeling because of the steep slope of the dose-response curve in the low-dose region and because the BMD 10 and BMDL 10 values were well outside the experimental ra nge of the data. The BMDS out put for the logistic model is presented below.

==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\LACRIMATION-MALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\LACRIMATION-MALE.plt Fri Jun 13 09:43:31 2008

====================================================================

BMDS MODEL RUN
B-6~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = 1/[1+EXP(-intercept-slope*dose)]

Dependent variable = #Lacrimation Independent variable = Dose(mg/kg-d) Slope parameter is not restricted

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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial Parameter Values background = 0 Specified intercept = 1.2157 slope = 14.6477

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

intercept 1 -0.38

slope -0.38 1

Parameter Estimates 95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit intercept -1.5506 0.416124 -2.36618 -0.735007 slope 561.879 138.394 290.632 833.127

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -22.5194 4 Fitted model -22.5194 2 3.27387e-008 2 1 Reduced model -64.1035 1 83.1684 3 <.0001

AIC: 49.0387

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.1750 7.000 7 40 -0.000 0.0080 0.9500 19.000 19 20 -0.000 0.0400 1.0000 20.000 20 20 0.000 0.2000 1.0000 20.000 20 20 0.000

Chi^2 = 0.00 d.f. = 2 P-value = 1.0000

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.000874909

BMDL = 0.000525133

B-70 0.2 0.4 0.6 0.8

1

0 0.05 0.1 0.15 0.2Fraction Affected

dose Logistic Model with 0.95 Confidence Level

09:43 06/13 2008 BMDLBMD Logistic

Table B-6. A summary of BMDS (version 1.4.1) modeling results based on incidence of exophthalmos in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.9995 66.7372 0.004381 0.0006528 Logistic 0.9987 64.7422 0.002070 0.001434 Log-logistic 0.9996 66.7372 0.006493 0.001206 Multistage (2°) 1.0000 64.7372 0.002991 0.0006528 Probit 1.0000 64.7372 0.001947 0.001373 Log-probit 0.9995 66.7372 0.005341 0.001283 Weibull 0.9980 66.7372 0.002766 0.0006528

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

This data set was not considered amenable to modeling because of the steep slope of the dose-response curve in the low-dose region and because the BMD 10 and BMDL 10 values were well outside the experimental range of the data. Of the models with the lowest AIC values, the
model with the lowest BMDL 10 (i.e., the two-stage multistage model) is presented below.

==================================================================== Multistage Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\EXOPHTHALMOS-MALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\EXOPHTHALMOS-MALE.plt Fri Jun 13 09:56:05 2008

====================================================================

BMDS MODEL RUN
B-8~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background + (1-background)*[1-EXP( -beta1*dose^1-beta2*dose^2)]

The parameter betas are restricted to be positive

Dependent variable = #Exophthalmos Independent variable = Dose(mg/kg-d)

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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial Parameter Values Background = 1 Beta(1) = 4.41266e+020 Beta(2) = 0

Asymptotic Correlation Matrix of Parameter Estimates ( *** The model parameter(s) -Beta(1) have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

Background Beta(2)

Background 1 -0.44

Beta(2) -0.44 1

Parameter Estimates 95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0.15 * * * Beta(1) 0 * * * Beta(2) 11777.7 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -30.3686 4 Fitted model -30.3686 2 2.2259e-007 2 1 Reduced model -68.0292 1 75.3212 3 <.0001

AIC: 64.7372

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.1500 6.000 6 40 0.000 0.0080 0.6000 12.000 12 20 -0.000 0.0400 1.0000 20.000 20 20 0.000 0.2000 1.0000 20.000 20 20 0.000

Chi^2 = 0.00 d.f. = 2 P-value = 1.0000

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95
B-9BMD = 0.00299095

BMDL = 0.000652821

BMDU = 0.00435446

Taken together, (0.000652821, 0.00435446) is a 90 % two-sided confidence interval for the BMD

0 0.2 0.4 0.6 0.8

1

0 0.05 0.1 0.15 0.2Fraction Affected

dose Multistage Model with 0.95 Confidence Level

09:56 06/13 2008 BMD BMDL Multistage

Table B-7. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of miosis in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.0924 88.4982 0.01341 0.009005 Logistic 0.0201 93.2144 0.03912 0.02853 Log-logistic 0.3209 86.2645 0.006854 0.003910 Multistage (1°) 0.0924 88.4982 0.01341 0.009005 Probit 0.0218 92.9002 0.03674 0.02777 Log-probit 0.0214 91.7394 0.02461 0.01547 Weibull 0.0924 88.4982 0.01341 0.009005

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

The log-logistic model was the only model that did not exhibit statistically significant lack of fit. The BMDS output fr om this model is provided below.

B-10==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\MIOSIS-MALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\MALE\MIOSIS-MALE.plt Fri Jun 13 10:00:04 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background+(1-background)/[1+EXP(-intercept-slope*Log(dose))]

Dependent variable = #Miosis Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

Default Initial Parameter Values background = 0.025 intercept = 2.67092 slope = 1

Asymptotic Correlation Matrix of Parameter Estimates

( *** The model parameter(s) -slope have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

background intercept

background 1 -0.38

intercept -0.38 1

Parameter Estimates 95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0.0342297 * * * intercept 2.78563 * * * slope 1 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -40.1186 4 Fitted model -41.1323 2 2.02732 2 0.3629 Reduced model -59.2953 1 38.3534 3 <.0001

AIC: 86.2645

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0342 1.369 1 40 -0.321 0.0080 0.1451 2.902 5 20 1.332 0.0400 0.4141 8.282 7 20 -0.582 0.2000 0.7723 15.447 15 20 -0.238

Chi^2 = 2.27 d.f. = 2 P-value = 0.3209

Benchmark Dose Computation

B-11Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0068545

BMDL = 0.00391034

0 0.2 0.4 0.6 0.8

0 0.05 0.1 0.15 0.2Fraction Affected

dose Log-Logistic Model with 0.95 Confidence Level

10:00 06/13 2008 BMDL BMD Log-Logistic

\ Table B-8. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of behavioral findings in male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.0271 108.74 0.06920 0.03038 Logistic 0.0251 109.105 0.09757 0.05538 Log-logistic 0.0283 108.549 0.05662 0.02007 Multistage (1°) 0.0271 108.74 0.06920 0.03038 Probit 0.0253 109.067 0.09430 0.05269 Log-probit 0.0191 109.827 0.1271 0.06223 Weibull 0.0271 108.74 0.06920 0.03038

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

This data set was not fit by any of the models in BMDS.

B-12B.2. Summary of BMDS Modeling Results for Clinical Observations in the Female Rat (MRI, 1988)

Table B-9. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of rough coat in female Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.6681 72.0684 0.02434 0.01597 Logistic 0.0831 75.8559 0.06646 0.04900 Log-logistic 0.8772 71.5823 0.01778 0.01008 Multistage (1°) 0.6681 72.0684 0.02434 0.01597 Probit 0.1033 75.4298 0.06033 0.04503 Log-probit 0.2042 74.1826 0.03935 0.02596 Weibull 0.6681 72.0684 0.02434 0.01597

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

All models with the exception of the logistic model provided adequate fits to the data as assessed by a chi-square goodness-of-fit test ( p ≥ 0.1) and visual inspect ion of the respective plots of observed versus predicted va lues from the various models. BMDL 10 estimates from
these models were within a factor of three of each other, suggesti ng no appreciable model dependence. Fitted models exhibiting adequate fit with AIC values within two units of the lowest AIC were considered indistingui shable from one another; thus, BMD 10 and BMDL 10 values from these models were averaged to deri ve the POD. Model fits that yielded the same mathematical model were counted as a single model for averaging purposes (these models included gamma, multistage, and Weibull). Therefore, the BMD 10 and BMDL 10 values for these models were averaged as follows:

Average BMD 10 = (0.02434 + 0.01778) ÷ 2 = 0.02106 mg/kg-day Average BMDL 10 = (0.01597 + 0.01008) ÷ 2 = 0.01303 mg/kg-day

BMDS outputs from the gamma and log-lo gistic models are presented below.

B-13==================================================================== Gamma Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ROUGH_COAT-FEMALE.(d)Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ROUGH_COATFEMALE.plt Fri Jun 13 08:50:07 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response]= background+(1-background)*CumGamma[slope*dose,power], where CumGamma(.) is the cummulative Gamma distribution function

Dependent variable = #RoughCoat Independent variable = Dose(mg/kg-d) Power parameter is restricted as power >=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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values Background = 0.0365854 Slope = 8.19903 Power = 1.3

Asymptotic Correlation Matrix of Parameter Estimates

( *** The model parameter(s) -Power have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

Background Slope

Background 1 -0.27

Slope -0.27 1

Parameter Estimates 95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0.0268981 0.0242052 -0.0205432 0.0743394 Slope 4.32831 1.21395 1.94901 6.70761 Power 1 NA

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 Log(likelihood) # Param's Deviance Test d.f. P-value Full model -33.6561 4 Fitted model -34.0342 2 0.756278 2 0.6851 Reduced model -47.1393 1 26.9666 3 <.0001

AIC: 72.0684

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0269 1.076 1 40 -0.074 0.0080 0.0600 1.200 1 20 -0.189 0.0400 0.1816 3.632 5 20 0.794 0.2000 0.5905 11.811 11 20 -0.369

B-14Chi^2 = 0.81 d.f. = 2 P-value = 0.6681

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0243422

BMDL = 0.0159664

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8

0 0.05 0.1 0.15 0.2Fraction Affected

dose Gamma Multi-Hit Model with 0.95 Confidence Level

08:50 06/13 2008 BMDL BMD Gamma Multi-Hit

==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ROUGH_COAT-FEMALE.(d)Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ROUGH_COATFEMALE.plt Fri Jun 13 08:52:08 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background+(1-background)/[1+EXP(-intercept-slope*Log(dose))]

Dependent variable = #RoughCoat Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

Default Initial Parameter Values background = 0.025 intercept = 2.22927 slope = 1.17797

Asymptotic Correlation Matrix of Parameter Estimates B-15( *** The model parameter(s) -slope have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

background intercept

background 1 -0.26

intercept -0.26 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0.0230944 * * * intercept 1.83236 * * * slope 1 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -33.6561 4 Fitted model -33.7911 2 0.270147 2 0.8737 Reduced model -47.1393 1 26.9666 3 <.0001

AIC: 71.5823

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0231 0.924 1 40 0.080 0.0080 0.0696 1.392 1 20 -0.345 0.0400 0.2184 4.369 5 20 0.342 0.2000 0.5658 11.315 11 20 -0.142

Chi^2 = 0.26 d.f. = 2 P-value = 0.8772

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0177817

BMDL = 0.0100819

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8

0 0.05 0.1 0.15 0.2Fraction Affected

dose Log-Logistic Model with 0.95 Confidence Level

08:52 06/13 2008 BMDL BMD Log-Logistic

B-16Table B-10. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of piloerection in female Spr ague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.8149 47.1544 0.03857 0.02426 Logistic 0.0484 54.0549 0.1008 0.07462 Log-logistic 0.6738 48.9941 0.03656 0.01845 Multistage (1°) 0.8149 47.1544 0.03857 0.02426 Probit 0.0600 53.5635 0.09134 0.06737 Log-probit 0.2492 49.1272 0.05296 0.03658 Weibull 0.8149 47.1544 0.03857 0.02426

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

All models with the exception of the logistic and probit models provided adequate fits to the data as assessed by a ch i-square goodness-of-fit test ( p ≥ 0.1) and visual inspection of the respective plots of observed versus predicte d values from the various models. BMDL 10 estimates from these models were within a f actor of three of each other, suggesting no appreciable model dependence. Fitted models e xhibiting adequate fit with AIC values within
two units of the lowest AIC were considered indistinguishable from one another; thus BMD 10 and BMDL 10 values from these models (gamma, log-logistic, multistage, log-probit, and Weibull) were averaged to deri ve the POD. Model fits that yielded the same mathematical model were counted as a single model for aver aging purposes (these models included gamma, multistage, and Weibull). Therefore, the BMD 10 and BMDL 10 values for these models were averaged as follows:

Average BMD 10 = (0.03857 + 0.03656 + 0.05296) ÷ 3 = 0.04270 mg/kg-day Average BMDL 10 = (0.02426 + 0.0184 + 0.03658) ÷ 3 = 0.02641 mg/kg-day

BMDS outputs from the gamma, log-logistic, and log-probit models are presented below.

==================================================================== Gamma Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\PILOERECTION-FEMALE.(d)Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\PILOERECTIONFEMALE.plt Fri Jun 13 08:56:08 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response]= background+(1-background)*CumGamma[slope*dose,power],

B-17where CumGamma(.) is the cummulative Gamma distribution function

Dependent variable = #Piloerection Independent variable = Dose(mg/kg-d) Power parameter is restricted as power >=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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values Background = 0.0121951 Slope = 5.0954 Power = 1.3

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

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0 NA Slope 2.73196 0.831275 1.10269 4.36123 Power 1 NA

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 Log(likelihood) # Param's Deviance Test d.f. P-value Full model -21.9144 4 Fitted model -22.5772 1 1.32556 3 0.7231 Reduced model -34.6515 1 25.4742 3 <.0001

AIC: 47.1544

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0000 0.000 0 40 0.000 0.0080 0.0216 0.432 0 20 -0.665 0.0400 0.1035 2.070 3 20 0.682 0.2000 0.4210 8.419 8 20 -0.190

Chi^2 = 0.94 d.f. = 3 P-value = 0.8149

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0385659

BMDL = 0.024262

B-180 0.1 0.2 0.3 0.4 0.5 0.6 0.7

0 0.05 0.1 0.15 0.2Fraction Affected

dose Gamma Multi-Hit Model with 0.95 Confidence Level

08:56 06/13 2008 BMDL BMD Gamma Multi-Hit

==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\PILOERECTION-FEMALE.(d)Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\PILOERECTIONFEMALE.plt Fri Jun 13 08:56:57 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background+(1-background)/[1+EXP(-intercept-slope*Log(dose))]

Dependent variable = #Piloerection Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

Default Initial Parameter Values background = 0 intercept = 1.35689 slope = 1.02772

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

intercept 1 0.92

slope 0.92 1

B-19Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0 * * * intercept 1.45513 * * * slope 1.10379 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -21.9144 4 Fitted model -22.497 2 1.16526 2 0.5584 Reduced model -34.6515 1 25.4742 3 <.0001

AIC: 48.9941

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0000 0.000 0 40 0.000 0.0080 0.0203 0.407 0 20 -0.644 0.0400 0.1093 2.186 3 20 0.583 0.2000 0.4203 8.407 8 20 -0.184

Chi^2 = 0.79 d.f. = 2 P-value = 0.6738

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0365553

BMDL = 0.018448

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7

0 0.05 0.1 0.15 0.2Fraction Affected

dose Log-Logistic Model with 0.95 Confidence Level

08:56 06/13 2008 BMDL BMD Log-Logistic

B-20==================================================================== Probit Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\PILOERECTION-FEMALE.(d)Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\PILOERECTIONFEMALE.plt Fri Jun 13 08:59:17 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = Background + (1-Background) * CumNorm(Intercept+Slope*Log(Dose)),

where CumNorm(.) is the cumulative normal distribution function

Dependent variable = #Piloerection Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

Default Initial (and Specified) Parameter Values background = 0 intercept = 1.70413 slope = 1

Asymptotic Correlation Matrix of Parameter Estimates

( *** The model parameter(s) -background -slope have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

intercept 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0 NA intercept 1.65675 0.228852 1.20821 2.10529 slope 1 NA

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 Log(likelihood) # Param's Deviance Test d.f. P-value Full model -21.9144 4 Fitted model -23.5636 1 3.29839 3 0.3479 Reduced model -34.6515 1 25.4742 3 <.0001

AIC: 49.1272

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------
B-210.0000 0.0000 0.000 0 40 0.000 0.0080 0.0008 0.015 0 20 -0.123 0.0400 0.0591 1.183 3 20 1.723 0.2000 0.5189 10.377 8 20 -1.064

Chi^2 = 4.12 d.f. = 3 P-value = 0.2492

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0529554

BMDL = 0.0365797

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7

0 0.05 0.1 0.15 0.2Fraction Affected

dose Probit Model with 0.95 Confidence Level

08:59 06/13 2008 BMDL BMD Probit

Table B-11. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of shedding in female Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.5894 59.2904 0.01968 0.01347 Logistic 0.1620 64.9908 0.06624 0.04847 Log-logistic 0.4442 61.4053 0.01453 0.008519 Multistage (1°) 0.5894 59.2904 0.01968 0.01347 Probit 0.1783 64.6269 0.05923 0.04416 Log-probit 0.1332 64.1872 0.03710 0.02449 Weibull 0.5894 59.2904 0.01968 0.01347

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

B-22All models provided adequate fits of the data as assessed by a chi-square goodness-of-fit test ( p ≥ 0.1) and visual inspection of the respective plots of observed vers us predicted values from the various models. The gamma, multistage, and Weibull models provided the same fit of the data and were judged to provide the best model fit based on the lowest AIC value. The BMDS output for the gamma model is provided below.

==================================================================== Gamma Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\SHEDDING-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\SHEDDING-FEMALE.plt Fri Jun 13 09:02:02 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response]= background+(1-background)*CumGamma[slope*dose,power], where CumGamma(.) is the cummulative Gamma distribution function

Dependent variable = #Shedding Independent variable = Dose(mg/kg-d) Power parameter is restricted as power >=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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values Background = 0.0121951 Slope = 8.9948 Power = 1.64923

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

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0 NA Slope 5.35376 1.30563 2.79477 7.91275 Power 1 NA

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 Log(likelihood) # Param's Deviance Test d.f. P-value Full model -27.9048 4 Fitted model -28.6452 1 1.48083 3 0.6867 Reduced model -47.1393 1 38.4691 3 <.0001

AIC: 59.2904

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

B-23------------------------------------------------------------------------ 0.0000 0.0000 0.000 0 40 0.000 0.0080 0.0419 0.839 2 20 1.296 0.0400 0.1928 3.855 3 20 -0.485 0.2000 0.6572 13.145 13 20 -0.068

Chi^2 = 1.92 d.f. = 3 P-value = 0.5894

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0196797

BMDL = 0.0134653

0 0.2 0.4 0.6 0.8

0 0.05 0.1 0.15 0.2Fraction Affected

dose Gamma Multi-Hit Model with 0.95 Confidence Level

09:02 06/13 2008 BMDL BMD

Gamma Multi-Hit

Table B-12. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of alopecia in female Spragu e-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.2392 111.335 0.02280 0.01389 Logistic 0.0863 113.282 0.04556 0.03215 Log-logistic 0.4922 109.978 0.01338 0.00665 Multistage (1°) 0.2392 111.335 0.02280 0.01389 Probit 0.0924 113.141 0.04351 0.03125 Log-probit 0.0695 113.624 0.04057 0.02328 Weibull 0.2392 111.335 0.02280 0.01389

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

B-24The gamma, log-logistic, multistage, and Weibul l models provided adequate fits to the data as assessed by a chi- square goodness-of-fit test ( p ≥ 0.1) and visual inspection of the respective plots of observed versus predicte d values from the various models. BMDL 10 estimates from these models were within a f actor of three of each other, suggesting no appreciable model dependence. Fitted models e xhibiting adequate fit with AIC values within
two units of the lowest AIC were considered indistinguishable from one another; thus, BMD 10 and BMDL 10 values from these models (gamma, log- logistic, multistage, and Weibull) were averaged to derive the POD. Model fits that yielded the same mathematical model were counted as a single model for averaging purposes (t hese models included gamma, multistage, and Weibull). Therefore, the BMD 10 and BMDL 10 values for these models were averaged as follows:

Average BMD 10 = (0.02280 + 0.01338) ÷ 2 = 0.01809 mg/kg-day Average BMDL 10 = (0.01389 + 0.00665) ÷ 2 = 0.01027 mg/kg-day

BMDS outputs from the gamma and log- logistic models are provided below.

==================================================================== Gamma Model. (Version: 2.8; Date: 02/20/2007)Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ALOPECIAFEMALE.(d)Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ALOPECIAFEMALE.plt Fri Jun 13 09:06:01 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response]= background+(1-background)*CumGamma[slope*dose,power], where CumGamma(.) is the cummulative Gamma distribution function Dependent variable = #Alopecia Independent variable = Dose(mg/kg-d) Power parameter is restricted as power >=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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values Background = 0.134146 Slope = 17.289 Power = 1.3

Asymptotic Correlation Matrix of Parameter Estimates

( *** The model parameter(s) -Power have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

Background Slope

Background 1 -0.42

B-25Slope -0.42 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0.160192 0.0508961 0.0604373 0.259946 Slope 4.62078 1.59736 1.49 7.75156 Power 1 NA

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 Log(likelihood) # Param's Deviance Test d.f. P-value Full model -52.3019 4 Fitted model -53.6677 2 2.73165 2 0.2552 Reduced model -61.0864 1 17.5691 3 0.0005397

AIC: 111.335

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.1602 6.408 5 40 -0.607 0.0080 0.1907 3.813 4 20 0.106 0.0400 0.3019 6.038 9 20 1.443 0.2000 0.6667 13.334 12 20 -0.633

Chi^2 = 2.86 d.f. = 2 P-value = 0.2392

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0228015

BMDL = 0.0138895

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8

0 0.05 0.1 0.15 0.2Fraction Affected

dose Gamma Multi-Hit Model with 0.95 Confidence Level

09:06 06/13 2008 BMDL BMD

Gamma Multi-Hit

B-26==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ALOPECIA-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\ALOPECIA-FEMALE.plt Fri Jun 13 09:07:14 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background+(1-background)/[1+EXP(-intercept-slope*Log(dose))]

Dependent variable = #Alopecia Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

Default Initial Parameter Values background = 0.125 intercept = 2.11821 slope = 1

Asymptotic Correlation Matrix of Parameter Estimates

( *** The model parameter(s) -slope have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

background intercept

background 1 -0.48

intercept -0.48 1

Parameter Estimates 95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0.138817 * * * intercept 2.11678 * * * slope 1 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -52.3019 4 Fitted model -52.9891 2 1.3744 2 0.503 Reduced model -61.0864 1 17.5691 3 0.0005397

AIC: 109.978

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.1388 5.553 5 40 -0.253 0.0080 0.1925 3.849 4 20 0.085 0.0400 0.3536 7.071 9 20 0.902 0.2000 0.6764 13.527 12 20 -0.730

Chi^2 = 1.42 d.f. = 2 P-value = 0.4922

Benchmark Dose Computation
B-27Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0133799

BMDL = 0.0066454

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8

0 0.05 0.1 0.15 0.2Fraction Affected

dose Log-Logistic Model with 0.95 Confidence Level

09:07 06/13 2008 BMDL BMD Log-Logistic

Table B-13. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of lacrimation in female Spr ague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 1.0000 54.4465 4.808 × 10 –5 BMD computation failed. Lower limit includes zero. Logistic 1.0000 54.4465 0.0001258 Log-logistic 1.0000 54.4465 1.517 × 10 –11 Multistage (2°) 0.9996 56.4465 4.882 × 10 –5 8.1310 × 10 –9 Probit 1.0000 54.4465 0.00024051 BMD computation failed. Lower limit includes zero. Log-probit 1.0000 54.4465 9.098 × 10 –6 Weibull 1.0000 54.4465 4.72 × 10 –5

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

All models in BMDS failed to estimate us eful BMD and BMDL values without excessive extrapolation.

B-28Table B-14. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of exophthalmos in female Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.9983 60.9941 0.0005049 0.0001719 Logistic 1.0000 58.9941 0.0006936 0.0004144 Log-logistic 0.9996 60.9941 0.004856 6.86 × 10 –6 Multistage (2°) 1.0000 60.9941 0.001588 0.0001719 Probit 1.0000 58.9941 0.0007337 0.0004976 Log-probit 0.9969 60.9941 0.002044 0.0001944 Weibull 0.9962 60.9942 0.0003155 0.0001719

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

The data set was not considered amenable to modeling because of the steep slope of the dose-response curve in the low-dose region and because the BMD 10 and BMDL 10 values were well outside the experimental range of the data. Of the models with the lowest AIC values, the
one with the lowest BMDL 10 (i.e., the logistic mode l) is presented below.

==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\EXOPHTHALMOS-FEMALE.(d)Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\EXOPHTHALMOSFEMALE.plt Fri Jun 13 09:15:35 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = 1/[1+EXP(-intercept-slope*dose)]

Dependent variable = #Exophthalmos Independent variable = Dose(mg/kg-d) Slope parameter is not restricted

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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial Parameter Values background = 0 Specified intercept = 1.43545 slope = 13.3397

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 )

B-29intercept slope

intercept 1 -0.33

slope -0.33 1

Parameter Estimates 95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit intercept -0.96942 0.354104 -1.66345 -0.275389 slope 489.249 135.678 223.326 755.172

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -27.4971 4 Fitted model -27.4971 2 3.53613e-007 2 1 Reduced model -61.0864 1 67.1787 3 <.0001

AIC: 58.9941

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.2750 11.000 11 40 0.000 0.0080 0.9500 19.000 19 20 -0.000 0.0400 1.0000 20.000 20 20 0.000 0.2000 1.0000 20.000 20 20 0.000

Chi^2 = 0.00 d.f. = 2 P-value = 1.0000

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.000693631

BMDL = 0.000414359

0.2 0.4 0.6 0.8

1

0 0.05 0.1 0.15 0.2Fraction Affected

dose Logistic Model with 0.95 Confidence Level

09:15 06/13 2008 BMDLBMD Logistic

B-30Table B-15. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of miosis in female Sprag ue-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.0010 109.237 0.03729 0.02024 Logistic 0.0004 111.015 0.07143 0.04590 Log-logistic 0.0028 107.854 0.02252 0.01048 Multistage (1°) 0.0010 109.237 0.03729 0.02024 Probit 0.0004 110.867 0.06780 0.04360 Log-probit 0.0001 112.733 0.08222 0.03640 Weibull 0.0010 109.237 0.03729 0.02024

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

This data set was not fit by any of the models in BMDS.

Table B-16. A summary of BMDS (versio n 1.4.1) modeling results based on incidence of behavioral findings in fe male Sprague-Dawley rats exposed to thallium sulfate via gavage for 90 days

Model Chi-square
p value a AIC BMD 10 (mg/kg-day) BMDL 10 (mg/kg-day) Gamma 0.1256 69.3045 0.1594 0.04585 Logistic 0.2197 67.905 0.1009 0.07207 Log-logistic 0.1256 69.3045 0.1736 0.04377 Multistage (2°) 0.2766 67.482 0.1081 0.04445 Probit 0.2100 68.0129 0.09473 0.06621 Log-probit 0.1256 69.3045 0.1541 0.05892 Weibull 0.1256 69.3045 0.1760 0.04585

a Chi-square p value = p value from the chi-square test for lack of fit. Values <0.1 fail to meet conventional goodness-of-fit criteria.

Data source: MRI (1988).

All of the models provided adequate fits of the data as assessed by a chi-square goodnessof-fit test ( p ≥ 0.1) and visual inspection of the respec tive plots of observed versus predicted
values from the various models. BMDL 10 estimates from these models were within a factor of three of each other, suggesting no appreciable model dependence. Fitted models exhibiting adequate fit with AIC values within two units of the lowest AIC were considered indistinguishable from one another; thus BMD 10 and BMDL 10 values from all seven models were averaged to derive the POD. Therefore, the BMD 10 and BMDL 10 values for these models were averaged as follows:

B-31Average BMD 10 = (0.1594 + 0.1009 + 0.1736 + 0.1081 + 0.09473 + 0.1541 + 0.1760) ÷ 7 = 0.1381 mg/kg-day Average BMDL 10 = (0.04585 + 0.07207 + 0.04377 + 0.04445 + 0.06621 + 0.05892 + 0.04585) ÷ 7 = 0.05387 mg/kg-day

BMDS outputs from each model are provided below.

==================================================================== Gamma Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.plt Fri Jun 13 09:23:45 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response]= background+(1-background)*CumGamma[slope*dose,power], where CumGamma(.) is the cummulative Gamma distribution function

Dependent variable = #Behavior Independent variable = Dose(mg/kg-d) Power parameter is restricted as power >=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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values Background = 0.109756 Slope = 7.56372 Power = 2.63978

Asymptotic Correlation Matrix of Parameter Estimates

Background Slope Power

Background 1 -0.0024 -0.0022

Slope -0.0024 1 1

Power -0.0022 1 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0.0625 0.0270635 0.00945649 0.115543 Slope 59.7049 4193.5 -8159.41 8278.82 Power 14.0574 900.278 -1750.45 1778.57

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -29.9226 4 Fitted model -31.6523 3 3.45942 1 0.06289 Reduced model -36.6925 1 13.5399 3 0.003603

AIC: 69.3045

B-32Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0625 2.500 4 40 0.980 0.0080 0.0625 1.250 0 20 -1.155 0.0400 0.0625 1.250 1 20 -0.231 0.2000 0.3500 7.000 7 20 0.000

Chi^2 = 2.35 d.f. = 1 P-value = 0.1256

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.159402

BMDL = 0.0458492

0 0.1 0.2 0.3 0.4 0.5 0.6

0 0.05 0.1 0.15 0.2Fraction Affected

dose Gamma Multi-Hit Model with 0.95 Confidence Level

09:23 06/13 2008 BMDL BMD Gamma Multi-Hit

==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.plt Fri Jun 13 09:24:08 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = 1/[1+EXP(-intercept-slope*dose)]

Dependent variable = #Behavior Independent variable = Dose(mg/kg-d) Slope parameter is not restricted

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: 1e-008

B-33Parameter Convergence has been set to: 1e-008

Default Initial Parameter Values background = 0 Specified intercept = -2.96071 slope = 11.6261

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

intercept 1 -0.74

slope -0.74 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit intercept -2.81211 0.489479 -3.77147 -1.85275 slope 10.758 3.50284 3.89255 17.6234

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -29.9226 4 Fitted model -31.9525 2 4.05988 2 0.1313 Reduced model -36.6925 1 13.5399 3 0.003603

AIC: 67.905

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0567 2.267 4 40 1.185 0.0080 0.0615 1.229 0 20 -1.144 0.0400 0.0846 1.691 1 20 -0.556 0.2000 0.3406 6.813 7 20 0.088

Chi^2 = 3.03 d.f. = 2 P-value = 0.2197

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.10089

BMDL = 0.0720676

B-340 0.1 0.2 0.3 0.4 0.5 0.6

0 0.05 0.1 0.15 0.2Fraction Affected

dose Logistic Model with 0.95 Confidence Level

09:24 06/13 2008 BMDL BMD Logistic

==================================================================== Logistic Model. (Version: 2.9; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.plt Fri Jun 13 09:24:29 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background+(1-background)/[1+EXP(-intercept-slope*Log(dose))]

Dependent variable = #Behavior Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

Default Initial Parameter Values background = 0.1 intercept = 0.293481 slope = 1

Asymptotic Correlation Matrix of Parameter Estimates

background intercept slope

background 1 0.0045 0.0046

intercept 0.0045 1 1

slope 0.0046 1 1

Parameter Estimates

B-3595.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0.0625 * * * intercept 14.8768 * * * slope 9.75033 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -29.9226 4 Fitted model -31.6523 3 3.45942 1 0.06289 Reduced model -36.6925 1 13.5399 3 0.003603

AIC: 69.3045

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0625 2.500 4 40 0.980 0.0080 0.0625 1.250 0 20 -1.155 0.0400 0.0625 1.250 1 20 -0.231 0.2000 0.3500 7.000 7 20 -0.000

Chi^2 = 2.35 d.f. = 1 P-value = 0.1256

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.173579

BMDL = 0.0437732

0 0.1 0.2 0.3 0.4 0.5 0.6

0 0.05 0.1 0.15 0.2Fraction Affected

dose Log-Logistic Model with 0.95 Confidence Level

09:24 06/13 2008 BMDL BMD Log-Logistic

B-36==================================================================== Multistage Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.plt Fri Jun 13 09:25:07 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background + (1-background)*[1-EXP( -beta1*dose^1-beta2*dose^2)]

The parameter betas are restricted to be positive

Dependent variable = #Behavior Independent variable = Dose(mg/kg-d)

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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial Parameter Values Background = 0.0459871 Beta(1) = 0 Beta(2) = 9.5796

Asymptotic Correlation Matrix of Parameter Estimates

( *** The model parameter(s) -Beta(1) have been estimated at a boundary point, or have been specified by the user, and do not appear in the correlation matrix )

Background Beta(2)

Background 1 -0.4

Beta(2) -0.4 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0.060443 * * * Beta(1) 0 * * * Beta(2) 9.01875 * * *

* - Indicates that this value is not calculated.

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -29.9226 4 Fitted model -31.741 2 3.63685 2 0.1623 Reduced model -36.6925 1 13.5399 3 0.003603

AIC: 67.482

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual
B-37------------------------------------------------------------------------ 0.0000 0.0604 2.418 4 40 1.050 0.0080 0.0610 1.220 0 20 -1.140 0.0400 0.0739 1.478 1 20 -0.409 0.2000 0.3450 6.900 7 20 0.047

Chi^2 = 2.57 d.f. = 2 P-value = 0.2766

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.108085

BMDL = 0.0444457

BMDU = 0.173737

Taken together, (0.0444457, 0.173737) is a 90 % two-sided confidence interval for the BMD

0 0.1 0.2 0.3 0.4 0.5 0.6

0 0.05 0.1 0.15 0.2Fraction Affected

dose Multistage Model with 0.95 Confidence Level

09:25 06/13 2008 BMD BMDL Multistage

==================================================================== Probit Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.plt Fri Jun 13 09:25:44 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = CumNorm(Intercept+Slope*Dose),

where CumNorm(.) is the cumulative normal distribution function

Dependent variable = #Behavior Independent variable = Dose(mg/kg-d) Slope parameter is not restricted

B-38Total number of observations = 4 Total number of records with missing values = 0 Maximum number of iterations = 250 Relative Function Convergence has been set to: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values background = 0 Specified intercept = -1.68929 slope = 6.25307

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

intercept 1 -0.66

slope -0.66 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit intercept -1.58535 0.232401 -2.04084 -1.12985 slope 5.83053 1.91096 2.08512 9.57595

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -29.9226 4 Fitted model -32.0065 2 4.16782 2 0.1244 Reduced model -36.6925 1 13.5399 3 0.003603

AIC: 68.0129

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0564 2.258 4 40 1.194 0.0080 0.0619 1.239 0 20 -1.149 0.0400 0.0882 1.763 1 20 -0.602 0.2000 0.3375 6.750 7 20 0.118

Chi^2 = 3.12 d.f. = 2 P-value = 0.2100

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.0947316

BMDL = 0.0662085

B-390 0.1 0.2 0.3 0.4 0.5 0.6

0 0.05 0.1 0.15 0.2Fraction Affected

dose Probit Model with 0.95 Confidence Level

09:25 06/13 2008 BMDL BMD Probit

==================================================================== Probit Model. (Version: 2.8; Date: 02/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.plt Fri Jun 13 09:26:03 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = Background + (1-Background) * CumNorm(Intercept+Slope*Log(Dose)),

where CumNorm(.) is the cumulative normal distribution function

Dependent variable = #Behavior Independent variable = Dose(mg/kg-d) 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: 1e-008 Parameter Convergence has been set to: 1e-008

User has chosen the log transformed model

Default Initial (and Specified) Parameter Values background = 0.1 intercept = 1.18165 slope = 1

Asymptotic Correlation Matrix of Parameter Estimates

background intercept slope

background 1 -0.001 -0.00087

intercept -0.001 1 1

slope -0.00087 1 1

B-40Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit background 0.0625 0.0270635 0.00945646 0.115544 intercept 4.28948 360.951 -703.162 711.741 slope 2.97918 224.271 -436.585 442.543

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -29.9226 4 Fitted model -31.6523 3 3.45942 1 0.06289 Reduced model -36.6925 1 13.5399 3 0.003603

AIC: 69.3045

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0625 2.500 4 40 0.980 0.0080 0.0625 1.250 0 20 -1.155 0.0400 0.0625 1.250 1 20 -0.231 0.2000 0.3500 7.000 7 20 0.000

Chi^2 = 2.35 d.f. = 1 P-value = 0.1256

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

Confidence level = 0.95

BMD = 0.154125

BMDL = 0.0589218

0 0.1 0.2 0.3 0.4 0.5 0.6

0 0.05 0.1 0.15 0.2Fraction Affected

dose Probit Model with 0.95 Confidence Level

09:26 06/13 2008 BMDL BMD Probit

B-41==================================================================== Weibull Model using Weibull Model (Version: 2.7; Date: 2/20/2007) Input Data File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.(d) Gnuplot Plotting File: M:\IRIS CHEMICALS\THALLIUM\BMD\POST EPR 6-2008\FEMALE\BEHAVIOR-FEMALE.plt Fri Jun 13 09:26:52 2008

====================================================================

BMDS MODEL RUN

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~

The form of the probability function is:

P[response] = background + (1-background)*[1-EXP(-slope*dose^power)]

Dependent variable = #Behavior Independent variable = Dose(mg/kg-d) Power parameter is restricted as power >=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: 1e-008 Parameter Convergence has been set to: 1e-008

Default Initial (and Specified) Parameter Values Background = 0.109756 Slope = 9.64766 Power = 2.10563

Asymptotic Correlation Matrix of Parameter Estimates

Background Slope Power

Background 1 0.017 0.017

Slope 0.017 1 1

Power 0.017 1 1

Parameter Estimates

95.0% Wald Confidence Interval Variable Estimate Std. Err. Lower Conf. Limit Upper Conf. Limit Background 0.0625 0.0270671 0.00944955 0.11555 Slope 2.30426e+006 1.04698e+011 -2.05203e+011 2.05207e+011 Power 9.72683 28231.5 -55323 55342.4

Analysis of Deviance Table

Model Log(likelihood) # Param's Deviance Test d.f. P-value Full model -29.9226 4 Fitted model -31.6523 3 3.45942 1 0.06289 Reduced model -36.6925 1 13.5399 3 0.003603

AIC: 69.3045

Goodness of Fit Scaled Dose Est._Prob. Expected Observed Size Residual

------------------------------------------------------------------------ 0.0000 0.0625 2.500 4 40 0.980 0.0080 0.0625 1.250 0 20 -1.155 0.0400 0.0625 1.250 1 20 -0.231 0.2000 0.3500 7.000 7 20 0.000

Chi^2 = 2.35 d.f. = 1 P-value = 0.1256

Benchmark Dose Computation

Specified effect = 0.1

Risk Type = Extra risk

B-42B-43 Confidence level = 0.95

BMD = 0.175955

BMDL = 0.0458492

0 0.1 0.2 0.3 0.4 0.5 0.6

0 0.05 0.1 0.15 0.2Fraction Affected

dose Weibull Model with 0.95 Confidence Level

09:26 06/13 2008 BMDL BMD Weibull