DI(2-ETHYLHEXYL)PHTHALATE
o
o
Agency for Toxic Substances and Disease Registry
U.S. Public Health Service
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ATSDR/TP-88/15
TOXICOLOGICAL PROFILE FOR
DI(2-ETHYLHEXYL)PHTHALATE
Date Published — April 1989
Prepared by:
Life Systems, Inc.
under Contract No. 68-02-4228
for
Agency for Toxic Substances and Disease Registry (ATSDR)
U.S. Public Health Service
in collaboration with
U.S. Environmental Protection Agency (EPA)
Technical editing/document preparation by:
Oak Ridge National Laboratory
under
DOE Interagency Agreement No. 1857-B026-A1
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DISCLAIMER
Mention of company name or product does not constitute endorsement by
the Agency for Toxic Substances and Disease Registry.
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FOREWORD
The Superfund Amendments and Reauthorization Act of 1986 (Public
Law 99-499) extended and amended the Comprehensive Environmental
Response, Compensation, and Liability Act of 1980 (CERCLA or Superfund)
This public law (also known as SARA) directed the Agency for Toxic
Substances and Disease Registry (ATSDR) to prepare toxicological
profiles for hazardous substances which are most commonly found at
facilities on the CERCLA National Priorities List and which pose the
most significant potential threat to human health, as determined by
ATSDR and the Environmental Protection Agency (EPA). The list of the 100
most significant hazardous substances was published in the Federal
Register on April 17, 1987.
Section 110 (3) of SARA directs the Administrator of ATSDR to
prepare a toxicological profile for each substance on the list. Each
profile must include the following content:
"(A) An examination, summary, and interpretation of available
toxicological information and epidemiologic evaluations on a
hazardous substance in order to ascertain the levels of significant
human exposure for the substance and the associated acute,
subacute, and chronic health effects.
(B) A determination of whether adequate information on the healch
effects of each substance is available or in the process of
development to determine levels of exposure which present a
significant risk to human health of acute, subacute, and chronic
health effects.
(C) Where appropriate, an identification of toxicological testing
needed to identify the types or levels of exposure that may present
significant risk of adverse health effects in humans."
This toxicological profile is prepared in accordance with
guidelines developed by ATSDR and EPA. The guidelines were published in
the Federal Register on April 17, 1987. Each profile will be revised and
republished as necessary, but no less often than every three years, as
required by SARA.
The ATSDR toxicological profile is intended to characterize
succinctly the toxicological and health effects information for the
hazardous substance being described Each profile identifies and reviews
the key literature that describes a hazardous substance's toxicological
properties. Other literature is presented but described in less detail
than the key studies. The profile is not intended to be an exhaustive
document; however, more comprehensive sources of specialty information
are referenced.
111
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foreword
Each toxicological profile begins with a public health statement,
which describes In nontechnical language a substance's relevant
toxicological properties. Following the statement Is material that
presents levels of significant human exposure and, where known,
significant health effects. The adequacy of Information to determine a
substance's health effects Is described in a health effects summary.
Research gaps in toxicologic and health effects information are
described in the profile. Research gaps that are of significance to
protection of public health will be Identified by ATSDR, the National
Toxicology Program of the Public Health Service, and EPA. The focus of
the profiles is on health and toxicological information; therefore, we
have included this information in the front of the document.
The principal audiences for the toxicological profiles are health
professionals at the federal, state, and local levels, interested
private sector organizations and groups, and members of the public. We
plan to revise these documents in response to public comments and as
additional data become available; therefore, we encourage comment that
will make the toxicological profile series of the greatest use.
This profile reflects our assessment of all relevant toxicological
testing and information that has been peer reviewed. It has been
reviewed by scientists from ATSDR, EPA, the Centers for Disease Control,
and the National Toxicology Program. It has also been reviewed by a
panel of nongovernment peer reviewers and was made available for public
review. Final responsibility for the contents and views expressed in
this toxicological profile resides with ATSDR.
James 0. Mason, M.D., Dr. P.H.
Assistant Surgeon General
Administrator, ATSDR
iv
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CONTENTS
FOREWORD llt
LIST OF FIGURES ix
LIST OF TABLES xi
1. PUBLIC HEALTH STATEMENT 1
1.1 WHAT IS DI(2-ETHYLHEXYL)PHTHALATE? '.'.'.'.'.'.'.'.'.' 1
1.2 HOW MIGHT I BE EXPOSED TO DI(2-ETHYLHEXYL)PHTHALATE? 1
1.3 HOW DOES DI(2-ETHYLHEXYL)PHTHALATE GET INTO MY BODY? ... 2
1.4 HOW CAN DI (2 - ETHYLHEXYL) PHTHALATE AFFECT MY HEALTH? 2
1.5 IS THERE A MEDICAL TEST TO DETERMINE IF I HAVE
BEEN EXPOSED TO DI(2-ETHYLHEXYL)PHTHALATE? 2
1.6 WHAT LEVELS OF EXPOSURE HAVE RESULTED IN HARMFUL
HEALTH EFFECTS? 3
1.7 WHAT RECOMMENDATIONS HAS THE FEDERAL GOVERNMENT MADE
TO PROTECT HUMAN HEALTH? 3
2. HEALTH EFFECTS SUMMARY 7
2.1 INTRODUCTION '.'.'.'.'.'.'.'.'.'.. 7
2.2 LEVELS OF SIGNIFICANT HUMAN EXPOSURE '.'.'.'.'.'.'.'.'.'. 8
2.2.1 Key Studies and Graphic Presentations 8
2.2.1.1 Lethality/decreased longevity 8
2.2.1.2 Systemic/target organ toxicity 13
2.2.1.3 Developmental toxicity 15
2.2.1.4 Reproductive toxicity . . . 16
2.2.1.5 Genotoxicity 17
2.2.1.6 Carcinogenicity 18
2.2.2 Biological Monitoring as a Measure of Exposure
and Effec ts 21
2.2.3 Environmental Levels as Indicators of Exposure
and Effects 22
2.2.3.1 Levels found in the environment 22
2.2.3.2 Human exposure potential 22
2.3 ADEQUACY OF DATABASE 22
2.3.1 Introduction 22
2.3.2 Health Effect End Points '.'.'.'.'. 23
2.3.2.1 Introduction and graphic summary 23
2.3.2.2 Description of highlights of graphs .... 23
2.3.2.3 Summary of relevant ongoing research .... 23
2.3.3 Other Information Needed for Human
Health Assessment 26
2.3.3.1 Pharmacokinetics and mechanisms
of action . . 26
2.3.3.2 Monitoring of human biological samples .. 26
2.3.3.3 Environmental considerations 26
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Concents
3. CHEMICAL AND PHYSICAL INFORMATION 29
3.1 CHEMICAL IDENTITY ' ' ' 29
3 . 2 CHEMICAL AND PHYSICAL PROPERTIES '.'.'.'.'.'. 29
4. TOXICOLOGICAL DATA 33
4.1 OVERVIEW 33
4.2 TOXICOKINETICS '.'.'.'.'.'.'.'.'.'.'.'.'.'.'.'. 34
4.2.1 Absorption 34
4.2.1.1 Inhalation '.'.'.'.'.'.'.'.'. 34
4.2.1.2 Oral 34
4.2.1.3 Dermal '.'.'.'.'. 35
4.2.2 Distribution 35
4.2.2.1 Inhalation " 36
4.2.2.2 Oral 36
4.2.2.3 Dermal 38
4.2.2.4 Systemic 38
4.2.3 Metabolism 39
4.2.3.1 Inhalation '.'.'.'.'.'.'.'. 39
4.2.3.2 Oral " 4!
4.2.3.3 Dermal '.'.'.'.'. 42
4.2.3.4 In vitro studies 43
4.2.4 Excretion 43
4.2.4.1 Inhalation '.'.'. 43
4.2.4.2 Oral 43
4.2.4.3 Dermal '//_ 45
4.2.4.4 Systemic 45
4.2.5 Discussion 45
4.3 TOXICITY '.'.'.'.'.'.'.'.'.'.'.'.'.'.'.'. 47
4.3.1 Lethality and Decreased Longevity 47
4.3.1.1 Inhalation 47
4.3.1.2 Oral 47
4.3.1.3 Dermal 49
4.3.1.4 Discussion 49
4.3.2 Systemic/Target Organ Toxicity 49
4.3.2.1 Hepatic toxicity 49
4.3.2.2 Testicular toxicity 53
4.3.3 Developmental Toxicity 53
4.3.3.1 Inhalation 53
4.3.3.2 Oral \\\ 53
4.3.3.3 Dermal 55
4.3.3.4 Discussion 55
4.3.4 Reproductive Toxicity 56
4.3.4.1 Inhalation 56
4.3.4.2 Oral '.'.'.'.'. 56
4.3.4.3 Dermal 60
4.3.4.4 General discussion 60
4.3.5 Genotoxicity 61
4.3.5.1 Genotoxicity studies 61
4.3.5.2 Discussion 61
4.3.6 Carcinogenicity 64
4.3.6.1 Inhalation 64
4.3.6.2 Oral 64
4.3.6.3 Dermal 67
vi
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Concents
4. 3.6.4 Possible mechanisms of carcinogenesis ... 68
4.3.6.5 General discussion 76
4.4 INTERACTION WITH OTHER CHEMICALS \ 75
5. MANUFACTURE, IMPORT, USE, AND DISPOSAL 77
5.1 OVERVIEW ' 77
5.2 PRODUCTION 77
5.3 IMPORT 77
5.4 USE '.'.'.'..'.'.'." 78
5.5 DISPOSAL 78
6. ENVIRONMENTAL FATE 79
6.1 OVERVIEW '.'.'.'.'.'.'.'.'.'.'. 79
6 . 2 RELEASES TO THE ENVIRONMENT '.'.'.'.'.'.'.'.'.'.'. 79
6.2.1 Anthropogenic Sources 79
6.2.2 Natural Sources 79
6 . 3 ENVIRONMENTAL FATE '.'.'.'.'.'.'. 81
6.3.1 Atmospheric Fate Processes 81
6.3.2 Surface Water/Groundwater Fate Processes 81
6.3.3 Soil Fate Processes 82
6.3.4 Biotic Fate Processes 82
7. POTENTIAL FOR HUMAN EXPOSURE 83
7.1 OVERVIEW ' 83
7.2 LEVELS MONITORED OR ESTIMATED IN THE ENVIRONMENT ........ 83
7.2.1 Levels in Air 83
7.2.2 Levels in Water 84
7.2.3 Levels in Soil 85
7.2.4 Levels in Food 85
7.2.5 Resulting Exposure Levels 85
7.3 OCCUPATIONAL EXPOSURES 86
7.4 POPULATIONS AT HIGH RISK 86
7.4.1 Above-Average Exposure 86
7.4.2 Above-Average Sensitivity 87
8. ANALYTICAL METHODS 89
8.1 Environmental Media 89
8.1.1 Air '.'.'.'.'.'.'.'.'. 89
8.1.2 Water 89
8.1.3 Soil ....... 91
8.1.4 Food ' 91
8 .2 BIOMEDICAL SAMPLES '.'.'.'.'.'.'.'.'. 91
8.2.1 Fluids and Exudates 91
8.2.2 Tissues 91
9. REGULATORY AND ADVISORY STATUS 93
9 .1 INTERNATIONAL ' ' 93
9 . 2 NATIONAL ' ' ." 93
9.2.1 Regulations 93
9.2.2 Advisory Guidance 95
9.2.3 Data Analysis 95
9.2.3.1 Reference dose 95
9.2.3.2 Carcinogenic potency 96
VI 1
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Concents
9.3 STATE 97
9.3.1 Regulations 97
9.3.2 Advisory Guidance // 97
10. REFERENCES 9g
11. GLOSSARY 115
APPENDIX: PEER REVIEW 119
viii
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LIST OF FIGURES
1.1 Health effects from breathing di(2-ethylhexyDphthalate 4
1.2 Health effects from Ingesting dl(2-ethylhexyl)phthalate 5
2.1 Effects of dl(2-ethylhexyl)phthalate--inhalatlon exposure 9
2.2 Effects of di(2-ethylhexyDphthalate--oral exposure 10
2.3 Levels of significant exposure for di(2-ethylhexyl)-
phthalate--Inhalation H
2.4 Levels of significant exposure for di(2-ethylhexyl)-
phthalate--oral 12
2.5 Availability of information on health effects of
dl(2-ethylhexyl)phthalate (human data) 24
2.6 Availability of information on health effects of
di(2-ethylhexyl)phthalate (animal data) 25
4.1 Proposed metabolism for di(2-ethylhexyl)phthalate
in the rat 40
4.2 Species-specific metabolism of di(2-ethylhexyl)phthalate 46
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LIST OF TABLES
2.1 Summary of ongoing research 27
3.1 Chemical identity of di(2-ethylhexyl)phthalate 30
3.2 Physical and chemical properties of
di(2-ethylhexyl)phthalate 31
4.1 Acute lethality of di(2-ethylhexyl)phthalate 48
4.2 Results of short-term assays 62
4.3 Incidence of hepatocellular carcinomas and neoplastic
nodules observed in DEHP-dosed rats 66
4.4 Incidence of hepatocellular carcinomas and hepatocellular
adenomas observed in DEHP-dosed mice 66
6.1 Releases of di(2-ethylhexyl)phthalate to the environment 80
8.1 Analytical methods for di(2-ethylhexyl)phthalate
in environmental samples 90
8.2 Analytical methods for di(2-ethylhexyl)phthalate
in biological samples 92
9.1 Regulations and guidelines applicable to
di(2-ethylhexyl)phthalate 94
XI
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1. PUBLIC HEALTH STATEMENT
1.1 WHAT IS DI(2-ETHYLHEXYL)FHTHALATE?
Di(2-ethylhexyl)phthalate, commonly referred Co as DEHP, is a
liquid widely used to make plastics more flexible. Plastics may contain
from 1 to 40% DEHP by weight and are widely used in consumer products
such as imitation leather, rainwear, footwear, upholstery, flooring,
tablecloths, shower curtains, food packaging materials, and children's
toys. Plastics containing OEHP are also used for Cubing and containers
for blood transfusions and blood products. OEHP is also used as a
hydraulic fluid and as a dielectric fluid (a nonconductor of electric
current) for use in electrical capacitors.
1.2 HOW MIGHT I BE EXPOSED TO DI(2-ETHYLHEXYL)PHTHALATE?
About 300 million pounds of DEHP are used each year to manufacture
plastic products for commercial, medical, and consumer use. Low-level
exposures to DEHP from the air or drinking water occur whenever these
products are used. Plasticizers such as DEHP do not become a permanent
part of the plastic matrix during the manufacturing process. Thus, under
certain use or disposal conditions, DEHP can migrate from plastic
products into the environment. As a result, DEHP is widely distributed
in the environment, and exposure can occur via air, water, and food.
Most exposures to DEHP are at very low levels, and exposures via air and
water are expected to be minimal.
A potential exists for DEHP contamination of food during
processing, handling, transportation, and packaging. DEHP is found in
animal products used for human consumption. The highest levels have been
detected in milk and cheese. The U.S. Food and Drug Administration (FDA)
currently allows the use of DEHP plasticized containers or wrappings
only for foods that primarily contain water; therefore, properly
packaged foods are unlikely to become contaminated. The FDA ruling,
however, does not preclude the possible misuse of DEHP plasticized
containers or wrappings by the consumer.
Acute exposures to relatively high levels (compared to DEHP levels
commonly found in food or drinking water) of DEHP can occur when DEHP
migrates from the plastics used in medical apparatus (such as storage
bags and tubing) used for blood transfusions or kidney dialysis.
Patients exposed to DEHP through medical procedures such as dialysis may
conceivably be at risk from its toxic effects, but confirmatory studies
are necessary to show this relationship.
Some exposure may occur from the evaporation of DEHP from
plasticized products during their use. The level of such exposures will
depend on the thickness of the plastic item, the temperature, and the
specific nature of the product. Therefore, it is difficult to estimate
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2 Section 1
the level of this kind of exposure. However, because the vapor pressure
of DEHP is very low, and essentially no evaporation occurs at normal
room temperature, inhalation exposure is expected to be minimal.
Most discarded plasticized products are disposed of either by
incineration or by dumping in a landfill. Incineration is not likely to
result in significant exposures to DEHP. After disposal of products in a
landfill, the DEHP will slowly leach from the plastic product and may
reach the groundwater. However, the low solubility of DEHP in water
limits the amounts that will enter the environment by this route.
Exposure from water nay also occur if DEHP migrates from some kinds
of flexible plastic tubing used to carry drinking water. Again, because
DEHP is not very soluble in water, the exposure level may be low but
could continue for a long period of time.
Estimation of human exposure to DEHP Is very complex because of the
wide range of items containing DEHP and the large number of variables
influencing how much of the DEHP content of an item would reach an
exposed individual and be absorbed into the body.
1.3 HOW DOES DI(2-ETHYLHEXYL)PHTHALATE GET INTO MY BODY?
There are several potential sources' of exposure to DEHP. Humans are
primarily exposed through foods that come into contact with packaging
materials containing DEHP. Once DEHP gets into the gastrointestinal
tract, it is quickly absorbed and gets into the blood and is rapidly
metabolized. Humans may also receive higher exposures during blood
transfusions and kidney dialysis because of the movement of DEHP into
blood from plastic bags and tubing.
1.4 HOW CAN DI(2-ETHYLHEXYL)FHTHALATE AFFECT MY HEALTH?
There are essentially no studies on the health effects of DEHP in
humans. However, DEHP causes cancer in rats and mice. It is also known
to produce liver damage and male reproductive system damage, affect
reproduction, and produce birth defects in laboratory animals. Because
none of these effects have been documented in humans, it is difficult to
estimate the kinds of health effects and exposure levels that may
actually affect humans. However, it is prudent to regard the animal data
as indicating some degree of concern for harmful human effects until
research can more reasonably conclude that no harm can occur.
1.5 IS THERE A MEDICAL TEST TO DETERMINE IF I HAVE
BEEN EXPOSED TO DI(2-ETHYLH£XYL)PHTHALATE?
The major method for determining whether a person has been exposed
to DEHP is by testing the urine. After exposure to DEHP, the body
metabolizes it into two major chemical products. mono(2-ethylhexyl)-
phthaiate (MEHP) and 2-ethylhexanol. Detection of DEHP and/or these
chemicals in the urine will indicate whether a person has been recently
exposed. In addition, the blood can be analyzed for DEHP, MEHP, and 2-
ethylhexanol shortly after exposure.
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Public Health Statement 3
L.6 WHAT LEVELS OF EXPOSURE HAVE RESULTED IN
HARMFUL HEALTH EFFECTS?
The graphs on Che following pages (Figs. 1.1 and 1.2) show the
relationship between exposure to DEHP and known health effects. In the
first set of graphs (Fig. 1.1). labeled "Health effects from breathing
DEHP - exposure is measured in milligrams of DEHP per cubic meter of air
(me/m3). In all graphs, effects In animals are shown on the left side,
and effects in humans are shown on the right. In the second set of
graphs (Fig 1.2), the same relationship is represented for the known
"Health effects from ingesting DEHP." Exposures are measured in
milligrams of DEHP per kilogram of body weight (mg/kg).
Figure 1 1 summarizes the effects resulting from short-term
inhalation of DEHP at levels up co -24,000 mg/m3. No adverse effects
were observed in the studies that were reviewed, and no longer-term
inhalation studies have been done. Figure 1.2 shows the general types of
adverse health effects resulting from ingestion of DEHP at various
exposure levels. In short-term animal studies, death has resulted from
ingestion of relatively high doses of DEHP. In other animal studies,
both short-term and long-term ingestion of DEHP has resulted in liver
toxicity, reproduction toxicity, and toxicity to the unborn or newborn
These effects have been observed only in animal studies. Similar
information based on human exposures to DEHP is not available.
DEHP has also been shown to cause liver tumors in both rats and
mice receiving DEHP in their diet throughout their entire life span.
Based on the results of these cancer studies, the excess risk to humans
has been estimated. The number of excess cases of cancer which would be
produced in populations of 10,000, 100.000, 1.000,000, or 10,000,000
individuals exposed to 1 microgram of DEHP per liter for their entire
lifetime is estimated to be 0.024. 0.24, 2.4, and 24. respectively. It
should be noted that these risk values are plausible upper-limit
estimates. Actual risk levels are unlikely to be higher and may be
lower.
1.7 WHAT RECOMMENDATIONS HAS THE FEDERAL GOVERNMENT
MADE TO PROTECT HUMAN HEALTH?
The federal government has taken a number of steps to protect
humans from DEHP. The Environmental Protection Agency (EPA) has
authority to regulate DEHP under the Resource Conservation and Recovery
Act (RCRA)- the Comprehensive Environmental Response. Compensation, and
Liability Act (CERCLA or "Superfund"); the Solid Waste Disposal Act; the
Clean Water Act; the Safe Drinking Water Act; the Clean Air Act; and the
Toxic Substances Control Act. The EPA has identified DEHP as a toxic
waste material and requires that persons who generate, transport treat.
store, or dispose of it comply with regulations of the EPA Federal
Hazardous Waste Management Program. The EPA has also identified DEHP as
a hazardous constituent of wastes from the cleaning equipment and tanks
used in paint manufacturing. The EPA has established human health
criteria for ingestion of water and contaminated aquatic organisms (such
as fish and shellfish) at 15 milligrams per liter of water for
noncancerous health effects. EPA has also calculated risk estimates for
the probability of developing cancer from ingesting DEHP. The
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Section 1
SHORT-TERM EXPOSURE
(LESS THAN OR EQUAL TO 14 DAYS)
LONG-TERM EXPOSURE
(GREATER THAN 14 DAYS)
EFFECTS
IN
ANIMALS
NO LETHALITY-
CONG. IN
AIR
(mg/m3)
100.000
10.000
1.000
100
10
EFFECTS
IN
HUMANS
QUANTITATIVE
DATA WERE
NOT AVAILABLE
EFFECTS CONC. IN EFFECTS
IN AIR IN
ANIMALS (mg/m3) HUMANS
QUANTITATIVE 100
DATA WERE
NOT AVAILABLE
10.
1.C
1C
1
,000 QUANTITATIVE
DATA WERE
NOT AVAILABLE
300
00
0
3
Fig. 1.1. Health effects from breathing di(2-etbylhexyl)plithalate.
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Public Health Statement
SHORT-TERM EXPOSURE
(LESS THAN OR EQUAL TO 14 DAYS)
LONG-TERM EXPOSURE
(GREATER THAN 14 DAYS)
EFFECTS
IN
ANIMALS
DEATH•
REPRODUCTIVE
TOXICITY
LIVER
TOXICITY
TOXIC EFFECTS
ON UNBORN OR
NEWBORN
DOSE
(mg/kg/day)
100.000
-^<
— ^«
10.(
I — 1.C
1(
00
00
0
EFFECTS
IN
HUMANS
QUANTITATIVE
DATA WERE
NOT AVAILABLE
EFFECTS
IN
ANIMALS
DOSE
(mg/kg/day)
100.000
EFFECTS
IN
HUMANS
10
QUANTITATIVE
DATA WERE
NOT AVAILABLE
10.000
1.000
REPRODUCTIVE
TOXICITY
TOXIC EFFECTS
ON UNBORN OR
NEWBORN
LIVER
TOXICITY
100
10
10
1 0
Fig. 1.2. Health effects from ingesting di(2-*thylhexyl)phthalate.
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Section 1
regulates the chemical as an unintentional food additive. Finally, the
Occupational Safety and Health Administration (OSHA) has set a
permissible exposure level of 5 milligrams DEHP per cubic meter of air
for occupational exposures to DEHP.
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2. HEALTH EFFECTS SUMMARY
2.1 INTRODUCTION
This section summarizes and graphs data on the health effects
concerning exposure to DEHP. The purpose of this section is to present
levels of significant exposure for DEHP based on key toxicological
studies, epidemiological investigations, and environmental exposure
data. The information presented in this section is critically evaluated
and discussed in Sect. 4, Toxicological Data, and Sect. 7, Potential for
Human Exposure.
This Health Effects Summary section comprises two major parts.
Levels of Significant Exposure (Sect. 2.2) presents brief narratives and
graphics for key studies in a manner that provides public health
officials, physicians, and other interested individuals and groups with
(1) an overall perspective of the toxicology of DEHP and (2) a
summarized depiction of significant exposure levels associated with
various adverse health effects. This section also includes information
on the levels of DEHP that have been monitored in human fluids and
tissues and information about levels of DEHP found in environmental
media and their association with human exposures.
The significance of the exposure levels shown on the graphs may
differ depending on the user's perspective. For example, physicians
concerned with the interpretation of overt clinical findings in exposed
persons or with the identification of persons with the potential to
develop such disease may be interested in levels of exposure associated
with frank effects (Frank Effect Level, FEL). Public health officials
and project managers concerned with response actions at Superfund sites
may want information on levels of exposure associated with more subtle
effects in humans or animals (Lowest-Observed-Adverse-Effect Level,
LOAEL) or exposure levels below which no adverse effects (No-Observed-
Adverse-Effect Level, NOAEL) have been observed. Estimates of levels
posing minimal risk to humans (Minimal Risk Levels) are of interest to
health professionals and citizens alike.
Adequacy of Database (Sect. 2.3) highlights the availability of key
studies on exposure to DEHP in the scientific literature and displays
these data in three-dimensional graphs consistent with the format in
Sect. 2.2. The purpose of this section is to suggest where there might
be insufficient information to establish levels of significant human
exposure. These areas will be considered by the Agency for Toxic
Substances and Disease Registry (ATSDR), EPA, and the National
Toxicology Program (NTP) of the U.S. Public Health Service in order to
develop a research agenda for DEHP.
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8 Section 2
2.2 LEVELS OF SIGNIFICANT HUMAN EXPOSURE
To help public health professionals address the needs of persons
living or working near hazardous waste sites, the toxicology data
summarized in this section are organized first by route of exposure--
inhalation, ingestion, and dermal--and then by toxicological end points
that are categorized into six general areas--lethality, systemic/target
organ toxicity, genetic toxicity, and carcinogenicity. The data are
discussed in terms of three exposure periods--acute, intermediate, and
chronic.
Two kinds of graphs are used to depict the data. The first.type is
a "thermometer" graph. It provides a graphical summary of the human and
animal toxicological end points (and levels of exposure) for each
exposure route for which data are available. The ordering of effects
does not reflect the exposure duration or species of animal tested. The
second kind of graph shows Levels of Significant Exposure (LSE) for each
route and exposure duration. The points on the graph showing NOAELs and
LOAELs reflect the actual doses (levels of exposure) used in the key
studies. No adjustments for exposure duration or intermittent exposure
protocol were made.
Adjustments reflecting the uncertainty of extrapolating animal data
to man, intraspecies variations, and differences between experimental vs
actual human exposure conditions were considered when estimates of
levels posing minimal risk to human health were made for noncancer end
points. These minimal risk levels were derived for the most sensitive
noncancer end point for each exposure duration by applying uncertainty
factors. These levels are shown on the graphs as a broken line starting
from the actual dose (level of exposure) and ending with a concave-
curved line at its terminus. Although methods have been established to
derive these minimal risk levels (Barnes et al. 1987), shortcomings
exist in the techniques that reduce the confidence in the projected
estimates. Also shown on the graphs under the cancer end point are low-
level risks (10'4 to 10'7) reported by EPA. In addition, the actual dose
(level of exposure) associated with the tumor incidence is plotted.
2.2.1 Key Studies and Graphic Presentations
Figures 2.1 and 2.2 are graphic presentations of the adverse health
effects that have been observed in studies with experimental animals
using inhalation and ingestion exposure routes, respectively. Animal
studies using dermal exposure were not located in the available
literature. Human data on the adverse health effects of DEHP were not
available.
Figures 2.3 and 2.4 present data for various toxicity end points
for the inhalation and oral routes, respectively. Details of these
studies are presented in the following subsections.
2.2.1.1 Lethality/decreased longevity
No animal studies provided appropriate data to evaluate lethality
or decreased longevity as a toxicity end point for DEHP exposure via
dermal exposure. In inhalation studies with rats, lethality was not
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Health Effects Summary 9
ANIMALS
(mgftn3)
HUMANS
100.000 |—
10.000
1.000
100
10
O RAT. ACUTE LETHALITY. 1 h
O NOAEL
QUANTITATIVE DATA
WERE NOT AVAILABLE
Fig. 2.1 Effects of di(2-ethylhexyl)phthalate—inhalation exposure.
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10 Section 2
ANIMALS
(mg/Vg/day)
HUMANS
100 0001-
10.000
100
10
RABBIT LDjo
RAT. LD,o
GUINEA PIG. LDM
MOUSE LDM
MOUSE. LIVER EFFECTS 14 DAYS
O MONKEY. TESTICULAR AND LIVER EFFECTS 14 DAYS
• MOUSE FEMALE. CANCER. 2 YEARS
• MOUSE. MALE. CANCER 2 YEARS
• RAT. TESTICULAR EFFECTS 14 DAYS
• RAT (SUCKLING). LDW 5 DAYS
• HAMSTER. UVER EFFECTS. 14 DAYS
• RAT. CANCER. 2 YEARS
\f» RAT. TESTICULAR EFFECTS. 2 YEARS
\O RAT. UVER EFFECTS. 2 YEARS
**• RAT. DEVELOPMENTAL EFFECTS. 20 DAYS DURING GESTATION
~"O RAT. TESTICULAR EFFECTS. 2 YEARS
;*;• RAT. TESTICULAH EFFECTS 60 DAYS
^O HAMSTER. LIVER EFFECTS. 14 DAYS
MOUSE. FERTILITY EFFECTS 106 DAYS (MATING AND POST MATING)
BAT I IIJPB fff^**^t* * * M.*..dh
f • RAT. LIVER EFFECTS. 14 DAYS
\ O RAT (SUCKLING). LD^ 5 DAYS
MOUSE DEVELOPMENTAL EFFECTS. DAYSO-17OF GESTATION
O RAT. TESTICULAR EFFECTS. 60 DAYS
RAT. LIVER EFFECTS. 3 AND 28 DAYS
MOUSE. DEVELOPMENTAL EFFECTS DAY 7 OF GESTATION
MOUSE. DEVELOPMENTAL EFFECTS. DAYSO-17OF GESTATION
-O RAT. UVER EFFECTS. 14 DAYS
• GUINEA PIG. LIVER EFFECTS. 1 YEAR
O MOUSE. FERTILITY EFFECTS. 105 DAYS (MATING AND POST MATING)
• LOAEL
O NOAEL
QUANTITATIVE DATA
WERE NOT AVAILABLE
Fig. 2.2. Effects of dK2-ettaylhexyl)pbthmUte—oral exposure.
-------
Health Effects Summary 11
(mg/m3)
100.000
10.000
1,000
100
10
ACUTE
(£14 DAYS)
LETHALITY
r RAT
INTERMEDIATE
(15-364 DAYS)
QUANTITATIVE
DATA WERE NOT
AVAILABLE
CHRONIC
.(2365 DAYS)
QUANTITATIVE
DATA WERE NOT
AVAILABLE
i MINIMAL RISK FOR EFFECTS O NOAEL
OTHER THAN CANCER
Fig. 2.3. Leveb of significant exposure for di(2-ethylhexyl)phthala(e—inhalation.
-------
ACUTE INTERMEDIATE CHROMC
(iUDAYS) (15-364 DAYS) (2365 DAYS)
DEVELOP REPHO TARGET TARGET REPRO- DEVELOP- TARGET REPHO
t— •
ro
LETHALITY MENTAL DUCTON ORGAN ORGAN DUCTON MENTAL ORGAN DUCTON CANCER !?
(ing 'kg/day)
100.000 r-
10000
1.000
100
10
1
01
001
0001
•
00001
000001
1000001
9 th
|i ' Om(UVER)
~
Ok Oh (LIVER) »'
'V
O
n
K-
o
a
n F
• r (SUCKLING) «r fs (LIVER) 9mM
T T Or(UVER) •- •'
i .. « 4
6 «r(LIVER) If1" ,m
• m •MUVER) »r (LIVER) <•> T
! r
-
-
^ ,0^,
vi/
io-» -
.0--
• LOAEL g GUINEA PIG 1 HAMSTER ! MNMALRBK '^
O NOAEL m MOUSE k MONKEY j LEVEL FOR EFFECTS
h RABBIT M MALE J, OTHER THAN CANCER
r RAT F FEMALE
ESTIMATED
UI'PER BOUND
HUMAN
CANCER
niSK LfcVfLS
Fig. 2.4. Levels of significant exposure for di(2-cthylbexyl)phthalate—oral.
-------
Healch Effects Summary 13
observed following a 1-h exposure at a dose level of 23,670 rng/nt^
(1,457 ppm) (WARF Institute 1976) or a 6-h exposure at a dose level of
600 mg/m3 (37 ppm) (Pegg 1979) No human data were available for either
of these routes of administration.
In an acute oral toxicity study reported by Shaffer et al (1945),
single doses of DEHP were administered by gavage to male Uistar rats.
The median lethal dose (LD50) was estimated to be 30,600 (95% confidence
limits, 20,800 to 45,200) mg/kg.
In other acute oral toxicity studies, Shaffer et al. (1945)
reported an LD50 of 33,900 mg/kg for rabbits, Fatty (1967) reported an
LD50 of 26,000 mg/kg for mice, and Krauskopf (1973) reported an LD50 of
26,300 mg/kg for guinea pigs. In a study by Dostal et al. (1987), DEHP
at a dose of 100 mg/kg was not lethal to either suckling or weanling
rats, whereas a dose of 1,000 mg/kg was lethal to suckling rats. These
findings indicate that DEHP has a low order of acute oral toxicity, but
that it may be more toxic during early postnatal development. Because
DEHP is a component of a variety of plastic consumer goods, there is a
potential for accidental ingestion. However, no human case studies of
this type were available. This suggests that such incidents are not
common or serious occurrences.
2.2.1.2 Systemic/target organ toxicity
Animal studies have shown the liver and testes to be the primary
end points for DEHP toxicity.
Hepatotoxicity (liver). A number of studies in laboratory animals
have demonstrated that oral exposure to DEHP results in adverse hepatic
effects. At high oral dose levels, DEHP causes functional hepatic damage
characterized by morphological changes and alterations in the activity
of hepatic enzyme systems. Several studies have also demonstrated that
DEHP is a peroxisome proliferator, which is of significance because of
an association between peroxisome proliferation and both liver
hyperplasia and liver cancer in laboratory animals. No animal studies
are available on Che hepatic effects of DEHP when administered either by
inhalation or by dermal exposure.
No human data are available on adverse hepatic effects following
inhalation, oral, or dermal exposures. However, Canning et al. (1984)
reported an increase in the number of peroxisomes in the livers of
patients receiving kidney dialysis treatments.
Rhodes et al. (1986) reported that oral and intraperitoneal
administration of DEHP to the marmoset monkey at doses up to 5
mmole/kg/day DEHP (equivalent to 1,950 mg/kg/day) for 14 days did not
induce morphological or biochemical changes in the liver comparable to
those obtained in rats administered the same amount of DEHP.
NTP (1982) performed single-dose and 14-day repeated-dose oral
studies using F344 rats and B6C3F1 mice. In the single-dose mouse study.
groups of five animals per sex were treated with various doses (1.25 to
20 g/kg) of DEHP by gavage. In the repeated-dose mouse study, groups of
five animals per sex were administered DEHP in the feed (6,300 to
100,000 ppm) for 14 days. No adverse hepatic effects were reported
-------
14 Section 2
following either single or repeated doses. These results indicate a
NOAEL of 20,000 mg/kg for acute hepatotoxicity in the mouse.
Lake et al. (1984) treated male Sprague-Dawley rats and Syrian
hamsters with 25, 100. 250, or 1,000 mg/kg/day DEHP orally for 14 days.
Liver weights in the rat were increased significantly at doses of 100
mg/kg and greater. In the hamster, liver weights were increased
significantly only at the 1,000-mg/kg dose. These results indicate a
marked species difference in this parameter of liver toxicity between
the rat (NOAEL - 25 mg/kg; LOAEL - 100 mg/kg) and the hamster (NOAEL -
250 mg/kg; LOAEL - 1,000 mg/kg).
F. E. Mitchell et al. (1985) evaluated acute and intermediate
hepatotoxicity in rats fed DEHP at doses of 50, 200, or 1,000 mg/kg/day
for 3, 7, 14, or 28 days or 9 months. Histological and biochemical
alterations were observed at all doses tested (including 50 mg/kg, the
LOAEL) for all acute (3, 7, and 14 days) and intermediate (28 days and 9
months) dosing regimens. Mitchell et al. (1985) regarded the liver
effects observed following the treatment with DEHP to be a compensatory
phenomenon rather than a pathological event. However, ATSDR considers
the effects observed in this study to be adverse rather than adaptive.
Dostal et al. (1987) investigated the relative sensitivity of
suckling rats as compared to adults to the effects of DEHP. Five daily
oral doses of 0, 10, 100, 1,000, or 2,000 mg/kg DEHP were administered
to male Sprague-Dawley rats beginning at 6, 14, 16, 21, 42, and 86 days
of age. Twenty-four hours after the last dose, rats were sacrificed, and
plasma cholesterol, triglyceride levels, and the activities of the
hepatic peroxisomal enzymes, palmitoyl CoA oxidase and carnitine
acetyltransferase, were determined. Relative liver weight (g/100 g body
wt) was increased by 30 to 58% at 1,000 mg/kg in all age groups. Smaller
but significant increases in relative liver weights were also observed
at doses of 100 mg/kg (LOAEL) in all but the 1-week-old rats. Absolute
liver weight was increased at doses of 100 mg/kg or more in all but the
1-week-old rats.
In a study by BIBRA (1987), Cynomolgus monkeys (2 males/group) were
administered 0, 100, or 500 mg/kg DEHP by gavage for 25 days. There were
no treatment-related changes in relative liver weight, palmitoyl-CoA
oxidation, carnitine acetyltransferase, or lauric acid 11- and 12-hydro-
zylation. Histological and electron-microscopic examination of the
livers revealed no treatment-related effects.
Carpenter et al. (1953) fed groups of 23 to 24 guinea pigs of each
sex diets containing 0, 400, or 1,300 ppm DEHP for a period of 1 year.
No treatment-related effects were observed on mortality, body weight,
kidney weight, or gross pathology and histopathology of the liver. No
changes in absolute liver weight were observed. However, statistically
significant increases in relative liver weights were observed in both
groups of treated females (19 and 64 ng/kg/day). Carpenter et al. (1953)
concluded that despite the significant increase in liver weights of the
high-dose females, 64 mg/kg/day was a NOAEL. However, ATSDR has
determined 19 mg/kg/day to be a LOAEL. based on the statistically
significant increases in relative liver weights of both groups of
treated females.
-------
Health Effects Summary 15
In a 2-year NTP (1982) bioassay of DEHP, male F344 rats were fed
diets which contained 0 (control). 6,000. or 12.000 ppm DEHP. This was
equivalent to doses of 0. 322. and 674 mg/kg/day DEHP. At the conclusion
of the study, the rats were sacrificed, and histological examinations
were performed. Hepatic tumors and peroxisome proliferation were
observed at the high- and low-dose levels; therefore, under the
conditions of this study the LOAEL was 322 mg/kg/day.
Jacobson et al. (1977) evaluated the effects of DEHP solubilized as
a result of leaching from polyvinyl chloride blood bags on the hepatic
function and histology of rhesus monkeys undergoing chronic blood
transfusions. The average cumulative amount of DEHP infused over 1 year
was 0.0584 mg/kg/day. Exposure to DEHP resulted in abnormal hepatic
scintillation liver scans and abnormal sulfobromophthalein (BSP)
clearance in four of seven animals, and abnormal morphologic changes in
liver histology in six of seven animals. Abnormal liver changes included
vacuolization of Kupffer cells, foci of parenchymal necrosis, chronic
inflammatory cell infiltrates, and binucleate cells.
Testicular toxicity. DEHP has also been shown to induce testicular
toxicity in numerous animal studies. Such effects, however, are
generally considered as evidence of reproductive toxicity and are
therefore discussed in detail in the section on reproductive toxicity
(Sect. 2.2.1.4).
2.2.1.3 Developmental toxicity
A number of studies in laboratory animals have demonstrated that
oral exposure to DEHP induces developmental toxicity. No animal studies
are available on the developmental toxicity of DEHP when administered
either by inhalation or by dermal exposure. No human data are available
on adverse developmental effects following inhalation, oral, or dermal
exposures.
Nakamura et al. (1979) evaluated teratogenicity in wy-Slc x CBA
mice administered a single oral dose of 0.05, 0.1, 1.0, 2.5, 5.0, or
10.0 raL/kg DEHP on day 7 of gestation. Based on a density of 0.9861 for
DEHP (IARC 1982), these doses were equivalent to 49.3, 98.6, 986, 2,455,
4,930, and 9,860 mg/kg. DEHP resulted in a decrease in body weight of
live fetuses at doses of 49.3 mg/kg (the LOAEL) and higher. At doses of
98.6 mg/kg and higher, DEHP also resulted in deformed or dead fetuses.
Wolkowski-Tyl et al. (1984a) fed timed pregnant Fischer 344 rats
nominal doses of 0, 5,000, 10.000, 15,000, or 20,000 ppm DEHP in the
diet on days 0 to 20 of gestation. Based on food consumption data, this
was equivalent to doses of approximately 0, 357. 666. 856, and 1.055
mgAg/day, respectively. Maternal toxicity, as evidenced by
significantly reduced body weight gains in the dams receiving 10,000.
15.000, and 20,000 ppm of DEHP, was observed. The number and percentage
of resorptions, nonlive fetuses, and nonlive malformed fetuses were
increased in a dose-related manner, with a statistically significant
increase in all three parameters in the high-dose group when compared
with the controls. The number of live fetuses per litter exhibited a
dose-related decrease, with the high-dose group significantly lower than
the controls. Among the live neonates, there was a dose-related decrease
in the body weights of both sexes, with a statistically significant
-------
16 Section 2
decrease in fetal weight at all doses tested. There was also a
significant dose-related trend in the percentage of malformed fetuses
per litter. However, there were no significant pairwise comparisons in
the number of fetuses malformed per litter. Under the conditions of this
study, DEHP produced significant developmental toxicity, exhibited as
decreased fetal weight, at doses of 10,000, 15,000, and 20,000 ppm.
ATSDR has concluded that the LOAEL was 5,000 ppm (357 mg/kg/day) and
that a NOAEL could not be determined from this study.
Wolkowski-Tyl et al. (1984b) reported that DEHP was developmentally
toxic to CD-I mice exposed during the entire gestational period (days 0
through 17) at dose levels that produced significant maternal and other
fetal toxicity. The mice received diets containing 0, 250, 500, 1,000,
or 1,500 ppm DEHP. Based on food consumption data, this was equivalent
to approximately 0, 44, 91, 191, or 292 mg/kg/day. Major malformations
included external, visceral, and skeletal effects, observed at doses of
91 mg/kg/day (the LOAEL) and higher. Under the conditions of this study
the NOAEL was determined to be 250 ppm (44 mg/kg/day) DEHP in the diet,
if no significant maternal or fetal toxicity had been observed.
2.2.1.4 Reproductive toxicity
A number of studies in laboratory animals have demonstrated that
oral exposure to DEHP induces adverse testicular effects, characterized
by decreased organ weights and histological changes in the seminiferous
tubules. Hono(2-ethylhexyl)phthalate (HEHP), a monoester metabolite
formed in the gastrointestinal tract, has also been reported to produce
testicular damage (Oishi and Hiraga 1980). No animal studies are
available on the testicular effects of DEHP when administered either by
inhalation or by dermal exposure. No human data are available on adverse
testicular effects following inhalation, oral, or dermal exposures.
Persons who may be at risk from reproductive and developmental
toxicity from DEHP exposure are individuals receiving hemodialysis
treatments. DEHP is frequently used as a plasticizer for medical vinyl
products, such as blood bags, intravenous administration sets, and
hemodialysis tubing. Trebbin (1979) discussed four case studies in which
pregnant women with chronic renal failure underwent hemodialysis. These
women successfully conceived and concluded the pregnancy with the birth
of apparently healthy, normal infants. Bailey (1977) discussed the
effect of hemodialysis on men with chronic renal failure. Prior to
treatment, the testicles become soft and atrophic, spermatogenesis
ceases, some Leydig cells are lost, plasma testosterone levels decrease,
and libido is lost. Hemodialysis resulted in restoration of their sex
drive and spermatogenesis. Conception and completely successful
pregnancies in their spouses followed. This suggests that intravenous
exposure to DEHP--if this was the plasticizer involved in these cases--
may not pose significant risks of reproductive toxicity in male dialysis
patients.
Sjoberg et al. (1985c) evaluated the testicular toxicity of 1.0
g/kg/day DEHP in 25-, 40-, and 60-day-old rats. Following 14 daily oral
doses, testicular damage was observed in the 25-day-old group but not in
the 40- or the 60-day-old group. These findings suggest an age-dependent
susceptibility to the testicular effects of DEHP. However, in a follow-
up study by Sjoberg et al. (1985b), age-dependent testicular effects
-------
Heal eh Effects Summary 17
were not evident following intravenous DEHP administration. This is
likely because of the differences in the route of administration. DEHP
is extensively converted to MEHP in the gastrointestinal tract HEHP
would not be formed to the same extent after intravenous administration.
Rhodes et al. (1986) evaluated the testicular toxicity of DEHP Ln
marmoset monkeys administered (orally or intraperitoneally) doses up to
5 mmole/kg/day (1,950 mg/kg/day). Significant biochemical and
morphological changes were not evident, even at the highest dose tested
(1,950 mg/kg/day, the NOAEL).
Aearwal et al. (1986) evaluated the reproductive (testicular)
toxicity of DEHP in sexually mature male F344 rats. DEHP was
administered in the diet at levels of 0, 320 1.250 5.000. o,: 20 000
ppm (equivalent to 0. 16, 62.4. 250. or 1,000 ^As/day) for 60 days.
Reduced body weight, testicular weight, and epididymal weight were
observed at doset of 250 mg/kg/day (the LOAEL) and higher but not at
doses of 62.5 mgAs/day (the NOAEL) and lower. A decrease in various sex
hormone levels was also observed at the two highest DEHP levels, with
degenerative testicular alterations and reduced epidiaymal sperm count
and motility noted only at the highest level.
In a study performed by Reel et al. (1982), groups of mice were fed
0 100, 1.000. or 3.000 ppm DEHP in the diet during a 7-day premating
oeriod and a 98-day cohabitation period. These dose levels are
equivalent to approximately 0, 13, 130, and 390 mg/kg/day. Fertility was
completely suppressed in the high-dose group and significantly
(P < 0 05) reduced in the intermediate-dose group. DEHP was a
reproductive toxicant in male and female CD-I mice, as evidenced by a
decreased fertility index, decreased number of litters decreased number
of live pups, and a lower proportion of live pups per litter. «"*««"•
conditions of this study, the NOAEL was determined to be 13 mg/kg/day.
and the LOAEL was determined to be 130 mg/kg/day.
In a 2-year NTP (1982) bioassay of DEHP (previously discussed).
male F344 rats were fed diets which contained 0 (control), 6,000, or
12 000 ppm DEHP. This was equivalent to doses of 0. 322, and &74
mg/kg/day DEHP. At the conclusion of the study the rats were sacrificed.
and historical examination revealed seminiferous tubule degeneration
of the testes in 1/43 controls (2.3%). 2/44 low-dose males (4.5%) and
43/48 high-dose males (89.6%). Therefore, under the conditions of this
study the LOAEL was 674 mg/kg/day and the NOAEL was 322 mg/kg/day.
2.2.1.5 Genotoxlcity
DEHP has been tested extensively in short-term genotoxicity assays
because of its reported carcinogenicity and its Inclusion in the
collaborative study program of the International Program on Chemical
Safety DEHP has not been shown to be mutagenic in most microbial and
mammalian assay systems. In addition, the majority of the data also
suggest that MEHP and 2-ethylhexanol. the major metabolites of DEHP. are
not mutagenic.
-------
18 Section 2
2.2.1.6 CareInogenic ity
There were no available data on the potential carcinogeniclty of
DEHP in humans exposed via inhalation, oral, or dermal exposures.
Neither were data available on inhalation or dermal exposures in
experimental animals.
The carcinogenicity of DEHP has been tested in a bioassay using
Fischer 344 rats and B6C3F1 mice (NTP 1982, Kluwe et al. 1982a,b).
Croups of 50 rats per sex were fed diets which contained 0 (control),
6.000, or 12,000 ppm DEHP, and groups of 50 mice per sex were fed diets
which contained 0 (control), 3,000, or 6,000 ppm of DEHP (purity 95.5%)
for 103 weeks. These dose levels were equivalent to 0, 322, and 674
mg/kg/day for the male rats; 0, 394, and 774 mgAg/day for the female
rats; 0, 672, and 1,325 mgAg/day for the male mice; and 0, 799. and
1,821 mgAg/day for the female mice, respectively. A 1- to 2-week
nontreatment observation period was allowed after the termination of
exposure, after which all survivors were sacrificed and examined both
grossly and microscopically. Histopathological examinations were
conducted on all animals sacrificed or discovered dead (if not precluded
by autolysis).
A summary of the incidence of hepatocellular carcinomas and
neoplastic nodules observed in the rats is presented below:
Dose
(ppm) Males Females
0 3/50 0/50
6,000 6/49 6/49
12,000 12/49 13/50
In female rats, the incidence of hepatocellular carcinomas at the
higher dose was greater than that in controls by pairwise comparisons,
and there was a significant (P < 0.05) dose-related trend. A dose-
related trend was also observed for hepatocellular carcinomas in the
males, but pairwise comparisons did not indicate a statistically
significant increase.
A summary of the incidence of hepatocellular carcinomas and
neoplastic nodules observed in the mice is presented below:
Dose
(ppm) Males Females
0 14/50 1/50
3,000 25/48 12/50
6,000 29/50 18/50
In mice, there was a statistically significant increase in the
incidence of hepatocellular carcinoma in both the high-dose (P - 0.022)
males and high-dose females (P < 0.001) when compared to the controls. A
significant (P < 0.05) dose-related trend of increasing hepatocellular
-------
Health Effaces Summary 19
carcinoma was also observed for Che mice of both sexes. Metastases of
the hepatocellular carcinomas were observed in the lungs of 12 treated
males and 8 treated females.
Northrup et al. (1982) have questioned the validity of the NTP
bioassay of DEHP. suggesting that the maximum tolerated dose (MTD) was
exceeded in both species because body weight gain was depressed by more
than 10% in several of the treatment groups. However, Kluwe et al.
(1983) have pointed out that the 10% weight differential is only a
guideline and that the primary reason for not exceeding the estimated
HTD is to avoid excessive early mortality which might prevent tumor
development, and to avoid pathological lesions other than neoplasia. In
this study, survival was not adversely affected by DEHP, and statistical
analyses failed to demonstrate any correlation between the occurrence of
nonneoplastic tissue lesions (testicular atrophy in the high-dose male
rats and mice and pituitary hypertrophy in the high-dose male rats) and
the development of hepatocellular tumors in rats and mice of both sexes
(Kluwe et al. 1982a,b 1983).
Northrup et al. (1982) also questioned the significance of the
DEHP-induced increase in liver tumors in the NTP bioassay because of
intralaboratory variations in incidences of liver tumors in controls in
bioassays conducted simultaneously in the same rooms as the DEHP study.
They reported that statistical analysis indicated that there was a
significantly lower incidence of tumor-bearing animals among the female
mice used in the DEHP control group.-In addition, they found that the
number of tumor-bearing male or female mice in the DEHP treatment groups
did not substantially differ from the average for all pooled controls.
They also found that in male rats, the incidence of tumor-bearing
animals had a highly negative dose response (? < 0.001), and female racs
had a marginally positive dose response (? - 0.045) when paired to the
pooled controls. However, Kluwe et al. (1983) reported that there was
relatively little variation in liver tumor incidences among NTP
historical controls, and that the data reported in the concurrent study
controls were quite similar to the historical controls. In addition,
Kluwe et al. (19B2a,b) have reported that replacement of the concurrent
study controls with concurrent laboratory controls or pooled controls
from bioassays performed at approximately the same time did not alter
the statistical significance of the DEHP effects. Replacement of the
concurrent study controls with historical controls strengthened the
statistical significance of the difference in liver tumor incidence.
In a draft document, the EPA (1986a) has classified DEHP as a
probable human carcinogen, B2, according to the criteria in the EPA
Guidelines for Carcinogen Risk Assessment (EPA 1986a). Evidence on
potential carcinogenicity from animal studies is "Sufficient," and
evidence from human studies is "No Data" (EPA 1986a).
Various types of mathematical extrapolation models can be fit to
the NTP carcinogenesis bioassay data of DEHP in F344 rats and B6C3F1
mice. Different high-to-low dose extrapolation methods may give a
reasonable fit to the observed data but may lead to differences in the
estimates of risk at low doses. The choice of the model depends largely
on the assumed mechanism of action If DEHP is a genotoxic carcinogen,
it is capable of direct reaction with genetic material. Under these
-------
20 Section 2
circumstances, the dose-response curve is expected to be linear at low
doses. However, if DEHP acts by an epigenetic mechanism, the dose-
response curve may be subject to individual tolerances, with each
individual having a specific threshold below which no effect would
occur, and the aggregation of these individual tolerances would fit a
specific statistical distribution for the general population. The issue
of whether DEHP acts by a genotoxic or nongenotoxic mechanism has not
been resolved at the present time. In keeping with the EPA Risk
Assessment Guidelines (EPA 1986) and the OSTP Cancer Principles (OSTP
1985), the choice of the low-dose extrapolation method is governed by
consistency with current understanding of the mechanism of
carcinogenesis and not solely on goodness-of-fit to the observed data.
When data are limited, and when uncertainty exists regarding the
mechanisms of carcinogenic action, the OSTP Principles suggest that
models or procedures which incorporate low-dose linearity are preferred
when compatible with the limited information available. The EPA
Guidelines recommend that the linearized multistage procedure be used in
the absence of adequate information to the contrary.
In evaluating the potential carcinogenicity of DEHP by the EPA (EPA
1987c), slope factors were estimated by fitting the liver tumor data in
male and female rats and mice with the linearized multistage model. The
unit risk value is estimated to be 4.0 x 10"? for drinking water
containing 1 pg/L DEHP. The drinking water concentrations of DEHP
corresponding to lifetime excess cancer risks of 10'4, 10'5, and 10'6
are 0.3, 0.03, and 0.003 mg/L, respectively.
Computations of the unit risk for the experimental animals were
made by the Consumer Product Safety Commission (CPSC) (1985) assuming
the generalized multistage model of carcinogenesis. Risk estimates were
performed using the Global 83 computer program of Howe and Crump in its
unrestricted form. The lifetime excess risk to the experimental animals
per mg/kg/day for hepatocellular carcinomas and neoplastic nodules
following exposure to DEHP was estimated by CPSC (1985) as follows:
Lifetime animal risk estimates
[(mg/kg/day)-1]
Species
Rat
Mouse
Sex
Male
Female
Male
Female
Maximum
likelihood
estimate
0.000154
0.000285
0.000678
0.000545
Upper 95%
confidence
limit
0.000471
0.000523
0.001122
0.000752
These estimated animal risks can be converted to human risk
estimates by applying animal-to-human extrapolation factors that are
based on comparative body surface areas of humans vs rats and mice.
These extrapolation factors are 5.6 for rats and 12.8 for mice. The
resulting human risk extrapolations are:
-------
Health Effaces Summary 21
Lifetime human risk estimates
«nig/kg/day)-1]
Animal on which based
Species
Rat
Mouse
Sex
Male
Female
Male
Female
Maximum
likelihood
estimate
0.000862
0.00160
0.00868
0.00698
Upper 95%
£ • 1
limit
0.00264
0.00293
0.0144
0.00963
Note that the most conservative (highest) risk estimate derived by
CPSC (1985) of 0.0144 is virtually identical to the 0.015 potency factor
derived by EPA (1986b). However, EPA based its risk estimates on
hepatocellular carcinomas in female rats, whereas the highest risk
estimate reported by CPSC (1985) is based on liver tumors in the male
mouse.
Because of what appeared to be excessive reported food consumption
by the mice (8.1 to 8.4 g/day), the CPSC (1985) converted the dose data
for mice to a milligram per kilogram basis using an estimated average
daily consumption of 3.9 g per animal. The CPSC's rationale for this was
that the food fed the mice was not in pelleted form, and some of the
reported food consumption was due to substantial scattering. The risk
estimate performed by EPA did not incorporate an adjustment for food
consumption; therefore, the lifetime extra carcinogenic risk may be
underestimated.
2.2.2 Biological Monitoring as a Measure of Exposure and Effects
As discussed in Sect. 4, Toxicological Data, DEHP (or its
metabolites) is well absorbed in the gastrointestinal tract following
oral exposure. Once absorbed, it is widely distributed in the body, with
the liver being the major repository organ. As discussed in the toxicity
review of Sect. 4, the major target organs of DEHP appear to be the
liver and testes.
DEHP and its metabolites. MEHP and 2-ethylhexanol. can be measured
in urine and blood in order to confirm prior recent exposure. Monitoring
urine for DEHP will not be a good marker for prior DEHP exposure because
of poor solubility. There is scant information on the relationship
between DEHP uptake and its elimination in the urine in humans. At
present, there are no data available for assessing the potential
correlation between the values obtained with these measurements and the
toxic effects observed in experimental animals. Also, because DEHP is
rapidly metabolized and excreted, these methods are not good indicators
of previous exposures. In addition, because MEHP and 2-ethylhexanol
undergo extensive metabolism, they are also poor markers of previous
exposure.
-------
22 Section 2
2.2.3 Environmental Levels as Indicators of Exposure and Effects
2.2.3.1 Levels found in the environment
There are no available case studies or epidemiological
investigations that suggest that levels of DEHP found in the
environmental media are associated with significant human exposure or
adverse health effects.
2.2.3.2 Human exposure potential
No data have been located to suggest that there are chemical-
specific issues relevant to estimating the body-dose, tissue levels, or
health effects from DEHP concentrations in soil, water, or food.
2.3 ADEQUACY OF DATABASE
2.3.1 Introduction
Section 110 (3) of SARA directs the Administrator of ATSDR to
prepare a toxicological profile for each of the 100 most significant
hazardous substances found at facilities on the CERCLA National
Priorities List. Each profile must include the following content:
"(A) An examination, summary, and interpretation of available
toxicological information and epidemiologic evaluations on a
hazardous substance in order to ascertain the levels of
significant human exposure for the substance and the
associated acute, subacute, and chronic health effects.
(B) A determination of whether adequate information on the health
effects of each substance is available or in the process of
development to determine levels of exposure which present a
significant risk to human health of acute, subacute, and
chronic health effects.
(C) Where appropriate, an identification of toxicological testing
needed to identify the types or levels of exposure that may
present significant risk of adverse health effects in humans."
This section identifies gaps in current knowledge relevant to
developing levels of significant exposure for DEHP. Such gaps are
identified for certain health effect end points (lethality,
systemic/target organ toxicity, developmental toxicity, reproductive
toxicity, and carcinogenicity) reviewed in Sect. 2.2 of this profile in
developing levels of significant exposure for DEHP, and for other areas
such as human biological monitoring and mechanisms of toxicity. The
present section briefly summarizes the availability of existing human
and animal data, identifies data gaps, and summarizes research in
progress that may fill such gaps.
Specific research programs for obtaining data needed to develop
levels of significant exposure for DEHP will be developed by ATSDR. NTP,
and EPA in the future.
-------
Health Effects Summary 23
2.3.2 Health Effect End Points
2.3.2.1 Introduction and graphic summary
The availability of data for health effects in humans and animals
is depicted on bar graphs In Figs. 2.5 and 2.6, respectively.
The bars of full height indicate that there are data to meet at
least one of the following criteria:
1. For noncancer health end points, one or more studies are available
that meet current scientific standards and are sufficient to define
a range of toxicity from no-effect levels (NOAELs) to levels that
cause effects (LOAELs or FELs).
2. For human carcinogenicity, a substance is classified as either a
"known human carcinogen" or a "probable human carcinogen" by both
EPA and IARC (qualitative), and the data are sufficient to derive a
cancer potency factor (quantitative).
3. For animal carcinogenicity, a substance causes a statistically
significant number of tumors in at least one species, and the data
are sufficient to derive a cancer potency factor.
4. There are studies which show that the chemical does not cause this
health effect via this exposure route.
Bars of half height indicate that "some" information for the end
point exists, but does not meet any of these criteria.
The absence of a column indicates that no information exists for
that end point and route.
2.3.2.2 Description of highlights of graphs
As shown in Figure 2.5, there were no adequate human data for DEHP.
There were no available case studies or epidemiological reports
providing information on exposure levels associated with any reported
effects.
In the data based on animal studies, as shown in Figure 2.6, there
were no data available via the dermal route. The few studies conducted
via inhalation were not sufficient to assess the effects
(lethality/decreased longevity) that were being tested. Studies using
the oral route of administration were adequate for acute, intermediate,
and chronic exposures to DEHP. with the liver being the most sensitive
target organ. The NTP carcinogenicity bioassay was also adequate for
assessing carcinogenicity. Data on lethality, developmental toxicity.
and reproductive toxicity following oral administration were considered
adequate.
2.3.2.3 Summary of relevant ongoing research
A search of the NIH CRISP (Computerized Retrieval of Information on
Scientific Projects) database revealed that several investigations of
the role of peroxisomal proliferation in DEHP-induced carcinogenicity
-------
HUMAN DATA
ORAL
01
r>
rt
>-.
§
J
^ SUFFICIENT
INFORMATION*
J
^ SOME
INFORMATION
NO
INFORMATION
INHALATION
DERMAL
LETHALITY ACUTE INTERMEDIATE CHRONIC DEVELOPMENTAL REPRODUCTIVE CARCINOOEMCITV
Z / TOXICITY TOXICITY
SYSTEMIC TOXICITY
'Sufficient information exists to meet at least one of the criteria for cancer or noncancer end points.
Fig. 2.5. Availability of information on health effects of d«2-«lhylhexyl)phih«late (human data).
-------
ANIMAL DATA
V SUFFICIENT
'INFORMATION*
V SOME
'INFORMATION
NO
INFORMATION
INHALATION
DERMAL
LETHALITY
ACUTE
INTERMEDIATE
-
CHRONIC DEVELOPMENTAL REPRODUCTIVE CAHCINOOENICITY
TOXICITT TOXICITV
Cki
t-4
n
(b
n
rt
to
SYSTEMIC TOXICITY
'Sufficient information exists to meet at least one of the criteria for cancer or noncancer end points.
Fig. 2.6. Availability of information on health effects of di(2-etnylhexyl)phthalate (animal data).
-------
26 Section 2
are being conducted by researchers at the National Institutes of Health.
These studies are summarized in Table 2.1. Investigations of DEHP are
being conducted by other research groups such as the Chemical Industry
Institute on Toxicology (CUT). In addition, a bioassay for the
metabolite 2-ethylhexanol is being planned by the National Toxicology
Program (NTP).
2.3.3 Other Information Needed for Human Health Assessment
2.3.3.1 Fharmacokinetics and mechanisms of action
The mechanism of DEHP-induced careinogenieity is currently not well
understood. Most of the evidence from genotoxicity assays indicates that
DEHP is nongenotoxic. DEHP appears to belong to a class of carcinogens
that induces proliferation of peroxisomes and liver cells. A
hypothetical, but unproven, mechanism for DEHP-induced carcinogenesis is
that peroxisome proliferation causes an imbalance in reactive oxygen
species that somehow initiates the carcinogenic process. In the NTP
bioassay, excess cancers were found only in the liver, the site of the
greatest peroxisome activity.
Additional research is necessary to elucidate the exact mechanism
of action responsible for the carcinogenic effects of DEHP.
A second area currently being investigated is that of the role of
zinc in the testicular effects of DEHP. DEHP-induced testicular injury
is accompanied by a decrease in the zinc content of the testes and an
increase in the urinary excretion of this element. At present, the
significance of these findings is unclear. Further research is necessary
to elucidate the role of DEHP in testicular effects.
Additional metabolic and pharmacokinetic data are needed for the
purpose of interspecies and high-to-low dose extrapolation of the toxic
effects of DEHP.
2.3.3.2 Monitoring of human biological samples
There are two methods to test for human exposure to DEHP. These
methods involve analysis of blood or urine for the presence of DEHP
and/or its major metabolites, HEHP and 2-ethylhexanol. Because DEHP is
rapidly metabolized and excreted (and apparently does not
bioaccumulate), these methods are appropriate only for measuring recent,
prior exposure.
2.3.3.3 Environmental considerations
Current methodologies for assessing the levels of DEHP in the
environment are adequate. DEHP is relatively nonvolatile at ambient
temperatures, so that partitioning into the atmosphere is extremely
limited. Because DEHP is readily degraded under aerobic conditions,
environmental accumulation of levels toxic to humans is not expected.
This has been confirmed by environmental monitoring and analytical
studies conducted under controlled conditions.
-------
Health Effects Summary 27
Table 2.1. Summary of ongoing research"
Principal
investigator
Melnick, R. L.
Institutional
affiliation
NIEHS
Description
of research
Investigate role
of peroxisomal
production in
DEHP-induced
hepatotoxicity
Data
deficiency
Mechanism of
DEHP-induced
hepatotoxicity
Melnick, R. L. NIEHS
Determine the role
of zinc in the
pathophysiology of
DEHP-induced
reproductive
effects
Mechanism of
DEHP-induced
reproductive
toxicity
Ward, J. M. NCI
Diwan, B. NCI
Investigate tumor
initiation and
promotion by DEHP
Investigate tumor
promotion by DEHP
Mechanism of
DEHP-induced
carcinogemcity
Mechanism of
DEHP-induced
carcinogemcity
"Adapted from NIH CRISP database.
-------
29
3. CHEMICAL AND PHYSICAL INFORMATION
3.1 CHEMICAL IDENTITY
The chemical formula, structure, synonyms, trade names, and
identification numbers for DEHP are listed in Table 3.1.
3.2 CHEMICAL AND PHYSICAL PROPERTIES
Important physical and chemical properties of DEHP are listed in
Table 3.2.
-------
30
Section 3
T.Me 3.1. Chemical identity of dK2-etbylbexyl)pfathalaMa
Chemical name
Synonyms
Trade name*
Chemical formula
Wuwesser line notation
Chemical structure
Di(2-ethylhexyl) phthalate
1.2-Bcnzenedicarboxylic acid, bis(2-ethylhexyl) ester
Phthahc acid. b»(2-ethylhexyl) ester
BEHP
1.2-Benzenedicarboxylic and. bis(ethylhexyl) ester
But 2-ethylhexyl )- 1 ,2-benzenedicarboxylate
Bis(2-ethylhexyl) phthalate
DEHP
Di(2-ethylbexyl) orthophthalate
Di(ethylhexyl) phthalate
Dioctyl phthalate
OOP
2-Eihylhexyl phthalate
Octyl phthalate
Phthalic acid, dioctyl ester
Bisoflex 81. Bisoflex OOP, Compound 889.
DAF 68. Ergoplast FDO, Evtplast 80.
Eviplast 81. Flexunel. Flexol OOP.
Good-Rite GP 264. Hatcol OOP. Kodaflex OOP,
Mollan O. Nuoplaz OOP, Octoil.
Palatinol AH. Palatinol OOP. Pittsburg PX-138.
Reomol OOP. Reomol D 79P. Sicol ISO.
Staflex OOP, Truflex OOP. Vestmol AH.
Vmicizer 80. Witcizer 312
4Y2&1OVRBV01Y4&2
COOCH2CH(C2H8)(CH2)!,CH3
Identification numbers
CAS Registry No. 117-81 -7
NIOSH RTECS No. TI0350000
EPA Hazardous Waste No. UO28
OHM-TADS No. 7216693
DOT/UN/NA/IMCO Shipping No NAI693
STCC No.
Hazardous Substances Data Bank No 339
National Cancer Institute No. CS2733
'References. HSDB 1987. I ARC 1982
-------
Chemical and Physical Information 31
Tabk 3.2. Physical and chemical properties of dK2-etbylbexyl)pfetkalate
Property
Molecular weight
Color
Physical state
Odor
Odor threshold
Melting pout
Boiling point
Autoignition temperature
Solubility
Water
Organic solvents
Density
Vapor density
Partition coefficients
Octanol-water (log !(„)
Soil-organic carbon-water
<*.), mL/g
Vapor pressure
At 200°C
At 25°C
Henry's law constant
Refractive index
Flash point (open cup)
Flammable limit
Conversion factor
Value
39054
Colorless
Liquid
Slight
NDa
-50°C
385°C at 760 mm Hg
390-C
0 285 mg/L at 24° C
0 40 mg/L at 20°C
0 34 mg/L at 2S°C
Miscible with mineral
oil and hexane;
soluble in most
organic solvents
09861 al20°C
160 (air - 1)
488
ND"
1 32 mm Hg
34 X I0~~mm Hg
1 X 10~4atm-mJ/mol
1 4836 at 20° C
215°C
0 3% at 245°C
1 ppm — 1 5 94 mg/m
References
ACCIH 1986
CHRIS 1978
IARC 1982
CHRIS 1978
Clayton and
Clayton 1981
Verschueren 1983*
NFPA 1978*
Verschueren 1983*
Branson 1980
CMA 1983
ACGIH 1986
IARC 1982
Clayton and
Clayton 1981
b
Fnssel 1956
Wolfe et al 1980*
IARC 1982
NFPA 1978*
NFPA 1978*
Clayton and
Clayton 1981
aND - no data.
*Cited in HSDB 1987.
Source Adapted from HSDB 1987
-------
33
4. TOXICOLOGICAL DATA
4.1 OVERVIEW
Phchalate esters such as DEHP are well absorbed from che
gastrointestinal tract following oral administration. Hydrolysis to the
corresponding monoester metabolite. MEHP. with release of an alcoholic
substituent, 2-ethylhexanol, largely occurs prior to intestinal
absorption. Once absorbed, DEHP and MEHP are widely distributed in the
body, with the liver being the major, initial repository organ.
Clearance from the body is rapid, and there is only a slight cumulative
potential. DEHP is converted principally to polar derivatives of the
monoesters by oxidative metabolism prior to excretion. In general, there
are only quantitative differences in the metabolite profiles of phase 1
oxidations between species. It had been previously thought that none of
the five major metabolites in rats, all of which involve oxidation of
methyl groups to carboxylic acid groups, occurred in man. Recently,
however, it has been shown that one of the two major metabolites in man
involves oxidation of a terminal methyl group to the corresponding acid.
However, there appears to be major interspecies differences in the
phase 2 metabolism of DEHP. Primates (man, African green monkey, and
marmoset) glucuronidate DEHP at the carboxylate moiety following
hydrolysis of a single ester linkage. Rats can glucuronidate MEHP, but
high 0-glucuronidase activity apparently converts this phase 2 conjugate
back to MEHP. Mice appear to be unable to glucuronidate the monoester
metabolite and, thus, oxidize the residual alkyl chain instead to
various hydroxy, ketone, and carboxylate derivatives. The major route of
DEHP elimination from the body is urinary excretion. The relationship of
DEHP pharmacokinetics to its toxicological actions is unknown at the
present time, largely due to a lack of elucidated mechanisms of toxic
action.
A number of studies have been conducted to investigate the acute
toxic effects of DEHP. When administered by the oral, intraperitoneal,
intravenous, and inhalation routes. DEHP has a low order of acute
toxicity.
The target organs for DEHP appear to be the liver and testes. DEHP
has been found to induce morphological and biochemical changes in the
liver of exposed rodents at relatively high dose levels. Similar effects
have been reported for a number of chemicals which induce hepatic
xenobiotic metabolizing capabilities. The testicular effects of DEHP are
characterized by a decrease in relative organ weight and damage to the
seminiferous tubules. Similar effects have been reported in animals
treated with MEHP, a major metabolite of DEHP.
Studies in rats and mice suggest that DEHP is developmentally
toxic. In the rat, a variety of congenital abnormalities have been
-------
34 Section 4
observed In the offspring of DEHP-treated dams. In the mouse, the
developing nervous system appears to be the major target site, producing
exencephaly and splna blflda. DEHP Is a reproductive toxicant In male
and female mice, resulting In reduced fertility and both production of
fewer litters by breeding pairs and decreased litter size.
A large database exists on the genotoxicity of DEHP. DEHP has been
subjected to extensive testing in bacterial and both In vitro and in
vivo mammalian assay systems. The weight of the evidence suggests that
DEHP is not mutagenic in bacterial or mammalian test systems.
In a study by NTP, DEHP induced an increase in the Incidence of
hepatocellular tumors in both sexes of Fischer 344 rats and B6C3F1 mice
There was a trend toward increasing numbers of tumors with Increasing
doses. In a draft report, the EPA has concluded that DEHP is a probable
human carcinogen classified as weight-of-evidence Group B2.
4.2 TOXICOKINETICS
4.2.1 Absorption
Studies of the absorption of DEHP following an oral dose Indicate
that the parent compound is initially hydrolyzed by a nonspecific lipase
in the gastrointestinal tract to produce mono(ethyIhexyl) phthalate
(MEHP) and 2-ethylhexanol. MEHP is readily absorbed from the
gastrointestinal tract. There is evidence for very low level absorption
of DEHP through the skin. Although quantitative information is not
available, animal studies indicate DEHP is absorbed by the lungs.
4.2.1.1 Inhalation
Human. No data are available on the absorption of DEHP by humans
exposed via inhalation.
Animal. No quantitative data are available on the absorption of
DEHP by animals exposed via inhalation. However, acute inhalation
toxicity studies with rats performed by the WARF Institute (1976) and
Pegg (1979) have demonstrated that DEHP is absorbed by the lung. In a
chronic inhalation study, Schmezer et al. (1988) have shown absorption
of saturated vapors of DEHP by hamsters.
4.2.1.2 Oral
Human. Information on the oral absorption of DEHP in humans is
limited. Shaffer et al. (1945) reported that a single oral dose of
10,000 mg of DEHP by a human subject was recovered as an unspecified
phthalate equivalent in the urine after 24 h. Approximately 4.5% of the
administered dose was recovered. Schmid and Schlatter (1985) found that
DEHP taken orally by two volunteers (30 mg each) was excreted as
derivatives of MEPH in the urine at 11 and 15% of the dose. DEHP taken
by the same volunteers for 4 days at 10 mg daily gave no evidence of
accumulation; 15 and 25% of the total dose was recovered in the urine.
No information of fecal recoveries of the parent compound or its
metabolites was reported.
-------
Tax Leo Logics I Data 35
Animal. DEHP appears to be efficiently absorbed from the
gastrointestinal tract of the rat. Williams and Blanchfield (1974)
reported that more than 90% of the radiolabel from an oral dose of 2,000
mgAg C-labeled DEHP was excreted in the urine. Very little DEHP was
absorbed intact. The diester was hydrolyzed by intestinal esterases, and
the MEHP is the primary form absorbed (Oishi and Hiraga 1982, White ec
al. 1980).
Schulz and Rubin (1973), who administered 250 mg L4C-DEHP to rats
in corn oil, found that approximately 61% of the administered dose was
found in the organic solvent extracts of urine, while 12.8% was found in
Che feces and contents of the large intestine.
Albro et al. (1982) and Albro (1986) have reported that there is a
relatively discrete absorption threshold for single oral gavage dosages
of DEHP in Fischer rats. This may be due to saturation of esterases in
the gastrointestinal tract. The animals received 1.8 to 1,000 mg/kg
C-DEHP in cottonseed oil. As the dosage increased, a threshold was
reached above which there was a steady increase in the amount of
unhydrolyzed DEHP reaching the liver. Additional studies utilizing
dietary administration demonstrated that intact DEHP reached the liver
at tissue level concentrations exceeding 4,300 ppm. In contrast to the
results observed in rats, Albro et al. (1982) did not detect an
absorption threshold in either CD-I or B6C3F1 mice administered up to
1.000 mg/kg DEHP.
Rhodes et al. (1986) reported that, in the marmoset, the excretion
profile and tissue levels of radioactivity following oral administration
of L**C-DEHP demonstrated considerably reduced absorption as compared co
the rat. The urinary metabolite pattern in the marmoset was, in many
respects, qualitatively similar to but quantitatively different from
that in the rat.
Chadwick et al. (1982) demonstrated that 14C-7-DEHP is almost
completely absorbed when administered in the diet to male F344 rats at
concentrations of 1,000, 6,000, and 12,000 ppm for 24 h, that is, after
pretreatment for 0, 6, or 20 days at these concentrations with unlabeled
DEHP.
4.2.1.3 Dermal
Human. No data are available on the absorption of DEHP in dermally
exposed humans.
Animal. DEHP appears to be poorly absorbed when applied to the
skin of rats. El Sisi et al. (1985) studied the dermal absorption of
lftC-DEHP In male Fischer 344 rats over a 5-day period. The site of
application was a shaved area of skin 1.8 cm in diameter that was
occluded with a perforated cap. The degree of absorption of DEHP was
estimated from the cumulative daily excretion of 14C-DEHP equivalents in
the urine and feces over 5 days. Only 5.1% of the administered dose of
30 mg/kg 14C-DEHP was excreted over a 5-day period, with 3.0% appearing
in the urine and 2.1% in the feces. An additional 1.8% of the
administered dose was found in the tissues, with muscle tissue
containing 1.2% of the administered dose.
-------
36 Section 4
The nature of the absorbed species was not determined. Onlv total
radioactivity was measured; therefore, it is unknown whether DEHP was
metabolized, and if so. to what products. At present, it is not known
what happens to DEHP after it is taken up by the skin. Mammalian skin
possesses cytochrome P-450 activity, epoxide hydrolase. and
glutathione-s-transferase. and has been shown to metabolize several
compounds (Bickers et al. 1982. Albro et al. 1984). However, the ability
of the mammalian skin to de-esterify xenobiotics in vivo has not been
established, and the existence of nonspecific lipases and esterases in
the skin has not been demonstrated (CPSC 1985).
4.2.2 Distribution
Absorbed DEHP and its metabolites are rapidly distributed to the
organs and tissues with no apparent accumulation. The tissue
distribution of DEHP and its metabolites after oral dosing appears to be
species dependent, with differences being reported for the rat, mouse.
and monkey.
4.2.2.1 Inhalation
Human. No data were available on the distribution of DEHP by
humans exposed via inhalation.
Animal. No data were available on the distribution of DEHP by
animals exposed via inhalation.
4.2.2.2 Oral
Human. No data were available on the distribution of DEHP in
orally exposed humans.
Animal. Eriksson and Darnerud (1985) performed a quantitative
retention study of ^C-labeled DEHP. Groups of mice aged 3, 10 and 20
days were orally administered 0.7 mg of l^C-7-DEHP by gavage in a fat
emulsion. The mice were killed 24 h or 7 days after treatment, and the
amount of radioactivity in the whole brain was measured and its
distribution studied. The retention of DEHP in the brain was minimal.
especially in the 10- and 20-day-old mice, where no radioactivity was
found 7 days after treatment. The amount of radioactivity in the liver
was two to three times higher than that in the brain. Mice exposed to
DEHP showed a significant decrease in the amount of retained
radioactivity between 24 h and 7 days after treatment in all three age
categories.
Pollack et al. (1985b) studied the pharmacokinetics of unlabeled
DEHP and its biologically active metabolite MEHP in rats following
single or multiple administration of DEHP by various routes. DEHP was
assayed in whole blood by a high performance liquid chromatographic
(HPLC) procedure. Following a single intraarterial injection, a large
apparent volume of distribution (5.390 mL/kg) and a high rate of
clearance (21.5 mL/minAg) were observed for DEHP. In contrast, the
systemic availability of DEHP was low following either a single oral
(13.6%) dose administered as a 40% solution in corn oil, or by
intraperitoneal (5.2%) administration A marked route dependency in the
formation of MEHP from DEHP was observed. The circulating concentrations
-------
ToxLcological Daca 37
of MEHP were substantially higher than those of DEHP, with a ratio of
the area under the curve (AUC) for concentration in blood vs time to
that of DEHP of -7 after oral administration. In contrast.
concentrations of the mono-de-esterified metabolite were much lower
relative to the parent diester concentration after intraarterial or
intraperitoneal administration, with an AUC ratio less than 0.4.
Pharmacokinetic modeling indicated that approximately 80% of an oral
dose of DEHP undergoes mono-de-esterification, as compared to only about
1% of the dose following either intraarterial or intraperitoneal
administration. This suggests that the low oral systemic availability of
DEHP may be largely attributed to presystemic hydrolysis of DEHP to MEHP
in the gut, whereas slow and/or incomplete absorption is the likely
cause of the poor bioavailability of DEHP after intraperitoneal
administration. No significant accumulation in the circulating
concentrations of DEHP or derived MEHP were observed following 7 days of
repetitive oral administration of DEHP. However, multiple
intraperitoneal injections resulted in an apparent decrease in the rate
and/or extent of DEHP absorption from the peritoneal cavity, while no
significant change in the oral absorption of the diester was observed
The striking difference in the MEHP to DEHP AUC ratio between oral and
intraperitoneal routes was still evident after multiple dosing. These
data suggest that previously reported differences in the biologic
effects of DEHP in rodents following different routes of administration
may be due to route dependency in the mono-de-esterification of the
diester.
Bratt and Batten (1982) observed clear species and sex differences
in the tissue retention of DEHP. Rats and marmosets were given 1,960
nig/kg/day 14C-DEHP orally for 14 days. In the rat, the females retained
higher concentrations of the L4C-radiolabel in the liver and kidney than
the males. A similar pattern was observed in the marmoset. The rats of
both sexes retained higher tissue concentrations of ^C-radiolabel than
did the marmosets.
Similar results were also reported by Rhodes et al. (1986) in a
comparison of the blood and tissue levels of DEHP and its metabolites in
the rat and marmoset. The animals were administered 2,000 mg/kg/day
l^C-DEHP for 14 days. The test animals were sacrificed 24 h after the
last treatment. The level of l^C-radiolabel in the marmoset tissues was
only 10 to 20% of that in the rat. In both the rat and marmoset, the
liver retained the highest level of 14C-radiolabel.
Lindgren et al. (1982) investigated the distribution and tissue
retention of 14C-DEHP (carbonyl-uC or 2-ethylhexyl-l-14C) in pregnant
and nonpregnant mice with whole-body autoradiography. Initially, a high
activity was observed in the brown fat, liver, gallbladder, intestinal
contents, kidney, and urinary bladder. Pretreatment with DEHP,
phenobarbital sodium, or 3-methylcholanthrene caused a relative increase
of the activity in the brown fat, indicating that induced metabolic
conversion of DEHP leads to an increased deposition of radioactivity in
brown fat. Lindgren et al. (1982) speculated that pretreatment with DEHP
and other known inducers of mixed-function oxidases caused an increase
in the production of DEHP metabolites with an affinity for brown fat
-------
38 Section 4
After administration of DEHP (carbonyl-14C), but not DEHP (2-
ethylhexyl-l-14C), marked retention was observed in the skin, cartilage
and tendons. The mechanism responsible for the slow accumulation in
these connective tissues is not known. In the early embryo, a high
concentration was observed in the neuroepithelium. This pronounced
uptake may be correlated to DEHP-induced exencephaly and spina bifida
observed in mice.
4.2.2.3 Dermal
Human. No data are available on the distribution of DEHP'in
derraally exposed humans.
Animal. No data are available on the distribution of DEHP in
dermally exposed animals.
4.2.2.4 Systemic
Human. Sjoberg et al. (1985a,d) studied the disposition of DEHP
and its primary metabolite MEHP in newborn infants subjected to exchange
transfusions. During a single exchange transfusion, the amounts of
unlabeled DEHP and MEHP infused ranged from 0.8 to 3.3 and 0.05 to 0.20
mg/kg, respectively. Approximately 30% of the infused amount of DEHP was
withdrawn during the course of repeated exchange transfusion.
Immediately after the transfusions, the plasma levels of DEHP ranged
between 5.8 and 19.6 ng/mL, and subsequently they declined rapidly. This
decline, probably reflecting distribution of DEHP within the body, was
followed by a slower elimination phase. The half-life of this phase was
approximately 10 h. The maximal plasma levels of MEHP were about 5
Mg/mL. In one preterra infant, the elimination of MEHP was slower than
its formation, whereas in one full-term newborn, the formation appeared
to be rate-limiting for elimination.
Pollack et al. (1985a) assessed the degree of exposure to DEHP in
11 patients undergoing maintenance hemodialysis for the treatment of
renal failure. The amount of DEHP leached from the dialyzer tubing
during a 4-h dialysis session was estimated by monitoring the DEHP blood
concentration gradient across the dialyzer. The plasma concentrations of
DEHP- and MEHP-derived metabolites were determined by gas
chromatography-chemical ionization spectrometry and gas chromatography-
electron impact mass spectrometry, respectively. Circulating
concentrations of the biologically active products of DEHP de-
esterificaclon, MEHP and phthalic acid, were also determined during the
dialysis session. On the average, an estimated 105 mg of DEHP was
extracted from the dialyzer during a single dialysis session, with a
range of 23.8 to 360 mg. The rate of extraction of DEHP from the
dialyzer was correlated with serum lipid content as expressed by the sum
of serum cholesterol and triglyceride concentrations (r - +0.65,
P < 0.05). Time-averaged circulating concentrations of MEHP during
dialysis (1.33 ± 0.58 Mg/mL) were similar to those of DEHP (1.91 ± 2.11
Mg/mL). Blood concentrations of phthalic acid (5.22 ± 3.94 /ig/mL) were
higher than those of the esters. The length of time patients had been
receiving regular dialysis treatment was not a determinant of
circulating concentrations of DEHP or MEHP In contrast, time-averaged
circulating concentrations of phthalic acid correlated strongly with the
-------
ToxicoLogical Daca 39
duration (In years) of dialysis treatment (r - +0.92, P < 0 001) The
results indicated substantial exposure to DEHP during hemodialysis and
that de-esterification products of DEHP are present in significant
concentrations in the systemic circulation.
Animal. Sjoberg et al. (1985b) studied the disposition of DEHP ard
four of its major metabolites in male rats. The test animals were given
single intravenous infusions of a DEHP emulsion in doses of 5, 50, or
500 mg/kg DEHP. Plasma concentrations of DEHP and metabolites were
followed for 24 h after the start of the infusion. The plasma
concentrations of DEHP- and MEHP-derived metabolites were determined by
gas chromatography-chemical ionization spectrometry and gas -
chromatography-electron impact mass spectrometry, respectively. The
concentrations of DEHP in plasma were at all times considerably higher
than those of MEHP, and the concentrations of MEHP were much higher than
those of the other investigated metabolites. In animals given 500 mg/kg
DEHP, the areas under the curves (AUCs) for concentration in plasma vs
time of the other investigated metabolites were less than 15% of that of
MEHP. The clearance of DEHP was much lower in the high-dose group than
in the lower-dose groups. This is probably due to saturation of enzymes
responsible for the hydrolysis of DEHP, product inhibition, or
saturation of biliary excretion. Parallel decreases in the plasma
concentrations of DEHP, MEHP, and the omega- and omega-1 oxidized
metabolites indicated that the plasma clearance of DEHP was the rate-
limiting step in the disposition of the metabolites. This was partly
supported by the observation that the clearance of MEHP was higher than
that of DEHP. Nonlinear increases in the AUCs of DEHP and MEHP indicated
saturation in the formation as well as the elimination of the
potentially toxic metabolite MEHP.
4.2.3 Metabolism
The metabolism of DEHP involves a complex series of primary and
secondary pathways. The proposed metabolic pathway for DEHP is shown in
Fig. 4.1. The first step in metabolism is the lipolytic cleavage of
DEHP, resulting in the formation of MEHP and 2-ethylhexanol. Oxidative
metabolism of MEHP occurs via omega- and omega-1 oxidation of the
aliphatic side chain. It is believed that this step is followed by a
dehydrogenase-dependent oxidation to the ketone or carboxylic acid, with
subsequent alpha- and beta-oxidation of the acids (Albro et al.
1983a,b). The metabolism of 2-ethylhexanol appears to be primarily via
the oxidative carboxylation leading to the formation of 2- and 4-oxy-3-
carboxyheptane. The primary urinary metabolites of 2-ethylhexanol are
2-ethylhexanoic acid and several keto acid derivatives that appear to be
beta-oxidation products of the primary metabolite.
4.2.3.1 Inhalation
Human. No data were available on the metabolism of DEHP by humans
exposed via inhalation.
Animal. No data were available on the metabolism of DEHP by
animals exposed via inhalation.
-------
OEHP
^^
COOCH2CH(C^)3CH3
HYOROUSB (LVMSES)
MEHP
l/l
(B
O
rt
\
OOOH
COOCH,CH|OH^OOCH(
OOOCH.CHCOH.Ip.,
\
CH,CH,OM
OOOCMjCMICMj^.OM
6-oxB/moN
CK.CH,
OOOH
OOOCHjCHCMgCHOHO^COOM
O^OM,
COOOMjCHCMjCOOM
Fig. 4.1. Proposed melaboliinn for di(2-ethylhexyl)phlhalale in the ml.
-------
Toxico logical Data £L
4.2.3.2 Oral
Human. Schraid and Schlatter (1985) found chat DEHP taken orally bv
two volunteers (30 mg each) was excreted in the urine at 11 and 15% of '
Cf%cSpr ter enzy»lc hydrolysis, the urinary metabolites (derivatives
of MEHP) were methylated and identified by gas liquid chromatography-
mass spectrometry, and the quantitative distribution of conjugated and
free metabolites was determined. Twelve metabolites were detected the
four major ones being free and conjugated forms of the monoester and its
6-carboxylic acid. 5-keto. and 5-hydroxy derivatives. The results of
this study suggest that the theory that none of the five major
metabolites of DEHP in rats (Sjoberg et al. 1985a. 1986a) are major
metabolites in man may be incorrect. Schraid and Schlatter (1985) found
that one of the two major metabolites in man involved oxidation of the
terminal (C-6) methyl group to the corresponding acid, a
biotransformation process that also occurs in the rat (Watts 1985) The
reason for this is not clear, but may reflect different dose levels or
different routes of administration.
P nc pLhu8enot ec al- <1985> investigated the in vivo metabolism
of DEHP and MEHP in rats after multiple dosing. Rats were orally
administered ^C-DEHP or ^C-MEHP at doses of 50 and 500 mg/kg for 3
consecutive days. Urine was collected at 24-h intervals, and metabolite
profiles were determined. After a single dose of either compound
urinary metabolite profiles were similar to those previously reported
However, after multiple administration of both DEHP and MEHP at 500
mgAg, increases in omega -/beta -oxidation products [raono(3-carboxy-2-
ethylpropyl) phthalate and mono(5-carboxy-2-ethylpentyl) phthalate
respectively] and decreases in omega -1- oxidation products [mono(2-
ethyl-5-oxohexyl) phthalate and mono (2-ethyl- 5 -hydroxyhexyl) -phthalate
respectively] were seen. At the low dose of 50 mg/kg, little or no
alteration in urinary metabolite profiles was observed. At 500 mg/kg
MEHP, a 4-fold stimulation of cyanide -insensitive palmitoyl-CoA
oxidation (a peroxisomal beta-oxidation marker) was seen after three
consecutive daily doses. At the low dose of 50 mg/kg, only a 1.8-fold
increase was noted. Similar observations were made by Lhugenot et al
(1985) with rat hepatocyte cultures.
Chadwick et al (1982) treated male F344 rats with 14C-7-DEHP in the
diet at concentrations of 1.000, 6.000, and 12,000 ppm for 24 h
following pretreatment for 0. 6. or 20 days with unlabeled DEHP. In the
animals without prior exposure to DEHP, there was a dose-related,
nonlinear, disproportionate increase in the fraction of the dose
converted to metabolites I and V, that is, the 3- and 5-carboxylic acid
derivatives formed via the omega-oxidation pathway. The percentage of
Increased urinary excretion of metabolites I and V was accompanied by a
reduction in the percentage of the administered dose excreted in the
feces. A reduction in the fecal excretion of metabolites IX and VI the
5-hydroxy and 5-keto metabolites formed via the omega- 1 oxidation
pathway, was also reported.
Prior exposure to DEHP resulted in nonlinear increases with dose in
the fraction of the administered dose metabolized via the omega- and
beta-oxidation pathways to metabolite I and excreted in the urine. Prior
exposure to 1,000-ppm DEHP resulted in a quantitative change in the
-------
42 Seccion 6
pattern of DEHP metabolism. After either 6 or 20 days of pretreatment,
the percentage of the administered dose excreted in the urine as
metabolite I was doubled. However, the percentage excreted as metabolite
V, the proposed precursor of metabolite I, remained unchanged. The
apparent increase in the conversion of metabolite V to metabolite I may
have resulted from hepatic enzyme induction. DEHP is known to induce
peroxisome proliferation, and has been shown to induce microsomal
cytochrome P-450 and 7-ethoxycoumarin 0-deethylase activity in the rat
(Mangham et al. 1981). The increased metabolism via the omega-oxidation
pathway was offset by a parallel decrease in the percentage of
administered dose converted to the omega-1 oxidation products,
metabolites IX and VI, and excreted in the urine and feces. At 6,000 and
12,000 ppm, prior exposure to DEHP for either 6 or 20 days resulted in a
major increase in the percentage of administered dose metabolized via
the omega-oxidation pathway. Urinary excretion of metabolite I was
significantly increased, accompanied by a parallel, dose-related
decrease in the urinary excretion of metabolite V. This was offset by an
overall decrease in the conversion of MEHP to omega-1 oxidation
products.
Agarwal et al. (1982) reported that the administration of DEHP as a
single oral dose of 2.6, 5.2, or 13.0 mL/kg or a single intraperitoneal
dose of 5.0, 10.0, or 25.0 mL/kg significantly increased the
pentobarbital-induced sleeping time in rats. Repeated intraperitoneal
administration of the same doses for 7 consecutive days also increased
the sleeping time, but repeated oral administration decreased it in a
dose-dependent manner. A single exposure to DEHP by either route
significantly inhibited the activity of aminopyrine tf-demethylase and
aniline hydroxylase without any change in the activities of
benzo(a)pyrene hydroxylase, glucose 6-phosphatase, or NADPH-cytochrome c
reductase or the concentration of cytochrome P-450 in rat liver.
Repeated intraperitoneal administration of DEHP had effects similar to
those observed with a single exposure. However, repeated oral doses of
DEHP resulted in a significant increase in the activities of aminopyrine
tf-demethylase, aniline hydroxylase, and benzo(a)pyrene hydroxylase, and
in the concentration of cytochrome P-450. The activity of glucose 6-
phosphatase was significantly Inhibited in a dose-dependent manner, but
that of NADPH-cytochrome c reductase remained unaffected after repeated
oral and intraperitoneal administration of DEHP. Treatment with DEHP
failed to affect total and ascorbate- or NADPH-dependent lipid
peroxidation as well as total and nonprotein sulfhydryl contents of rat
liver. Under in vitro conditions, DEHP had no effect on the activity of
aminopyrine tf-demethylase or aniline hydroxylase, whereas mono-(2-
ethylhexyl) phthalate and 2-ethylhexanol significantly inhibited the
activity of both enzymes at concentrations ranging from 2.5 to 15.0 mM.
the effect of 2-ethylhexanol being more pronounced.
4.2.3.3 Dermal
Human. No data were available on the metabolism of DEHP in
dermally exposed humans.
Animal. No data were available on the metabolism of DEHP in
dermally exposed animals.
-------
Toxicological Data U1
4.2.3.4 In vitro studies
Lhugenoc et al. (1985) reported that MEHP at concentrations of 50
and 500 jiM was extensively metabolized in the rat hepatocyte cultures.
Metabolic profiles similar to those seen after in vivo administration of
MEHP were observed. At the higher concentration of MEHP, changes in the
relative proportions of omega- and omega-1 oxidized metabolites were
seen. Over the 3-day experimental period, omega-/beta-oxidation produces
increased in a time-dependent manner at the expense of omega-1 oxidation
products. At a concentration of 500 jiM MEHP, a 12-fold increase of
cyanide-insensitive palmitoyl CoA oxidation (a peroxisomal beta-
oxidation marker) was observed. At the low concentration of MEHP
(50 /^M), only a 3-fold increase in cyanide-insensitive palmitoyl-CoA
oxidation was noted, and little alteration in the metabolite profile of
MEHP was observed with time. Biotransformation studies of the
metabolites of MEHP confirmed the postulated metabolic pathways.
Mono(3-carboxy-2-ethylpropyl)phthalate and mono(2-ethyl-5-
oxohexyl)phthalate appeared to be end points of metabolism, while
mono(5-carboxy-2-ethylpentyl)phthalate was converted to mono(3-carboxy-
2-ethylpropyl)phthalate, and mono(2-ethyl-5-hydroxyhexyl) phthalate to
mono(2-ethyl-5-oxohexyl)phthalate.
Gollamudi et al. (1983) studied the impairment of DEHP metabolism
in young and aged rats. Liver, kidney, and lung homogenates from 45- and
630-day-old rats were incubated with 14C-labeled DEHP.
Radiochromatograms of ether extracts of the incubated mixtures from
young animals showed peaks corresponding to the parent compound and the
hydrolytic product MEHP. Preparations from old animals revealed a
dramatic reduction in the formation of MEHP in the liver homogenates
This difference may be attributable to differences in Km values of the
enzymes from adult and old rats, respectively. Protein content in the
three tissues did not differ between young and old animals. Because the
formation of MEHP is a major step in the metabolism of DEHP, impairment
of this conversion could possibly alter the rate of its excretion and
its toxicologic significance.
4.2.4 Excretion
DEHP is converted principally to polar derivatives of the
monoesters by oxidative metabolism prior to excretion. Clearance from
the body is rapid, and there is only a slight cumulative potential. The
major route of DEHP elimination from the body is urinary excretion.
4.2.4.1 Inhalation
Human. No data were available on the excretion of DEHP by humans
exposed via inhalation.
Animal. No data were available on the excretion of DEHP by animals
exposed via inhalation.
4.2.4.2 Oral
Human. Schmid and Schlatter (1985) found that DEHP taken orally by
two volunteers (30 mg each) was excreted in the urine at 11 and 15% of
the dose. DEHP taken by the same volunteers for 4 days at 10 mg daily
-------
44 Section 6
gave no evidence of accumulation; 15 and 25% of the total dose was
recovered in the urine. On the basis of this, the investigators
estimated a half-life of 12 h and concluded that accumulation of DEHP in
the body is unlikely to occur. Unfortunately, fecal analysis that could
have supported this hypothesis was not performed. Generally, the data
compare well with those of Albro et al. (1982) for human leukemia
patients who received infusions of DEHP-contaminated blood.
Animal. Rhodes et al. (1986) studied the comparative
pharmacokinetics of DEHP in the Wistar rat and the marmoset monkeys
following oral, intravenous, or intraperitoneal administration. In both
the rat and marmoset, multiple oral administrations of l^C-DEHP at 2,000
mg/kg did not modify the proportion of dose excreted in the urine or
feces in either male or female animals.
Chadwick ec al. (1983) administered 100 mg/kg 14C-7-DEHP by gavage
to three male Cynomolgus monkeys, five male F344 rats, and five groups
of five B6C3F1 mice. Urinary and fecal excretion of the radiolabel was
almost complete after 24 and 48 h, respectively. The radiolabel in the
urine and feces was resolved into as many as 12 components. The
monoester derivative of DEHP was found in the monkey and mouse urine,
but was not detected in the rat urine. Metabolite V, the 5-carboxylic
acid omega-oxidation product, was a major metabolic product in rat and
monkey urine. Metabolite IX, the 5-hydroxy omega-1 oxidation product,
was a major component in the urine of all three species. Metabolite VI,
a 5-keto derivative formed from metabolite IX, was a major component in
the urine of mice and rats but not monkeys. Metabolite I, the 3-
carboxylic acid derivative formed via beta oxidation, was also a major
metabolite in the urine of mice and rats but not monkeys. These data
suggest that beta oxidation is a major metabolic pathway in rodents but
not in nonhuman primates.
Chadwick et al. (1982) reported that urinary excretion of
radioactivity increased with dose whereas fecal excretion decreased
with dose in male F344 rats fed L4C-7-DEHP in the diet at concentrations
of 1.000, 6,000, and 12,000 ppm for 24 h following pretreatment for 0,
6, or 20 days with unlabeled DEHP. The time course patterns of
elimination of DEHP metabolites via the urinary and fecal routes were
different. Urinary excretion of metabolites primarily occurred during
the first 24 h after dosing, whereas fecal excretion occurred primarily
during the second 24-h phase. The differences in clearance of
metabolites are probably due to the enterohepatic recycling of
metabolites secreted via the bile and reabsorbed in the gastrointestinal
tract.
Teirlynck and Belpaire (1985) studied the disposition of DEHP and
MEHP in male Wistar rats. Three hours after a single oral dose of 2,800
mg/kg DEHP, plasma concentrations of 8.8 /ig/mL DEHP and 63.2 jig/mL MEHP
were observed. The ratio of the area under the curve (AUC) for
concentration in plasma vs time of MEHP to that of DEHP was 16.1. MEHP
had a half-life of 5.2 h. When 14C-DEHP was administered, 19.3% of the
radioactivity was excreted in the urine within the first 72 h. The
remaining radiolabel was excreted in che feces. The urinary excretion
rate indicated a half-life of 7.9 h Multiple dosing with 2.800 rag/kg
-------
lexicological Daca ^5
DEHP for 7 consecutive days produced no accumulation of either DEHP or
MEHP.
4.2.4.3 Dermal
Human. No data were available on the excretion of DEHP in dermal!,
exposed humans.
Animal. No data were available on the excretion of DEHP in
dermally exposed animals.
4.2.4.4 Systemic
Human. No data were available on the excretion of DEHP in
systemically exposed humans.
Animal. Metabolites of DEHP found in urine from African green
monkeys after intravenous administration of ^C-labeled parent compound
were isolated and identified by Albro et al. (1981). Urinary metabolices
(80%) were excreted in the form of glucuronide conjugates. This was
analogous to that reported for the urinary metabolites of DEHP from
humans, but in contrast to metabolites found in rat urine. Rat urinary
metabolites of DEHP were excreted unconjugated and consisted primarily
of derivatives more highly oxidized than the major metabolites produced
by monkey or human. This suggests that the African green monkey may be a
better model for human metabolism of DEHP than the rat (Peck and Albro
1982). Interspecies differences in the metabolism of DEriP are summarized
in Fig. 4.2.
4.2.5 Discussion
Animal experiments have shown that DEHP is readily absorbed after
ingestion. In the one study available (El Sisi et al. 1985), it was
shown that substantially less DEHP is absorbed through the skin, and che
nature of the absorbed species is unknown.
Oral exposure to less than very high dose levels of DEHP does noc
result in tissue exposure to the intact diester, which is rapidly and
efficiently hydrolyzed in the gastrointestinal tract by microflora and
hydrolytic enzymes.
The primary metabolites, MEHP and 2-ethylhexanol, are rapidly
oxidized to a variety of more polar products. The biological activity of
the metabolic products has not been investigated extensively. There are
major interspecies differences in the metabolism of DEHP.
In rodents (rats and mice), the excreted metabolites of DEHP
primarily consist of diacids and ketoacids, which are terminal oxidation
products. However, in both humans and the African green monkey, the
major excretion metabolites are MEHP and minimally oxidized hydroxyacid
products.
In addition to interspecies differences, there are major
qualitative differences in metabolism and major quantitative differences
in excretion rates between young and old animals of the same species
-------
— OR
DEHP
HYDROLYSIS
RODENTS
AND
PRMATES
MEHP
GLUCURONIDATION
AND
CO andcj-1
OXDATIONS
— OH
CH2CH3
I
R - -CH2CH2(CH2);jCH3
R'-OXDATION PRODUCTS OF R
RATS
RODENTS (MICE)
AND
PRIMATES
— OH
— OR
— O-GLUCURONIDE
10
(6
n
n
F-
o
3
Fig. 4.2. Species-specific metabolism of dK2-ethylhexyl)phlluilate.
-------
ToxicologicaL Daca ^.7
4.3 TOXICITY
4.3.1 Lethality and Decreased Longevity
4.3.1.1 Inhalation
Human. Thiess et al. (1978a) conducted a morbidity study and a
mortality study (1978b) on 101 workers who had been employed for periods
ranging from 4 months to 35 years in a DEHP production facility Currenc
exposures in work areas ranged from 0.0006 to 0.01 ppm. but may have
been higher prior to 1966 when a process change was made. Blood, liver,
and urinary conditions were examined, and for the six worker-s with more
than 20 years of service, a neurologic examination was administered
Analyses of absenteeism, of accidents, and of a reproduction
questionnaire were conducted. In-house control groups (with possible
exposure to styrene and dimethyl carbamoyl chloride) were compared with
the DEHP workers. No findings of morbidity attributable to DEHP exposure
were reported. The lack of good exposure data and of a control group not
exposed to industrial chemicals limits the utility of this morbidity
study.
In the mortality study (Thiess 1978b), which involved 221 workers
with average time in service of 11.5 years, no increase in mortality
with increase in exposure duration was seen. A single case of bladder
papilloma resulted in a higher-than-expected rate for that phenomenon.
but this was not considered sufficient.
Thiess et al. (1979) also performed chromosomal analysis on blood
lymphocytes from a subset of this same study population. Lymphocytes
were cultured from 10 exposed production workers according to a modified
method of Moorhead et al. (1960). The workers' duration of exposure
ranged from 10 to 34 years (mean - 22.1 years). Lymphocytes from 20
age-matched workers served as controls. It was not stated whether these
controls were also exposed to styrene and DMCC, as mentioned previously
One hundred metaphases were scored for abnormalities on lymphocytes from
each worker. The specific structural abnormalities were not defined, buc
were categorized with and without gaps. Neither category appeared to be
different from the control, although statistical analyses were not
stated.
None of the above three studies by Thiess et al. can be used to
establish a NOAEL.
Animal. Acceptable lethality data were not available for animals
exposed to DEHP. In an acute rat study. 1 h exposure to 23,670 mg/m3
(1,457 ppm) DEHP did not result in any deaths (WARF Institute 1976). In
a study by Pegg (1979). no deaths were observed in rats exposed to 600
mg/mj (37 ppm) DEHP for 6 h.
4.3.1.2 Oral
Human. No data were available on the effects of ingestion exposure
to DEHP on lethality or decreased longevity in humans.
Animal. A number of animal studies, which are summarized in Table
4.1. have been performed to investigate the acute toxicity of DEHP The
LD50 for DEHP ranges from 26.000 to 49.000 mg/kg. which indicates thac
-------
Section
Table 4.1. Acute lethality of diiZ-ethylhexvliphthalaie
Route of
administration
Inhalation
(1 h)°
Inhalation
(6h)°
Oral
•
Inirapentoneal
Intravenous
Intravenous
(sonicated in
rat serum)
Species
Rat
Rai
Rat
Rat
(Wisiar. male)
Rat
(Wistar. male)
Mouse
Mouse
Mouse
Guinea pig
Rabbit
Rat
Rat
(Wistar. male)
Mouse
Mouse
(ICR. male)
Mouse
Rat
LD50
(mg/kg)
26.000
>34000
30.600
49.000
26.000
33.300
26.300
33.900
49.000
30.600
4.200
38.000
1.060
2.080
LC50
(mg/m ) References
>23.670 WARF Institute 1976
(1457 ppm)
>600 Pegg 1979
(37 ppm)
Patty 1967
Hodge 1943
Shaffer et al I94S
Yamada 1974
Patty 1967
Krauskopf 1973
Krauskopf 1973
Shaffer et al I94S
Singh etal 1972
Shaffer ei al I94S
Galley etal 1966
Lawrence et al 1973
Petersen et al 1974
Petersen et al 1974
'Duration of exposure
Source. Adapted from NTP 1982
-------
ToxicologicaL Daca 69
DEHP has a low order of acute oral coxicicy. Studies using the
intravenous and intraperitoneal routes of administration, which are also
summarized in Table 4.1. provide additional evidence of the low order of
the acute toxicity of DEHP.
The NTP (1982) evaluated the carcinogenicity of DEHP in a standard
bioassay using F344 rats and B6C3F1 mice of both sexes. In the rats, no
significant trends in mortality were observed in the treated animals
when compared to the controls. In the low-dose female mice, a
significantly shortened survival was observed compared with the
controls. However, no positive trend was observed since there was a
somewhat longer survival in the high-dose females.
4.3.1.3 Dermal
Human. No data were available on the effects of dermal exposure co
DEHP on lethality or decreased longevity in humans except for those
reported in Sect. 4.3.1.1.
Animal. There were no available data on the potential lethality or
decreased longevity associated with dermal exposure of animals to DEHP
4.3.1.4 Discussion
The available data suggest that DEHP has a low order of acute oral
toxicity. In oral studies in rodents, the LDso ranges from 26,000 to
49.000 mg/kg. Data from a 2-year oral carcinogenicity bioassay indicate
that DEHP does not induce dose-related increases in mortality in male
rats or mice at doses of 322 or 674 mg/kg/day and 672 or 1,325
mg/kg/day, respectively, or in female rats or mice at doses of 394 or
774 mg/kg/day and 799 or 1,821 mg/kg/day. respectively.
4.3.2 Systemic/Target Organ Toxicity
The primary target organs of DEHP have been shown to be the liver
and testes.
4.3.2.1 Hepacic toxicity
Certain phthalate esters, including DEHP. and hypolipidemic agents
are known to induce morphological and biochemical changes in the liver
of rodents. Such changes have been associated with increased incidences
of hepatocellular tumors in these species. While there is evidence that
some hypolipidemic agents do not induce these effects in either subhuman
primates or man (Cohen and Grasso 1981. Lake and Gray 1985). Jacobson ec
al. (1977) and Canning et al. (1984) contradict this.
Inhalation, human. No data were available on the hepatic toxicity
of DEHP in humans exposed via inhalation.
Inhalation, animal. No data were available on the hepatic toxicity
of DEHP in animals exposed via inhalation.
Oral, human. No data were available on the hepatic toxicity of
DEHP in orally exposed humans.
Oral, animal. Gollamudi et al (1985) reported that DEHP inhibited
UDP-glucuronyl-transferase activity of rat liver in vitro and in vivo
-------
50 Section 4
Diechyl phthalate and dimethoxyethyl phthalate also Inhibited this
enzyme in vitro. Conversely, DEHP did not inhibit the activity of the
cytosolic enzyme N-acetyltransferase or alter the levels of rat liver
microsomal cycochrorae P-450 in vitro. It is suggested that DEHP may
alter the composition of microsomal phospholipids.
Rhodes et al. (1986) reported that oral and intraperitoneal
administration of DEHP to the marmoset monkey at doses up to 5 mmole
(1950 mg) DEHP/kg/day for 14 days did not induce morphological or
biochemical changes in the liver comparable with those obtained in rats
administered the same amount of DEHP.
In a study of the hepatic effects of DEHP (CMA 1984), groups of
five male and five female rats were treated with 0. 0.1, 0.6, 1.2, or
2.5% DEHP in the diet for 3 weeks. In rats fed 2.5% DEHP, body weight
and food intake were significantly reduced. Statistically significant
increases in liver weight were observed at dose levels of 0.6, 1.2, and
2.5%. There was a dose-related increase in cyanide-insensitive palmitoyl
CoA oxidation (pCoA) at dose levels of 0.6% and above. An increase in
lauric acid 12-hydroxylation was seen at doses of 0.1% and above in the
males and at the 1.2 and 2.5% dose levels in the females. A less-marked
increase in 11-hydroxylation was also observed. Electron microscopy
revealed a dose-related increase in peroxisomes at levels of 0.1% and
above in males and at 0.6% and above in the females.
The Chemical Manufacturers Association (CMA) (1985a,b,c,d;
1986a,b,c,d) also evaluated the hepatic effects of eight other phthalate
esters in rats. Groups of five male and five female rats fed 1.2% DEHP
in the diet for 3 weeks served as positive controls in each study.
Analysis of the DEHP-positive control values between studies indicates
that male rats treated with 1.2% DEHP in the diet gained less weight
than concurrent untreated control male rats. However, the body weight
values observed in the DEHP-treated rats were generally not more than
10% less than untreated control values. There was substantial
variability (2.38-fold) between the values obtained for pCoA, a primary
marker for peroxisome proliferation. Values obtained for 11- and 12-
lauric acid hydroxylase were also variable (2.28- and 2.33-fold,
respectively).
Parmar et al. (1985) investigated the effects of DEHP on the
detoxification mechanisms of xenobiotics in neonatal rat pups of mothers
who received 2,000 mg/kg phthalate daily from day 1 of birth to day 21.
The nursing rat pups of mothers exposed to DEHP showed decreases in body
weight gain and activities of aniline hydroxylase, ethylmorphine N-
demethylase, and arylhydrocarbon hydroxylase, and decreased levels of
cytochrome P-450 at 21 days of age. Significant quantities of DEHP were
also detected In the liver of pups In this Interval. These results
suggest that the livers of developing animals can be affected by the
lactational transfer of DEHP.
F. E. Mitchell et al. (1985) fed groups of male and female Wistar
albino rats levels of 50, 200, or 1.000 mg/kg/day DEHP in the diet. Four
rats from each experimental group and six control rats of the same sex
were killed 3, 7, 14, and 28 days and 9 months after the start of
treatment. At all time points the major abdominal organs were removed
and subjected to histological examination A more extensive necropsy was
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TOXLCOlogical Daca 5L
performed on chose rats killed after 9 months of treatment. Two early,
transient alterations were observed. There were morphologic changes in
the bile canaliculi of male rats treated with 1,000 mg/kg/day DEHP. In
addition, the liver cells exhibited a burst of mitosis immediately afcer
the start of administration of the compound. The time course of this
mitotic burst varied; the increase in mitosis was greatest at 3 days in
rats treated with 1,000 mg/kg/day DEHP, and was smaller, but more
prolonged, in rats treated with 50 or 200 mg/kg/day. Other changes,
including a midzonal to periportal accumulation of fat, induction of
peroxisomal enzymes, and induction of the P-450 isoenzyme, also
developed rapidly and were observed throughout the study. The maximal
change was usually attained within 7 days of the start of treatment.
More slowly developing changes were hypertrophy of the hepatocytes,
centrilobular loss of glycogen, and a fall in glucose-6-phosphatase
activity. Maximal changes were not attained until 28 days after
treatment began. These effects were clearly observed in rats treated
with 200 or 1,000 mg/kg/day DEHP, but were only marginally altered in
rats treated with 50 mg/kg/day. Finally, accumulation of lipid-loaded
lysosomes assessed by light and electron microscopy and by assay of
beta-galactosidase activity was only apparent in rats treated with DEHP
for 9 months with 200 or 1.000 mg/kg/day DEHP. Changes in female rats
were qualitatively similar to those observed in male rats. The
alterations were, however, less pronounced than in male rats treated
with an equal dose of DEHP, and the degree of liver enlargement was much
less because, although the initial hyperplasia was clearly apparent,
there was a much smaller degree of hypertrophy.
Melnick and Schiller (1985) exposed isolated rat liver mitochondria
to mono- and di-n-butyl phthalate (MBP and DBP) and MEHP and DEHP. These
were examined for effects on mitochondrial energy-dependent processes,
including oxidative phosphorylation and active K+ uptake. Additional
studies on the effects of these phthalate esters on succinate oxidation
and on mitochondrial membrane integrity were also performed. DEHP had no
apparent effect on any of these processes except for slight impairment
of ATP-dependent £+-valinomycin-induced swelling. The MEHP was found co
act as a noncompetitive inhibitor of succinate dehydrogenase activity
At concentrations which uncoupled energy linked reactions, MEHP produced
slight energy-dependent swelling and the release of soluble proteins
from isolated mitochondria, probably due to an increase in membrane
permeability to H+ and other small ions.
In a study by Carpenter et al. (1953), groups of 23 to 24 guinea
pigs of each sex were fed diets containing DEHP at dose levels of 19 or
64 mg/kg/day for a period of 1 year. No treatment-related effects were
observed on mortality, body weight, kidney weight, or gross pathology
and histopathology of kidney, liver, lung, spleen, or testes. A
statistically significant increase in relative liver weight was observed
in both groups of treated females.
Carpenter et al. (1953) also performed a study with groups of male
and female Sherman rats. However, this study was inadequate because of
high mortality in the control group, thus limiting its utility.
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52 Section 4
Doscal et al. (1987) investigated the relative sensitivity of
suckling rats, as compared to adults, to the effects of DEHP. Five daily
oral doses of 0, 10, 100, 1,000, or 2,000 mg/kg DEHP were administered
to male Sprague-Dawley rats beginning at 6, 14, 16, 21, 42, and 86 days
of age. Twenty-four hours after the last dose, rats were sacrificed, and
plasma cholesterol and triglyceride levels and the activities of the
hepatic peroxisomal enzymes, palmitoyl CoA oxidase and carnitine
acetyltransferase, were determined.
Suckling rats (1 to 3 weeks of age) suffered severe growth
retardation at doses of 1,000 mg/kg and death at 2,000 mg/kg, while
older rats only showed decreased weight gain at 2,000 mg/kg. Of '
particular interest was the lethality at doses of 1,000 mg/kg at 14 days
of age, but not at 16 days or at other ages. Increases in relative liver
weight and hepatic peroxisomal enzyme activities were similar in all age
groups except the 14-day-old group, in which the increases were greater.
Relative kidney weight was increased in 21-, 42-, and 86-day-old rats,
but not in younger rats, at the highest doses. Hypolipidemia was
observed only in 21-, 42-, and 86-day-old rats at doses of 1,000 and
2,000 mg/kg, whereas elevated plasma cholesterol levels were observed in
6- and 14-day-old rats at the 1,000-rag/kg dose, possibly due to the
dietary differences between suckling and weaned rats. The results
suggest that neonatal and suckling rats are more sensitive to the lethal
and growth retardation effects of DEHP than are adult rats, but the
hepatic peroxisome proliferation was similar at all ages with the
exception of a greater increase at 14 days of age.
Dermal, human. No data were available on the hepatic toxicity of
DEHP in dermally exposed humans.
Dermal, animal. No data were available on the hepatic toxicity of
DEHP in dermally exposed animals.
Systemic, human. Canning et al. (1984) reported that liver
biopsies taken from dialysis patients showed peroxisome proliferation.
Systemic, animal. Walseth et al. (1982) administered DEHP via
intraperitoneal injection (3.8 mM/kg) to Sprague-Dawley rats for 5 days.
DEHP increased significantly the liver concentration of cytochrome
P-450 but decreased the lung concentration of cytochrome bS and NADPH-
cytochrome-c-reductase activity. The direction of benzo(a)pyrene
metabolism was changed, and the formation of 2- and 3-hexanol
metabolites was increased in liver microsomes after treatment. The
cytochrome P-450 enzyme system in the lung was 10-fold more effective
than that in the liver as far as metabolism of n-hexane was concerned.
Only minor effects were observed in serum enzyme activities, but a
significant decrease in the serum level of albumin was observed after
treatment with DEHP. No relationship was found between the carbon chain
length of the investigated chemicals and effects on microsomal enzymatic
activities.
Discussion. Exposure to DEHP has resulted in adverse hepatic
effects in laboratory test animals DEHP at high concentrations can
cause functional hepatic damage, as reflected by morphological changes.
and alterations in the activity of energy-linked enzymes and metabolism
of llpids and carbohydrates. Several studies have demonstrated that DEHP
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TaxLCOLogical Data 53
is a peroxisome proliferator. These observations are significant because
of an association between peroxisome proliferation and both liver
hyperplasia and carcinogenesis in laboratory animals Administration of
DEHP orally ac sufficient doses has been shown to induce proliferation
of peroxisomes, increase mitochondria and endoplasmic reticulum
membranes, induce cytochrome P-450, and induce hepatomegaly associated
with hepatic hyperplasia. Studies performed to date have not established
any single metabolite or metabolic pathway as essential for peroxisome
proliferation. In addition, there have been no long-term studies that
allow a precise estimate of the relationship between DEHP dose and
peroxisome proliferation.
4.3.2.2 Testicular toxicity
The testicular effects produced by DEHP are discussed in the
section on reproductive toxicity.
4.3.3 Developmental Toxicity
The available data indicate that DEHP is developmentally toxic in
rats at relatively high doses. In mice, DEHP is developmentally toxic ac
levels where no effects were observed in the treated dams.
4.3.3.1 Inhalation
Human. No data were available on the developmental toxicity of
DEHP in humans exposed via inhalation.
Animal. No data were available on the developmental toxicity of
DEHP in animals exposed via inhalation.
4.3.3.2 Oral
Human. No data are available on the developmental toxicity of DEHP
in orally exposed humans.
Animal. Nakamura et al. (1979) investigated the teratogenicity of
DEHP in ddy-Slc x CBA mice. The animals received a single oral
administration of 0.05, 0.1, 1.0, 2.5, 5.0, or 10.0 mL/kg DEHP by gavage
on day 7 of gestation. A dose of 0.05 mL/kg DEHP resulted in a decrease
in body weight of live fetuses. However, there were no fetal deaths or
soft-tissue or skeletal abnormalities. At doses of 0.1 mL/kg and above,
DEHP decreased fetal body weight, and the fetuses were deformed or dead
Wolkowski-Tyl et al. (1984a) fed timed pregnant Fischer 344 rats
diets which contained 0, 5,000. 10.000. 15.000, or 20,000 ppm DEHP on
days 0 to 20 of gestation. Based on food consumption data, this was
equivalent to doses of approximately 0, 357. 666, 856, or 1.055
nig/kg/day. There was no maternal mortality in any of the dosed animals
There was a significant dose-response trend toward maternal body weight
loss, decreased gravid uterine weight, and increased absolute and
relative liver weights. Clinical signs of toxicity, including
piloerection, rough coat, and reduced food intake, were observed in a
dose-related manner. There were no dose-related differences observed in
the number of corpora lutea or implantation sites per dam or percentage
of preimplantation losses. The number and percentage of resorptions,
nonlive fetuses, and affected (nonlive and malformed) fetuses were
-------
54 Section 4
Increased In a dose-related manner, with a statistically significant
increase in all three parameters in the high-dose group when compared
with the controls. The number of live fetuses per litter exhibited a
dose-related decrease, with the high-dose group significantly lower than
the controls. Among the live litters, there was a dose-related decrease
in the body weights of both sexes at all doses tested. There was a
significant upward dose-related trend in the percentage of malformed
fetuses per litter. However, there were no significant differences in
pairwise comparisons in the number of malformed fetuses per litter. The
results of this study demonstrate that DEHP is maternally and
developmentally toxic. DEHP has been shown to cross the placenta in rats
(Singh et al. 1975), and the fetotoxicity observed in this study may
have been the result of direct contact to the parent compound and/or its
metabolites with embryonic or fetal tissues.
Wolkowsky-Tyl et al. (1984b) exposed timed pregnant CD-I mice on
days 0 to 17 of gestation to diets containing 0, 250, 500, 1,000. or
1,500 ppm DEHP. Based on food consumption data, this was equivalent to
approximately 0, 44, 91, 191, or 292 mg/kg/day. There was no maternal
mortality in any dose group. There was a significant dose-response trend
toward reduced maternal body weight on-days 12, 16, and 17 of gestation
in the groups receiving 1.000 or 1,500 ppm DEHP. Gravid uterine weight
exhibited a dose-related decrease in the 1,000- and 1,500-ppm dose
groups. Maternal liver weight exhibited a dose-related increase in the
two highest dose groups when compared to the controls. Clinical signs of
toxicity, which included piloerection, lethargy, and rough coat, were
seen in all of the DEHP groups in a dose-related manner.
There were no dose-related differences observed in the number of
corpora lutea or implantation sites per dam or in the percentage of
preimplantation losses. The number and percentage of resorptions [dead.
nonlive (dead plus resorbed), and affected (nonlive plus malformed)] per
litter were increased in a dose-dependent manner. These parameters were
significantly higher than the controls for the 1,000- and 1,500-ppm dose
groups. In the 1,000- and 1,500-ppm dose groups, there was a
statistically significant decrease in the number of live fetuses and che
number of males and females per litter. There was a dose-related
decrease in the body weights of the male and female fetuses. For the
1,000- and 1,500-ppm dose groups, the differences were statistically
significant when compared to the controls. There were significant dose-
related increases at the 500-, 1.000-. and 1,500-ppm doses in the number
and percentage of fetuses malformed per litter. The major malformations
included external, visceral, and skeletal defects. The NOAEL for DEHP in
this study was 250 ppm, where there was no significant maternal or
developmental toxicity.
Tomita et al. (1986) have shown that MEHP, one of the major
metabolites of DEHP, exhibits developmentally toxic effects. Mono(2-
ethylhexyl)phthalate was given to mice as a single oral dose of 0.1,
0.5. or 1 raL/kg MEHP on day 7, 8. or 9 of gestation. High-dose mice
treated on days 7 or 8 exhibited only 18.5 and 31.8% fetal survival at
parturition, respectively. In the latter group. 100% of the live fetuses
were malformed.
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ToxicoLogLcaL Data 55
Shioca et al. (1980) demonstrated that DEHP is developmentally
toxic in ICR-JCL mice. Pregnant animals were given diets containing 500
1.000. 2.000, 4.000. or 10.000 ppra DEHP throughout gestation. The mean
daily intake calculated from food consumption was 70, 190, 400. 830, and
2.200 rag/kg, respectively. Treatment with 2,000, 4,000, and 10,000 ppm
DEHP resulted in decreased maternal weight gain and increased resorpcion
rate. At the 4,000- and 10,000-ppm levels, all of the implants died. A
dose-related pattern of intrauterine growth retardation and delayed
ossification was observed. The malformation rate of fetuses at the
2,000- and 10.000-ppm dose levels was increased significantly compared
to controls. The most predominant malformations were neural tube defects
(exencephaly and spina bifIda). which suggests that DEHP may interfere
with the closure of neuropores in the developing embryo.
Shiota and Mima (1985) reported that DEHP was developmentally toxic
in ICR mice when administered orally, but not when administered by
intraperitoneal injection. Groups of pregnant mice were administered
250, 500, 1,000. or 2.000 mg/kg orally or 500, 1,000. 2,000, 4,000, or
8,000 mg/kg by injection on days 7, 8. or 9 of gestation. In the groups
treated orally, resorptions and malformations were significantly
increased at the two highest dose levels. Anterior neural tube defects
(anencephaly and exencephaly) were the most predominant abnormalities
Fetal weights were also significantly reduced. In the animals treated by
intraperitoneal injection, DEHP was abortifacient and lethal to the
pregnant females at the higher dose levels. However, no teratogenic
effects were observed. Differences in the metabolism, disposition, or
excretion resulting from the different routes of administration may be
responsible for the differences observed.
Yagi et al. (1980) evaluated the teratogenic potential of DEHP in
ddY-Slc mice. Groups of mice received a single oral administration of L
to 30 mL/kg DEHP on days 6. 7. 8. 9. or 10 of gestation. Gross and
skeletal abnormalities occurred in live fetuses from the 2.5- and 7 5-
mLAg groups treated on days 7 or 8 of gestation. The gross
abnormalities included exencephaly and club foot. Skeletal abnormalities
occurred in the skull, cervicle, and thoracic bones. Similar effects
were observed in animals treated with 0.5 or 1.0 mL/kg DEHP on day 8 of
gestation. •
4.3.3.3 Dermal
Human. No data were available on the developmental toxicity of
DEHP in dermally exposed humans.
Animal. No data were available on the developmental toxicity of
DEHP in dermally exposed animals.
4.3.3.4 Discussion
The potential developmentally toxic effects of DEHP have been
evaluated in several species. DEHP is developmentally toxic in both the
rat and mouse. In addition, the metabolite MEHP is developmentalIv
toxic. y
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56 Section 4
4.3.4 Reproductive Toxicity
DEHP has been shown to be a reproductive toxicant in CD-I mice and
Fischer 344 rats. In other studies. DEHP has been shown to damage the
cestes. The testicular effects produced by DEHP in the rat are
characterized by a decrease in the relative organ weight and
histological changes in the seminiferous tubules (Gangolli 1982).
Mono(2-ethylhexyl)phthalate, the corresponding raonoester of DEHP formed
in vivo as a result of the action of nonspecific esterases in the
intestinal mucosa and other tissues, is equally effective in inducing
testicular damage. DEHP-induced testicular injury is accompanied by a
decrease in the zinc content in the gonads and in increased urinary
excretion of this element. Exposure of preparations of rat seminiferous
tubule cells in culture to MEHP results in a dose-related detachment of
germinal cells from Sertoli cells in a manner similar to the effect seen
in the intact animal. The testicular effects observed in the rat and
mouse have not been seen in the hamster, ferret, or marmoset (Gray et
al. 1982. Lake et al. 1976, Rhodes et al. 1986, Jaeckh et al. 1984).
4.3.4.1 Inhalation
Human. No data were available on the reproductive effects of DEHP
in humans exposed via inhalation.
Animal. No data were available on the reproductive effects of DEHP
in animals exposed via inhalation.
4.3.4.2 Oral
Human. No data were available on the reproductive effects of DEHP
in orally exposed humans.
Animal. In a study performed by Reel et al. (1982), DEHP was a
reproductive toxicant in male and female CD-I mice, as evidenced by a
decreased fertility index, decreased number of litters, decreased number
of live pups, and a lower proportion of live pups per litter. Groups of
mice were given 0, 100, 1,000, or 3,000 ppm DEHP in the diet during a
7-day premating period and a 98-day cohabitation period. Fertility was
completely suppressed in the high-dose group and significantly reduced
in the intermediate-dose group. In animals receiving 1,000 ppm, the
breeding pairs produced fewer litters and had fewer male and female live
pups per litter than did the controls. In a crossover mating trial, the
proportion of detected mating did not differ significantly among
treatment groups. However, fertility was significantly reduced
(P < 0.01) in both the high-dose male/control female crossover group and
the control male/high-dose female group. Histological examination of che
high-dose males showed that the percentage of motile sperm and the sperm
concentration were significantly (P < 0.01) reduced compared with the
controls. In addition, testicular and epididymal weights were reduced.
and extensive destruction of the seminiferous tubules was observed. In
the high-dose females, reproductive tract weights (ovaries, oviducts,
uterus, and vagina) were significantly (P < 0.05) decreased when
compared with controls.
Price et al. (1986) evaluated the reproductive effects of DEHP
administered to time-mated Fischer 344 rats on gestational days 0 to 20
-------
ToxicoLogical Data 57
The treatment groups received 0. 2,500. 5.000, or 10.000 ppm DEHP in the
diet, with resultant average doses of 0, 162.5, 310.4, or 573.4
mg/kg/day DEHP. During treatment, maternal food consumption and weight
gain were reduced in a dose-related manner. In the mid- and high-dose
groups, food consumption was significantly lower than controls. However,
maternal body weight gain was significantly reduced only in the high-
dose animals.
No treatment-related effects were observed in the number of
implantation sites per dam, in the percentage of fertile matings, the
percentage of live litters, or the percentage of viable litters.
Postimplantation mortality was increased in the mid- and high-dose
groups, but this effect was statistically significant only at the middle
dose. On postnatal day 1. average litter size and average pup body
weight per litter were decreased in a dose-related manner. In the high-
dose group, the effect on pup body weight was statistically significant.
During the period between postnatal day 4 through sacrifice, the
postnatal growth, viability, age of acquisition for developmental
landmarks, and levels of spontaneous locomotor activity of the Fl
litters from DEHP-treated dams were comparable to controls. Treatment
with DEHP did not produce any adverse effects in the reproductive
performance of the Fl generation or upon the growth and viability of the
F2a litters.
Agarwal et al. (1986) investigated the sensitivity of sexually
mature rats to the toxic responses induced by the dietary administration
of DEHP. Adult male F344 rats were exposed to 0. 320, 1.250, 5.000, or
20,000 ppm DEHP for 60 days. Administration of DEHP resulted in a dose-
dependent reduction in total body weight, testis, epididymis, and
prostate weights at the two highest dose levels. In the testis,
degenerative changes, decreased testicular zinc content, reduced
epididymal sperm density and motility, and increased occurrence of
abnormal sperm were observed at 20,000 ppra DEHP. At doses of 5,000 and
20,000 ppm, there was a trend towards reduced testosterone and increased
luteinizing hormone and follicle-stimulating hormone.
On day 61 of the study, the treated males were returned to a normal
diet and mated with untreated females. The incidence of pregnancy, mean
litter weight on day 1, frequency of stillbirths and neonatal deaths,
and mean litter growth up to 7 days of age were unaffected by DEHP
treatment. However, litter size was significantly reduced in the
20,000-ppm group. Cessation of treatment resulted in partial to complete
recovery from toxicity. The magnitude of recovery was variable, with
that of Che gonads being slower than other systems. Agarwal et al.
(1986) concluded that these data suggest a lack of reproductive
dysfunction in the treated male after exposure to DEHP at levels below
20,000 ppm, which produced measurable testicular degeneration and
afflicted epididymal sperm morphology.
Parmar et al. (1986) reported that oral administration of DEHP to
adult rats at dose levels of 250. 500. 1.000, and 2,000 mg/kg for 15
days caused a significant dose-dependent decrease in the sperm count of
the epididymal spermatozoa. The activity of gamma-glutamyl
transpeptidase and lactate dehydrogenase was significantly increased in
the animals of the treated groups An increase in the activity of beta-
-------
58 Seccion 4
glucuronidase and decrease in the activity of acid phosphatase were also
observed at the highest dose of DEHP. The activity of sorbitol
dehydrogenase was found to be decreased in the animals exposed to 1,000
and 2,000 mg/kg DEHP. These results suggest that DEHP can affect
spermatogenesis by altering the activities of the enzymes responsible
for the maturation of spermatozoa. The reduced number of spermatozoa may
be responsible for the antifertilic effects of DEHP.
Gray and Gangolli (1986) found that DEHP produced seminiferous
tubular atrophy and reductions in seminal vesicle and prostate weight in
4-week-old (but not 15-week-old) rats. Coadministration of testosterone
or gonado-trophins did not protect against DEHP-induced testicular
toxicity, but did partly reverse the depression of seminal vesicle and
prostate weight. Secretion of seminiferous tubule fluid and androgen
binding protein by the Sertoli cells was markedly suppressed within 1 h
of a dose of MEHP, the major metabolite of DEHP, in immature rats. This
occurred less rapidly in mature rats. However, in distribution studies,
^C-MEHP penetrated the blood testis barrier only to a very limited
extent. These findings suggest that DEHP, rather than MEHP, act
initially to cause Sertoli cell injury, the subsequent loss of germ
cells occurring as a consequence of this. Some features of the
testicular lesion could be reproduced in primary cocultures of rat
Sertoli and germ cells. The initial Sertoli cell lesion may explain
testicular toxicity despite the limited extent to which the monoester
can penetrate the blood testis barrier. Three metabolites of MEHP were
much less toxic in culture than MEHP itself, suggesting that MEHP may be
the active testicular toxin from DEHP.
Sjoberg et al. (198Sc) studied the kinetics of DEHP in immature and
mature rats. DEHP was not found to any significant extent in the
peripheral plasma after an oral dose of 1,000 mg/kg. High plasma levels
of MEHP were found, with maximal plasma concentrations ranging from 48
to 152 ng/mL. An in vitro assessment of plasma protein binding indicated
that MEHP was approximately 98% bound in all age groups, and no age-
related difference in the elimination half-life was observed. The amount
of DEHP-derived material excreted in urine was twice as high in 25- as
in SO-day-old rats. The mean area under the curve for MEHP concentration
in plasma vs time was also significantly larger in 25- than in 40- and
60-day-old rats. These observations suggest that the extent of
absorption and/or formation of MEHP from DEHP, and hence total exposure
to MEHP and its metabolites, is higher in young than in more mature rats
after oral administration of DEHP. It is probable that this finding is
relevant to the age-related difference in the toxic effects of DEHP on
the testis.
Rhodes et al. (1986) reported that oral and intraperitoneal
administration of DEHP to the marmoset monkey at doses up to
5 irunole/kg/day DEHP for 14 days did not induce morphological or
biochemical changes in the tescls comparable with those obtained in rats
given the same amount of DEHP. This interspecies difference in
testicular effects reflects that seen in hepatic effects and may be
related to the postulated difference Ln metabolism of DEHP between rats
and primates.
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ToxicoLogical Daca 59
Sjoberg et al. (1985c) also studied the testicular response of DEH?
In immature and mature rats and effect on testis. After 14 daily oral
doses of 1.0 g/kg DEHP to 25-, 40-. and 60-day-old rats, testicular
damage was observed in the youngest age group only.
Saxena et al. (1985) reported that DEHP produced significant
histochemical and hlstological alterations in the testes of rats given
daily oral doses of 2.000 mg/kg DEHP for 7 days. Histological
examination revealed marked degeneration of the seminiferous tubules
Histochemical examination revealed patchy loss in the activity of
succinic dehydrogenase, NADH-diaphorase, and acid phosphatase, and
increases in adenosine triphosphate, glucose-6-dehydrogenase, and
alkaline phosphatase. This suggests that inhibition of enzymes involved
in energy synthesis may result in cellular damage to the target tissue
Mangham et al. (1981) administered 2,500 mg/kg/day orally to male
Wistar rats for 7 or 21 days. DEHP produced a significant reduction in
the relative weight of the testes. Light microscopic examination of che
testes of treated animals revealed that 50 to 100% of the germinal cells
were absent from the seminiferous tubules. No effects were observed in
the interstitial cells or on the Sertoli cells.
Several studies have shown that in rodents administered DEHP,
decreases in testicular zinc concentrations and increases in
testosterone accompany testicular atrophy. Oishi and Hiraga (1980) fed
JCL:ICR mice 2,000 ppm DEHP in the diet for 1 week. Treatment with DEHP
increased relative testes weight and decreased testicular testosterone
concentrations. Oishi and Hiraga (1983) found that young male rats
orally administered 2.0 mL/kg of DEHP developed testicular atrophy and
lost testicular zinc. Coadministration of 2 mL/kg of DEHP and 30 ppm
zinc sulfate in the diet for 10 days did not result in the prevention of
testicular atrophy. Despite increases in the zinc concentrations in che
liver and serum, testicular zinc concentrations were not increased This
suggests that the testicular effects of DEHP are not related to
inhibition of gastrointestinal absorption of zinc.
Oishi (1985) reported that the reversibility of DEHP-induced
testicular atrophy was limited in the Wistar rat. Animals were orally
administered 2,000 mg/kg DEHP for 14 days, and then half of the animals
were sacrificed. The testicular weights of the rats sacrificed at 14
days were significantly less than those of the controls. In addition,
testicular testosterone and zinc concentrations were reduced. The
remaining rats were allowed 45 days to recover and then were sacrificed
At the end of the recovery period the serum testosterone levels in the
treated rats were comparable to those of controls, but the testicular
zinc and testosterone levels were lower than the control values. In
addition, testicular weight of the treated animals was decreased and,
histologically, only a small number of seminiferous tubules showed
spermatogenesis.
Gray and Buttervorth (1980) found that DEHP-induced testicular
atrophy was age dependent in Uistar rats. Oral administration of 2,800
mg/kg/day DEHP for 10 days produced tubular atrophy in 4-week-old racs
but had no effect on the testes of 15-week-old rats. In rats that were
10 weeks old, testicular weight was not affected, but histological
examination revealed that 5 to 50% of che tubules were atrophlc compared
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60 Section 4
to the controls. In both the 4- and 10-week-old animals, advanced
germinal cells were lost, with only spermatogonia, Sertoli cells, and
occasional primary spermatocytes remaining. The testicular effects of
DEHP were reversible whether treatment was stopped prior to, or
continued until after, the time at which rats normally become sexually
mature.
In an attempt to further elucidate the age-dependent response of
DEHP, Sjoberg et al. (1986b) treated groups of 25-, 40- and 60-day-old
Sprague-Dawley rats by oral administration with DEHP at dose levels of
1,000 or 1,700 mg/kg/day for 14 days. Body weight gain was retarded in
all groups, and testicular weight was markedly reduced in the 25- and
40-day-old rats in the high-dose group. Repeated administration of 500
mg/kg DEHP intravenously (3-h infusions every other day for 11 days; six
infusions per animal) to 25- and 40-day-old rats did not cause age-
dependent effects. This suggests that the testicular response of younger
rats is not greater than that of older rats. It may be that the age-
related difference in testicular response after oral administration is
due to age differences in the pharmacokineti.es of DEHP (Sjoberg et al.
1985c). However, because the pharmacokinetics of DEHP after intravenous
administration has not been investigated, this possibility cannot be
verified.
Gray et al. (1982) studied species differences in the testicular
toxicity of DEHP. Groups of Sprague Dawley rats and DSN Syrian hamsters
were orally administered 2,800 or 4,200 mg/kg DEHP by oral intubation
for 9 days. In the rats, DEHP produced severe seminiferous tubular
atrophy. In contrast, the hamster showed only minor changes in response
to DEHP. The rate of intestinal hydrolysis of DEHP was significantly
slower in the hamsters than in the rats, which may account for the
observed species differences. In the same study, MEHP, the major
metabolite of DEHP, did cause focal seminiferous tubular atrophy in
hamsters.
4.3.4.3 Dermal
Human. • No data were available on the reproductive effects of DEHP
in dermally exposed humans.
Animal. No data were available on the reproductive effects of DEHP
in dermally exposed animals.
4.3.4.4 General discussion
DEHP has been shown to be a reproductive toxicant in both male and
female mice. Its effects were characterized by reduced fertility.
production of fewer litters by the breeding pairs, and the birth of
fewer male and female live pups per litter in the treated groups than in
controls. However, DEHP did not affect libido or mating performance at
dietary doses of up to 3,000 ppm (approximately 390 mg/kg/day).
At high doses, exposure to DEHP has been demonstrated to cause
testicular atrophy, characterized by degeneration of the seminiferous
tubules, in several species. The testicular effects of DEHP appear to be
related both to dose and duration of exposure. Mechanistic studies of
DEHP-induced testicular effects have focused on the role of zinc, which
-------
TOXLCOLogical Data 61
is normally found at relatively high concentrations in testicular
tissue. DEHP-induced testicular injury is accompanied by a decrease in
the zinc content in the gonads, and an increase in the urinary excretion
of this element. Although an association between testicular atrophy and
zinc depletion has been demonstrated, it is not known whether a cause-
effect relationship exists, or what role zinc has on this effect. The
age of the animals tested in studies with DEHP has been the focus of
several studies. Oral administration of DEHP at high doses has been
shown to produce testicular atrophy in immature (but not in mature)
rats. Data from metabolism studies suggest that gastrointestinal
absorption of the metabolite MEHP is greater in young animals,
suggesting that larger amounts of this active toxicant reach the target
tissues.
Numerous studies indicate that orally administered DEHP induces
dose-dependent testicular lesions. Moreover, these lesions have been
shown to be both species and age dependent. The available evidence
indicates that these species- and age-dependent differences may be
related to differences in the pharmacokinetics of DEHP, primarily
through differences in rate of intestinal hydrolysis of DEHP to MEHP
However, age- and species-dependent differences in the metabolic
conversion of MEHP to glucuronidates and/or oxidation products cannot be
discounted.
4.3.5 Genotoxicity
DEHP has been extensively tested in short-term genotoxicity assays
due to its reported carcinogenicity and its inclusion in the
collaborative study program of the International Program on Chemical
Safety (Ashby et al. 1985, Ashby 1986). The major metabolites of DEHP,
MEPH and 2-ethylhexanol, have also been studied sufficiently to permit a
determination of their genotoxicity. DEHP has not been shown to be
mutagenic in most microbial and mammalian assay systems. Most of the
data also suggest that MEHP and 2-ethylhexanol are not mutagenic.
4.3.5.1 Genotoxicity studies
The genotoxicity of DEHP has been studied extensively (Butterworth
1984, CMA 1982, Douglas et al. 1986, Hopkins 1983, Kozumbo et al. 1982.
Parry et al. 1985, Phillips et al. 1982, Priston and Dean 1985, Probst
and Hill 1985, Sanner and Rivedal 1985, Toraita et al. 1982, Vogel 1985,
Von Halle 1985, Williams et al. 1985, Wurgler et al. 1985, Yoon et al.
1985, Yoshokawa eC al. 1983, Zeiger and Haworth 1985, Zeiger et al.
1982). An overall summary of the results in specific test systems is
presented in Table 4.2.
4.3.5.2 Discussion
The overall weight of evidence indicates that DEHP is not mutagenic
in microbial or in in vivo and in vitro mammalian test systems. The
available data have not shown evidence of direct damage by DEHP to DNA
or chromosomes. Although there are several equivocally positive studies.
most of the evidence suggests that MEHP is not genotoxic. The database
for 2-ethylhexanol is less extensive, but the results suggest that it is
not mutagenic.
-------
62 Seccion <4
Tabte 4.2. Results of short-twin assays
OEHP
MEHP
Description of test
Positive Negative Positive Negative
2-Ethylhexanol
Positive Negative
Bacteria
Salmonella mutagemcity
£ co/i mutagemcity
B subnlu DNA repair
Yeast
Sacctiaromyces cerevisiae
Mutation
Crossing-over
Gene conversion
Aneuploidy
Schuosaccharomycti pombt
Mutation
Aiptrgilliu
Crossing-over
Aneuploidy
In fitro i
geootoxlciry
Chromosomal damage
Sister chromatid exchanges
Polyploidy
Aneuploidy
Mitotic spindle
Unscheduled DNA synthesis
DNA damage
la ritro
Mouse lymphoma
Mouse Balb/c3T3
Chinese hamster ovary
Chinese hamster V79
Human lymphoblasu TK
Human lymphoblasu AHH
to fifoi
genotoxktty
Micronucleus test
Human lymphocyte
chromosome damage
Sperm morphology-mouse
Sperm morphology-rat
Rat bone marrow
chromosome damage
Syrian hamster traaspla-
cental chromosome damage
Body I\w4»/Salmonella
Unscheduled DNA synthesis
DNA damage
Dominant lethality in mice'
\"
0
0
0
0
0
0
0
I
K')'
0
0
0
0
0
0
0
0
0
0
0
I
0
0
1C)
I
16
2
2
17
7
3
0
0
8
S
0
0
0
0
0
0
2
I
0
0
0
0
0
0
0
0
2
0
0
0
0
0
0
0
I
0
ll
0
0
0
0
0
0
0
0
I
0
0
0
2
I
0
I
1m
0
0
I
0
0
0
1
0
0
Im"
0
0
0
0
0
0
1m
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
I
l(')
0
0
0
0
0
0
0
0
0
0
1
0
I
0
0
0
0
0
I
0
0
0
I
0
0
2
1
-------
ToxicoLogLcaL Daca 63
Table 4.2 (cootiooed)
DEHP
Description of test
In >itT« niainnulUfl
cell transformation
Syrian hamster embryo
Mouse C3H
Mouse BALB/e 3T3
Virus-enhanced
transformation
Virus-enhanced survival
Drotofkila
Sex-linked recessive
lethal
Recombination
Somatic mutations/deletions
Recombination/somatic
mutation/aneuploidy
Test of promotion In rltro
Metabolic cooperation
Anchorage independence
Promotion of cell
transformation
Hepatocellular
hyperplastic foci
Mouse
Rats
Skin painting
SENCAR mouse
CD-I mouse
Positive
3
1
0
1
1
0
0
1m
0
2 + 1m
1
0
1
0
1
0
Negative
0
1
3
0
0
1
|
0
1
1
0
1
0
3
0
1
MEHP
Positive
1
0
0
0
0
0
0
0
0
0
1
0
0
0
0
0
Negative
0
1
1
0
0
0
0
0
0
0
0
1
0
0
0
0
2-Ethylhexanol
Positive
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
Negative
2
0
1
0
0
0
0
0
2
0
1
0
0
0
0
0
"Numeric values are number of studies.
m — marginal.
e(') ~ equivocal.
i — inconclusive.
'The positive result seen in this dominant lethal assay has not been clarified as being induced by a genetic
effect or a developmental effect caused by a nonchromosomal mechanism of action.
Source Modification (update) of table presented in CPSC 1985.
-------
64 SecCion 4
4.3.6 Carcinogenicity
DEHP is a probable human carcinogen, classified as weight-of-
evidence Group B2 under the EPA Guidelines for Carcinogen Risk
Assessment (EPA 1986a). Evidence on potential carcinogenicity from
animal studies is "Sufficient," and the evidence from human studies is
"No Data" (EPA 1986b).
4.3.6.1 Inhalation
Human. No data were available on the carcinogenic effects of DEHP
in humans exposed via inhalation.
Animal. No data were available on the carcinogenic effects of DEHP
in animals exposed via inhalation.
4.3.6.2 Oral
Human. No data were available on the carcinogenic effects of DEHP
in orally exposed humans.
Animal. The carcinogenicity of DEHP has been tested in a bioassay
using Fischer 344 rats and B6C3F1 mice (NTP 1982. Kluwe et al. 1982a,b)
Groups of 50 rats per sex were fed diets that contained 0 (control),
6,000, or 12,000 ppm DEHP, and groups of 50 mice per sex were fed diets
that contained 0 (control), 3,000, or 6,000 ppm DEHP (purity >95.5%) for
103 weeks. A 1- to 2-week nontreatment observation period was allowed
after the termination of exposure, after which all survivors were
sacrificed and examined both grossly and microscopically.
Histopathological examinations were conducted on all animals sacrificed
or discovered dead (if not precluded by autolysis). The tissues examined
included skin, lungs, bronchi, trachea, larynx, bones, bone marrow,
spleen, lymph nodes, heart, liver, pancreas, esophagus, stomach, small
intestine, large intestine, kidney, bladder, pituitary, adrenal, thymus.
thyroid, parathyroid, salivary gland, mammary gland, testis or ovary,
prostate and seminal vesicles or uterus, and brain.
A dose-related decrease in body weight gain was observed in the
male rats throughout the study. Body weight gain was also reduced in che
high-dose females. Daily mean food consumption was slightly decreased in
the low- and high-dose rats of both sexes when compared to the controls
No other compound-related clinical signs of toxicity were reported No
statistically significant dose-related trends in mortality were
observed. In male rats, 60% of the control, 56% of the low-dose, and 66%
of the high-dose animals lived to the end of the study. In female rats.
72% of the control. 68% of the low-dose, and 76% of the high-dose
animals were alive at the end of the study. A sufficient number of racs
were alive at the end of the study to be at risk from late-developing
tumors.
The incidence of female rats wich hepatocellular carcinomas at the
higher dose was greater than that in controls by pairwise comparisons,
and there was a significant (P < 0 05) dose-related trend. A dose-
related trend was also observed for hepatocellular carcinomas in the
males, but pairwise comparisons did not indicate a statistically
significant increase.
-------
Toxicological Data 65
A summary of the incidence of hepatocellular carcinomas and
neoplastic nodules observed in the rats is presented in Table 4.3.
The incidence of female rats with neoplastic nodules was greater
for the higher-dose group than in the controls and displayed a
significant (P < 0.05} dose-related trend. The incidence of male rats
with neoplascic nodules was not significantly increased when evaluated
by either pairwise comparisons or trend tests When combined, the
incidence of rats with either hepatocellular carcinomas or neoplastic
nodules exhibited a significant (P < 0.01) dose-related trend in both
sexes, and was significantly (P < 0.05) greater than the controls for
female rats at both doses and for male rats at the higher dose level by
pairwise comparisons.
The incidences of male rats with pituitary tumors, thyroid C-cell
tumors, or testicular interstitial-cell tumors were all significantly
(P < 0.05) reduced in treated animals by pairwise comparisons or by
trend test. Myeloraonocytic leukemia, mammary gland fibroadenoma,
clitoral gland carcinoma, and uterine endometrial stroraal polyps were
also observed in one or more rats in this study, but their incidences in
treated animals did not differ significantly from chose in the controls.
In mice, there was a dose-related decrease in the body weights of
the female from week 25 to the end of the study. Daily mean food
consumption was similar in the treated and control groups throughout the
study. No other compound-related clinical signs of toxicity were
reported. In male mice, 68% of the control, 76% of the low-dose, and 70%
of the high-dose animals lived to the end of the study. In female mice,
72% of the control, 68% of the low-dose, and 76% of the high-dose
animals were alive at the end of the study. Low-dose females had
significantly shortened survival compared with that of the controls
(P - 0.006), but no positive trend was observed with respect to survival
when all groups were compared. A sufficient number of mice were alive at
the end of the study to be at risk from late-developing tumors.
There was a statistically significant increase in the incidence of
hepatocellular carcinoma in both the high-dose males and high-dose
females when compared with the controls. A significant (P < 0.05),
dose-related trend of Increasing hepatocellular carcinoma was also
observed for the alee of both sexes. A summary of the incidence of
hepatocellular carcinomas and hepatocellular adenomas observed in the
mice is presented in Table 4.4.
Metastases of the hepatocellular carcinomas were observed in the
lungs of 12 treated males and B treated females. The incidence of
neoplastic lesions of other sites in the treated animals did not differ
significantly from that of controls.
Northrup et al. (1982) have questioned the validity of the NTP
bioassay of DEHP. They suggested that the MTD was exceeded in the low-
and high-dose female mice, low- and high-dose male rats, and high-dose
female rats because body weight gain was depressed by more than 10% in
these groups. However, Kluwe et ai. (1983) have replied that the 10%
weight differential was only a guideline and that the primary reason for
not exceeding the estimated MTD was to avoid excessive early mortality
which might preclude tumor development and to avoid pathological lesions
-------
66 Seccion
Table 4.3. Incidence of hepatocellular carcinomas and
neoplastic nodules observed in DEHP-dosed rats"
Dose Hepatocellular Neoplastic All liver
(ppm) carcinomas nodules tumors
Males
0 1/50 2/50 3/50
6.000 1/49 5/49 6/49
12.000 5/49 7/49 12/49
Females
0
6.000
12.000
0/50
2/49
8/50
0/50
4/49
5/50
0/50
6/49
13/50
"References- NTP 1982; Kluwe et al. 1982a,b.
Table 4.4. Incidence of hepatocellnlar carcinomas and
hepatocellular adenomas observed in DEHP-dosed mice"
Dose Hepatocellular Hepatocellular All liver
(ppm) carcinomas adenomas tumors
Males
0 9/50 6/50 14/50
3000 14/48 11/48 2S/48
6000 19/50 10/50 29/50
Females
0
3000
6000
0/50
7/50
17/50
1/50
5/50
1/50
1/50
12/50
18/50
"References: NTP 1982: Kluwe et al. 1982a.b.
-------
Toxicological Data 67
other than neoplasia (Haseman 1985). In the case of the DEHP bloassay.
NTP (1982) concluded that both of these goals had been met. Survival was
not adversely affected by DEHP, and statistical analyses failed to
demonstrate any correlation between the occurrence of nonneoplastlc
tissue lesions (testicular atrophy In the high-dose male rats and mice
and pituitary hypertrophy in the high-dose male rats) and the
development of hepatocellular tumors in rats and mice of both sexes
(Kluwe et al. 1982a,b, 1983). However, there is a second issue with
regard to the use of the MTD in this study. As discussed below, there is
some evidence that suggests that peroxisorae proliferation, which occurs
in both mice and rats at the dose levels used in the NTP bioassay, is
involved in a secondary mechanism of cancer induction (Reddy et al.
1986). If this hypothesis is correct, it suggests that at low doses,
where peroxisome proliferation does not occur, one would not expect
cancer to develop (Turnbull and Rodricks 1985, Warren et al. 1982).
Northrup et al. (1982) also questioned the significance of the
DEHP-induced increase in liver tumors in the NTP bioassay because of
intralaboratory variations in incidences of liver tumors in controls in
bioassays of guar gum, butyl benzyl phthalate, and di(2-ethylhexyl)
adipate, which were conducted simultaneously in the same rooms as the
DEHP study. They reported that statistical analysis indicated that there
was a significantly lower incidence of tumor-bearing animals among the
female mice used in the DEHP control group. In addition, they suggest
that the number of tumor-bearing male or female mice in the DEHP
treatment groups did not substantially differ from the average for all
pooled controls. They also report that in male rats, the incidence of
tumor-bearing animals had a highly negative dose response (P < 0.001),
and female rats had a marginally positive dose response (P - 0.045)
when paired to the pooled controls. However, Kluwe et al. (1983)
reported that there was relatively little variation in liver tumor
incidences among both NTP historical controls and concurrent laboratory
controls, and that the data reported in the concurrent study controls
were quite similar to the historical controls (Haseman 1983). In
addition, Kluwe et al. (1983) reported that replacement of the
concurrent study controls with either concurrent laboratory controls or
pooled controls from bioassays performed at approximately the same time
did not alter the statistical significance of the DEHP effects.
Replacement of the concurrent study controls with historical controls
strengthened the statistical significance of the difference in liver
tumor incidence.
Carcinogenic effects were not reported in 2-year bioassays with
Sherman or Wistar rats of both sexes fed diets which contained up to
5,000 ppa DEHP (Carpenter et al. 1953, Harris et al. 1956). However, the
design and reporting of these studies were not adequate to fulfill the
stringent Good Laboratory Practices and Good Management Practices
required of today's toxicity testing guidelines.
4.3.6.3 Dermal
Human. No data were available on the carcinogenic effects of DEHP
in dermally exposed humans.
-------
68 Section 4
Animal. No data were available of the carcinogenic effects of DEHP
In dermally exposed animals except for those reported in Sect. 4.3.6 4.
The carcinogenic risk due to dermal exposure to DEHP is likely to be
low, based on observations of poor dermal absorption (El Sisi et al.
1985) and an absence of initiation or promotion activity in mouse skin
painting studies (Ward et al. 1986).
4.3.6.4 Possible mechanisms of carcinogenesls
Several initiation-promotion studies have been performed with DEHP
Ward and coworkers (Diwan et al. 1983, 1985; Ward 1983, 1986) conducted
skin-painting studies to assess the promotional activity of DEHP. Single
topical applications of 50 pg of 7,12-dimethylbenz(a)anthracane (DMBA),
an initiator, were applied to the skin of CD-I mice, followed by DEHP
(98.1 pg in acetone) twice weekly for 40 weeks. A known promoter, 12-0-
tetradecanoyl-phorbol-13-acetate (TPA), was used as a positive control
Under the conditions of this study, DEHP was not a promoter. No tumors
were observed in animals treated with DMBA followed by DEHP. In
contrast, 97% of the mice treated with DMBA followed by TPA developed
skin tumors.
In a second study performed by Ward and coworkers, female SENCAR
mice received a single topical application of 20 /jg of DMBA, followed by
2 pg of TPA twice weekly for 2 weeks, followed by 100 /jg of DEHP twice
weekly. Animals treated with TPA, mezerein, or acetone served as
controls. Under the conditions of the study, DEHP showed some late
promotional activity, producing 6.4 papillomas per mouse. In contrast,
DMBA + TPA + mezerein produced 23 papillomas per mouse, DMBA + TPA + TPA
gave 26.4 papillomas per mouse, DMBA + DEHP + DEHP gave 0.9 papillomas
per mouse, and DMBA alone produced no tumors.
Albro et al. (1982) reported that radioactivity from carbonyl-
labeled DEHP did not bind to purified protein, RNA, or DNA from rat
liver in vivo. 0-ethyl-(l-14C)-hexyl-labeled DEHP and MEHP did appear to
bind strongly with purified DNA, but the label from free 14C-labeled 2-
ethylhexanol did not. The apparent binding of DEHP and MEHP was not
exchangeable and was not demonstrated to be covalent.
DeAngelo et al. (1985a) examined the protein kinase activity of
isolated plasma membranes from the livers of rats treated with three
promoting regimens. Male rats first received either an initiating dose
(30 rag/kg) of the hepatocarcinogen diethylnitrosamine or the 0.9% NaCl
solution vehicle by intraperitoneal injection at 18 h following partial
hepatectomy (to induce cell proliferation). Ten days later, the three
promoting regimens were begun. These consisted of 10 weeks of treatment
with either (a) a choline-deficient (CD) diet, (b) a choline-
supplemented (CS) diet containing 0.06% phenobarbital (PHB) (CS plus
PHB), or (c) a CD diet containing 0.06% PHB (CD plus PHB). In addition,
two other groups of rats received either (a) a CS diet containing 2%
DEHP (CS plus DEHP) or (b) a CD diet containing 2% DEHP (CD plus DEHP)
DEHP had been shown previously to inhibit the development of putative
preneoplastic gamma-glutamyl transpeptldase (GOT) positive foci in rat
liver. Total liver plasma membrane protein kinase (PK) activity, using
both protamine sulfate andhistone. was cyclic adenosine 3':5'-
monophosphate independent and did not appear to be a marker of
-------
ToxLcologicaL Daca 69
promotion. Protein kinase activity, however, was increased by both DEHP
(which suppresses the development of GGT positive foci) and a CD diet
(which promotes the appearance of GGT positive foci). The CD, CS plus
PHB, and CD plus PHB dietary regimens, which promote the appearance of
GGT positive foci, induced the phosphorylation of a 40,000 molecular
weight (raw) plasma membrane protein in vitro by endogenous protein
kinases. Plasma membranes from DEHP-treated rats did not demonstrate
phosphorylation of this 40,000-mw protein. DEHP dietary treatment also
blocked the ability of the epidermal growth factor to enhance the
phosphorylation of its 175.000-mw receptor protein in isolated liver
plasma membranes. These results suggest that the phosphorylation of a
40,000-raw plasma membrane protein may be important to the early
promotional phase of liver carcinogenesis, and that one mechanism by
which DEHP inhibits the emergence of GGT positive foci may be by
blocking the response of initiated cells to stimulation by epidermal
growth factor.
As discussed above, DEHP has been demonstrated to cause an increase
in the incidence of liver tumors in the F344 rat and the B6C3F1 mouse
(NTP 1982). The weight of evidence suggests that DEHP is not genotoxic
in either in vitro or in vivo assay systems. Several hypotheses have
been proposed to explain the mechanism(s) by which DEHP induces liver
tumors in rodents. Reddy and Lalwani (1983) suggested that DEHP belongs
to a novel class of nonmutagenic chemical carcinogens which alter the
level and function of peroxisomes in rodent liver cells. This hypothesis
assumes that the hepatocarcinogenesis of DEHP is not related to a direct
initiating effect of DEHP (or its metabolites), but is linked to
metabolic disturbances emanating from a sustained increase in the number
of peroxisomes in liver cells (Rao and Reddy 1987). The terra "peroxisome
proliferation" refers to an increase in the number of subcellular
organelles called peroxisomes in response to administration of a series
of compounds collectively called "peroxisome proliferators" (CPSC 1985)
The peroxisome proliferating compounds are structurally diverse. The
only thing these compounds have in common is the capability to induce
increased numbers of peroxisomes and to produce hypolipidemia. They have
been characterized by the CPSC (1985) as having the following features
• Induction of increased numbers of peroxisomes in several species,
• Induction of variable degrees of hepatocyte hyperplasia,
• Lowering of the concentrations of plasma lipids,
• Induction of liver carcinogenesis, and
• Weak or absent mutagenicity in in vitro rautagenicity bioassays.
The phenomenon of peroxisome proliferation and its relationship co
the carcinogenicity of DEHP has been studied extensively and has been
the subject of several recent reviews (Reddy et al. 1986, Rao and Reddy
1987, CPSC 1985).
Liver enlargement occurs within a few days of dietary
administration of peroxisome proliferators such as DEHP, rapidly reaches
a steady-state level (in 7 to 10 days), and continues as long as
administration of the proliferator continues (Reddy and Lalwani 1983)
The peroxisome proliferators are not equipotent in the degree of
-------
70 Section 4
observed hepatomegalic effect chey induce. DEHP is a relatively weak
peroxisome proliferates when compared to compounds such as ciprofibrate
(by a factor of 1.000) (Reddy et al. 1986). The liver enlargement
associated with the administration of peroxisome proliferators is due to
both hyperplasia and hypertrophy of hepatocytes (Rao and Reddy 1987)
The hyperplasia is evident as early as 24 h after treatment, reaching a
maximum in 5 to 7 days. Hypertrophy of the hepatic cells is secondary co
marked increases in the number and volume of peroxisomes.
The development of hepatocellular carcinomas due to administration
of a peroxisome proliferator was first reported by Reddy et al. (1976)
Since then, several hypolipidemic compounds and plasticizers that are
peroxisome proliferators have been shown to induce liver tumors in mice
and rats (Rao and Reddy 1987). The latency period and incidence of
tumors appear to correlate well with the ability to induce peroxisome
proliferation. Potent peroxisome proliferators such as Wy-14643,
ciprofibrate, and methyl clofenapate induce liver tumors in nearly 100%
of treated rats after SO to 60 weeks of administration (Rao and Reddy
1987). Less potent peroxisome proliferators such as clofibrate and DEHP
induce liver tumors after 70 and 104 weeks, respectively (Svoboda and
Azarnoff 1979, NTP 1982). The majority of the liver tumors induced by
the peroxisome proliferators are well-differentiated hepatocellular
carcinomas, with predominantly trabecular histological patterns (Rao and
Reddy 1987). Metastases are encountered in 20 to 40% of rats and mice
with peroxisome-proliferator-induced hepatocellular carcinomas (Rao and
Reddy 1987).
The lack of mutagenicity of the peroxisome proliferators in in
vitro and in vivo assays has generated interest in the mechanism of
action by which these compounds induce liver tumors. Several hypotheses
have been posed stating that the carcinogenicity of these compounds is
due to biologically active products of the proliferated peroxisomes
rather than direct DNA damage. The evidence supporting this hypothesis.
which was recently summarized by Rao and Reddy (1987), includes (1) a
consistent association between the induction of peroxisome proliferation
and liver carcinogenesis; (2) sustained and specific induction of H202-
producing beta-oxidation enzymes; (3) increased intracellular levels of
H202 in livers of rats with peroxisome proliferation; (4) increases of
hepatic lipofuscin and increased hepatic levels of diene conjugates,
indicative of increased lipid proliferation; and (5) marked inhibition
of peroxisome proliferator-induced hepatocarcinogenesis by antioxidants
The importance of oxidative stress on the process of initiation and
promotion of carcinogenesis remains unknown. The peroxisomes contain a
variety of enzymes associated with llpid metabolism (CPSC 1985). Several
oxidation reactions can be catalyzed by the enzymes residing in the
peroxisomes. These reactions employ as their terminal oxidase an H202-
generating flavin oxidase. The H202 produced is converted to H20 by
catalase, either by directly converting H202 to H20 plus 1/2 02 or by
utilizing two hydrogen atoms from suitable molecular donors to convert
H202 to two molecules of H20 plus an oxidized substrate.
The administration of drugs such as hypolipidemic agents and
several phthalates results in a sudden proliferation of peroxisomes
followed by a significant increase in parcicle-bound catalase activity
-------
Toxicological Data 7L
The mechanism by which peroxisome proliferators exert their effects is
not well understood at present and has been the subject of considerable
research. Several investigators have proposed that these compounds
induce peroxisome proliferation by increasing the influx of lipids, such
as long-chain fatty acids, into hepatocytes (Reddy and Lalwani 1983.
Havel and Kane 1973). This results in an increase in the number of
peroxisoraes, presumably due to the induction and increased synthesis of
the enzymes and other components that constitute these organelles (CPSC
1985). This hypothesis is supported by the finding that feeding rats a
diet high in fats results in a slight increase in the number of
peroxisomes (CPSC 1985). However, this increase is very slight compared
to the increase following administration of peroxisome proliferating
compounds. The results of several studies suggest that there is no
direct lipolytic effect by peroxisome proliferators. Lake et al. (1983)
reported that in a comparison of the effects of several phthalate
esters, both DEHP and mono-n-octyl phthalate had hypolipidemic effects,
while peroxisome proliferation was seen only with DEHP and its
metabolites. Similar findings were reported in a study by Lazarow and
DeDuve (1976) with bezafibrate and clofibrate. in which both compounds
had hypolipidemic effects, but only clofibrate induced peroxisomes.
An alternative hypothesis is that peroxisome proliferators act as
substrates for the peroxisomal beta-oxidation system (Reddy and Lalwani
1983, Reddy 1987). This hypothesis is supported by the fact that some
components of the molecular structure of the peroxisome proliferators
appear as suitable candidates for metabolism by the peroxisomal enzymes
(CPSC 1985). The combined effect of fatty acid influx with influx of the
peroxisorae proliferator is sufficient to result in peroxisome
proliferation. Phthalate esters appear to undergo beta oxidation, and
this could occur in the peroxisomes. A study by Lhuguenot et al. (1985)
suggests that at high doses, MEHP is partly metabolized by peroxisomes
A. M. Mitchell et al. (1985) reported that peroxisome proliferation is
associated with concomitant increase of a cytochrome P-450 species that
is associated with omega oxidation of lauric acid and other long-chain
fatty acids. They proposed that peroxisome proliferation and cytochrome
P-450 induction are attempts by the rat hepatocyte to cope with a
disturbed lipid metabolism (CPSC 1985). Because of the inability of the
rat to conjugate these compounds, their presence leads to a substrate
load that is alleviated by the creation of these alternative pathways
(CPSC 1985). At high doses of DEHP. excess H202 or other oxygen species
are produced in excessive amounts because catalase production does not
increase as rapidly as peroxide production.
Recently, Rao and Reddy (1987) have proposed that peroxisome
proliferacors exert their effect by a ligand-receptor-mediated
mechanism. According to this hypothesis, the peroxisome proliferators
interact with a receptor molecule present in the cytoplasm or plasma
membrane of hepatocytes. The formation of a receptor-peroxisome
proliferator complex results in the generation of a message that is
transmitted to the cell nucleus, initiating the process for production
of peroxisomal components (CPSC 1985). The interaction between the
ligand and the receptor results in a measurable effect only after a
sufficient number of receptors are occupied by the ligand. Rao and Reddy
(1987) have stated that indirect evidence in favor of this hypothesis
-------
72 Section 6
includes (1) tissue-specific predictable biological response, (2)
response of extrahepatic hepatocytes to the inductive effects of these
compounds, (3) inducibility of peroxisome proliferation in hepatocytes
in primary cultures maintained in defined medium, (4) induction of
specific changes in protein composition in the livers of rats treated
with peroxisome proliferators which differ in chemical structure, (5) a
rapid and significant increase in the rate of synthesis of mRNAs for
peroxisomal beta-oxidation enzymes in the liver, (6) the rapidity of
transcriptional response of these genes, and (7) detection of a specific
binding moiety in liver cytosol for nafenopin, a clofibrate-like
peroxisome proliferator. In a study published by Lalwani et al -(1983),
3H-nafenopin, a known inducer of liver peroxisomal enzymes, was shown to
bind to a specific, saturable pool of binding sites in rat liver and rat
kidney cortex cytosols. Clifibrate and ciprofibrate, which are
structurally similar to nafenopin, competitively inhibited the specific
binding of 3H-nafenopin. Phenobarbital (a noninducer of peroxisomes),
4-chloro-6-(2,3-xylidino)-2-pyrimidinylthio acetic acid, and 4-chloro-
6-(2,3-xylidino)-2-pyrimidinylthio (Af-beta-hydroxyethyl) acetamide did
not compete for the specific 3H-nafenopin binding sites. This study
provides some evidence for the presence of such a receptor. Additional
studies are being conducted to identify and purify the putative receptor
protein.
DEHP was identified as a peroxisome proliferator in 1976 (Reddy et
al. 1976). DEHP induces a stronger response in the rat compared to the
hamster (Turnbull and Rodricks 1985, Rodricks and Turnbull 1987). In a
report by the Council European of the Federations of the Industry
Chemical (CEFIC) (1982) presented to the CPSC (1985), it was reported
that the induction of peroxisome proliferation occurs in the marmoset to
a much lower degree than in the rat. However, in the same report, it was
also stated that the extent of absorption of DEHP in the marmoset was
much lower than in the rat. Peroxisome proliferation, such as that seen
in the rodent, has not been quantitatively demonstrated in higher
primates or in humans (De La Iglesai et al. 1982, Staubli and Hess 1975,
Sternlieb 1979). Peroxisome nucleoids are absent from the liver of
humans, are rudimentary in some primates, and are well developed in the
liver of rodents (De La Iglesia et al. 1982). Differences in the
relative proportions of metabolites of DEHP have been described for the
rat, nonhuman primates, and humans. Relatively high quantities of
omega-1 oxidation products of MEHP are formed in primates and humans
when compared to the rat. These metabolites are excreted largely as
conjugated products in humans, primates, and the mouse, but not in the
rat (Albro et al. 1981, 1982). The differences between the species in
terms of metabolite profiles and conjugation pathways preclude
definitive conclusions as to the expected sensitivity to DEHP
carcinogenicity (CPSC 1985). Most of the available evidence does suggest
that primates in general are less sensitive to the induction of
peroxisome proliferation than are rodents (Lake et al. 1984).
Although the absence of mutagenic activity is consistent with other
evidence, which suggests that DEHP exerts its carcinogenic effect by an
epigenetic mechanism, there are not sufficient data to allow a
determination of a dose of DEHP below which peroxisome proliferation
should not be expected to occur in a long-term study (CPSC 1985). All of
-------
Toxicological Daca 73
nr«MfM °f ^ mechanisffl of induction of peroxisome
proliferation suggest an effect will occur only after sufficient
concentrations of the inducing compound are reached The available
studies (CMA 1984; 1985a.b.C.d; 1986a,b.c(d; Morton I™?; Red^y 1985)
Torecfj rF-^'f^0", ^'^ f™ b"n "° ^"ft""-" «udl.. that allow
a precise definition of the relationship between DEHP dose and
peroxisome proliferation. In addition, although there is much indirect
evidence relating peroxisome proliferation to carcinogenesis. this
Uln-M " rt* T ?6n Pr°Ved directlv- Uncil ^ch evidence becomes
available, the theories regarding the mechanism of action of DEHP-
induced carcinogenicity must be regarded as plausible but unproven
Continuing research is necessary to resolve this issue.
Lake et al. (1984) administered 1,000 mg/kg/day DEHP by gavage
daily to male Sprague-Dawley rats for 14 days. Concurrent control
TP ITnSn^C? °nly,Che Corn o11 vehicl.. DEHP produced significant
(P < 0.01) increases in relative liver weight, marked increases in the
hepatic activities of cyanide-insensitive palmitoyl-CoA oxidation a
specific peroxisomal marker enzyme, carnitine acetyltransferase (an
enzyme located in mitochondrial, peroxisomal. and microsomal fractions)
and both total (mitochondrial and peroxisomal) and heat labile
(peroxisomal) enoyl-CoA hydratase activities. In addition DEHP
treatment resulted in inhibition of D-amino acid oxidase activity. The
effect of DEHP on hepatic microsoraal mixed-function oxidase parameters
was also investigated. DEHP markedly induced the combined omega- and
omega-l-hydroxylation of lauric acid, with smaller increases observed in
the activities of ethylmorphine tf-demethylase and 7-ethoxycouraarin O-
deethylase. In contrast, the activity of 7-ethoxyresorufin O-deethylase
was significantly inhibited. DEHP also induced cytochrome P-450 content
and produced spectral changes in the properties of hepatic microsomal
hemoproteins. Histological studies revealed marked peroxisome
proliferation in liver sections from treated rats.
DeAngelo et al. (1985b) studied the effect of diets containing 0.1
0.5. 1.0. and 2.0% DEHP as equimolar amounts of its two major
metabolites. MEHP and 2-ethylhexanol. on the induction of liver
peroxisomes and pre-induced GGT positive (GGT+) foci GCT+ foci were
initiated in the livers of Sprague-Dawley male rats with a single dose
of di.thylnleroM.in. (DEN) following partial hepatectomy. Controls were
fed a semipurified diet for 10 weeks; experimental groups received the
semipurified diet containing the respective compounds. Induction of
/rr^lfOBe Proliferaclon w« monitored by carnitine acetyl-transferase
(CAT) levels. The 2-ethylhexanol produced essentially no effect with
nrSrd ?u^b?r °f f°Cl> Peroxiso«"e proliferation, or liver weight.
DEHP and MEHP induced significant peroxisome proliferation and
hepatomegaly.
Toraaszewski et al. (1986) investigated the hypothesis that
hepatocarcinogenesis resulting from treatment of rats and mice with
peroxisome proliferators is linked to increased cellular levels of
hydrogen peroxide from peroxisomal beta-oxidation. Hale F344 rats and
female B6C3F1 nice were treated for 14 days with DEHP di(2-
ethylhexyl)adipate (DEHA). or nafenopln. a hypolipidemic drug.
Activities of enzymes responsible for the production [peroxisomal
palmitoyl CoA oxidase (PCO)] and degradation [catalase (Cat) and
-------
74 Section 4
glutathione peroxidase (GSHPx)] of H202 were assayed in liver
homogenaces prepared from created animals. The activities of the
peroxisomal enzymes PCO and Cat were enhanced 5- to 25-fold and 1.5- to
3-fold, respectively, by treatment with DEHP, DEHA, and nafenopin. The
activity of GSHPx, a cytoplasmic enzyme, was decreased 40 to 60% in
liver homogenates prepared from treated animals compared to control
animals. A kinetic analysis of the rates of formation of hydrogen
peroxide by PCO, and of degradation of hydrogen peroxide by catalase,
was used to estimate steady-state hydrogen peroxide concentrations
[(H202)] during peroxisomal oxidation of palmitoyl CoA. Increases in
peroxisomal steady state (H202) for the F344 rat liver homogenates
correlated well with the carcinogenic potential of these chemicals,
determined in previous carcinogenicity studies. Increases in the steady
state (H202) were also calculated for liver homogenates prepared from
mice treated with these compounds. Decreases in liver lipid peroxidation
were observed after treatment with each chemical in both species. The
results of these studies are consistent with an involvement of increased
peroxisomal hydrogen peroxide in the hepatocarcinogenesis of these
compounds.
Lake et al. (1986) investigated the hepatic effects of DEHP on
peroxisomal and microsomal enzyme activities in the intact animal and in
primary hepatocyte cultures. In the in vivo studies, DEHP produced large
increases in liver size and peroxisomal enzyme activities in Sprague-
Dawley rats and Chinese hamsters, but had less effect in Syrian
hamsters. These effects were largely reproduced in vitro, where good
responses were obtained with rat and Chinese hamster hepatocytes, but
either little or no effect was observed with Syrian hamster and Dunkin-
Hartley guinea pig hepatocytes. These results demonstrate a good
relationship between the in vivo and in vitro effects of DEHP and
indicate that hepatocyte cultures may be useful for further studies on
the hepatic effects of DEHP on peroxisome proliferation.
DeAngelo et al. (1986) studied the ability of DEHP to suppress the
development of rat liver preneoplastic lesions. GGT+ foci were initiated
in the livers of Sprague-Dawley male rats with a single dose of
diethylnitrosaraine (DEN) following partial hepatectomy. Promotion of
foci was induced by feeding the rats a choline-deficient diet (CD).
Control rats were fed a choline-supplemented diet (CS). The ability of
DEHP to suppress the emergence of GGT+ foci was evaluated by feeding
groups of rats the CD diet containing either 0.1. 0.5. 1.0, or 2.0%
DEHP. The CD diet promoted the number of GGT+ foci above levels in
control livers. Inclusion of DEHP in the CD diet at levels of 0.5. 1.0,
and 2.0% effectively inhibited the appearance of the foci. However, DEHP
was unable to inhibit the promoting effect of the CD diet at a
concentration of 0.1%. DEHP's ability to block development of GGT+ foci
correlated with its ability to increase liver weight and to induce
carnitine acetyltransferase, a marker of peroxisome proliferation.
A. M. Mitchell et al. (1985) used a primary rat hepatocyte culture
system to determine the proximate peroxisome proliferator(s) derived
from DEHP. DEHP was administered orally to rats, and the urinary
metabolites were isolated and identified The major metabolites were
those resulting from initial omega- or omega-1-carbon oxidation of the
-------
TOXI.COLogical Data 75
mono(2-ethylhexyl) moiety. These metabolites, together with MEHP and 2-
ethylhexanol, were added to primary rat hepatocyte cultures, and the
effect on peroxisomal enzyme activity was determined. The omega-carbon
oxidation products (mono(3-carboxy-2-ethyl-propyl) phthalate and
mono(5-carboxy-2-ethylpentyl) phthalate] and 2-ethylhexanol produced
little or no effect on cyanide-insensitive palmitoyl-CoA oxidation, a
peroxisomal marker. Mono(2-ethyl-hexyl) phthalate and the omega-1-carbon
oxidation products [mono-(2-ethyl-5-oxohexyl) phthalate and mono(2-
ethyl-5-hydroxyhexyl) phthalate] produced a large (7- to 11-fold)
induction of peroxisomal enzyme activity. Similar structure-activity
relationships were observed for the induction of cytochrome P-450-
mediated lauric acid hydroxylase and the increase in cellular coenzyrae A
content. In contrast. A. M. Mitchell et al. (1985) reported that oral
administration of MEHP (150 or 250 mg/kg) to male guinea pigs did noc
produce hepatic peroxisome proliferation. Addition of MEHP (0 to 0.5 mM)
or one of the active proliferators in the rat to primary guinea pig
hepatocyte cultures also failed to produce an induction of peroxisomal
beta oxidation.
Kornbrust et al. (1984) performed experiments to test the
hypothesis that the hepatocarcinogenicity of DEHP is due to its ability
to produce DNA damage, either directly or as a result of the
proliferation of peroxisomes and accompanying increased production of
H202 and other DNA-damaging oxygen radicals induced by sustained
exposure to this chemical. DEHP did not appear to directly produce
repairable DNA damage in rat hepatocytes. The DNA damage, as assessed by
the autoradiographic measurement of unscheduled DNA synthesis, was not
observed in primary rat hepatocytes exposed in vitro to DEHP or in vivo
by a single gavage dose of 5 ^g/kg DEHP administered 2, 15, or 24 h
prior to the isolation of hepatocytes.
Sustained feeding of DEHP at a dietary concentration of 20,000 ppm
led to a marked proliferation of peroxisomes in the liver after 4 weeks
of treatment. Additional administration of a single gavage dose of 5
g/kg DEHP to animals fed the 20,000-ppm diet for 4 or 8 weeks, as well
as to 4-week-fed animals that were also pretreated with 3-amino-l,2,4-
triazole (to inhibit endogenous catalase activity), did not induce any
detectable. DNA repair in hepatocytes isolated 15 h following the single
gavage dose of DEHP. Lipid peroxidation in livers of animals treated
either with a single dose of 5 g/kg DEHP or with a 20,000 ppm DEHP diet
for 6 weeks, plus a single dose of 5 g/kg, did not differ from controls
These findings suggest that DEHP does not elicit DNA damage or llpid
peroxidation in liver consequent to the proliferation of peroxisomes
resulting from 4 to 8 weeks of administration.
Lak*-et al. (1984) observed a marked species difference in response
to peroxisome proliferators. Male Sprague-Dawley rats and Syrian
hamsters were treated with 25, 100, 250. and 1,000 mg/kg/day DEHP orally
for 14 days. Liver enlargement was observed in both species, with more
marked effects being observed in the rat. Liver weights were increased
significantly at doses of 100 mg/kg and greater in the rat, but only at
1,000 ag/kg in the hamster. In the rat there was a marked dose-dependent
induction of the peroxisomal marker cyanide-insensitive palmitoyl-CoA
oxidation and also of carnitine acetyltransferase. Little effect was
observed on the mitochondria! markers carnitine palmitoyltransferase and
-------
76 Section 4
succinace dehydrogenase. Increased peroxisomal enzyme activities were
observed after treatment with 100 and 250 mg/kg/day DEHP.
In contrast, much Less effect was observed in the hamster, even
after 1,000 mg/kg/day DEHP. Parallel morphological investigations
demonstrated a greater increase in hepatic peroxisome numbers in the rat
than in the hamster. ^C-labeled DEHP was found to be more rapidly
hydrolyzed by rat than hamster hepatic and small intestinal mucosal cell
preparations, and differences were also observed in the absorption and
excretion of oral doses of 14C-DEHP. Studies with MEHP and a
hypolipidemic drug, clofibrate, also resulted in a greater increase in
hepatic peroxisomal enzymes in the rat compared to the hamster.
4.3.6.5 General discussion
Based on the weight of evidence, there is sufficient reason to
conclude that DEHP is a carcinogen in experimental animals. DEHP,
administered in the diet, was carcinogenic in rats and mice, producing
increased incidence of liver cancers in female rats and in male and
female mice. Because DEHP is not mutagenic and has been demonstrated to
be a peroxisome proliferator, the excess liver cancers may be due to
peroxisome proliferation. It is unclear whether this effect is likely Co
occur in man, because of differences in'metabolism and liver cell
susceptibility to peroxisome induction, and because structural features
of peroxisomes in humans make these organelles more difficult to assay.
If peroxisomal induction is the only mechanism of action for tumor
induction in rodents (which is not known), a dose level may exist below
which no tumors would occur. However, current methods of assessing the
carcinogenic potential of chemicals and methods for high-to-low dose and
interspecies extrapolation are not sufficiently sensitive to estimate
such a dose.
4.4 INTERACTION VITH OTHER CHEMICALS
No studies have been identified which have investigated the effects
of DEHP when administered with other chemicals.
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77
5. MANUFACTURE. IMPORT. USE. AND DISPOSAL
5.1 OVERVIEW
Eighc major plants, locaced mostly along the Gulf Coast and in che
northeastern states, produced approximately 181,700 metric tons (kkg) of
DEHP in the United States during 1977. After accounting for imports and
exports, most of this total production, or about 167,600 kkg, was
available for consumptive use during that year. By 1982, total DEHP
production dropped to about 114,000 kkg; in 1986, production was back up
to 131,543 kkg.
The major uses of DEHP are as a plasticizer in polyvinyl chloride
(PVC) resins and vinyl copolymer resins. Together, these uses accounted
for more than 90% of the DEHP produced in the United States in 1979
Most of the processing wastes and by-products containing DEHP are
disposed of on land; relatively small amounts are found in wastewater
5.2 PRODUCTION
DEHP Is produced commercially by the esterification of phthalic
anhydride with 2-ethylhexanol (octyl alcohol). The reaction is catalyzed
by p-toluene sulfonic acid or by an amphoteric catalyst. The acid-
catalyzed process involves a water-wash purification step, whereas che
amphoteric process does not involve water washing. The excess alcohol LS
recovered and recycled. The product is purified by vacuum distillation
and/or with activated charcoal, and the yield is >90% (EPA 1981, HSDB
1987, IARC 1982).
There were eight major producers of DEHP in the United States in
1977, operating plants in six states. Most of the total 1977 production
of about 181,700 kkg came from two sites (EPA 1981).* Currently, there
are eight-major DEHP production plants (SRI 1987). DEHP accounted for
about one-third of the total phchalate ester production in the United
States in 1977. In 1982, DEHP production was reduced to 114,000 kkg
(HSDB 1987), but, by 1986, production was 131,543 kkg (EPA 1987b).
5.3 IMPORT
No data were located on importation of DEHP into the United States
in 1977. Approximately 42,500 kkg of phthalate esters were exported in
1977. It is estimated that about 14,000 kkg of these exports consisted
of DEHP (EPA 1981. HSDB 1987). The United States imported 2,722 kkg of
DEHP in 1986 and exported 5,443 kkg (EPA 1987b).
*This citation erroneously states that Monsanto Company was a 1977
producer of DEHP. Monsanto discontinued DEHP in the early 1970s.
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78 Seccion 5
5.4 USE
The major consumptive use of DEHP is as a plasticizer for PVC
resins (EPA 1981, HSDB 1987). It is also used as a plasticizer for vinyl
chloride copolymer resins. Together, these uses consumed 94% of the DEHP
produced in the United States in 1979 (HSDB 1987). Other reported uses
for DEHP include inert ingredient in pesticides, component of dielectric
fluids (replacing PCBs) in electrical capacitors, solvent for erasable
ink, acaricide in orchards, vacuum pump oil, and as a testing agent for
air filtration systems (EPA 1981, IARC 1982, HSDB 1987). Of these uses,
the use of DEHP in dielectric fluids is the only one which is.
commercially important and documented.
Consumer products manufactured from PVC using DEHP as a plasticizer
include vinyl upholstery, tablecloths, shower curtains, raincoats, and
food wrap. Other products using this material are adhesives, resins and
polymeric coatings, components of paper and paperboard, vinyl gloves for
medical examinations, medical tubing, and flexible bags for intravenous
fluids. Until recently, DEHP was used as a plasticizer in the
manufacture of teething rings, pacifiers, and squeeze toys. Most
manufacturers have discontinued the use of DEHP in these products
(USDHHS 1985).
5.5 DISPOSAL
Approximately 800 kkg of DEHP were lost to water during the
production processes in 1977. However, most wastes from DEHP processing
and the manufacture of products containing DEHP are disposed of in
landfills. About 97% of these wastes are disposed of in landfills, and
most of the remaining 3% is incinerated. In 1977, it is estimated that
materials containing 154,200 kkg of DEHP were disposed of in landfills
(EPA 1981).
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6. ENVIRONMENTAL FATE
6.1 OVERVIEW
DEHP enters the environment as a result of releases during
production and use and after disposal. With annual U.S. consumption of
-130 million kg (1986), large quantities of DEHP are available for
potential loss to the environment. Most DEHP is used as a plascicizer LP
flexible PVC products, typically at levels of 1 to 40% by weight
Applications in a large range of products such as household items,
packaging, and medical products ensures widespread distribution of
plastics containing DEHP. Losses of DEHP during manufacture are
considered to be around 0.5%. Most DEHP-containing plastics are disposed
of eventually, with 97% going to landfills and 3% to incineration
Typically, DEHP has been detected in the environment at very low levels
The environmental fate of DEHP is influenced strongly by its
physical and chemical properties. It is a high-boiling liquid (385°C at
760 nun Hg) with a low water solubility of only 50 to 285 ^g/L and a
vapor pressure of 3.4 x 10"^ nun Hg (both at 25"C). The compound is
lipophilic and has a log octanol/water partition coefficient of 4 88
Its biodegradation occurs under aerobic conditions, with a half-life of
several weeks to a month. Under anaerobic conditions, DEHP degrades much
more slowly. Reaction with hydroxyl radical followed by further
atmospheric chemical reactions, along with deposition and rainout,
removes DEHP from the atmosphere. In the aquatic environment, DEHP
accumulates in sediments, on suspended solid materials, and in the lip id
tissues of aquatic biota. In soil, DEHP is strongly held by soil solids
and organic material such as fulvic acid, from which it is not readily
leached.
6.2 RELEASES TO THE ENVIRONMENT
6.2.1 Anthropogenic Sources
Table 6.1 summarizes estimates of releases of DEHP to the
environment (EPA 1980) based on the estimated 1986 U.S. production of
130 million kg.
6.2.2 Natural Sources
Although there have been reports suggesting natural sources of
DEHP, they would be negligible compared to anthropogenic sources (Mathur
1974. as cited in EPA 1980).
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80 Section 6
Table 6.1. Releases of di(2-ethylhexyl)phlhalate to the environment"
Amount released*
Category (millions of kilograms)
U.S. supply
Production waste to water
Transportation loss
Production of products
Vaporized during compounding (loss to air)
Vaporized during compounding (loss to water)
Fabrication waste to landfill
Fabrication waste to incineration
Product inventory
Loss to air
Loss to water
To landfill
To incineration
(130)
0.5
0.1
(129.4)
1.4
2.5
73
0.3
(117.9)
1.8
0.7
112
3.4
"Based on EPA 1980 adjusted to reflect 1986 U.S. supply of DEHP
of 130 million kg.
•*Each figure in parentheses is for a supply/product/inventory
category: other figures give losses and disposal. For example, of a total
U.S. supply of 130 million kg, 0.5 million kg is lost as production waste
to water and O.I million kg is transportation loss, leaving 129.4 million
kg for production of products.
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Environmental Face 81
6.3 ENVIRONMENTAL FATE
6.3.1 Atmospheric Fate Processes
The volatilization of DEHP from plastics and releases during
manufacture and incineration can lead to che presence of chis compound
in the atmosphere Giam et al. (1978) reported combined levels of DEHP
of 0 4 ng/ra^ in the troposphere over the Gulf of Mexico and 2 0 ng/m3
over the North Atlantic. Levels of 300 ng/m3 were reported by Thomas
(1973) near an incinerator in Hamilton, Ontario, Canada. Because of LCS
adsorption properties (log p - 4.88), atmospheric DEHP should have a
strong tendency to adhere to atmospheric particulate matter, especially
organic matter and soot, and to be removed from the atmosphere by
rainout. Chemical hydrolysis of DEHP might occur to a very limited
extent. Direct photolytic reactions are very unlikely. As with most
organic compounds, reaction with the hydroxyl radical 'OH can occur,
such as by abstraction of a hydrogen atom:
(DEHP)-H + -OH - (DEHP)- + H20 .
The free radical product, (DEHP)-, would undergo further reactions
leading to the eventual removal of DEHP from the atmosphere.
6.3.2 Surface Water/Groundwaeer Fate Processes
Reports abound in the scientific literature of the occurrence of
DEHP in surface water and to a lesser extent in groundwater. In the
absence of a direct pollution source, reported levels in water tend to
be of the order of a mlcrogram per liter.
The removal of DEHP from water occurs primarily through uptake by
suspended matter, sediments, and biota. Its biodegradation occurs under
aerobic conditions, with a half-life in the range of weeks to a month
Under anaerobic conditions, the compound appears to be biodegraded much
more slowly.
Because of the very low volatility of DEHP, its volatilization to
the atmosphere from water is likely to be negligible. This is true even
under conditions (such as aeration) conducive to contaminant
evaporation.
Sediments play a major role in determining the fate and perhaps
concentrations of DEHP in surface waters. As noted in EPA (1980), the
sediment/water distribution coefficient for DEHP is of the order of
4 x 103, indicating a very strong tendency of the compound to adhere to
sediments. Reported values of DEHP concentrations in sediments vary
widely. Gian and Atlas (1980), reporting on values obtained from
sediments In the Gulf of Mexico, the Rhine and Ijssel rivers, and Lake
Superior, cited values ranging from about 3 Mg/kg Co around 100 mg/kg of
sediment. The more heavily laden sediments likely provide a steady
source of DEHP contamination to the overlying surface water. The fate of
DEHP will be influenced by several factors. The race of biodegradation
of DEHP in sediments is variable and uncertain, and chemical processes
(e.g., as hydrolysis) leading to the destruction of DEHP in sediments
are probably negligible. If those factors favoring biodegradation are
not present, then DEHP may be transported in the stream sediments.
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82 Section 6
6.3.3 Soil Fate Processes
The phthalate esters, including DEHP, have a strong affinity for
soil solids, including the organic, humic fraction of soil. The
degradation of DEHP in soil has been summarized by Callahan (1979). In
soil under anaerobic conditions, DEHP degrades, but only slowly. Under
aerobic conditions in soil, DEHP has been shown to degrade with a half-
Life of several days. DEHP degrades in soil under aerobic conditions,
but only slowly, if at all, under anaerobic conditions. As expected, the
degradation rate increases with increasing temperature. Studies with
^C-carbonyl-labeled DEHP in aerobic freshwater hydrosoil yielded a
degradation half-life of -14 days. The degradation was found to be
mediated by microorganisms through an initial hydrolysis to the phthalic
acid monoester and 2-ethylhexyl alcohol by the action of esterase
enzymes. The next step in the biodegradation process was found to be
oxidative decarboxylation of the exposed carboxyl group. An intermediate
product in the total degradation process is 1,2-dihydroxybenzene.
Optimal conditions for the degradation of DEHP were found to be within a
range of pH 7 to 9 and at a temperature of -20"C
Both the mineral and organic fractions of solid soil particles tend
to bind DEHP. The high octanol/water partition coefficient of this
compound is indicative of strong binding by soil humus, and van der
Waals-type bonds with soil minerals are favored by the aromatic ring and
carbonyl group on the DEHP molecule.
6.3.4 Biotic Fate Processes
The biotic fate of DEHP has been summarized by Callahan et al.
(1979). Bioaccumulation followed by metabolization and biodegradation by
numerous organisms constitutes the biotic fate of DEHP. A relatively
high log octanol/water partition coefficient is indicative of the high
lipid solubility of DEHP. This has been confirmed by direct observation
of partitioning into the lipids of both plants and animals.
DEHP is biosorbed by both single- and multicellular organisms.
Because it is degraded by microorganisms and metabolized by
invertebrates, fish, and other animals, the tendency for DEHP to undergo
bioraagnification is lessened. However, very rapid bioaccumulation and
concentration factors, ranging from several hundred to several thousand
times the concentration of DEHP in water, have been observed for various
aquatic organisms. The concentration factors appear Co be larger for
smaller aquatic invertebrates (such as Crustacea and midge larvae) than
for fish. Biomagnification of DEHP appears to be much less than that of
organochlorine compounds.
DEHP is less biodegradable than short (i.e., <6 carbons) carbon-
chain-length phthalate esters, but more so than several other long-
carbon-chain phthalates (i.e., >7 carbons), and much more so than
persistent organic compounds such as the chlorinated hydrocarbon
pesticides and PCBs. Several microorganisms, including the saprophytic
bacterium Serratia, Penicillium Lilaanuns, and fnterobaccer aerogenes.
have been shown to degrade DEHP, and Serracia can use the compound as a
sole source of carbon and energy Animals shown to metabolize DEHP
include fish and rats. Rates of metabolism are much less than for
short-chain phthalate esters.
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7. POTENTIAL FOR HUMAN EXPOSURE
7.1 OVERVIEW
DEHP has been detected In air, water, soil, and foods, and in blooc
and plasma handled in tubing and containers that contain DEHP- as a
plasticizer. For the general population, the most likely route of
exposure is through contaminated food, which provides an average of
about 0.3 og/day and a maximum of about 2 mg/day per individual. Because
of its low solubility in water and its low vapor pressure, exposure co
OEHP in either water or air appears to be minimal for most individuals
However, at low levels, DEHP is a ubiquitous contaminant,and this face
combined with the fact that it is an experimental carcinogen in animals
is cause for some concern. The greatest potential for reducing exposure
of the general population lies in preventing contact of food with high
fat or oil content with containers and wrappings containing DEHP
Because of the compound's high lipophilicity, such foods may tend co
extract it, thereby enabling exposure to humans ingesting the food The
FDA currently allows the use of DEHP plasticized containers or wrappings
only for foods that primarily contain water; therefore, properly
packaged foods are unlikely to become contaminated. The FDA ruling.
however, does not preclude the possible misuse of DEHP plasticized
containers or wrappings by the consumer.
A high-risk segment of the population consists of individuals
receiving dialysis treatments or large quantities of blood that has
contacted DEHP-containing tubing or containers. Among this population
are hemophiliacs and dialysis patients.
Workers employed at plants that formulate and process plastics are
also subject to exposure from DEHP. The total pool of such workers
numbers about 600,000 In the United States, although only a small
percentage of these workers will likely be highly exposed.
7.2 LEVELS MONITORED OR ESTIMATED IN THE ENVIRONMENT
7.2.L Levels in Air
Information is limited regarding the levels of DEHP in air. Because
DEHP's vapor pressure is very low, exposure from air is expected co be
minimal. Levels of 0.4 ng/m^ were reported in the air over the Gulf of
Mexico and 2.0 ng/m3 over the North Atlantic by Giam et al. (1978)
Levels of 300 ng/m^ were reported by Thomas (1973) in the atmosphere
near an incinerator in Hamilton. Ontario, Canada. It is generally
accepted that low levels of phthalates occur in the atmosphere
throughout the United States, with higher levels near release sources
-------
84 Seccion 7
The Inhalation Toxicology Research Institute (ITRI 1983) conducted
a vaporization study of eight household consumer products for the
Consumer Product Safety Commission (CPSP) in 1983. The products tested
included window curtain, shower curtain, nursery pad, tablecloth, floor
tile, a segment of vinyl flooring, and two types of wallpaper. A very
low rate of vaporization was reported for each product.
7.2.2 Levels in Water
In general, concentrations of DEHP in freshwaters lie within a
range of a fraction of a Mg/L to 10 Mg/L, although occasionally much
higher values have been observed. Among the levels that have been
reported are 0.1 to 2.19 Mg/L in Japanese waters, 1 to 85 pg/L in U.S
water basins surrounded by heavy concentrations of industry, and 0.02 to
0.5 Mg/L in three German estuaries. In a surveillance study of phthalate
esters in surface waters and sediments in the United States, Michael et
al. (1984) used rigorously controlled sampling techniques, chemical
analysis, and statistical analysis to determine butylbenzylphthalate in
31 U.S. lakes, rivers, and estuaries in the period 1980 to 1982. Their
detection limit was 0.3 ^g/L. Only three sites yielded levels above the
detection limit, and these ranged from 0.3 to 0.9 Mg/L. Because of its
widespread use and lesser tendency to degrade, it may be anticipated
that somewhat higher levels of DEHP could have been found at these
sites. In a study of the determination of phthalate esters in samples
from the marine environment, using combined gas chromatography/mass
spectrometry, Waldock (1983) found 0.058 to 0.078 Mg/L of OEHP in an
estuarine water in the United Kingdom. The values cited above are well
within the solubility limit of DEHP, now generally accepted to be about
50 Mg/L at 20"C (a value difficult to determine because of the tendency
of the compound to form colloidal suspensions and yield erroneously high
values of solubility when determined experimentally). Low aquatic
concentrations of DEHP are consistent with the well known strong
affinity of this compound for suspended particulate matter, biota, and
sediment.
Hazardous waste site leachate that enters either surface water or
groundwater supplies must certainly be considered as a potential source
of DEHP in water. Leachate from such sites often contains significant
quantities of organohalide compounds such as methylene chloride, 1,2-
dichloroethane, and trichloroethene. Much less information is available
regarding the levels of phthalate esters in such leachate, a situation
due in pare to the greater difficulties associated with analysis of
phthalate esters compared to organohalides. In addition, the low water
solubility* of DEHP may impede its appearance in certain leachates A
study by Ghasserai et al. (1984) showed the presence of DEHP in one of 30
leachates at 11 disposal sites at a level of 200 Mg/L. The availability
of a standard method for determining phthalate esters, including DEHP,
in solid wastes and leachates (EPA 1986d) should give additional impetus
to obtaining more data on DEHP levels in hazardous waste site leachates
Because most DEHP-containing plastics are discarded to landfills
after use, monitoring leachate from municipal landfills for DEHP may
yield useful information regarding che exposure potential from such
sources. Prior to the enactment of RCRA. ic was possible that DEHP
-------
PocenCLaL for Human Exposure =;
concaminanc wastes not bound in a polymer matrix were disposed of n
ordinary landfills.
7.2.3 Levels In Soil
A significant fraction of DEHP entering the atmosphere should end
up in soil as a result of deposition and rainout Most of che compound
should remain with the soil because of strong binding by organic soil
huraus and van der Waals-type bonds with soil minerals that are favored
by the aromatic ring and carbonyl group on the DEHP molecule
Furthermore, the aerobic conditions and abundance of saprophytic
microorganisms In soil favor degradation in a soil medium
Because of the factors mentioned above, accurate determination of
DEHP In soil is very difficult, and this is reflected by a dearth of
data on the subject. Despite the large amounts of DEHP deposited in
landfills, little information is available on levels of this chemical
(or other phthalate esters) in soils near landfill sites Soil analyses
have been reported for samples taken In the vicinity of a DEHP factory
and near an industrial dump by Persson et al. (1978). Levels of 0 to 0 5
mg/kg DEHP in soil were reported, although these values were not
corrected for the reported analytical recovery (60% of known analyte
concentration in quality control samples was recovered).
7.2.4 Levels in Food
A potential exists for DEHP contamination of food during
processing, handling, transportation, and packaging. This potential is
especially high for DEHP-plastlclzed wrappings on fatty foods As a
consequence, by 1980. FDA had allowed such wrappings only for foods •-!-_-
a high water content containing minimal fat or oil (EPA 1980). In
addition to packaging materials, a potential source of DEHP in food is
plastic that contacts the food during some stage of processing, for
example, in PVC milk tubing.
A 1974 FDA survey, which included margarine, cheese, meat, cereal.
eggs, milk, white bread, canned corn, corn meal, and baked beans, showed
DEHP levels in most foods of less than 1 mg/kg.
In a recent migration study sponsored by the CMA (1986), it was
shown that the levels of DEHP found in fatty foods such as milk, cheese
and meat were not significantly different from background levels.
7.2.5 Resulting Exposure Levels
DEHP is the phthalate to which humans are most likely to be
exposed. Exposure of the general population to DEHP Is low, with an
average of about 0.3 mg/day and a maximum of about 2 rag/day through
ingestion of phthalate-contarainated food. For the general population.
relatively small amounts of DEHP are taken in from drinking water, che
inhalation of contaminated air. and skin absorption (EPA 1980) Higher
exposures occur to people receiving blood handled in plastics, such as
hemophiliacs and dialysis patients.
-------
86 Section 7
7.3 OCCUPATIONAL EXPOSURES
A high potential exists for the release of DEHP to the workplace
environment from manufacturing operations. This potential is apparent
from an examination of the four basic methods of compounding DEHP
plasticizer with resins. The methods include (1) hot compounding; (2)
dry blending; (3) formation of a stable liquid dispersion of resin in
plasticizer, known as a plasticol. and (4) dissolution of the resin in a
solvent followed by mixing with a plasticizer to make solvent-cast film
and surface-coating materials. As noted in Sect. 6. Table 6.1, about 1 4
million kilograms of DEHP are vaporized to the atmosphere during
plastics manufacturing processes (compounding) each year. An additional
1 8 million kilograms are lost to air from plastic product inventory
each year.
Occupational exposures to DEHP may be relatively high (Liss et al.
1985). Approximately 600,000 U.S. workers are exposed to DEHP in its
manufacture, in plastics formulation, and in manufacturing p.rocesses
that use plastics. In a study of workers in a DEHP formulation plant
(Liss et al. 1985), 6 of 50 personal air monitoring samples showed
levels of DEHP above the analytical limit of detection (10 ng/sample).
These air samples corresponded to time-weighted average concentrations
ranging from 20 to 4,110 /ig/m3, with a mean of 71 /ig/»3- The urine of
workers was also monitored for total phthalate, showing detectable
levels in the more heavily exposed workers and significant increases in
samples taken after each shift as compared to pre-shift samples. It
should be noted that the DEHP levels observed by Liss et al. (1985) were
wichin the OSHA Permissible Exposure Level (PEL) of 5 mg/m3.
Acute toxic responses are unlikely in workers exposed to DEHP. A
publication (Gosselin 1984) on the clinical toxicology of commercial
products assigns to this compound a toxicity rating of I — practically
nontoxic. However, the compound has been shown to cause liver tumors in
Fischer 344 rats and B6C3F1 mice (Menzer and Nelson 1986, NTP 1982);
therefore, it is considered to be an experimental animal carcinogen.
7.4 POPULATIONS AT HIGH RISK
7.4.1 Above-Average Exposure
Exposure of the general population to DEHP tends to be low and is
mainly through food sources. Despite low exposures, the ubiquitous
presence of residues of this substance and the fact that it is an
experimental animal carcinogen are reasons for some concern.
Some segments of the population, such as dialysis patients and
hemophiliacs receiving large quantities of blood, receive appreciably
higher levels of DEHP than does the general population, if their
treatment involves the use of DEHP-plasticized plastics.
The largest segment of the population at risk of DEHP exposure
consists of approximately 600.000 workers engaged in activities such as
plastics formulation. The low volatility of DEHP is helpful in
controlling worker exposure, especially through inhalation.
-------
Potential for Human Exposure *
7.4.2 Above-Avarage Sensitivity
No Information is available on populations with above-average
sensitivity to DEHP. However, results from animal studies indicate chat
an age-dependent susceptibility to testicular and hepatic effects ma/
exist, with immature animals being more susceptible.
-------
8. ANALYTICAL METHODS
Gas chromatography (GC) is Che mosc common analytical method for
detecting and measuring DEHP in environmental and biological samples
High performance liquid chromatography (HPLC) may also be used Co
measure DEHP in biological samples (Pollack et al. 1985a)
Samples may be prepared by several methods, depending on the macr'
being sampled. The sample preparation procedures for analyzing
environmental samples for DEHP include extraction with carbon dlsulf-de
extraction with methylene chloride, soxhlet extraction, and sonicac'on
extraction (EPA 1983. 1986c; NIOSH 1977). Sample preparation for
biological samples often involves extraction with hexane, ethyl acecace
or chloroform/methanol (Sjoberg et al. 1985a. Pollack et al 1985a
Jaeger and Rubin 1972. Hillman et al. 1975, Overturf et al. 1979)
Detectors used to identify DEHP include the flame ionization
detector (FID) and the electron capture detector (ECD) (EPA 1983, 1986c
NIOSH 1977). When unequivocal identification is required, a mass
spectrometer (MS) coupled to the GC column (GC/MS) may be used
(EPA 1983).
Contamination of laboratory apparatus and solvents with DEHP is
very common, since it is a component of many plastic and rubber procuc-s
and is ubiquitous in the environment. Florisil and alumina column
clean-up procedures are recommended to help eliminate interferences, arc
plastics should not be used in phthalate ester analyses (EPA 1983)
8.1 ENVIRONMENTAL MEDIA
Representacive methods appropriate for measuring DEHP in
environmental media are listed in Table 8.1.
8.1.1 Air
The NIOSH method for measurement of DEHP in air involves collector
of vapors in the air onto a cellulose membrane filter, extraction wi-n
carbon disulfide. and injection of an aliquot of the extract into a CC
equipped with an FID. The resulting peaks are measured and compared .it-
standards (NIOSH 1977. as cited in IARC 1982).
8.1.2 Water
The EPA describes two methods (606. 625) for analyzing wastewacer
samples for DEHP. Both of these methods employ packed column GC for
separation of the organic pollutants, however, the methods differ in
sample preparation procedures and detection instrumentation.
-------
90
Section 8
Tabte«.l. Analytical
for
Sample
matrix
Air
Sample preparation
Collection on cellulose
membrane niter.
extraction with carbon
disulfide
Analytical
method"
GC/FI
Sample
detection
limit
1 27-6 82 pom
Accuracy
NAe
Reference*
NIOSH 1977.
cited in (ARC
1982
Water Extraction with methylene
chlonde: exchange to
hexane
Extraction with methylene
chloride
Soil Soxhlet or somcation
extraction: exchange to
hexane
Food Extraction with
acetonitnle and petroleum
ether
Extraction with
acetonitnle. methylene
chlonde. and petroleum
ether
Extraction with hexane.
acetonitnle, and
petroleum ether
EPA method 606- 2.0 Mg/L
GC/EC
EPA method 625- 2 5 Mg/L
GC/MS
EPA method 8060 20 Mg/L to
GC/MS or GC/FI 2 g/L*
GC/EC 0.1 ng
85%
EPA 1983
82% EPA 1983
O.S3C+2.02* EPA I986c
GC/EC
GC/FI
ppb
ISppb
90.7%
70-100%
65-70%
Thuren 1986
Giametal 197$
Williamt 1973
"GC. gai chromatography; Fl. name lonuatnn; EC. electron capture; MS, man ipectromctry.
* Average percent recovery, unlest otherwue indicated.
' Not available.
rf Depending on the matrix.
'Here, C - concentration in Mg/L, in an equation directly relating accuracy to concentration.
-------
Analytical Mechods 9
Measuring method 606 is optimized for phthalate esters with a
detection limit of 2.0 pg/L for DEHP Method 625. which employs MS for
detection and quantification, is more broadly applicable to a large
number of base/neutral and acid extractable organics Sample detection
limits and accuracy for DEHP are similar for both methods
8.1.3 Soil
Soil samples can be analyzed for DEHP by GC with an ECD or FID
Sample preparation is by soxhlet or sonication extraction, with an
exchange to hexane. This method (8060) is used by the EPA for analyzing
solid waste for phthalate esters. The method detection limit for DEHP
ranges from 20 to 2 x 106 ng/L, depending on the matrix (EPA 1986c)
8.1.4 Food
The most common method for analyzing foods for DEHP is by GC/ECD
(Thuren 1986. Giam et al. 1975). Analysis may also be by GC/FID
(Williams 1973, Ishida et al. 1981) or gel filtration with GC or HPLC
(Baker 1978).
8.2 BIOMEDICAL SAMPLES
Methods for the analysis of DEHP in biological samples are listed
in Table 8.2.
8.2.1 Fluids and Exudates
Blood levels of DEHP are measured using HPLC with ultraviolet
absorbance monitoring at 254 run. The samples are prepared by extraction
with ethyl acetate (Pollack et al. 1985a). Plasma levels of DEHP may be
measured with GC/MS after extraction with hexane (Sjoberg et al. 19S5a)
Analysis of urine for DEHP is by GC/MS (Liss et al. 1985)
8.2.2 Tissues
Human tissues, including lung, liver, spleen, kidney, heart, and
adipose, have been analyzed for DEHP by GC methods (Jaeger and Rubin
1972, Hillman et al. 1975, Overturf ec al. 1979).
-------
92 Section 8
Table 8.2. Analytical
for dM2-ethylh*xyl)pfctBaltt* !• Matofkal
Sample
matrix
Unne
Plasma
Whole blood
Lung, liver.
spleen, adipose
Heart
Kidney
Sample preparation
NR*
Extraction with hexane
Extraction with ethyl
acetate
Extraction with
chloroform / methanol
Extraction with
chloroform/methanol
Extraction with
chloroform/methanol
Analytical
method0
GC/MS
GC/MS
HPLC/UV
GC
GC/MS
GC/MS
Sample
detection
limit
NR nmol/mL
3 8 Mg/mL
0 345 Mg/mL
5 0 Mg/gf
dry weight
002Mg/g
0 1 ng/mgc
Accuracy
NR
NR
NR
60-90%
NR
NR
Rcferenca
List et aL 1985
Sjoberg et aL I985»
Pollack et al. 198Sa
Jaeger and Rubin 1972
Rubin 1972
Hillmuetal I97S
Overturf et al. 1979
"GC. gas chromatography: MS. mass spectrometry. HPLC. high-performance liquid chromatography. UV,
ultraviolet spectroscopy
*NR - not reported.
r Lowest concentration reported.
-------
9. REGULATORY AND ADVISORY STATUS
9.1 INTERNATIONAL
The World Health Organization (WHO) has not recommended a drin.ki.rg
water guideline value for DEHP
9.2 NATIONAL
9.2.1 Regulations
Regulations applicable to DEHP address occupational exposure
concentrations, spill quantities, presence in hazardous wastes and
reporting rules (Table 9.1).
OSHA sets Permissible Exposure Limits (PELs) for occupational
exposures to chemicals based on the recommendations of the National
Institute for Occupational Safety and Health (NIOSH). The OSHA PEL for
DEHP is 5 mg/m-} in workplace air for a time-weighted average (TWA) (3
h/day. 40 h/week).
CERCLA requires that persons in charge of facilities from which a
hazardous substance has been released in quantities equal to or greater
than its reportable quantity (RQ) immediately notify the National
Response Center of the release. The statutory RQ for DEHP set by che E?-
Office of Emergency and Remedial Response (OERR) is 1 Ib. However the
EPA has proposed revising the RQ to 100 Ib, based on the assignment of
DEHP to Category B in the hazard-ranking, weight-of-evidence groups for
potential carcinogens.
Chemicals are included on the RCRA Appendix VIII list of hazardous
constituents (40 CFR Part 261) if they have toxic, carcinogenic,
mutagenic, or teratogenic effects on humans or other life forms. DEHP is
included on this list. Vastes containing DEHP are subject to the RCRA
regulations promulgated by the EPA Office of Solid Waste (OSW).
The Office of Toxic Substances (OTS) promulgates regulations
related to manufacturers and/or processors of chemicals which may
present an unreasonable risk to health or the environment. Manufacturers
of DEHP are required to submit to EPA information on the quantity of :he
chemical manufactured or imported, the amount directed to certain uses.
and the potential exposure and environmental release of the chemical
under the Preliminary Assessment Information Rule. Unpublished health
and safety studies on DEHP must be submitted to EPA by manufacturers or
processors under the Health and Safety Data Reporting Rule.
-------
Seccion 9
Tabfc9.l. Regolaitoas aad gvidetiM applicable
to tfM2-etkylk«iyl)Dejihalau
Agency
Description
Value
References
OSHA
EPA OERR
EPAOSW
EPAOTS
Nadosal regulation*
Permissible exposure limit (PEL)
Time-weighted average (TWA) 5 mg/m
Reponable quantity (RQ)
Statutory I l»
Proposed 100 Ib
Luting as a Hazardous Waste NAa
Constituent (Appendix VIII)
Preliminary Assessment Information NA
Rule
Health and Safety Data Reporting NA
Rule
29 CFR 1910
(1971)
40 CFR 302.4
50 FR 13456
(04/04/85)
52 FR 8140
(03/16/87)
40 CFR 261
45 FR 33084
(05/19/80)
40 CFR 712
47 FR 26992
(06/22/82)
40 CFR 716
47 FR 38780
(09/02/82)
NIOSH Recommended exposure limit (RED Lowest NIOSH 1985
for occupational exposure feasible
limit
ACGIH Threshold limit value (TLV) ACGIH 1986
TWA 3 mg/m*
Short-term exposure limit (STED 10 mg/m1
NAS Chronic suggested no adverse 4.2 mg/L NAS 1977
response level (SNARL)
EPA OWRS Ambient water quality cntena to 45 FR 79318
protect human health (11/28/80)
Ingesting water and organisms 15 mg/L
Ingesting organisms only 50 mg/L
EPA OWRS Draft water quality cntena EPA I987e
1-day HA for 10-kg child' 20 mg/L
1-day HA for 70-kg adult 70 mg/L
10-day HA for 10-kg child S mg/L
10-day HA for 70-kg adult 17 S mg/L
Lifetime DWEL for 70-kg adult' I 33 mg/day
IARC Carcinogenic rank Group 2B IARC 1982
EPA Carcinogenic rank Group B2 EPA I986b
State
environmental
agencies
Scat* rtfmtadoei
Water quality standards for
some states
s
3 mg/L Environment
Reporter
"NA - not available
0
HA - health advisory
f DWEL - drinking water equivalent level
-------
Regulatory and Advisory Scacus '•:
922 Advisory Guidance
Advisory guidance levels are environmental concentrations
recommended by either regulatory agencies or other organizations which
are protective of human health or aquatic life Although not
enforceable, these levels may be used as the basis for enforceable
standards. Advisory guidance for DEHP is summarized in Table 9 1 and
includes the following a NIOSH Recommended Exposure Limit (REL) for
occupational exposure, a threshold limit value (TLV). ambient water
quality criteria, a chronic Suggested No Adverse Response Level (SVARL.
calculated by the National Academy of Sciences (NAS); and carclnoaen-c
ranking by EPA and IARC
The NIOSH REL for occupational exposure to DEHP in air is the
lowest feasible limit. The NIOSH does noc specify an Immediately
Dangerous to Life or Health (IDLH) value, but recommends that DEHP be
treated as a potential human carcinogen.
The American Conference of Governmental Industrial Hygienists
(ACGIH) recommends a TLV TWA of 5 mg/m3 and a Short-Term Exposure Limic
(STEL) of 10 mg/ra-5 per 15 rain per 8 h based on the toxicity of DEHP
The ambient water quality criteria are guidelines set by the EPA
Office of Water Regulations and Standards (OWRS) to protect human healch
from potential adverse effects from the ingestion of water and/or
organisms, or from ingestion of organisms only from surface water
sources. The values for DEHP are 15 mg/L for ingesting water and
organisms and 50 mg/L for ingesting only organisms.
The NAS calculated a chronic SNARL for DEHP of 4 2 mg/L based on
chronic toxicity data. The calculation was based on an Acceptable Dail
Intake (ADI) of 0 6 mg/kg/day, assuming 20% of total intake is from
water, as follows:
0.6 mg/kg/day x 70 kg x 0.2
4.2 mg/L .
2 L/day
9.2.3 Data Analysis
9.2.3.1 Reference dose
A reference dose (RfD) has been calculated for DEHP (EPA 1987) It
was based on a 1-year study in guinea pigs by Carpenter et al. (1953)
In this study, groups of 23 to 24 animals of each sex were fed diets
containing 0, 400, or 1,300 ppm DEHP. This is equivalent to doses of -0
19. and 64 mg/kg/day. Statistically significant Increases in relative
liver weights were observed in both groups of treated animals. An RfD
was determined as follows:
(19 mg/kg/day)
2 x ID'2 mg/kg/day .
1.000
-------
96 Seccion 9
where
19 mg/kg/day - NOAEL
1,000 - uncertainty factor
9 2.3.2 Carcinogenic potency
Data from the NTP (1982) bioassay (Sect. 4.3 6) were used by EPA
(1987c) to calculate the upper-bound incremental unit carcinogenic risk
to humans. Using the linearized multistage model, potency slope factors
were estimated. These are shown below.
Animals on which based
Slope factor3
Species Sex [(mg/kg/day)'1]
Rat
Mouse
Male
Female
Male
Female
0.0045
0.0035
0.0084
0.01
^Calculations were based on combined carcinomas
and neoplastic nodules (or adenomas).
The unit risk value is estimated to be 4.0 x 10"7 for drinking
water containing 1 Atg/L DEHP The drinking water concentrations of DEH?
corresponding to lifetime excess cancer risks of 10'4, 10'5, and 10'6
are 0.3, 0 03, and 0.003 mg/L, respectively.
In the NTP (1982) bioassay, an adequate number of animals were
observed, and a statistically significant increase in incidence of liver
tumors was seen in both sexes and was dose dependent in both sexes of
mice and in female rats.
The CPSC (1985) estimated the lifetime excess risk per mg/kg/day
for hepatocellular carcinomas and neoplastic nodules observed in the NTP
(1982) study. The estimates of risk to humans using the linearized
multistage model with body-surface-area correction factors of 5.6 for
rats and 12.8 for mice are as follows:
Lifetime human risk estimates
[(mg/kg/day)"I]
Animal on which based
Species
Rat
Mouse
Sex
Male
Female
Male
Female
Maximum
likelihood
estimate
0 000862
0 00160
0 00868
0 00698
Upper 95%
confidence
limit
0 00264
0.00293
0.0144
0.00963
-------
Regulacory and Advisor:-
The differences between the potency slope factors calculated b/ £?-
(1987c) and CPSC's (1985) upper 95% confidence limit values are due :o
differences in doses used in the two agencies' calculations
9.3 STATE
Regulations and advisory guidance from the states were still be:-g
compiled at the time of printing
9.3.1 Regulations
Phthalate esters are specified in a few State Water Quality
Standards; the standard for public water systems is 3 Mg/L. .'-."hen not
specified, DEHP is included in the category "toxic substances" in
narrative form in the Water Quality Standards of most states. Specific
narrative standards protect the use of surface waters for public -acer
supply and contact recreation.
9.3.2 Advisory Guidance
No data were available on state advisory guidance
-------
10. REFERENCES
ACGIH (American Conference of Government Industrial Hygienists). 1986
Documentation of the Threshold Limit Values and Biological Exposure
Indices. 5th ed. Cincinnati, OH.
Agarwal DK, Eustis S. Lamb JC. Reel JR. Kluwe WM 1986. Effects of di-
(2-echylhexyDphchalace on the gonadal pathophysiology, sperm
morphology, and reproductive performance of male rats. Environ Healch
Perspect 65:343-350.
Agarval DK, Agarwal S, Seth PK. 1982. Effect of DEHP on drug metabolism.
lipid peroxidation. and sulfhydryl content of rat liver. Drug Metab
Dispos 10:77-80.
* Albro PW. 1986. Absorption, metabolism, and excretion of DEHP by ra-s
and mice. Environ Health Perspect 65:293-298.
Albro PV, Corbett JT, Schroeder JL, Jordan S, Matthews HB. 1982.
Pharmacokinetics, interactions with macromolecules, and species
differences in metabolism of DEHP. Environ Health Perspect 45:19-25
Albro PW, Tondeur I, Marbury D, Jordan S, Schroeder J, Corbect JT.
1983a. Polar metabolites of DEHP in the rat. Biochim. Biophys. Acca
760: 283-292.
Albro PW, Hass JR, Peck CC, Jordan ST. Corbett JT. Schroeder J. 1983b
Applications of isotope differentiation for metabolic studies with DEK?
J Environ Scl 17:701-714.
Albro PU, Hass JR, Peck CC, et al. 1981. Identification of the
metabolites of DEHP In urine from the African green monkey. Drug Metab
Dispos 9:223-225.
Albro PV, Thomas R, Fishbein L. 1973. Metabolism of diethylhexyl
phthalaCe by rats: Isolation and characterization of the urinary
metabolites. J Chromatogr 76-321-330.
Albro PW, Tondeur I, Marbury D. Jordan S, Schroeder J, Corbett JT. 198^
Polar metabolites of DEHP in che rat. Biochim Biophys Acta 760:283-292
Aldrich. 1987. Aldrich Chemical Catalog. Aldrich Chemical Company.
*Key studies.
-------
100 Seccion 10
APHA (American Public Health Association) 1985 Standard Methods for
the Examination of Water and Wastewater 16th ed Washington, DC.
Ashby 1986. The prospects for a simplified and internationally
harmonized approach to the detection of possible human carcinogens and
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LL5
11. GLOSSARY
Acute Exposure--Exposure to a chemical for a duration of 14 days or
less, as specified in Che Toxicological Profiles.
Bioconcentration Factor (BCF)--The quotient of the concentration of a
chemical in aquatic organisms at a specific time or during a discrete
time period of exposure divided by the concentration in the surrounding
water at the same time or during the same time period.
Carcinogen--A chemical capable of inducing cancer.
Ceiling value (CL)--A concentration of a substance that should not be
exceeded, even instantaneously.
Chronic Exposure--Exposure to a chemical for 365 days or more, as
specified in the Toxicological Profiles.
Developmental Toxicity--The occurrence of adverse effects on the
developing organism that may result from exposure to a chemical prior to
conception (either parent), during prenatal development, or postnatally
to the time of sexual maturation. Adverse developmental effects may be
detected at any point in the life span of the organism.
Embryotoxicity and Fetotoxicity--Any toxic effect on the conceptus as a
result of prenatal exposure to a chemical; the distinguishing feature
between the two terms is the stage of development during which the
insult occurred. The terms, as used here, include malformations and
variations, altered growth, and In utero death.
Frank Effect Laval (FED--That level of exposure which produces a
statistically or biologically significant increase in frequency or
severity of unmistakable adverse effects, such as irreversible
functional impairment or mortality, in an exposed population when
compared with its appropriate control.
EPA Health Advisory—An estimate of acceptable drinking water levels for
a chemical substance based on health effects information. A health
advisory is not a legally enforceable federal standard, but serves as
technical guidance to assist federal, state, and local officials.
Immediately Dangerous to Life or Health (IDLH)--The maximum
environmental concentration of a contaminant from which one could escape
within 30 min without any escape-impairing symptoms or irreversible
health effects.
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116 Section 21
Intermediate Exposure--Exposure to a chemical for a duration of 15-364
days, as specified in the Toxicological Profiles
ImmunoLoglc Toxlcity--The occurrence of adverse effects on the immune
system that may result from exposure to environmental agents such as
chemicals
In vitro--Isolated from the living organism and artificially maintained.
as in a test tube
In vivo--Occurring within the living organism.
Key Study--An animal or human toxicological study that best illustrates
the nature of the adverse effects produced and the doses associated with
those effects.
Lethal Concentration(LO) (LCLO)--The lowest concentration of a chemical
in air which has been reported to have caused death in humans or
animals
Lethal Concentration(SO) (LCSO)--A calculated concentration of a
chemical in air to which exposure for a specific length of time is
expected to cause death in 50% of a defined experimental animal
population.
Lethal Dose(LO) (LDLO)--The lowest dose of a chemical introduced by a
route other than inhalation that is expected to have caused death in
humans or animals
Lethal Dose(50) (LDSO)--The dose of a chemical which has been calculated
to cause death in 50% of a defined experimental animal population.
Lowest-Observed-Adverse-Effect Level (LOAEL)--The lowest dose of
chemical in a study or group of studies which produces statistically or
biologically significant increases in frequency or severity of adverse
effects between the exposed population and its appropriate control.
Lowest-Observed-Effect Level (LOEL)--The lowest dose of chemical in a
study or group of studies which produces statistically or biologically
significant increases in frequency or severity of effects between the
exposed population and its appropriate control.
Malformations--Permanent structural changes that may adversely affect
survival, development, or function.
Minimal Risk Level — An estimate of daily human exposure to a chemical
that is likely to be without an appreciable risk of deleterious effects
(noncancerous) over a specified duration of exposure.
Mutagen--A substance that causes mutations. A mutation is a change in
the genetic material in a body cell. Mutations can lead to birth
defects, miscarriages, or cancer.
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Glossary 117
Neurotoxicity--The occurrence of adverse effects on the nervous system
following exposure to a chemical.
No-Observed-Adverse-Effect Level (NOAEL)--That dose of chemical at which
there are no statistically or biologically significant increases in
frequency or severity of adverse effects seen between the exposed
population and its appropriate control Effects may be produced at this
dose, but they are not considered to be adverse.
No-Observed-Effect Level (NOEL) --That dose of chemical at which there
are no statistically or biologically significant increases in frequency
or severity of effects seen between the exposed population and its
appropriate control.
Permissible Exposure Limit (PEL) --An allowable exposure level in
workplace air averaged over an 8-h shift.
q, --The upper-bound estimate of the low-dose slope of the dose- response
curve as determined by the multistage procedure. The q * can be used co
calculate an estimate of carcinogenic potency, the incremental excess-
cancer risk per unit of exposure (usually /*g/L for water, mg/kg/day for
food, and Mg/m^ for air).
Reference Dose (RfD)--An estimate (with uncertainty spanning perhaps an
order of magnitude) of the daily exposure of the human population to a
potential hazard that is likely to be without risk of deleterious
effects during a lifetime. The RfD is operationally derived from the
NOAEL (from animal and human studies) by a consistent application of
uncertainty factors that reflect various types of data used to estimate
RfDs and an additional modifying factor, which is based on a
professional judgment of the entire database on the chemical. The RfDs
are not applicable to nonthreshold effects such as cancer.
Reportable Quantity (RQ)--The quantity of a hazardous substance that is
considered reportable under CERCLA. Reportable quantities are: (1) 1 Ib
or greater or (2) for selected substances, an amount established by
regulation either under CERCLA or under Sect. 311 of the Clean Water
Ace. Quantities are measured over a 24 -h period.
Reproductive Toxic ity- -The occurrence of adverse effects on the
reproductive system that may result from exposure to a chemical. The
toxicity may be directed to the reproductive organs and/or the related
endocrine system. The manifestation of such toxicity may be noted as
alterations in sexual behavior, fertility, pregnancy outcomes, or
modifications in other functions that are dependent on the integrity of
this system.
Short-Term Exposure Limit (STEL)--The maximum concentration to which
workers can be exposed for up to 15 min continually. No more than four
excursions are allowed per day, and there must be at least 60 min
between exposure periods. The daily TLV-TWA may not be exceeded.
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118 Section 11
Target Organ Toxlcity--This term covers a broad range of adverse effects
on target organs or physiological systems (e.g., renal, cardiovascular)
extending from those arising'through a single limited exposure to those
assumed over a lifetime of exposure to a chemical.
Teratogen--A chemical that causes structural defects that affect che
development of an organism.
Threshold Limit Value (TLV)--A concentration of a substance to which
most workers can be exposed without adverse effect. The TLV may be
expressed as a TWA, as a STEL, or as a CL.
Time-weighted Average (TWA)--An allowable exposure concentration
averaged over a normal 8-h workday or 40-h workweek.
Uncertainty Factor (UF)--A factor used in operationally deriving the RfD
from experimental data. UFs are Intended to account for (1) the
variation in sensitivity among the members of the human population,
(2) the uncertainty in extrapolating animal data to the case of humans,
(3) the uncertainty in extrapolating from data obtained in a study that
is of less than lifetime exposure, and (4) the.uncertainty in using
LOAEL data rather than NOAEL data. Usually each of these factors is set
equal to 10.
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119
APPENDIX: PEER REVIEW
A peer review panel was assembled for DEHP. The panel consisted of
the following members: Dr. W. H. Lawrence, University of Tennessee.
Dr. G. Michalopoulos, Duke University; and Dr. G. M. Pollack. University
of North Carolina. These experts collectively have knowledge of DEHP's
physical and chemical properties, toxicokinetics. key health end poincs.
mechanisms of action, human and animal exposure, and quantification of
risk to humans. All reviewers were selected in conformity with the
conditions for peer review specified in the Superfund Amendments and
Reauthorization Ace of 1986, Section 110.
A joint panel of scientists from ATSDR and EPA has reviewed the
peer reviewers' comments and determined which comments will be included
in the profile. A listing of the peer reviewers' comments not
incorporated in the profile, with a brief explanation of the rationale
for their exclusion, exists as part of the administrative record for
this compound. A lisC of databases reviewed and a list of unpublished
documents cited are also included in the administrative record.
The citation of the peer review panel should not be understood co
imply their approval of the profile's final content. The responsibility
for the content of this profile lies with the Agency for Toxic
Substances and Disease Registry.
*US COVaNMOtrnUNTlNCOmCI.! tl« -13 ••>•<•«/ UCIONNO4
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