Significant New Alternatives Policy Program

Solvent Cleaning Sector

Risk Screen on Substitutes for CFC-113 in Electronics, Metal, and
Precision Cleaning and Drying Applications

Substitute: Perfluorobutyl Iodide (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.



Introduction

Ozone-depleting substances (ODSs) are being phased out of production in
response to a series of diplomatic and legislative efforts 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 New Alternatives Policy (SNAP) Program,
which identifies 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 largely 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 ODS-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 health 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
substitute 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 CFC-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. 

Table   SEQ Table \* ARABIC  1 : Composition of Capstone® 4-I

Component	Chemical Formula	CAS No.	Weight Percent (%)

Perfluorobutyl Iodide	C4F9I	423-39-2	99-100

Potential Impurities

Perfluoroethyl Iodide	C2F5I	354-64-3	0 - 0.5

Perfluorohexyl Iodide	C6F13I	355-43-1	0 - 0.4

Iodine	I2	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 used to conduct this risk screen. Occupational 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 results 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 Population Assessment 

Section 7: Volatile Organic Compound Assessment

Section 8: References

Summary 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. No 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.   

Atmospheric Assessment

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. 

Table   SEQ Table \* ARABIC  2 :  Atmospheric Impacts of PFBI and the
Remaining Constituents of Capstone® 4-I Compared to CFC-113

 Chemical	

ODPa	

GWP (relative to CO2)

100 Year	

ALT

PFBI	<0.005b	<5b	~daysb

Perfluoroethyl Iodide	NA	<5c	NA

Perfluorohexyl Iodide	NA	NA	NA

Iodine	NA	NA	~minutesd

CFC-113	0.8e	4,800f	85f

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 developed
through consultation with Dr. Don Wuebbles).

dThe 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:  HYPERLINK
"http://www.epa.gov/ozone/ods.html" http://www.epa.gov/ozone/ods.html .

f IPCC 4th 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 formulation
possesses no Class I or Class II ozone-depleting substances, it is
expected to have a non-zero ODP. The 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 magnitude 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.  

Flammability Assessment

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

Occupational Exposure Assessment

This section presents an assessment of potential health risks to workers
exposed to PFBI during end-use application. 

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.

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 temperatures
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 byproduct. However, the submitter
has also noted that hydrofluoric acid is not expected to form during
normal use of the substitute. 

Occupational Exposure during End-Use 

Occupational exposure modeling was performed for the proposed substitute
to ensure that 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 wipe cloths and in hand-pump bottles for
assorted cleaning applications. To minimize potential exposures, the
submitter has indicated that open top degreasers will be used under
ventilation controls.  

 

Because actual worker exposure data for this 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 exposure 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 not 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 personnel involved
during use of PFBI.  

Table   SEQ Table \* ARABIC  4 : PFBI Occupational Exposure Assessment

Maximum Calculated Exposure Concentration	AEL

83.3 ppm	375 ppm1

1See Attachment 1. ICF International,  2010. “Determination of an AEL
for Perfluorobutyl Iodide.”

General Population Exposure Assessment

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 listed 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
general population when manufactured or used for electronics, metal, and
precision cleaning and drying applications. 

Ambient Air

If the accidental release measures, handling and storage, engineering
controls, and waste disposal are followed per the substitute’s MSDS,
significant releases to air are not expected to occur. According to the
submitter, releases to the air of Capstone® 4-I are expected to occur
during precision 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. 

Surface Water 

High concentrations of PFBI in surface water could pose a concern to the
environment as an LC50 of 2.3 mg/L was estimated for fathead minnow in
a study performed to assess the acute aquatic toxicity of PFBI. However,
due 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, the submitter estimates that 1-10 kg/day of
spent Capstone® 4-I will be collected 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.  

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 containers (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 solvent to
display characteristics 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. 

Volatile Organic Compound Assessment 

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 emissions 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 were 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 3.5x10-4 percent of all annual
anthropogenic VOC emissions.  As these emissions of Capstone® 4-I are
several orders of magnitude less than other anthropogenic emissions, the
environmental impacts of these VOCs are not considered a threat.  

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. 2010a.  Significant New Alternatives Policy
Program Submission to the United States Evironmental Protection Agency
for Capstone® 4-I.  January 20, 2010.

Confidential submitter.  2010b.  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 updated 09 June 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. 

IPCC/TEAP. 1995. Second Assessment Report: Climate Change: Working Group
I: The Science of Climate Change. Chapter 2: Radiative 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
 HYPERLINK "http://www.osha.gov/dts/osta/otm/otm_toc.html"
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 Warming 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 CF3I and CH3I 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.

Appendix A:  Determination of an AEL for Perfluorobutyl 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 concentration)

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))

Introduction

The compound, perfluoro-n-butyl iodide (PFBI) has been proposed for use
as a solvent for precision cleaning, electronics cleaning, 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.

Summary of Toxicity 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 20,000,
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 any changes in weight gain in
the lowest concentration group, and did not cause any organ effects
identifiable by gross necropsy. The combined sex LC50 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 occurred 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 weight 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.
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 below.

Table 1.  Changes in Serum Thyroid Hormone Levels Following 4-Week PFBI
Exposure

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, 500 (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 (13th
week only), motor activity (13th week only), thyroid function (serum TSH
[thyroid stimulating hormone], triiodothyronine [T3], 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 ability

Body temperature

Body weight

Statistical analyses were performed on the following parameters:  

Weekly body weights/body weight changes

Food consumption

Hematology

Coagulation

Clinical chemistry

Organ weights

Forelimb and hindlimb grip strength measurements

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 study; 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% decrease 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, but 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. 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 PFBI for
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
exposure 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 of 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
kidneys; 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 kidney
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 organ 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
recovery 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 tissue, 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 changes, 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 presence 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
OPPTS). 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 in bacterial cells either in the presence or absence of
activating enzymes.

The clastogenicity of PFBI was tested by culturing human lymphocytes
with increasing concentrations of PFBI; concentrations tested in
Experiments 1 and 2, -S9, 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 levels.

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-lives 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 contrast, 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 control 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 added 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 toxicants 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/homeostasis. 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 iodine into varying tumor cell
lines. They found that 7 human cancer and one monkey cancer cell lines
transfected with and positively expressing the rat NIS gene took up 125I
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.  

It is 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 that 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 study. 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 model 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.  

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 effects 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.  

References

Capen CC.  1994. 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 effective 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.

 Based on 2010 projections calculated using 2008 EPA annual VOC
emissions data (EPA 2009) and ICF assumptions.

 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, which is the active form of the
hormone.  TSH, thyroid stimulating hormone, stimulates thyroid cells to
take up iodine and begin the iodine organification process.

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%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 metabolizing 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 was tested for efficacy
using two mutagenic polycyclic aromatic hydrocarbons. ‘+S9’
indicates that experiments were undertaken with the fraction and
‘–S9’ indicates without the fraction. 

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	  DATE \@ "MMMM d, yyyy"  September 27, 2011 

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	September 26, 2010

