EPA/635/R-14/333
Preliminary Materials
www.epa.gov/iris
Preliminary Materials for the Integrated Risk Information System (IRIS)
Toxicological Review of Diisobutyl Phthalate (DIBP)
(CASRN No. 84-69-5)
September 2014
NOTICE
This document is comprised of preliminary materials. This information is distributed solely for
the purpose of pre-dissemination review under applicable information quality guidelines. It has
not been formally disseminated by EPA. It does not represent and should not be construed to
represent any Agency determination or policy. It is being circulated for review of its technical
accuracy and science policy implications.
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Washington, DC
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
DISCLAIMER
This document is comprised of preliminary materials for review purposes only. This
information is distributed solely for the purpose of pre-dissemination review under applicable
information quality guidelines. It has not been formally disseminated by EPA. It does not represent
and should not be construed to represent any Agency determination or policy. Mention of trade
names or commercial products does not constitute endorsement or recommendation for use.
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
CONTENTS
PREFACE viii
1. INTRODUCTION 1-1
1.1. DIBP IN THE ENVIRONMENT 1-1
1.1.1. Production and Use 1-1
1.1.2. Environmental Fate 1-2
1.1.3. Human Exposure Pathways 1-2
1.2. SCOPE OF THE ASSESSMENT 1-3
2. METHODS FOR IDENTIFYING AND SELECTING STUDIES 2-1
2.1. DRAFT LITERATURE SEARCH AND SCREENING STRATEGY 2-1
2.2. SELECTION OF CRITICAL STUDIES IN EARLY STAGES OF DRAFT DEVELOPMENT 2-14
2.2.1. General Approach 2-14
2.2.2. Exclusion of Studies 2-15
2.3. STUDY CHARACTERISTICS THAT WILL BE CONSIDERED IN THE FUTURE EVALUATION AND
SYNTHESIS OF THE CRITICAL EPIDEMIOLOGICAL STUDIES FOR DIBP 2-16
2.4. STUDY CHARACTERISTICS THAT WILL BE CONSIDERED IN THE FUTURE EVALUATION AND
SYNTHESIS OF THE CRITICAL EXPERIMENTAL STUDIES FOR DIBP 2-30
3. PRELIMINARY EVIDENCE TABLES AND EXPOSURE-RESPONSE ARRAYS 3-1
3.1. DATA EXTRACTION FOR EPIDEMIOLOGICAL AND ANIMAL STUDIES: PREPARATION OF
PRELIMINARY EVIDENCE TABLES 3-1
3.2. EPIDEMIOLOGICAL STUDIES 3-2
3.2.1. Sexual Differentiation Measures 3-2
3.2.2. Male Reproductive Effects in Humans 3-4
3.2.3. Male Pubertal Development in Humans 3-7
3.2.4. Female Pubertal Development in Humans 3-9
3.2.5. Female Reproductive Effects in Humans 3-12
3.2.6. Pregnancy Outcomes in Humans 3-15
3.2.7. Immune Effects in Humans 3-18
3.2.8. Neurodevelopmental Effects in Humans 3-25
3.2.9. Thyroid Hormone Effects in Humans 3-30
3.2.10.Obesity and Metabolic Effects in Humans 3-31
3.2.11. Male Reproductive Effects 3-62
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.2.12. Female Reproductive Effects 3-73
3.2.13. Liver Effects 3-79
3.2.14. Kidney Effects 3-83
3.2.15. Hematopoietic Effects 3-87
3.2.16.Other Effects 3-90
3.4. PRELIMINARY MECHANISTIC INFORMATION FOR DIBP 3-94
4. REFERENCES 4-1
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
TABLES
Table 2-1. Database search strategy for DIBP 2-2
Table 2-2. Summary of additional search strategies for DIBP 2-3
Table 2-3. Inclusion criteria used to identify animal studies of health-related endpoints,
supporting data, or secondary literature 2-8
Table 2-4. Summary of search terms: targeted epidemiology search 2-9
Table 2-5. Inclusion criteria used to identify epidemiology studies of health-related endpoints 2-11
Table 2-6. Summary of additional search strategies for epidemiology studies of phthalate
exposure in relation to health-related endpoints 2-12
Table 2-7. Primary source epidemiological studies examining health effects of DIBP 2-12
Table 2-8. General and outcome-specific considerations for DIBP study evaluation 2-28
Table 2-9. Questions and relevant experimental information for the evaluation of experimental
animal studies 2-31
Table 3-1. Evidence pertaining to DIBP and sexual differentiation effects in humans 3-2
Table 3-2. Evidence pertaining to DIBP and semen parameters or infertility in adult men or
couples 3-4
Table 3-3. Evidence pertaining to DIBP and reproductive hormones in adult men 3-6
Table 3-4. Evidence pertaining to DIBP and the timing of male puberty or sex hormones in boys 3-7
Table 3-5. Evidence pertaining to DIBP and timing of female puberty or sex hormones in girls 3-9
Table 3-6. Evidence pertaining to DIBP and reproductive hormones in adult women 3-12
Table 3-7. Evidence pertaining to DIBP and gynecological conditions in humans 3-13
Table 3-8. Evidence pertaining to DIBP and pregnancy outcomes in humans 3-15
Table 3-9. Evidence pertaining to DIBP and allergy/immune effects in humans 3-18
Table 3-10. Evidence pertaining to DIBP and asthma/wheezing and hypersensitivity in humans 3-22
Table 3-11. Evidence pertaining to DIBP and neurodevelopmental effects in humans 3-25
Table 3-12. Evidence pertaining to DIBP and thyroid hormones in humans 3-30
Table 3-13. Evidence pertaining to DIBP and obesity in humans 3-31
Table 3-14. Evidence pertaining to DIBP and diabetes/insulin resistance in humans 3-36
Table 3-15. Evidence pertaining to DIBP and cardiovascular disease risk factors in humans 3-40
Table 3-16. Evidence pertaining to DIBP and cancer in humans 3-43
Table 3-17. Evidence pertaining to developmental effects in animals following oral exposure to
DIBP 3-44
Table 3-18. Evidence pertaining to male reproductive effects in animals following oral exposure
to DIBP 3-62
Table 3-19. Evidence pertaining to female reproductive effects in animals following oral
exposure to DIBP 3-73
Table 3-20. Evidence pertaining to hepatic effects in animals following oral exposure to DIBP 3-79
Table 3-21. Evidence pertaining to renal effects in animals following oral exposure to DIBP 3-83
Table 3-22. Evidence pertaining to hematopoietic effects in animals following oral exposure to
DIBP 3-87
Table 3-23. Evidence pertaining to other toxicity effects in animals following oral exposure to
DIBP 3-90
Table 3-24. Summary of mechanistic outcomes evaluated following DIBP or MIBP
administration 3-95
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
FIGURES
Figure 1-1. Chemical structure of DIBP 1-1
Figure 2-1. Literature search approach for DIBP 2-7
Figure 3-1. Exposure-response array of effects on developmental growth and survival following
developmental oral exposure to DIBP 3-59
Figure 3-2. Exposure-response array of effects on postnatal and adult body weight following
developmental oral exposure to DIBP 3-60
Figure 3-3. Exposure-response array of effects on fetal morphological developmental following
developmental oral exposure to DIBP 3-61
Figure 3-4. Exposure-response array of effects on male reproductive development following
developmental oral exposure to DIBP 3-70
Figure 3-5. Exposure-response array of effects on fetal testosterone (T) following
developmental oral exposure to DIBP 3-71
Figure 3-6. Exposure-response array of male reproductive effects following oral exposure to
DIBP 3-72
Figure 3-7. Exposure-response array of female reproductive effects, maternal weight and
toxicity, following oral exposure to DIBP 3-77
Figure 3-8. Exposure-response array of female reproductive effects, fertility and fetal survival,
following oral exposure to DIBP 3-78
Figure 3-9. Exposure-response array of liver effects following oral exposure to DIBP 3-82
Figure 3-10. Exposure-response array of kidney effects following oral exposure to DIBP 3-86
Figure 3-11. Exposure-response array of hematopoeitic effects following oral exposure to DIBP 3-89
Figure 3-12. Exposure-response array of effects on other toxicities following oral exposure to
DIBP 3-93
Figure 3-13. Summary of in vivo and in vitro mechanistic data for DIBP and MIBP by mechanistic
category 3-96
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
ABBREVIATIONS
AGO anogenital distance IQR
aOR adjusted odds ratio IRIS
BASC-PRS Behavior Assessment System for Koc
Children—Parent Rating Scales LDL
BBP butyl benzyl phthalate LH
BMI body mass index LMW
BP blood pressure LOD
BPA bisphenolA LOQ
BRIEF Behavior Rating Inventory of Executive MBzP
Function MBP
BW body weight MCPP
CASRN Chemical Abstracts Service Registry MDI
Number MEHP
CHAP Chronic Hazard Advisory Panel MEP
CI confidence interval MHBP
CPSC Consumer Product Safety Commission MIBP
DBF dibutyl phthalate MMP
DEP di-ethyl phthalate MOA
DEHP di(2-ethylhexyl)phthalate MOINP
DHEAS dehydroepiandrosterone MRI
DIBP diisobutyl phthalate NCEA
DINP diisononyl phthalate
DnBP dibutyl phthalate NHANES
DNA deoxyribonucleic acid
DPP dipentyl phthalate NHS
DXA dual energy x-ray absorptiometry NRC
EPA Environmental Protection Agency OR
FBG fasting blood glucose ORD
FDA Food and Drug Administration PAH
FSH follicle stimulating hormone PCO
GD gestational day PCOS
HbAlc glycosolated hemoglobin PDI
HCG human chorionic gonadotropin PND
HDL high-density lipoprotein PPS
HERO Health and Environmental Research PVC
Online RBC
Hgb hemoglobin SD
HOMA homeostatic model assessment SE
HOMA-IR homeostatic model assessment of SHBG
insulin resistance T3
HOME Health Outcomes and Measures of the T4
Environment TSH
IgE immunoglobulin E VO
ICC intra-class correlation coefficient VOC
IM-GSM grey scale media of the intima media WBC
complex WHO
IMT intima media thickness
interquartile range
Integrated Risk Information System
partition coefficient
low-density lipoprotein
luteinizing hormone
low molecular weight
level of detection
level of quantification
mono-benzyl phthalate
monobutyl phthalate
mono-(3-carboxypropyl) phthalate
mental delay index
mono-(2-ethylhexyl) phthalate
monoethyl phthalate
mono-3-(3-carboxypropyl)phthalate
monoisobutyl phthalate
monomethyl phthalate
mode of action
oxo-(mono-oxoisononyl) phthalate
magnetic resonance imaging
National Center for Environmental
Assessment
National Health and Nutrition
Examination Survey
Nurses' Health Study
National Research Council
odds ratio
Office of Research and Development
polycyclic aromatic hydrocarbon
polycystic ovarian morphology
polycystic ovarian syndrome
psychomotor delay index
postnatal day
preputial separation
polyvinyl chloride
red blood cell
standard deviation
standard error
sex-hormone binding globulin
triiodothyronine
thyroxine
thyroid stimulating hormone
vaginal opening
volatile organic compound
white blood cell
World Health Organization
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
PREFACE
3 This draft document presents preliminary materials for an assessment of diisobutyl
4 phthalate (DIBP) prepared by the U.S. Environmental Protection Agency's (EPA's) Integrated Risk
5 Information System (IRIS) Program. These preliminary materials include a planning and scoping
6 summary, information on the approaches used to identify pertinent literature, results of the
7 literature search, approaches for selection of studies for hazard identification, presentation of
8 critical studies in evidence tables and exposure-response arrays, and mechanistic information for
9 DIBP. This material is being released for public review and comment prior to a public meeting,
10 providing an opportunity for the IRIS Program to engage in early discussions with stakeholders and
11 the public on data that may be used to identify adverse health effects and characterize dose-
12 response relationships.
13 The planning and scoping summary includes information on the uses of DIBP, occurrence of
14 DIBP in the environment, and the rationale and scope for the development of the assessment This
15 information is responsive to recommendations in the 2009 National Research Council (NRC) report
16 Science and Decisions: Advancing Risk Assessment [NRC. 2009] related to planning and scoping in
17 the risk assessment process.
18 The preliminary materials are also responsive to the 2011 NRC report Review of the
19 Environmental Protection Agency's Draft IRIS Assessment of Formaldehyde [NRC, 2011]. The IRIS
20 Program's implementation of the NRC recommendations is following a phased approach that is
21 consistent with the NRC's "Roadmap for Revision" as described in Chapter 7 of the formaldehyde
22 review report The NRC stated that "the committee recognizes that the changes suggested would
23 involve a multi-year process and extensive effort by the staff of the National Center for
24 Environmental Assessment and input and review by the EPA Science Advisory Board and others."
25 Phase 1 of implementation has focused on a subset of the short-term recommendations, such as
26 editing and streamlining documents, increasing transparency and clarity, and using more tables,
27 figures, and appendices to present information and data in assessments. Phase 1 also focused on
28 assessments near the end of the development process and close to final posting. Phase 2 of
29 implementation is focused on assessments that are in the beginning stages of assessment
30 development The IRIS DIBP assessment is in Phase 2 and represents a significant advancement in
31 implementing the NRC recommendations. In the development of this assessment, many of the
32 recommendations are being implemented in full, while others are being implemented in part
33 Achieving full and robust implementation of certain recommendations will be an evolving process
34 with input and feedback from the public, stakeholders, and independent external peer review.
35 Phase 3 of implementation will incorporate the longer-term recommendations made by the NRC,
36 including the development of a standardized approach to describe the strength of evidence for
37 noncancer effects.
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 In May 2014, the NRC released their report reviewing the IRIS assessment development
2 process. As part of this review, the NRC reviewed current methods for evidence-based reviews and
3 made several recommendations with respect to integrating scientific evidence for chemical hazard
4 and dose-response assessments. In their report, the NRC states that EPA should continue to
5 improve its evidence-integration process incrementally and enhance the transparency of its
6 process. The committee did not offer a preference but suggests that EPA consider which approach
7 best fits its plans for the IRIS process. The NRC recommendations will inform the IRIS Program's
8 efforts in this area going forward. This effort is included in Phase 3 of EPA's implementation plan.
9 The literature search strategy, which describes the processes for identifying scientific
10 literature, screening studies for consideration, and identifying primary sources of health effects
11 data, is responsive to NRC recommendations regarding the development of a systematic and
12 transparent approach for identifying the primary literature for analysis. The preliminary materials
13 also describe EPA's approach for the selection of critical studies to be included in the evidence
14 tables, as well as the approach for evaluating methodological features of studies that will be
15 considered in the overall evaluation and synthesis of evidence for each health effect. The
16 development of these materials is in response to the NRC recommendation to thoroughly evaluate
17 critical studies with standardized approaches that are formulated and based on the type of research
18 (e.g., observational epidemiology or animal bioassays). In addition, NRC recommendations for
19 standardized presentation of key study data are addressed by the development of the preliminary
20 evidence tables and preliminary exposure-response arrays for primary health effect information.
21 EPA welcomes all comments on the preliminary materials in this document, including the
22 following:
23 • the clarity and transparency of the materials;
24 • the approach for identifying pertinent studies;
25 • the selection of critical studies for data extraction to preliminary evidence tables and
26 exposure-response arrays;
27 • any methodological considerations that could affect the interpretation of or confidence in
28 study results; and
29 • any additional studies published or nearing publication that may provide data for the
30 evaluation of human health hazard or dose-response relationships.
31 The preliminary evidence tables and exposure-response arrays should be regarded solely as
32 representing the data on each endpoint that have been identified as a result of the draft literature
33 search strategy. They do not reflect any conclusions as to hazard identification or dose-response
34 assessment.
35 After obtaining public input and conducting additional study evaluation and data
36 integration, EPA will revise these materials to support the hazard identification and dose-response
37 assessment in a draft Toxicological Review that will be made available for public comment
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
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2
1. INTRODUCTION
3 This introduction contains a planning and scoping summary for the Integrated Risk
4 Information System (IRIS) assessment of diisobutyl phthalate (DIBP). The planning and scoping
5 summary includes information on the properties, sources, and uses of DIBP, occurrence and fate of
6 DIBP in the environment, potential for human exposure, and the rationale for the development of
7 this assessment
8 1.1. DIBP IN THE ENVIRONMENT
9 1.1.1. Production and Use
10 DIBP (Figure 1-1) is used as a plasticizer (HSDB. 2013) in a wide range of materials
11 including polyvinyl chloride (PVC) formulations; paints; lacquers; varnish; paper, pulp and board
12 industry; as a softener; in viscosity adjustment; nail polish; cosmetics; lubricants; carpets; clothing
13 treatments; rubber dentistry settings; as a fuel stabilizer; as a concrete additive; explosive
14 materials; and printing inks. DIBP has also been classified by the Food and Drug Administration
15 (FDA) as an indirect food additive through its use as a component of adhesives. Because DIBP has
16 similar properties to di-n-butyl phthalate (DBF), it can be used as a substitute for DBF (HSDB.
17 2013). Approximately 500,000 pounds were manufactured in the United States in 2012
18 (http://www.epa.gov/oppt/cdr/index.html). In July 2014, the Consumer Product Safety
19 Commission's (CPSC) Chronic Hazard Advisory Panel (CHAP) recommended that DIBP be
20 permanently banned from use in children's toys and child care articles at levels greater than 0.1%
21 fCHAP. 20141
22
H;C
23
24 Figure 1-1. Chemical structure of DIBP (HSDB. 2013).
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 1.1.2. Environmental Fate
2 If released to air, DIBP will exist in both the vapor and particulate phases in the atmosphere.
3 Vapor-phase DIBP will be photolytically degraded with a half-life of about 1.2 days, and particulate-
4 phase DIBP will be removed from the atmosphere by wet or dry deposition [HSDB. 2013). In soil,
5 DIBP is expected to have low mobility due to a moderately high organic carbon partition coefficient
6 (Koc). Biodegradation in aerobic soil and water is expected to occur over days or weeks. Anaerobic
7 biodegradation rates are expected to be slower. Volatilization from moist soil or water is expected
8 to be an important fate process for DIBP, but volatilization from dry soil is not expected. If released
9 into water, DIBP is expected to adsorb to sediments and solids, and volatilization from water
10 surfaces is expected to be an important process. An estimated bioconcentration factor of 240
11 suggests that there is a potential for the chemical to concentrate in aquatic organisms, but
12 metabolism in the organisms can reduce accumulation [HSDB. 2013). As noted by Wormuth et al.
13 [2006]. the majority of phthalates that are found in the environment come from their slow releases
14 from plastics and other phthalate-containing articles. Certain waste streams, sludges, and
15 industrially contaminated sites, however, may contain higher levels of phthalates than other sites.
16 1.1.3. Human Exposure Pathways
17 The routes by which humans are exposed to phthalates and the magnitude of individual
18 phthalate exposures have changed over time as the quantities and uses of the various phthalates
19 have changed. Human exposure to phthalates occurs mainly in occupational or household settings
20 because they are used and released from products in the home environment. Environmental
21 concentrations of phthalates are typically the highest in house dust, and they may be present in
22 food due to the use of phthalates in packaging and food preparation materials. For most phthalates,
23 food ingestion is the dominant pathway of exposure, with dust exposures (ingestion and dermal
24 contact) and inhalation also being important in some circumstances. Infant and toddler exposures
25 occur due to teething and playing with plastic toys that contain phthalates [Wormuth etal.. 2006).
26 The presence of parent phthalates or their metabolites in a body matrix, such as blood or
27 urine, provides evidence of exposure to that chemical. The predominant metabolite of DIBP in
28 humans is monoisobutyl phthalate (MIBP). Zotaetal. [2014] evaluated the prevalence and
29 temporal trends of MIBP in urine samples collected as part of the National Health and Nutrition
30 Examination Survey [NHANES] conducted between 2001 and 2010. MIBP was found in 72% of the
31 samples in the 2001-2002 cycle and 96% of the samples in the 2009-2010 cycle, and increased in
32 concentration over time, starting at about 2.4 ng/mL in the 2001-2002 cycle, and rising to about
33 7.8 ng/mL in the 2009-2010 cycle.
34 Intake exposures can be estimated on a pathway-basis by combining exposure media
35 concentrations and contact rates. Using this approach, Clark etal. [2011] estimated median intakes
36 of DIBP for various lifestages as defined by the authors: between 0.75 and 1.0 [ig/kg-day for teens
37 (12-19 years of age] and adults (20-70 years of age], based on ingestion of food, drinking water,
38 dust/soil, and inhalation of air; and between 1.3 and 2.6 [ig/kg-day for infants (0-0.5 years of age],
39 toddlers (ages 0.5-4 years of age], and children (5-11 years of age]. The exposure was found to be
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 dominated by food, with inhalation of indoor air also important. The intakes determined by Clark
2 etal. [2011] were higher than those found by Wormuth etal. [2006], who determined intakes for
3 these age ranges at about <0.5 [ig/kg-day. Clark etal. [2011] attributed this difference to use of
4 higher food concentrations in the estimates.
5 Wittassek et al. [2011] reported median intakes of DIBP in the range of 0.1-1.7 |ig/kg-day
6 based on a literature survey of urinary biomonitoring data and intake estimates provided therein.
7 Their review included a single study in the United States of a cohort of pregnant woman that found
8 median intakes at 0.1 [ig/kg-day. Three other studies from Germany had median intakes ranging
9 from 1.1 to 1.7 [ig/kg-day. Qianetal. [2014] used NHANES 2007-2008 and found a median intake
10 of 0.2 [ig/kg-day and a 95th percentile intake of 0.9 [ig/kg-day. Christensen et al. [2014] combined
11 the data from NHANES 2005-2008 and found similar results to Qianetal. [2014], with a median
12 over that time span of 0.2 [ig/kg-day and a 95th percentile intake of 0.8 [ig/kg-day.
13 1.2. SCOPE OF THE ASSESSMENT
14 The National Research Council [NRC] has recommended that, "[Cumulative risk assessment
15 based on common adverse outcomes is a feasible and physiologically relevant approach to the
16 evaluation of the multiplicity of human exposures and directly reflects EPA's mission to protect
17 human health" [NRC. 2008. pll]. They envisioned facilitating the process by "defining the groups
18 of agents that should be included for a given outcome" [NRC. 2008. p!2]. In humans, the NRC cited
19 results from NHANES that demonstrate exposure to multiple phthalates in most people [NRC. 2008.
20 p23-25]. A recent review of human exposure to eight phthalates estimated that indoor air
21 contributed to approximately 25% of DIBP exposure in children [CHAP, 2014, Appendix El, p35].
22 The unique exposure scenarios and potential sensitivities of children contribute to the need for an
23 assessment of phthalate toxicity. This IRIS assessment will help to inform EPA programs and
24 regions of the potentially unique vulnerabilities of children to DIBP exposure and enable future
25 cumulative risk assessments that assess effects on human health outcomes that might be associated
26 with DIBP and other phthalates. There is currently no IRIS assessment of DIBP.
27
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
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2 2. METHODS FOR IDENTIFYING AND SELECTING
3 STUDIES
4 The NRCf20111 recommended that the U.S. Environmental Protection Agency (EPA)
5 develop a detailed search strategy utilizing a graphical display documenting how initial search
6 findings are narrowed to the final studies that are selected for further evaluation on the basis of
7 inclusion and exclusion criteria. Following these recommendations, a literature search and
8 screening strategy was applied to identify literature related to characterizing the health effects of
9 diisobutyl phthalate (DIBP). This strategy consisted of a search of online scientific databases and
10 other sources, casting a wide net in order to identify all potentially pertinent studies. In subsequent
11 steps, references were screened to exclude papers not pertinent to an assessment of the health
12 effects of DIBP, and remaining references were sorted into categories for further evaluation.
13 Section 2.1 describes the literature search and screening strategy in detail. The NRG [2011] further
14 recommended that after studies are identified for review by utilizing a transparent search strategy,
15 the next step is to summarize the details and findings of the most pertinent studies in the evidence
16 tables. The NRC suggested that such tables should provide a link to the references, and include
17 details of the study population, methods, and key findings. This approach provides for a systematic
18 and concise presentation of the evidence. The NRC also recommended that the methods and
19 findings should then be evaluated with a standardized approach. The approach that was outlined
20 identified standard issues for the evaluation of epidemiological and experimental animal studies.
21 Section 2.2 describes the approach taken for DIBP for selecting studies to be included in the
22 preliminary evidence tables and exposure-response arrays. Section 3 presents the selected studies
23 in preliminary evidence tables and exposure-response arrays, arranged by health effect
24 2.1. DRAFT LITERATURE SEARCH AND SCREENING STRATEGY
25 The literature search for DIBP was conducted in four online scientific databases (PubMed,
26 Web of Science, Toxline, and Toxic Substances Control Act Test Submissions (TSCATS2)) in
27 February of 2013; the search was repeated in March of 2014. This document is complete through
28 March 2014. Additional updates will be performed at regular (e.g., 6-month) intervals. The
29 detailed search approach, including the search strings and number of citations identified per
30 database, is presented in Table 2-1. The search strings and search terms described for DIBP
31 captured studies using the parent compound and metabolites (i.e., the active metabolite,
32 monoisobutyl phthalate [MIBP]). This search of online databases identified 504 citations (after
33 electronically eliminating duplicates). The computerized database searches were also
34 supplemented by a manual search of citations from other regulatory documents (Table 2-2);
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review of Diisobutyl Phthalate
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2
3
343citations were obtained using these additional search strategies. In total, 809 citations were
identified using online scientific databases and additional search strategies.
Table 2-1. Database search strategy for DIBP
Database
(search date)
Keywords3
PubMed
03/2014
02/2013
dibp OR (mibp AND phthalate) OR "diisobutylphthalate" OR "di-isobutyl phthalate" OR "84-69-
5" OR "diisobutyl phthalate" OR "di(i-butyl)phthalate" OR "di-iso-butyl phthalate" OR "isobutyl
phthalate" OR "phthalic acid diisobutyl ester" OR ("diisobutyl ester" AND phthalate) OR "1,2-
benzenedicarboxylic acid bis(2-methylpropyl) ester" OR "1,2-benzenedicarboxylic acid 1,2-
bis(2-methylpropyl) ester" OR "monoisobutyl phthalate" OR "mono(i-butyl)phthalate" OR
"mono-iso-butyl phthalate" OR "phthalic acid monoisobutyl ester" OR "1,2-
benzenedicarboxylic acid, mono(2-methylpropyl) ester" OR "2-[(2-
methylpropoxy)carbonyl]benzoic acid" OR "1,2-benzenedicarboxylic acid, mono(2-
methylpropyl) ester (9CI)" OR "isobutyl hydrogen phthalate" OR "1,2-benzenedicarboxylic acid
l-(2-methylpropyl) ester"
Web of Science
03/2014
02/2013
TS=dibp OR (TS=mibp AND TS=phthalate) OR TS="diisobutylphthalate" OR TS="di-isobutyl
phthalate" ORTS="84-69-5" ORTS="diisobutyl phthalate" ORTS="di(i-butyl)phthalate" OR
TS="di-iso-butyl phthalate" ORTS="isobutyl phthalate" ORTS="phthalicacid diisobutyl ester"
OR (TS="diisobutyl ester" AND TS=phthalate) OR TS="l,2-benzenedicarboxylic acid bis(2-
methylpropyl) ester" ORTS="l,2-benzenedicarboxylicacid l,2-bis(2-methylpropyl) ester" OR
TS="monoisobutyl phthalate" ORTS="mono(i-butyl)phthalate" ORTS="mono-iso-butyl
phthalate" ORTS="phthalicacid monoisobutyl ester" ORTS="l,2-benzenedicarboxylicacid,
mono(2-methylpropyl) ester" OR TS="2-[(2-methylpropoxy)carbonyl]benzoic acid" OR
TS="l,2-benzenedicarboxylic acid, mono(2-methylpropyl) ester (9CI)" OR TS="isobutyl
hydrogen phthalate" ORTS="l,2-benzenedicarboxylicacid l-(2-methylpropyl) ester"
Toxline
03/2014
02/2013
Split into 4 separate search strings:
@TERM+@rn+84-69-5
@AND+mibp+phthalate
@AND+"diisobutyl ester"+phthalate
@OR+(dibp+"diisobutylphthalate"+"di-isobutyl+phthalate"+"diisobutyl+phthalate"+"di(i-
butyl)phthalate"+"di-iso-
butyl+phthalate"+"isobutyl+phthalate"+"phthalic+acid+diisobutyl+ester"+"l,2-
benzenedicarboxylic+acid+bis(2-methylpropyl)+ester"+"l,2-benzenedicarboxylic+acid+l,2-
bis(2-methylpropyl)+ester"+"monoisobutyl+phthalate"+"mono(i-butyl)phthalate"+"mono-iso-
butyl+phthalate"+"phthalic+acid+monoisobutyl+ester"+"l,2-
benzenedicarboxylic+acid,+mono(2-methylpropyl)+ester"+"2-[(2-
methylpropoxy)carbonyl]benzoic+acid"+"l,2-benzenedicarboxylic+acid,+mono(2-
methylpropyl)+ester+(9CI)"+"isobutyl+hydrogen+phthalate"+"l,2-
benzenedicarboxylic+acid+l-(2-methylpropyl)+ester")
TSCATS2
03/2014
(2000-) 84-69-5
5
6
7
aThe search strings and search terms described above captured studies using the parent compound and the
metabolite MIBP.
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review of Diisobutyl Phthalate
Table 2-2. Summary of additional search strategies for DIBP
System used
Selected key reference(s) or sources
Date
Additional
references
identified
Manual search of
citations from
regulatory
documents
CPSC (2010). Toxicity Review for Diisobutyl phthalate
(DIBP). Bethesda, MD: Consumer Product Safety
Commission.
3/2014
9 citations added
Web of Science,
forward search
Hannas et al. (2011). Dose-response assessment of fetal
testosterone production and gene expression levels in
rat testes following in utero exposure to diethylhexyl
phthalate, diisobutyl phthalate, diisoheptyl phthalate,
and diisononyl phthalate. Toxicol Sci. 123(1):206-16.
Saillenfait et al. (2008). Diisobutyl phthalate impairs the
androgen-dependent reproductive development of the
male rat. Reprod Toxicol. 26(2):107-15.
Rayet al. (2012). Ovarian development in Wistar rat
treated prenatally with single dose diisobutyl phthalate.
Bratisl Lek Listy. 113(10):577-82.
Kleinsasser et al. (2001b). Genotoxicity of di-butyl-
phthalate and di-iso-butyl-phthalate in human
lymphocytes and mucosal cells. Teratog Carcinog
Mutagen. 21(3):189-96.
3/2014
2 citations added
3/2014
3/2014
3/2014
1 citation added
0 citations added
1 citation added
Web of Science,
backward search
Hannas et al. (2011). Dose-response assessment of fetal
testosterone production and gene expression levels in
rat testes following in utero exposure to diethylhexyl
phthalate, diisobutyl phthalate, diisoheptyl phthalate,
and diisononyl phthalate. Toxicol Sci. 123(1):206-16.
Saillenfait et al. (2008). Diisobutyl phthalate impairs the
androgen-dependent reproductive development of the
male rat. Reprod Toxicol. 26(2):107-15.
Rayet al. (2012). Ovarian development in Wistar rat
treated prenatally with single dose diisobutyl phthalate.
Bratisl Lek Listy. 113(10):577-82.
Kleinsasser et al. (2001b). Genotoxicity of di-butyl-
phthalate and di-iso-butyl-phthalate in human
lymphocytes and mucosal cells. Teratog Carcinog
Mutagen. 21(3):189-96.
3/2014
1 citation added
3/2014
3/2014
3/2014
1 citation added
4 citations added
2 citations added
Snowball search
DIBP references in previous assessment or previously
added to the HERO project page
4/2014
45 citations added
Background Check
Searched a combination of CASRNs and synonyms on the
following databases:
ACGIH (http://www.acgih.org/home.htm)
ATSDR (http://www.atsdr.cdc.gov/substances/index.asp)
CalEPA Office of Environmental Health Hazard Assessment
(http://www.oehha.ca.gov/risk.html)
2/2013,
update
3/2014
17 citations added
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
System used
Selected key reference(s) or sources
Date
Additional
references
identified
OEM HA Toxicity Criteria Database
(http://www.oehha.ca.gov/tcdb/index.asp)
Biomonitoring California-Priority Chemicals
(http://www.oehha.ca.gov/multimedia/biomon/pdf/Priori
tvChemsCurrent.pdf)
Biomonitoring California-Designated Chemicals
(http://www.oehha.ca.gov/multimedia/biomon/pdf/Desig
natedChemCurrent.pdf)
Cal/Ecotox database
(http://www.oehha.ca.gov/scripts/cal ecotox/CHEMLIST.
ASP)
OEHHA Fact Sheets
(http://www.oehha.ca.gov/public info/facts/index.html)
Non-cancer health effects Table (RELs) and Cancer
Potency Factors (Appendix A and Appendix B)
(http://www.oehha.ca.gov/air/hot spots/index.html)
CPSC (http://www.cpsc.gov)
eChemPortal
(http://www.echemportal.Org/echemportal/participant/page.a
ction?pagelD=9)
Environment Canada - Search entire site if not found below:
(http://www.ec.gc.ca/default.asp?lang=En&n=ECD35C36)
Toxic Substances Managed under CEPA
(http://www.ec.gc.ca/toxiques-
toxics/Default.asp?lang=En&n=98E80CC6-l)
Screening Assessment reports
Risk Management reports
Final Assessments (http://www.ec.gc.ca/lcpe-
cepa/default.asp?lang=En&xml=09F567A7-BlEE-lFEE-73DB-
8AE6C1EB7658)
Draft Assessments (http://www.ec.gc.ca/lcpe-
cepa/default.asp?lang=En&xml=6892C255-5597-C162-95FC-
4B905320F8C9)
EPA Acute Exposure Guideline Levels
(http://www.epa.gov/oppt/aegl/pubs/chemlist.htm)
EPA - IRISTrack/New Assessments and Reviews
EPA NSCEP (http://www.epa.gov/ncepihom/)
EPA RfD/RfC and CRAVE meeting notes
EPA Science Inventory (http://cfpub.epa.gov/si/)
FDA (http://www.fda.gov/)
Federal Docket (www.regulations.gov)
Health Canada First Priority List Assessments (http://www.hc-
sc.gc.ca/ewh-semt/pubs/contaminants/psll-lspl/index-
Health Canada Second Priority List Assessments
(http://www.hc-sc.gc.ca/ewh-semt/pubs/contaminants/psl2-
Isp2/index-eng.php)
IARC (http://monographs.iarc.fr/htdig/search.html)
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
System used
Selected key reference(s) or sources
ITER (TERA database)
(http://iter.ctcnet.net/publicurl/pub search list.cfm)
NAP - Search Site (http://www.nap.edu/)
NRC - AEGLs via NAP search for "Acute Exposure Guideline
Level" and the chemical
NCI (http://www.cancer.gov)
National Institute for Environmental Health Sciences (NIEHS)
http://www.niehs.nih.gov/
NICNAS (PEC only covered by eChemPortal)
(http://www.nicnas.gov.au/industrv/aics/search.asp)
NIOSH (http://www.cdc.gov/niosh/topics/)
NIOSHTIC2 (http://www2a.cdc.gov/nioshtic-2/)
NTP - RoC, status, results, and management reports
(http://ntpsearch.niehs.nih.gov/auerv.html)
OSHA
(http://www.osha.gov/dts/chemicalsampling/toc/toc chemsa
mp.html)
RTECS http://www.ccohs.ca/search.html
Date
Additional
references
identified
1
2 These citations were screened using the title, abstract, and in limited instances, full text for
3 pertinence to examining the health effects of DIBP exposure. The citations were then screened
4 using inclusion criteria (Table 2-3) describing specific information to help identify primary source
5 health effect data and mechanistic and/or genotoxic data, as well as resources useful in preparation
6 of the DIBP package. The process for screening the literature search is described below and is
7 shown graphically in Figure 2-1:
8 • 31 references were identified as animal studies with health effects data and were
9 considered for data extraction to evidence tables and exposure-response arrays.
10 • 54 references were identified as supporting studies; of these, 12 were toxicokinetic studies
11 and 43 were mechanistic and genotoxicity studies.
12 • 97 references were identified as secondary literature (e.g., reviews and editorials, risk
13 assessments, meta analyses, and regulatory documents); these references were kept as
14 additional resources for development of the Toxicological Review.
15 • 632 references were excluded because these studies did not include primary source data
16 evaluating DIBP in relation to any kind of toxicity or health endpoint, and did not provide
17 either supporting information (e.g., toxicokinetic or mechanistic/genotoxicity data) or
18 secondary literature information (see Figure 2-1 and Table 2-3 for inclusion categories and
19 criteria).
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 Note that some studies were identified as belonging to multiple categories. As a result, the
2 total number of studies in a given category may be less than the sum of the individual studies listed
3 in subcategories. For example, the category "Studies with Supporting Data" included one study that
4 contained information relevant to both the toxicokinetics and mechanistic and/or genotoxicity
5 subcategories.
6 Among the studies identified in the DIBP literature searches, there were a number of
7 foreign language studies. Based on a review of the English titles and, when available, English
8 abstracts, two of the foreign language articles, Maetal. [2013b] and lijo [1975], were tagged as
9 toxicity studies and four foreign language articles, Ma etal. [2010], Maetal. [2013c], Kleinsasser et
10 al. [1999], and Kleinsasser et al. [2001a], were tagged as mechanistic and genotoxicity studies. The
11 other foreign language articles were excluded (tagged to Excluded: No primary data on toxic
12 effects]. Maetal. [2013b] is a report of neurotoxicological effects after DIBP exposure. With the
13 exception of one study [University of Rochester. 1954] that assessed brain weight, the Ma et al.
14 [2013b] article was the only available neurotoxicological study; this article was translated into
15 English (certified translation, Maetal., 2013a]. The remaining five foreign language articles
16 [above], tagged to toxicity studies or mechanistic and genotoxicity studies, have not yet been
17 translated or considered for inclusion in either evidence or mechanistic tables. These studies will
18 be further evaluated and considered during the development of the draft assessment of the
19 available evidence of DIBP-induced health effects.
20
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
3
Database Searches
(see Table 2-1 for keywords and limits)
PubMed
n = 243
Web of
Science
n = 310
Toxline
(incl.TSCATS)
(After duplicates removed electronically)
n=504
Additional Search
Strategies
(see Table 2-2 for
methods and results)
n = 343
Phthalates - Epidemiological
Studies Search
(see Table 2-4 for keywords and limits)
Primary Source Human Data
n = 185
(See Table 2-5 for Inclusion Criteria)
Combined Dataset
(After all duplicates removed)
n=809
Manual Screening For Pertinence
(Title/Abstract/Full Text)
(see Table 2-3 for inclusion criteria)
Excluded: No Primary Data on Toxic Effects
(n=632)
14 Abstract Only
187 Not Chemical Specific
66 Manufacture/use
23 Chemical Treatment/Disposal/Remediation
12 Use in sample prep or assay
59 Measurement Methods
16 Miscellaneous
14 Ecosystem Effect
179 Exposure levels
52 Fate and Transport
25 Chemical/physical properties
9 Mixtures only
Other Studies:
Studies with Supporting Data (n=54)
12 Toxicokinetics
43 Mechanistic and Genotoxicrty Studies
Secondary Literature (n=97)
49 Reviews/editorials
26 Regulator,' documents
27 Risk assessments
1 Meta analyses
selection of studies
that include DIBP
Animal Primary
Source Health
Effects Studies
(n = 31)
Human Primary
Source Health
Effects Studies
(n = 52)
(See Table 2-7 for a listing
of DIBP-specific
epidemiological studies)
Note: Studies containing multiple information categories were sorted into multiple tags. For this reason, the
subcategory numbers do not always add up to the category total.
Figure 2-1. Literature search approach for DIBP.
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
2 Table 2-3. Inclusion criteria used to identify animal studies of health-related
3 endpoints, supporting data, or secondary literature
Inclusion criteria3
Did the study evaluate effects of DIBP or its metabolites known to be formed in humans?
Did the study evaluate effects in a tissue (organ) or cells derived from a tissue (organ)?
Did the study evaluate cellular, biochemical or molecular effects relevant to any mode of action?
or
Does the study include information from other agencies, risk assessments, or reviews that would aid in
the development of a toxicological review of DIBP?
4
5 alf the answer is "no" to any of these criteria questions, the study was placed under "No Primary Data on Toxic
6 Effects."
7
8 Thirty-six human studies were also identified from the initial literature search using the
9 search strings presented in Table 2-1. However, work being done concurrently on the development
10 of other phthalate preliminary materials revealed that this set of DIBP epidemiology studies was
11 incomplete. Epidemiology studies frequently examine multiple compounds (e.g., metabolites of
12 several different phthalates). The indexing terms and abstracts may not include a comprehensive
13 list of all of the specific phthalates examined, resulting in the inappropriate exclusion of studies and
14 the potential for introduction of bias in the selection process. Specifically, "negative" studies (i.e.,
15 studies that did not demonstrate an association between exposure and disease) are potentially
16 more likely to be missed than "positive" studies. This issue did not arise in the search process for
17 experimental (animal toxicology) studies, for which the test compound is virtually always identified
18 through search terms or key word searches of abstracts.
19 Another issue encountered in the development of the search and screening process for the
20 phthalate epidemiology studies relates to the duplication of efforts involved in the development of
21 EPA's health assessments for several individual phthalates (e.g., dibutyl phthalate [DBF], DIBP,
22 butyl benzyl phthalate [BBP], di(2-ethylhexyl)phthalate [DEHP], di-ethyl phthalate [DEP],
23 diisononyl phthalate [DINP], and dipentyl phthalate [DPP]). In contrast to animal toxicology
24 studies, most of the epidemiology studies examine more than one phthalate, resulting in
25 considerable overlap in the sets of studies identified using individual-phthalate search terms. Full
26 text screening of the same studies identified in multiple searches results is an inefficient use of
27 resources.
28 For these reasons, EPA developed a process for identifying epidemiological studies
29 evaluating phthalates by performing a single broad search to create a listing of epidemiological
30 studies of all phthalates mentioned above, from which the selection of studies examining potential
31 health effects of an individual phthalate could be drawn. This list records each of the phthalates
32 included in the study, based on information in the methods section of the paper, and the outcome(s)
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 examined. This literature search for epidemiological studies examining phthalates in relation to
2 health-related endpoints (from which the DIBP studies were drawn) was conducted in PubMed,
3 Web of Science, and ToxNet databases in June 2013, using keywords and limits described in
4 Table 2-4; the search was updated in December 2013 and in June 2014. For this search, "phthalate"
5 (and related terms) rather than names of specific phthalates was used as the foundation of the
6 search, along with terms designed specifically to identify epidemiological studies. These terms
7 were based on terms used in previously identified epidemiology studies of six different phthalates.
Table 2-4. Summary of search terms: targeted epidemiology search
Database,
search date
Terms
Hits
June 2013 search
PubMed
06/2013
No date restriction
(phthalate OR phthalates OR phthalic acid) AND
(human OR case-control OR pregnancy OR cohort OR
workers OR children OR survey)
Imported: 2,505
After duplicates deleted: 2,482
Web of Science
06/2013
No date restriction
(TS="phthalic acid" ORTS="phthalate" OR
TS="phthalates") AND (TS="humans" ORTS="human"
ORTS="case-control" ORTS="pregnancy" OR
TS="cohort" ORTS="workers" ORTS="child" OR
TS="children" OR TS="survey")
Imported: 1,840
After duplicates deleted: 1,836
ToxNet
06/2013
No date restriction
(phthalate OR phthalates OR phthalic acid) AND
(human OR case-control OR pregnancy OR cohort OR
workers OR children OR survey)
Imported: 2,505
After duplicates deleted: 2,426
Merged
Reference Set
Merged dataset, with duplicates eliminated through
electronic screen
4,127
Epidemiology articles meeting inclusion criteria
127
December 2013
search
PubMed
Web of Science
ToxNet
Merged Reference Set
Additional epidemiology articles meeting inclusion
criteria
155
249
114
350
22
June 2014
search
PubMed
Web of Science
ToxNet (was not searched because no articles have
been found solely through this source in all the
previous searches)
Merged Reference Set
Additional epidemiology articles meeting inclusion
criteria
184
409
0
494
24
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 More than 4,000 citations were identified through this search. These were then screened
2 using inclusion criteria describing specific population (i.e., human), exposure measures,
3 comparison, and health effects (Table 2-5). Note that other studies obtained in the search, for
4 example mechanistic and pharmacokinetic studies, are excluded from consideration with respect to
5 the specific objective of this search (i.e., identification of epidemiology studies), but could be
6 included in other steps in the assessment. Duplicate citations of the same article were excluded,
7 and articles written in a language other than English were retained for subsequent review. Earlier
8 analyses that are updated in a subsequent paper (e.g., with a larger sample size) are not included as
9 a primary paper, but may be used as background material regarding study methods.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 Table 2-5. Inclusion criteria used to identify epidemiology studies of health-
2 related endpoints
Inclusion criteria
Is the study population humans?
and
Is exposure to one or more phthalate (parent compound or metabolite(s)a...
- measured in air, dust, or biological tissue?
- based on knowledge of industrial hygiene (occupational settings)?
- based on knowledge of specific contamination sites or accidental exposure?
and
Does the study compare a health effect in higher versus lower or no exposure?
and
Does the study include a measure of one or more primary health effect endpoints relating to...
- sexual differentiation measures (e.g., male genital malformations, anogenital distance, gender-related
play behavior)
- male reproductive effects (e.g., steroidal and gonadotropin hormone levels, measures of male-
mediated infertility)?
- female reproductive effects (e.g., steroidal and gonadotropin hormone levels, measures of female-
mediated infertility, gynecological conditions)?
- pregnancy outcomes (e.g., birth weight, gestation age)?
- puberty (male and female) (e.g., timing of development, precocious puberty, gynecomastia)?
- neurodevelopment (infants and children) (e.g., standardized tests of reflexes, behavior, and
intelligence)?
- thyroid effects (e.g., thyroid stimulating hormone and thyroid hormones, subclinical and clinical thyroid
disease)?
- immune system effects (e.g., asthma, allergies, immunoglobulin E (IgE) levels, skin prick tests)?
- pulmonary function (e.g., standardized test of lung volume, diffusing capacity)?
- neurological effects (adults) (e.g., peripheral neuropathy, vision or hearing or other sensory tests)?
- liver effects (e.g., cholestasis, biomarkers of liver function)?
- kidney effects (e.g., end stage renal disease, biomarkers of kidney function)?
- diabetes and measures of insulin resistance?
- obesity (and other measures of adiposity)?
- cardiovascular disease (cause-specific incidence or mortality)?
- cardiovascular risk factors (e.g., triglyceride and lipid levels, blood pressure or hypertension)?
- cancer (cause-specific incidence or mortality)?
or
Does the study include a measure of one or more secondary health effect endpoints (to be considered
within context of mechanistic evidence) relating to...
- oxidative stress?
- inflammation?
-gene expression?
3
4 aFor DIBP, the primary metabolite of interest is MIBP.
5
6 One hundred and seventy-three epidemiological studies examining one or more phthalates
7 in relation to one or more endpoints were identified by the searches conducted through June 2014
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 (127 in the initial search, 22 in the December 2013 update, and 24 in the June 2014 update;
2 Figure 2-1). Other strategies were also used to supplement this broad search for epidemiology
3 studies of phthalates), resulting in the identification of 12 additional publications (Table 2-6), for a
4 total of 185 epidemiological studies. From this set of all of the epidemiological studies examining
5 any phthalate, 52 studies analyzed one or more health effects in relation to a measure of DIBP
6 (Table 2-7).
7
8
Table 2-6. Summary of additional search strategies for epidemiology studies
of phthalate exposure in relation to health-related endpoints
Approach used
Testing and refinement of search terms based on terms used for the
identified articles within each category
Review of references cited in the identified list of epidemiology studies
("backward" search)
Electronic forward search through Web of Science of one to three
studies within each health endpoint category (early studies within each
category generally selected to maximize potential for citation in
subsequent publications)3
Date
performed
June 2014
July 2014
July 2014
Number of additional
citations identified
6
1
5
9
10
11
12
13
14
15
16
aThe following studies were used to conduct the forward searches: Trasande et al. (2013b); James-Todd et al.
(2012); Lind and Lind (2011); Boas et al. (2010); Cho et al. (2010); Engel et al. (2010); Lopez-Carrillo et al. (2010);
Wolff etal. (2010); Adibietal. (2009); Chouetal. (2009); Hatch et al. (2008); Wolff etal. (2008); Meeker et al.
(2007); Stahlhut et al. (2007); Mauser et al. (2006); Reddyet al. (2006); Jonssonetal. (2005); Swan et al. (2005);
Bornehagetal. (2004); Hoppin etal. (2004); Aschengrau et al. (1998); Heineman etal. (1992); Nielsen etal.
(1989); Nielsen etal. (1985).
17
18
Table 2-7. Primary source epidemiological studies examining health effects of
DIBP
Outcome category
Sexual differentiation measures
(Table 3-1)
Male reproductive (semen
parameters, infertility, and
hormones)
(Tables 3-2 and 3-3)
Male pubertal development
(Table 3-4)
Female pubertal development
(Table 3-5)
Reference3
Swan (2008)
Swan et al. (2010)
Buck Louis etal. (2014)
Joensen etal. (2012)
Kranvogl et al. (2014)
Mendiola etal. (2011)
Wirth et al. (2008)
Mieritzetal. (2012)
Mouritsen etal. (2013b)
Frederiksen et al. (2012)
Hart et al. (2013)
Lomenicketal. (2010)
Mouritsen etal. (2013b)
DIBP measure
MIBP (maternal urine)
MIBP (maternal urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (maternal urine)
MIBP (urine)
Sum MIBP +MBP (urine)3
Sum MIBP+ MBP (urine)3
MIBP (maternal serum)
MIBP (urine)
Sum MIBP+MBP(urine)3
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Outcome category
Female reproductive (infertility,
hormones, gynecological conditions)
(Tables 3-6 and 3-7)
Pregnancy outcomes (fetal growth,
preterm birth)
(Table 3-8)
Immune: allergy (rhinitis, eczema)
(Table 3-9)
Immune: asthma
(Table 3-10)
Neurodevelopment
(Table 3-11)
Thyroid
(Table 3-12)
Obesity
(Table 3-13)
Diabetes and insulin resistance
(Table 3-14)
Other cardiovascular disease risk
factors
(Table 3-15)
Reference3
Buck Louis etal. (2013)
Hart et al. (2013)
Sathyanarayana et al. (2014)
Upson etal. (2013)
Ferguson etal. (2014a)
Ferguson et al. (2014b)
Huang etal. (2014b)
Meeker et al. (2009)
Philippatetal. (2012)
Wolff etal. (2008)
Ait Bamai etal. (2014)
Bornehag et al. (2004)
Callesen et al. (2014a)
Callesen et al. (2014b)
Hoppinetal. (2013)
Sun etal. (2009)
Ait Bamai etal. (2014)
Bertelsen et al. (2013)
Callesen et al. (2014a)
Callesen et al. (2014b)
Hoppinetal. (2013)
Sun etal. (2009)
Braunetal. (2014)
Engel etal. (2010)
Kobrosly et al. (2014)
Tellez-Roio et al. (2013)
Whyatt et al. (2012)
Dirtu et al. (2013)
Meeker and Ferguson (2011)
Buseretal. (2014)
Dirtu et al. (2013)
Hart et al. (2013)
Kasper-Sonnenberg et al. (2012)
Lindetal. (2012a)
Olsen et al. (2012)
Svensson et al. (2011)
Teitelbaumetal. (2012)
Trasandeetal. (2013a)
Wang et al. (2013)
Huang etal. (2014a)
James-Todd etal. (2012)
Lindetal. (2012b)
Olsen et al. (2012)
Svensson et al. (2011)
Trasandeetal. (2013c)
Lind and Lind (2011)
Shiue (2014)
Trasandeetal. (2013b)
DIBP measure
MIBP (urine)
MIBP (maternal serum)
MIBP (maternal urine)
MIBP (urine)
MIBP (maternal urine)
MIBP (maternal urine)
DIBP (cord blood)
MIBP (maternal urine)
MIBP (maternal urine)
MIBP (maternal urine)
DIBP (dust)
DIBP (dust)
MIBP (urine)
DIBP (dust)
MIBP (urine)
DIBP (dust)
DIBP (dust)
MIBP (urine)
MIBP (urine)
DIBP (dust)
MIBP (urine)
DIBP (dust)
MIBP (maternal urine)
MIBP (maternal urine)
MIBP (maternal urine)
MIBP (maternal urine)
MIBP (maternal urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (maternal serum)
Sum MIBP + OH-MIBP (urine)
MIBP (serum)
MIBP (serum)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (urine)
MIBP (serum)
MIBP (serum)
MIBP (urine)
MIBP (urine)
MIBP (serum)
MIBP (urine)
MIBP (urine)
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Outcome category
Cancer
(Table 3-16)
Reference3
Olsen et al. (2012)
Lopez-Carrillo et al. (2010)
DIBP measure
MIBP (serum)
MIBP (urine)
1
2 Included in DIBP tables because in this population, at this time, MIBP concentrations were greater than
3 monobutyl phthalate (MBP) concentrations.
4
5 The literature for both epidemiological and animal studies will be regularly monitored for
6 the publication of new studies. The documentation and results for this supplementary search can
7 be found on the Health and Environmental Research On-line (HERO) website1
8 [http://hero.epa.gov/DIBP and http://hero.epa.gov/phthalates-humanstudies].
9 2.2. SELECTION OF CRITICAL STUDIES IN EARLY STAGES OF DRAFT
10 DEVELOPMENT
11 2.2.1. General Approach
12 Each study retained following the literature search and screen was evaluated for aspects of
13 design, conduct, or reporting that could affect the interpretation of results and the overall
14 contribution to the synthesis of evidence for determination of hazard potential. Much of the key
15 information for conducting this evaluation can generally be found in the study's methods section
16 and in how the study results are reported. Importantly, this evaluation does not consider study
17 results or, more specifically, the direction or magnitude of any reported effects. For example,
18 standard issues for evaluation of experimental animal data identified by the NRC and adopted in
19 this approach include consideration of the species and sex of animals studied, dosing information
20 (dose spacing, dose duration, and route of exposure), endpoints considered, and the relevance of
21 the endpoints to the human endpoints of concern. Similarly, observational epidemiologic studies in
22 this approach for evaluation should consider the following:
23 • Approach used to identify the study population and the potential for selection bias.
24 • Study population characteristics and the generalizability of findings to other populations.
iHERO is a database of scientific studies and other references used to develop EPA's risk assessments aimed
at understanding the health and environmental effects of pollutants and chemicals. It is developed and
managed in EPA's Office of Research and Development (ORD) by the National Center for Environmental
Assessment (NCEA). The database includes more than 1,400,000 scientific articles from the peer-reviewed
literature. New studies are added continuously to HERO.
Note: The HERO database will be regularly updated as additional references are identified during assessment
development. Therefore, the numbers of references (by tag) displayed on the HERO webpage for DIBP may
not match the numbers of references identified in Figure 2-1 (current through March 2014).
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1 • Approach used for exposure assessment and the potential for information bias, whether
2 differential (nonrandom) or nondifferential (random).
3 • Approach used for outcome identification and any potential bias.
4 • Appropriateness of analytic methods used.
5 • Potential for confounding to have influenced the findings.
6 • Precision of estimates of effect
7 • Availability of an exposure metric that is used to model the severity of adverse response
8 associated with a gradient of exposures.
9 To facilitate the evaluation outlined above, evidence tables are constructed that
10 systematically summarize the important information from each study in a standardized tabular
11 format as recommended by the NRG [2011]. In general, the evidence tables include all studies that
12 inform the overall synthesis of evidence for hazard potential. At this early stage of study
13 evaluation, the goal is to be inclusive. Exclusion of studies may unnecessarily narrow subsequent
14 analyses by eliminating information that might later prove useful. Premature exclusion might also
15 give a false sense of the consistency of results across the database of studies by unknowingly
16 reducing the diversity of study results. However, there may be situations in which the initial review
17 of the available data will lead to a decision to focus on a particular set of health effects and to
18 exclude others from further evaluation.
19 2.2.2. Exclusion of Studies
20 After the literature search was manually screened for pertinence, studies were excluded if
21 fundamental flaws were identified in their design, conduct, or reporting. The DIBP experimental
22 animal database consists of studies designed to examine repeat-dose intraperitoneal or oral toxicity
23 (including subchronic and short-term duration studies) and endpoint-specific toxicities (including
24 reproductive and developmental toxicity). Four studies administered DIBP via the intraperitoneal
25 route of exposure. These studies were excluded from the DIBP evidence tables because the
26 intraperitoneal route of exposure is generally considered less relevant to human health exposure.
27 The remaining studies involved administration of DIBP in the diet or via gavage administration.
28 Acute studies are generally less pertinent for characterizing health hazards associated with chronic
29 exposure. There was one acute study that was excluded from the evidence tables. Two BASF
30 reports identified in the literature searches could not be obtained and thus, could not be evaluated
31 for inclusion in the evidence tables (BASF, 2003,1961). For these reasons, these studies are not
32 summarized in the preliminary evidence tables. Nevertheless, with the exception of the studies that
33 could not be obtained, the studies will still be evaluated as possible sources of supporting health
34 effects information during assessment development. Experimental animal studies that were
35 sources of short-term, subchronic, or chronic health effects were evaluated for potential flaws in
36 their design, reporting, or conduct. As a result, one study, Maetal. (2013b) (English translation
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1 cited as Maetal. [2013a]], was removed from consideration in the assessment because of
2 incomplete description of experimental methods that leads to uncertainty in the results. Another
3 study, Eastman Kodak [1978], a one-page data summary, was excluded because it does not provide
4 detailed data reporting.
5 The remaining studies are all sources of health effects data that may be used in the
6 assessment The 20 studies summarized in the evidence tables are considered the "critical"
7 studies from which the study methods and results are presented in preliminary evidence tables
8 and exposure-response arrays (Section 3). There were also a few cases of the same study data
9 being contained in multiple reports; in those cases, the studies are listed together in the evidence
10 tables.
11 2.3. STUDY CHARACTERISTICS THAT WILL BE CONSIDERED IN THE
12 FUTURE EVALUATION AND SYNTHESIS OF THE CRITICAL
13 EPIDEMIOLOGICAL STUDIES FOR DIBP
14 Several considerations will be used in EPA's evaluation of epidemiological studies of human
15 health effects of DIBP. These considerations include aspects of the study design affecting the
16 internal or external validity of the results (e.g., population characteristics and representativeness,
17 exposure and outcome measures, confounding, data analysis), focusing on specific types of bias
18 (e.g., selection bias; information bias due to exposure misclassification) and other considerations
19 that could otherwise influence or limit the interpretation of the data. A study is externally valid if
20 the study results for the study population can be extrapolated to external target populations. An
21 internally valid study is free from different types of biases, and is a prerequisite for generalizing
22 study results beyond the study population. These issues are outlined in the IRIS Preamble, and are
23 described below.
24 Study Population
25 Evaluation of study population characteristics (including key socio-demographic variables
26 and study inclusion criteria) can be used to evaluate external validity (i.e., generalizability) and to
27 facilitate comparison of results across different study populations. Some aspects of the selection
28 process may also affect the interval validity of a study, resulting in a biased effect estimate.
29 The general considerations for evaluating issues relating to the study population include
30 adequate documentation of participant recruitment, including eligibility criteria and participation
31 rates, missing data, and loss to follow-up. This information is used to evaluate internal study
32 validity related to selection bias. Different types of selection bias that may occur include the
33 healthy worker effect, differential loss to follow up, Berkson's bias (relating to selection of
34 participants in hospital-based case-control studies), and participation bias. It is important to note
35 that low participation rates, or differences in participation rates between exposed and non-exposed
36 groups or between cases and controls, is not evidence of selection bias. Rather, selection bias arises
37 from a differential pattern of participation with respect to both the exposure and the outcome, i.e.,
38 patterns of participation that would result in a biased effect estimate. An example of differential
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1 participation would be when people with high levels of exposure and the outcome of interest are
2 more likely to participate than people with low levels of exposure and the outcome.
3 The available DIBP studies have generally examined metabolites from many different
4 phthalates within the context of research on environmental exposures. Most of these studies rely
5 on objective exposure measures (e.g., biomonitoring data), some of which are collected prior to
6 onset of the outcomes being examined (e.g., in the prospective pregnancy cohort studies). Study
7 participants generally do not have knowledge of the study hypothesis or their exposure to DIBP and
8 thus, knowledge of exposure or exposure level is unlikely to result in differential participation with
9 respect to outcomes. These study features should minimize the potential for selection bias.
10 However, EPA will consider the possibility that a particular concern about the specific sources of
11 DIBP, in conjunction with knowledge of specific health outcomes, may motivate people to
12 participate in a study or to continue participation throughout a follow-up period. In the absence of
13 evidence that any of these scenarios is likely to occur in a study, EPA will not consider selection bias
14 as a limitation of a study.
15 Exposure Considerations
16 General considerations for evaluating exposure include: (1) identifying how exposure can
17 occur (e.g., exposure sources, routes, and media); (2) determining appropriate critical exposure
18 period(s) for the outcomes under study; (3) evaluating variability in the exposure metrics of
19 interest (e.g., temporal and spatial variability for environmental measures or inter-individual
20 variability for biomonitoring data) that can impact different types of exposure metrics (e.g.,
21 cumulative, average, or peak exposure); (4) determining if an appropriate analytical methodology
22 was employed (e.g., choice of biological matrix, sampling protocol, quantification approach);
23 (5) evaluating the choice of exposure surrogate evaluated (e.g., constituent chemical or
24 group/mixture); and (6) evaluating the classification of individuals into exposure categories. These
25 six considerations help determine the accuracy and precision of the exposure estimates, and the
26 likelihood of measurement error with respect to the exposure metrics used. Nondifferential
27 misclassification of exposure categories, for example, can also result from measurement error and
28 is expected to predominantly result in attenuated effect estimates (Blair etal., 2007).
29 Some common sources of exposure to DIBP include cosmetics, food, and food packaging
30 (Zota etal., 2014) with the primary route of exposure occurring through ingestion and some
31 exposure occurring via inhalation and dermal routes (see Section 1.1.3). Thus, exposure to DIBP is
32 typically from multiple sources, and occurs episodically on a daily basis. Exposure to DIBP may be
33 increasing; a recent study of the U.S. general population found that urinary concentrations of the
34 DIBP metabolite MIBP have increased over time and were 206% higher in 2009-2010 compared to
35 2001-2002 (Zota etal.. 2014).
36 Urine provides an integrated measure of phthalate exposure from all sources.
37 Measurement of DIBP metabolites, rather than the parent compound, is preferred because the
38 parent compound is metabolized very quickly and does not provide an accurate measure of
39 exposure. The simple monoester metabolite, monoisobutyl phthalate (MIBP) is the most commonly
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1 measured DIBP metabolite in epidemiologic studies. MIBP accounts for an estimated 70.3% of the
2 urinary excretion of DIBP; this value is based on human data from a controlled dosing study of a
3 single volunteer [Kochetal., 2012]. EPA considers the use of MIBP to be a good proxy for total
4 DIBP exposure.
5 Although urine measures are most commonly used in epidemiological studies of phthalate
6 exposure, measures in serum, semen, and breast milk have also been used. Studies examining DIBP
7 metabolites in breast milk or serum have generally reported low levels of detection. One study in
8 Taiwan reported that MIBP above the limit of detection was found in 33.3% of breast milk samples
9 from 30 women. The detection rate in 30 cord blood samples in this study was 100%, but the
10 correlation between MIBP measured in cord blood and maternal urine was -0.11 (Pearson
11 correlation of log-transformed levels] [Linetal., 2011]. Hogberg et al. [2008] reported that few
12 breast milk (2 out of 42] or serum (3 out of 36] samples in a study in Sweden had detectable MIBP
13 concentrations. Another study conducted among 60 men ages 18-26 years found that 33.3% of
14 serum samples and 16.9% of seminal plasma samples had MIBP concentrations above the limit of
15 detection [Frederiksenetal., 2010]. The Spearman correlation coefficient between urine and
16 serum concentrations was 0.39; the correlation between urine and seminal plasma concentrations
17 was not calculated because of the low detection rate for the latter samples [Frederiksenetal.,
18 2010]. The lower detection rate in tissues other than urine reduces EPA's confidence in DIBP
19 metabolite measures in these biological matrices.
20 Given their first-order kinetics with half-lives on the order of hours [3.9 hours for MIBP in
21 [Koch and Angerer. 2007]]. urinary phthalate metabolite concentrations peak shortly after
22 exposure. Thus, for single-time exposure scenarios (rather than multi-source, multiple time
23 exposure scenarios], urine sampled during this time of peak concentration could lead to
24 overestimates of average daily intake, and conversely, measurements made after concentrations
25 have peaked and declined could lead to underestimates of intake. One study conducted among
26 139 pregnant women in Puerto Rico included measurement of MIBP found that specific gravity
27 adjusted concentrations were lower in samples collected from 9 am to noon (geometric mean 9.4]
28 compared with samples collected in early morning, early afternoon, or evening (geometric means
29 13-14] (Cantonwine etal., 2014]. Urinary measures of DIBP metabolite concentrations in
30 epidemiological studies are generally conducted using spot urine samples (i.e., collected at time of a
31 clinic or study examination visit] rather than at a specified time (e.g., first morning void] or in 24-
32 hour urine samples. Although the time of sample collection described above may affect the
33 accuracy of an estimated intake for a single individual, studies of other phthalates (e.g., DEHP] have
34 demonstrated that on a group level, spot urine samples provide a reasonable approximation of
35 concentrations that would have been observed using full-day urine samples (Christensenetal..
36 2014] and that a single spot sample was reliable in ranking subjects according to tertile of MIBP
37 (Teitelbaum etal., 2008]. Based on this information, EPA does not consider the reliance on spot
38 urine samples for exposure estimation (including ranking of individuals into different DIBP
39 categories] to be a major limitation for epidemiological studies. However because of the potential
40 for greater inaccuracy of estimates in the "tails" of the distribution, EPA will include additional
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1 considerations (e.g., discussion of analysis of residuals, outliers) when evaluating analyses based on
2 use of DIBP metabolites as continuous measures.
3 Another potential limitation of measurement of DIBP metabolites in urine is the
4 reproducibility of phthalate metabolite concentrations over time; that is, how well does a single
5 measure reflect the key exposure metric (average, peak) for the critical exposure window of
6 interest For many short-lived chemicals, considerable temporal variability in exposure level is
7 expected, and thus, repeated measures in the critical exposure window are preferred over a single
8 measurement Reproducibility is usually evaluated with the intraclass correlation coefficient (ICC),
9 a measure of the 'between-individual' variance divided by the total variance (between and within
10 individuals). A higher ICC indicates greater reproducibility (i.e., lower within-person variance). An
11 ICC of 0.51 for MIBP was reported in a study of 25 Hmong women ages 19-51 years with samples
12 collected 2-4 weeks apart (Pecketal.. 2010). In studies of reproducibility of measures during
13 pregnancy, Cantonwine etal. (2014) reported ICCs of 0.35 and 0.34 (unadjusted and specific gravity
14 adjusted) when comparing urine samples taken at approximately 18, 22, and 26 weeks of gestation.
15 ICCs of 0.36 and 0.38, respectively, were seen before pregnancy and in early pregnancy (Braun et
16 al., 2012), and an ICC of approximately 0.5 was seen over a 6-week period in the last trimester
17 (Adibi etal.. 2008). Among women participating in the Nurses' Health Study (NHS) (in 2000-2001
18 for NHS and in 1996-1999 for NHS II), the ICC for samples collected 1-3 years apart was 0.30 for all
19 samples, and was 0.29 for first-morning samples (Townsendetal.. 2013). Data for children are
20 sparse, limiting the ability to examine this source of uncertainty in this population. One study
21 evaluated variability in children aged 6-10 years old over a 6-month period (Teitelbaum etal..
22 2008) and found a relatively low ICC (0.21 unadjusted, 0.28 creatinine-adjusted). The available
23 data highlight the value of repeated exposure measures collected during the appropriate critical
24 period for the outcome(s) under study. Based on these studies, however, EPA does not consider the
25 use of a single measurement to be a major limitation in studies in adults in which the measure of
26 exposure is closely aligned with the relevant window(s) of exposure, if known, for the effect under
27 study. EPA has greater uncertainty, however, about measurements taken outside of the relevant
28 time window (e.g., several years after diagnosis, or the difference between first and third trimesters
29 of pregnancy), and about measurements taken in children.
30 Some studies present analyses using a combined measure based on summation of MIBP and
31 monobutyl phthalate (MBP), as a measure of both DIBP and DBF, respectively. The relative
32 contribution of DIBP to this total has varied over time (as the use of DIBP has increased), and can
33 vary between populations (e.g., greater use of DIBP compared with DBF in some countries). EPA
34 includes studies in the DIBP evidence tables using this summed exposure measure in situations in
35 which the concentration of MIBP is greater than that of MBP, but recognizes that this measure
36 introduces an additional source of exposure misclassification. Other studies present analyses using
37 a combined "low molecular weight" phthalate measure based on the summation of MIBP, MBP, and
38 monoethyl phthalate (MEP) (reflecting exposure to the parent compounds of DIBP, DBF, and DEP,
39 respectively). Because MIBP does not represent a major contributor to this summation
40 measurement, EPA has not included data from these studies in the DIBP evidence tables.
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1 EPA will also consider the potential for differential misclassification of biomarker measures
2 of exposure; for example, in situations in which a health outcome (e.g., diagnosis with diabetes or
3 cancer) could lead to a behavioral change that results in a change in DIBP exposure. This type of
4 scenario adds an additional challenge to the interpretation of the DIBP metabolites as valid
5 measures of exposure in a relevant time window(s) with respect to disease development
6 The distribution of exposure will also be considered in evaluating individual studies and
7 when comparing results among groups of studies. One consideration is the contrast of exposure
8 levels (i.e., the difference between "high" and "low"): a study with a very narrow contrast may not
9 have sufficient variability to detect an effect that would be seen over a broader range. Another
10 consideration is the absolute level of exposure, as different effect estimates may be expected in
11 studies examining different exposure levels even if they had similar exposure contrasts.
12 Prim ary Outcom e Measures
13 The general considerations for evaluating issues relating to accuracy, reliability, and
14 biological relevance of outcomes include adequate length of follow-up to evaluate the outcomes of
15 interest, and use of appropriate ascertainment methods to classify individuals with regard to the
16 outcome (e.g., high sensitivity and specificity). With respect to continuous measures, such as
17 hormone concentrations or semen parameters, EPA will consider, in addition to assessing whether
18 reported parameters are outside normal physiological range, evidence of smaller changes in the
19 distribution of a parameter that may represent an effect on a population level [e.g., as is the case for
20 early childhood exposure to lead and decrements in intelligence as measured by IQ (U.S. EPA.
21 2013).
22 Issues relating to assessment of the specific primary health effects are discussed below and
23 summarized in Table 2-8 at the end of Section 2.3.
24 Sexual differentiation
25 Cryptorchidism and hypospadias are two disorders of the development of the male
26 reproductive system. Cryptorchidism, or undescended testes, can be present at birth (congenital
27 Cryptorchidism) or can occur later during infancy and childhood (acquired Cryptorchidism).
28 Surgical correction (orchiopexy) is recommended in cases of Cryptorchidism that do not resolve
29 during infancy because long-term complications include impaired sperm production and increased
30 risk of testicular cancer (Virtanen et al., 2007). Retractile testes can move back and forth between
31 the scrotum and the abdomen; this condition usually resolves by puberty and is not associated with
32 reproductive or other complications. Classification criteria for Cryptorchidism that involve
33 testicular positioning are commonly used in clinical research (Tohn Radcliffe Hospital
34 Cryptorchidism Study Group, 1988: Scorer, 1964). EPA will consider the definition used and age
35 range in interpreting studies of Cryptorchidism or related outcomes.
36 In animal toxicology studies, anogenital distance (AGD) is a routine marker to assess
37 endocrine disruption; this marker has only recently been adapted for use in epidemiological
38 studies. One study in adult men reported associations between decreased AGD and measures
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1 relating to infertility [Eisenbergetal., 2011]: most studies have used this measure in infants,
2 however, as a marker of endocrine environment during development. It is important to consider
3 general size, in addition to sex, in the evaluation of AGD, for example by incorporating birth weight
4 or length (e.g., calculation of "anogenital index" by dividing anogenital distance by weight). With
5 regard to reproducibility of this measure, a low degree of between-observer variability was found
6 using a standardized protocol and trained observers [Romano-Riqueraetal.. 2007: Salazar-
7 Martinez etal.. 2004). Because of the importance of size and age in the interpretation of this
8 measure, EPA has greater confidence in studies with measures taken at birth or over a narrow age
9 range and lesser confidence in studies among a group spanning a larger age range.
10 Gender-related behaviors, as measured by the Pre-School Activities Inventory [Golombok
11 and Rust, 1993] or other scales, has been examined in relation to direct or indirect measures of
12 fetal testosterone levels, including studies of DIBP. This outcome measure has been examined in
13 studies of relatively rare genetic conditions (e.g., congenital adrenal hyperplasia and complete
14 androgen insensitivity syndrome], as well as in studies focusing on the normal variability seen in
15 the general population (reviewed in Hines, 2006]. EPA will consider evidence pertaining to the
16 reliability and validity of the Pre-School Activities Inventory in its evaluation of studies using this
17 scale.
18 Male and female reproductive outcomes
19 The DIBP literature includes studies of reproductive and gonadotropin hormone levels in
20 men and studies of semen parameters that can be indicative of reduced fertility. The details of the
21 laboratory procedures, including information on the basic methods, level of detection, and
22 coefficient of variation, are important considerations for hormone assays and measures of semen
23 parameters. The World Health Organization (WHO] laboratory methods for analysis of sperm
24 counts and semen parameters (see, for example. WHO. 1999] are generally recognized as standards
25 in this field. EPA will consider studies that reference these methods, regardless of which revision
26 used, to be reliable measures.
27 Much of the focus of the research on male steroidal and gonadotropin hormones in the DIBP
28 database concerns testosterone. One issue with respect to these measures is the estimation method
29 used for free testosterone. Based on the analysis by Vermeulen et al. (1999], EPA will consider
30 estimates based on total testosterone divided by immunoassay-derived sex-hormone binding
31 globulin (SHBG] levels to be most reliable.
32 The DIBP literature also includes studies of reproductive hormones in women. In addition
33 to the general considerations regarding hormone assays noted above, timing within a menstrual
34 cycle for studies of pre- and peri-menopausal women, and timing with respect to gestational age for
35 studies of women during pregnancy, are also be an important considerations for interpretation of
36 reproductive hormone concentrations.
37 Another female reproductive outcome included in the DIBP literature is endometriosis.
38 Endometriosis can be symptomless, or can lead to surgical intervention; it is often diagnosed as
39 part of a work-up for infertility. Variability in clinical presentation and in access and use of health
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1 care services present considerable challenges to conducting epidemiological studies of this
2 condition [Holt and Weiss, 2000]. Confirmation of "case" and "control" status (i.e., presence or
3 absence of endometriosis) by ultrasound or clinical evaluation is recommended to reduce outcome
4 misclassification, and representation of the source population should be carefully considered.
5 Infertility is generally defined clinically and for research purposes as the inability to
6 conceive a clinically-recognized pregnancy after 12 months of intercourse of regular frequency
7 without use of contraceptives. Fecundity or fecundability are terms for the capacity for
8 reproduction. "Time to pregnancy" (i.e., the number of cycles of unprotected intercourse before
9 conception) has been used as a measure of fecundability in studies of environmental and
10 occupational exposures (Bairdetal., 1986: Baird and Wilcox, 1985]. Time to pregnancy is a
11 measure of a couple's fecundability, incorporating effects that can be manifested through the male
12 or female (or both]. Considerations in time to pregnancy studies include the source of data (i.e.,
13 retrospective or prospective designs] and incorporation of information on "non-pregnancy
14 planners" (Weinberg et al.. 1994].
15 Timing of male and female puberty, and conditions of unusual pubertal development
16 Pubertal development in humans is often assessed using timing of peak height velocity
17 ("growth spurt"] and secondary markers of sexual development Secondary markers for females
18 include breast development (thelarche] and pubic hair development (pubarche], and age at first
19 period (menarche]. Secondary markers for males include gonadal development (gonadarche] and
20 pubic hair development, and age at first sperm emission (spermarche].
21 Evaluation of breast, pubic hair, and gonadal development is frequently performed using
22 the Tanner stages (Marshall and Tanner, 1970,1969], which places the individual in one of five
23 stages, ranging from pre-pubertal (stage 1] to adult maturation (stage 5). However, the process of
24 this staging is not straightforward, and is most reliable when performed by trained personnel
25 (rather than by the individual or a parent, for example] (Slough etal.. 2013: Schlossberger et al..
26 1992: Espeland etal.. 1990]. Age at menarche is considered to more reliable when assessed via
27 self-report (Koprowskietal.. 2001], although reliability may decrease with increasing time since
28 menarche (Cooper et al., 2006]. Additionally, hormone levels may sometimes be used to evaluate
29 pubertal development Individuals may vary widely in the timing of these developmental
30 milestones.
31 Several clinical syndromes are known to disrupt the timing and order of markers of
32 pubertal development Considerations in the diagnosis of either precocious or delayed puberty
33 include the diagnostic criteria used and the source of the information (e.g., whether collected from
34 medical records or from self- or parental report]. For females, precocious puberty is usually
35 defined as the onset of puberty before the age of 8 years, while delayed puberty is usually defined
36 as the lack of pubertal development by the age of 13 years (Marshall and Tanner, 1969]:
37 corresponding ages in males are before the age of 9 years for precocious puberty and lack of
38 pubertal development by the age of 14 years for delayed puberty (Marshall and Tanner. 1970].
39 Clinical evaluation would involve hormone assays to distinguish between gonadotropin dependent
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1 ("central"), gonadotropin independent ("peripheral"), or a combination of both (Traggiai and
2 Stanhope, 2003) forms of these conditions.
3 Pregnancy-related outcomes
4 Infant birth weight and gestational age are two outcomes commonly used in reproductive
5 epidemiology studies. EPA considers analyses of the various indices for both outcomes (fetal
6 growth and gestational age) to be informative with respect to hazard identification, but will
7 consider each separately as they address different issues. Gestational duration can be measured as
8 a continuous outcome or dichotomous outcome such as preterm birth. Preterm births include
9 infants delivered earlier than 37 gestational weeks, and those delivered earlier than 32 gestational
10 weeks are classified as very preterm births. Different measures of fetal growth restriction are often
11 examined in epidemiological studies. In addition to the continuous measure of birth weight,
12 another commonly used measure of fetal growth restriction is the categorical variable of low birth
13 weight (defined as <2,500 g). Small for gestational age (defined as birth weight less than the 10th
14 percentile for the gestational birth weight distribution) is considered a better measure of fetal
15 growth rate as it takes into consideration gestational duration, and would be preferred over a
16 measure of birth weight in a study that includes preterm births. Birth weight and gestational
17 duration can also be examined as continuous variables, often in analysis that excludes preterm or
18 low birth weight births, so that the focus of the analysis is on variability within the "normal" range.
19 EPA considers birth weight obtained from medical records to be a reliable source as this is a
20 very accurate and precise measurement Although more prone to measurement error than birth
21 weight measures, gestational age can be estimated from several approaches. Some of these include
22 ultrasonography, estimates based on date of last menstrual period based on maternal recall, or
23 from clinical examination based on antenatal or newborn assessments (which may include an
24 ultrasound). Menstrual dating of gestational age dependent on maternal recall of the last menstrual
25 period can be subject to considerable measurement error in some cases, so ultrasonography-based
26 estimates may be considered more accurate (Savitzetal.. 2002: Taipale and Hiilesmaa. 2001).
27 Immune-related outcomes: allergy and asthma
28 Skin prick testing is a standard method for assessing atopy (allergic disease) used in some
29 epidemiologic studies. Other studies use an assessment protocol based on reported history of
30 symptoms (e.g., rhinitis, hay fever) or specific types of allergies. These can be considered
31 complementary types of measures: skin prick tests provide information on a defined set of
32 potential antigens to which a person may be exposed, and symptom-based evaluations provide
33 information on experiences of individuals and the variety of exposures they encounter. Studies
34 comparing questionnaire responses with skin prick tests in children have reported relatively high
35 specificity (89-96%) and positive predictive value (69-77%) for self-reported history of pollen or
36 pet dander allergy or for answers to a combination of questions incorporating itchy eyes with nasal
37 congestion in the absence of a cold or flu (Braun-Fahrlander etal.. 1997: Dotterudetal.. 1995). The
38 validity was somewhat lower for a more restricted set of questions (nasal congestion in the absence
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1 of a cold or flu; specificity 83%, positive predictive value 52%) [Braun-Fahrlander etal., 1997].
2 Based on these data, EPA considers allergy history based only on rhinitis symptoms to have a
3 greater likelihood of outcome misclassification compared with those based on a combination of
4 symptoms.
5 Epidemiologic studies of asthma typically use a questionnaire-based approach to define
6 asthma based on symptoms relating to wheezing episodes or shortness of breath, reported history
7 of asthma attacks, or use of asthma medication, usually for a period defined as "current" or in the
8 past year. Much of this work is based upon the American Thoracic Society questionnaire [Ferris,
9 1978] or subsequent instruments that built upon this work, including the International Society of
10 Arthritis and Allergies in Children Questionnaire and the European Community Respiratory Health
11 Survey. These questionnaire-based approaches have been found to have an adequate level of
12 specificity and positive predictive value for use in etiologic research [Ravault and Kauffmann. 2001:
13 Pekkanen and Pearce. 1999: Burneyetal.. 1989: Burney and Chinn. 1987]. EPA considers
14 outcomes defined over a recent time period (e.g., symptoms in the past 12 months] to be more
15 relevant within the context of concurrent exposure measurements compared with outcomes
16 defined over a lifetime (e.g., ever had asthma].
17 Neurodevelopment
18 With respect to neurodevelopmental outcomes, a major consideration is the assessment
19 tool(s] used by the study investigators; details of the assessment method, or references providing
20 this information, should be provided. In addition, EPA also looks for discussion of (or reference to]
21 validation studies and the appropriateness of the tool for evaluation in the specific study population
22 (e.g., age range, language].
23 Thyroid
24 Thyroid-related endpoints examined in epidemiological studies of DIBP include thyroid
25 hormones (triiodothyronine, T3, and thyroxine, T4] and thyroid stimulating hormone (TSH] (or
26 thyrotropin] produced by the pituitary.
27 As with other hormone assays, the details of the laboratory procedures, including
28 information on the basic methods, limit of detection, and coefficient of variation, are important
29 considerations for the hormone assays. Thyroid hormones are generally measured in serum,
30 although they may also be measured in dried blood spots, such as are collected from newborn
31 infants in screening for congenital hypothyroidism. Studies in older age groups have also shown a
32 very high correlation (r = 0.99] between thyroid hormone levels measured in dried blood spots and
33 levels in serum (Hofman et al.. 2003].
34 With respect to thyroid hormones, time of day and season of sampling are two main
35 potential sources of variability. For example, serum TSH measured shortly after midnight may be
36 as much as twice as high as the value measured in late afternoon (Brabant etal., 1991: Weeke and
37 Gundersen. 1978]. The evidence with respect to seasonal variability is mixed (Plasqui etal.. 2003:
38 Nicolau etal.. 1992: Simonietal.. 1990: Behalletal.. 1984: Postmes etal.. 1974] and this effect is
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1 likely to be smaller than that of time of day. The impact of these sources of variation will depend on
2 whether they are also related to DIBP (i.e., whether DIBP levels vary diurnally or seasonally). If this
3 is the case, failure to address these factors in the design or analysis could result in confounding of
4 the observed association, with the direction of this bias determined by the direction of the
5 association between these factors and DIBP. If this is not the case, the lack of consideration of time
6 of day or seasonality would result in greater variability in the hormone measures, and would thus
7 result in more imprecise (but not biased) estimates was located. EPA has not found studies
8 examining seasonal variation in DIBP levels. With respect to variability relating to time of day, as
9 noted previously, one study of 139 pregnant women in Puerto Rico reported lower concentrations
10 of specific gravity-adjusted MIBP in samples collected from 9 am to noon (geometric mean of 9.4)
11 compared with samples collected in early morning, early afternoon, or evening (geometric means of
12 13-14) (Cantonwine etal.. 2014). Based on these data, EPA has greater confidence in thyroid
13 hormone studies that consider time of sample collection in the analysis, but recognizes the limited
14 nature of the available data pertaining to this issue.
15 Obesity
16 Most of the studies of obesity measures in the DIBP database are based on body mass index
17 (BMI, calculated as kg/m2) or waist circumference using measurements taken as part of the data
18 collection protocol. BMI is highly correlated with body fat, and standardized cut-points have been
19 established for characterization of "normal" (BMI between 18.5 and 24.9 kg/m2), "overweight"
20 (BMI between 25.0 and 29.9 kg/m2) and "obese" (BMI > 30.0 kg/m2) categories. Waist
21 circumference is also highly correlated with body fat, and is a more direct measure of abdominal
22 obesity. EPA notes that use of self-reported weight (e.g., report of pre-pregnancy weight) would
23 not be considered to be as reliable as actual measurements.
24 Diabetes and measure of insulin resistance
25 In the DIBP database, diabetes has been assessed by a variety of biomarkers of glucose and
26 insulin and by self-report of diabetes diagnosis. Oral glucose tolerance testing and glycosolated
27 hemoglobin (HbAlc) are used clinically and in epidemiological research (Selvinetal., 2011). Self-
28 report of prevalent diabetes can have high sensitivity and specificity in comparison to diagnosed
29 diabetes based on validated medical record data (Oksanenetal., 2010: Leikauf and Federman,
30 2009). The biomarker-based classifications, however, offer an added advantage of being able to
31 include undiagnosed disease. EPA will consider these points in assessing the reliability and validity
32 of the diabetes measures used in the studies. None of the currently available studies assessed
33 diabetes through cause of death data; sensitivity of diabetes assessed using cause of death data is
34 low, even if underlying and other contributing cause of death fields are included (Cheng etal.,
35 2008).
36 Insulin resistance, a marker of diabetes risk, can be measured using the homeostatic model
37 assessment (HOMA) method, a physiologically-based structural model, using fasting glucose and
38 insulin or C-peptide concentrations. HOMA is a validated tool for the estimation of insulin
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1 resistance in epidemiology studies, and requires a single measurement of fasting glucose and
2 insulin [Wallace etal., 2004]. Although the mean of three samples taken at 5-minute intervals
3 results in a more precise estimate, insulin resistance estimated using a single baseline
4 measurement is well correlated with that using the mean of three measurements when used to
5 estimate a group mean. Therefore, EPA does not consider the use of a single measurement as an
6 input to the HOMA model to be a limitation.
7 Cancer
8 With respect to studies of cancer, EPA considers the source of the outcome data (e.g., cause
9 of death data, hospital cancer registry data, hospital discharge data, histopathology reports) in its
10 evaluation of the accuracy of the data. An additional issue is the validity of mortality data as a
11 representation of cancer incidence; mortality data for cancer types with a high survival rate may
12 underrepresent disease incidence, require additional considerations with respect to determining
13 appropriate time windows of exposure, and may lead to biased risk estimates if survival is related
14 to exposure.
15 Confounding
16 The general considerations for evaluating issues relating to potential confounding include
17 consideration of which factors may be potential confounders (i.e., those which are strongly related
18 to both the exposure and the outcome under consideration, and are not intermediaries on a causal
19 pathway), adequate control for these potential confounders in the study design or analysis, and
20 where appropriate, quantification of the potential impact of mismeasured or unmeasured
21 confounders. Uncontrolled confounding by factors that are positively associated with both the
22 exposure (e.g., DIBP) and health endpoint of interest, and those that are inversely associated with
23 both exposure and health endpoint, will result in an upward bias of the effect estimate.
24 Confounding by factors that are positively associated with exposure and inversely associated with
25 the health endpoint (or vice versa) will result in a downward bias of the effect estimate.
26 Potential confounding by other phthalates
27 Few studies have reported results of analyses evaluating the correlation between MIBP and
28 metabolites of other phthalates. In an analysis conducted by EPA of 5,109 samples from the
29 2003-2008 National Health and Nutrition Examination Survey (NHANES) participants aged >6
30 years, the pairwise Spearman correlation coefficient between MIBP and MEP (the primary
31 metabolite of DEP) was low (0.33). A more moderate correlation was seen with the DEHP
32 metabolites (correlations of approximately 0.5); higher correlations were seen with MBzP (the
33 primary metabolite of BBP, correlation coefficient = 0.58) and MBP (the primary metabolite of DBF;
34 correlation = 0.72). Similar or some what lower correlations were seen between MIBP and other
35 phthalate metabolites in a small study (n = 45) of men seen in an infertility clinic (Wirthetal.,
36 2008], in 319 pregnancy women (Whyattetal., 2012], and in 600 reproductive age women in a
37 study of endometriosis (Buck Louis etal.. 2013). EPA will evaluate the potential for confounding by
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1 examining the similarity of the results seen with different metabolites. Thus, for example, lack of
2 adjustment for mono-benzyl phthalate (MBzP) would not be considered a limitation in a study in
3 which an association was seen with MIBP that was not seen with MBzP; however this lack of
4 adjustment would be considered a limitation if an association of similar or higher magnitude was
5 seen for both of metabolites.
6 Potential confounding by demographic factors
7 Age, race/ethnicity, and sex are considered important explanatory factors for most types of
8 outcomes measured in epidemiological research. In NHANES 2009-2010 data, urinary MIBP levels
9 decreased with age (geometric means of 13.2, 8.63, and 7.45 |ig/g-creatinine, respectively, in ages
10 6-11,12-19 and >20 years) [CDC, 2013]. Concentrations were lower levels in males compared
11 with females (geometric means of 6.99 and 9.05 |ig/g-creatinine, respectively, in males and
12 females), and variability by ethnicity was also observed, with lower levels in non-Hispanic whites
13 (geometric mean of 7.12 |ig/g-creatinine) compared with non-Hispanic blacks and Mexican
14 Americans (geometric means of 10.1 and 9.27 [ig/g-creatinine, respectively). EPA will consider
15 these differences in assessing the potential influence of demographic factors on observed effect
16 estimates for DIBP.
17 Potential confounding by other factors
18 Some of the health effects under consideration may have strong associations with other risk
19 factors. For example, smoking is associated with increased risk of low birth weight and preterm
20 births, and with infertility. Abstinence time is strongly related to sperm concentration measures.
21 In evaluating the potential for confounding by any of these factors, EPA will review evidence
22 pertaining to the strength and direction of its association with DIBP (or its metabolites).
23 Data An alysis
24 The general considerations for evaluating issues relating to data analysis include adequate
25 documentation of statistical assumptions and analytic approach (including addressing skewness of
26 exposure or outcome variable and shape of exposure-response), consideration of sample size and
27 statistical power, and use of appropriate statistical methods for the study design.
28 One other issue, specific to much of the DIBP literature, concerns the optimal approach to
29 addressing urinary volume or dilution in the analysis of spot urine or first morning void samples.
30 Options include use of creatinine- or specific gravity-adjusted metabolite concentrations, or use of
31 unadjusted concentrations. Although use of some kind of correction factor has been advocated for
32 studies of obesity (Goodman etal.. 2014). a simulation study reported that creatinine-adjusted
33 exposure measures may produce biased effect estimates for outcomes that are strongly related to
34 factors affecting creatinine levels, of which obesity is a prime example (Christensenetal., 2014).
35 EPA recognizes the lack of consensus at this time, as well as the need for continued research into
36 the potential bias introduced by different analytic approaches. Based on current understanding of
37 this issue, EPA prefers results using unadjusted concentration for outcomes strongly related to
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1 creatinine levels; for other outcomes, EPA does not have a basis for preferring one type of analysis
2 over another.
3
4
Table 2-8. General and outcome-specific considerations for DIBP study
evaluation
General considerations
Study population
Exposure
Analysis
• Study population and setting: geographic area, site, time period, age and sex
distribution, other details as needed (may include race/ethnicity,
socioeconomic status)
• Recruitment process; exclusion and inclusion criteria, knowledge of study
hypothesis; knowledge of exposure and outcome
• Participation rates: total eligible; participation at each stage and for final
analysis group and denominators used to make these calculations
• Length of follow-up, loss to follow-up
• Comparability: participant characteristic data by group, data on non-
participants
• Biological matrix or target tissue/organ (e.g., urine, serum, semen, breast
milk)
• Level of detection (LOD) or level of quantitation (LOQ)
• Exposure distribution (e.g., central tendency, interquartile range), proportion
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Steroidal and
gonadotropin
hormones (adults; sex-
specific)
Measures
Consideration of
confounding
Relevant exposure
time window(s)
• Type of assay
• Sensitivity/detection limits, coefficient of variation; number of samples
below LOD
• Age, day or phase of menstrual cycle (if cycling)
• Up to 6 months preceding hormone sample collection
Sperm parameters
Measures
Consideration of
confounding
Relevant exposure
time window(s)
Type of assay (e.g., WHO protocol)
• Age, smoking, BMI, abstinence time (consider if these are related to
exposure)
• Up to 6 months preceding semen sample collection
Infertility
Measures
Consideration of
confounding
Relevant exposure
timewindow(s)
• Definition, source of data
• Age, smoking, alcohol use, heavy metal exposure, radiation time (consider if
these are related to exposure)
• Time preceding and during attempt to become pregnant
Timing of puberty
Measures
Consideration of
confounding
Relevant exposure
timewindow(s)
• Source of data (e.g., self-report, physician assessment)
• Age, sex, ethnicity, body size, nutritional status (consider if these are related
to exposure)
• In utero? Up to 12 months preceding transition from one stage to another
stage?
Gestational age
Measures
Consideration of
confounding
Relevant exposure
timewindow(s)
• Source of data and estimation procedure (ultrasound; last menstrual period
or clinical assessment)
• Smoking, pregnancy complications, assisted reproduction technologies
(consider if these are related to exposure)
• In utero
Birth weight
Measures
Consideration of
confounding
Relevant exposure
timewindow(s)
• Source of data (e.g., medical records, birth certificate)
• Gestational age, maternal age, ethnicity, nutritional intake, smoking,
maternal height/BMI, (consider if these are related to exposure)
• In utero
Immune - allergy and
asthma
Measures
Consideration of
confounding
• Number of allergens used in skin prick testing or allergen-specific IgE assay;
sensitivity/specificity of specific questions used in history assessment
• Age, family history (consider if these are related to exposure)
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Relevant exposure
time window(s)
• For current conditions (e.g., asthma in past 12 months): up to 12 months
preceding outcome assessment
Ne urob ehavioral
Measures
Consideration of
confounding
Relevant exposure
time window(s)
• Standardized assessment tool, validation studies for specific study
population (e.g., age group, geographic location)
• Blinding of assessor to exposure
• Age, sex, socioeconomic status
• In utero; early childhood
Thyroid
Measures
Consideration of
confounding
Relevant exposure
time window(s)
• Assay used and evidence from validation studies, if available
• Sensitivity/detection limits, coefficient of variation; number of samples
below LOD
• Time of day and season when samples for thyroid hormone (and TSH)
collected
• Age, sex, smoking, iodine, radiation exposure (consider if these are related
to exposure)
• Varies by lifestage (i.e., infants, children, adults)
Obesity
Measures
Consideration of
confounding
Relevant exposure
time window(s)
• Source of data (e.g., measured or self-reported weight and height)
• Age, sex, ethnicity, caloric intake, physical activity (consider if these are
related to exposure)
• Not established (likely to be more than one, including in utero)
Diabetes and insulin
resistance
Measures
Consideration of
confounding
Relevant exposure
time window(s)
• Source of data (e.g., biomarkers of insulin or glucose, medical records, self-
report)
• Age, sex, ethnicity
• Not established (likely to be more than one, including in utero)
2 2.4. STUDY CHARACTERISTICS THAT WILL BE CONSIDERED IN THE
3 FUTURE EVALUATION AND SYNTHESIS OF THE CRITICAL
4 EXPERIMENTAL STUDIES FOR DIBP
5 Beyond the initial methodological screening described above in Section 2.2.2,
6 methodological aspects of a study's design, conduct, and reporting will be considered again in the
7 overall evaluation and synthesis of the pertinent data that will be developed for each health effect.
8 Some general questions that will be considered in evaluating experimental animal studies are
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1 presented in Table 2-9. These questions are, for the most part, broadly applicable to all
2 experimental studies.
3
4
Table 2-9. Questions and relevant experimental information for the
evaluation of experimental animal studies
Methodological
feature
Test animal
Experimental setup
Exposure
Endpoint evaluation procedures
Outcomes, data, and reporting
Question(s) considered
Based on the endpoint(s) in question, are concerns raised regarding the
suitability of the species, strain, or sex of the test animals on study?
Are the timing, frequency and duration of exposure, as well as animal age
and experimental group allocation procedures/group size for each endpoint
evaluation, appropriate for the assessed endpoint(s)?
Are the exposure conditions and controls informative and reliable for the
endpoint(s) in question, and are they sufficiently specific to the compound
of interest?
Do the procedures used to evaluate the endpoint(s) in question conform to
established protocols, or are they biologically sound? Are they sensitive for
examination of the outcome(s) of interest?
Were data reported for all pre-specified endpoint(s) and study groups, or
were any data excluded from presentation/analyses?
5
6
7
8
9
10
11
12
13
14
15
16
17
18
Note: "Outcome" refers to findings from an evaluation (e.g., steatosis), whereas "endpoint" refers to the
evaluation itself (e.g., liver histopathology).
Evaluation of some specific methodological features identified in Table 2-9 such as
exposure, is likely to be relatively independent of outcome. Other methodological features, in
particular those related to experimental setup and endpoint evaluation procedures, are generally
outcome specific (i.e., reproductive and developmental toxicity). In general, experimental animal
studies will be compared against traditional assay formats (e.g., those used in guideline studies),
with deviations from the protocol evaluated in light of how the deviations could alter interpretation
of the outcome in question. A full evaluation of all critical studies will be performed as part of the
critical review and synthesis of evidence for hazard identification for each of the health endpoints
identified in the evidence tables presented in Section 3.
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1
2 3. PRELIMINARY EVIDENCE TABLES AND
3 EXPOSURE-RESPONSE ARRAYS
4 3.1. DATA EXTRACTION FOR EPIDEMIOLOGICAL AND EXPERIMENTAL
5 STUDIES: PREPARATION OF PRELIMINARY EVIDENCE TABLES
6 The evidence tables present data from studies related to a specific outcome or endpoint of
7 toxicity. At a minimum, the evidence tables include the relevant information for comparing key
8 study characteristics such as study design, exposure metrics, and dose-response information.
9 Evidence tables will serve as an additional method for presenting and evaluating the suitability of
10 the data to inform hazard identification for DIBP during the analysis of hazard potential and utility
11 of the data for dose-response evaluation. For each critical study selected, key information on the
12 study design, including characteristics that inform study quality, and study results pertinent to
13 evaluating the health effects from subchronic and chronic oral exposure to DIBP are summarized in
14 preliminary evidence tables.
15 Epidemiological studies are presented first where each study per table is listed in reverse
16 chronological order. Animal studies are then presented where each study per health endpoint is
17 presented in alphabetical order by study author, followed by species and strain. Most results are
18 presented as the percent change from the control group; an asterisk (*) indicates a result that has
19 been calculated and reported by study authors to be statistically significant compared to controls
20 (p < 0.05). Unless otherwise noted in a footnote, doses presented in the animal evidence tables
21 were those reported by the study authors.
22 The information in the preliminary evidence tables is also displayed graphically in
23 preliminary exposure-response arrays. In these arrays, a significant effect (indicated by a filled
24 circle) is based on statistical significance by the study authors. The complete list of references
25 considered in preparation of these materials can be found on the Health and Environmental
26 Research Online (HERO) website at (https://hero.epa.gov/DIBP and
27 http://hero.epa.gov/phthalates-humanstudies).
28
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l 3.2. EPIDEMIOLOGICAL STUDIES
2 3.2.1. Sexual Differentiation Measures
3 Table 3-1. Evidence pertaining to DIBP and sexual differentiation effects in
4 humans
Reference and study design
Results
Anogenital distance (AGD)
Swan (2008) (United States; Minnesota, Missouri,
California)
Population: 106 boys from birth cohort study (Study
for Future Families), 2000-2002, mean age 12.8 mo
(0-36 mo)
Outcome: AGD (to posterior genitalia) measured at
0-36 mo (mean 70.4 mm, 7.1 mm/kg)
Exposure: Maternal urine sample, 3rd trimester
MIBP in urine (ng/mL):
Median 75th percentile
Unadjusted 2.5 5.1
Analysis: Regression analysis using mixed model
adjusting for age and weight percentile
Related references: Swan et al. (2005) (exposure
data and analysis of smaller sample size with less
robust method of adjustment for variation by size)
Percent change in AGD per interquartile increase in MIBP
concentration (p-value)
MIBP
-3.5 (0.097)
Cryptorchidism or testicular position
Swan (2008) (United States; Minnesota, Missouri,
California)
Population: 106 boys from birth cohort study (Study
for Future Families), 2000-2002, mean age 12.8 mo
(0-36 mo)
Outcome: Incomplete testicular descent assessed at
clinical exam (10% prevalence)
Exposure: Maternal urine sample, 3rd trimester
MIBP in urine (ng/mL):
Median 75th percentile
Unadjusted 2.5 5.1
Analysis: Logistic regression, adjusting for age and
weight percentile
Related references: Swan et al. (2005) (exposure
data)
MIBP reported as not associated with testicular position
(quantitative results not reported)
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Reference and study design
Results
Gender-related play
Swan et al. (2010) (United States; Minnesota,
Missouri, California, Iowa)
Population: 145 children from birth cohort study
(Study for Future Families), 2000-2002 and
2002-2005 (Iowa), ages 4-7 yrs; second follow-up
Outcome: Gender-specific play based on Pre-School
Activities Inventory (24 items completed by parent or
caregiver; subscores of male-oriented items and
female-oriented items and a composite score
consisting of male summation minus the female
summation scores)
Exposure: Maternal urine sample, 3rd trimester
Unadjusted MIBP in urine (ng/mL):
Median 75th percentile
Boys 2.4 5.1
Girls 2.8 5.0
Analysis: Regression analysis using Generalized
Linear Models, considering creatinine, sex and age of
child, maternal age, parental education, number of
same and opposite sex siblings, ethnicity, clinic
location, and parental attitude as potential covariates
Related references: Swan et al. (2005) (exposure
data)
Regression coefficient (95% Cl) for pre-school activities
index scores and log-transformed MIBP (adjusted for
child's age, mother's age, mother's education, parents'
attitude toward boy's play, and interaction between
education and attitude; negative value indicates less
masculine play behavior with higher metabolite level)
Boys Girls
Masculine -1.65 (-4.57, 1.28) 1.04 (-1.75, 3.82)
Composite -4.53 (-8.12,-0.94) 0.38 (-3.86, 4.63)
1
2
Cl = confidence interval; MIBP = monoisobutyl phthalate
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3.2.2. Male Reproductive Effects in Humans
Table 3-2. Evidence pertaining to DIBP and semen parameters or infertility in
adult men or couples
Reference3 and study design
Results
Kranvogl et al. (2014) (Slovenia)
Population: 136 men from couples seeking
infertility treatment (mean age 36.2 yrs, range
24-54 yrs), 2012
Outcome: Semen analysis
Exposure: Urine sample, collected at same time
as semen sample
MIBP in urine:
Median Maximum
Unadjusted (u.g/L) 21.6 161.8
Cr-adjusted (u.g/g Cr) 20.8 119.2
Analysis: Spearman correlation
Spearman correlation coefficient, MIBP and sperm
parameters:
Sperm concentration
Sperm motility
(p > 0.05 for both parameters)
-0.044
-0.075
Joensen et al. (2012) (Denmark)
Population: 881 men from general population,
assessed at military conscript exam*, 2007-2009,
median age 19.1 yrs (5th-95th percentile:
18.4-22.0 yrs)
Outcome: Semen analysis
Exposure: Urine sample, collected at same time
as semen sample
MIBP in urine (ng/mL):
Median 95th percentile
Unadjusted 58 173
Analysis: Linear regression, considering age,
BMI, smoking, alcohol consumption, ethnicity,
BMI squared, in utero exposure to tobacco
smoke, previous or current diseases, recent
fever, recent use of medication, abstinence time,
and time from ejaculation to analysis as potential
covariates
*As reported by Ravnborg et al. (2011)
Results for individual phthalate metabolites (including MIBP)
reported as "few significant associations" with sperm volume,
count, or percentage progressively motile sperm (quantitative
results not reported). Sperm concentration analysis adjusted
for abstinence time (volume, concentration, and count);
sperm motility analysis adjusted for time from ejaculation to
analysis (progressively motile); analysis of percent of
morphologically normal sperm was unadjusted
Wirth et al. (2008) (United States, Michigan)
Population: 45 male partners seen in infertility
clinic, time period not reported; mean age 34 yrs
Outcome: Semen analysis
Exposure: Urine sample, collected at same time
as semen sample (all between 7 and 11 am)
MIBP in urine (ng/mL) (percentile):
Median 75th percentile 95th percentile
5.8 10.0 17.9
Analysis: Dichotomized outcomes (above and
below WHO reference values), MIBP
dichotomized at median; age, education (three
levels), income (three levels), race, BMI (three
levels), current smoking status, and alcohol use
The combined measure for MIBP and MBP was not associated
with any sperm parameter, nor was MIBP when analyzed
individually (data not shown)
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference3 and study design
Results
(two levels) considered as potential confounders;
specific gravity also included in all models
Infertility
Buck Louis et al. (2014) (United States; Michigan
and Texas)
Population: 501 couples discontinuing
contraception and attempting to achieve
pregnancy; recruited from 16 counties using
population sampling; women's mean age
30.0 yrs, men's mean age 31.8 yrs; 2005-2009
Outcome: Time to pregnancy as assessed by
diaries recording intercourse and menstruation,
home-fertility monitoring to detect ovulation,
and home pregnancy tests
Exposure: Urine samples from both partners,
collected at enrollment (beginning of pregnancy
attempt)
Unadjusted MIBP in urine (ng/mL) among
couples achieving pregnancy:
Geometric mean (95% Cl)
Women 5.11(4.58-5.70)
Men 3.44 (3.09-3.83)
Analysis: Fecundability OR calculated using Cox
models, adjusting for variables shown in results
column
Fecundability OR (95% Cl) for increase in log-transformed
MIBP scaled by standard deviation (adjusted for female age,
difference in couple's ages, research site, and both partners'
urinary creatinine, BMI, and serum cotinine; in addition,
results for exposure in each partner adjusted for exposure in
the other partner, and models accounted for left truncation or
time off contraception)
Women
Men
0.97 (0.80,1.18)
0.91 (0.76, 1.09)
1
2
3
BMI = body mass index; MBP = monobutyl phthalate; OR = odds ratio; WHO = World Health Organization
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
Table 3-3. Evidence pertaining to DIBP and reproductive hormones in adult
men
Reference and study design
Results
Joensen et al. (2012) (Denmark)
Population: 881 men from general population, assessed at
military conscript exam*, 2007-2009, median age 19.1 yrs
(5th-95th percentile: 18.4-22.0 yrs)
Outcome: Serum steroidal and gonadotropin hormones
Exposure: Urine sample, collected at same time as serum
sample for hormone analysis
MIBP in urine (ng/mL):
Median 95th percentile
Unadjusted 58 173
Analysis: Linear regression considering age, BMI, smoking,
alcohol consumption, time of blood sampling, assay type,
ethnicity, BMI squared, in utero exposure to tobacco smoke,
previous or current diseases, recent fever, and recent use of
medication as potential covariates
*As reported by Ravnborg et al. (2011)
Results for individual phthalate metabolites
(including MIBP) reported as "few significant
associations" with free testosterone, estradiol,
SHBG, LH, inhibin-B, or FSH (quantitative results
not reported); analyses adjusted for age, BMI,
smoking, alcohol consumption, and time of blood
sampling (and assay type for inhibin-B only)
Mendiola et al. (2011) (United States; Minnesota, Missouri,
California, Iowa, New York)
Population: 425 men whose partners enrolled in birth
cohort study (Study for Future Families), 1999-2005, mean
age 32 yrs
Outcome: Serum steroidal and gonadotropin hormones
Exposure: Urine sample, collected at same time as serum
sample for hormone analysis
MIBP in urine (ng/mL) (distribution not reported)
Analysis: Pearson correlation of log(10)-transformed MIBP
and hormone measures; linear regression considering age,
age square, BMI, smoking status, ethnicity, urinary
creatinine concentration, time of sample collection, time of
collection squared, season, educational level, center, and
stressful life events)
Authors reported "little or no association with
metabolites of phthalate other than DEHP"
[including MIBP] with testosterone, estradiol,
SHBG, LH, inhibin-B, or FSH (quantitative results
not reported)
3
4
5
6
DEHP = diethylhexyl phthalate; FSH = follicle-stimulating hormone; LH = luteinizing hormone; MOINP = oxo-(mono-
oxoisononyl) phthalate; SD = standard deviation; SHBG = sex hormone binding globulin
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.2.3. Male Pubertal Development in Humans
Table 3-4. Evidence pertaining to DIBP and the timing of male puberty or sex
hormones in boys
Reference and study design
Results
Ferguson et al. (2014c) (Mexico)
Population: 115 boys ages 8-14 yrs from a birth
cohort (Early Life Exposure in Mexico to
Environmental Toxicants, participants enrolled
during first trimester 1994-2004), follow-up
initiated in 2010
Outcome: Adrenarche or puberty, based on Tanner
staging by physician (pubic hair stage >2; genitalia
stage >2 or testicular volume >3 mL); serum
hormone level
Exposure: Maternal urine sample (n = 107) from
third trimester or child's urine sample (n = 113)
collected at time of Tanner staging and serum
collection
Unadjusted MIBP in urine (ng/mL):
Median 95th percentile
Maternal sample 1.83 6.64
Child's sample 9.61 36.1
Analysis: Logistic regression for analysis of puberty
onset, adjusting for variables shown in results
column; linear regression for analysis of hormone
levels, considering age, BMI z-score, socioeconomic
status, and maternal smoking as potential
covariates
OR (95% Cl) for adrenarche or puberty per interquartile
increase in In-transformed MIBP (adjusted for child age,
BMI z-score, and urine specific gravity)
Exposure basis
Tanner stage or
testicular
volume
Maternal urine
(prenatal)
Pubic hair
(stage >2)
0.29 (0.07, 1.30)
Genitalia (stage 0.71 (0.37,1.35)
Testicular 1.60 (0.70, 3.65)
volume (>3 mL)
Child urine
0.76 (0.32,1.81)
0.76 (0.39, 1.49)
2.17(0.81,5.82)
Percent change (95% Cl) in serum hormone level per
interquartile increase in In-transformed MIBP (adjusted for
urine specific gravity, child age, and BMI z-score)
Exposure basis
Serum Maternal urine
hormone (prenatal) Child urine
Testosterone 5.12 (-23.3, 44.0) -26.2 (-45.6, 0.16)
Free 1.69 (-26.7, 41.1) -27.9 (-47.8,-0.60)
testosterone
SHBG 5.72 (-5.18, 17.9) 2.20 (-8.41, 14.1)
DHEAS -2.02 (-15.9, 14.1) 3.02 (-11.4, 19.8)
Estradiol -1.94 (-11.2, 8.23) -12.3 (-20.2, -3.54)
Inhibin B -1.98 (-12.7,10.1) 2.73 (-8.24,15.0)
Mouritsen et al. (2013b) (Denmark)
Population: Boys from population-based cohort
(COPENHAGEN Puberty Study), 2006-2010; age
11 yrs (53 boys) or 13 yrs (31 boys)
Outcome: Adrenarche or puberty, based on Tanner
staging by physician (pubarche = pubic hair stage >2
and testicular volume >3 mL); serum hormone level
Exposure: Urine sample, first morning sample; data
reported in Mouritsen et al. (2013a, Supplemental
Material)
Median age (yrs) at development by ZMIBP + MBP level
(evaluation at 11 yrs)
Pubarche
Testicular volume >3 mL
Low
12.3
11.5
High
11.0 (p< 0.05)
11.1
Median hormone concentration by MIBP + MBP level
(evaluation at 11 yrs)
Testosterone (nmol/L)
DHEAS (nmol/L)
Low
<0.23
2.02
High
<0.23
1.61
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
MIBP + MBP in urine (ng/mL)a:
Geometric mean Maximum
118 676
(based on larger sample of 84 boys)
Analysis: Two-tailed Mann-Whitney U-test for
comparisons between groups, comparing median
hormone levels and pubertal stage in "high" and
"low" phthalate groups (based on above or below
group mean excretion)
Mieritz et al. (2012) (Denmark)
Population: 38 boys with pubertal gynecomastia
and 190 age-matched controls drawn from 555 boys
from population-based cohort (COPENHAGEN
Puberty Study), 2006-2008; ages 6-19 yrs
Outcome: Anthropometry, pubertal stage (pubic
hair and genital development), presence of
gynecomastia, and serum testosterone
Exposure: Urine sample, first morning sample
MIBP in urine (ng/mL):
Median 95th percentile
Groups 74.88 229.1
(boys without gynecomastia, all ages)
Analysis: Two-tailed Mann-Whitney U-test for
comparisons between groups; linear regression with
age adjustment for association with serum
testosterone; probit analysis with phthalate
concentrations divided in quartiles for analysis of
puberty timing
Results
Adione (nmol/L) 1.28 1.22
Estradiol (pmol/L) <18 <18
FSH (IU/L) 1.28 1.68
LH (IU/L) 0.28 0.27
Median age (yrs) at development by MIBP + MBP level
(evaluation at 13 yrs)
Low High
Pubarche 12.5 12.1
Testicular volume >3 mL 11.6 11.6
Median age (yrs) at development by MIBP + MBP level
(evaluation at 13 yrs)
Low High
Testosterone (nmol/L) 5.1 7.7
DHEAS (u.mol/L) 2.61 3.64
Adione (nmol/L) 2.96 3.85
Estradiol (pmol/L) 19 37
FSH (IU/L) 2.4 2.5
LH (IU/L) 1.8 1.4
MIBP concentration (ng/mL) by group
Group 1 Group 2 Group 3
(n = 38) (n = 189) (n = 517)
Median 68.50 73.96 74.88
95th percentile 178.8 199.5 229.1
Group 1 = boys with palpable gynecomastia
Group 2 = boys without palpable gynecomastia (age-
matched)
Group 3 = boys without palpable gynecomastia (all ages)
No association between MIBP concentration and timing of
puberty or serum testosterone level (quantitative results
not reported)
1
2
3
4
5
6
7
aln this population at this time, MIBP tended to be present at higher concentrations than MBP; EPA includes these
studies in the DIBP tables, but recognizes the exposure misclassification introduced by the use of the summed
concentration exposure measure.
DHEAS = dehydroepiandrosterone; EPA = Environmental Protection Agency
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2
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.2.4. Female Pubertal Development in Humans
Table 3-5. Evidence pertaining to DIBP and timing of female puberty or sex
hormones in girls
Reference and study design
Results
Precocious puberty and premature thelarche
Frederiksen et al. (2012) (Denmark)
Population: 24 girls with precocious puberty (n = 13
with central precocious puberty, n = 6 with early
normal puberty, n = 5 with premature thelarche)
from outpatient clinic, 2008-2009 and 184* age-
matched controls from population-based cohort
(COPENHAGEN Puberty Study), recruited from high
schools 2006-2008; age 7.4-9.9 yrs
Outcome: Precocious puberty, early normal
puberty, or premature thelarche based on Tanner
staging by physician
Exposure: Urine sample (child's), first morning
sample collected at clinical evaluation
MIBP and MBP in urine (ng/mL)a, controls (analysis
based on sum of these two metabolites):
Median 95th percentile
MIBP 81 241
MBP 51 153
(based on larger sample of 725 controls)
Analysis: Urine concentrations in cases and controls
compared with Mann-Whitney U test
*Study reports number of controls inconsistently;
text reports 164 controls, while Table 4 reports 184
Median (range) ZMIBP and MBP metabolites in urine
(ng/mL) in cases and controls
Controls
147 (22-2,195)
Precocious
puberty
94 (32-383)
(p-value)
(p<0.01)
Lomenick et al. (2010) (United States, Ohio and
Kentucky)
Population: 28 girls with central precocious
puberty, 28 age- and race-matched controls; all
recruited from pediatric endocrinology clinic,
2005-2008; mean age 7 yrs
Outcome: Central precocious puberty defined
based on clinical standards (appearance of physical
characteristics of puberty before 8 yrs of age, with
laboratory confirmation of central origin of breast
development); no cases had received medical
treatment prior to urine sample collection
Exposure: Urine sample (child's), collected at
clinical evaluation
MIBP in urine of controls:
MeaniSE
Unadjusted (ng/mL) 22.6 ± 7.6
Cr-adjusted (ng/g Cr) 20.2 ± 4.9
Analysis: MIBP concentrations in cases and controls
compared with Wilcoxon rank-sum test
Unadjusted
(ng/mL)
Cr-adjusted
Cr)
Central
precocious
Controls puberty (p-value)
22.6 ±7.6 15.4 ±2.9 (0.77)
20.2 ±4.9 16.5 ±2.1 (0.96)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Pubertal development (general population)
Hart et al. (2013) (Australia)
Population: 121 girls from birth cohort study
(Western Australian Pregnancy Cohort), whose
mothers were recruited at 18 wks of gestation,
1989-1991; follow-up at ages 14-16 yrs
Outcome: Age at menarche
Exposure: Maternal serum samples (n = 123)
collected at 18 and 34-36 wks of gestation
(combined aliquot from both time periods)
MIBP in serum (ng/mL):
Median 90th percentile
Unadjusted 1.77 6.16
Analysis: Correlation between log-transformed
MIBP and age at menarche
Authors reported no association between MIBP and age at
menarche (quantitative results not reported)
Authors reported no correlation between MIBP and serum
SHBG, FSH, total testosterone, free androgen index, anti-
Miillerian hormone, or inhibin B in adolescents
(quantitative results not reported by study authors)
Mouritsen et al. (2013b) (Denmark)
Population: Girls from population-based cohort
(COPENHAGEN Puberty Study), 2006-2010; age
10 yrs (47 girls) and 13 yrs (33 girls)
Outcome: Adrenarche or puberty, based on Tanner
staging by physician (pubarche = breast stage >2 and
pubic hair stage >2); serum hormone level
Exposure: Urine sample, first morning sample; data
reported in Mouritsen et al. (2013a, Supplemental
Material)
MIBP + MBP in urine (ng/mL)a:
Geometric mean Maximum
122 904
(based on larger sample of 84 girls)
Analysis: Two-tailed Mann-Whitney U-test for
comparisons between groups, comparing median
hormone levels and pubertal stage in "high" and
"low" phthalate groups (based on above or below
group mean excretion)
Median age (yrs) at development by MIBP + MBP level
(evaluation at 10 yrs)
Pubarche (pubic hair stage
Pubarche (breast stage >2)
Low
10.7
10.6
High
11.2
10.3
Median hormone concentration by MIBP + MBP level
(evaluation at 10 yrs)
Testosterone (nmol/L)
DHEAS (u.mol/L)
Adione (nmol/L)
Estradiol (pmol/L)
FSH (IU/L)
LH (IU/L)
Low
<0.23
1.1
2.03
20
1.86
0.06
High
<0.23
0.83
1.29
22
2.25
0.1
Median age (yrs) at development by MIBP + MBP level
(evaluation at 13 yrs)
Pubarche (pubic hair stage
Pubarche (breast stage >2)
Low
10.7
10.7
High
11.2
10.5
Median hormone concentration by MIBP + MBP level
(evaluation at 13 yrs)
Testosterone (nmol/L)
DHEAS (u.mol/L)
Adione (nmol/L)
Low
1.1
2.23
6.40
High
0.5 (p< 0.05)
1.27 (p< 0.05)
3.91
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
3
4
5
Reference and study design
Frederiksen et al. (2012) (Denmark)
Population: 725 healthy girls ages 5.6-19.1 yrs from
COPENHAGEN Puberty Study cohort, recruited from
high schools during 2006-2008
Outcome: Stage of breast or pubic hair
development based on Tanner staging by physician;
Serum steroid and gonadotropin hormones
Exposure: Urine sample (child's), collected at time
of pubertal stage assessment
Unadjusted MIBP and MBP in urine (ng/mL)a, all
725 participants:
Median 95th percentile
MIBP 81 241
MBP 51 153
Analysis: Probit analysis, results verified using Pool-
Adjacent-Violators algorithm
Results
Estradiol (pmol/L) 194 131
FSH(IU/L) 4.9 5.8 (p< 0.05)
LH (IU/L) 3.8 3.8
Mean age (95% Cl) (yrs) at entry into breast stage 2 or
pubic hair stage 2, by quartile of JMIBP + MBP
metabolites:
IMIBP +
MBP Breast stage 2 Pubic hair stage
quartile (n = 394) (n not reported)
l(low) 10.12(9.61,10.62) 10.83(10.54,11.12)
2 9.97 (9.48, 10.46) 10.97 (10.67, 11.28)
3 9.89 (9.40, 10.37) 11.22 (10.93, 11.52)
4 (high) 9.79 (9.30, 10.30) 11.54*(11.21, 11.88)
*Significantly different from quartile 1; p < 0.05
Levels of FSH, LH, estradiol, and testosterone were similar
across JMIBP + MBP metabolite exposure groups when
adjusted for age distribution (quantitative results not
reported)
aln this population at this time, MIBP tended to be present at higher concentrations than MBP; EPA includes these
studies in the DIBP tables, but recognizes the exposure misclassification introduced by the use of the summed
concentration exposure measure.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.2.5. Female Reproductive Effects in Humans
Table 3-6. Evidence pertaining to DIBP and reproductive hormones in adult
women
Reference and study design
Results
Maternal hormones during pregnancy
Sathvanaravana et al. (2014) (United States; Minnesota,
Missouri, California)
Population: 180 mothers from birth cohort (Study for
Future Families), recruited during pregnancy, 1999-2002
Outcome: Serum hormone levels, samples collected
during prenatal clinic visit
Exposure: Maternal urine sample, collected during 2nd or
3rd trimester
MIBP in urine (ng/mL):
Median 75th percentile
Unadjusted 2.7 4.85
Analysis: Linear regression, log-transformed MIBP and log-
transformed hormone level
Regression coefficient (95% Cl) for change in
maternal log-transformed serum hormone level
with unit increase in log-transformed MIBP,
stratified by sex of fetus
Testosterone
(total)
Testosterone
(free)
Estradiol
Mothers with
male fetus
(n = 94)
-0.03
(-0.18,0.13)
-0.03
(-0.20,0.14)
0.003
(-0.12,0.12)
Mothers with
female fetus
(n = 86)
-0.10
(-0.28, 0.07)
-0.11
(-0.30, 0.08)
0.03
(-0.14,0.20)
Hart et al. (2013) (Australia)
Population: 123 mothers from birth cohort (Western
Australian Pregnancy Cohort), whose mothers were
recruited at 18 wks of gestation between 1989 and 1991
Outcome: Reproductive and gonadotropin hormone levels
in maternal serum collected at 18 and 34-36 wks of
gestation
Exposure: Maternal serum samples (n = 123) collected at
18 and 34-36 wks of gestation (combined aliquot from
both time periods)
MIBP in serum (ng/mL):
Median 90th percentile
MIBP 1.77 6.16
Analysis: Correlation between quartiles of serum MIBP
and log-transformed hormone levels
Correlation coefficient between log-transformed
maternal serum hormone level and quartiles of
MIBP in maternal serum
Androstene-
dione (nmol/L)
DHEAS (u.mol/L)
Testosterone
(pmol/L)
SHBG (nmol/L)
Free
testosterone
(pmol/L)
Free
testosterone
index
At 18 wks of
gestation
(n = 119)
0.023
-0.042
0.003
0.108
-0.061
-0.051
At 34-36 wks of
gestation
(n = 114)
-0.060
-0.084
-0.101
-0.020
-0.063
-0.064
p > 0.10 for all correlations
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
Table 3-7. Evidence pertaining to DIBP and gynecological conditions in
humans
Reference and study design
Results
Endometriosis
Buck Louis et al. (2013) (United States, California and Utah)
Population: 473 women undergoing laparoscopy or
laparotomy and 127 population age- and residence-matched
referents, 2007-2009; ages 18-44 yrs; confirmed cases of
endometriosis matched to women without endometriosis
within each cohort: operative cohort 190 cases,
238 controls; population cohort 14 cases, 127 controls
Outcome: Endometriosis confirmed by surgery (operative
cohort) or MRI (population cohort)
Exposure: Urine sample
MIBP in urine (ng/mL), unadjusted:
Geometric mean
Operative cohort-controls 6.82
Population cohort-controls 7.59
Analysis: Student's t-test or Wilcoxon test for continuous
data; logistic regression, adjusting for age, BMI, and
creatinine; sensitivity analyses conducted restricting cohort
to endometriosis stages 3 and 4 diagnoses or visually and
histologically confirmed endometriosis, and referent group
consisting of women with postoperative diagnosis of normal
pelvis
OR (95% Cl) for endometriosis per unit increase in
In-MIBP, by cohort (adjusted for age, BMI, and
creatinine)
Operative cohort
Population cohort
1.02 (0.80,1.29)
2.22 (0.98, 5.04)
Adjusted OR (95% Cl) for endometriosis per unit
increase in In-MIBP in operative cohort (sensitivity
analysis)
Endometriosis stage 3 and 4
(n = 339)
Visual/histological confirmed
endometriosis (n = 473)
Comparison with women with
postoperative diagnosis
normal pelvis (n = 320)
0.96 (0.67, 1.38)
1.08 (0.77, 1.51)
1.09 (0.82, 1.46)
Note: Concentrations were log transformed and
rescaled by their SDs for analysis
Upson et al. (2013) (United States, Washington)
Population: 92 incident endometriosis cases, 195 controls
frequency-matched on age, all members of a large health
care system and enrolled in Women's Risk of Endometriosis
Study, 1996-2001; ages 18-49 yrs
Outcome: Endometriosis confirmed by surgery; for each
case, reference date assigned by date of first visit for
symptoms leading to diagnosis; reference dates randomly
assigned to controls based on case distribution
Exposure: Urine sample, collected after enrollment
(2001-2002)
MIBP in urine, controls:
Median 75th percentile
Unadjusted (ng/mL) 1.5 3.1
Analysis: Logistic regression (quartiles of exposure),
covariates considered based on directed acyclic graph; final
model adjusted for variables shown in results column
OR (95% Cl) for endometriosis by quartile MIBP
(adjusted for In-transformed urinary creatinine,
age, and reference yr)
MIBP quartile (ng/mL)
1 (<0.7)
2 (0.7-1.5)
3(1.5-3.1)
4(>3.1)
(trend p-value)
OR (95% Cl)
1.0 (referent)
0.9 (0.4, 2.0)
0.8 (0.3, 2.2)
0.8 (0.3, 2.6)
(0.84)
Adjustment for education, smoking status and
alcohol consumption did not alter the results;
similar results in analyses based on summation of
MIBP and MBP
Polycystic ovarian syndrome
Hart et al. (2013) (Australia)
Population: 121 girls from birth cohort study (Western
Australian Pregnancy Cohort), whose mothers were
recruited at 18 wks of gestation between 1989 and 1991;
follow-up at ages 14-16 yrs
Correlation coefficient (p-value) between log-
transformed MIBP and pubertal development
parameter
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Outcome: Uterine volume, ovarian volume, and antral
follicle count measured by ultrasound; PCO defined as
>1 ovary more than 10 cm3 or >12 follicles between 2 and
9 mm in diameter; PCOS defined either as (1) presence of at
least two of: polycystic ovarian morphology, clinical or
biochemical hyperandrogenism, or oligo-anovulation; or
(2) oligo-anovulatory menstrual cycles with either clinical or
biochemical hyperandrogenism; all clinical assessments
conducted on d 2-5 of menstrual cycle
Exposure: Maternal serum samples (n = 123) collected at
18 and 34-36 wks of gestation (combined aliquot from both
time periods)
MIBP in serum (ng/mL):
Median 90th percentile
MIBP 1.77 6.16
Analysis: Correlation between log-transformed MIBP and
uterine volume, ovarian volume, and antral follicle counts;
MIBP concentrations in PCO or PCOS cases and controls
compared calculated using t-tests or Mann-Whitney U-tests
Uterine volume (mL)
Ovarian volume (cm3)
Antral follicle count
r < 0.20 (p> 0.17)
r < 0.10 (p> 0.29)
r < 0.12 (p> 0.20)
Authors reported no association between MIBP
and polycystic ovarian syndrome using either
definition (quantitative results not reported).
1
2
3
PCO = polycystic ovarian morphology; PCOS = polycystic ovarian syndrome
This document is a draft for review purposes only and does not constitute Agency policy,
3-14 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 3.2.6. Pregnancy Outcomes in Humans
2 Table 3-8. Evidence pertaining to DIBP and pregnancy outcomes in humans
Reference and study design
Results
Fetal growth (birth weight, birth length, head circumference)
Huang etal. (2014b) (China)
Population: 207 women delivering at one hospital
in Chongqing between 2011 and 2012, aged
18-35 yrs and with no history of tobacco or alcohol
use; mean age 28 yrs
Outcome: Standard clinical measures at birth
Exposure: Cord blood sample
DIBP in cord blood (ug/L)
Median 75th percentile 95th percentile
All samples 16.7 26.9 114
Analysis: Linear regression, adjusting for variables
shown in results column
Regression coefficient (95% Cl) for change in clinical
measurement at birth per unit increase in In-transformed
DIBP (ug/L) (adjusted for gestational age):
Birth weight (g)
Birth length
(cm)
Head
circumference
(mm)
Girls
-27 (-90, 36)
-0.06 (-0.45, 0.33)
Boys
-87 (-195, 200)
-0.75 (-1.35,
-0.15)
-3.85 (-9.47, 1.76) -2.76 (-7.62, 2.11)
Philippat et al. (2012) (France)
Population: 72 cases with undescended testis or
hypospadias, 215 matched controls from two birth
cohorts (EDEN and PELAGIE), 2002-2006
Outcome: Standard clinical measurements at birth
Exposure: Maternal urine sample, collected
between 6 and 19 (PELAGIE) or between 24 and
30 (EDEN) wks of gestation
MIBP in urine (ng/mL):
Median 95th percentile
Measured 45.9 219.0
Standardized* 64.7 365.3
Analysis: Cases and controls combined for this
analysis; weighted linear regression using tertiles or
In-transformed urine concentrations, adjusting for
variables shown in results column; analysis by
tertiles for evaluation of possible non-monotonic
relationship; analyses corrected for oversampling of
malformation cases
*Standardized for sampling conditions and
gestational age at collection
Regression coefficient (95% Cl) for change in birth outcome
by MIBP tertile and per unit change in In-MIBP
(standardized, ng/mL) (adjusted for gestational duration,
maternal pre-pregnancy weight and height, maternal
smoking, maternal education, parity, recruitment center,
urine creatinine, and mode of delivery as potential
covariate; head circumference model also adjusted for
mode of delivery)
MIBP tertile
(Ug/L)
1 (<48.2)
2 (48.2-97.9)
3 (>97.9)
(trend p-value)
In (MIBP)
Birth
weight (g)
0
(referent)
61
(-77, 200)
-31
(-190, 129)
(0.48)
-44
(-110, 23)
Birth
length
(cm)
0
(referent)
0.4
(-0.3,1.1)
0.3
(-0.4, 1.0)
(0.54)
0.0
(-0.3, 0.3)
Head
circumference
(cm)
0
(referent)
-0.1
(-0.6, 0.4)
0.2
(-0.5, 0.9)
(0.40)
-0.1
(-0.4,0.1)
Wolff et al. (2008) (United States, New York City)
Population: 382 singleton live births without
medical complications from birth cohort (Mt. Sinai
Children's Environmental Health study), 1998-2002
Outcome: Standard clinical measurements at birth
Exposure: Maternal urine sample, third trimester
MIBP in urine (ng/mL):
Median 75th percentile
Unadjusted 6.2 12
Analysis: Linear regression, adjusting for variables
shown in results column
Regression coefficient (95% Cl) for change in birth outcome
with unit increase in In-MIBP (ng/mL) (adjusted for
race/ethnicity, infant sex, gestational age at delivery, In-
creatinine, prenatal smoking, pre-pregnancy BMI, maternal
education, and marital status)
Birth weight (g)
Birth length (cm)
Head circumference (cm)
-14 (-57, 28)
0.04 (-0.19, 0.28)
0.05 (-0.11, 0.21)
Restricted to observations with creatinine >20 mg/dL
This document is a draft for review purposes only and does not constitute Agency policy,
3-15 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Preterm birth (<37 wks) and gestational age
Ferguson et al. (2014b): Ferguson et al. (2014a)
(United States; Boston)
Population: 130 cases, 352 controls from
pregnancy cohort (study of predictors of pre-
eclampsia, enrolled during first trimester,
2006-2008); controls randomly selected from
among those delivering >37 wks of gestation; mean
age 33 yrs
Outcome: Preterm birth (<37 wks of gestation;
gestation estimated from first trimester ultrasound);
additional analysis of subgroup with spontaneous
preterm labor or preterm premature rupture of
membranes ("spontaneous preterm," n = 57)
Exposure: Maternal urine sample, one to three
samples collected at median times of 9.7,17.9, or
26.0 wks of gestation; geometric mean of results
from visits 1-3 used in analyses.
MIBP in urine, SG-adjusted (ng/L):
Geometric mean 75th percentile
Controls 6.71 10.3
All cases 6.85 10.5
Analysis: Logistic regression (In-transformed
metabolites), considering average specific gravity,
maternal age, race/ethnicity, education level, health
insurance provider, BMI at first study visit, smoking
status, alcohol use, parity, use of assisted-
reproductive technology, and sex of infant as
potential covariates
Related reference: Ferguson et al. (2014a) (analysis
by individual sample results for the four visits)
OR (95% Cl) for preterm birth per unit increase in In-
transformed MIBP (adjusted for average specific gravity,
maternal age, race/ethnicity, education level, and
insurance provider)
All preterm 0.98 (0.72,1.34)
Spontaneous preterm 1.52 (0.97, 2.38)
[Results weaker than those seen with DEHP metabolites]
Results by study visit from Ferguson et al. (2014a), all pre-
term births
Visit 1
Visit 2
Visit 3
Visit 4
0.92
0.88
0.75
0.66
(0.57, 1.47)
(0.54, 1.41)
(0.50, 1.13)
(0.28, 1.55)
Huang etal. (2014b) (China)
Population: 207 women delivering at one hospital in
Chongqing between 2011 and 2012; aged 18-35 yrs
and with no history of tobacco or alcohol use; mean
age 28 yrs
Outcome: Preterm birth (<37 wks of gestation;
gestational age estimated from last menstrual
period)
Exposure: Cord blood sample
DIBP in cord blood (ng/L)
Median 75th percentile 95th percentile
All samples 16.7 26.9 114
Analysis: Logistic and linear regression, adjusting
for variables shown in results column
OR (95% Cl) for preterm delivery comparing In-DIBP above
and below the median (adjusted for maternal age, BMI,
frequency of prenatal exam, and pregnancy history), with
additional stratification by history of intravenous infusions
Total sample (n = 207)
No intravenous infusions (n = 154)
Intravenous infusions (n = 53)
6.01 (3.24, 11.17)
4.78 (1.68, 13.57)
6.07 (2.66, 13.83)
[History of intravenous infusions present in 26% of total
and 55% of preterm birth group]
Regression coefficient (95% Cl) for change in gestational
age (wks) per unit increase in In-transformed DIBP (ng/L)
(adjusted for maternal age, BMI, frequency of prenatal
examination, history of intravenous infusions therapy, and
pregnancy history)
-0.75 (-1.03, -0.46)
This document is a draft for review purposes only and does not constitute Agency policy,
3-16 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Meeker et al. (2009) (Mexico)
Population: 30 cases, 30 controls (term births) from
pregnancy cohort, 2001-2003.
Outcome: Preterm birth (<37 wks of gestation),
determined using maternal recall of last menstrual
period
Exposure: Maternal urine sample, third trimester
MIBP in urine, among term births
Median 75th percentile
Unadjusted 2.0 4.1
SG-adjusted (jjg/L) 2.3 5.0
Cr-adjusted (ng/g Cr) 3.7 6.6
Analysis: Logistic regression, considering maternal
age, pre-pregnancy BMI, parity, education, marital
status, infant's sex, and gestational age at urine
sample as potential covariates
OR (95% Cl) for preterm birth by MIBP above compared
with below the median (adjusted for marital status,
maternal education, and infant sex and gestational age at
time of urine sample)
Cr-unadjusted (ng/L)
SG-adjusted (ng/L)
Cr-adjusted (ng/g Cr)
3.6(1.1,12.2)
2.0 (0.7, 6.0)
1.5 (0.5, 4.5)
Wolff et al. (2008) (United States, New York City)
Population: 382 singleton live births without
medical complications from birth cohort (Mt. Sinai
Children's Environmental Health study), 1998-2002
Outcome: Standard clinical measurements at birth
Exposure: Maternal urine sample, third trimester
MIBP in urine (ng/mL):
Median 75th percentile
Unadjusted 6.2 12
Analysis: Linear regression, adjusting for variables
shown in results column
Regression coefficient (95% Cl) for change in birth outcome
with unit increase in In-MIBP (ng/mL) (adjusted for
race/ethnicity, infant sex, gestational age at delivery, In-
creatinine, prenatal smoking, pre-pregnancy BMI, maternal
education, and marital status)
Gestational age (wks) 0.03 (-0.20, 0.14)
Restricted to observations with creatinine >20 mg/dL
1
2
This document is a draft for review purposes only and does not constitute Agency policy,
3-17 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 3.2.7. Immune Effects in Humans
2 Table 3-9. Evidence pertaining to DIBP and allergy/immune effects in humans
Reference and study design
Results
Ait Bamai et al. (2014)a (Japan)
Population: Children (n = 122, ages
<15 yrs) and adults (n = 374, ages >15 yrs)
living in 148 detached dwellings in which
at least 25 mg of dust was collected; 2006
follow-up of 2003 baseline survey
Outcome: Allergic condition assessed by
self-administered questionnaire (positive
response to: in the past 2 yrs have you
been seen at a hospital for allergic rhinitis,
allergic conjunctivitis, or atopic
dermatitis?); parents completed
questionnaires for children <6 yrs
Exposure: Dust samples
DIBP in dust (ng/g dust):
Median 75th
percentile
Floor dust (n = 148) 2.4 5.5
Multi-surface dust (n = 120) 1.9 3.5
Analysis: Generalized linear mixed effects
model, considering gender, age strata
(<15, >15 yrs), smoking status (personal
and environmental tobacco smoke), furry
pets in home, signs of dampness, Der 1
(not defined by authors), other phthalates
dust, airborne fungi, formaldehyde, total
VOC, and building characteristics as
potential covariates
OR (95% Cl) for allergic condition by tertile of DIBP in floor dust (ng/g
dust)(adjusted for adjusted for gender, age strata, smoking status,
dampness index, furry pets inside the home, Der 1, and sum of other
phthalates)
DIBP
tertile
1 (low)
2
3 (high)
(trend
p-value)
1 (low)
2
3 (high)
(trend
p-value)
1 (low)
2
3 (high)
Full sample Children Adults
Allergic rhinitis
1.0 (referent) 1.0 (referent) 1.0 (referent)
1.87 (0.83, 4.22) 3.54 (0.86, 14.5)
1.05 (0.47, 2.32) 2.30 (0.60, 8.89)
(0.91) (0.23)
0.99 (0.47,
2.05)
0.48 (0.22,
1.02)
(0.06)
Allergic conjunctivitis
1.0 (referent) 1.0 (referent) 1.0 (referent)
1.07 (0.38, 3.01) 1.97 (0.35, 11.1) 0.59 (0.19, 1.8)
1.64(0.64,4.18) 3.27(0.68,15.7) 0.82(0.31,2.2)
(0.30) (0.14) (0.69)
Atopic dermatitis
1.0 (referent) 1.0 (referent) 1.0 (referent)
5.52 (1.68, 18.1) 11.95 (1.37, 104) 2.55 (0.89,
7.31)
4.84 (1.46, 16.0) 15.0 (1.91,118)
(0.01) (0.01)
p-value for age interaction >0.05 for all endpoints
(trend
p-value)
1.56 (0.44,
5.53)
(0.49)
No increased aORs (either in the full sample or stratified by age)
were observed in analyses using DIBP measurements in multisurface
dust.
This document is a draft for review purposes only and does not constitute Agency policy,
3-18 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Callesen et al. (2014b): Callesen et al.
(2014a)a (Denmark)
Population: 81 rhinoconjunctivitis cases,
88 atopic dermatitis cases, 242 healthy
controls group from population-based
survey (Indoor Environment and Children's
Health); ages 3-5 yrs
Outcome: Clinical exam and parent
interview; allergic rhinoconjunctivitis:
recurrence of at least two or more nasal
symptoms (pruritus, runny nose, sneezing
spells >20, nasal stenosis/mouth
breathing) and ocular symptoms (itching,
conjunctival injection, or watery secretion
in both eyes) when exposed to allergens;
atopic dermatitis: presence of at least 3 of
4 major features and 3 of 23 minor
features; 70% of rhinoconjunctivitis and
50% of atopic dermatitis cases were IgE
positive based on 20 allergen tests
Exposure: DIBP concentrations in dust
samples from bedroom and day care
centers; total DIBP exposure estimated as
a weighted mass fraction
DIBP in dust among controls (ng/g):
Median
Home 27.0
Day care 22.6
Area-weighted 27.2
(weighted by assumed hrs in each
environment)
Analysis: Mann-Whitney U-test
Related study: Callesen etal. (2014a)
(same study population, with exposure
measured in urine sample from
participants
MIBP in urine: median 74.2 ng/mL
(controls)
Median DIBP in dust (ng/g), by case-control status assessed by
clinical examination
Cases
Home
Day care
Area-
weighted
Controls
(n = 242)
27.0
22.6
27.2
Rhinoconjunctivitis
(n = 81)
30.4
22.3
26.8
Atopic dermatitis
(n = 88)
33.4
22.5
33.1
Similar results when based on case status defined by parent-
questionnaire data (n = 56 rhinoconjunctivitis, n = 83 atopic
dermatitis)
Results from Callesen etal. (2014a):
OR (95% Cl) by quartile of MIBP (urine sample), adjusting for sex,
breastfeeding less than 3 mo, smoking in the home, and single
allergic predisposition
Rhinoconjunctivitis
(76 cases, 222 controls)
1 1.0 (referent)
2 1.18(0.54,2.55)
3 0.89 (0.39, 2.02)
4 1.07 (0.52, 2.22)
Atopic dermatitis
(76 cases, 216 controls)
1.0 (referent)
1.11 (0.53, 2.34)
0.88 (0.41, 1.91)
0.97 (0.48, 1.94)
This document is a draft for review purposes only and does not constitute Agency policy,
3-19 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Hoppin et al. (2013)a (United States,
NHANES)
Population: 2,325 participants in
population-based survey (NHANES),
2005-2006; ages >6 yrs
Outcome: Self-administered
questionnaire current allergy symptoms
(hay fever, allergy, itchy rash, rhinitis) in
past yr; allergic sensitization as measured
by serum IgE (19 allergen specific IgEs,
>0.35kU/L)
Exposure: Urine sample collected same
day as serum sample
MIBP in urine (ng/L): Percentile
Median 75th 95th
Children 8.93 16.38 45.97
Adults 5.42 10.53 28.98
Analysis: Logistic regression, adjusting for
variables shown in results column and
sampling weights; separate analyses for
children (ages 6-17 yrs) and adults
(>17yrs)
Prevalence and OR (95% Cl) for allergy symptoms and allergic
sensitization per unit change in log-transformed urinary MIBP level
(adjusted for age, race/ethnicity, gender, BMI, creatinine, and
cotinine)
Children (n = 779)
Hay fever (n = 23)
Rhinitis (n = 188)
IgE sensitization
(any)
Adults (n = 1,546)
Hay fever (n = 88)
Rhinitis (n = 498)
IgE sensitization
(any)
3.6%
27.6%
46.1%
7.4%
35.4%
44.0%
0.12(0.04,0.39)
0.84 (0.53, 1.33)
0.93 (0.51, 1.70)
0.93 (0.46, 1.87)
0.99 (0.76, 1.29)
1.32 (0.99, 1.76)
Authors reported that adjustment for poverty income ratio did not
alter ORs.
Sunetal. (2009)a (China)
Population: Cases of rhinitis (n = 240) or
eczema (n = 61) and controls (n = 204 and
119 for rhinitis and eczema analysis,
respectively), all students of Tianjin
University who had participated in a cross-
sectional study of allergic symptoms and
environmental factors; 2006-2007
Outcome: Self-reported symptoms from
questionnaire: rhinitis = in past 12 mo, had
a problem with sneezing, or a runny, or a
blocked nose when not having a cold or
the flu, or sneezing, or a runny, or a
blocked nose, or itchy-watery eyes after
contact with furred animals or after
contact with pollen; eczema = in past
12 mo, had an itchy rash; controls
responded no to questions on
asthma/wheeze, rhinitis, and eczema
Exposure: Surface dust sample in dorm
rooms
DIBP in dust (ng/g):
Median 75th percentile
20.24 34.77
Analysis: Logistic regression for OR
considering age, gender, passive smoking,
smoking, pet raising, atopy, and building
age as potential covariates; Mann-
Whitney U-test for comparison between
DIBP concentrations of cases and controls;
OR for rhinitis and eczema comparing DIBP in dust (|Jg/g dust) above
and below the median (adjusted for age, gender, smoking, atopy and
building age) reportedly did not reach statistical significance
(quantitative results not reported)
Median Concentration DIBP in dust (|Jg/g dust)
Cases Controls
Rhinitis 20.17 28.76*
Eczema 28.68 22.56
*p = 0.019 by Mann-Whitney test; p = 0.051 by t-test.
This document is a draft for review purposes only and does not constitute Agency policy,
3-20 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
t-test for comparisons between log-
transformed concentrations
Bornehag et al. (2004) (Sweden)
Population: 198 cases, 202 controls from
population-based cohort (Dampness in
Buildings and Health cohort); n = 10,852;
2001-2002; ages 3-8 yrs
Outcome: Eczema, wheezing, or rhinitis
(Cases report at least two incidents of
eczema, or wheezing or rhinitis without a
cold, in the preceding yr, and at follow-up
1.5 yrs later)
Exposure: Surface dust samples from
children's bedrooms
DIBP in dust (mg/g):
Median
All homes 0.045
Analysis: Mann-Whitney U-test for
comparing concentrations in all homes;
t-test for comparing log-transformed
concentrations in homes with
concentrations above detection limit.
Concentration in dust (mg/g dust)
Median, all
homes
(n = 346)
Controls 0.048
Cases (all) 0.042
p > 0.4 in both tests
Geometric mean (95% Cl), homes
with phthalate > detection limit
(n = 290)
0.055 (0.046, 0.065)
0.058 (0.048, 0.070)
1
2
3
4
5
6
Additional results for this study presented in asthma table.
aOR = adjusted odds ratio; IgE = immunoglobin E; NHANES = National Health and Nutrition Examination Survey;
VOC = volatile organic compound
This document is a draft for review purposes only and does not constitute Agency policy,
3-21 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
Table 3-10. Evidence pertaining to DIBP and asthma/wheezing and
hypersensitivity in humans
Reference and study design
Results
Ait Bamai et al. (2014)a (Japan)
Population: Children (n = 122, ages <15 yrs)
and adults (n = 374, ages >15 yrs) living in
148 detached dwellings in which at least
25 mg of dust was collected; 2006 follow-up of
2003 baseline survey.
Outcome: Bronchial asthma assessed by self-
administered questionnaire (positive response
to: in the past 2 yrs have you been seen at a
hospital for bronchial asthma?); parents
completed questionnaires for inhabitants
<6yrs
Exposure: Dust samples from floor and other
surfaces
DIBP (ng/g dust):
Median 75th
percentile
Floor dust (n = 148) 2.4 5.5
Multi-surface dust (n = 120) 1.9 3.5
Analysis: Generalized linear mixed effects
model, considering gender, age strata (<15,
>15 yrs), smoking status (personal and
environmental tobacco smoke), furry pets in
home, signs of dampness, Der 1 (not defined
by authors), other phthalates dust, airborne
fungi, formaldehyde, total VOC, and building
characteristics as potential covariates.
OR (95% Cl) for bronchial asthma by tertile of DIBP in floor dust
(adjusted for adjusted for gender, age strata, smoking status,
dampness index, furry pets inside the home, Der 1, and sum of
other phthalate dusts)
1 (low)
2
3 (high)
(trend
p-value)
Full sample
1.0 (referent)
2.25 (0.48,
10.57)
5.09(1.17,
22.15)
(0.03)
Children Adults
1.0 (referent) 1.0 (referent)
4.37(0.36,53.6) 1.16(0.16,8.17)
8.94 (0.86, 93.0) 2.90 (0.52, 16.2)
(0.067) (0.22)
p-value for age interaction = 0.51
No significantly increased aORs (either in the full sample or
stratified by age) were observed in analyses of bronchial asthma
using DIBP measurements in multisurface dust.
This document is a draft for review purposes only and does not constitute Agency policy,
3-22 DRAFT—DO NOT CITE OR QUOTE
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Callesen et al. (2014b): Callesen et al.
(2014a)a (Denmark)
Population: 72 asthma cases, 242 healthy
controls group from population-based survey
(Indoor Environment and Children's Health);
ages 3-5 yrs
Outcome: Clinical exam and parent interview;
asthma: recurrence of at least two of the
three symptoms: cough, wheeze, and
shortness of breath within the previous 12 mo
(symptoms other than those
triggered by respiratory infections); and
doctor diagnosis of asthma in combination
with ongoing treatment; 47% of asthma cases
were IgE positive based on 20 allergen tests
Exposure: DIBP concentrations in dust
samples from bedroom and day care centers;
total DIBP exposure estimated as a weighted
mass fraction
DIBP in dust among controls (ng/g):
Median
Home 27.0
Day care 22.6
Area-weighted 27.2
(weighted by assumed hrs in each
environment)
Analysis: Mann-Whitney U-test
Related study: Callesen etal. (2014a) (same
study population, with exposure measured in
urine sample from participants
MIBP in urine: median 74.2 ng/mL (controls)
Median DIBP in dust (ng/g), by case-control status assessed by
clinical examination
Home
Day care
Area-weighted
Controls (n = 242)
27.0
22.6
27.2
Asthma (n = 72)
25.8
21.5
25.7
Similar results when based on case status defined by parent-
questionnaire data (n = 110 asthma cases)
Results from Callesen etal. (2014a):
OR (95% Cl) by quartile of MIBP (urine sample), adjusting for sex,
breastfeeding <3 mo, smoking in the home, and single allergic
predisposition
Bronchial asthma
(60 cases, 216 controls)
1.0 (referent)
0.49 (0.22, 1.09)
0.91(0.41,1.69)
0.61 (0.27, 1.34)
Bertelsen et al. (2013) (Norway)
Population: 623 children from birth cohort
(Environment and Childhood Asthma study),
born 1992-1993; children with current asthma
over-sampled (follow-up 2001-2004); ages
10 yrs
Outcome: Current asthma (parental report of
history of asthma plus >1 of the following:
dyspnea, chest tightness, and/or wheezing in
previous 12 mo; use of asthma medications in
previous 12 mo; positive exercise challenge
test)
Exposure: First morning urine sample
(child's), collected at study examination
MIBP in urine (ng/L) Percentile
Median 75th 95th
Unadjusted 49.2 88.4 231.0
SG-adjusted 50.1 90.5 239.6
Analysis: Logistic regression, adjusting for
variables shown in the results column
OR (95% Cl) for current asthma by quartile of MIBP (ng/L)
(adjusted for urine specific gravity, sex, parental asthma, and
household income)
1: <31.4 (referent)
2: >31.4-49.2
3: >49.2-88.4
4: >88.4
1 (referent)
1.3 (0.74, 2.4)
1.4(0.73,2.5)
1.5 (0.80, 2.7)
Increase in odds of current asthma per logio IQR MIBP
(95% Cl) = 1.1 (0.87,1.5)
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Hoppin et al. (2013)a (United States, NHANES)
Population: 2,325 participants in population-
based survey (NHANES), 2005-2006; ages
>6yrs
Outcome: Self-administered questionnaire
(asthma, wheeze in past yr)
Exposure: Urine sample collected same day as
serum sample
Unadjusted MIBP in urine (ng/L):
Percentile
Median 75th 95th
Children 8.93 16.38 45.97
Adults 5.42 10.53 28.98
Analysis: Logistic regression, adjusting for
variables shown in results column and
sampling weights; separate analyses for
children (ages 6-17 yrs) and adults (>17 yrs)
Prevalence and OR (95% Cl) for asthma symptoms per unit
change in log-transformed urinary MIBP level (adjusted for age,
race/ethnicity, gender, BMI, creatinine, and cotinine)
Children (n = 779)
Asthma (n = 65)
Wheeze (n = 80)
Adults (n = 1,546)
Asthma (n = 116)
Wheeze (n = 219)
8.4%
10.7%
7.4%
16.6%
0.92 (0.26, 3.29)
1.08 (0.49, 2.35)
1.39 (0.77, 2.50)
0.92 (0.57, 1.48)
Authors reported that adjustment for poverty income ratio did
not alter ORs.
Sun et al. (2009)a (China)
Population: 92 cases asthma/wheezing, cases
and 346 controls, all students of Tianjin
University who had participated in a cross-
sectional study of allergic symptoms and
environmental factors; 2006-2007
Outcome: Self-reported symptoms from
questionnaire; asthma/wheezing = in past
12 mos, have you had wheezing or whistling
the in the chest; have you had dry cough at
night for more than 2 wks, apart from a cough
associated with a cold or chest infection;
controls responded no to questions on
asthma/wheeze, rhinitis, and eczema
Exposure: Surface dust sample in dorm rooms
DIBPin dust (|jg/g):
Median 75th percentile
20.24 34.77
Analysis: Logistic regression for OR,
considering age, gender, passive smoking,
smoking, pet raising, atopy, and building age
as potential covariates; Mann-Whitney U-test
(nonparametric) for comparison between
DIBP concentrations of cases and controls;
t-test for comparisons between log
transformed concentrations
OR for asthma comparing DIBP in dust (ng/g dust) above and
below the median (adjusted to age, gender, smoking, atopy, and
building age) reportedly did not reach statistical significance
(quantitative results not reported)
Median concentration DIBP in dust (ng/g dust)
Cases
Wheezing 23.13
(p > 0.46 by Mann Whitney or t-test)
Controls
22.73
1
2
3
4
Additional results for this study presented in allergy/immune table.
IQR = interquartile range
This document is a draft for review purposes only and does not constitute Agency policy,
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2
3
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.2.8. Neurodevelopmental Effects in Humans
Table 3-11. Evidence pertaining to DIBP and neurodevelopmental effects in
humans
Reference and study design
Results
Attention and executive function in school-aged children
Kobroslv et al. (2014) (United States; Minnesota,
Missouri, California, Iowa)
Population: 153 children (n = 76 girls, n = 77 boys)
from birth cohort study (Study for Future Families),
born 2000-2005, ages 6-10 yrs in 2010 follow-up
Outcome: Child Behavior Checklist (completed by
parent)
Exposure: Maternal urine sample, 3rd trimester
(mean 26.6 wks)
Unadjusted MIBP in urine (ng/mL):
Geometric mean (95% Cl)
2.3 (2.0,2.8)
Analysis: Linear regression, considering sex, age,
mother's education, urinary creatinine, family stress
measure, and race/ethnicity, as potential covariates.
Related references: Swan et al. (2005) (exposure
data)
Regression coefficient (95% Cl) for change in raw score on
child behavior checklist per unit increase in In-transformed
MIBP (adjusted for sex, age, mother's education and
urinary creatinine, and family stress score)
Anxiety/
depression
Withdrawn
Somatic
complaints
Social problems
Thought
problems
Attention
problems
Rule-breaking
behavior *
Aggressive
behavior
Internalizing
behavior
Externalizing
behavior
Boys Girls
0.11 (-0.13, 0.34) -0.03 (-0.29, 0.22)
-0.01 (-0.21, 0.18) -0.04 (-0.25, 0.17)
-0.03 (-0.23, 0.16) -0.07 (-0.28, 0.13)
0.18 (-0.02, 0.37) -0.06 (-0.27, 0.16)
0.15 (-0.05, 0.35) 0.07 (-0.15, 0.29)
0.27 (0.04, 0.50)
0.20 (0.01, 0.38)
0.34 (0.09, 0.59)
0.12 (-0.12, 0.36)
-0.04 (-0.23, 0.16)
0.12 (-0.14, 0.39)
0.09 (-0.18, 0.37) -0.07 (-0.37, 0.22)
0.32 (0.06, 0.58) 0.06 (-0.22, 0.34)
Total problems 0.42 (0.05, 0.80) 0.07 (-0.33, 0.47)
*Sex interaction p-value = 0.04; all other interaction
p-values >0.05
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Engel et al. (2010) (United States; New York City)
Population: 177 children from original birth cohort
studied by Engel etal. (2009), 54% boys, three
follow-up exams at ages 4.5-5.5, 6-6.5, 7-9 yrs
Outcome: Behavior assessed by maternal reporting
on BRIEF and BASC-PRS
Exposure: Maternal urine sample, 25-40 wks
gestation*
Median 75th percentile
MIBP (ng/L)* 2.6 12.2
Sum LMW (U.M/L) 1.88 4.59
(sum of MBP, MEP, MIBP, and MMP)
Analysis: Generalized linear regression model,
adjusting for variables shown in results column;
other variables (not specified) were considered
Related references: Engel et al. (2009) (exposure
data for n = 295 children in the cohort)
*MIBP concentrations not reported in (Engel et al.,
2010); values reported here are from an earlier
analysis of this cohort described in Engel et al.
(2009)
Regression coefficient for change in behavioral score
(BASC-PRS) per unit increase in In-phthalate level (nM/L) in
boys (adjusted for race, educational level and marital
status of the primary caretaker, and urinary creatinine)
MIBP
Low molecular
weight phthalate
sum
Clinical scales (higher score = more problem behaviors)
Aggression -0.12
Anxiety -0.25
Attention 0.66
problems
Atypicality 0.53
Conduct problems 0.23
Depression 0.29
Hyperactivity 0.85
Somatization 1.04
1.24*
0.78
1.29*
0.95
2.40*
1.18*
1.03
0.36
Withdrawal -0.01 0.46
Adaptive scales (lower score = more problem behaviors)
Adaptability -1.32* -1.08*
Leadership -1.30 -0.88
Social skills -0.93 -1.04
Composite scales (higher score = more problem behaviors)
0.33 1.75*
Externalizing
problems
Internalizing
problems
Adaptive skills
Behavioral
Symptoms Index
0.46
-1.17
0.47
0.99
-0.98
1.55*
Significant sex-phthalate interactions (p < 0.05) for
aggression, conduct problems, hyperactivity, externalizing
problems, and behavioral symptoms index, as reported by
study authors.
Regression coefficient for change in behavioral score
(BRIEF scores; higher score = worse executive functioning)
per unit increase in In-phthalate level (nM/L) in boys and
girls (adjusted for race, sex, educational level and marital
status of the primary caretaker, and urinary creatinine)
Emotional control
0.09
1.33*
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Behavioral 0.30 1.13
regulation index
Initiate 0.83 0.81
Working memory 1.11 1.03
Plan/organize 0.76 1.02
Metacognition 0.70 1.05
index
Global executive 0.56 1.23*
composite
*p < 0.05
Study authors reported that there were few significant
associations between phthalate concentration and
behavior among girls (quantitative results not reported).
Neurobehavioral outcomes in infants and preschool-aged children
Braun et al. (2014) (United States)
Population: 175 children from birth cohort in Ohio
(HOME cohort, recruited during pregnancy,
2003-2006); follow-up at ages 4-5 yrs
Outcome: Autistic behaviors based on Social
Responsiveness Scale completed by mother; 65 item
scale, higher score = more autistic behaviors
Exposure: Maternal urine samples, 16-26 wks of
gestation
MIBP in urine (u.g/g Cr):
Percentile
Median 75th 95th
Cr-adjusted 5.6 8.6 17
Analysis: Semi-Bayesian hierarchical regression
model
Regression coefficient (95% Cl) for change in total score
per unit increase in log-transformed Cr-adjusted MIBP
(adjusted for maternal demographic and perinatal factors,
depressive symptoms, caregiving environment, and serum
cotinine):
0.7 (-1.4, 2.8)
Tellez-Roio et al. (2013) (Mexico)
Population: 135 children from birth cohort (Early
Life Exposure in Mexico to Environmental Toxicants
cohort; mothers recruited during first trimester,
1997-2003)
Outcome: Mental and psychomotor development
based on Bayley Scales of Infant Development-ll
(assessed by trained examiner, videotaped for
quality control assessment) tested at 24, 30, and
36 mo of age
Exposure: Maternal urine sample, 3rd trimester
MIBP in urine (ng/mL):
Geometric mean (95% Cl)
SG-adjusted 2.30 (1.92, 2.76)
Analysis: Linear regression for longitudinal data,
stratified by sex and adjusted for variables shown in
results column
Related reference: Ettinger et al. (2009)
Regression coefficient (95% Cl) for change in
neurodevelopment score per unit increase in maternal In-
MIBP (adjusted for birthweight, breastfeeding practices,
weight-for-age, child's age, mother's age, mother's
education, and laboratory)
Total sample
(n = 135)
Boys
(n = 64)
Girls (n = 71)
MDI
PDI
0.53 0.32 -0.12
(-0.85, 1.91) (-1.62, 2.28) (-1.94, 1.69)
0.57 0.63 0.37
(-0.67, 1.82) (-0.68, 1.95) (-1.67, 2.43)
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Whvattetal. (2012) (United States, New York City)
Population: 297 children from birth cohort
(Columbia Center for Children's Environmental
Health), born 1999-2006; 3-yr follow-up, mean age
36 mo (range 27-42 mo)
Outcome: Mental, psychomotor and behavioral
development at 3 yrs based on Bayley Scales of
Infant Development-ll (assessed by trained
examiners) and Child Behavior Checklist (completed
by parent)
Exposure: Maternal urine sample, 3rd trimester
MIBP in urine (ng/mL):
Geometric mean
Unadjusted 9.3
Analysis: Linear and logistic regression adjusting for
variables shown in results column; Wald test used to
detect sex differences
Regression coefficient (95% Cl) for change in
neurodevelopment score per unit increase in maternal In-
MIBP (adjusted for specific gravity, race/ethnicity,
maternal marital status and prenatal alcohol consumption,
child's gestational age and sex, and quality of care-taking
environment)
Boys (n = 140) Girls (n = 157)
MDI
PDI
0.59
(-1.40, 2.58)
-2.21
(-4.61,0.19)
-1.33
(-3.20, 0.54)
-2.33
(-4.59, -0.08)
OR (95% Cl) for risk of mental or psychomotor delay (score
<85) per In-unit increase in maternal In-MIBP (each model
adjusted for one or more of the following: specific gravity,
race/ethnicity, maternal marital status and prenatal
alcohol consumption, child's gestational age and sex, and
quality of care-taking environment)
Boys (n = 140) Girls (n = 157)
MDI
PDI
0.87
(0.60, 1.28)
1.80
(1.13, 2.87)
0.98
(0.62, 1.56)
1.98
(1.02, 3.83)
Regression coefficient (95% Cl) for change in
neurobehavior per unit increase in maternal In-MIBP
(adjusted for specific gravity; ethnicity; maternal IQ,
demoralization, hardship, satisfaction during pregnancy
and prenatal exposure to PAH and BPA; and child's sex and
age at testing)
Boys (n = 129) Girls (n = 148)
Emotionally reactive
Anxious/depressed
Somatic complaints
Withdrawn behavior
0.42
(-0.005, 0.85)
0.12
(-0.38, 0.61)
0.31
(-0.18,0.81)
0.36
(-0.05, 0.77)
0.34
(-0.11,0.78)
0.16
(-0.34, 0.66)
0.24
(-0.22, 0.70)
0.47
(-0.007, 0.94)
1.21
(-0.16,2.56)
Internalizing behavior
No effect modification by gender was observed (p-values
1.20
(-0.15, 2.55)
OR (95% Cl) for child's score in the borderline or clinical
range (compared to normal) per unit increase in maternal
In-MBP (adjusted for specific gravity, maternal
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
demoralization and satisfaction during pregnancy, and
child's sex and age at testing)
Somatic complaints
Withdrawn behavior
Internalizing behavior
Borderline
1.29
Clinical
0.76
(0.84, 1.99) (0.42, 1.36)
0.81
(0.44, 1.51)
1.98
1.62
(0.97, 2.73)
1.41
(1.24, 3.23) (0.91, 2.18)
1
2
3
4
5
BASC-PRS = Behavior Assessment System for Children —Parent Rating Scales; BPA = bisphenol A; BRIEF = Behavior
Rating Inventory of Executive Function; HOME = Health Outcomes and Measures of the Environment; LMW = low
molecular weight; MDI = mental delay index; MMP = monomethyl phthalate; PAH = polycyclic aromatic
hydrocarbon; PDI = psychomotor delay index
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1
2
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.2.9. Thyroid Hormone Effects in Humans
Table 3-12. Evidence pertaining to DIBP and thyroid hormones in humans
Reference and study design
Results
Dirtuetal. (2013) (Belgium)
Population: 152 overweight or obese adults from
weight loss cohort (ENDORUP) seen at weight
management clinic, 43 age- and sex-matched
controls from hospital staff and other volunteers,
enrolled 2009-2012; among obese/overweight
group, 65 received bariatric surgery and 87
received standard diet and lifestyle counseling;
follow-up 3, 6, and 12 mo
Outcome: Serum thyroid hormone levels (details of
blood collection were not reported)
Exposure: Urine sample (24-hr)
MIBP in urine (ng/mL):
Median
Controls 65
Obese (at baseline) 58
Percentile
75th 90th
93 133
89 129
Regression coefficient (p-value) for change in hormone level
with unit change in In-MIBP (adjusted for age, weight loss,
and sex, or stratified by sex) (0.0 = no effect)
Full sample Men Women
Overweight/obese group
FreeT4 0.07(0.41) 0.11(0.47) 0.05(0.66)
TSH -0.01(0.93) 0.09(0.58) -0.01(0.94)
Referent group
FreeT4 0.24(0.14) 0.49(0.12) 0.16(0.44)
TSH 0.23(0.16) -0.43(0.19) 0.32(0.10)
Analysis: Linear regression, adjusting for variables
shown in results column
3
4
5
Meeker and Ferguson (2011) (United States)
Population: Participants in population-based
survey (NHANES), 2007-2008; 1,346 ages >20 yrs
and 329 adolescents ages 12-19 yrs
Outcome: Serum thyroid hormone levels
Exposure: Urine sample collected same day as
serum sample
Cr-adjusted MIBP in urine (u.g/g Cr):
Percentile
Median 75th 95th
Adults 6.67 11.1 24.1
Adolescents 8.24 13.73 28.78
Analysis: Linear regression adjusting for variables
shown in results column.
Regression coefficient (95% Cl) for change in hormone level
with unit increase in In-MIBP (adjusted for age, sex, race,
BMI, In-serum cotinine, In-urinary creatinine, and In-urinary
iodine, and weighted for sampling strategy)
TotalT3(ng/dL)
Ln(FreeTS)
(pg/mL)
TotalT4(u.g/mL)
Ln(FreeT4)
(ng/dL)
Ln (TSH)
(ulU/mL)
Ln (Tg) (ng/mL)
Adults
0.77
(-0.59, 2.12)
-0.0012
(-0.0074, 0.0051)
0.020
(-0.075,0.11)
0.0010
(-0.0094, 0.011)
-0.013
(-0.054, 0.028)
-0.018
(-0.081, 0.045)
Adolescents
2.30
(-0.81, 0.52)
0.0083
(-0.0062, 0.023)
-0.034
(-0.25,0.19)
-0.0001
(-0.021, 0.021)
0.003
(-0.076, 0.081)
-0.047
(-0.12,0.074)
T3 = triiodothyronine; T4 = thyroxine; TSH = thyroid stimulating hormone
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 3.2.10. Obesity and Metabolic Effects in Humans
2 Table 3-13. Evidence pertaining to DIBP and obesity in humans
Reference and study design
Results
Buser et al. (2014) (United States,
NHANES)
Population: Participants in population-
based survey (NHANES), 2007-2010ages
>6 yrs [sample size not reported]
Outcome: BMI measured at exam; divided
into obese (BMI z-score >95th percentile in
children, BMI >30 in adults) and
overweight (BMI z-score 85th-95th
percentiles in children, BMI 25-29.9 in
adults).
Exposure: Urine sample, collected at same
time as exam
Unadjusted MIBP in urine (ng/mL)
Geometric mean (SE)
Ages 6-19 yrs 10.43 (0.39)
Ages >20 yrs 6.75(0.23)
Analysis: Logistic regression, considering
age, race/ethnicity, sex, urinary creatinine,
poverty income ratio, calorie intake, and
serum cotinine as potential covariates in
analyses of ages 6-19 yrs; or age,
race/ethnicity, sex, education, diabetes,
alcohol consumption, cigarette smoking,
calorie intake, vigorous recreational
activities, urinary creatinine, and serum
cotinine as potential covariates in analysis
of ages >20yrs
OR (95% Cl) in children (6-19 yrs of age) for obesity or overweight
comparing highest quartile urinary MIBP (>20.84 ng/mL) with lowest
quartile (<5.38 ng/mL) (adjusted for age, race/ethnicity, calorie
intake, serum cotinine, urinary creatinine, income level)
All
Boys
Girls
Obese
1.82 (0.73, 4.57)
4.26 (1.32, 13.74)
0.57 (0.18, 1.83)
Overweight
1.85 (0.78, 4.40)
2.22 (0.78, 6.28)
1.57 (0.58, 4.25)
OR (95% Cl) in adults (>20 yrs of age) for obesity or overweight
comparing highest quartile urinary MIBP (>14.40 ng/mL) with lowest
quartile (<3.49 ng/mL) (adjusted for age, gender, race/ethnicity,
calorie intake, recreational activity, serum cotinine, education level,
smoking status, alcohol intake, diabetes)
All
Men
Women
Obese
1.40(0.90,2.16)
0.98 (0.57, 1.67)
1.81 (0.94, 3.48)
Overweight
1.18 (0.79,1.78)
1.06 (0.60, 1.89)
1.25 (0.66, 2.36)
Hart et al. (2013) (Australia)
Population: 121 girls from birth cohort
study (Western Australian Pregnancy
Cohort), whose mothers were recruited at
18 wks of gestation between 1989 and
1991; follow-up at ages 14-16 yrs
Outcome: Offspring BMI (height and
weight measured at clinic visit on d 2-5 of
menstrual cycle)
Exposure: Maternal serum samples
(n = 123) collected at 18 and 34-36 wks of
gestation (combined aliquot from both
time periods)
MIBP in serum (ng/mL):
Median 90th percentile
Unadjusted 1.77 6.16
Analysis: Correlation between log-
transformed MIBP and BMI
Authors reported no association between adolescent BMI (either as
absolute value or as age- and gender-adjusted z-score) and any
phthalate metabolite in maternal serum (r = -0.10-0.04,
p = 0.345-0.931)
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Trasande et al. (2013a) (United States,
NHANES)
Population: 2,884 participants in
population-based survey (NHANES),
2003-2008; 6-19 yrs old
Outcome: BMI z-score, obesity (BMI
z-score >95th percentile), and overweight
(BMI z-score >85th percentile) (measured)
Exposure: Urine sample, collected at same
time as BMI measurement
ZLMW phthalates in urine (nM):
Geometric mean
Not obese 0.701
Obese 0.855
ZLMW phthalates = sum of MEP, MBP,
MIBP, andMCPP
Analysis: Logistic regression for
overweight and obese classification; linear
regression of BMI z-score as continuous
variable; adjusted for variables shown in
results column
Full sample results, no association with In-LMW phthalates: OR or
regression coefficient (95% Cl) per one unit increase in £LMW
phthalates (nM) (adjusted for urinary creatinine, sex, poverty-
income ratio, parental education, serum cotinine, age, and
race/ethnicity, caloric intake, and television watching)
Overweight
Obese
BMI z-score
OR (95% Cl)
OR (95% Cl)
P (95% Cl)
1.01(0.90,1.13)
1.02(0.90,1.17)
0.03 (-0.03, 0.09)
Interaction by ethnicity seen, with associations seen between In-
LMW phthalates and each of the obesity measures in blacks, but not
in whites or Hispanics. The patterns seen with ZLMW phthalates
were also seen in analyses for MIBP. Using same adjustment factors
as above, the associations with In-MIBP are:
ILMW phthalates
Hispanic White
MIBP
Black Black
Over-
weight OR
(95% Cl)
Obese OR
(95% Cl)
BMIz-
score P
(95% Cl)
0.88
(0.72, 1.08)
0.97
(0.78, 1.22)
1.21
(1.05, 1.39)
1.16
(0.99, 1.37)
0.97 0.94
(0.83, 1.14) (0.69, 1.29)
1.22 1.17
(1.07, 1.39) (0.97, 1.41)
-0.04
(-0.15,
0.06)
0.02
(-0.08,0.12)
0.09
(0.003,0.18)
0.08
(-0.01,
0.17)
Wang et al. (2013) (China)
Population: 259 primary and middle
school students, 8-15 yrs old, stratified
sample from six schools, selected based on
sex and BMI
Outcome: BMI, waist circumference
(measured)
Exposure: First morning urine sample,
collected at same time as BMI
measurement
MIBP in urine (ng/mL):
Geometric mean (SD)
38.9(1.1)
Low molecular weight phthalate
metabolites included MMP, MEP, MBP,
MIBP, and MHBP
Analysis: Linear regression, sampling
weights applied to adjust for sampling
strategy; adjusted for variables shown in
the results column
Regression coefficient (95% Cl) for change in BMI or waist
circumference per unit increase in SG-adjusted InMIBP (adjusted for
age and sex in Model 1; plus sum of DBP, MMP, and MEP in Model
2)
BMI
Waist
circumference
Model 1
0.027 (0.006, 0.048)
0.022(0.005, 0.038)
Model 2
0.020 (-0.005, 0.045)
0.019 (-0.001, 0.038)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Dirtuetal. (2013) (Belgium)
Population: 152 overweight or obese
adults from weight loss cohort (ENDORUP)
seen at weight management clinic, 43 age-
and sex-matched controls from hospital
staff and other volunteers, enrolled
2009-2012; among obese/overweight
group, 65 received bariatric surgery and
87 received standard diet and lifestyle
counseling; follow-up 3, 6, and 12 mo
Outcome: Waist circumference measured
at each follow-up visit
Exposure: Urine sample (24-hr sample)
MIBP, in urine (ng/mL):
Percentile
Median 75th 90th
Controls 65 93 133
Obese 58 89 129
(at baseline)
Analysis: Linear regression, adjusting for
variables shown in results column;
treatment of repeated urinary phthalate
measures was not specified
Regression coefficient (p-value) for change in waist circumference
with unit change in In-MIBP (adjusted for age, weight loss, and sex,
or stratified by sex) (0.0 = no effect)
Overweight/ obese
group
Referent group
Full sample
0.07 (0.40)
Men
-0.16(0.30)
Women
0.03 (0.76)
-0.16(0.30) 0.07(0.81) -0.01(0.98)
Lind et al. (2012a) (Sweden)
Population: 1,016 (507 men, 509 women),
from population-based cohort (Prospective
Investigation of Vasculature in Uppsala
Seniors study), 2001-2003; age 70 yrs at
enrollment
Outcome: BMI, waist circumference
measured at enrollment; DXA (n = 890
participated) and MRI of abdominal region
(n = 287 randomly selected) 2 yrs later
Exposure: Serum sample (fasting),
collected at baseline
MIBP in serum (ng/mL):
Median 75th percentile
Women 13.4 24.5
Men 13.5 33.3
Analysis: Linear regression, adjusted for
variables shown in results column
Related reference: Olsen etal. (2012)
reports cross-sectional analysis of BMI
from this study population, see Table 14
Regression coefficient (95% Cl) for change in body metric per unit
increase in In-MIBP (ng/mL) (adjusted for serum cholesterol and
triglycerides, education, exercise, and smoking)
Outcome
BMI (kg/m2)
Waist circumference
(cm)
DXA total fat (kg)
MRI visceral adipose
tissue (cm2)
Males
P (95% Cl)
-0.083 (-0.35,
0.19)
-0.025 (-0.80,
0.75)
-73 (-754, 608)
-5.9 (-24, 13)
Females
P (95% Cl)
0.39 (0.002, 0.79)
1.3 (0.425, 2.3)
1,079 (283, 1875)
14 (1.4, 26)
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Teitelbaum et al. (2012) (United States,
New York City)
Population: 387 children (80 boys, 307
girls) in child development cohort (Growing
Up Healthy Study), 2004-2008; Hispanic
and black), 6-8 yrs at enrollment
Outcome: BMI and waist circumference
measured 1 yr after enrollment; normal
weight = BMI <85th percentile (n = 2,284);
overweight = BMI >85th percentile (n = 578)
Exposure: Urine sample, collected at
enrollment
Cr-adjusted phthalates in urine (u.g/g Cr),
median:
MIBP ILMW phthalates
Boys 22.7 253.2
Girls 22.2 294.0
Low molecular weight phthalate
metabolites included MEP, MBP, MIBP, and
MCPP.
Analysis: Linear regression, considering
sex, age at baseline, sedentary hrs,
metabolic equivalent hrs, caloric intake,
race, ethnicity, season of urine collection,
family income, and parent education as
potential covariates; restricted to children
with creatinine >10 mg/dL
Full sample results, regression coefficient (95% Cl) for change in
body metric per unit change in In-MIBP (ng/g Cr) (adjusted for
creatinine, age, sex, sedentary hrs, metabolic equivalent hrs,
Hispanic ethnicity, caloric intake, season, and parental education
level)
BMI (kg/m2)
Waist circumference (cm)
-0.27 (-0.73, -0.18)
-0.62 (-1.84, -0.61)
Olsenetal. (2012) (Sweden)
Population: 1,016 (507 men, 509 women),
from population-based cohort (Prospective
Investigation of Vasculature in Uppsala
Seniors study), 2001-2003; age 70 yrs at
enrollment
Outcome: BMI measured at study visit
Exposure: Serum sample, collected at time
of examination; results not shown
Analysis: Linear regression, adjusted for
the variables shown in results column
Regression coefficient for change in outcome per unit increase in In-
MIBP (adjusted for sex, smoking, diabetes (except for glucose) and
the other variables in the table; model for Framingham Risk Score
only adjusted for sex)
BMI
0.094 (-0.13, 0.32)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Kasper-Sonnenberg et al. (2012)
(Germany)
Population: 104 mothers (and children)
enrolled in birth cohort study, children
born between 2000 and 2002, follow-up in
2007-2009; mean age 39.2 yrs (mothers),
6.8yrs (children)
Outcome: BMI based on questionnaire
(mothers) and measurements (children)
Exposure: Urine sample (first morning),
collected on same day as exam
Cr-adjusted MIBP and OH-MIBP in urine
(ug/gCr):
Geometric mean (95% Cl)
Children
64.6 (55.2, 75.7)
34.2 (28.5, 41.0)
101 (87.2, 118)
Spearman correlation coefficient between £DIBP anc| BMI jn
Children -0.035 (p> 0.05)
Mothers -0.137 (p> 0.05)
MIBP
OH-MIBP
IDIBP
Adults
MIBP
OH-MIBP
IDIBP
37.2(31.8,43.5)
17.4(15.1,20.0)
55.9 (48.4, 64.5)
Analysis: Spearman's rank correlation
analysis
1
2
3
4
Svensson et al. (2011) (Mexico)
Population: 182 women; healthy controls
without diabetes from case-control study
of breast cancer, 2007-2008; mean age
54 yrs
Outcome: BMI, waist circumference, and
waist:height ratio
Exposure: First morning urine sample
collected at time of clinical evaluation
Cr-adjusted MIBP in urine (u.g/g Cr):
Geometric mean (SD)
No diabetes 9.1(2.3)
Analysis: Spearman correlation coefficient
Related references: Lopez-Carrillo et al.
(2010)
Spearman correlation coefficient between anthropometric measure
and In-MIBP in urine (u.g/g Cr)
BMI (kg/m2)
Waist circumference (cm)
Waist/height ratio
(p > 0.05 for all parameters)
0.0457
0.0151
-0.0156
DXA = dual energy x-ray absorptiometry; MCPP = mono-(3-carboxypropyl) phthalate; MHBP = mono-
(S-hydroxybutyl)phthalate; MRI= magnetic resonance imaging; SE = standard error
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
Table 3-14. Evidence pertaining to DIBP and diabetes/insulin resistance in
humans
Reference and study design
Results
Diabetes diagnosis
James-Todd et al. (2012) (United States,
NHANES)
Population: 215 cases, 1,235 controls from
population-based survey (NHANES),
2001-2008; women age 20-79 yrs
Outcome: Positive response to, "Other
than during pregnancy, have you ever been
told by a doctor or health professional that
you have diabetes or sugar diabetes?"
Exposure: Urine sample, collected at time
of survey
MIBP in urine (units not reported):
Geometric mean
Unadjusted 3.7
(based on larger sample of 2,350 women)
Analysis: Logistic regression, adjusting for
variables shown in the results column
OR (95% Cl) for diabetes by quartile of MIBP (adjusted for urinary
creatinine, age, race/ethnicity, education, poverty status, fasting
time, total caloric intake, total fat intake, smoking status, and
physical activity; little change with additional adjustment for BMI
and waist circumference)
MIBP quartile
1 (low)
2
3
4 (high)
1.0 (referent)
1.04 (0.66-1.67)
1.69 (0.93-3.06)
1.95 (0.99-3.85)
Lind et al. (2012b) (Sweden)
Population: 1,003 (501 men, 502 women),
from population-based cohort (Prospective
Investigation of Vasculature in Uppsala
Seniors study), 2001-2003; age 70 yrs at
enrollment
Outcome: Diabetes (n = 88; history of
diabetes or fasting glucose >7.0 mmol/L,
mean duration 8.9 yrs);
Exposure: Serum sample (fasting),
collected at time of clinical assessment
MIBP in serum (ng/mL):
Median 75th percentile
Women 13.4 24.5
Men 13.5 33.3
Analysis: Logistic regression for diabetes
classification, adjusting for variables shown
in results column
OR (95% Cl) per unit increase in serum In-MIBP (adjusted for sex,
serum cholesterol and triglycerides, BMI, smoking, exercise, and
education)
1.30(1.10,1.55)
OR (95% Cl) by quintile of In-MIBP (adjusted for sex, serum
cholesterol and triglycerides, BMI, smoking, exercise, and education)
MIBP quintile
1 (low)
2
3
4
5 (high)
(trend p)
1.0 (referent)
1.19(0.59,2.38)
0.84 (0.41, 1.76)
1.37 (0.7, 2.66)
2.00 (1.03, 3.99)
(0.038)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Svensson et al. (2011) (Mexico)
Population: 221 women with diabetes,
182 healthy without diabetes from case-
control study of breast cancer, 2007-2008;
mean age 54 yrs
Outcome: Self-reported diabetes
Exposure: First morning urine samples
MIBP in urine (u.g/g creatinine):
Geometric mean (SD)
No diabetes 9.1(2.3)
Diabetes 7.9(2.1)
Analysis: Logistic regression, adjusted for
variables shown in the results column (age
and waist-height ratio not found to be
potential confounders)
OR (95% Cl) per unit increase in In-MIBP (adjusted for creatinine and
education)
1.01 (0.65,1.55)
Markers of insulin resistance
Huang etal.(2014a) (United States,
NHANES)
Population: 3,083 participants in
population-based survey (NHANES),
2001-2008; ages 12-<80 yrs; self-reported
non-diabetic, non-pregnant participants
Outcome: Fasting blood glucose; fasting
insulin; HOMA-IR
Exposure: Urine sample at time of clinical
exam
Cr-adjusted MIBP in urine (ng/g Cr):
Median 75th percentile
Men 3.8 6.6
Women 4.9 8.9
Analysis: Logistic regression, adjusting for
variables shown in the results column
Median change (95% Cl) in biomarkers for diabetes by quartile of
MIBP (adjusted for age, gender, race/ethnicity, fasting time, urinary
creatinine, total caloric intake, triglycerides, education, and poverty
and smoking status)
MIBP
quartile
1 (low)
2
3
4 (high)
(pfor
trend)
Fasting glucose
1.0 (referent)
1.87 (0.83, 2.92)
2.77 (1.75, 3.80)
3.69 (2.60, 4.78)
(<0.0001)
Fasting insulin HOMA-IR
1.0 (referent) 1.0 (referent)
1.45 (0.85, 2.04) 0.38 (0.23, 0.52)
1.23 (0.57, 1.89) 0.35 (0.19, 0.51)
1.73 (0.92, 2.54) 0.53 (0.33, 0.72)
(0.0028) (0.0002)
Trasande et al. (2013c) (United States,
NHANES)
Population: 766 participants in the
2003-2008 NHANES, 12-19 yrs old
Outcome: HOMA, calculated as fasting
glucose (mmol/L) multiplied by fasting
insulin (nU/mL divided by 22.5.
Exposure: Urine sample, collected at same
time as insulin resistance measurements.
ZLMW phthalates in urine (nM):
Median 75th percentile
Unadjusted 0.83 1.89
ZLMW phthalates = sum of MEP, MBP, and
MIBP
Urinary concentration of MIBP alone not
reported.
Analysis: HOMA-IR assessed as continuous
or categorical variable; categorical analysis
OR (95% Cl) for insulin resistance and In-urinary metabolite
concentration (nM), adjusted for urinary creatinine, BMI category,
continuous age, race/ethnicity, caregiver education, poverty-income
ratio, gender, serum cotinine, and caloric intake
Ln-MIBP
Ln-ZLMW
1.57(1.18,2.09)
0.92 (0.71, 1,19)
Regression coefficient (95% Cl) for increase in In-HOMA-IR per unit
increase in In-urinary metabolite concentration (nM), adjusted for
urinary creatinine, BMI category, continuous age, race/ethnicity,
caregiver education, poverty-income ratio, gender, serum cotinine,
and caloric intake.
Ln-MIBP
Ln-ZLMW
0.15(0.04,0.26)
-0.07 (-0.18, 0.04)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
used cut point of 4.39, reflecting >2 SD
above the mean HOMA-IR for normal
weight adolescents with normal fasting
glucose in NHANES 1999-2002. Linear and
logistic regression analyses, adjusting for
variables shown in results column. HOMA-
IR and urinary phthalate measures natural-
log transformed for analysis.
James-Todd et al. (2012) (United States,
NHANES)
Population: 2,092 women without history
of diabetes with various measures of
insulin resistance from population-based
survey (NHANES), 2001-2008; women age
20-79 yrs
Outcome: Among women without history
of diabetes, FBG (n = 985), HOMA-IR
(n = 971), glycosolated hemoglobin Ale
(n = 2,092)
Exposure: Urine sample, collected at time
of survey
MIBP in urine (units not reported):
Geometric mean
Unadjusted 3.7
Analysis: Logistic regression, adjusting for
variables shown in the results column
Results
Among women
without diabetes, difference (from first quartile) in
median value (95% Cl) of glucose and insulin parameters by quartile
of MIBP (Model
education level,
total fat intake,
1 adjusted for urine creatinine, age, race/ethnicity,
poverty status, fasting time, total caloric intake,
smoking status, and physical activity; Model 2 also
adjusted for BMI and waist circumference)
MIBP Quartile
FBG (mg/dL)
1 (low)
2
3
4 (high)
Ln (HOMA)
1 (low)
2
3
4 (high)
Ale (%)
1 (low)
2
3
4 (high)
Model 1 Model 2
(referent) (referent)
3.08 (1.22, 4.93) 3.03 (1.05, 5.00)
3.50(1.45,5.54) 3.17(1.17,5.17)
5.86 (3.55, 8.17) 6.04 (3.81, 8.28)
(referent) (referent)
0.13 (-0.02, 0.28) 0.13 (0.01, 0.25)
0.08 (-0.08, 0.25) 0.10 (-0.01, 0.21)
0.22(0.06,0.38) 0.18(0.06,0.31)
(referent) (referent)
0.03 (-0.01, 0.08) 0.03 (-0.01, 0.08)
0.03 (-0.02, 0.09) 0.04 (0.00, 0.09)
0.01 (-0.05, 0.07) 0.01 (-0.04, 0.07)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
Lind et al. (2012b) (Sweden)
Population: 1,003 (501 men, 502 women),
from population-based cohort (Prospective
Investigation of Vasculature in Uppsala
Seniors study), 2001-2003; age 70 yrs at
enrollment
Outcome: Ratio of fasting proinsulin to
insulin; HOMA
Exposure: Serum sample (fasting),
collected at time of clinical assessment
MIBP in serum (ng/mL):
Median 75th percentile
Women 13.4 24.5
Men 13.5 33.3
Analysis: Linear regression for continuous
outcomes (proinsulin/insulin and HOMA-
IR); adjusting for variables shown in results
column
Related reference: Olsen etal. (2012)
presents blood glucose data for this study
population; the regression coefficient per
unit increase in serum In-MIBP was 0.024
(0.01, 0.04) (see Table 14)
Regression coefficient (95% Cl) for insulin measures per unit
increase in serum In-MIBP (adjusted for sex, serum cholesterol and
triglycerides, BMI, smoking, exercise, and education)
Proinsulin/insulin
HOMA
0.06 (0.03, 0.089)
0.014 (-0.015, 0.043)
The magnitude of the association between proinsulin/insulin and
MIBP was similar to that for MEHP, but in the opposite direction of
MEP and MMP (-0.05 and -0.005, respectively). The magnitude of
the association between HOMA-IR and MIBP was lesser than that
for MEP and MMP. The magnitude of the association between
prevalent diabetes and MIBP was greater than that for MEHP, and
less than that for MEP and MMP in the highest quintile.
Olsen etal. (2012) (Sweden)
Population: 1,016 (507 men, 509 women),
from population-based cohort (Prospective
Investigation of Vasculature in Uppsala
Seniors study), 2001-2003; age 70 yrs at
enrollment
Outcome: Fasting serum sample for
glucose
Exposure: Serum sample, collected at time
of examination; results not shown
Analysis: Linear regression, adjusted for
the variables shown in results column.
Regression coefficient for change in outcome per unit increase in In-
MIBP (adjusted for sex, smoking, diabetes (except for glucose), and
the other variables in the table; model for Framingham Risk Score
only adjusted for sex)
Fasting serum glucose
0.024 (0.01, 0.04; p = 0.0001
1
2
FBG = fasting blood glucose; HOMA-IR = homeostatic model assessment of insulin resistance
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
Table 3-15. Evidence pertaining to DIBP and cardiovascular disease risk
factors in humans
Reference and study design
Results
Shiue (2014) (United States, NHANES)
Population: 2,489 participants in
population-based survey (NHANES),
2011-2012; ages >20 yrs
Outcome: High blood pressure (systolic
blood pressure >140 mmHg and diastolic
blood pressure >90 mmHg)
Exposure: Urine sample collected at time
of clinical exam
MIBP in urine (units not given)
Mean ±SD
Normal BP 13.13 ±22.17
High BP 15.71 ±25.15
Analysis: Survey-weighted logistic
regression, adjusting for variables shown in
results column; t-test for comparison
between concentrations
OR (95% Cl) for high blood pressure with increased log-transformed
MIBP (adjusted for urinary creatinine, age, sex, ethnicity, BMI, and
sampling weights)
1.14 (0.92,1.41)
Mean ± SD MIBP in urine (units not given) in participants with
normal and high BP
Normal BP (n = 2,180)
High BP (n = 309)
13.13 ±22.17
15.71 ±25.15
Trasande et al. (2013b) (United States,
NHANES)
Population: 2,447 children in population-
based survey (NHANES), 2003-2008; ages
8-19 yrs old
Outcome: Systolic BP and diastolic BP
z-score (based on height-, sex-, and age-
normalized values); prehypertension (BP
>90th percentile for age/height/sex); fasting
serum triglycerides (n = 906; high =
>100 mg/dL); nonfasting high density
cholesterol (HDL; n = 2,555;
low = <40 mg/dL)
Exposure: Urine sample, collected at time
of BMI measurement
ILMW phthalates in urine (nM):
Geometric mean
BP <90th percentile 0.817
BP >90th percentile 1.002
ILMW phthalate = sum of MEP, MBP, and
MIBP
Analysis: Logistic regression for pre-
hypertension (BP >90th percentile)
classification; linear regression for systolic
BP and diastolic BP z-score and triglycerides
and HDL as continuous variable; all models
adjusted for variables shown in results
column
Changes in z-score (95% Cl) per unit increase in In-phthalates
(adjusted for sex, caloric intake, television watching,
poverty:income, parental education, serum cotinine, urinary
creatinine, BMI, race/ethnicity, and age)
Systolic BP
Diastolic BP
Triglycerides
HDL
ILMW phthalates
0.03 (-0.02, 0.07)
0.02 (-0.04, 0.07)
-0.22 (-4.40, 0.07)
0.13 (-0.60, 0.85)
MIBP
0.03 (-0.02, 0.08)
-0.02 (-0.09, 0.04)
not reported
not reported
OR (95% Cl) for BP >90th percentile per unit increase in In-
phthalates
BP >90th percentile
High triglycerides
Low HDL
ILMW phthalates
1.19 (0.96, 1.47)
0.85 (0.71, 1.01)
1.00(0.87,1.15)
MIBP
1.00 (0.74, 1.35)
not reported
not reported
Interactions with covariates examined in supplemental analyses;
stratified analyses showed no statistically significant associations
between ILMW phthalates and systolic BP for gender, age,
race/ethnicity, cotinine level, or BMI
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Olsen et al. (2012) (Sweden)
Population: 1,016 (507 men, 509 women),
from population-based cohort (Prospective
Investigation of Vasculature in Uppsala
Seniors study), 2001-2003; age 70 yrs at
enrollment
Outcome: Blood pressure measured at
study visit; fasting serum sample for LDL
and HDL cholesterol, and triglycerides;
Framingham risk score
Exposure: Serum sample, collected at time
of examination; results not shown
Analysis: Linear regression, adjusted for
the variables shown in results column
Lind and Lind (2011) (Sweden)
Population: 1,016 (507 men, 509 women),
from population-based cohort (Prospective
Investigation of Vasculature in Uppsala
Seniors study), 2001-2003; age 70 yrs at
enrollment
Outcome: Carotid artery intima media
thickness (IMT); grey scale media of the
intima media complex (IM-GSM); plaque in
carotid artery
Exposure: Serum sample (fasting),
collected at time of clinical assessment
MIBP in serum (ng/mL):
Median 75th percentile
13.5 29.3
Analysis: Linear regression for continuous
outcomes (IMT, IM-GSM) and ordinal
logistic regression for number of carotid
arteries with plaques (0, 1, 2), adjusted for
variables shown in results column
Results
Regression coefficient for change in outcome per unit increase in In-
MIBP (adjusted for sex, smoking, diabetes and the other variables in
the table; model for Framingham Risk Score only adjusted for sex)
/D rc ci \
(P [SE])
LDL 0.044 (-0.01, 0.09)
HDL 0.017 (-0.01, 0.09)
Triglycerides -0.009 (-0.03, 0.01)
Systolic BP -0.05 (-1.28, 1.18)
Diastolic BP 0.35 (-0.20, 0.90)
Framingham risk score 0.13 (-0.05, 0.31)
Median IMT by quintile of MIBP (adjusted for sex, BMI, fasting
blood glucose, systolic BP, diastolic BP, HDL and LDL cholesterol,
triglycerides, smoking, antihypertensive treatment, statin use)
MIBP
quintile IMT IM-GSM
Median IM-
Median IMT (p-value) GSM (p-value)
1 (low) 0.87 (referent) 80 Referent
2 0.89 (0.91) 72 (0.0001)
3 0.86 (0.13) 68 (0.0001)
4 0.89 (0.91) 69 (0.0001)
5 (high) 0.85 (0.074) 102 (0.0001)
Regression coefficient (P [p-value]) per unit increase in serum MIBP
(adjusted for sex, BMI, fasting blood glucose, systolic BP, diastolic
BP, HDL and LDL cholesterol, triglycerides, smoking,
antihypertensive treatment, statin use)
IMT -0.0045(0.14)
IM-GSM 5.5 (O.OOOlp
OR for presence of plaques and median value of plaque GSM by
quintile of MIBP (adjusted for sex, BMI, fasting blood glucose,
systolic BP, diastolic BP, HDL and LDL cholesterol, triglycerides,
smoking, antihypertensive treatment, statin use)
MIBP
quintile Plaque prevalence Plaque GSM
OR (p-value) Median (p-value)
1 (low) 1.0 (referent) 65 (referent)
2 0.70 (0.059) 69 (0.37)
3 0.74 (0.17) 59 (0.11)
4 1.00 (0.78) 62 (0.074)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
5 (high) 0.64 (0.011) 99 (0.0001)
OR or regression coefficient per unit increase in serum MIBP
Plaque
prevalence
Plaque GSM
OR (95% Cl)
(p-value)
0.88 (0.79, 0.98)
8.0 (0.0001)
The regression models did not show evidence of interaction by
gender, except for IMT (interaction term p-value = 0.030).
1
2
3
4
BP = blood pressure; HDL= high-density lipoprotein; IM-GSM = grey scale media of the intima media complex;
IMT = intima media thickness; LDL = low-density lipoprotein
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Table 3-16. Evidence pertaining to DIBP and cancer in humans
Reference and study design
Results
Geometric mean (95% Cl) DBP in urine (u.g/g Cr), all subjects
and by menopausal status
Lopez-Carrillo et al. (2010) (Mexico)
Population: 233 incident cases, 221 population
controls matched by age and residency, >18 yrs
of age, >1 yr in study area, 2007-2008; mean
age 53 yrs; participation rates: 94.8% of cases
and 99.5% of controls
Outcome: Histologically-confirmed breast
cancer
Exposure: Urine sample (for cases, urine
collected on average 2 mo after diagnosis, but
before treatment)
MIBP in urine, controls:
Geometric mean
Cr-adjusted (u.g/g Cr) 8.85
Analysis: Logistic regression, adjusting for
variables shown in results column
All
Pre-menopause
Post-menopause
Controls
8.85 (7.95, 9.84)
9.99 (8.42, 11.85)
8.32 (7.27, 9.52)
Cases
7.81(6.93,8.81)
8.31 (6.85, 10.09)
7.53 (6.45, 8.78)
OR (95% Cl) for breast cancer, by tertile of MIBP (adjusted for
current age, age at menarche, parity, menopausal status, and
other phthalate metabolites)
MIBP tertile
(Hg/g Cr)
1 (0.23-7.44)
2 (7.45-12.07)
3 (12.08-86.22)
(trend p)
1.0 (referent)
0.59 (0.35, 0.98)
0.73 (0.43, 1.24)
(0.365)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 3.3. Experimental Studies
2 3.3.1. Developmental Effects
3 Table 3-17. Evidence pertaining to developmental effects in animals following
4 oral exposure to DIBP
Reference and study design
Results3
Fetal survival
See Table 3-19
Fetal growth
BASF (2007)
Rat (Wistar); 22-23 dams/group
0, 88, 363, 942 mg/kg-day
Diet
CDs 6-20 (GD 20 c-section)
Borch et al. (2006)
Rat (Wistar); 11-12 dams/group
0, 600 mg/kg-day
Gavage
GDs 7-19 (GD 19 c-section) or
G Ds 7-20/2 1(GD 20/21
c-section); 5-6 dams/group per
time point
Saillenfait et al. (2006)
Rat (Sprague-Dawley);
20-22 dams/group
0, 250, 500, 750, 1,000 mg/kg-day
Gavage
GDs 6-20
(GD 21 c-section)
Fetal body weight (percent
Doses
MBW
FBW
Fetal body weight (percent
Doses (M)
BW(GD19)
(data presented in graphb)
B W (GD 20/21)
(data presented in graphb)
Doses (F)
BW(GD19)
(data presented in graphb)
B W (GD 20/21)
(data presented in graphb)
change compared to control)
0 88
0% -3%
0% -3%
change compared to control)
0
0%
0%
0
0%
0%
363 942
-3% -5%**
-3% -6%**
600
-27%*
-12%
600
-28%*
12%
Fetal body weight (mean percent change compared to control)
Doses
M and F (all fetuses) BW
MBW
FBW
0 250 500
0% 0% -7%**
0% 0% -6%*
0% -1% -8%**
750 1,000
-17%** -24%**
-17%** -25%**
-18%** -26%**
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Postnatal survival
Saillenfait et al. (2008)
Rat (Sprague-Dawley);
11-14 dams/group
0, 125, 250, 500, 625 mg/kg-day
GDs 12-21 (dams allowed to
deliver)
Pup survival (percent change compared to control [litter means])
Doses 0 125 250 500
percentage pup survival 0% -1% -1% -1%
PNDs 1-4
percentage pup survival 0% 2% 5% 3%
PNDs 4-21
625
-7%
5%
Postnatal and adult growth
Eastman Kodak (1954)
Rat (no strain designation);
5 male and 5 females/group
0, 0.1, 1, 5% DIBP (0, 97, 1,000,
7,800 mg/kg-day for males; 0,
110, 1,100, 6,400 mg/kg-day for
females)0
Diet
Weaning to 8 weeks
post-weaning
Body weight gain (percent change compared to control)
Doses (M) 0 97 1,000
BW gain (weaning to 0% 3% -2%
4 weeks post-weaning)
Doses (F) 0 110 1,100
BW gain (weaning to 0% -9% 1%
4 weeks post-weaning)
7,800
-61%
6,400
-34%
Body weight (percent change compared to control)
Doses (M) 0 97 1,000
4 weeks post-weaning 0% 2% -1%
BW
Doses (F) 0 110 1,100
4 weeks post-weaning 0% -5% 1%
BW
7,800
-41%
6,400
-19%
Body weight gain (percent change compared to control)
Doses (M) 0 97 1,000
BW gain (weaning to 0% 3% -3%
8 weeks post-weaning)
Doses (F) 0 110 1,100
BW gain (weaning to 0% -11% 0%
8 weeks post-weaning)
7,800
-58%
6,400
-34%
Body weight (percent change compared to control)
Doses (M) 0 97 1,000
8 weeks post-weaning 0% 2% -3%
BW
Doses (F) 0 110 1,100
8 weeks post-weaning 0% -7% 0%
BW
Note: Statistical analysis not reported in study.
7,800
-44%
6,400
-22%
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Hazleton Laboratories (1992,
1987): NIOSH (1983)
Mouse (CD-I); 50 control
females; 10 females/treated
group (6.5-9 weeks old)
0, 1,000, 1,795, 3,225, 5,790,
10,400 mg/kg-day
Gavage
8 days
Oishi and Hiraga (1980d)
MIBP
Rat (Wistar) (JCL);
10 males/group
0, 2% in diet (0,
l,100mg/kg-day)d
Diet
1 week
Oishi and Hiraga (1980a)
Mouse (JCLICR); 10 males/group
0, 2%(0,2,100mg/kg-day)d
Diet
1 week
Saillenfait et al. (2008)
Rat (Sprague-Dawley);
11-14 dams/group
0, 125, 250, 500, 625 mg/kg-day
Gavage
Results3
Body weight change (g)
Doses (F) 0
BW change 0
(days 1-8 of study)
1,000 1,795 3,225
010
5,790 10,400
1 1
Body weight change (g)
Doses (F) 0
BW change (days 1
1-12 of study)
1,000 1,795 3,225
110
5,790 10,400
1 1
Body weight change (g)
Doses (F) 0
1,000 1,795 3,225
BW change (days 1111
1-16 of study)
Note: Statistical analysis not reported in study.
Body weight and weight gain
Doses (M)
5,790 10,400
1 1
(percent change compared to control)
0
B Wat 6 weeks 0%
B W gain (5-6 weeks* ) 0%
Note: Statistical analysis was not performed on BW gain.
Body weight and weight gain
Doses (M)
B W at 6 weeks
BW gain (5-6 weeks6)
1,100
-10%*
-31%
(percent change compared to control)
0
0%
0%
2,100
-13%*
-54%
Note: Statistical analysis was not performed on BW gain.
Body weight (percent change compared to control [litter means])
Doses 0
M postnatal (PND 1) 0%
BW
M postnatal (PND 21) 0%
BW
F postnatal (PND 21) 0%
BW
125 250
-1% -2%
-1% -3%
-3% -5%
500 625
-2% -10%**
-6% -10%*
-3% -10%
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
GDs 12-21 (dams allowed to
deliver)
University of Rochester (1953)
Rat (Albino; no other strain
designation); 5 males/group
0, 0.01, 0.1, 1, 2, 5% (0, 15, 140,
1,400, 3,000, 8,900 mg/kg-day)f
Diet
Weaning to 1 month
post-weaning
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and
5 females/group
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for
females)h
Diet
Weaning to 4 months
post-weaning
Results3
M BW at day of PPS 0% -8%* -5%* 7%
M adult (PNDs 77-84) 0% -6% -4% -7%*
BW
2%
-9%**
Body weight gain (percent change compared to control)
Doses (M) 0 15 140 1,400 3,000
BWgaine(weaningto 0% -22% -19% -22% -27%
1 month post-weaning)
Note: Statistical analysis was not performed on BW gain.
8,900
-51%
Body weight (percent change compared to control)
Doses (M) 0 15 140 1,400 3,000
1 month post-weaning 0% -16% -14% -16% -20%
BW
Note: Statistical analysis not reported in study.
8,900
-38%
Body weight gain (percent change compared to control)
Doses (M) 0 15 140 1,400 3,000
BW gain (weaning to 0% -20% -18% -21% -27%
PND 49 [-after PPS^])
Note: Statistical analysis was not performed on BW gain.
8,900
-49%
Body weight (percent change compared to control)
Doses (M) 0 15 140 1,400 3,000
PND 49 Rafter PPSg) 0% -14% -13% -15% -19%
BW
Note: Statistical analysis not reported in study.
8,900
-35%
Body weight gain (percent change compared to control)
Doses (M) 0 65 710
BW gain (weaning to 0% 5% -6%
1 month
post-weaning)8
Doses (F) 0 82 770
BW gain (weaning to 0% -7% 3%
1 month
post-weaning)8
Note: Statistical analysis was not performed on BW gain.
5,800
-60%
4,700
-27%
Body weight (percent change compared to control)
Doses (M) 0 65 710
BW at 1 month 0% 4% -4%
post-weaning
5,800
-42%
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Doses (F) 0 82 770
BW at 1 month 0% -4% 2%
post-weaning
Note: Statistical analysis not reported in study.
4,700
-16%
Body weight gain (percent change compared to control)
Doses (M) 0 65 710
BW gain (weaning to 0% 3% -2%
PND 49 [-after PPS^]f
Doses (F) 0* 82 770
BW gain (weaning to 0% -8% 3%
PND 49 [-after V(y]e
Note: Statistical analysis was not performed on BW gain.
5,800
-61%
4,700
-33%
Body weight (percent change compared to control)
Doses (M) 0 65 710
BW at PND 49 (-after 0% 2% -1%
PPSQ)
Doses (F) 0 82 770
BW at PND 49 (-after 0% -4% 1%
VCft)
Note: Statistical analysis not reported in study.
5,800
-41%
4,700
-18%
Body weight gain (percent change compared to control)
Doses (M) 0 65 710
BW gain (weaning to 0% 5% -11%
4 months
post-weaning)
Doses (F) Oy 82 770
BW gain (weaning to 0% -1% 10%
4 months
post-weaning)
Note: Statistical analysis was not performed on BW gain.
5,800
-53%
4,700
-19%
Body weight (percent change compared to control)
Doses (M) 0 65 710
BW at 4 months 0% 4% -9%
post-weaning
Doses (F) 0 82 770
BW at 4 months 0% -1% 7%
post-weaning
Note: Statistical analysis not reported in study.
5,800
-43%
4,700
-13%
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Fetal morphological development
Saillenfait et al. (2006)
Rat (Sprague-Dawley) rats;
20-22 dams/group
0, 250, 500, 750, 1,000 mg/kg-day
Gavage
CDs 6-20 (GD 21 c-section)
Malformations
External malformations (incidence; number of affected fetuses [litters])
Doses 0 250 500
total fetuses (litters) 281 276 237
examined for external (22) (21) (21)
malformations
anasarca 00 0
exophthalmos 00 0
(unilateral) and
absence of eyelids
(bilateral)
exencephaly 00 0
meningoencephalocele 00 0
microstomia 00 0
ectopia cordis 00 0
omphalocele 00 0
750 1,000
212 111
(21) (18)
0 1(1)
1(1) 0
2(2) 0
3 (3) 3 (2)
0 1(1)
0 1(1)
0 1(1)
Combined total with external malformations (incidence [percent])
Doses 0 250 500
total number (%) 000
fetuses with external
malformations
total number (%) litters 000
with external
malformations
mean % fetuses with 0% 0% 0%
external
m a If arm ations/litter
750 1,000
5 (2%)* 6 (5%)**
4 (19%) 4 (22%)
2% 4%
Visceral malformations (incidence; number of affected fetuses [litters])
Doses 0 250 500
total fetuses (litters) 141 138 119
examined for visceral (22) (21) (21)
malformations
anophthalmia, uni- or 0 0 0
bilateral
aorta and/or 000
pulmonary artery
transposed
diaphragmatic hernia 0 2(1) 2 (2)
kidney and ureter, 000
absent, uni- or bilateral
750 1,000
106 56
(21) (18)
6 (4) 4 (3)
6 (5) 3 (3)
2 (2) 1 (1)
KD 3(3)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
kidney, small, uni- or
bilateral
0
0
Combined total with visceral malformations
Doses
total number (%)
fetuses with visceral
malformations
total number (%) litters
with visceral
malformations
mean % fetuses with
visceral
malformations/litter
Skeletal malformations
Doses
total number of fetuses
(litters) examined for
skeletal malformations
mandible, small
stern ebrae, fused
sternebrae, fused and
scrambled
sternebrae, total
cleft sternum
sternebrae,
checkerboard
ribs, fused
cervical arches, fused
thoracic or lumbar
vertebral arches, fused
thoracic or lumbar
vertebral centra, fused
thoracic or lumbar
centrum, hemicentric
thoracic or lumbar
vertebral centra,
misaligned
0
0
0
0%
250
2 (1%)
1
(5%)
1%
0
(incidence
500
2 (2%)
KD
[percent])
750
13
(12%)**
2 8(38%)**
(10%)
2%
13%*
KD
1,000
10 (18%)**
8(44%)**
16%*
(incidence; number of affected fetuses [litters])
0
140
(22)
0
0
0
0
0
0
0
0
0
0
0
0
250
138
(21)
0
0
0
0
0
0
0
0
0
0
0
0
500
118
(21)
0
0
0
0
KD
0
0
0
1(1)
1(1)
1(1)
2(2)
750
106
(21)
0
7(6)
5(3)
12 (7)*
KD
2(2)
0
3(3)
2(2)
0
4(3)
3(2)
1,000
55
(18)
KD
14(9)**
12(7)
26 (13)**
2(2)
0
2(2)
3(3)
2(2)
4(3)
3(3)
5(4)
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Combined total with skeletal malformations (incidence [percent])
Doses
total number (%)
fetuses with skeletal
malformations
total number (%) litters
with skeletal
malformations
mean % fetuses with
skeletal
malformations/litter
0
0
0
0%
250 500
0 4 (3%)
0 4
(19%)
0% 3%
750
18
(17%)**
11
(52%)**
18%**
1,000
34 (62%)**
15 (83%)**
67%**
Variations
External variations (incidence; number of affected fetuses [litters])
Doses
total fetuses (litters)
examined for external
variations
clubfoot
tail, curly
tail tip, haemorrhage
0
281
(22)
0
0
0
250 500
276 237
(21) (21)
2(1) 0
0 0
1(1) 0
750
212
(21)
0
KD
0
1,000
111
(18)
0
0
0
Visceral variations (incidence; number of affected fetuses [litters])
Doses
total fetuses (litters)
examined for visceral
variations
dilated cerebral
ventricle, slight
dilated renal pelvis
ureter (all)
hydroureter
distended ureter
ovaries, displaced
testis, ectopic
degree of trans-
abdominal testicular
migration (mean)
0
141
(22)
0
1(1)
3(3)
0
3(3)
0
0
2.6
250 500
138 119
(21) (21)
0 0
0 0
0 2(2)
0 0
0 2(2)
0 0
0 3(2)
3.8 13.6**
750
106
(21)
KD
2(2)
10(8)
4(4)
6(5)
5(4)
30(16)**
42.2**
1,000
56
(18)
0
5(4)
12 (8)*
6(5)
6(4)
2(2)
30(16)**
58.1**
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
BASF (2007V
Rat (Wistar); 25 dams/group
0, 88, 363, 942 mg/kg-day
Diet
CDs 6-20 (GD 20 c-section)
Results3
Skeletal variations (incidence; number of affected fetuses [litters])
Doses
total fetuses (litters)
examined for skeletal
variations
parietals or
supraoccipital,
incomplete ossification
hyoid, absent or
incomplete ossification
stern ebrae, fused,
1st and 2nd only
stern ebrae, bipartite
stern ebrae, incomplete
ossification
ribs, cervical,
rudimentary
ribs, 14th, any
supernumerary
ribs, 14th, long
supernumerary
ribs, short or reduced
ossification (unilateral)
thoracic or lumbar
vertebral centra,
incomplete ossification
vertebrae, 27 presacral
Note: A single fetus may
variations.
0 250 500 750
140 138 118 106
(22) (21) (21) (21)
0 0 0 3(2)
0 0 0 1(1)
1(1) 0 8(4) 29(11)**
0 1(1) 2(2) 7(5)
0 1(1) 5(5) 9(6)
0 0 2(2) 12(9)*
23(11) 32(14) 42(18) 72(20)**
1(1) 1(1) 2(2) 15(9)*
0 0 0 1(1)
3(2) 8(6) 7(7) 18(14)**
000 0
1,000
55
(18)
1(1)
8(7)
5(4)
4(4)
6(5)
9(6)
52 (18)**
9(9)*
KD
16 (8)*
2(2)
be represented more than once in the individual
Malformations
External malformations (incidence; number of affected fetuses [litters])
Doses
total fetuses (litters)
examined for external
malformations and
variations
malformed head
anophthalnia
0 88 363
208 (23) 197 (22) 182 (22)
0 1(1) 0
0 1(1) 0
942
211(23)
0
0
Combined total with external malformations (incidence [percent])
Doses
fetuses
0 88 363
0 1 (1%) 0
942
0
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
litters 0 1 (5%)
0 0
Combined total with soft tissue malformations
No fetuses affected at any dose
Skeletal malformations (incidence; number of affected fetuses [litters])
Doses 0 88
total fetuses (litters) 109 (23) 101 (22)
examined for skeletal
malformations/
variations
severely malformed 0 1(1)
skull bones
shortened scapula 0 0
(cartilage present)
malpositioned and 1(1) 1(1)
bipartite sternebral
(unchanged cartilage)
branched rib 1(1) 0
misshapen humerus 0 2 (2)
shortened humerus 0 0
363 942
97 (22) 110 (23)
0 0
1(1) 0
0 0
0 0
0 2(2)
2(1) 0
Combined total with skeletal malformations (incidence [percent])
fetuses 1 (1%) 4 (4%)
litters 1 (4%) 4 (18%)
2 (2%) 2 (2%)
1 (5%) 2 (9%)
Variations
Combined total with external variations
No fetuses affected at any dose
Soft tissue variations (incidence; number of affected fetuses [litters])
Doses 0 88
total fetuses (litters) 99 (23) 96 (22)
examined for external
soft tissue
malformations and
variations
dilated renal pelvis 10 (7) 7 (5)
dilated ureter 2 (2) 2 (1)
363 942
85 (22) 101 (23)
9 (8) 7 (5)
1 (1) 1 (1)
Combined total with soft tissue variations (incidence (percent))
Doses 0 88
fetuses 10 (10%) 7 (7%)
litters 7 (30%) 5 (23%)
363 942
9 (11%) 7 (7%)
8 (36%) 5 (22%)
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Skeletal variations (incidence; number of affected fetuses [litters])
Doses
supraoccipital hole(s)
incomplete ossification
of basisphenoid
incomplete ossification
of interparietal
(unchanged cartilage)
incomplete ossification
of parietal (unchanged
cartilage)
incomplete ossification
of supraoccipital
(unchanged cartilage)
incomplete ossification
of skull (unchanged
cartilage)
incomplete ossification
ofhyoid (cartilage
present)
incomplete ossification
of cervical arch
(cartilage present)
incomplete ossification
of thoracic centrum
(unchanged cartilage)
dumbbell ossification of
thoracic centrum
(unchanged cartilage)
dumbbell ossification of
thoracic centrum
(dumbbell-shaped
cartilage of centrum)
bipartite ossification of
thoracic centrum
(dumbbell-shaped
cartilage of centrum)
supernumerary thoracic
vertebra
unossified thoracic
centrum (dumbbell-
shaped cartilage of
centrum)
0
31(15)
7(3)
24 (15)
16 (10)
9(7)
5(4)
KD
KD
0
4(3)
14(9)
2(2)
KD
KD
88
23 (13)
2(2)
11(7)
13(9)
14 (11)
2(1)
0
0
3(3)
3(3)
13(9)
2(2)
2(1)
0
363
15 (12)
6(5)
22 (12)
22 (13)
14 (12)
7(4)
KD
0
3(3)
2(2)
13 (13)
0
KD
0
942
38 (19)
8(4)
13(9)
13(7)
13(6)
2(2)
KD
0
0
9(7)
17 (14)
KD
3(2)
0
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
dumbbell ossification of
lumbar centrum
(dumbbell-shaped
cartilage of centrum)
incomplete ossification
of lumbar arch
(cartilage present)
misshapen sacral
vertebra
fused sacral centrum
and arch (unchanged
cartilage)
incomplete ossification
of sacral arch (cartilage
present)
unossified sternebra
(unchanged cartilage)
incomplete ossification
of sternebra
(unchanged cartilage)
misshapen sternebral
(unchanged cartilage)
unilateral ossification of
sternebra (unchanged
cartilage)
extra sternebral
ossification site
(unchanged cartilage)
bipartite ossification of
sternebral (unchanged
cartilage)
supernumerary rib
(14th) (cartilage
present)
supernumerary rib
(14th) (cartilage not
present)
cervical rib (cartilage
present)
cervical rib (cartilage
not present)
wavy rib
incomplete ossification
ofpubis (cartilage
present)
KD
0
KD
3(2)
5(4)
4(4)
42 (17)
32 (19)
0
0
KD
6(5)
50 (15)
0
5(5)
6(5)
0
0
0
1(1)
7(3)
2(2)
11(7)
44 (17)
26 (17)
0
0
0
KD
33 (15)
KD
5(4)
2(2)
KD
0
KD
KD
5(4)
0
KD
44 (19)
20 (12)
0
KD
0
6(6)
40 (17)
0
5(3)
10(4)
0
0
0
4(4)
5(3)
0
6(4)
75(22*)
28 (16)
4(4)
0
2(2)
6(5)
62(22*)
0
4(4)
9(9)
0
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Combined total with skeletal variations (incidence (percent))
Doses 0 88 363
fetuses 101 87(86%) 85(88%)
(93%)
litters 23 22(100%) 22(100%)
(100%)
942
107 (97%)
23 (100%)
Unclassified observations
Unclassified external observations (incidence; number of affected fetuses
[litters])
Doses 0 88 363
total fetuses (litters) 208(23) 197(22) 182(22)
examined for
unclassified
observations
discolored amniotic 00 0
fluid
942
211(23)
KD
Combined total with external unclassified observations (incidence [percent])
Doses 0 88 363
fetuses 000
litters 000
942
1 (1%)
1 (4%)
Combined total with unclassified soft tissue observations
No fetuses affected at any dose
Skeletal unclassified cartilage observations (incidence; number of affected
fetuses [litters])
Doses 0 88 363
total fetuses (litters) 109(23) 101(22) 97(22)
examined for skeletal
unclassified
observations
notched cartilage 2(2) 0 0
between basiphenoid
and basioccipital
fused cervical arch 1(1) 0 0
cartilage
dumbbell-shaped 0 1(1) 0
cartilage of cervical
centrum
hole in processus 0 1(1) 0
coracoideus
bipartite processus 36(14) 30(15) 30(15)
xiphoideus
942
110 (23)
KD
0
0
0
40 (14)
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Borch et al. (2006)
Rat (Wistar); 11-12 dams/group
0, 600 mg/kg-day
Gavage
GDs 7-19 (GD 19 c-section) or
CDs 7-20/21 (GD 20/21
c-section); 5-6 dams/group per
time point
Results3
notched manubrium
fused rib cartilage
branched rib cartilage
5 (5) 7 (5)
0 0
0 0
3 (3) 1 (1)
1(1) 0
1(1) 0
Combined total with skeletal unclassified cartilage observations (incidence
[percent])
Doses
fetuses
litters
AGO change in females
Doses
AGO at GD 19 (data
shown in graphb)
AGO at GD 20/21) (data
shown in graphb)
0 88
39 (36%) 35 (35%)
16 (70%) 17 (77%)
363 942
32 (33%) 41 (37%)
16 (73%) 15 (65%)
(percent change compared to control)
0
0%
0%
600
16%
26%*
AGD/cubic root of BW change in females (percent change compared to
control)
Doses
at GD 19 (data shown
in graphb)
at GD 20/21 (data
shown in graphb)
0
0%
0%
600
27%**
27%*
1
2 Response is % control (indicated by %) or in cases when % control was not possible to present (e.g., if control
3 value was 0), response levels are presented. Equation used to calculate percent change compared to control:
4 treated value - control value x 100
5 control value
6 bGrablt Software used to estimate % control from graph.
7 °Dose conversions were performed using this information: For the Eastman Kodak (1954) study, average BWs were
8 183,186,180, and 115 g for males, and 132,126,133, and 110 g for females at 0, 0.1,1.0, and 5.0%, respectively.
9 Reference values for food consumption of 0.018 and 0.014 kg/day for male and female rats of an unspecified
10 species (U.S. EPA, 1988) were used.
11 dDose conversions were performed using this information: In Oishi and Hiraga (1980d), average BWs for rats and
12 mice in these studies were 145 and 25 g, respectively, and the default food consumption rates of 0.008 kg/day for
13 male Wistar rats and 0.0025 kg/day for male B6C3Fi mice (U.S. EPA, 1988) were applied. In Oishi and Hiraga
14 (1980a), average BWs over the week-long studies were 132 and 24 g for rats and mice, respectively, and the
15 default food consumption rates of 0.008 kg/day for male Wistar rats and 0.0025 kg/day for male B6C3Fi mice
16 were applied (U.S. EPA, 1988).
17 eChange in body weight was calculated by EPA.
18 fDose conversions were performed using this information: For University of Rochester (1953), average BWs were
19 139,124,127,127,121, and 101 g at 0, 0.01, 0.1,1.0, 2.0, and 5.0%, respectively). Reference values for food
20 consumption of 0.018 and 0.014 kg/day for male and female rats of an unspecified strain (U.S. EPA, 1988) were
21 used.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 gPND 49 was selected among the periodic weight measurement ages to present in this table because it
2 corresponds to the age when VO and PPS, the developmental markers of puberty, would be expected to have
3 completed in the male and female rat.
4 hDose conversions were performed by EPA using this information: For University of Rochester (1954), average BWs
5 were 269, 277, 252, and 155 g for males, and 178,170,182, and 148 g for females at 0, 0.1,1.0, and 5.0%,
6 respectively. Reference values for food consumption of 0.018 and 0.014 kg/day for male and female rats of an
7 unspecified species (U.S. EPA, 1988) were used.
8 'Male reproductive organs were not evaluated in the BASF study.
9
10 * = Statistically significant difference at p < 0.05 from control value, as reported by study authors; ** = Statistically
11 significant difference at p < 0.01 from control value, as reported by study authors; *** = Statistically significant
12 difference at p < 0.001 from control value, as reported by study authors; BW = body weight; GD = gestation day;
13 PND = postnatal day; PPS = preputial separation; VO = vaginal opening
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
O
(5
OJ
Q.
s
Q
fetal body weight
BASF, 2007; rat, male
fetal body weight
BASF, 2007; rat, female
fetal body weight; GD 19
Borch et al., 2006; rat, male
fetal body weight; GD 19
Borch et al., 2006; rat, female
fetal body weight; GD 20/21
Borch et al., 2006; rat, male
fetal body weight; GD 20/21
Borch et al., 2006; rat, female
fetal body weight
Saillenfait et al., 2006; rat, male
fetal body weight
Saillenfait et al., 2006; rat, female
ro pup survival; PND 1-4
'> Saillenfait et al., 2008; rat, male & female
i:
i/l
£ pup survival; PND 4-21
S. Saillenfait et al., 2008; rat, male & female
• significantly changed
O not significantly changed
G.
•
•
0
O
10 100
Doses (mg/kg-day)
1000
2 Figure 3-1. Exposure-response array of effects on developmental growth and
3 survival following developmental oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Eastman Kodak, 1954; 4 wk; rat, male
Eastman Kodak, 1954; 4 wk; rat, femah
Eastman Kodak, 1954; 8 wk; rat, mal<
Eastman Kodak, 1954; 8 wk; rat, femahr
NIOSH, 1983; day 1-8; mouse, female
NIOSH, 1983; day 1-12; mouse, female
NIOSH, 1983; day 1-16; mouse, female
Oishi and Hiraga, 1980d; 6 wk; rat
Oishi and Hiraga, 1980d; 5-6 wk; rat
Oishi and Hiraga, 1980a; 6 wk; mice
Oishi and Hiraga, 1980a; 5-6 wk; mice
Saillenfait et al., 2008; PND 1, PND 21; rat, male
Saillenfait et al., 2008; pps; rat, male
Saillenfait et al., 2008; PND 21; rat, female
Saillenfait et al., 2008; PND 77-84; rat, male
University of Rochester, 1953; 1 mo; rat, male
University of Rochester, 1953; PND 49; rat, male
University of Rochester, 1954; 1 mo; rat, male
University of Rochester, 1954; 1 mo; rat, female
University of Rochester, 1954; PND 49; rat, male
University of Rochester, 1954; PND 49; rat, female
University of Rochester, 1954; 4 mo; rat, male
University of Rochester, 1954; 4 mo; rat, female
10
3
C
• significantly changed
O not significantly changed
D statistical analysis not
reported
^
^
t
3
3
3
3
3
3
3
[
[
[
0/-\ i^Y^i
•
3
•
0
100
Doses (mg/kg-day)
1000
10000
100000
2 Figure 3-2. Exposure-response array of effects on postnatal and adult body weight
3 following developmental oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
total malformations
Saillenfait et al., 2006; rat, male & female
total variations
Saillenfait et al., 2006; rat, male & female
total malformations
BASF, 2007; rat, male & female
total variations
BASF, 2007; rat, male & female
total unclassfied observations
BASF, 2007; rat, male & female
total malformations
Saillenfait et al., 2006; rat, male & female
variation: ureter (all)
Saillenfait et al., 20061; rat, male & female
varaiation trans abdominal testicular migration
Saillenfait et al., 20061; rat, male
variation: testis, etopic
Saillenfait et al., 20061; rat, male
total malformations
Sailenfait et al., 2006; rat, male & female
variation: sternebrae, fused
Saillenfait et al., 20061; rat, male & female
variation: ribs, cervical, rudimentary
Saillenfait et al., 20061; rat, male & female
variation: ribs, 14th, any supernumentary
Saiilenfait et al., 20061; rat, male & female
variation: ribs, 14th, long supernumentary
Saiilenfait et al., 20061; rat, male & female
variation: thoracic/ lumbar
Saillenfait et al., 20061; rat, male & female
total malformations
BASF, 2007; rat, male & female
total variations
BASF, 2007; rat, male & female
total unclassified cartilage observations
BASF, 2007; rat, male & female
total malformations
BASF, 2007; rat, male & female
total variations
BASF, 2007; rat, male & female
total unclassified observations
BASF, 2007; rat, male & female
AGO change; GD 19
Borch et al., 2006 (female); rat, male & female
AGO change; GD 20/21
Borch et al., 2006 (female); rat, male & female
1
1 Individual variation incidences shown here were included because statistical
significance was reported. Incidence for other individual malformations and
variations are shown in Table 3-1 (Saillenfait et al., 2006).
_L
• significantly changed
O not significantly changed
X-N X">
O
•
10
Doses (mg/kg-day)
100
1000
2
3
Figure 3-3. Exposure-response array of effects on fetal morphological
developmental following developmental oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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2
3
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.3.2. Male Reproductive Effects
Table 3-18. Evidence pertaining to male reproductive effects in animals
following oral exposure to DIBP
Reference and study design
Results3
Morphological development (assessed in fetal or postnatal development or adults)
Borch et al. (2006)
Rat (Wistar); 11-12 dams/group
0, 600 mg/kg-day
Gavage
CDs 7-19 (GD 19 section) or CDs
7-20/21 (GD 20/21 c-section); 5-6
dams/group per time point
AGO change in fetus (percent
Doses
At GD 19 (data shown
in graphb)
At GD 20/21 (data
shown in graphb)
change compared to control)
0 600
0% -15%**
0% -11%**
AGD/cubic root BW change in fetus (percent change compared to
control)
Doses (M)
GD 19 (data shown in
graph")
GD 20/21 (data
shown in graphb)
0 600
0% -5%
0% -9%**
Histologic lesions in fetal testis
Borch et al. (2006)
Rat (Wistar); 11-12 dams/group
0, 600 mg/kg-day
Gavage
CDs 7-19 (GD 19 section) or CDs
7-20/21 (GD 20/21 c-section); 5-6
dams/group per time point;
1-3 males/litter
Testicular histological changes (incidence; percentage incidence in
fetuses)
Doses
Fetuses GD 19
clustering of small
Leydig cells
Sertoli cell
vacuolization
central localization of
gonocytes
multinucleated
gonocytes
Fetuses GD 20/21
0 600
2/13 9/9***
15% 100%***
0/13 1/9
0% 11%
0/13 2/9
0% 22%
1/13 0/9
8% 0%
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
clustering of small
Ley dig cells
Sertoli cell
vacuolization
central localization of
gonocytes
multinucleated
gonocytes
0/10 13/15***
0% 87%***
0/10 14/16***
0% 88%***
0/10 14/16***
0% 88%***
1/10 10/16*
10% 63%*
Fetal testicular testosterone production
Borch et al. (2006)
Rat (Wistar); 11-12 dams/group
0, 600 mg/kg-day
Gavage
GDs 7-19 (GD 19 c-section) or
CDs 7-20/21 (GD 20/21 c-section);
5-6 dams/group per time point
Hannasetal. (2011)
Rat (Harlan Sprague-Dawley);
3 dams/group; 3 males/dam
0, 100, 300, 600, 900 mg/kg-day
Gavage
GDs 14-18
Testicular testosterone (T)
control)
Doses
T content (GD 19 M)
(data shown in
graphb)
T content (GD 20/21
M) (data shown in
graph")
Testicular testosterone (T)
to control)
Doses
testicular T production
ex vivo (GD 19 M)
(data shown in
graph")
testicular T production
ex vivo (GD 20/21 M)
(data shown in
graph")
content (percentage change compared to
0 600
0% -70%
0% -90%***
production (percentage change compared
0 600
0% -21%
0% -96%***
Fetal testicular testosterone (T) production (percentage change
compared to control)
Doses 0
Tproduction 0%
100 300 600 900
10% -56%** -80%** -87%**
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Hannas et al. (2012)
Rat (Sprague-Dawley); 3 dams/group;
3 males/dam
0, 500 mg/kg-day
Gavage
CDs 14-18
Howdeshell et al. (2008)
Rat (Sprague-Dawley); 5-8 dams/group;
3 males/dam
0, 100, 300, 600, 900 mg/kg-day
Gavage
GDs 8-18; c-section on GD 18
Results3
Fetal testicular testosterone (T) production (percentage change
compared to control)
Doses 0 500
T production (data 0% -73%**
shown in graphb)
Fetal testicular testosterone (T) production (percentage change
compared to control)
Doses 0 100 300 600 900
T production (litter 0% -5% -40%** -59%** -63%**
mean)
Morphological development assessed in postnatal development and adults
Saillenfait et al. (2008)
Rat (Sprague-Dawley);
11-14 dams/group
0, 125, 250, 500, 625 mg/kg-day
Gavage
GDs 12-21 (dams allowed to deliver)
Postnatal effects (percent change in litter mean compared to control)
Doses 0 125 250 500 625
AGD(PNDl) 0% -4% -11%* -21%** -22%**
ageatPPS 0% -4%* -1% 10%** 6%*
Postnatal effects (incidence; percentage incidence)
Doses 0 125 250 500 625
retained 0/76 0/78 8/96 47/79 56/76
nipples or
areolas at
PNDs 12-14
0% 0% 8% 59% 74%
Note: No statistical analysis was reported by the authors for this
endpoint.
Male adult effects at necropsy (PNDs 77-84 or 112-119; percentage
incidence)
Doses 0 125 250 500 625
retained 0/46 0/40 4/55 24/44 29/38
nipples or
areolas
0% 0% 7% 55% 76%
hypospadias 0/46 0/40 0/55 5/44 22/39
0% 0% 0% 11% 56%
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
exposed os 0/46 0/40 0/55
penis
0% 0% 0%
cleft prepuce 0/46 0/40 0/55
0% 0% 0%
nonscrotal 0/46 0/40 0/55
testis
0% 0% 0%
Note: No statistical analysis was reported by the
endpoint.
4/44
9%
0/44
0%
11/44
25%
11/39
28%
10/39
26%
30/39
77%
authors for this
Histopathologic lesions in adult testis and epididymis
Saillenfait et al. (2008)
Rat (Sprague-Dawley);
11-14 dams/group
0, 125, 250, 500, 625 mg/kg-day
Gavage
GDs 12-21 (dams allowed to deliver)
Adult effects0 (PNDs 77-84; incidence)
Doses 0 125 250
number of males 24(12) 20(10) 28(14)
(litters)
examined
500
22(11)
625
20 (10)
Epididymides (number of males with effect)
Doses 0 125 250
oligospermia 013
azoospermia 013
granulomatous 000
inflammation
500
2
10
4
625
1
18
3
Testes (number of males with effect)
Doses 0 125 250
tubular 227
degeneration-
atrophy/
hypoplasia
tubular necrosis 001
interstitial cell 000
hyperplasia
Note: No statistical analysis was reported by the
endpoints.
500
16
3
1
625
20
5
9
authors for these
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Testes weight
Zhu et al. (2010)
Rat (Sprague-Dawley)
Mouse (C57BI/6N) (number of animals
not specified)
0, 100, 300, 500, 800, 1,000 mg/kg-day
Gavage
7 days
Foster et al. (1982); Foster et al. (1981)
MIBP
Rat (Sprague-Dawley); 6 males/group
0, 800 mg/kg-day
Gavage
6 days
Oishi and Hiraga (1980c)
Rat (JCLWistar); 10 males/treated group;
20 control males
0, 2% (0, 1,200 mg/kg-day)d
Diet
1 week
Oishi and Hiraga (1980d)
MIBP
Rat (JCLWistar); 10 males/group
0, 2% in diet (0, 1,100 mg/kg-day)d
Diet
1 week
Oishi and Hiraga (1980a)
Mouse (JCLICR); 10 males/group
0, 2%(0,2,100mg/kg-day)d
Diet
1 week
Testes weight at
Doses
7 days (percent change compared to control)
0 100 300 500 800 1,000
Rat, absolute 0% -3% -10% -22%*** -32%*** -44%***
weight (data
shown in
graph")
Mouse, 0% 0% 10% 6% 4% -22%**
absolute
weight(data
shown in
graph")
Note: Relative weight not reported by study authors.
Testes weight (percent change compared to control)
Doses
absolute weight
relative weight
0 800
0% -28%***
0% -27%***
Testes weight (percent change compared to control)
Doses
absolute weight
relative weight
0 1,200
0% -37%*
0% -33%*
Testes weight (percent change compared to control)
Doses
absolute weight
relative weight
0 1,100
0% -47%*
0% -40%*
Testes weight (percent change compared to control)
Doses
relative weight
0 2,100
0% 29%*
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Oishi and Hiraga (1980b)
MIBP
Mouse (JCLICR); 10 males/group
0, 2% (0, 2,000 mg/kg-day)d
Diet
1 week
Saillenfait et al. (2008)
Sprague-Dawley rats; 11-14 dams/group
0, 125, 250, 500, 625 mg/kg-day
Gavage
GDs 12-21 (dams allowed to deliver)
Assessed PNDs 77-84 (adults) after in
utero exposure
University of Rochester (1954)
Rat (Albino; no other strain designation);
5 males/group
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day)e
Diet
Weaning to 4 months post-weaning
Results3
Testes weight (percent change compared to control)
Doses
relative weight
0
0%
2,000
45%*
Male reproductive organ weights (percent change compared to
control)
Doses 0
right testis 0%
weight
right 0%
epididymal
weight
left testis 0%
weight
left 0%
epididymal
weight
seminal 0%
vesicles
prostate 0%
Testes weight at 4 months
Doses
absolute weight
relative weight
125 250
1% 0%
-2% -6%
-2% -1%
-4% -8%
1% -6%
-10% -11%*
500 625
-22% -52%**
-22%** -49%**
-13% -59%**
-16%** -49%**
-18%** -33%**
-16%** -30%**
(percent change compared to control)
Oh 65
0% 2%
0% 1%
710 5,800
-1% -70%
12% -45%
Note: Statistical analysis not reported in study.
Seminal vesicle weight
Foster et al. (1982); Foster et al. (1981)
MIBP
Rat (Sprague-Dawley); 6 males/group
0, 800 mg/kg-day
Gavage
6 days
Seminal vesicle weight (percent change compared to control)
Doses
absolute weight
relative weight
0
0%
0%
800
-18%
-11%
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Prostate weight
Foster et al. (1982); Foster et al. (1981)
MIBP
Rat (Sprague-Dawley); 6 males/group
0, 800 mg/kg-day
Gavage
6 days
Prostate weight (percent change compared to control)
Doses
absolute weight
relative weight
0 800
0% -13%
0% 4%
Testosterone concentration in adults
Oishi and Hiraga (1980c)
Rat (JCLWistar); 10 treated males;
20 control males
0, 2% (0, 1,200 mg/kg-day)d
Diet
1 week
Oishi and Hiraga (1980d)
MIBP
Rat (JCLWistar); 10 males/group
0, 2% (0, 1,100 mg/kg-day)d
Diet
1 week
Oishi and Hiraga (1980a)
Mouse (JCLICR); 10 males/group
0, 2%(0,2,100mg/kg-day)d
Diet
1 week
Oishi and Hiraga (1980b)
MIBP
Mouse (JCLICR); 10 males/group
0, 2% (0, 2,000 mg/kg-day)d
Testosterone (T) concentration (percent change compared to control)
Doses
serum T concentration
(data shown in graphb)
testicular T concentration
(data shown in graphb)
0 1,200
0% 19%
0% 158%*
Dihydrotestosterone (DHT) concentration (percent change compared
to control)
Doses
serum DHT concentration
(data shown in graphb)
0 1,200
0% 40%
Testosterone (T) concentration (percent change compared to control)
Doses
serum T concentration
(data shown in graphb)
testicular T concentration
(data shown in graphb)
Testicular testosterone (T)
to control)
Doses
T concentration
Testicular testosterone (T)
to control)
Doses
T concentration
0 1,100
0% 61%*
0% 161%*
concentration (percent change compared
0 2,100
0% 7%
concentration (percent change compared
0 2,000
0% -83%*
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results3
Diet
1 week
Testes histology in adults
Foster et al. (1982): Foster et al. (1981)
MIBP
Rat (Sprague-Dawley); 6 males/group
0, 800 mg/kg-day
Gavage
6 days
Number of animals with atrophy of seminiferous tubules
Doses
0
800
0% atrophic tubules
6/6
0/6
<50% atrophic tubules 0/6 3/6
>50% atrophic tubules 0/6 3/6
The study authors noted marked atrophy of the majority of the
seminiferous tubules with decreased spermatocytes and decreased
spermatogonia (data not shown).
Oishi and Hiraga (1980c)
Rat (JCLWistar); 10 treated males;
20 control males
0, 2% (0,1,212 mg/kg-day)d
Diet
1 week
The testes showed decreased spermatocytes and decreased
spermatogonia compared to control (quantitative results not
provided).
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
aPercent change compared to control = treated value - control value x 100
control value
bGrablt Software used to estimate % change from graph.
cPNDs 112-119 males were also evaluated for these endpoints (see Saillenfait et al., 2008, Table 4).
dDose conversions were performed by EPA using this information: Oishi and Hiraga (1980a, c) average BWs over
the week-long studies were 132 and 24 g for rats and mice, respectively, and the default food consumption rates
of 0.008 kg/day for male weanling Wistar rats and 0.0025 kg/day for male weanling B6C3Fi mice were applied
(U.S. EPA, 1988). Oishi and Hiraga (1980b, d) average BWs for rats and mice in these studies were 145 and 25 g,
respectively, and the default food consumption rates of 0.008 kg/day for male weanling Wistar rats and
0.0025 kg/day for male weanling B6C3Fi mice (U.S. EPA, 1988) were applied. Note that Table 1-6 of U.S. EPA
(1988) listed the default food consumption rate for male weanling Wistar rats as 0.080 kg/day. However, it was
later determined using an equation in Table 1-3 of the document that this value was actually supposed to be
0.008 kg/day.
eDose conversions were performed by EPA using this information: University of Rochester (1954) average BWs
(measured at least once weekly) of the rats were 269, 277, 252, and 155 g for males, and 178,170,182, and 148 g
for females at 0, 0.1,1.0, and 5.0%, respectively; and the default food consumption rates of 0.018 kg/day for male
rats and 0.014 kg/day for female rats (U.S. EPA, 1988) for an unspecified strain in a sub-chronic study were
applied.
* = Statistically significant difference at p < 0.05 from control value, as reported by study authors; ** = Statistically
significant difference at p < 0.01 from control value, as reported by study authors; *** = Statistically significant
difference at p < 0.001 from control value, as reported by study authors.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
o
Q.
leydig cell clusters; GD19
Borch et al., 2006; rat
sertoli cell vacilization; GD19
Borch et al., 2006
multinucleated gonocytes; GD19
Borch et al., 2006; rat
leydig cell clusters; GD20/21
Borch et al., 2006; rat
sertoli cell vacilization; GD20/21
Borch et al., 2006; rat
multinucleated gonocytes; GD 20/21
Borch et al., 2006; rat
nipple retention
Saillenfait et al.,2008; rat
hypospadis
Saillenfait et al., 2008; rat
cleft prepuce
Saillenfait et al.,2008; rat
nonscrotal testis
Saillenfait et al., 2008; rat
exposed os penis
Saillenfait et al.,2008; rat
aligospermia & azoospermia
Saillenfait et al., 2008; rat
granulomatous inflammation
Saillenfait et al., 2008; rat
tubular degeneration & necrosis
Saillenfait et al.,2008; rat
interstitial hyperplasia
Saillenfait et al., 2008; rat
AGO; PND 1
Saillenfait et al.,2008; rat
AGO change; GD 19
Borch et al., 2006; rat
AGO change; GD 20/21
Borch et al., 2006; rat
• significantly changed
O not significantly changed
1
0
Doses (mg/kg-day)
-e-e
-e-e
-e-e
-e-e
-e-e
-e-e
-e-e
100
1000
2
3
Figure 3-4. Exposure-response array of effects on male reproductive
development following developmental oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
o
tt
TesticularT content; GD 19
Borch etal., 2006; rat
TesticularT content; GD 20/21
Borch et al., 2006; rat
T production; GD 19
Borch etal., 2006; rat
T production; GD 20/21
Borch etal., 2006; rat
T production; GD 18
Hannas etal., 2011; rat
T production; GD 18
Hannas etal., 2012; rat
T production; GD 18
Howdeshell et al., 2008; rat
• significantly changed
O not significantly changed
10
100
1000
Doses (mg/kg-day)
2
3
Figure 3-5. Exposure-response array of effects on fetal testosterone (T)
following developmental oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Foster et al., 1981; Foster et al., 1982; rat
Oishi and Hiraga, 1980c; rat
Oishi and Hiraga, 1980d; rat
"Si Oishi and Hiraga, 1980b; mouse
'oj
£ University of Rochester, 1954; rat
Zhu et el., 2010; rat
Zhu et el., 2010; mouse
testis weight
Saillenfait et al., 2008; rat
•gj epididymal weight
| Saillenfait et al., 2008; rat
^ Saillenfait et al., 2008; rat
_c
op
Foster et al., 1981; Foster et al., 1982; rat
_c
•5 Saillenfait etal., 2008; rat
g
OJ
ra
g Foster et al., 1981; Foster et al., 1982; rat
Q_
testicular T concentration
Oishi and Hiraga, 1980c; rat
c
O
'•£ serum DHT concentration
•£ Oishi and Hiraga, 1980c; rat
OJ
u
2 serum & testicular T concentration
£ Oishi and Hiraga, 1980d; rat
o
y testicular T concentration
& Oishi and Hiraga, 1980a; mouse
£
testicular T concentration
Oishi and Hiraga, 1980b; mouse
ao atrophic tubules
J Foster et al., 1981; Foster et al., 1982; rat
ra
o" decreased spermatocytes
~ Oishi and Hiraga, 1980c; rat
,
• significantly changed
O not significantly changed
D statistical analysis not reported
c
\.
B(~\ (~\ f~\l
e/~\ 1^^
o
o
o
9
I
I
o
o
o
10
100
1000
10000
Doses (mg/kg-day)
2
3
Figure 3-6. Exposure-response array of male reproductive effects following
oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy.
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2
3
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.3.3. Female Reproductive Effects
Table 3-19. Evidence pertaining to female reproductive effects in animals
following oral exposure to DIBP
Reference and study design
Results
Maternal weight"
Howdeshell et al. (2008)
Rat (Sprague-Dawley);
5-8 dams/group
0, 100, 300, 600,
900 mg/kg-day
Gavage
CDs 8-18 (GD 18 c-section)
Borch et al. (2006)
Rat (Wistar);
11-12 dams/group
0, 600 mg/kg-day
Gavage
GDs 7-19 (GD 19 c-section) or
G Ds 7-20/2 1(GD 20/21
c-section); 5-6 dams/group per
time-point
BASF (2007)°
Rat (Wistar);
22-23 dams/group
0, 88, 363, 942 mg/kg-day
Diet
GDs 6-20 (GD 20 c-section)
Saillenfait et al. (2008)
Rat (Sprague-Dawley);
11-14 dams/group
0, 125, 250, 500,
625 mg/kg-day
Gavage
GDs 12-21 (dams allowed to
deliver)
Maternal body weight (percent change compared to control)
Doses 0 100 300 600
maternal BW gain GDs 8-18 0% 9% 4% -35%
900
-42%*
Maternal body weight
Doses 0 600
— Wo significant effect on maternal
weight gain during pregnancy
(quantitative data not reported by
study authors)
Maternal body weight (percent change compared to control)
Doses 0 88 363
BW change GDs 6-20 0% -3% -6%
gravid uterine weight 0% -3% -8%
corrected BW gain GDs 6-20? 0% -2% -2%
942
-11%*
-3%
-25%*
Maternal body weight (percent change compared to control)
Doses 0 125 250 500
BW gain GDs 12-21 0% 4% 6% 6%
625
-3%
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Saillenfait et al. (2006)
Rat (Sprague-Dawley);
20-22 dams/group
0, 250, 500, 750,
1,000 mg/kg-day
Gavage
CDs 6-20 (GD 21 c-section)
Results
Maternal body weight (percent
Doses
BW gain GDs 6-21
gravid uterine weight
corrected BW gain GDs 6-21e
change compared to control13)
0 250 500 750
0% -1% -14% -14%
0% -2% -19%* -28%**
0% 0% 0% 19%
1,000
-39%**
-61%**
19%
Maternal food consumption
Doses
food consumption every
4-6 days, GDs 0-21
0 250 500 750
Wo statistically significant
change from control
1,000
Maternal toxicity
BASF (2007)c
Rat (Wistar); 25 females/group
0, 88, 363, 942 mg/kg-day
Diet
CDs 6-20 (GD 20 c-section)
Abnormalities in dams examined at necropsy
Doses
0 88 363
abnormalities (incidence) 0/25 2/25 0/25
Observed: hemorrhagic thymus,
diaphragmatic hernia, and
dilated renal pelvis
abnormalities (percent 0% 8% 0%
incidence)
Note: Statistical analysis was not performed on these data
942
2/25
8%
Fertility/fetal survival
BASF (2007)c
Rat (Wistar); 25 dams/group
0, 88, 363, 942 mg/kg-day
Diet
CDs 6-20 (GD 20 c-section)
Note: BW and food
consumption measured every
1-3 days through GD 20
Fertility (percent change compared to control13)
Doses
percentage pregnant
0 88 363
23/25 22/25 22/25
942
23/25
Fetal survival (incidence)
Doses
dams with all resorptions
0 88 363
0/23 0/22 0/22
942
0/23
Fetal survival (percent change compared to control)
Doses
percentage preimplantation
loss/litter
percentage postimplantation
loss/litter
percentage resorptions/litter
number of live fetuses/litter
number of live male
fetuses/litter
0 88 363
0% -19% 25%
0% 36% 105%
0% 36% 105%
0% 0% -8%
0% 5% 2%
942
-30%
16%
16%
2%
-2%
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Borch et al. (2006)
Rat (Wistar); 16 dams/group
0, 600 mg/kg-day
Gavage
GDs 7-19 (GD 19 c-section) or
G Ds 7-20/2 1(GD 20/21 c-
section); 5-6 dams/group per
time-point)
Howdeshell et al. (2008)
Rat (Sprague-Dawley);
5-8 dams/group
0, 100, 300, 600,
900 mg/kg-day
Gavage
GDs 8-18 (GD 18 c-section)
Saillenfait et al. (2006)
Rat (Sprague-Dawley);
23-24 dams/group
0, 250, 500, 750,
1,000 mg/kg-day
Gavage
GDs 6-20 (GD 21 c-section)
Results
Fertility (incidence)
Doses 0 600
number pregnant/dams mated 11/16 12/16
Fetal survival
Doses 0 600
— Wo significant effect on litter size,
fetal viability, or number of
resorptions (quantitative data not
reported by study authors)
Fetal survival (n litters evaluated for endpoint)
Doses 0 100 300 600 900
dams with whole litter loss/total 0/5 0/8 0/5 0/5 1/5
dams
number of implantations/litter 13.7(3) 14.8(4) 16.0(3) 12.7(3) 13.3(5)
number of live fetuses/litter 13.3(3) 13.5(4) 15.3(3) 9.3(3) 5.0* (3)
total resorptions/litter 0.2(5) 1.0(8) 0.4(5) 2.0(5) 7.8* (5)
percentage fetal mortality per 1.3% 4.6% 2.7% 17.2% 59.0%*
litter (3) (4) (3) (5) (5)
Fertility
Doses 0 250 500 750 1,000
number pregnant/mated dams 22/24 22/24 22/23 21/23 20/24
(percent) (91.7%) (91.7%) (95.7%) (91.3%) (83.3%)
Fetal survival (percent incidence)
Doses 0 250 500 750 1,000
percentage postimplantation 6.7% 11.0% 13.9% 28.2%** 59.6%**
loss/litter
percentage dead fetuses per 0% 0% 0.3% 0.7% 0.3%
litter
percentage resorptions/litter 6.7% 11.0% 13.6% 27.6%** 59.3%**
Fetal survival (percent change compared to control)
Doses 0 250 500 750 1,000
percentage live litters 0% -5% -5% 0% -10%
number of live fetuses/litter 0% 2% -12% -21%* -52%**
percentage male fetuses/litter 0% -4% -5% 0% 17%
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Saillenfait et al. (2008)
Rat (Sprague-Dawley);
11-14 dams/group
0, 125, 250, 500,
625 mg/kg-day
Gavage
GDs 12-21 (dams allowed to
deliver)
Results
Fetal survival (percent change compared to control13)
Doses
gestation length
percentage postimplantation
loss per litter
percentage pups born alive per
litter
live pups/litter at PND 1
0 125
0% 1%
0% 41%
0% -1%
0% 5%
250
0%
-35%
-4%
0%
500
0%
-31%
-1%
8%
625
1%
-13%
-8%
1%
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
aSome studies measured BW at multiple timepoints/lifestages and not all of these data are presented here. For
the sake of comparability of data across the available studies, BW data measures presented are similar across
studies and/or measures of BW change over the dosing period or greatest time period.
bPercent change compared to control = treated value - control value x 100
control value
CBASF (2007): Dams in the 952 mg/kg-day group showed significantly decreased food consumption on days 10-13
and 15-17 (<10% decreased compared to control); however, overall food consumption did not differ between
groups.
dCorrected weight gain = carcass weight (GD 20 body weight - gravid uterine weight) - GD 6 body weight.
Corrected weight gain = BW gain GDs 6-21 - gravid uterine weight.
* = Statistically significant difference at p < 0.05 from control value, as reported by study authors; ** = Statistically
significant difference at p < 0.01 from control value, as reported by study authors; *** = Statistically significant
difference at p < 0.001 from control value, as reported by study authors.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
I
CD
3=
BASF, 2007; rat
gravid uterine weight
Saillenfait et al., 2006; rat
BW AGO 6-21
Saillenfait et al., 2006; rat
BW AGO 12-21
Saillenfait et al., 2008; rat
gravid uterine weight
BASF, 2007; rat
BW AGO 6-20
BASF, 2007; rat
body weight; GD7-21
Borch etal., 2006; rat
body weight; GD8-18
Howdeshell et al. 2008; rat
• significantly changed
O not significantly changed
Dstatistical analysis not
reported
&
&
O-
-e—e-
o
-e—•
10
100
1000
Doses (mg/kg-day)
2
3
Figure 3-7. Exposure-response array of female reproductive effects, maternal
weight and toxicity, following oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
£ I
•p */)
pre/postimplantation loss
BASF, 2007; rat
% resorptions
BASF, 2007; rat
o # of resorptions
Q. Borchetal., 2006; rat
o
J] # of implantations
^ Howdeshell et al., 2008; rat
£ total resorptions
'•>= Howdeshell et el., 2008; rat
_§ % postimplantation loss
|- Saillenfait et al., 2006; rat
% resorptions
Saillenfait etal., 2006; rat
% postimplantation loss
Saillenfait et al., 2008; rat
# of fetuses/litter
BASF, 2007; rat
fetal viability
Borchetal., 2006; rat
litter size
£ Borch et al., 2006; rat
;§ # of fetuses/litter
- Howdeshell et al. 2008; rat
TO
2 % live litters
Saillenfait etal., 2006; rat
# of fetuses/litter
Saillenfait et al., 2006; rat
# of fetuses/litter
Saillenfait et al., 2008; rat
>- Howdeshell et el., 2008; rat
~3
£ whole litter loss
2 Howdeshell et el., 2008; rat
2
^ Saillenfait etal., 2006; rat
BASF, 2007; rat
Saillenfait et al., 2006; rat
• significantly changed
O not significantly changed
-•-HP
-e—•—«
-e-e
o
o
-•-HI
-e-e
-e—o
10 100
Doses (mg/kg-day)
1000
2
3
Figure 3-8. Exposure-response array of female reproductive effects, fertility
and fetal survival, following oral exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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2
3
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.3.4. Liver Effects
Table 3-20. Evidence pertaining to hepatic effects in animals following oral
exposure to DIBP
Reference and study design
Results
Liver weight
Foster et al. (1982); Foster et al. (1981)
MIBP
Rat (Sprague-Dawley); 6 males/group
0, 800 mg/kg-day
Gavage
4 days
Oishi and Hiraga (1980c)
Rat (JCLWistar); 10 treated males;
20 control males
0, 2% (0, 1,200 mg/kg-day)b
Diet
1 week
Oishi and Hiraga (1980a)
Mouse (JCLICR); 10 males/group
0, 2%(0,2,100mg/kg-day)b
Diet
1 week
Oishi and Hiraga (1980b)
MIBP
Mouse (JCLICR); 10 males/group
0, 2% (0, 2,000 mg/kg-day)b
Diet
1 week
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82, 770,
4,700 mg/kg-day for females)0
Diet
Weaning to 4 months post-weaning
Liver weight (percent change compared to control")
Doses 0
relative weight 0
800
30%***
Liver weight (percent change compared to control0)
Doses 0
absolute weight 0%
relative weight 0%
1,200
27%*
35%*
Liver (with gallbladder) weight (percent change compared to control")
Doses 0
relative weight 0%
2,100
45%*
Liver weight (percent change compared to control0)
Doses 0
relative weight 0%
2,000
30%*
Liver weight (percent change compared to control")
Doses (M) 0 65 710
absolute weight 0% 6% 11%
relative weight 0% 2% 22%
Doses (F) Of 82 770
absolute weight 0% 0% 16%
5,800
5%
84%
4,700
41%
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
University of Rochester (1953)
Rat (Albino; no other strain
designation); 5 males/dose
0, 0.01, 0.1, 1, 2, 5% (0, 15, 140, 1,400,
3,000, 8,900 mg/kg-day)d
Diet
Weaning to 1 month post-weaning
Results
relative weight 0% 0% 8%
62%
Note: Statistical analysis not reported in study.
Liver weight (percent change compared to control")
Doses 0 15 140 1,400 3,000
absolute weight 0% -17% -12% 5% 15%
relative weight 0% -1% 5% 26% 43%
Note: Statistical analysis not reported in study.
8,900
13%
79%
Liver histopathology
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82, 770,
4,700 mg/kg-day for females)0
Diet
Weaning to 4 months post-weaning
University of Rochester (1953)
Rat (Albino; no other strain
designation); 5 males/dose
0, 0.01, 0.1, 1, 2, 5% (0, 15, 140, 1,400,
3,000, 8,900 mg/kg-day)d
Diet
Weaning to 1 month post-weaning
Liver histopathology
Wo treatment-related differences from control were observed at any
dose group.
Note: 4-5 animals per dose group were assessed. Statistical analysis
not reported in study.
Liver histopathology
Histopathological findings only noted in the control group. No
treatment-related differences were observed.
Notes: Number of animals assessed is unclear. Findings limited
"filled with coarse granular cytoplasm." Statistical analysis not
reported in study.
to
1
2
3
4
5
6
7
8
9
10
11
12
13
14
aPercent change compared to control = treated value - control value x 100
control value
bDose conversions were performed using this information: Oishi and Hiraga (1980a, c) average BWs over the week-
long studies were 132 and 24 g for rats and mice, respectively, and the default food consumption rates of
0.008 kg/day for male weanling Wistar rats and 0.0025 kg/day for male weanling B6C3Fi mice were applied (U.S.
EPA, 1988). Oishi and Hiraga (1980b) average BWs for rats and mice in these studies were 145 and 25 g,
respectively, and the default food consumption rates of 0.008 kg/day for male weanling Wistar rats and
0.0025kg/day for male weanling B6C3Fi mice (U.S. EPA, 1988) were applied. Note that Table 1-6 of U.S. EPA
(1988) listed the default food consumption rate for male weanling Wistar rats as 0.080 kg/day. However, it was
later determined using an equation in Table 1-3 of the document that this value was actually supposed to be
0.008 kg/day.
°Dose conversions were performed using this information: University of Rochester (1954) average BWs (measured
at least once weekly) were 269, 277, 252, and 155 g for male rats, and 178,170,182, and 148 g for female rats at
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 0, 0.1,1.0, and 5.0%, respectively; and the default food consumption of 0.018 for male and 0.014 kg/day for
2 female rats (U.S. EPA, 1988) of an unspecified strain in a subchronic study were applied.
3 dDose conversions were performed using this information: University of Rochester (1953) average BWs (measured
4 weekly) of the rats were 139,124,127, 127,121, and 101 g at 0, 0.01, 0.1,1.0, 2.0, and 5.0%, respectively; and
5 the default food consumption of 0.018 for male rats and 0.014 kg/day for female rats (U.S. EPA, 1988) of an
6 unspecified strain in a subchronic study were applied.
7
8 * = Statistically significant difference at p < 0.05 from control value, as reported by study authors; ** = Statistically
9 significant difference at p < 0.01 from control value, as reported by study authors; *** = Statistically significant
10 difference at p < 0.001 from control value, as reported by study authors.
11
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
10
100
1000
10000
University of Rochester, 1954; rat, male
University of Rochester, 1954; rat, female
University of Rochester, 1953; rat, male
00 OJ
1 5
u
-§) Oishi & Hiraga, 1980b; mouse, male
'(U
£
Oishi & Hiraga, 1980a; mouse, male
Oishi & Hiraga, 1980c; rat, male
0)
00
3
5 Foster et al, 1982; rat, male
0
• sign fican
Onot signif
D statist ca
ly changed
cantly changed
analysis not reported
•
1
•
i
»
Doses (mg/kg-day)
2
3
Figure 3-9. Exposure-response array of liver effects following oral exposure to
DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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2
3
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.3.5. Kidney Effects
Table 3-21. Evidence pertaining to renal effects in animals following oral
exposure to DIBP
Reference and study design
Results
Kidney weight
Foster et al. (1982); Foster et al.
(1981)
MIBP
Rat (Sprague-Dawley);
6 males/group
0, 800 mg/kg-day
Gavage
4 days
Oishi and Hiraga (1980c)
Rat (JCLWistar); 10 treated males;
20 control males
0, 2% (0, 1,200 mg/kg-day)b
Diet
1 week
Oishi and Hiraga (1980b)
MIBP
Mouse (JCLICR); 10 males/group
0, 2% (0, 2,000 mg/kg-day)b
Diet
1 week
Oishi and Hiraga (1980a)
Mouse (JCLICR); 10 males/group
Diet
0, 2%(0,2,100mg/kg-day)b
1 week
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for females)0
Diet
Kidney weight (percent change compared to control")
Doses 0
relative weight 0
800
396%***
Kidney weight (percent change compared to control")
Doses 0
absolute weight 0%
relative weight 0%
1,200
-5%
2%
Kidney weight (percent change compared to control0)
Doses 0
relative weight 0%
2,000
-5%
Kidney weight (percent change compared to control")
Doses 0
relative weight 0%
2,100
-10%*
Kidney weight (percent change compared to control")
Doses (M) 0 65 710
absolute weight 0% 10% 9%
relative weight 0% 7% 20%
5,800
-31%
22%
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Weaning to 4 months post-weaning
University of Rochester (1953)
Rat (Albino; no other strain
designation); 5 males/dose
0, 0.01, 0.1, 1, 2, 5% (0, 15, 142,
1,417, 2,975, 8,911 mg/kg-day)d
Diet
Weaning to 1 month post-weaning
Results
Doses (F) 0 82 770 4,
700
absolute weight 0% 4% 11% -2%
relative weight 0% 5% 4% 13%
Note: Statistical analysis not reported in study.
Kidney weight (percent change compared to control")
Doses 0 15 140 1,400 3,000
absolute weight 0% -14% -11% -11% -11%
relative weight 0% 3% 7% 7% 12%
Note: Statistical analysis not reported in study.
8,900
-23%
23%
Kidney histopathology
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for females)0
Diet
Weaning to 4 months post-weaning
University of Rochester (1953)
Rat (Albino; no other strain
designation); 5 males/dose
0, 0.01, 0.1, 1, 2, 5% (0, 15, 142,
1,417, 2,975, 8,911 mg/kg-day)d
Diet
Weaning to 1 month post-weaning
Kidney histopathology
Wo treatment-related differences in males or females were observed
compared with control.
Note: 4-5 animals per dose group were assessed. Findings in males
limited to pyelitis, granuloma, and pyelonephritis; Findings in females
limited to pyelitis and pyelonephritis.
Kidney histopathology
Wo treatment-related differences were observed.
Note: Number of animals assessed is unclear. Findings limited to
eosinophils and inflammatory cells. Statistical analysis not reported
study.
in
1
2
3
4
5
6
7
8
9
10
aPercent change compared to control = treated value - control value x 100
control value
bDose conversions were performed using this information: Oishi and Hiraga (1980a, c) average BWs over the week-
long studies were 132 and 24 g for rats and mice, respectively, and the default food consumption rates of
0.008 kg/day for male weanling Wistar rats and 0.0025 kg/day for male weanling B6C3Fi mice were applied (U.S.
EPA, 1988). Oishi and Hiraga (1980b) average BWs for rats and mice in these studies were 145 and 25 g,
respectively, and the default food consumption rates of 0.008 kg/day for male weanling Wistar rats and 0.0025
kg/day for male weanling B6C3Fi mice (U.S. EPA, 1988) were applied. Note that Table 1-6 of U.S. EPA (1988)
listed the default food consumption rate for male weanling Wistar rats as 0.080 kg/day. However, it was later
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 determined using an equation in Table 1-3 of the document that this value was actually supposed to be 0.008
2 kg/day.
3 °Dose conversions were performed using this information:University of Rochester (1954) average BWs (measured
4 at least once weekly) were 269, 277, 252, and 155 g for male rats, and 178,170,182, and 148 g for female rats at
5 0, 0.1,1.0, and 5.0%, respectively; and default food consumption of 0.018 kg/day for male rats and 0.014 kg/day
6 for female rats (U.S. EPA, 1988) of an unspecified strain in a subchronic study were applied.
7 dDose conversions were performed using this information: University of Rochester (1953) average BWs (measured
8 weekly) were 139,124,127,127,121, and 101 g at 0,0.01, 0.1,1.0, 2.0, and 5.0%, respectively; and default food
9 consumption of 0.018 for male rats and 0.014 kg/day for female rats (U.S. EPA, 1988) of an unspecified strain in a
10 subchronic study were applied.
11
12 * = Statistically significant difference at p < 0.05 from control value, as reported by study authors.
13
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Foster et al., 1982; rat, male
Oishi and Hiraga, 1980c; rat, male
Oishi and Hiraga, 1980a; mouse, male
Oishi and Hiraga, 1980b; mouse, male
University of Rochester, 1953; rat, male
University of Rochester, 1954; rat, male
University of Rochester, 1954; rat, female
• significantly changed
O not significantly changed •
D statistical analysis not reported
B_
O
•
O
10 100
Doses (mg/kg-day)
1000
10000
2
3
Figure 3-10. Exposure-response array of kidney effects following oral
exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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3
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.3.6. Hematopoietic Effects
Table 3-22. Evidence pertaining to hematopoietic effects in animals following
oral exposure to DIBP
Reference and study design
Results
Hematology
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82, 770,
4,700 mg/kg-day for females)b
Diet
Weaning to 4 months post-weaning
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82, 770,
4,700 mg/kg-day for females)b
Diet
Weaning to 4 months post-weaning
Hematology at 4 months (percent change compared to control")
Doses (M)
RBCs
WBCs
Hgb
Doses (F)
RBCs
WBCs
Hgb
Note: Statistical
0
0%
0%
0%
0
0%
0%
0%
analysis not
65
-1%
-37%
-4%
82
6%
-19%
0%
reported in study.
Differential counts at 4 months (percent of each
Doses (M)
neutrophils
eosinophils
basophils
lymphocytes
monocytes
myeloids
blast forms
plasma cells
Doses (F)
neutrophils
eosinophils
basophils
lymphocytes
monocytes
0
20%
1%
0%
79%
1%
0%
0%
0%
0
15%
2%
0%
83%
0%
65
18%
5%
0%
76%
0%
0%
0%
0%
82
14%
4%
0%
82%
0%
710
-5%
-15%
-5%
770
1%
-8%
3%
type of WBC)
710
19%
5%
1%
75%
0%
0%
0%
0%
770
22%
4%
0%
73%
0%
5,800
-16%
38%
-9%
4,700
13%
29%
-6%
5,800
17%
2%
1%
80%
0%
0%
0%
0%
4,700
13%
3%
1%
84%
0%
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
myeloids 0% 0%
blast forms 0% 0%
Note: Statistical analysis not reported in study.
0%
0%
0%
0%
Spleen weight
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for females)b
Diet
Weaning to 4 months post-weaning
Spleen weight (percent change compared to control")
Doses (M) 0 65
absolute weight 2% 9%
relative weight 0% 5%
Doses (F) 0 82
absolute weight 0% 8%
relative weight 0% 11%
Note: Statistical analysis not reported in study.
710
7%
17%
770
93%
80%
5,800
-13%
52%
4,700
-6%
9%
1
2
3
4
5
6
7
8
9
aPercent change compared to control = treated value - control value x 100
control value
bDose conversions were performed by EPA using this information: University of Rochester (1954) average BWs
(measured at least once weekly) were 269, 277, 252, and 155 g for male rats, and 178,170,182, and 148 g for
female rats at 0, 0.1,1.0, and 5.0%, respectively; and the default food consumption of 0.018 kg/day for male
rats and 0.014 kg/day for female rats (U.S. EPA, 1988) of an unspecified strain in a subchronic study were applied.
Hgb = hemoglobin; RBC = red blood cell; WBC = white blood cell
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
= University of Rochester, 1954; rat, male
£1 University of Rochester, 1954; rat, female
University of Rochester, 1954; rat, male
>. oj University of Rochester, 1954; rat, female
on ^
o -n
£ E
U OJ
differential counts*
University of Rochester, 1954; rat male
differential counts*
University of Rochester, 1954; rat, female
c University of Rochester, 1954; rat, male
University of Rochester, 1954; rat, female
University of Rochester, 1954; rat, male
University of Rochester, 1954; rat, female
* differential counts include neutrophil, eosinophil,
basophil, lymphocyte, monocyte, myelocyte, blast forms
and plasma cell measurements
D statistical analysi
reported
not n
10 100
Doses (mg/kg-day)
1000
10000
2
3
Figure 3-11. Exposure-response array of hematopoeitic effects following oral
exposure to DIBP.
This document is a draft for review purposes only and does not constitute Agency policy,
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2
3
Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
3.3.7. Other Effects
Table 3-23. Evidence pertaining to other toxicity effects in animals following
oral exposure to DIBP
Reference and study design
Results
Neurotoxicity effects
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for females)b
Diet
Weaning to 4 months post-weaning
Brain weight (percent change compared to control")
Doses (M) 0 65 710
absolute weight 0% 1% 0%
relative weight 0% -2% 11%
Doses (F) 0 82 770
absolute weight 0% 3% 1%
relative weight 0% 4% -6%
Note: Statistical analysis not reported in study.
5,800
-3%
72%
4,700
2%
17%
Cardiac effects
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for females)b
Diet
Weaning to 4 months post-weaning
Heart weight (percent change compared to control)
Doses (M) 0 65 710
absolute weight 0% 2% -6%
relative weight 0% -3% 3%
Doses (F) 0 82 770
absolute weight 0% 10% -4%
relative weight 0% 10% -11%
Note: Statistical analysis not reported in study.
5,800
-28%
24%
4,700
11%
28%
Lung effects
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for females)b
Diet
Lung weight (percent change compared to control")
Doses (M) 0 65 710
absolute weight 0% 10% 4%
relative weight 0% 5% 16%
5,800
-30%
23%
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Weaning to 4 months post-weaning
Results
Doses (F)
absolute weight
relative weight
Note: Statistical
0
0%
0%
analysis not
reported
82
-20%
-19%
in study.
770
-6%
-12%
4,700
-27%
-16%
Stomach effects
University of Rochester (1954)
Rat (Albino; no other strain
designation); 5 males and 5
females/dose
0,0.1, 1,5% (0,65, 710,
5,800 mg/kg-day for males; 0, 82,
770, 4,700 mg/kg-day for females)b
Diet
Weaning to 4 months post-weaning
Stomach weight (percent change compared to
Doses (M)
absolute weight
relative weight
Doses (F)
absolute weight
relative weight
Note: Statistical
0
0%
0%
0
0%
0%
analysis not
reported
65
10%
7%
82
8%
9%
in study.
control")
710
6%
18%
770
7%
0%
5,800
1%
80%
4,700
18%
35%
Clinical signs
Hazleton Laboratories (1992, 1987);
NIOSH(1983)
Mouse (CD-I); 50 control females,
10 females/treated group
0, 1,000, 1,795, 3,225, 5,790,
10,400 mg/kg-day
Gavage
8 days
Clinical signs of toxicity (incidence /total animals)
Doses
languid
prostrate
ataxia
hunched
tremors
head tilt
thin
wheezing
dyspnea
urine stains
alopecia
rough hair coat
sores
piloerection
opaque eyes
0
1/50
0/50
0/50
0/50
0/50
0/50
1/50
0/50
0/50
0/50
0/50
0/50
0/50
0/50
N/A
1,000
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
1,795
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
3,225
2/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
5,790
5/10
4/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
1/10
10,400
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
0/10
10/10
0/10
10/10
0/10
0/10
0/10
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Reference and study design
Results
discoloration N/A 0/10 0/10 0/10 1/10 0/10
(yellow hair)
Note: Statistical analysis not reported in study for clinical signs data.
1
2
3
4
5
6
7
8
aPercent change compared to control = treated value - control value x 100
control value
bDose conversions were performed using this information: University of Rochester (1954) average BWs (measured
at least once weekly) were 269, 277, 252, and 155 g for male rats, and 178,170,182, and 148 g for female rats at
0, 0.1,1.0, and 5.0%, respectively; and default food consumption of 0.018 kg/day for male rats and 0.014 kg/day
for female rats (U.S. EPA, 1988) of an unspecified strain in a subchronic study were applied.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
University of Rochester, 1954; rat, male
D no statistical analysis
reported
University of Rochester, 1954; rat, female
University of Rochester, 1954; rat, male
University of Rochester, 1954; rat, female
University of Rochester, 1954; rat, male
University of Rochester, 1954; rat, female
University of Rochester, 1954; rat, male
University of Rochester, 1954; rat, female
H, 1983; Hazleton Laboratories, 1987;
Laboratories, 1992; mouse, male & female
1 1
0 1C
Doses (mg/kg-d
i
)0 10
ay)
00 IOC
100000
2
3
Figure 3-12. Exposure-response array of effects on other toxicities following
oral exposure to DIBP.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
l 3.4. PRELIMINARY MECHANISTIC INFORMATION FOR DIBP
2 The systematic literature search for DIBP also identified studies evaluating mechanisms of
3 action considered potentially relevant to effects observed following exposure to DIBP. Studies were
4 included if they evaluated mechanistic events following exposure to DIBP or the metabolite, MIBP,
5 or contained information relevant to the mechanistic understanding of DIBP toxicity. Reviews or
6 analyses that do not contain original data are not included here, but may be considered in later
7 stages of assessment development
8 The diverse array of mechanistic studies presented here includes investigations of the
9 cellular, biochemical, and molecular mechanisms underlying toxicological outcomes. For this
10 preliminary evaluation, information reported in each study was extracted into a database (in the
11 form of an Excel spreadsheet) that will facilitate future evaluation of mechanistic information. This
12 information is being made available to provide an opportunity for stakeholder input, including the
13 identification of relevant studies not captured here.
14 The information extracted from each study and included in the database, corresponds to the
15 column headings in the spreadsheet, and is as follows: link to the HERO record (contained within a
16 URL that links to the study abstract in the HERO database), HERO ID, author(s), year, molecular
17 formulation, in vitro/in vivo, species, cell type, endpoint(s) (i.e., mechanistic outcomes), assay, and
18 mechanistic category. The database supports sorting capabilities, e.g., data can be organized by
19 assay. The database is available through HERO at
20 [http://hero.epa.gov/index.cfm?action=reference.details&reference_id=2508641]. To access the
21 database, click on the link at the top of the web page and select "download" and then "ok" to view
22 the spreadsheet in Excel. This spreadsheet may also be saved to your desktop by downloading and
23 selecting "save." The resulting inventory of DIBP mechanistic studies consists of 32 mechanistic
24 outcomes from 13 identified in vivo studies, as well as 28 mechanistic outcomes from 23 in vitro
25 assays. Table 3-24 presents a summary of the mechanistic outcomes recording in the database
26 from each study identified.
27 The mechanistic categories developed here are not mutually exclusive and are designed to
28 facilitate the analysis of similar studies and experimental observations in a systematic manner.
29 This process will allow the identification of mechanistic events that contribute to mode(s) of action
30 (MOAs) and/or adverse outcome pathways (AOPs) following DIBP exposure. The mechanistic
31 categories assigned to each mechanistic outcome reported by an individual study are as follows:
32 (1) mutation, including investigations of gene and chromosomal mutation; (2) DNA damage,
33 including indicator assays of genetic damage; (3) DNA repair; (4) oxidative stress; (5) cell death and
34 division (this captures a broad range of assays, but it is useful to consider them together as
35 observations resulting from cell cycle alterations; (6) pathology, which includes morphological
36 evaluations pertaining to the dysfunction of organs, tissues, and cells; (7) epigenetic effects, which
37 are observations of heritable changes in gene function that cannot be explained by changes in the
38 DNA sequence; (8) receptor-mediated and cell signaling effects; (9) immune system effects;
39 (10) cellular differentiation and transformation; (11) cellular energetics; and (12) "other," to
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1
2
3
4
capture those mechanistic outcomes not easily assigned to a defined category. Mechanistic
outcomes in the "other" category include sex steroid hormone (e.g., testosterone) production and
gene expression.
5
6
Table 3-24. Summary of mechanistic outcomes evaluated following DIBP or
MIBP administration
Mechanistic
category
Mutation3
DNA damage
Total #
outcomes/
# studies
5/5
6/4
Total
0
0
In vivo (# outcomes/
# studies)
Human
0
0
Rat
0
0
Mouse
0
0
In vitro (# outcomes/ft studies)
Total
5/5
6/4
Human
0
6/4
Primate
0
0
Rat
0
0
Mouse
0
0
DNA repair
Oxidative stressb
Cell death and
division
Pathology
1/1
7/5
2/2
1/1
3/2
2/2
0
0
0
0
2/2
2/2
0
1/1
0
0
4/3
N/A
0
3/2
0
0
0
0
0
1/1
N/A
Epigenetics
Receptor-
mediated and
cell signaling0
Immune system
Cellular
differentiation
and
transformation
Cellular
energetics
Otherd
Total
14/9
5/3
1/1
1/1
18/11
60/35
8/4
1/1
1/1
0
16/10
1/1
0
1/1
0
0
7/3
1/1
0
0
14/8
0
0
0
0
2/2
32/13
6/5
4/2
0
1/1
2/2
0
1/1
0
0
0
1/1
0
0
0
0
0
2/1
0
1/1
2/2
0
1/1
0
0
0
28/23
7
8
9
10
11
12
13
14
15
16
17
18
aDatabase included five outcomes in five studies utilizing Salmonella typhimuhum.
bDatabase included one outcome from one study utilizing Caenorhabditis elegans.
°Database included two outcomes from one study utilizing cultured hamster cells, two outcomes from two studies
utilizing yeast, and one cell-free system.
dDatabase primarily composed of hormone (testosterone, estradiol) content or production in tissues from rats and
mice.
Notes: The number in rows may not sum to "total" amounts as several studies evaluated multiple species or
employed both in vivo and in vitro models. The mechanistic categories in italics and in gray shading had no DIBP-
specific information available.
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1
2
3
4
5
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7
8
9
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
Information summarized in Table 3-24 and Figure 3-13 and detailed in the mechanistic
database can be used to ascertain the breadth and scope of available mechanistic studies. At this
preliminary stage, study results are not presented. Additionally, the inclusion of a study in the
spreadsheet does not reflect conclusions reached as to mechanistic study quality or relevance.
After the epidemiological and experimental studies on each health effect have been synthesized,
mechanistic studies will be reviewed and findings synthesized to evaluate potential MOAs and/or
AOPs, which can be used to inform hazard identification and dose-response assessment, specifically
addressing questions of human relevance, susceptibility, and dose-response relationships.
11
12
13
14
Mechanistic Outcomes
(60 outcomes from 35 reports)
I n vivo
I In vitro
Mutation
DNA damage
Oxidative stress
Cell death and division
Pathology
Receptor-mediated and cell
signaling
Immune system
Cellular differentiation and
transformation
Cellular energetics
Other
10
Number of endpoints
15
20
Figure 3-13. Summary of in vivo and in vitro mechanistic data for DIBP and
MIBP by mechanistic category.
This document is a draft for review purposes only and does not constitute Agency policy,
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
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26 of Age. Environ Health Perspect 120: 290-295. http://dx.doi.org/10.1289/ehp.1103705
27 Wirth. J: Rossano. M: Potter. R: Puscheck. E: Daly. D: Paneth. N: Krawetz. S: Protas. B:
28 Diamond, M. (2008). A pilot study associating urinary concentrations of phthalate
29 metabolites and semen quality. Sys Biol Reprod Med 54: 143-154.
30 http://dx.doi.org/10.1080/19396360802055921
31 Wittassek, M: Koch, HM; Angerer, J: Bruning, T. (2011). Assessing exposure to phthalates - the
32 human biomonitoring approach [Review]. Mol Nutr Food Res 55: 7-31.
33 http://dx.doi.org/10.1002/mnfr.201000121
34 Wolff. MS: Engel SM: Berkowitz. GS: Ye. X: Silva. MJ: Zhu. C: Wetmur. J: Calafat AM.
35 (2008). Prenatal phenol and phthalate exposures and birth outcomes. Environ Health
36 Perspect 116: 1092-1097. http://dx.doi.org/10.1289/ehp.11007
37 Wolff. MS: Teitelbaum. SL: Pinnev. SM: Windham. G: Liao. L: Biro. F: Kushl LH: Erdmann.
38 C: Hiatt RA; Rvbak, ME; Calafat, AM. (2010). Investigation of relationships between
39 urinary biomarkers of phytoestrogens, phthalates, and phenols and pubertal stages in
40 girls. Environ Health Perspect 118: 1039-1046. http://dx.doi.org/10.1289/ehp.0901690
This document is a draft for review purposes only and does not constitute Agency policy.
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Preliminary Materials for the IRIS Toxicological Review ofDiisobutyl Phthalate
1 Wormuth, M; Scheringer, M; Vollenweider, M; Hungerbuhler, K. (2006). What are the sources
2 of exposure to eight frequently used phthalic acid esters in Europeans? Risk Anal 26:
3 803-824. http://dx.doi.0rg/10.llll/i.1539-6924.2006.00770.
4 Zhu, XB; Tay, TW: Andriana, BB; Alam, MS: Choi, EK; Tsunekawa, N; Kanai, Y; Kurohmaru,
5 M. (2010). Effects of di-iso-butyl phthalate on testes of prepubertal rats and mice.
6 Okajimas Folia Anat Jpn 86: 129-136.
7 Zota, AR; Calafat AM; Woodruff, TJ. (2014). Temporal trends in phthalate exposures: findings
8 from the national health and nutrition examination survey, 2001-2010. Environ Health
9 Perspect 122: 235-241. http://dx.doi.org/10.1289/ehp.1306681
10
11
This document is a draft for review purposes only and does not constitute Agency policy.
4-15 DRAFT—DO NOT CITE OR QUOTE
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