**September 27, 2011** 

## Significant New **Alternatives Policy Pr ogram** 

**Solvent Cleaning Sector Risk Screen on Substitutes for CFC**  **- 113**  **in Electronics, Metal, and Precision**

**Cleaning and Drying Applications Substitute:**  **Perfluorobutyl I**  **odide ( Capstone**  **® 4**  **- I**  **) This risk screen**  **does not contain Clean Air Act (CAA) Confidential Business**  **Information (CBI) and, therefore, may**  **be disclosed to the public. 1.**  **Introduction** Ozone - depleting substances (ODSs) are being phased out of production in response to a series of diplomatic and legislative e fforts that have taken place over the past few years, including the Montreal Protocol and the Clean Air Act Amendments of 1990 (CAAA). The U.S. Environmental Protection Agency (EPA), as authorized by Section 612 of the CAAA , administers the Significant N ew Alternatives Policy (SNAP) P rogram, which identif ies acceptable and unacceptable substitutes for ODS in specific end -uses based on assessment of their health and environmental impacts . EPA's decision on the acceptability of a substitute is based large ly on the findings of a screening assessment of potential human health and environmental risks posed by the substitute in specific applications. EPA has already screened a large number of substitutes in many end-use applications within all of the major OD S -using sectors, including refrigeration and air conditioning; solvent cleaning; foam blowing; aerosols; fire extinguishing; adhesives, coatings and inks; and sterilization. The results of these risk screens are presented in a series of Background Documents that are available in EPA’s docket. The purpose of this risk screen is to supplement EPA's Background Document on the solvent cleaning sector ( EPA 1994 ) (hereinafter referred to as the Background Document) . This risk screen discusses potential human h ealth and environmental risks posed by perfluorobutyl iodide (PFBI) when used as a replacement for CFC - 113 in electronics, metal, and precision cleaning and drying applications. PFBI was submitted under the trade name, Capstone ® 4 - I , as a potential substi tute for CFC - 113 in electronics, metal, and precision cleaning and drying applications, with primary application in the flushing of oxygen supply lines on aircraft. In addition, the substitute is nonflammable and may therefore serve as a replacement for C FC - 113 in vapor degreasing applications. Based on information from the submitter, Capstone ® 4 - I is expected to gain only a small portion of the solvent cleaning market . Table 1 provides the composition of Capstone ® 4 - I. The formulation proposed by the submitter consists of impurities which total at most one percent of the formulation. In addition to PFBI, this risk screen also discusses the risks associated with use the of the PFBI impurities indicated in the Capstone ® 4 - I formulation .September 27, 2011 2 **Table 1**  **: Composition of Capstone**  **® 4**  **- I**  **Component Chemical Formula**  **CAS No. Weight Percent**  **(%)** P erfluoro butyl I odide C 4 F 9 I 423 - 39 - 2 99 - 100 Potential Impurities P erfluoroethyl I odide C 2 F 5 I 354 - 64 - 3 0 - 0.5 P erfluorohexyl I odide C 6 F 13 I 355 - 43 - 1 0 - 0. 4 I odine I 2 7553 - 56 - 2 0 - 0. 1 The potential risks associated with use of substitutes in solvent cleaning have been examined at length in the Background Document. The reader is referred to this reference for a detailed discussion of the methodologies us ed to conduct this risk screen . O ccupational and general population exposure modeling was performed in this risk screen to ensure that use of the proposed substitute did not pose unacceptable risk to workers. Section 2 of this report summarizes the resul ts of the risk screen for the proposed substitute. The remainder of the report is organized into the following sections:

 Section 3: Atmospheric Assessment  Section 4: Flammability Assessment  Section 5 : Occupational Exposure Assessment  Section 6 : General P opulation Assessment  Section 7 : Volatile Organic Compound Assessment  Section 8 : References 

### 2. **S ummary** 

**of Results** PFBI ( as well as Capstone ® 4 - I) is recommended for SNAP approval for electronics, metal, and precision cleaning and drying applications, provided that proper disposal procedures and appropriate personal protective equipment (PPE) , such as gloves and goggles, are used (see Section 5 and Section 6 ). EPA’s risk screen indicates that the use of the proposed substitute will be less harmful to the atmosphere than the continued use of CFC - 113. N o significant toxicity risks to the general population are expected. In addition, the characteristics of the substitute indicate that flammability is not a concern at end -use. 

### 3. **A tmospheric A** 

**ssessment** This section presents an assessment of the potential risks to atmospheric integrity posed by the use of PFBI , as well as the remaining constituents of Capstone ® 4 - I, in electronics, metal, and precision cleaning and drying applications. The ozone depletion potential ( ODP ) , global warming potential ( GWP ) , and atmospheric lifetime (ALT ) of the proposed substitute are presented in Table 2.September 27, 2011 3 **Table 2**  **: Atmospheric Impacts of**  **PFBI and the Remaining Constituents of Capstone ® 4**  **- I**  **Compared to CF**  **C -**  **113 Chemical**  **ODP a**  **GWP (relative to CO 2**  **) 100 Year**  **ALT** PFBI <0.005 b < 5 b ~ days b Perfluoroethyl Iodide NA < 5 c NA Perfluorohexyl Iodide NA NA NA Iodine NA NA ~ minutes d CFC - 113 0.8 e 4,800 f 85 f a ODPs for these compounds are expected to vary by location of release, based on their very short lifetimes. b Confidential submitter 2011. The ALT for PFBI is estimated to be a few days. c The GWP value for perfluoroethyl iodide is assumed to be roughly equal to the calculated GWP value of PFBI (ICF estimate develop ed through consultation with Dr. Don Wuebbles). d T he ALT value for iodine is expected to be very short, on the order of a few minutes (ICF estimate developed through consultation with Dr. Don Wuebbles). e Montreal Protocol and available at: http://www.epa.gov/ozone/ods.html . f IPCC 4 th Assessment Report (Forster et al. 2007). NA – Not Available. In the case of perfluorhexyl iodide, the values are not publicly available. PFBI (including the Capstone ® 4 - I formulation) presents a viable alternative to CFC - 113. The GWP and ODP of PFBI are estimated to be less than 5 and less than 0.005 , respectively, significantly lower than these for CFC - 113 and comparable to or lower than other alternatives for CFC - 113 in this end use . Even though the Capstone® 4 - I f ormulation possesses no Class I or Class II ozone - depleting substances, it is expected to have a non - zero ODP. T he three remaining constituents of Capstone® 4 - I (perfluorohexyl iodide, perfluoroethyl iodide, and iodine ), which comprise a maximum 1 % of the formulation, do not have reported atmospheric values . However, because of the chemical similarities of the remaining constituents to PFBI, the ODP and GWP values are expected be roughly on the same order of magnitud e as PFBI. Moreover, because of the small percentage of the three remaining constituents in the formulation, the overall GWP and ODP of the Capstone formulation is not expected to vary significantly. Therefore , the use of PFBI, including the Capstone® 4 - I formulation, is expected to be less harmful to the ozone layer than the continued use of CFC - 113. 

### 4. **Flammability**

**A ssessment**  As indicated in the substitute’s MSDS, the substitute is non - flammable and thus not expected to pose flammability concerns. 

### 5. **Occupat ional Exposure A**  **ssessment** This section presents an assessment of potential health risks to workers exposed to PFBI during end -use application . **5.1 Toxicity** 

**of the Substitute** For this occupational exposure analysis, potential risks from worker exposure were evaluated by comparing estimated exposure concentrations to the available occupational exposure limit , which is listed in Table 4 . EPA’s approach for identifying or developing these values is discussed in Chapter 3 of the Background Document.September 27, 2011 4 Current information regarding the decomposition of the substitute indicates that breakdown products , including perfluorooctane, iodine, carbon dioxide, carbon monoxide, and hydrofluoric acid , can form when the substitute is exposed to ultraviolet (UV) light or temperatu res above 250°C. When exposed to UV light, a maximum concentration of 0.1% per byproduct may be generated, although generation of hydrofluoric acid is not likely. In addition, degradation during fire conditions may generate unspecified concentrations of by

product. However, the submitter has also noted that hydrofluoric acid is not expected to form during normal use of the substitute. **5.2 Occupational Exposure during End**  **- Use**  Occupational exposure modeling was performed for the proposed substitute to ensure tha t its use does not pose an unacceptable risk to workers during solvent cleaning applications. For end -uses , the submitter has indicated that the proposed substitute will be used primarily for flushing of oxygen supply lines used in aircraft . In addition, the substitute may be used in shallow dipping pans, open top vapor degreasers, or for use with wip e cloths and in hand -pum p bottles for assorted cleaning applications. To minimize potential exposure s , the submitter has indicated that open top degreasers wi ll be used under ventilation controls . Because actual worker exposure data for th is proposed substitute is generally unavailable, potential exposures were estimated using the surrogate method. The methodology used for this screening assessment is based on the one used in the occupational exposure and hazard analysis described in Chapter 5 of the Background Document. Using this method, the potential occupational exposures to PFBI range from 23.0 to 83.3 ppm . As summarized in Table 4, the maximum exposur e is almost five times lower than the recommended exposure limit for PFBI , 375 ppm . Since the maximum occupational exposure to the proposed substitute during end -use is significantly below the acceptable exposure limit ( AEL ) , the proposed substitute is no t considered a significant toxicity threat. However, to ensure that exposures to PFBI remain below the recommended AEL, EPA recommends that workers wear appropriate PPE (e.g., goggles, gloves) and that proper ventilation controls be established when using the proposed substitute for solvent cleaning applications. Section VIII of the Occupational Safety & Health Administration (OSHA) Technical Manual (OSHA 1999) should be consulted for information on selecting the appropriate types of PPE to be worn by perso nnel involved during use of PFBI . **Table 3**  **: PFBI**  **Occupational Exposure**  **Assessment Maximum Calculated Exposure**  **Concentration AEL**  83.3 ppm 375 ppm 1 1 See Attachment 1. ICF International, 2010. “Determination of an AEL for Perfluorobutyl Iodide.” 

### 6. **General Population Exposure A ssessment**  This section screens potential risks to the general population from exposure to releases of PFBI to ambient air , surface water, and solid waste . When the proper safety and disposal precautions, as li sted in the remainder of this section, are followed , PFBI is not expected to cause a significant threat to the environment and human health in the g eneral population when manufactured or used for electronics, metal, and precision cleaning and drying applic ations .September 27, 2011 5 **6.1 Am**  **bient A ir**  If the accidental release measures, handling and storage, engineering controls , and waste disposal are followed per the substitute’s MSDS , significant release s to air are not expected to o ccur . According to the submitter, releases to the air of Capstone ® 4 - I are expected to occur during preci sion cleaning uses in aircraft and estimated to be 1 - 2 kg per day. Other releases to the air during precision, electronics, and metal cleaning and drying applications are expected to occur indoors, and therefore not expected to cause exposure to the general population. Because of the minimal releases of PFBI to the air , use of PFBI is not expected to generate concentrations of concern to the health of the general population. **6.2 Surface Water**  High conc entrations of PFBI in surface water could pose a concern to the environment as an LC 50 of 2.3 mg/L was estimated for fathead minnow in a study performed to assess the acute aquatic toxicity of PFBI . However, d

ue to PFBI’s high vapor pressure (114.3 mm Hg) and high percent volatiles (>98% WT), PFBI is not likely to accumulate in surface water at concentrations high enough to be considered toxic to aquatic organisms. In addition, th e submitter estimates that 1 - 10 kg/day of spent Capstone ® 4 - I will be collecte d as wastewater when the substitute is used for precision cleaning. For electronics and metal cleaning , the product will be handled indoors with wastes scrubbed through ventilation systems and primarily collected as wastewater, estimated to be 10 - 20 kg per day of the spent solvent . For facilities which collect the wastewater for treatment, the wastewater must be sent to an onsite industrial wastewater treatment facility that employs sufficient controls in order to remove halogenated materials. Domestic ( sanitary) wastewater treatment facilities do not meet these requirements. EPA believes that treated wastewater which meets the discharge concentration requirements of an industrial treatment facility is sufficient to control human health and environmental risks , and thus recommends that all spent solvent collected as wastewater be sent to an industrial treatment facility. **6.3 Solid Waste**  The Capstone ® 4 - I PFBI formulation is estimated to be used at less than 100 sites ; where the spent solvent can be collected, the spent solvent can also be reclaimed through distillation. EPA recommends that reclamation of the substitute through distillation be performed to the extent feasible . Residuals from the distillation process and spent solvent collected in container s (and therefore not collected as wastewater and sent to an onsite wastewater treatment facility) are generally considered to be a RCRA hazardous waste (40 CFR Part 261.31). However, not all applications of Capstone ® 4 - I may generate hazardous wastes. Although the constituents of Capstone ® 4 - I are not listed as RCRA hazardous wastes, solvents used for electronics, metal, and precision cleaning applications generally pick up metals and other chemical compounds during the process, which causes the spent solv ent to display characteristic s of hazardous waste (ignitability, corrosivity, reactivity, or toxicity); any solid waste displaying these characteristics or having a specific waste code (e.g., F001) is considered to be a hazardous waste (40 CFR Part 261). Waste regulated as RCRA hazardous wastes is subject to the requirements of the Subtitle C program (including storage, treatment, and disposal requirements), which EPA believes are sufficient to control human health and environmental risks.September 27, 2011 6 

### 7. **Volatile Organ ic Compound A** **ssessment** PFBI is considered to be a VOC for purposes of local air quality. In addition, perfluoroethyl iodide and perfluorohexyl iodide , of the remaining constituents of Capstone ® 4 - I , are also considered to be VOCs. In general, VOC emission s should be sufficiently controlled through regulations and standard industry practices . Analysis shows that even if all Capstone ® 4 - I produced by the submitter in one year at market penetration w ere to be released to the atmosphere over the course of the year (extremely unlikely), the resulting annual VOC emissions would be approximately 1.3x10 - 3 percent of all VOC emissions from all solvent uses, and moreover, only 3 .5 x10 - 4 percent of all annual anthropogenic VOC emissions. 1 As these emissions of Capsto ne ® 4 - I are several orders of magnitude less than other anthropogenic emissions, the environmental impacts of these VOCs are not considered a threat. 

### 8. **References** Confidential submitter . 2011. Follow -up Data for Significant New Alternatives Policy Program Submission to the United States Environmental Protection Agency for Capstone® 4 - I. September 22, 2011. Confidential submitter . 2010 a . S ignificant New Alternatives Policy Program Submission to the United States Evironmental Protection Agency for Capston e ® 4 - I . January 20, 2010 . Confidential submitter . 2010 b . Follow -up Data for Significant New Alternatives Policy Program Submission to the United States Evironmental Protection Agency for Capstone ® 4 - I . April 24, 2010. Confidential submitter . 2010c. Follow -up Data for Significant New Alternatives Policy Program Submission to the United States Evironmental Protection Agency for Capstone ® 4 - I. April 16, 2010. EPA. 2009. 2008 National Emissions Inventory Air Pollutant Emissions Trends Data. Last update d 09 Ju ne 2009. Available online at <http://www.epa.gov/ttn/chief/trends/index.html#tables >. EPA. 1994. Risk Screen on the Use of Substitutes for Class I Ozone - Depleting Substances: Solvent Cleaning. Stratospheric Protection Division. February 1994. ICF International. 2010. “Determination of an AEL for Perfluorobutyl Iodide.” Prepared by Mukhi , N., Prusiewicz, C., Rubenstein, R., and Wagner, M. under EPA Contract No. EP - W - 06 - 008 Task Order 038, Task 06. IPCC/TEAP. 2001. Third Assessment Report: Climate Change: Working Group I: The Scientific Basis. Chapter 6: Radiative Forcing of Climate Change. Cambridge University Press, UK.

1 Based on 2010 projections calculated using 2008 EPA annual VOC emissions data (EPA 2009) and ICF assumptions.September 27, 2011 7 IPCC/TEAP. 1995. Second Assessment Report: Climate Change: Working Group I: The Science of Climate Change. Chapter 2 : Radiativ e Forcing of Climate Change. Cambridge University Press, UK. OSHA. 1999. OSHA Technical Manual. Department of Labor. Occupational Safety and Health Administration. January 20, 1999. Available online at http://www.osha.gov/dts/osta/otm/otm_toc.html Salamone, J. 1996. Polymeric Materials Encyclopedia. Volume 10 Q - S. CRC Press. 1996. Solomon, S., Burkholder, J. B., Ravishankara, A. R., and Garcia, R. R. 1994. Ozone Depletion and Global Warmin g Potentials of CF3I. Journal of Geophysical Research. Vol. 99, pp. 20929 - 20935. October 20, 1994. Youn, D., Patten, K.O., Wuebbles, D.J., Lee, H., So, C.W.. 2010. Potential impact of iodinated replacement compounds CF 3 I and CH 3 I on atmospheric ozone: A three - dimensional modeling study. Atmospheric Chemistry and Physics Discussions. July 2, 2010. WMO (World Meteorological Organization) . 2003. Scientific Assessment of Ozone Depletion: 2002, Global Ozone Research and Monitoring Project

— Report No. 47, 498 pp., Geneva, 2003.**September 26 , 2010** 



## Appendix A: Determination of an AEL for P **erfluorobutyl Iodide AEL Derivation**  Recommended AEL: 375 ppm (8 -hour Time Weighted Average) Basis and Endpoints: NOAEL 1,500ppm (no body weight changes were noted at this exposure c oncentration) Study: A 13 - week nose - only inhalation toxicity study in rats with a 4 -week recovery period Protocol: Nose - only inhalation, 6 hours/day, 5 days/week for 13 weeks Concentrations: 0; 500; 1,500; 5,000 ppm NOAEL: 1,500 ppm Uncertainty Factors: 3 (interspecies extrapolation) AEL 375 ppm (= 1,500 * 6 hours (exposure of rats)/8 hours (workday))* (1/3)) **I. Introduction**  The compound, perfluoro -n -butyl iodide (PFBI) has been proposed for use as a solvent for precision cleaning, electronics cl eaning, metals cleaning, and associated drying end uses. Several studies have been performed for this compound including an acute, 4 -week, and 13 -week series of inhalation studies in rats, a cardiac sensitization study in beagle dogs, pharmacokinetic studies, and genotoxicity studies in bacteria and human cells in culture. The results of these studies are discussed in the following sections to inform an assessment of the potential health risks from human exposure in the intended end uses. **II. Summary of Toxi**  **city Studies Acute Toxicity**  In a published article summarizing the series of toxicity studies, Dodd and coauthors (2004) report the findings of an acute toxicity study performed in Fischer 344 rats. Young adult rats (5/sex/group) were exposed (nose - only) to 0, 10,000, 20,000, 35,000, and 100,000 ppm of vaporized study compound for 4 hours. The rats were kept and observed for 14 - days post - exposure; toxicity measurements were taken pre - , post - , and during exposure, and during the 2 -week post - exposure period. Body weights were obtained prior to exposure and then weekly during the observation period. Gross necropsy was performed on any animals dying prematurely, and at scheduled sacrifice. No mortality was observed at 10,000 ppm; however, all animals at 20,0 00, 35,000, and 100,000 ppm died prematurely, before the end of the 4 -hour exposure. Symptoms observed in rats during or immediately after exposure included clear or red nasal discharge, chromodacryorrhea (red tears), and labored breathing; recovery by the rats was noted within 48 hours post - exposure. The exposure did not induce anySeptember 26, 2011

changes in weight gain in the lowest concentration group, and did not cause any organ effects identifiable by gross necropsy. The combined sex LC 50 was calculated as 14,000 ppm. **Cardiac Sensitization** Six pure -bred beagle dogs were exposed to increasing concentrations of PFBI following an individualized epinephrine dose (ranging from 2 - 12 μg/kg) that had been previously determined optimal. A positive response was identified as the appearance of a burst of multifocal ventricular ectopic activity or ventricular fibrillation (tachycardia alone was not sufficient to signal a positive response). None of the dogs exhibited a positive response at 900 or 3,900 ppm PFBI, but 2/6 dogs exhibited a positive response at 6,200 ppm. The LOAEL for cardiac sensitization was thus identified as 6,200 ppm. **Four -**  **Week Inhalation Toxicity** Five rats per sex per group were exposed to the following concentrations of vaporized PFBI: 0, 100, 1,000, or 10 ,000 ppm. Exposures were nose - only, for 6 hours/day, 5 days/week. The animals were observed for clinical signs of toxicity (e.g., straining/twisting against the exposure apparatus) during exposures. Animals were also inspected for signs of toxicity twice prior to exposure and weekly after exposure started. Body weights and food consumption were measured similarly, before exposure began and weekly thereafter. Body weights were measured and blood samples were taken just prior to scheduled sacrifice; standard hematology and clinical chemistry analyses, as well as determination of circulating thyroid hormone levels, were performed on the blood samples. Urine samples were taken prior to sacrifice for iodide analysis. Select organs (adrenal glands, brain, kidneys , liver, lungs with bronchi, testes with epididymides, and thyroid/parathyroids) were weighed at terminal sacrifice. Tissues preserved for histopathology included all the weighed tissues, as well as esophagus, heart, larynx, nasopharyngeal tissue, ovaries, spleen, trachea, urinary bladder, and any tissues exhibiting abnormalities on gross necropsy. Histopathology was performed on the control and 10,000 ppm groups, and adrenal glands, liver, and testes were examined histologically in the 100 - and 1,000 -ppm groups. Means and standard deviations were determined for all measurements; statistical analyses of the data were performed appropriately. The only clinical sign observed during treatment was reduced activity in the 10,000 -ppm group. No mortalities occurre d in any treatment group during the four -week study. Body weight gains were significantly decreased in males, but not females, at the end of each week of the study (weight gain at the end of week 4 in high - concentration males was just over 50% of control w eight gain). Food consumption was unaffected. According to the study authors, hematology parameters were unaffected by exposure; the data were not published in the reference for confirmation. Triglycerides were significantly increased in males at the top two concentrations, and in females at 10,000 ppm. Urinary iodide levels were significantly increased in males and females at 10,000 ppm, and in males at 1,000 ppm. Interestingly, males excreted greater concentrations of iodide via the urine than did the females at all concentrations. Although serum triglycerides were increased in mid - and high - concentration males and high concentration females, other parameters of liver disruption/damage, including serum levels of liver enzymes, namely aspartate and alanine transaminases; ALT and AST, respectively were unchanged and there was no notable liver histopathology. Significant thyroid hormone changes were limited to increases in T4 levels only; these changes were noted in males at each exposure concentration and in females at the 1,000 and 10,000 ppm concentrations.September 26, 2011

T3 levels in females show a decline of the same general magnitude across all exposure concentrations, but the differences did not reach statistical significance. These changes are shown in Table 1 b elow. **Table 1. Changes in Serum Thyroid Hormone Levels Following 4 -**  **Week PFBI Exposure Hormone**  Exposure Group, ppm#

0 100 1,000 10,000 **TSH** males 0.60±0.08 0.62±0.35 0.74±0.19 0.72±0.09 females 0.52±0.18 0.79±0.45 0.58±0.12 0.77±0.25 **T3** male s 61.4±11.5 60.9±5.3 72.5±2.8 55.0±9.5 females 59.5±13.4 50.0±5.9 49.5±1.2 48.2±9.2 **T4** males 2.95±0.44 6.07±0.52* (206%)^ 8.78±0.72* (298%)^ 7.86±1.10* (266%)^ females 2.47±0.56 3.38±0.46 (137%)^ 4.07±0.22* (165%)^ 5.60±0.95* (227%)^ #mean±SD; n=5 /sex/group; *, statistically significant (p ≤0.01); ^, percent increase over controls **Subchronic (13 -**  **week exposure) Toxicity** In this study, male (15/group) and female (10/group) were exposed 6 hr/day, 5 days/week for 13 weeks (nose - only) to the following concentrations of PFBI: 0, 50 0 (low), 1,500 (mid), or 5,000 (high) ppm. Ten male and female rats were sacrificed for necropsy at the end of the exposure period; the remaining 5 males/group were maintained for a 4 -week recovery period involving no exposure to the test compound. During exposures the following endpoints were measured: mortality and clinical observations, ophthalmology (pre exposure and at sacrifice), weekly food consumption and body weights, clinical pathology, neurotoxicity as determined by a functional observational battery (13 th week only), motor activity (13 th week only), thyroid function (serum TSH [thyroid stimulating hormone], triiodothyronine [T3] 2 , and thyroxine [T4] levels), organ weights and gross necropsy and histopathology. A Functional Observational Battery (FOB) was performed on 10 main study animals/sex/exposure group and consisted of the following analyses:  Home cage evaluations  Handling evaluations  Open field evaluations  Reflex assessments  Grip strength  Landing foot splay  Proprioception  Air righting ab ility  Body temperature  Body weight Statistical analyses were performed on the following parameters: 2 T3 and T4 are the primary iodine - containing hormones responsible for mediating several functions, including metabolism, within the body. T4 is deiodinated in the body to form T3, w hich is the active form of the hormone. TSH, thyroid stimulating hormone, stimulates thyroid cells to take up iodine and begin the iodine organification process 

### .September 26, 2011

 Weekly body weights/body weight changes  Food consumption  Hematology  Coagulation  Clinical chemistry  Organ weights  Forelimb and hindlimb grip strength meas urements  Landing foot splay measurements  Body temperature and body weights for FOB evaluations **Results for Subchronic Toxicity** Atmospheric Exposure Concentrations Measured concentrations of PFBI were very close to nominal concentrations 0.00±0.00, 500±22 , 1,489±83, and 4,931±359 ppm, for the control, low - , mid - , and high - concentration groups, respectively. Analyses indicated that the test compound was completely vaporized during the exposures. Mortality, Clinical Effects, Eye Effects, Food Consumption, Body Weights One control male died on Day 29 during loading into the inhalation tube; all other animals survived to scheduled sacrifice. Clinical effects were generally absent with the exception of increased ano - genital staining in high - concentration rats during the final few weeks of exposure; this effect was not present during the recovery period in males. Ophthalmoscopy revealed no adverse exposure -related effects on the eyes. No exposure related effects in food consumption were noted throughout the s tudy; any changes in food consumption were minor and transient. Body weights in the control, low - and mid - concentration males and females, and high - concentration females were unaffected by treatment, but the high - concentration males exhibited a 9.4% decre ase in mean absolute body weight compared to controls at the end of the exposure period. Statistically decreased body weight gains and absolute body weights were evident as early as week 3 and 4, respectively in high - concentration males, and these changes persisted until the end of exposure. However, for most measurements in the first 8 weeks of the study, the differences in absolute body weight were around 5 - 6%. Body weights of all males during the recovery period were comparable, indicating that weight gain resumed to normal within the first week after exposure ceased. Functional Observational Battery Endpoints measured in the FOB were not significantly different in treated animals compared to controls, with the exception of group effect for male rats; however, because subsequent comparisons between treated groups and controls were not significant, the difference was not considered biologically relevant. Increases in motor activity were noted in the low - and mid - concentration rats compared to controls, b ut these increases were not statistically significant until combined. Because the changes were not dose responsive, they were not considered biologically relevant. Hematology/Clinical Chemistry There were also no treatment -related effects in measured hematology parameters that were clinically significant. Dose -related and statistically significant decreases in prothrombin time (up to 1.3 seconds) were not considered relevant because they did not exceed 3 seconds difference from controls. Treated males and females at all concentrations had increased hemoglobin levels; the increases were only 10% and were consistent across exposure levels, they were not considered adverse or clinically relevant.September 26, 2011

Decreases in aspartate and alanine transaminases (ALT/AST) and alkaline phosphatase (ALP) levels and a non - dose -related increase in serum phosphorus concentrations in males in all exposed groups and in females in the mid - and high - concentration groups were not deemed clinically relevant. There were no clinical chemistry results that were indicative of potential liver or kidney damage. Other changes in clinical chemistry values (e.g., decreases in glucose, blood urea nitrogen, creatinine and cholesterol) were small or were only noted in one sex, and thus, were not considered significant. Thyroid Function The thyroid, as well as some other organs/tissues, possesses an enzyme known as the sodium iodide symporter (NIS), which is responsible for taking up iodine, a necessary element for thyroid function. Because of this, it is expected that the thyroid will respond to the excess presence of free iodide in the body, or the disruption of thyroid hormone stasis, either at the histopathological level, or in the production of hormones that incorporate the iodine. Exposure to PF BIfor 13 weeks did affect the levels of circulating thyroid hormones in both male and female rats. The results are shown in Table 2 below. In the absence of detail in the original study, it was assumed that the hormone levels were bound hormone, not free hormone. Only free hormone is biologically active (again, only T3 activates other biological processes), and therefore, most circulating hormone is bound with some protein (in the case of rats, this protein is albumin). TSH was increased in females only at concentrations ≥500 ppm, while T3 and T4 levels were increased in both sexes at all exposure concentrations. T4 levels were affected the most, and the effect was most evident in male rats. It is of note that T4 levels were comparably elevated at all expos ure concentrations in male rats, and that TSH levels did not increase in exposed males, similar to the response in the 28 - day study. Again, as with the 28 - day study, the male rats were more sensitive to thyroid hormone changes. **Table 2. Changes in Serum Thyroid Hormone Levels Following PFBI Exposure**  Hormone Exposure Concentration, ppm 500 1,500 5,000 Male Female Male Female Male Female TSH +6% +32%* +11% +42%* +10% +53%* T3 -- +27%* +20%* +31%* +33%* +31%* T4 +270%* +65%* +300%* +97%* +310%* +210%* Percent increases were relative to control values; *, statistically significant The hormone levels in exposed male rats were comparable to those of control rats at the end of the recovery period, indicating that the changes were transient and a result o f thyroid responding to increased iodine load in the body. The study authors did not consider the effects as necessarily adverse. Organ Weights Slight decreases, some statistically significant, in select absolute organ weights were noted in males at 5,000 ppm compared to controls, most likely as a result of the slight decrease in overall body weights of these animals. These included brain, epididymides, heart, kidneys, prostate, seminal vesicles, spleen, and thymus. Exceptions to this were absolute adrenal weights, which were increased in both mid - and high concentration males (the latter exhibited a 24% increase relative to controls). The absolute weights of adrenal glands were also significantly increased in females at 1,500 and 5,000 ppm, as were the kid neys; no other organs showed any dose response. Relative adrenal gland weights were increased in males and females at 1,500 and 5,000 ppm, and relative kidney, liver, seminal vesicle and testes weights were increased in males at 5,000 ppm. Relative kidneySeptember 26, 2011

weights were increased in all treated females. In the recovery group of males, absolute organ weights were unchanged relative to control animals, with the exception of the prostate, which was increased in 1,500 -ppm

males, the thyroid/parathyroid gland, which was significantly increased in 500 - and 5,000 - ppm males, but not in 1500 -ppm males, and the liver and spleen, which were increased in 5,000 -ppm males only. The increased relative liver weight in the 5,000 -ppm males was the only persistent change in o rgan weight following cessation of exposure. Gross Necropsy and Histopathology There were no notable gross findings in any organs/tissue systems of any of the treated groups at necropsy. The only remarkable histopathological finding was a minimal level of follicular cell hypertrophy with/without minimal hyperplasia in 6/10, 6/10, and 7/10 male rats at 500 - , 1,500 - , and 5,000 -ppm , respectively, and in 1/10 female rats at 5,000 ppm only. This finding was not observed in any of the treated males in the rec overy group at sacrifice, indicating the effect was completely reversible. It also did not result in an increase in thyroid weight. No effects were noted in any other tissue/organ that has NIS but does not engage in iodide organification (e.g., mammary tis sue, uterine tissue, etc.). There were no notable histopathological findings in any of the organs with absolute or relative weight changes in the study, indicating that the organ weight changes may have been related to stress of exposure or body weight cha

nges, and not the test compound itself. **Pharmacokinetic Studies** As a complement to the 13 - week inhalation toxicity study, Mattie and coworkers at the Air Force Research Laboratory (2010) evaluated in vitro partition coefficients of PFBI using blood, fat, and liver samples from ten previously unexposed Fischer 344 rats. Blood:air partition coefficients for PFBI were evaluated also using human blood samples obtained from excess blood samples provided by a local hospital. The partition coefficients are provided below in Table 3. **Table 3. In Vitro Partition Coefficients for Rats and Humans Rat**  **Human** Blood:air Liver:air Fat:air Blood:air **0.95±0.45** (16*) 0.44±0.31 (12*) 41±3.9 (12*; 16 hrs#) 41±6.0 (15*; 24 hrs#) **0.67±0.34** (18*) * Number of samples tested; # equilibrium of fat samples tested at 16 & 24 hrs As shown above, the blood:air partition coefficient for PFBI was 42% greater in the rat than the human, indicating that inhaled PFBI will partition more readily into the blood of a rat than in the blood of a human. **Genotoxicity** PFBI was tested in an Ames bacterial reversion assay in Salmonella typhimurium ( S. typhimurium ) strains TA98, TA100, TA1535, TA1537, and E. coli strain WP2 uvrA (measuring either base pair or frame shift mutations) in the presenc e and absence of S9 fraction from induced rat livers (metabolizing enzymes including mixed function oxidases) at concentrations ranging from 313 to 5,000 μg/plate (5000 ug is the maximum recommended by harmonized guidelines for this assay published by OPPT S). Appropriate negative and positive controls were analyzed in the same assays. The number of mutants induced by PFBI did not reach the criteria for a positive response in this assay (e.g., a dose of study compound should induce a 2 - fold increase in the numbers of revertants over controls and a dose -response should be seen), indicating that PFBI does not cause either base pair exchange mutations or frame shift mutations inSeptember 26, 2011

bacterial cells either in the presence or absence of activating enzymes. The clasto genicity of PFBI was tested by culturing human lymphocytes with increasing concentrations of PFBI; concentrations tested in Experiments 1 and 2, - S9 3 , were 39 - 5,000 μg/mL, and 20 - 500 μg/mL; concentrations used in both experiments, +S9 were 39 - 313 μg/mL. Cultures were incubated for either 21 hours (experiments 1 and 2) or 45 hours (experiment 2). Concentrations ≥625 μg/mL PFBI, - S9, were highly toxic to the cultured cells. Concentrations ranging from 39 - 200 μg/mL were selected for metaphase analysis, for which 39 μg/mL was the highest non - toxic concentration. In the presence of S9, PFBI was toxic at 313 μg/mL, prompting metaphase analysis at dose levels of 78 - 263 μg/mL. No increases in chromosomal aberrations were observed at any of the analyzed dose le vels. **III. Disruptions in Iodide Homeostasis** 
**— Potential Differences in Sensitivity between**  **Humans and Rats** There are distinct differences between animal species in thyroid hormone economy and metabolism. For example, T3 and T4 in serum have much shorter half - l ives in rats than humans and other primates (Capen 1994) as a result of differences in the type of proteins with which these enzymes bind and how tightly they bind. In rats, T3 and T4 only have albumin with which to bind; in human and monkey serum, by cont rast, T4 and T3 can bind to albumin, as well as T4 - and T3 -binding globulins. Binding to thyroglobulin by T3 and T4 is stronger than binding to albumin, which increases the half - life of these hormones in the human, and means that they are under tighter con trol following perturbations in iodine homeostasis. Also, T3 is bound less tightly to thyroglobulin than T4, resulting in faster turnover and shorter half

- life in most species evaluated (Capen 1994). There are also species - specific differences in sensitivity of the thyroperoxidase enzyme to therapeutic agents and other toxicants (Capen 1994). Thyroperoxidase is responsible for oxidizing iodide, which is the predominant form of iodine to which humans are exposed, to elemental iodine, which can then be add ed to the tyrosine residues of the thyroglobulin in a process called iodine organification, which results in thyroxine (T4), and triiodothyronine (T3). Studies show that thyroperoxidase from the rat, mouse, and dog is more susceptible to inhibition by toxi cants than the same enzyme from the human, monkey, guinea pig, or chicken (Capen 1994). Studies also show that TSH levels are naturally higher in male rats than female rats, and that male rats are more sensitive to disruptions in thyroid hormone economy/h omeostasis. The toxicity studies currently under consideration support this finding. More recently, the human and rat NIS (sodium iodide symporter) genes have been identified, isolated, and transfected into varying tumor cell types to determine if they can be useful in chemotherapy. For example, Heltemes and coworkers (2003) determined the abilities of the two symporters to concentrate radiolabeled 3 S9 fraction is the supernatant fraction of homogenized liver that has been centrifuged at 9000xg -- the supernatant contains cytosol and microsomes which contain metabolic enzymes from the liver, such as cytochrome P450 and others responsible for metabolizin g toxicants. In this specific case, the S9 was isolated from male rats induced with Aroclor 1254 (dissolved in Arachis oil) 5 days prior to liver isolation and supernatant preparation. The S9 fraction wa s tested for efficacy using two mutagenic polycyclic aromatic hydrocarbons. ‘ +S9 ’ indicates that experiments were undertaken with the fraction and ‘

– S9 ’ indicates without the fraction.September 26, 2011

iodine into varying tumor cell lines. They found that 7 human cancer and one monkey cancer cell lines transfe cted with and positively expressing the rat NIS gene took up 125 I consistently better than those transfected with and expressing the human variant (uptake ratios of cells with rat NIS to those with human NIS ranged from 1.9 to 3.4, e.g., 190% to 340% more uptake with the rat symporter than the human symporter). Comparison of the thyroid hormone changes in the 4 -week and the 13 -week study indicate that T4 was more susceptible to change, and that these changes occurred even at the lowest exposure concentration used (100 ppm) and at the shortest exposure duration (28 days). TSH and T3 levels changed slightly, but not significantly, in exposed males and females in the 28 - day study, but the trends were inconsistent. Itis not possible to determine without additional study what cellular mechanisms in the PFBI - exposed rats in the 13 -week study may be causing the increased values of TSH in female rats, and T4 and T3 in male and female rats. Increased release of stored T3 and T4 is possible but unlikely, given th at histopathology of the thyroid did not indicate any notable degradation of the colloid within the thyroid that would be necessary for hormone release. Hepatic microsomal enzymes play a significant role in thyroid hormone regulation, since they are involved in turnover and excretion of both T3 and T4. However, the data do not indicate that increased excretion of thyroid hormones is responsible, as there was no reported increase in hepatic microsomal enzymes in the histopathology tables provided in the stu dy. Disruption of the deiodination of T4 (to form T3) is a possible cause of the changes noted, but cannot be confirmed without additional mechanistic studies, which are not warranted at this time. Taken together, the data do indicate that the rodent mode l is not the most appropriate one for trying to determine what effects on iodine homeostasis might be induced by exposure to PFBI. They also indicate that the human is likely to be less sensitive than the rat to excess iodide intake, and suggest that an uncertainty factor for intraspecies extrapolation (protection for sensitive subpopulations) is overly conservative. **IV. Development of the AEL for PFBI**  A NOAEL (point of departure) of 1,500 ppm from the 13 -week inhalation study was chosen because at this exposure concentration, no body weight changes were noted. Thyroid histopathology and hormone changes observed at this level were completely resolved after exposure cessation and were considered adaptive, rather than adverse effects. The AEL was calculated in the following manner: 1,500 x (6/8) ÷ 3 = **375 ppm** A human equivalent concentration (HEC) was determined by adjusting the NOAEL of 1,500 ppm by the ratio of rat exposure duration per day to that of an occupational worker (6 hours/8 hours). The guidelines for developing Reference Concentration (RfC) values (EPA 1994) were generally followed, which suggest the use of a combined uncertainty factor of 10 for interspecies extrapolation (3 for pharmacokinetic [PK] differences, and 3 for pharmacodynamic [PD] differences). Because the rat blood:air partition coefficient is larger than that of a human, and because the rat sodium iodide symporter is more effective at uptake than that of the human, a UF of 1 was used for PK differences, and a UF of 3 was used for PD differences. An intraspecies UF was deemed not necessary because thyroid effects are specific to existing iodine status which varies significantly between individuals, and because the effectsSeptember 26, 2011

are adaptive and reversible. Because subchronic studies are typically used to develop AEL values, no additional UF has been added to account for study duration. Similarly, the database is considered comprehensive, and no additional UF has been added to account for database limitations.September 26, 2011

**References** Capen CC. 199 4. Mechanisms of Chemical Injury of Thyroid Gland. In Receptor- Mediated Biological Processes: Implications for Evaluating Carcinogenesis , eds. HL Spitzer, TJ Slaga, WF Greenlee, and M McClain. Hoboken, NJ: Wiley - Liss, Inc. pp. 173 - 191. Dodd DE, Hoffmann GM, and Hardy CJ. 2004. Perfluoro -n -butyl iodide: Acute toxicity, subchronic toxicity and genotoxicity evaluations. Int J Toxicol 23:249 - 258. Heltemes LM, Hagan CR, Mitrofanova EE, et al. 2003. The rat sodium iodide symporter gene permits more effecti ve radioisotope concentration than the human sodium iodide symporter gene in human and rodent cancer cells. Cancer Gene Ther 10:14 - 22. Mattie DR, Dodd DE, John PE, et al. 2006. Perfluoro - n -butyl iodide (PFBI): A 13 - week nose - only inhalation toxicity study in rats with a 4 -week recovery period. Air Force Research Laboratory, Wright Patterson AFB, OH, September 2006. Mattie D, G Hoffman, L Narayanan, et al. 2010. A 13 -week nose - only inhalation toxicity study for perfluoro -n -butyl iodide (PFBI) in rats with recommended occupational exposure levels. Inhalation Toxicology 22 (10): 847 - 860. August 2010. U.S. EPA. 1994. Methods for Derivation of Inhalation Reference Concentrations (RfCs) and Application of Inhalation Dosimetry. Office of Research and Development. EPA/600 - 8 - 90/066F. October 1994.