DIETHYLPHTHALATE/media/Files/Report Files/2007/Long-Term-Health-Effects...rocket propellants and...

Contract No. IOM-2794-04-001

The National Academies

HEALTH EFFECTS OF

PROJECT SHAD

CHEMICAL AGENT:

DIETHYLPHTHALATE

[CAS # 84-66-2]

Prepared for the National Academies

by

The Center for Research Information, Inc. 9300 Brookville Rd

Silver Spring, MD 20910

http:// www.medresearchnow.com

(301) 346-6501

[email protected]

2004


Health Effects of Diethylphthalate

ii

ACKNOWLEDGEMENTS

Submitted to Dr. William Page, Program Officer, Advisory Panel for

the Study of Long-term Health Effects of Participation in Project SHAD

(Shipboard Hazard and Defense), Institute of Medicine, the National

Academies.

This report is subject to the copyright and reproduction arrangements defined in

Contract No. IOM-2794-04-001 of the National Academies.

This report and any supplements were prepared by the Center for Research

Information, Inc. which is solely responsible for its contents.

Although this draft is the definitive submission on its subject matter, the Center for

Research Information recognizes its ethical and contractual obligation to update,

revise, or otherwise supplement this report if new or necessary information on its

subject matter should arise, be requested, or be ascertained during the contract

period.

The Principal Investigator wishes to acknowledge and thank Matthew Hogan, Linda

Roberts, Lawrence Callahan, Judith Lelchook and Emnet Tilahun for research

assistance, editorial content assistance, and project input.

Principal Investigator: Victor Miller

Text Draft & Editing: Victor Miller & Matthew Hogan

Project Manager: Matthew Hogan

Administration: Linda Roberts



iii

SPECIAL NOTE ON PSYCHOGENIC SEQUELAE OF PERCEIVED

EXPOSURE TO BIOCHEMICAL WARFARE AGENTS

This report deals primarily with the biological health challenges engendered by the agent

that is the subject of the report. Nevertheless, this report also incorporates, by reference

and attachment, a supplement entitled "Psychogenic Effects of Perceived Exposure to

Biochemical Warfare Agents".

The supplement addresses and describes a growing body of health effects research and

interest centered upon the psychogenic sequelae of the stress experienced personally from

actual or perceived exposure to chemical and biological weaponry. Because awareness

of exposure to agents in Project SHAD logically includes the exposed person also

possessing a perception of exposure to biochemical warfare agents, the psychogenic

health consequences of perceived exposure may be regarded as additional health effects

arising from the exposure to Project SHAD agents. This reasoning may also apply to

simulants and tracers. Therefore, a general supplement has been created and submitted

under this contract to address possible psychogenic effects of perceived exposure to

biological and chemical weaponry.

Because such health effects are part of a recent and growing public concern, it is expected

that the supplement may be revised and expanded over the course of this contract to

reflect the actively evolving literature and interest in the issue.



iv

TABLE OF CONTENTS

I. EXECUTIVE SUMMARY……………………..… 1

II. BACKGROUND DATA……………………….… 3 Identification & Physical Chemistry…………………………………… 4

Manufacture & Use……………………………………………………… 4

Kinetics…………………………………………………………………… 4

III. HEALTH EFFECTS/TOXICITY…………………… 6

Overview…………………………………………………………………. 6

Acute/Subchronic/Chronic Concerns………………………………….. 7

Reproductive Toxicity…………………………………………………… 8

Carcinogenicity……………………………………………………….…. 9

Genotoxicity……………………………………………………………… 10

IV. PSYCHOGENIC EFFECTS……………….……… 11

V. TREATMENT & PREVENTION…………………….. 12

VI. SECONDARY SOURCE COMMENT………… 13

VII. BIBLIOGRAPHY WITH ABSTRACTS…….. 14



1

I. EXECUTIVE SUMMARY

Diethylphthalate (more commonly rendered in the scientific literature as two words

“diethyl phthalate”) is a phthalic acid ester with the chemical formula C12H14O4 , and

commonly identified by Chemical Abstracts Service (CAS) registry number 84-66-2. It

ordinarily appears as a bitter-tasting colorless or water-white liquid with no odor, or a

slight aromatic odor. It is slightly soluble in water, while also soluble in alcohol, ether,

benzene, and acetone. Diethylphthalate is miscible with vegetable oils, esters, and

aromatic hydrocarbons. It is manufactured by refluxing one equivalent of phthalic

anhydride with a greater than two-fold excess of ethanol in the presence of one percent of

concentrated sulfuric acid. It is also classed as a phthalic anhydride ester (PAE).

Diethylphthalate is a widely encountered compound in daily life. Automobile parts,

toothbrushes, tools, and food packaging are ordinary products in which one can

frequently find diethylphthalate. Aspirin, insecticides, and cosmetics can also contain it.

The most common industrial use for diethylphthalate is as a “plasticizer” -- an agent for

making plastics more flexible. In Project SHAD, diethylphthalate was used as a simulant

for VX Nerve Agent. Because of its common use in so many household and personal

consumer products, exposure through many pathways (oral, dermal, respiratory) has been

studied.

The Threshold Limit Value for diethylphthalate of the American Conference of

Governmental Industrial Hygienists (ACGIH) is 5.0mg/m3 based on an 8-hour workday

time-weighted average. The pharmacology and kinetics of diethylphthalate exposure

indicate slow absorption by the skin, the metabolic conversion of absorbed

diethyphthalate into ethanol and the monoester monoethyl phthalate, followed by rapid

excretion, mostly in the urine.

The effects of diethylphthalate are fairly extensively studied. The chemical shares with

other phthalates the characteristic of being among the least toxic of substances in

industrial use. In vivo human studies or case reports of serious direct physiological

insult as a result of diethylphthalate exposure are not to be found, with the exception of

mucous membrane/pulmonary irritation, or a general anesthetic effect at very high

concentrations/doses, along with unusual sensitive skin reactions in exceptional

sensitized individual cases. An in vitro study on a human skin model did produce a

strong cytotoxic reaction but this has not been duplicated in vivo.

Animal studies provide powerful corroboration of diethylphthalate’s low toxicity. Only

very high acute oral doses have produced lethality in animals. Otherwise, non-toxic

systemic effects usually seen in animal testing are decreased weight gain with alterations

in liver and kidney size, likely attributable to hypertrophy. Animal studies indicate that

diethylphthalate is only mildly or moderately irritating when applied to the skin or the

eye.



2

Evidence of carinogenicity is at best equivocal. In rodent studies a carcinoma/adenoma

positive dose-response versus control results was found in only one sex of one species,

and the response did not differ significantly from a historical mean for the species and

gender. Evidence of genotoxicity is also weak, with only in vitro sister-chromatid

exchanges (SCE) a confirmed effect, but these occurred only in the presence of an S9

fraction from a sensitive species in which a correlation between SCEs and

carcinogenicity is regarded as tenuous. Both the Environmental Protection Agency (EPA)

and ACGIH regard diethylphthalate to be a substance without evidence of cancer risk

[EPA class D; ACGIH class A4]; human case reports or epidemiological study of

carcinogenesis from diethylphthalate have not been found.

Some concern may exist for toxicity in the reproductive/developmental area. Skeletal

abnormalities in rodent offspring have been seen after maternal administration of high

doses. Chicken embryos die at a faster rate after direct injection of diethylphthalate. A

lowering of testosterone levels in rodents has been seen following diethylphthalate

exposure, though no fertility or testicular damage was seen. A lowering of human sperm

motility was observed after direct in vitro administration of diethylphthalate. Concerns

have been raised on risks to pregnant human females and offspring in light of the

detected presence of significant amounts of diethylphthalate in the blood of pregnant

women in urban areas.

One comprehensive and relatively recent (2001) review of diethylphthalate toxicity

concludes that there are ultimately “no toxic endpoints of concern” for the substance in

regard to acute toxicity, eye irritation, dermal irritation, dermal sensitization,

phototoxicity, photoallergenicity, percutaneous absorption, subchronic toxicity,

teratogenicity, reproductive toxicity, genetic toxicity, chronic toxicity, carcinogenicity,

and potential human exposure.

Psychogenic effects specifically of diethylphthalate exposure have not been found in the

literature, but the general effects of a perceived exposure to chemical warfare agents are

treated in the supplement provided under this contract entitled “Psychogenic Effects of

Perceived Exposure to Biochemical Warfare Agents.”

Secondary literature tends to be comprehensive. It appears that the similarity in names

and characteristics of the PAE class may cause confusion in reportage of effects,

however.



3

II. BACKGROUND DATA

Identification & Physical Chemistry

Project SHAD Chemical Agent Name: Diethylphthalate.

CAS#: 84-66-2

More Commonly Appearing Name: Diethyl Phthalate

Abbreviation: DEP (Common Use), D (Project SHAD)

Alternate Names: anozol; 1,2-benzenedicarboxylic acid diethyl ester; o-

benzenedicarboxylic acid diethyl ester; carboxylic acid, diethyl ester; diethyl ester

phthalic acid; diethyl o-phthalate; diethyl-o-phenylenediacetate; DPX-F5384; estol 1550;

ethyl phthalate; NCI-C60048; neantine; palatinol; phthalol; phthalsaeurediaethylester;

placidol E (HSDB 2004, RTECS 2004)

Chemical Formula: C12H14O4

Chemical Structure (CHEMIDplus 2004):

Molecular Weight: 222.23

Specific gravity: 1.232 (14 oC)

Vapor Pressure: 14mm Hg (163 oC)

Conversion rate: 9.07 mg/m3 = 1 ppm

Boiling Point: 298 oC

Melting Point: -40.5 oC

Sources: HSDB 2004, RTECS 2004; Chem ID/TOXNET 2004.

Diethylphthalate is classified among the phthalic anhydride esters (PAEs) (Kamrin 1991).



4

Diethylphthalate is produced by refluxing one equivalent of phthalic anhydride with a

greater than two-fold excess of ethanol in the presence of one percent of concentrated

sulfuric acid (Fed. Reg. 60(171): 46076-9 (September 5, 1995) 46076-46079).

Diethylphthalate is soluble in alcohol, ether, acetone, benzene. It is miscible with

vegetable oils, ketones, esters, and aromatic hydrocarbons; it is partly miscible with

aliphatic solvents. Diethylphthalate is soluble in water at a rate of 1000 mg/l (25 oC).

Diethylphthalate in pure form is usually encountered as a stable, usually colorless (but

sometimes white), oily liquid with a bitter taste (HSDB 2004, RTECS 2004).

Manufacture and Use

Eastman Kodak Company and Unitex Chemical Company are the major American

manufacturers of diethylphthalate (HSDB 2004).

During Project SHAD, diethylphthalate was used as a simulant for VX Nerve Agent

(Project 65-17 2004).

Diethylphthalate is a widely used chemical in industrial and consumer products. It chief

use is as a “plasticizer” -- an agent for making plastics more flexible. Automobile parts,

toothbrushes, tools, and food packaging are ordinary products in which one can often find

diethylphthalate. Aspirin and insecticides may also contain it (HSDB 2004).

Because of diethylphthalate’s common use in so many household and personal consumer

products, human exposure is likely to occur through many pathways (oral, dermal,

respiratory). Cosmetic products using diethylphthalate include bath preparations, eye

shadows, hair sprays, wave sets, nail polish, nail polish remover, nail extenders,

detergents, aftershave lotions, and skin care preparations (HSDB 2004; Api 2001).

Diethylphthalate is also used to manufacture celluloid. It has been used as a solvent for

cellulose acetate in varnishes; as a fixative for perfumes; as a wetting agent; as a camphor

substitute (HSDB 2004; Api 2001).

Diethylphthalate has served also as a diluent in polysulfide dental impression materials;

and as a solvent for nitrocellulose and cellulose acetate. It is used as a plasticizer in solid

rocket propellants and cellulose ester plastics such as photographic films and sheets,

blister packaging, and tape applications (HSDB 2004; Api 2001).

Kinetics

Diethylphthalate can be absorbed through the skin, the lungs, and the digestive tract (Api

2001).

Dermal absorption has been extensively studied. In vitro testing by the Research Institute

on Fragrance Materials has lead to establishing a human skin steady-state abortion rate of



5

1.27 + 0.11 mg/cm2/hr (Api 2001). Dermal absorption by human skin is about 30 times

slower than that of experimental in vitro tissue (Api 2001).

Data on inhalation absorption cannot be found.

After absorption, diethylphthalate tends to accumulate most in the kidneys and liver, with

blood, spleen, and fat cells. Diethylphthalate is usually rapidly metabolized into its

monoester, monoethyl phthalate, and ethyl alcohol (ethanol) (Kamrin 1991). The

hydrolysis enzymes of diethylphthalate are not well-characterized (Api 2001).

Excretion occurs primarily through the urine (WHO 2003; Api 2001).



6

III. HEALTH EFFECTS/TOXICITY

Overview

The toxicology of diethylphthalate has been the subject of extensive study. With other

phthalates, it shares the characteristic of being among the least toxic of substances in

industrial use. As the discussion in this section will indicate, human or clinical studies

reporting serious direct physiological insult as a result of diethylphthalate exposure show

only rare, transient, and mostly minor occurrences in high dose situations or with

sensitive individuals. These include irritation from heated diethylphthalate, central

nervous system depression after heavy exposure, along with rare dermal sensitivity in

individuals with a pre-disposition for dermal sensitization

A recent (2001) comprehensive review of diethylphthalate toxicity concludes that there

are “no toxic endpoints of concern” in the following areas: acute toxicity, eye irritation,

dermal irritation, dermal sensitization, phototoxicity, photoallergenicity, percutaneous

absorption, subchronic toxicity, teratogenicity, reproductive toxicity, genetic toxicity,

chronic toxicity, carcinogenicity, and potential human exposure (Api 2001).

In many experimental tests, the lowest observed effect level exceeds the maximum tested

dose. A table in the next subsection below will provide toxicity values found in key

studies. Indications of genotoxicity or carcinogenicity have been authoritatively deemed

to be at most equivocal and key authorities do not consider diethylphthalate a cancer risk

The Threshold Limit Value/Time Weighted Average (TLV-TWA) for diethyphthalate of

the American Conference of Government Industrial Hygienists (ACGIH) is 5 mg/m3

as

an 8-hour time weighted average. (WHO 2003). This TLV level in practice amounts to

about 50mg inhaled per person each work day for a lifetime (Api 2001). The highest

consumer food with diethylphthalate content (presumably from contamination from

packaging) in a UK study were quiches, averaging 2-3 mg/kg (human). Nevertheless 4

mg (absolute amount) was found to be the average ordinary human exposure/intake per

person per day (Kamrin 1991). Indoor air concentration of diethylphthalate recorded in

Tokyo recently was 0.1 µg/m3

(Otake 2004).

In general, significant systemic toxicity is not indicated -- high dose oral subchronic and

chronic tests on rats shows only gains in liver weight as an effect, which was attributable

to hypertophy and not toxicity, along with moderate decreased weight gain. (Api 2001;

Kamrin 1991) Guinea pigs experienced only slight liver and kidney damage when

exposed orally to a high dose of 1 mg/kg daily for up to 12 consecutive days.



7

Acute/Subchronic/Chronic Concerns

There is a broad consensus in the literature that diethylphthalate is generally of low acute

toxicity. (NTP 1995; Api 2001; WHO 2003). The following Lethal Dose 50 percent kill

(LD50) values tend to support this:

ROUTE ANIMAL LD50

Oral guinea pig 8,600 mg/kg

Oral mouse 6,172 mg/kg

Oral rat 8,600 mg/kg

Oral rabbit 1 gm/kg

Intraperitoneal mouse 2,749 mg/kg

Intraperitoneal rat 5,058 µL/kg

Subcutaneous guinea pig 3 gm/kg

Unreported route guinea pig 3 gm/kg

Unreported route mouse 8,600 mg/kg

Unreported route rat 9,500 mg/kg

(Adapted from RTECS 2004)

Human toxic effect lowest levels (Lowest Observed Adverse Effect Levels -- LOAELs)

have been reported to show at these concentrations:

Oral Human 357 µL/kg

Inhalation Human 1,000 mg/m3

(Adapted from RTECS 2004)

In humans, coughing and lacrimation occur at the inhalation dose 1.0 g/m3 (note: 200

times the Threshold Limit Value (TLV)). Miosis, dyspnea, amd general anesthetic effects

followed human oral exposure at the dose level above. Central nervous system

depression signs were seen in the animals prior to death. (RTECS 2004; HSDB 2004)

Acute doses in the range of .07-0.3g/kg delivered intravenously are the lowest levels for

lethality in mice and rabbits. Dermal exposure of rats at levels lower than 11g/kg, the

highest dose administered, failed to kill any (Api 2001).

No studies or record of human fatality from acute or chronic exposure have been found.

Animal and human studies indicate at most mild to moderate irritant toxicity to skin and

eyes. An administration to clipped rat skin of 2ml/kg at 6 hour intervals each day for two

weeks evoked erythema, desquamation, and mild epidermal thickening. Some irritation

was found in a 24 hour epicutaneous test on guinea pigs; moderate irritation only was



8

found on closed patch tests on rabbit skin. No effects occurred in a semiocclusive patch

test of exposure to 0.5 ml of diethylphthalate for four hours (Api 2001).

Intradermal administration to test animals have produced significant irritation. Ocular

exposure in one standard rabbit test elicited no obvious irritation (Lawrence 1975). Other

rabbit tests have shown some transitory severe conjunctival irritation, but the use of the

solvent ethanol may have been to blame (Api 2001).

Humans showed no primary irritation from applications of undiluted diethylphthalate

epicutaneously. An in vitro model of human skin showed marked cytotoxicity but this

could not be duplicated in vivo (Api 2001). A computer “mouse” containing

diethylphthalate appears to have induced dermatitis in the hands of two women according

to one case study (WHO 2003).

No dermal sensitization was elicited in guinea pigs and humans not prone to allergic

reactions or previously exposed have shown no little or no sensitization in volunteer

testing and case studies. A small proportion (1/30) workers previously exposed to

diethylphthalate exhibited some dermatitis (WHO 2003).

No significant photoxicity or photoallerginicity was elicited in tests on human volunteers

(Api 2001).

Chronic studies reveal that at very high doses (> 25g/kg/day) organ weight loss and

decreased weight gain are typically noted (RTECS 2004; Api 2001). Reports describing

cases of human toxic effects from chronic exposures were not found. Chronic rodent

testing also cited below in the subsection on carcinogenesis indicated no general toxic

effects from exposure (WHO 2003; NTP 1995).

Study or reports of neurological toxicity have not been found, other than a general central

nervous system depression at high toxic doses (Api 2001). Two studies found no toxic

pathology in the brain after administration orally of up to 3.7 g/kg in rats or mice (Brown

et al 1978; WHO 2003).

Reproductive Toxicity

Concerns may exist in the area of reproductive toxicity. Administration of

diethylphthalate to rats and mice has led to the increase in skeletal defects or rib number

alteration in offspring. (WHO 2003; Field et al 1993; Kamrin 1991; Singh et al 1972).

Testing of rats yielded no embryotoxicty or teratogenicity except an extra rib when

administration of up to 3.2g/kg (oral) diethylphthalate and 5.6 g/kg (percutaneous)

diethylphthalate occurred at the time of organogenesis. Fetal weight was lower in the

high-dose group (WHO 2003).



9

Direct injections of diethylphthalate (0.025 ml) into chicken eggs prior to embryonic

stage development has induced an increased level of deaths, with a small percentage of

malformations among the survivors (Api 2001). . A multigeneration developmental study of mice noted no signs of toxicity/defects in the

F0 generation after high oral dosing of 3.6g/kg for 14 weeks after cohabitation, and none

also in the subsequent (unexposed) generation F1, except for a reduced number of pups

per litter, mild inhibition of body weight gain, reduction by 30% in epididymal sperm

concentration, and moderate increase in liver and prostate weight (WHO 2003; NTP

1984). A NOAEL (no observed adverse effect level) for reproductive toxicity after

administration for over 14 weeks after gestation in pregnant SD rats was established at

750 mg/kg per day (WHO 2003).

No study indicating human maternal or developmental toxicity have been found.

Concerns have been raised, nonetheless, by the substantial presence of diethylphthalate in

the blood of pregnant women in urban areas (Adibi et al. 2003).

There has been some indication of possible detrimental effects by diethylphthalate on the

male reproductive system. In the multigenerational study cited above a reduction of 30%

in epididymal sperm concentration and an increase in prostate weight were noted. Human

sperm treated in vitro showed impairment in motility after less than 18 minutes of

exposure (Fredricsson et al 1993). Testosterone levels in rat testes and serum decreased

after an oral exposure to 2% diethylphthalate in the diet. Nevertheless, no other toxic

male reproductive injury was found anywhere in the rat, including no testicular damage

or Sertoli cell impairment (Api 2001). Mice showed no reproductive system effects of

any kind at the same level of dosing (Api 2001).

Leydig cells showed ultrastructural changes in male rats receiving 2g/kg daily for two

days. Smooth endoplasmic reticulum focal dilation and vesiculation, mitochondrial

swelling, and increased macrophage activity associated with the Leydig cell surface were

noted (Jones et al1993; WHO 2003).

Enormous oral doses induced some testicular weight changes in rats These occurred only

at a level of 44g-354g/kg delivered daily over 2-16 weeks (Brown et al 1978; RTECS

2004).

In vivo human studies or reports of testicular or male reproductive toxicity from

diethylphthalate have not been found.

Carcinogenicity

Key authorities deem the available evidence insufficient to declare diethylphthalate

carcinogenic. The EPA classifies diethyl phthalate as class D, unclassifiable as to human

carcinogenicity. The ACGIH classifies diethylphthalate as A4, not classifiable as a

human carcinogen (WHO 2003; US EPA 2004).



10

Evidence of carcinogenicity remains at best “equivocal” (WHO 2003; Api 2001). A

National Toxicology Program dermal study found no evidence of site-specific dermal

carcinogenicity in male and female rats in 2 year dermal administration studies applying

up to 1.6 g/kg of diethylphthalate per day (Api 2001).

Another dermal test of up to 1.1 g/kg per day of diethylphthalate in acetone for 103

weeks in both rats and mice turned up no neoplasia at the site of application, but at high

doses there was some increase in combined hepatocellular adenoma or carcinoma in male

mice. A non-dose-related increase in carcinomas occurred in female mice. This has been

regarded as equivocal as the rate among males was similar to the historical neoplasm

mean rate in males of that mice strain and because the female response was not dose-

related. Additionally the results were not duplicated in the tested rat species. (NTP

1995; Api 2001; WHO 2003)

Diethylphthalate was also dermally tested over a period of one year with cancer promoter

12-O-tetradecanoylphorbol-13-acetate and with initiator 7,12-dimethylbenz[a]anthracene.

No initiator or promoter activity was observed (NTP 1995).

Genotoxicity

Diethylphthalate was found not to be mutagenic in Salmonella strains TA 98, TA 100,

TA 1535 or TA 1537 with or without liver fraction S9 (NTP 1995). No in vivo studies

have been reported (WHO 2003). Chromosomal aberrations were also absent in Chinese

hamster ovary cells with and without S9 liver fraction. Nevertheless, sister-chromatid

exchanges (SCEs) were noted at 167-750 µg/ml concentrations but only with rat liver S9.

While this was regarded as some evidence for potential DNA damage in vivo, the NTP

cautioned that the endpoint is highly sensitive and it does not correlate well with

carcinogenicity in rodents (NTP 1995).



11

IV. PSYCHOGENIC EFFECTS

There are no known studies addressing psychogenic effects of exposure to

diethylphthalate. The general effects of perceived exposure to chemical or biological

warfare agents are treated in the supplement “Psychogenic Effects of Perceived Exposure

to Biochemical Warfare Agents.”



12

V. TREATMENT/PREVENTION

Standard protection procedures, and limitations of them, regarding diethyl phthalate

overexposure is provided below (Mallinckrodt Baker Inc. 2004):

Ventilation System:

A system of local and/or general exhaust is recommended to keep employee exposures

below the Airborne Exposure Limits. Local exhaust ventilation is generally preferred

because it can control the emissions of the contaminant at its source, preventing dispersion

of it into the general work area….

Personal Respirators (NIOSH Approved):

If the exposure limit is exceeded and engineering controls are not feasible, a half facepiece

particulate respirator (NIOSH type P95 or R95 filters) may be worn for up to ten times the

exposure limit or the maximum use concentration specified by the appropriate regulatory

agency or respirator supplier, whichever is lowest.. A full-face piece particulate respirator

(NIOSH type P100 or R100 filters) may be worn up to 50 times the exposure limit, or the

maximum use concentration specified by the appropriate regulatory agency, or respirator

supplier, whichever is lowest. Please note that N filters are not recommended for this

material. For emergencies or instances where the exposure levels are not known, use a full-

facepiece positive-pressure, air-supplied respirator. WARNING: Air-purifying respirators

do not protect workers in oxygen-deficient atmospheres.

Skin Protection:

Wear protective gloves and clean body-covering clothing.

Eye Protection: Use chemical safety goggles and/or full face shield where dusting or splashing of solutions

is possible. Maintain eye wash fountain and quick-drench facilities in work area.

Exposure to diethylphthalate has rarely been reported as harmful so no systematic

treatment methods specific to diethylphthalate are reported. TOXNET suggests applying

the emergency medical treatment protocol for dibutyl phthalate (HSDB 2004).



13

VI. SECONDARY SOURCE COMMENT

Secondary literature tends to be comprehensive.

The International Agency for Research on Cancer (IARC) provides no evaluation for

diethylphthalate (CAS 84-66-2). (See http://www.iarc.fr/).

Occasional references to gastrointestinal irritation (nausea) from oral exposure arise in

the literature, including Project SHAD’s Glossary (Project 112 2004; Mallinckrodt Baker

2004). The Merck Index claims polyneuritis as a possible effect (O’Neil 2001). But no

primary information on such toxic effects has been found, nor have they been reported in

recent reviews (WHO 2003; Api 2001; NTP 1995). It is suggested that the insertion of

dibutyl phthalate’s medical handling procedures in TOXNET’s discussion of

diethylphthalate treatment inside its diethylphthalate HSDB monograph, and which

includes statements describing dibutyl phthalate’s toxic effects of nausea and

polyneuritis, may be responsible for such confusion, or at least reflect a pattern of easy

confoundment that can give rise to such errors (HSDB 2004).

One author notes aptly that the similarity in common abbreviations and in the productive

uses of the two phthalic acid esters DEP (diethylphthalate) and DEHP (di (ethylhexyl)

phthalate) can lead to confusion (Kamrin 1991).



14

VII. BIBLIOGRAPHY WITH ABSTRACTS

{The following bibliography may contain supplementary material beyond those

cited in the text. Abstracts are reproduced without alteration from how they are

provided in their original source or database. Errors and defects of form, content,

or style are strictly those of the original source.}

Anonymous 1997. Reproductive toxicology. Diethylphthalate. Environ.Health Perspect.

105 Suppl 1: 245-246.

Adibi, et al. 2003. Prenatal exposures to phthalates among women in New York City and

Krakow, Poland. Environ.Health Perspect. 111(14): 1719-1722.

Experimental evidence has shown that certain phthalates can disrupt endocrine function

and induce reproductive and developmental toxicity. However, few data are available on

the extent of human exposure to phthalates during pregnancy. As part of the research

being conducted by the Columbia Center for Children's Environmental Health, we have

measured levels of phthalates in 48-hr personal air samples collected from parallel

cohorts of pregnant women in New York, New York, (n = 30) and in Krakow, Poland (n

= 30). Spot urine samples were collected during the same 48-hr period from the New

York women (n = 25). The following four phthalates or their metabolites were measured

in both personal air and urine: diethyl phthalate (DEP), dibutyl phthalate (DBP),

diethylhexyl phthalate (DEHP), and butyl benzyl phthalate (BBzP). All were present in

100% of the air and urine samples. Ranges in personal air samples were as follows: DEP

(0.26-7.12 microg/m3), DBP (0.11-14.76 microg/m3), DEHP (0.05-1.08 microg/m3), and

BBzP (0.00-0.63 microg/m3). The mean personal air concentrations of DBP, di-isobutyl

phthalate, and DEHP are higher in Krakow, whereas the mean personal air concentration

of DEP is higher in New York. Statistically significant correlations between personal air

and urinary levels were found for DEP and monoethyl phthalate (r = 0.42, p < 0.05),

DBP and monobutyl phthalate (r = 0.58, p < 0.01), and BBzP and monobenzyl

phthalate (r = 0.65, p < 0.01). These results demonstrate considerable phthalate

exposures during pregnancy among women in these two cohorts and indicate that

inhalation is an important route of exposure.

Agarwal, et al. 1985. Mutagenicity evaluation of phthalic acid esters and metabolites in

Salmonella typhimurium cultures. J.Toxicol.Environ.Health. 16(1): 61-69.

The mutagenic potential of dimethyl phthalate (DMP), diethyl phthalate (DEP), dibutyl

phthalate (DBP), and di-2-ethylhexyl phthalate (DEPH), as well as metabolites of DEHP-



15

-i.e., mono-2-ethylhexyl phthalate (MEHP), 2-ethylhexanol (2-EH), and phthalic acid

(PA)--were tested in Salmonella typhimurium cultures using the Ames test procedure.

The compounds were tested on strains TA98, TA100, TA1535, TA1537, TA1538, and

TA2637 for base-pair substitution or frameshift-type mutations. Spot tests yielded

negative responses for all compounds with the strains tested. Each compound was tested

for a dose-effect relationship in the TA98, TA100, TA1535, and TA1538 systems. DEP

and DBP exhibited a mildly positive response in both TA100 and TA1535 cultures, and

DMP showed a similar response in TA1535. Normalization of the data for cytotoxicity of

DMP suggests TA100 has a mildly positive effect. The higher doses of these compounds

exhibited some cytotoxic effects. The mutagenic effects were apparently abolished by the

addition of S9 fraction in TA100 and TA1535 cultures, while no effect, other than

cytotoxicity, was observed in the TA98 and TA1538 systems. DEHP, MEHP, 2-EH, and

PA exhibited no mutagenicity in any of the strains of Salmonella typhimurium tested,

with or without S9 metabolic activation. MEHP and 2-EH, however, exhibited a

moderate cytotoxic effect in most cultures.

Api. 2004. Evaluation of the dermal subchronic toxicity of phenoxyethyl isobutyrate in

the rat. Food Chem.Toxicol. 42(2): 307-311.

Phenoxyethyl isobutyrate (PEIB) is a fragrance and food ingredient that has been granted

GRAS status and approved by the FDA for food use. The present studies investigated the

dermal absorption parameters and subchronic toxicity of PEIB. For the absorption,

distribution and elimination study, Sprague-Dawley rats received a dermal application of

2-[ring U 14C]-PEIB under occlusion for 6 h. PEIB was diluted in diethyl phthalate

(DEP) to administer, a total application volume of 2 ml/kg, concentrations of 0.5, 5 and

50% ( congruent with 10, 100 and 1000 mg PEIB/kgBW). Approximately 61-69% of the

applied dose was recovered from the dressing and skin surface washing procedure

performed after 6-h exposure. By 72 h post dose, systemic elimination of radioactivity

was congruent with 18 to 19% of the absorbed dose via the urine with small amounts also

found in the feces (<1.0%). Terminal (72 h) tissue analysis showed that 0.35-0.72% of

the applied dose of radioactivity was retained in the carcass with low levels

(</=0.03%) measured also in the liver, kidney and gastrointestinal tract. Plasma levels

increased in a dose-related manner, with concentrations equal to 0.02, 0.2 and 2.0 microg

equiv/ml from low to high dose, respectively. The total recovery for these studies ranged

from 92.2 to 96.2% of the dermally applied radioisotope. In a 13-week subchronic rat

toxicity study, daily dermal applications of PEIB were made under occlusion for 6 h. All

groups were dosed at a constant 2 ml/kgBW volume of PEIB in the DEP vehicle at

concentrations calculated to administer 0, 100, 300 or 1000 mg PEIB/kgBW/day. Clinical

observations, assessments of skin irritation, hematology, and blood chemistry, necropsy,

and gross and histopathologic evaluation of tissues demonstrated no treatment-related



16

effects. The local skin irritation and systemic toxicity no-observed-effect-levels (NOELs)

for PEIB in this study were determined to be >1000 mg/kgBW/day.

Api. 2002. Sensitization methodology and primary prevention of the research institute for

fragrance materials. Dermatology. 205(1): 84-87.

The Research Institute for Fragrance Materials Inc. (RIFM) has approached sensitization

studies with fragrance materials as primary prevention of sensitization in the healthy,

normal population. Secondary prevention, or avoidance of elicitation, most often

suggested by dermatologists for patients presenting with dermatitis, has not been part of

its program effort. Historically, RIFM evaluated the sensitization potential of fragrance

materials using the human maximization test method; no animal models were used. In

general, petrolatum was used as the vehicle. This is a harsh procedure whose main use

may provide a measure of the uppermost limits of sensitization. Treating skin with

sodium lauryl sulfate may be problematic and finding a laboratory to conduct the study

may also be difficult. In addition, using a human predictive test method for both hazard

and safety assessments is not ideal. The current practice involves a hazard assessment

using an animal model, followed by a safety assessment in a human repeated-insult patch

test (HRIPT). The animal test method is used to identify the sensitization potential and a

no-effect level. Following a review of the no-effect level and the maximum skin level, a

safety assessment in humans can be conducted. RIFM also modified the original vehicle

used in sensitization testing, since petrolatum presents two major difficulties: solubility

and inconsistent effects on skin penetration. Since the greatest exposure to fragrance

materials is considered to be from a cologne-type product, ethanol was chosen as a more

realistic vehicle. Further modification resulted in combinations of ethanol and diethyl

phthalate, due to diethyl phthalate's use in many perfume formulations as a solvent and

fluidizer. Human testing should not be conducted as a hazard assessment. If conducted as

a safety study, induction of sensitization should be a rare occurrence. Thus, follow-up

studies are not meaningful since the number of sensitized volunteers would be low.

However, following a series of the RIPTs with various concentrations of

hydroxycitronellal, RIFM identified a group of 41 individuals who became sensitized. An

extensive 3-phase use study, with 3 diagnostic patch tests and 4 whole-body

dermatological examinations showed that most subjects were able to use a bar soap, a

moisurizing lotion and cologne-type products with up to 1% hydroxycitronellal. In

subjects where sensitization was induced by predictive testing, no serious recurring

adverse dermatological conditions developed.

Api. 2001. Toxicological profile of diethyl phthalate: a vehicle for fragrance and

cosmetic ingredients. Food Chem.Toxicol. 39(2): 97-108.

Diethyl phthalate (DEP; CAS No. 84-66-2) has many industrial uses, as a solvent and



17

vehicle for fragrance and cosmetic ingredients and subsequent skin contact. This review

focuses on its safety in use as a solvent and vehicle for fragrance and cosmetic

ingredients. Available data are reviewed for acute toxicity, eye irritation, dermal

irritation, dermal sensitization, phototoxicity, photoallergenicity, percutaneous

absorption, kinetics, metabolism, subchronic toxicity, teratogenicity, reproductive

toxicity, estrogenic potential, genetic toxicity, chronic toxicity, carcinogenicity, in vitro

toxicity, ecotoxicity, environmental fate and potential human exposure. No toxicological

endpoints of concern have been identified. Comparison of estimated exposure (0.73

mg/kg/day) from dermal applications of fragrances and cosmetic products with other

accepted industrial (5 mg/m(3) in air) and consumer exposures (350 mg/l in water; 0.75

mg/kg/day oral exposure) indicates no significant toxic liability for the use of DEP in

fragrances and cosmetic products. .

Berg, et al. 1991. Diethyl phthalate not dangerous. Am.J.Hosp.Pharm. 48(7): 1448-1449.

Beving, et al. 1990. Increased isotransferrin ratio and reduced erythrocyte and platelet

volumes in blood from thermoplastic industry workers. Ann.Occup.Hyg. 34(4): 391-397.

Ten women (aged 31-61 years) and five men (aged 20-59 years) occupationally exposed

to welding fumes of polyacetate containing diethylphthalate in a thermoplastic industry

were studied. They had been employed 1-33 years (median: 11 years). Seven women

(aged 35-55) and eight men (aged 26-73) acted as unexposed controls. The exposed

persons showed increased isotransferrin ratio in blood serum and reduced volumes of

erythrocytes and platelets in blood.

Blount, et al. 2000. Levels of seven urinary phthalate metabolites in a human reference

population. Environ.Health Perspect. 108(10): 979-982.

Using a novel and highly selective technique, we measured monoester metabolites of

seven commonly used phthalates in urine samples from a reference population of 289

adult humans. This analytical approach allowed us to directly measure the individual

phthalate metabolites responsible for the animal reproductive and developmental toxicity

while avoiding contamination from the ubiquitous parent compounds. The monoesters

with the highest urinary levels found were monoethyl phthalate (95th percentile, 3,750

ppb, 2,610 microg/g creatinine), monobutyl phthalate (95th percentile, 294 ppb, 162

microg/g creatinine), and monobenzyl phthalate (95th percentile, 137 ppb, 92 microg/g

creatinine), reflecting exposure to diethyl phthalate, dibutyl phthalate, and benzyl butyl

phthalate. Women of reproductive age (20-40 years) were found to have significantly

higher levels of monobutyl phthalate, a reproductive and developmental toxicant in

rodents, than other age/gender groups (p < 0.005). Current scientific and regulatory

attention on phthalates has focused almost exclusively on health risks from exposure to



18

only two phthalates, di-(2-ethylhexyl) phthalate and di-isononyl phthalate. Our findings

strongly suggest that health-risk assessments for phthalate exposure in humans should

include diethyl, dibutyl, and benzyl butyl phthalates.

Brown, et al. 1978. Short-term oral toxicity study of diethyl phthalate in the rat. Food

Cosmet.Toxicol. 16(5): 415-422.

Cafmeyer, et al. 1991. Possible leaching of diethyl phthalate into levothyroxine sodium

tablets. Am.J.Hosp.Pharm. 48(4): 735-739.

The possible leaching of diethyl phthalate into four currently marketed brands of

levothyroxine sodium tablets was investigated. Several strengths of levothyroxine sodium

tablets and sizes of containers were used. Samples were analyzed by high-performance

liquid chromatography (HPLC) to determine the levothyroxine sodium content and to

determine if any unidentified compounds were present. The packaging for the four brands

of tablets was also analyzed by using the same HPLC system to determine if any

extractable compounds could be detected in the tablets. The potencies of the four brands

of tablets were comparable. The tablets from the 100-count container of one brand (brand

A) were the only tablets found to contain an unidentified peak in the chromatogram. The

desiccants from the bottle showed the same unidentified compound, while the bottle and

closure did not yield the peak. Thin-layer chromatography and HPLC identified the peak

as diethyl phthalate, a plasticizer in the desiccant. Tablets, bottles, closures, and

desiccants for the 1000-count brand A product and all sizes of the other brands were

negative for the presence of diethyl phthalate. The desiccants in those containers were

from a different manufacturer than the desiccant in the brand A 100-count bottle. Diethyl

phthalate in the desiccant in 100-count bottles of brand A levothyroxine sodium tablets

appeared to have leached into the tablets.

Call, et al. 2001. An assessment of the toxicity of phthalate esters to freshwater benthos.

2. Sediment exposures. Environ.Toxicol.Chem. 20(8): 1805-1815.

Seven phthalate esters were evaluated for their 10-d toxicity to the freshwater

invertebrates Hyalella azteca and Chironomus tentans in sediment. The esters were

diethyl phthalate (DEP), di-n-butyl phthalate (DBP), di-n-hexyl phthalate (DHP), di-(2-

ethylhexyl) phthalate (DEHP), diisononyl phthalate (DINP), diisodecyl phthalate (DIDP),

and a commercial mixture of C7, C9, and C11 isophthalate esters (711P). All seven esters

were tested in a sediment containing 4.80% total organic carbon (TOC), and DBP alone

was tested in two additional sediments with 2.45 and 14.1% TOC. Sediment spiking

concentrations for DEP and DBP were based on LC50 (lethal concentration for 50% of

the population) values from water-only toxicity tests, sediment organic carbon

concentration, and equilibrium partitioning (EqP) theory. The five higher molecular



19

weight phthalate esters (DHP, DEHP, DINP, DIDP, 711P), two of which were tested and

found to be nontoxic in water-only tests (i.e., DHP and DEHP), were tested at single

concentrations between 2,100 and 3,200 mg/kg dry weight. Preliminary spiking studies

were performed to assess phthalate ester stability under test conditions. The five higher

molecular weight phthalate esters in sediment had no effect on survival or growth of

either C. tentans or H. azteca, consistent with predictions based on water-only tests and

EqP theory. The 10-d LC50 values for DBP and H. azteca were >17,400, >29,500,

and >71,900 mg/kg dry weight for the low, medium, and high TOC sediments,

respectively. These values are more than 30x greater than predicted by EqP theory and

may reflect the fact that H. azteca is an epibenthic species and not an obligative burrower.

The 10-d LC50 values for DBP and C. tentans were 826, 1,664, and 4.730 mg/kg dry

weight for the low, medium, and high TOC sediments, respectively. These values are

within a factor of two of the values predicted by EqP theory. Pore-water 10-d LC50

values for DBP (dissolved fraction) and C. tentans in the three sediments were 0.65, 0.89,

and 0.66 of the water-only LC50 value of 2.64 mg/L, thereby agreeing with EqP theory

predictions to within a factor of 1.5. The LC50 value for DEP and C. tentans was

>3,100 mg/kg dry weight, which is approximately 10x that predicted by EqP theory. It

is postulated that test chemical loss and reduced organism exposure to pore water may

have accounted for the observed discrepancies with EqP calculations for DEP

Call, et al. 2001. An assessment of the toxicity of phthalate esters to freshwater benthos.

1. Aqueous exposures. Environ.Toxicol.Chem. 20(8): 1798-1804.

Tests were performed with the freshwater invertebrates Hyalella azteca, Chironomus

tentans, and Lumbriculus variegatus to determine the acute toxicity of six phthalate

esters, including dimethyl phthalate (DMP), diethyl phthalate (DEP), di-n-butyl phthalate

(DBP), butylbenzyl phthalate (BBP), di-n-hexyl phthalate (DHP), and di-2-ethylhexyl

phthalate (DEHP). It was possible to derive 10-d LC50 (lethal concentration for 50% of

the population) values only for the four lower molecular weight esters (DMP, DEP, DBP,

and BBP), for which toxicity increased with increasing octanol-water partition coefficient

(Kow) and decreasing water solubility. The LC50 values for DMP, DEP, DBP, and BBP

were 28.1, 4.21, 0.63, and 0.46 mg/L for H. azteca; 68.2, 31.0, 2.64, and > 1.76 mg/L

for C. tentans; and 246, 102, 2.48, and 1.23 mg/L for L. variegatus, respectively. No

significant survival reductions were observed when the three species were exposed to

either DHP or DEHP at concentrations approximating their water solubilities.

CHEMIDplus 2004. Diethyl Phthalate. TOXNET [NLM Database].

[http://chem.sis.nlm.nih.gov/chemidplus/ProxyServlet?objectHandle=DBMaint&actionH

andle=default&nextPage=jsp/chemidlite/ResultScreen.jsp&TXTSUPERLISTID=000084

662]



20

Elsisi, et al. 1989. Dermal absorption of phthalate diesters in rats. Fundam.Appl.Toxicol.

12(1): 70-77.

This study examined the extent of dermal absorption of a series of phthalate diesters in

the rat. Those tested were dimethyl, diethyl, dibutyl, diisobutyl, dihexyl, di(2-ethylhexyl),

diisodecyl, and benzyl butyl phthalate. Hair from a skin area (1.3 cm in diameter) on the

back of male F344 rats was clipped, the [14C]phthalate diester was applied in a dose of

157 mumol/kg, and the area of application was covered with a perforated cap. The rat

was restrained and housed for 7 days in a metabolic cage that allowed separate collection

of urine and feces. Urine and feces were collected every 24 hr, and the amount of 14C

excreted was taken as an index of the percutaneous absorption. At 24 hr, diethyl phthalate

showed the greatest excretion (26%). As the length of the alkyl side chain increased, the

amount of 14C excreted in the first 24 hr decreased significantly. The cumulative

percentage dose excreted in 7 days was greatest for diethyl, dibutyl, and diisobutyl

phthalate, about 50-60% of the applied 14C; and intermediate (20-40%) for dimethyl,

benzyl butyl, and dihexyl phthalate. Urine was the major route of excretion of all

phthalate diesters except for diisodecyl phthalate. This compound was poorly absorbed

and showed almost no urinary excretion. After 7 days, the percentage dose for each

phthalate that remained in the body was minimal and showed no specific tissue

distribution. Most of the unexcreted dose remained in the area of application. These data

show that the structure of the phthalate diester determines the degree of dermal

absorption. Absorption maximized with diethyl phthalate and then decreased significantly

as the alkyl side chain length increased.

Field, et al. 1993. Developmental toxicity evaluation of diethyl and dimethyl phthalate in

rats. Teratology. 48(1): 33-44.

Diethyl phthalate (DEP) and dimethyl phthalate (DMP), phthalic acid ester (PAE)

plasticizers, were evaluated for developmental toxicity because of reports in the literature

that some PAE were embryotoxic and teratogenic. A previous study (Singh et al., '72)

suggested that an increased incidence of skeletal defects in rats might result from

gestational exposure to DEP (0.6-1.9 g/kg) or DMP (0.4-1.3 g/kg), ip, on gestational days

(gd) 5, 10, and 15. In the current study DEP (0, 0.25, 2.5, and 5%) or DMP (0, 0.25, 1,

and 5%) in feed (approximately 0.2-4.0 g/kg/day) were supplied to timed-mated rats from

gd 6 to 15. Treatment with 5% DMP resulted in increased relative maternal liver weight.

Also, animals exhibited reduced body weight gain during treatment (5% DEP or DMP)

and during gestation (5% DEP). Weight gain corrected for gravid uterine weight was also

reduced in animals fed 5% DEP. However, high-dose treatment with either DEP or DMP

resulted in changes in food and water consumption paralleling the body weight

reductions, suggesting that apparent toxic effects on maternal body weight may reflect

PAE/feed unpalatability. Treatment with 2.5% DEP resulted in only transient changes in



21

body weight during early treatment. The only maternal effects at 0.25 or 1% DMP were

minor changes in food and/or water consumption, and there were no effects at 0.25%

DEP. Thus, the NOAELs for maternal toxicity were 1% DMP and 0.25% DEP. In

contrast to the observed maternal toxicity, there was no effect of DEP or DMP treatment

on any parameter of embryo/fetal development, except an increased incidence of

supernumerary ribs (a variation) in the 5% DEP group. These results do not support the

conclusion of other investigators that DEP and DMP are potent developmental toxicants.

Rather, they suggest that the short-chain PAE are less developmentally toxic than PAE

with more complex substitution groups, e.g., di(2-ethylhexyl) phthalate, mono(2-

ethylhexyl) phthalate, and butyl benzyl phthalate.

Foster, et al. 1983. Effect of DI-n-pentyl phthalate treatment on testicular steroidogenic

enzymes and cytochrome P-450 in the rat. Toxicol.Lett. 15(2-3): 265-271.

Treatment of young male rats with dipentyl phthalate (DPP) produced significant

decreases in testicular cytochrome P-450, cytochrome P-450 dependent microsomal

steroidogenic enzymes (17 alpha-hydroxylase, 17-20 lyase) and in the maximal binding

of a natural substrate (progesterone) to testis microsomes. No effect was demonstrated by

this compound on hepatic cytochrome P-450 content. Treatment of animals with a

phthalate ester not causing testicular atrophy (diethyl phthalate; DEP) produced no

significant changes in any of the parameters measured. This effect on the enzymes

responsible for androgen production may be important as a mechanism of action involved

in the development of phthalate-induced testicular damage.

Fredricsson et al 1993. Human sperm motility is affected by plasticizers and diesel

particle extracts. Pharmacol Toxicol. 72(2):128-33.

In order to test various drugs and possibly hazardous compounds on living cells in vitro a

system with human spermatozoa was employed. A population of human spermatozoa was

transferred into a defined medium by a swim-up procedure or by separation on a Percoll

gradient. Such a population is rather homogenous with respect to motility characteristics

and was found to be useful for this purpose. Different modes of response were recorded,

indicating various effect mechanisms. Effects of various phthalates used as plastic

softeners in the production of medical equipment, and extracts from diesel particulate

material were recorded. All these compounds interfered with sperm motility in a dose-

response fashion. Immediate effects of phthalates were modest, but upon prolonged

exposure effects became more evident. Sperm motility was more affected by diethyl-

hexyl and dibutyl phthalates. Significant effects were noted for the different phthalates

with regard both to percent motility and to some of the various qualities of motility, such

as velocity, linearity and amplitude of the track. Thus, the pattern of response considering

the motion variables was not the same with the different phthalates. With regard to the

effects on sperm motion di-n-octyl phthalate seemed to be the least toxic, followed by

dibutyl phthalate. The initial effects of diesel particulate extracts were moderate and

mainly restricted to percent motile sperm but upon exposure for 18 hr the effects became

more pronounced for all the movement variables.



22

Fromme, et al. 2004. Occurrence of phthalates and musk fragrances in indoor air and

dust from apartments and kindergartens in Berlin (Germany). Indoor Air. 14(3): 188-195.

In this study, the occurrence of persistent environmental contaminants room air samples

from 59 apartments and 74 kindergartens in Berlin were tested in 2000 and 2001 for the

presence of phthalates and musk fragrances (polycyclic musks in particular). These

substances were also measured in household dust from 30 apartments. The aim of the

study was to measure exposure levels in typical central borough apartments,

kindergartens and estimate their effects on health. Of phthalates, dibutyl phthalate had the

highest concentrations in room air, with median values of 1083 ng/m(3) in apartments

and 1188 ng/m(3) in kindergartens. With around 80% of all values, the main phthalate in

house dust was diethylhexyl phthalate, with median values of 703 mg/kg (range: 231-

1763 mg/kg). No statistically significant correlation could be found between air and dust

concentration. Musk compounds were detected in the indoor air of kindergartens with

median values of 101 ng/m(3) [1,3,4,6,7,8-hexahydro-4,6,6,7,8,8- hexamethylcyclopenta-

(g) 2-benzopyrane (HHCB)] and 44 ng/m(3) [7-acetyl-1,1,3,4,4,6-hexamethyl-tetraline

(AHTN)] and maximum concentrations of up to 299 and 107 ng/m(3) respectively. In

household dust HHCB and AHTN were detected in 63 and 83% of the samples with

median values of 0.7 and 0.9 mg/kg (Maximum: 11.4 and 3.1 mg/kg) each. On

comparing the above phthalate concentrations with presently acceptable tolerable daily

intake values (TDI), we are talking about only a small average intake [di(2-ethylhexyl)

phthalate and diethyl phthalate less than 1 and 8% of the TDI] by indoor air for children.

The dominant intake path was the ingestion of foodstuffs. For certain subsets of the

population, notably premature infants (through migration from soft polyvinyl chloride

products), children and other patients undergoing medical treatment like dialysis,

exchange transfusion, an important additional intake of phthalates must taken into

account. PRACTICAL IMPLICATIONS: The phthalate and musk compounds load in a

sample of apartments and kindergartens were low with a typical distribution pattern in air

and household dust, but without a significant correlation between air and dust

concentration. The largest source of general population exposure to phthalates is dietary.

For certain subsets of the general population non-dietary ingestion (medical and

occupational) is important.

Frosch, et al. 1995. Patch testing with fragrances: results of a multicenter study of the

European Environmental and Contact Dermatitis Research Group with 48 frequently used

constituents of perfumes. Contact Dermatitis. 33(5): 333-342.



23

The objective of this study was to determine the frequency of reactivity to a series of

commonly used fragrances in dermatological patients. A total of 48 fragrances (FF) were

chosen, based on the publication of Fenn in 1989 in which the top 25 constituents of 3

types (1. perfumes, 2. household products, 3. soaps) of 400 commercial products on the

US market had been determined. In a pilot study on a total of 1069 patients in 11 centres,

the appropriate test concentration and vehicle were examined. For most fragrances, 1%

and 5% were chosen, and petrolatum proved to be the best vehicle in comparison to

isopropyl myristate and diethyl phthalate. In the main study, a set of 5 to 10 fragrances at

2 concentrations was patch tested in each centre on a minimum of 100 consecutive

patients seen in the patch test clinic. These patients were also patch tested to a standard

series with the 8% fragrance mix (FM) and its 8 constituents. In patients with a positive

reaction to any of the 48 FF, a careful history with regard to past or present reactions to

perfumed products was taken. A total of 1323 patients were tested in 11 centres. The 8%

FM was positive in 89 patients (8.3% of 1072 patients). Allergic reactions to the

constituents were most frequent to oak moss (24), isoeugenol (20), eugenol (13),

cinnamic aldehyde (10) and geraniol (8). Reactions read as allergic on day 3/4 were

observed only 10X to 7 materials of the new series (Iso E Super (2), Lyral (3), Cyclacet

(1), DMBCA (1), Vertofix (1), citronellol (1) and amyl salicylate (1)). The remaining 41

fragrances were negative. 28 irritant or doubtful reactions on day 3/4 were observed to a

total of 19 FF materials (more than 1 reaction: 5% citronellol (2), 1% amyl salicylate (2),

1% isononyl acetate (3), 0.1% musk xylol (2), 1% citral (2), and 1% ionone beta (2)).

Clinical relevance of positive reactions to any of the FF series was not proved in a single

case. This included the 4 reactions in patients who were negative to the 8% FM. In

conclusion, the top 25 fragrances commonly found in various products caused few

reactions in dermatological patients and these few appeared to be clinically irrelevant,

with the possible exception of Lyral. However, this data should be interpreted in the light

of the relatively small number of patients tested (only 100 in most centres).

Ghorpade, et al. 2002. Toxicity study of diethyl phthalate on freshwater fish Cirrhina

mrigala. Ecotoxicol.Environ.Saf. 53(2): 255-258.

Diethyl phthalate (DEP) is used as a plasticizer, a detergent base, in aerosol sprays, as a

perfume binder in incense sticks and after-shave lotions. It is known to be a contaminant

of freshwater and marine ecosystems. Therefore, a study was designed to determine the

toxic effects of DEP on a freshwater fish, Cirrhina mrigala. The fish was treated with 25,

50, 75, and 100 ppm (w/v) DEP dissolved in acetone to determine the LC50. Positive

controls were treated with acetone only. There was 100% mortality observed within 24 h

in 75 and 100 ppm, and 50% mortality in 50 ppm treated fish in 72 h. Those treated at 25

ppm showed only 10% mortality within 72 h and remaining fish continued to survive.

The surviving fish were treated with 25 ppm DEP once daily for 3 days with every



24

change of water (Group III). One group was maintained as negative control in

dechlorinated water (Group I) and the other group received acetone once daily for 3 days

with every change of water and was used as positive control (Group II). Fish were killed

by cold narcosis on an ice block and dissected to obtain liver, muscle, and brain samples;

10% homogenates in ice-cold saline were prepared. Brain and muscle

acetylcholinesterase (AchE) activity was measured. Liver aspartate (AST) and alanine

aminotransferase (ALT), and liver and muscle succinate dehydrogenase (SDH) alkaline

and acid phosphate (ALP and ACP) were measured. There was a significant increase in

liver and muscle ACP and ALP in DEP-treated fish compared with positive and negative

controls. There was a significant increase in muscle SDH and liver ALT (ALT) in DEP-

treated fish compared with positive and negative controls. Brain AchE level was

significantly decreased in DEP-treated fish compared to positive and negative controls.

These results indicate that DEP brings about significant changes in the activity of certain

liver and muscle enzymes. These alterations in enzyme activity may have long-term

effects on that are continuously exposed to low doses of DEP in the aquatic environment.

Gollamudi, et al. 1985. Effects of phthalic acid esters on drug metabolizing enzymes of

rat liver. J.Appl.Toxicol. 5(6): 368-371.

Di(2-ethylhexyl)phthalate (DEHP) inhibited UDP-glucuronyltransferase activity of rat

liver in vitro and in vivo. Diethyl phthalate and dimethoxyethyl phthalate also inhibited

this enzyme in vitro. On the other hand, DEHP did not inhibit the activity of the cytosolic

enzyme N-acetyltransferase; it also did not alter the levels of rat liver microsomal

cytochrome P-450 in vitro. It is suggested that DEHP may alter the composition of

microsomal phospholipids.

Gray, et al. 2000. Perinatal exposure to the phthalates DEHP, BBP, and DINP, but not

DEP, DMP, or DOTP, alters sexual differentiation of the male rat. Toxicol.Sci. 58(2):

350-365.

In mammals, exposure to antiandrogenic chemicals during sexual differentiation can

produce malformations of the reproductive tract. Perinatal administration of AR

antagonists like vinclozolin and procymidone or chemicals like di(2-ethylhexyl) phthalate

(DEHP) that inhibit fetal testicular testosterone production demasculinize the males such

that they display reduced anogenital distance (AGD), retained nipples, cleft phallus with

hypospadias, undescended testes, a vaginal pouch, epididymal agenesis, and small to

absent sex accessory glands as adults. In addition to DEHP, di-n-butyl (DBP) also has

been shown to display antiandrogenic activity and induce malformations in male rats. In

the current investigation, we examined several phthalate esters to determine if they

altered sexual differentiation in an antiandrogenic manner. We hypothesized that the

phthalate esters that altered testis function in the pubertal male rat would also alter testis



25

function in the fetal male and produce malformations of androgen-dependent tissues. In

this regard, we expected that benzyl butyl (BBP) and diethylhexyl (DEHP) phthalate

would alter sexual differentiation, while dioctyl tere- (DOTP or DEHT), diethyl (DEP),

and dimethyl (DMP) phthalate would not. We expected that the phthalate mixture

diisononyl phthalate (DINP) would be weakly active due to the presence of some

phthalates with a 6-7 ester group. DEHP, BBP, DINP, DEP, DMP, or DOTP were

administered orally to the dam at 0.75 g/kg from gestational day (GD) 14 to postnatal day

(PND) 3. None of the treatments induced overt maternal toxicity or reduced litter sizes.

While only DEHP treatment reduced maternal weight gain during the entire dosing

period by about 15 g, both DEHP and DINP reduced pregnancy weight gain to GD 21 by

24 g and 14 g, respectively. DEHP and BBP treatments reduced pup weight at birth

(15%). Male (but not female) pups from the DEHP and BBP groups displayed shortened

AGDs (about 30%) and reduced testis weights (about 35%). As infants, males in the

DEHP, BBP, and DINP groups displayed femalelike areolas/nipples (87, 70, and 22% (p

< 0.01), respectively, versus 0% in other groups). All three of the phthalate treatments

that induced areolas also induced a significant incidence of reproductive malformations.

The percentages of males with malformations were 82% (p < 0.0001) for DEHP, 84%

(p < 0.0001) for BBP, and 7.7% (p < 0.04) in the DINP group. In summary, DEHP,

BBP, and DINP all altered sexual differentiation, whereas DOTP, DEP, and DMP were

ineffective at this dose. Whereas DEHP and BBP were of equivalent potency, DINP was

about an order of magnitude less active.

Hansen, et al. 2001. 1H NMR of compounds with low water solubility in the presence of

erythrocytes: effects of emulsion phase separation. Eur.Biophys.J. 30(1): 69-74.

When lipophilic compounds like diethyl phthalate (DEP) were added to water, two sets of

resonances appeared in the 1H NMR spectrum, whereas when added in concentrations

above approximately 3.5 mM to erythrocytes in a high haematocrit suspension, only one

set of resonances was observed at the low-frequency position. The appearance of one set

of resonances at lower frequency was found to be common to a series of lipophilic

compounds in erythrocytes. The appearance of the NMR spectra is ascribed to the

existence of an emulsion, meaning two different phases of a compound: a

"droplet" (resonances to lower frequency) and aqueous dissolved phase

(resonances to higher frequency). The absence of the resonances from the dissolved

phase in erythrocyte solution is ascribed to exchange broadening. The absolute chemical

shift of the compound in its "droplet" phase was also measured using a

cylindrical/spherical microcell. This arrangement mimicked the geometry of the

dissolved versus the phase-separated species and thus obviated the effect of a difference

in magnetic susceptibility between the "droplet" solute and its aqueous

solution. Factors influencing the formation of emulsion phases such as erythrocytes,



26

haemoglobin and smaller proteins were investigated; they are found to be effective in the

order given.

Harris, et al. 1997. The estrogenic activity of phthalate esters in vitro. Environ.Health

Perspect. 105(8): 802-811.

A large number of phthalate esters were screened for estrogenic activity using a

recombinant yeast screen. a selection of these was also tested for mitogenic effect on

estrogen-responsive human breast cancer cells. A small number of the commercially

available phthalates tested showed extremely weak estrogenic activity. The relative

potencies of these descended in the order butyl benzyl phthalate (BBP) > dibutyl

phthalate (DBP) > diisobutyl phthalate (DIBP) > diethyl phthalate (DEP) >

diisiononyl phthalate (DINP). Potencies ranged from approximately 1 x 10(6) to 5 x

10(7) times less than 17beta-estradiol. The phthalates that were estrogenic in the yeast

screen were also mitogenic on the human breast cancer cells. Di(2-ethylhexyl) phthalate

(DEHP) showed no estrogenic activity in these in vitro assays. A number of metabolites

were tested, including mono-butyl phthalate, mono-benzyl phthalate, mono-ethylhexyl

phthalate, mon-n-octyl phthalate; all were wound to be inactive. One of the phthalates,

ditridecyl phthalate (DTDP), produced inconsistent results; one sample was weakly

estrogenic, whereas another, obtained from a different source, was inactive. analysis by

gel chromatography-mass spectometry showed that the preparation exhibiting estrogenic

activity contained 0.5% of the ortho-isomer of bisphenol A. It is likely that the presence

of this antioxidant in the phthalate standard was responsible for the generation of a dose-

response curve--which was not observed with an alternative sample that had not been

supplemented with o,p'-bisphenol A--in the yeast screen; hence, DTDP is probably not

weakly estrogenic. The activities of simple mixtures of BBP, DBP, and 17beta-estradiol

were assessed in the yeast screen. No synergism was observed, although the activities of

the mixtures were approximately additive. In summary, a small number of phthalates are

weakly estrogenic in vitro. No data has yet been published on whether these are also

estrogenic in vitro. No data has yet been published on whether these are also estrogenic in

vivo; this will require tests using different classes of vertebrates and different routes of

exposure.

Hauser, et al. 2004. Medications as a source of human exposure to phthalates.

Environ.Health Perspect. 112(6): 751-753.

Phthalates are a group of multifunctional chemicals used in consumer and personal care

products, plastics, and medical devices. Laboratory studies show that some phthalates are

reproductive and developmental toxicants. Recently, human studies have shown

measurable levels of several phthalates in most of the U.S. general population. Despite

their widespread use and the consistent toxicologic data on phthalates, information is



27

limited on sources and pathways of human exposure to phthalates. One potential source

of exposure is medications. The need for site-specific dosage medications has led to the

use of enteric coatings that allow the release of the active ingredients into the small

intestine or in the colon. The enteric coatings generally consist of various polymers that

contain plasticizers, including triethyl citrate, dibutyl sebacate, and phthalates such as

diethyl phthalate (DEP) and dibutyl phthalate (DBP). In this article we report on

medications as a potential source of exposure to DBP in a man who took Asacol [active

ingredient mesalamine (mesalazine)] for the treatment of ulcerative colitis. In a spot urine

sample from this man collected 3 months after he started taking Asacol, the concentration

of monobutyl phthalate, a DBP metabolite, was 16,868 ng/mL (6,180 micro g/g

creatinine). This concentration was more than two orders of magnitude higher than the

95th percentile for males reported in the 1999-2000 National Health and Nutrition

Examination Survey (NHANES). The patient's urinary concentrations of monoethyl

phthalate (443.7 ng/mL, 162.6 micro g/g creatinine), mono-2-ethylhexyl phthalate (3.0

ng/mL, 1.1 micro g/g creatinine), and monobenzyl phthalate (9.3 ng/mL, 3.4 micro g/g

creatinine) were unremarkable compared with the NHANES 1999-2000 values. Before

this report, the highest estimated human exposure to DBP was more than two orders of

magnitude lower than the no observable adverse effect level from animal studies. Further

research is necessary to determine the proportional contribution of medications, as well

as personal care and consumer products, to a person's total phthalate burden.

HSDB [Hazardous Substances Data Bank] 2004. Diethyl Phthalate. TOXNET.

[http://toxnet.nlm.nih.gov/cgi-bin/sis/search/f?./temp/~qbM8dG:1].

Hu, et al. 2003. Survey of phthalate pollution in arable soils in China. J.Environ.Monit.

5(4): 649-653.

The problem of pollution by phthalates is of global concern due to their widespread

occurrence, toxicity and endocrine disruption properties. The contamination by phthalates

such as dimethyl phthalate (DMP), diethyl phthalate (DEP), di-n-butyl phthalate (DBP)

and di(2-ethylhexyl) phthalate (DEHP) in 23 arable soils throughout China was

investigated to evaluate the present pollution situation. The survey results demonstrated

that phthalates were ubiquitous pollutants in soils in China. The total concentrations of

phthalates differed from one location to another, and ranged from 0.89 to 10.03 mg kg(-

1) with a median concentration of 3.43 mg kg(-1). Among the phthalates, DEHP was

dominant and detected in all 23 soils. DEP and DBP were also in abundance, and DMP

was rarely detected. Similar contamination patterns were observed in all 23 soils. A

distinct feature of phthalate pollution in China was that the average concentration in

northern China was higher than that in southern China. In addition, a close relationship

was observed between the concentration of phthalates in soils and the consumption of



28

agricultural film. The correlation showed that the application of agriculture film might be

a significant pollution source of phthalates in arable soils of China. The potential risk of

phthalates in soils was assessed on the basis of current guide values and limits.

Jones, et al. 1993. The influence of phthalate esters on Leydig cell structure and function

in vitro and in vivo. Exp.Mol.Pathol. 58(3): 179-193.

Phthalate esters are widely used in the manufacture of plastics and have been shown to

cause testicular toxicity, purportedly, by targeting the Sertoli cell alone. Recent evidence,

however, indicates that a paracrine control exists between Sertoli and Leydig cells and

the breakdown of one component of this relationship is therefore detrimental to normal

function. However, no data that explore the influence of testicular toxins on Leydig cell

structure and function have been published hitherto. The preliminary studies reported

here were initiated to test the hypothesis that phthalate intoxication may adversely alter

Leydig cell structural and functional integrity. Four phthalate esters, namely, di(2-

ethylhexyl) phthalate (DEHP, di-n-pentyl phthalate (DPP)., di-n-octyl phthalate (DOP),

and diethyl phthalate (DEP) were investigated in vivo and their monoesters (MEHP,

MPP, MOP, and MEP, respectively) in vitro for indications of Leydig cell toxicity in the

rat. Rats were dosed by oral gavage with 2 g phthalate diester/kg/day in corn oil vehicle

for 2 days, while Leydig cell primary cultures were incubated with 1,000 microM

monoester for 2 hr. Light and electron microscopy were undertaken to determine the type

and degree of any changes. Phthalate esters exerted a direct effect on Leydig cell

structure and function (as determined by testosterone output) with correlation of the in

vitro and in vivo effects of MEHP (DEHP) and MOP (DOP). No effects on Leydig cell

structure or function were seen with MPP (DPP), although Sertoli cell cytoplasmic

rarefaction and vacuolation were observed in vivo. DEP produced Leydig cell

ultrastructural alterations in vivo. We conclude that individual phthalate esters may exert

effects on both Sertoli and Leydig cells or one cell type alone.

Jonsson, et al. 2003. Toxicity of mono- and diesters of o-phthalic esters to a crustacean,

a green alga, and a bacterium. Environ.Toxicol.Chem. 22(12): 3037-3043.

The degradation of phthalic acid diesters may lead to formation of o-phthalic acid and

phthalic acid monoesters. The ecotoxic properties of the monoesters have never been

systematically investigated, and concern has been raised that these degradation products

may be more toxic than the diesters. Therefore, the aquatic toxicity of phthalic acid, six

monoesters, and five diesters of o-phthalic acid was tested in three standardized toxicity



29

tests using the bacteria Vibrio fischeri, the green algae Pseudokirchneriella subcapitata,

and the crustacean Daphnia magna. The monoesters tested were monomethyl, monoethyl,

monobutyl, monobenzyl, mono(2-ethylhexyl), and monodecyl phthalate, while the

diesters tested were dimethyl, diethyl, dibutyl, butylbentyl, and di(2-ethylhexyl)phthalate,

which were assumed to be below their water solubility. The median effective

concentration (EC50) values for the three organisms ranged from 103 mg/L to >4.710

mg/L for phthalic acid, and corresponding values for the monoesters ranged from 2.3

mg/L (monodecyl phthalate in bacteria test) to 4,130 mg/L (monomethyl phthalate in

bacteria test). Dimethyl and diethyl phthalate were found to be the least toxic of the

diesters (EC50 26.2-377 mg/L), and the toxicity of the other diesters (butylbenzyl and

dibutyl phthalate) ranged from 0.96 to 7.74 mg/L. In general, the phthalate monoesters

(degradation products) were less toxic than the corresponding diesters (mother

compounds).

Kamrin, et al. 1991. Diethyl phthalate: a perspective. J.Clin.Pharmacol. 31(5): 484-489.

Keire, et al. 2001. Diethyl phthalate, a chemotactic factor secreted by Helicobacter

pylori. J.Biol.Chem. 276(52): 48847-48853.

The structure of a small-molecule, non-peptide chemotactic factor has been determined

from activity purified to apparent homogeneity from Helicobacter pylori supernatants. H.

pylori was grown in brucella broth media until one liter of solution had 0.9 absorbance

units. The culture was centrifuged, and the bacteria re-suspended in physiological saline

and incubated at 37 degrees C for 4 h. A monocyte migration bioassay revealed the

presence of a single active chemotactic factor in the supernatant from this incubation. The

chemotactic factor was concentrated by solid phase chromatography and purified by

reverse phase high pressure liquid chromatography. The factor was shown to be

indistinguishable from diethyl phthalate (DEP) on the basis of multiple criteria including

nuclear magnetic resonance spectroscopy, electron impact mass spectroscopy, UV visible

absorption spectrometry, GC and high pressure liquid chromatography retention times,

and chemotactic activity toward monocytes. Control experiments with incubated culture

media without detectable bacteria did not yield detectable DEP, suggesting it is

bacterially derived. It is not known if the bacteria produce diethyl phthalate de novo or if

it is a metabolic product of a precursor molecule present in culture media. DEP produced

by H. pylori in addition to DEP present in man-made products may contribute to the high

levels of DEP metabolites observed in human urine. DEP represents a new class of

chemotactic factor.

Kelman, et al. 1999. Chemical components of shredded paper insulation: a preliminary

study. Appl.Occup.Environ.Hyg. 14(3): 192-197.



30

We conducted an evaluation of shredded paper insulation to identify potentially toxic

components. The study was to provide a preliminary characterization of a few samples of

insulation currently in use. The following samples were analyzed: previously produced

insulation (PPI) containing fire retardants, shredded recycled paper (PPI feedstock),

freshly produced insulation (FPI), and insulation which had been installed in a residence

(II). Volatile constituents were analyzed by GC-MS from headspace air of samples held

at room temperature or heated to 90 degrees C. Extractable constituents were sampled by

extracting with methylene chloride, and analyzing by GC-MS. Formaldehyde analysis

was done according to EPA Method TO11. Headspace air at room temperature contained

no detectable quantities of volatile constituents for any sample measured. In headspace

air at 90 degrees C, only PPI contained traces of aliphatic and aromatic hydrocarbons and

higher aldehydes, and FPI traces of toluene. Extracts of PPI contained traces of

octadecadienoic acid methyl ester and aliphatic and aromatic hydrocarbons and higher

aldehydes. Extracts of PPI feedstock contained traces of a substituted

cyclohexenecarboxylic acid. FPI contained extractable diethyl phthalate (30-50

micrograms/g). Extracts of II contained traces of methyl palmitate, an octadecenoic acid

methyl ester, and a phthalate plasticizer. No formaldehyde was detected. PPI was

composed of approximately 98 percent paper fiber and 2 percent pre-gelatinized starch.

PPI samples agglomerated together with less than 0.01 percent separating from clumps as

fine dust. Boron and sodium were expected and confirmed because they were added to

PPI and FPI as fire retardants. Chromium, copper, iron, potassium, magnesium,

manganese, phosphorus, and silicon were present at detectable concentrations. Study

calculations indicate that an occupant would have to completely consume all the fine

particles produced from 3.3 kg of insulation per day to have an intake of boron equivalent

to the EPA RfD. No other constituent appeared to be present even close to toxicologically

relevant amounts.

Kozumbo, et al. 1982. Assessment of the mutagenicity of phthalate esters.

Environ.Health Perspect. 45: 103-109.

The Ames assay was used to investigate the mutagenicity of several phthalate esters as an

approximation of their carcinogenic potential. The ortho diesters, dimethyl phthalate

(DMP) and diethyl phthalate (DEP) produced a positive dose-related mutagenic response

with Salmonella TA100, but only in the absence of S-9 liver enzymes. Dibutyl, di(2-

ethylhexyl), mono(2-ethylhexyl), and butyl benzyl phthalate as well as the dimethyl

isophthalate and terephthalates and the trimethyl ester, trimellitate, were not mutagenic

with TA100 or TA98 in the presence or absence of S-9. In a host-mediated assay, extracts

of 24-hr urines of rats injected IP with DMP (2 g/kg) were not mutagenic to TA100 at

levels up to 8 equivalent-ml of urine/plate (representing 30% of their daily urinary

output). In vitro studies revealed that S-9 associated esterase hydrolyzed DMP to the



31

monoester and methanol and eliminated its mutagenicity. Whole rat skin was shown to

have about 1.5% of the DMP-esterase activity of liver, when compared on a wet weight

basis. An in vitro binding study indicated that epidermal macromolecules bound DMP at

a severalfold greater rate than hepatic macromolecules. Thus, both the mutagenicity and

binding of DMP are inversely related to the metabolism of this compound. These results

suggest that skin could be at high risk for a mutagenic/carcinogenic insult.

Lamb, et al. 1987. Reproductive effects of four phthalic acid esters in the mouse.

Toxicol.Appl.Pharmacol. 88(2): 255-269.

These studies compared the reproductive toxicity of four phthalates by a continuous

breeding protocol. Mice were given diets with diethyl phthalate (DEP) (0.0, 0.25, 1.25, or

2.5%), di-n-butyl phthalate (DBP) (0.0, 0.03, 0.3, or 1.0%), di-n-hexyl phthalate (DHP)

(0.0, 0.3, 0.6, or 1.2%), or di(2-ethylhexyl) phthalate (DEHP) (0.0, 0.01, 0.1, or 0.3%).

Both male and female CD-1 mice were dosed for 7 days prior to and during a 98-day

cohabitation period. Reproductive function was evaluated during the cohabitation period

by measuring the numbers of litters per pair and of live pups per litter, pup weight, and

offspring survival. There was no apparent effect on reproductive function in the animals

exposed to DEP, despite significant effects on body weight gain and liver weight. DBP

exposure resulted in a reduction in the numbers of litters per pair and of live pups per

litter and in the proportion of pups born alive at the 1.0% amount, but not at lower dose

levels. A crossover mating trial demonstrated that female mice, but not males, were

affected by DBP, as shown by significant decreases in the percentage of fertile pairs, the

number of live pups per litter, the proportion of pups born alive, and live pup weight.

DHP in the diet resulted in dose-related adverse effects on the numbers of litters per pair

and of live pups per litter and proportion of pups born alive at 0.3, 0.6, and 1.2% DHP in

the diet. A crossover mating study demonstrated that both sexes were affected. DEHP (at

0.1 and 0.3%) caused dose-dependent decreases in fertility and in the number and the

proportion of pups born alive. A crossover mating trial showed that both sexes were

affected by exposure to DEHP. These data demonstrate the ability of the continuous

breeding protocol to discriminate the qualitative and quantitative reproductive effects of

the more and less active congeners as well as the large differences in reproductive

toxicity attributable to subtle changes in the alkyl substitution of phthalate esters.

Lawrence et al 1975 A toxicological investigation of some acute, short-term and chronic

effects of administering di-2-ethylhexyl phthalate (DEHP) and other phthalate esters.

Environ. Res.9:1-11.

Mallinckrodt-Baker, Inc. 2004. DIETHYL PHTHALTE. MSDS [Material Safety Data

Sheet] [http://www.jtbaker.com/msds/englishhtml/D3688.htm].



32

Mihovec-Grdic, et al. 2002. Phthalates in underground waters of the Zagreb area.

Croat.Med.J. 43(4): 493-497.

AIM: To determine whether and in what concentrations the underground waters, stream

waters, spring water, and tap water from the Zagreb area contain phthalates -- compounds

used as plastic softeners, which have recently been ascribed carcinogenic, mutagenic, and

teratogenic effects. METHOD: The presence of dimethyl phthalate (DMP), diethyl

phthalate (DEP), dibutyl phthalate (DBP), benzylbutyl phthalate (BBP), diethylhexyl

phthalate (DEHP), and dioctyl phthalate (DOP) was determined in a total of 96 samples

of underground waters, stream waters, and tap water from the Zagreb area between

February and June 1998. Identification and quantification of phthalates were performed

by the method of gas chromatography (GC-ECD), with a detection limit of 0.005

microg/L. RESULTS: The presence of one or more phthalates was demonstrated in 93

out of 96 (97%) water samples. The measured values ranged from 0.005 to 18.157

microg/L. Phthalates were detected in 76 out of 77 (98%) underground water samples.

The mean level of all phthalates present in the water samples was 4.879 microg/L.

Median test yielded a significantly increased level of phthalates in the underground

waters from Jakusevac (sampled in February 1998) and Trebe , which are Zagreb and

Samobor city waste dumps, as compared with other sites in the study (overall

median=3.785; chi-square=22.682; p<0.001). Phthalates were found at a mean

concentration of 3.363 microg/L in all 10 water samples from the Sava river, the major

source of the Zagreb alluvium underground waters. In case of drinking water, phthalates

were detected in 7 out of 9 (78%) samples, at a mean concentration of 0.887 microg/L.

As expected, DEHP was the most commonly detected phthalate, found in 78 (81%) water

samples. CONCLUSION: The highest phthalate concentrations were recorded in

underground waters directly related to the proximity of a waste dump. The levels of

phthalates recorded in this study were lower than those reported from other countries and

did not present a threat to human health. Environmental phthalate monitoring should be

continued and their maximum allowed concentrations should be prescribed by

regulations.

National Toxicology Program. 1995. NTP Toxicology and Carcinogenesis Studies of

Diethylphthalate (CAS No. 84-66-2) in F344/N Rats and B6C3F1 Mice (Dermal Studies)

with Dermal Initiation/ Promotion Study of Diethylphthalate and Dimethylphthalate

(CAS No. 131-11-3) in Male Swiss (CD-1(R)) Mice.

Natl.Toxicol.Program.Tech.Rep.Ser. 429: 1-286.

Diethylphthalate and dimethylphthalate are used as phthalate plasticizers, in an extensive

array of products. The chronic dermal toxicity of diethylphthalate was evaluated in male



33

and female F344/N rats and B6C3F1 mice in 2-year studies. In a series of special studies,

the tumor initiation or promotion potential of diethylphthalate or dimethylphthalate was

evaluated in male Swiss (CD-1(R)) mice by an initiation/promotion model of skin

carcinogenesis. The genetic toxicity of diethylphthalate and dimethylphthalate in

Salmonella typhimurium and cultured Chinese hamster ovary cells was also evaluated. 4-

WEEK STUDY IN F344/N RATS: Groups of 10 male and 10 female rats were dermally

administered diethylphthalate at volumes of 0, 37.5, 75, 150, or 300 &mgr;L (0, 46,

92, 184, or 369 &mgr;g) applied neat, 5 days per week for 4 weeks. All male and

female rats survived to the end of the study. No evidence of dermatotoxicity was

observed, with no adverse clinical signs observed and no effects on weight gain or feed

consumption. Relative liver weights of 300 &mgr;L males and females and 150

&mgr;L females were greater than those of controls. Relative kidney weights of 150

and 300 &mgr;L males and 150 &mgr;L females were greater than those of

controls. No other adverse effects were observed in this study. 4-WEEK STUDY IN

B6C3F1 MICE: Groups of 10 male and 10 female mice were dermally administered

diethylphthalate at volumes of 0, 12.5, 25, 50, or 100 &mgr;L (0, 15, 31, 62, or 123

&mgr;) applied neat, five days per week for 4 weeks. One control female died

before the end of the study; all other mice survived. No evidence of dermatotoxicity or

other adverse clinical signs were observed, and no clear adverse effects on weight gain or

feed consumption were seen. Absolute and relative liver weights of 25 and 100

&mgr;L females were greater than those of the controls. Based on these 4-week

study results, doses of 0, 35, and 100 &mgr;L diethylphthalate were recommended

for the 2-year mouse studies. A chronic study in male and female B6C3F1 mice at 0, 35,

and 100 &mgr;L (applied neat, once per day, 5 days per week) was started and

subsequently stopped after 32 weeks when significant body weight reductions were noted

in treated animals (males and females, 100 &mgr;L groups: 19% lower; males, 35

&mgr;L group: 12% lower; females, 35 &mgr;L group: 10% lower than

controls). Based on these body weight reductions, doses of 0, 7.5, 15, and 30

&mgr;L in 100 &mgr;L acetone were recommended for the restart of the 2-year

mouse study. 2-YEAR STUDY IN F344/N RATS: Based upon the results of the 4-week

study, doses of 0, 100, or 300 &mgr;L diethylphthalate (0, 123, or 369 &mgr;)

were chosen for the 2-year rat study. Groups of 60 male and 60 female rats received the

doses applied neat 5 days per week for 103 weeks and up to 10 animals per group were

evaluated after 15 months. Survival, Body Weights, and Clinical Findings: Survival of

dosed rats during the first 15 months was similar to that of controls. However, 2-year

survival was significantly reduced in all groups of male rats (survival probabilities,

males: 0 &mgr;L, 8%; 100 &mgr;L, 12%; and 300 &mgr;L, 12%). The

mean body weights of 300 &mgr;L males were slightly less than those of the



34

controls throughout the study. No adverse clinical signs were observed, including no

evidence of dermatotoxicity. Pathology Findings: No morphological evidence of dermal

or systemic toxicity was observed in male or female rats. Skin neoplasms were not

observed in female rats and were only rarely observed in male rats. A high incidence of

anterior pituitary adenoma occurred in all groups of male and female rats. The incidence

of anterior pituitary adenomas in the 0, 100, and 300 &mgr;L groups were: males,

39/44, 41/49, 41/49; females, 38/50, 33/49, 33/48. The incidence of this benign tumor in

control males (84%) exceeded the historical control mean incidence [feed controls,

(28.7%)] and range (12% to 60%). Anterior pituitary adenomas were considered a

primary contributing factor in the increased mortality observed in all grtor in the

increased mortality observed in all groups, regardless of treatment. A dose-related

decreasing trend in the incidence of mammary gland fibroadenomas was observed in

female rats (21/50, 12/48, 7/50). The incidence of mononuclear cell leukemia in male rats

in this study was lower than the historical incidence and may be attributable to the

shortened life span of male rats. Similarly, the incidence of interstitial cell tumors of the

testes was markedly decreased in all groups of males (4/50, 3/50, 8/50), relative to

historical control rates (90.1&percnt;; range 74&percnt;-98&percnt;). The

incidence of fatty liver degeneration was notably lower in dosed rats than in controls

(males: 26/50, 8/50, 4/51; females: 23/50, 11/50, 3/50). 2-YEAR STUDY IN B6C3F1

MICE: Groups of 60 male and 60 female mice received doses of 0, 7.5, 15, or 30

μL diethylphthalate (0, 9, 18, or 37 μ) in 100 μL acetone 5

days per week for 103 weeks with a 1 week recovery period, and up to 10 animals per

group were evaluated after 15 months. Survival, Body Weights, and Clinical Findings:

Two-year survival of dosed mice was similar to that of controls: 43/50, 41/48, 46/50, and

43/50 (males), and 41/50, 38/51, 37/49, and 36/49 (females). Mean body weights of

dosed male and female mice were similar to those of the controls throughout the study.

No adverse clinical signs were observed in mice, including no gross evidence of

dermatotoxicity. Feed consumption by male and female mice was similar to or up to 13%

greater than that by controls. Pathology Findings: No morphological evidence of dermal

toxicity was observed in male or female mice. No skin neoplasms were observed in dosed

male mice. In female mice receiving 30 μL, one squamous cell carcinoma and

one basal cell carcinoma were seen at the site of application. An increased incidence of

liver neoplasms was observed in dosed male and female mice. The incidence of

hepatocellular adenoma or carcinoma (combined) in B6C3F1 mice in the 0, 7.5, 15, and

30 μL groups were: (males) 9/50, 14/50, 14/50, and 18/50; (females) 7/50,

16/51, 19/50, and 12/50. The incidence of adenoma or carcinoma (combined) was

increased in 30 μL male mice and the incidences of adenoma and of adenoma or

carcinoma (combined) were increased in 7.5 and 15 μL females. A positive



35

dose-related trend in the incidence of adenoma or carcinoma (combined) was also

observed in male mice. The incidence of basophilic hepatic foci was increased in 15

μL male mice (0/50, 1/50, 9/50, 3/50). The increased incidence of liver

neoplasms in this study was considered equivocal because the incidence of hepatocellular

neoplasms in control and dosed males was within the historical range and because there

was no clear dose-response relationship in females. No other treatment-related findings

were observed in this study. 1-YEAR INITIATION/PROMOTION STUDY IN MALE

SWISS (CD-1®) MICE: Groups of 50 male mice were dosed dermally with

diethylphthalate or dimethylphthalate to study their effect as initiators and promoters.

Diethylphthalate and dimethylphthalate were tested as initiators with and without the

known skin tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). Diethyl

phthalate and dimethylphthalate were tested as promoters with and without the known

skin tumor initiator 7,12-dimethylbenzanthrancene (DMBA). Comparative control groups

used during the study of diethylphthalate and dimethylphthalate included: vehicle control

(acetone/acetone); initiation/promotion control (DMBA/TPA); initiator control

(DMBA/acetone); and promoter control (acetone/TPA). Based on the incidence of skin

neoplasms diagnosed histologically and the multiplicity of skin neoplasms, there was no

suggestion that either diethylphthalate or dimethylphthalate was able to initiate skin

carcinogenesis when chronically promoted by TPA. Further, there was no evidence that

either diethylphthalate or dimethylphthalate was able to promote skin carcinogenesis in

skin previously initiated with DMBA. High incidences of both squamous cell papillomas

and squamous cell carcinomas occurred among the initiation/promotion control animals

initiated with DMBA and promoted with TPA. All TPA-dosed groups had significantly

greater incidences of dermal acanthosis, ulceration, exudation, and hyperkeratosis than

controls. GENETIC TOXICOLOGY: Neither diethylphthalate (10-10,000

μ/plate) nor dimethylphthalate (33-6,666 μ/plate) induced gene

mutations in Salmonella typhimurium strains TA98, TA100, TA1535, or TA1537, with

or without rat and hamster liver S9. In cultured Chinese hamster ovary cells, both

diethylphthalate and dimethylphthalate induced sister chromatid exchanges in the

presence of S9. Neither induced sister chromatid exchanges in the absence of S9. Neither

chemical induced chromosomal aberrations, with or without S9, in cultured Chinese

hamster ovary cells. CONCLUSIONS: Under the conditions of these 2-year dermal

studies, there was no evidence of carcinogenic activity of diethylphthalate in male or

female F344/N rats receiving 100 or 300 μL. The sensitivity of the male rat

study was reduced due to low survival in all groups. There was equivocal evidence of

carcinogenic activity of diethylphthalate in male and female B6C3F1 mice based on

increased incidences of hepatocellular neoplasms, primarily adenomas. In an

initiation/promotion model of skin carcinogenesis, there was no evidence of initiating



36

activity of diethylphthalate or dimethylphthalate in male Swiss (CD-1®) mice.

Further, there was no evidence of promotion activity of diethylphthalate or

dimethylphthalate in male Swiss (CD-1®) mice. The promoting activity of TPA

following DMBA initiation was confirmed in these studies. Minor dermal acanthosis was

observed following dermal application of diethylphthalate in male and female F344/N

rats dosed for 2 years and in male Swiss (CD-1®) mice dosed for 1 year.

Synonyms: Diethylphthalate (CAS No. 84-66-2): 1,2-benzenedicarboxylic acid, diethyl

ester; DEP; diethyl 1,2-benzenedicarboxylate; diethyl o-phthalate; diethyl phthalate; ethyl

phthalate; o-benzenedicarboxylic acid diethyl ester; phthalic acid, diethyl ester; RCRA

U088 Dimethylphthalate (CAS No. 131-11-3): 1,2-benzenedicarboxylic acid, dimethyl

ester; dimethyl 1,2-benzenedicarboxylate; dimethyl benzene-o-dicarboxylate; dimethyl

benzeneorthodicarboxylate; dimethyl o-phthalate; dimethyl phthalate; DMP; FIFRA

028002; methyl phthalate; go-dimethyl phthalate; phthalic acid, dimethyl ester; phthalic

acid methyl ester; RCRA U102

Oliwiecki, et al. 1991. Contact dermatitis from spectacle frames and hearing aid

containing diethyl phthalate. Contact Dermatitis. 25(4): 264-265.

Olsen, et al. 1982. Nephrotoxicity of plasticizers investigated by 48 hours hypothermic

perfusion of dog kidneys. Scand.J.Urol.Nephrol. 16(2): 187-190.

The possible nephrotoxicity of the plasticizers diethyl phthalate and di-2-ethylhexyl

phthalate was tested in vitro using a 48 hours continuous pulsatile hypothermic perfusion

of canine kidneys with a human albumin perfusion medium. Since polysorbate 80 was

used to facilitate the solution of the plasticizers in the perfusion medium, this substance

was also tested. Six groups containing 9--15 kidneys were perfused with different

amounts of plasticizers and/or polysorbate 80 added. In perfusates containing polysorbate

80 either alone or with one of the two plasticizers, the LDH activity and the potassium

concentrations rose significantly higher than in the control group (p less than 0.001). The

kidney weight gain was also significantly greater in these groups. A "blind"

histological examination of needle biopsies by light microscopy revealed no differences

among the groups. Although the biochemical evidence of tissue damage was not tested by

re-implantation of the kidneys, we suggest that caution should be exercised in the use of

polysorbate 80 in organ perfusion systems.

O'Neil et al 2001. Merck Index 13th ed. (White House Station, NJ. Merck and Co.)

Otake et al 2004. Exposure to phthalate esters from indoor environment. J Expo Anal

Environ Epidemiol. Mar 24 [Epub ahead of print]

Phthalate esters and phosphate esters in samples of indoor air from 27 houses in the



37

Tokyo Metropolitan area were quantified using gas chromatograph/mass spectrometer

and gas chromatograph/flame photometric detector after adsorption on to charcoal and

solvent extraction. The median concentrations of diethyl phthalate, dibutyl phthalate

(DBP), butylbenzyl phthalate, dicyclohexyl phthalate and diethylhexyl phthalate were

0.10, 0.39, 0.01, 0.07 and 0.11 microg/m(3), respectively. The median concentrations of

tributyl phosphate, tris(2-chloroethyl) phosphate, triphenyl phosphate and tris(2-

butoxyethyl) phosphate were less than 0.001 microg/m(3). DBP was detected at the

highest concentration (6.18 microg/m(3)) in a new residential housing. This research

indicated that exposure to phthalate esters through inhalation of air from the indoor

environment is as important as dietary intake of phthalate esters, and can contribute to

daily intake to a much greater extent than has been assumed hitherto.Journal of Exposure

Analysis and Environmental Epidemiology advance online publication, 24 March 2004;

doi:10.1038/sj.jea.7500352

Parthasarathy, et al. 2002. Ethyl cellulose and polyethylene glycol-based sustained-

release sparfloxacin chip: an alternative therapy for advanced periodontitis. Drug

Dev.Ind.Pharm. 28(7): 849-862.

This study reports the development of a sustained-release system of sparfloxacin for use

in the treatment of periodontal disease. A sustained-release sparfloxacin device was

formulated, based on ethyl cellulose (EC) 10 cps, polyethylene glycol (PEG) 4000, and

diethyl phthalate (DEPh). It will hereafter be called the sparfloxacin chip (SRS chip). The

chip has dimensions of 10 mm length, 2 mm width, and 0.5 mm thickness. The in vitro

drug release pattern and clinical evaluation of the formulations were studied. Reports of

the short-term clinical study show that the use of the SRS chip may cause complete

eradication of the pathogenic bacteria in the periodontal pockets of patients who have

chronic generalized periodontitis. In this clinical study, the baseline and follow-up

measurements of various clinical indices, such as oral hygiene index(es), plaque index,

sulcular depth component of periodontal disease index, gingival crevicular fluid flow

measurement, and dark field microscopic examinations of oral pathogens in plaque

samples were studied. Significant improvements were observed in many parameters of

the treatment group compared with the placebo group.

Pragst, et al. 2000. Are there possibilities for the detection of chronically elevated

alcohol consumption by hair analysis? A report about the state of investigation. Forensic

Sci.Int. 107(1-3): 201-223.

The analysis of suitable ethanol markers in hair would be an advantageous tool for

chronic alcohol abuse control because of the wide diagnostic window allowed by this

specimen and the possibility of segmental investigation. Between the markers practically

used or thoroughly investigated in blood or urine, ethylglucuronide, fatty acid ethylesters,

phosphatidylethanol, acetaldehyde adducts to protein and 5-hydroxytryptophol can be

regarded as possible candidates also in hair, but preliminary data were found in the



38

literature only for ethylglucuronide and acetaldehyde modified proteins. By using

headspace gas chromatography and headspace solid phase microextraction in

combination with gas chromatography-mass spectrometry (SPME-GC/MS), in alkaline

hydrolysates of hair it was possible to determine between 17 and 135 ng/mg of ethanol

beside acetone and several other volatile compounds with slightly higher ethanol values

for alcoholics than for social drinkers and teetotalers. A part of this is ethanol only

absorbed in the hair matrix from the surrounding environment and consequently is not

applicable as a diagnostic criterion. By extraction with aqueous buffer, methanol or a

methanol/chloroform mixture and subsequent alkaline hydrolysis it was found that

another part is generated from ethylesters, which are preferentially deposited in the lipid

fraction of hair. In a specific search for ethylesters of 17 carboxylic acids by GC/MS-SIM

in most cases ethyl 4-hydroxybenzoate (0.1 to 5.9 ng/mg, a preservative in hair

cosmetics) and in four cases traces of indolylacetic acid ethylester were found.

Furthermore, diethyl phthalate (a softening agent, present also in many cosmetic

products) was identified in the hair of alcoholics as well as of children. As potential

markers of alcohol intake, ethyl palmitate, ethyl stearate and ethyl oleate were detected in

hair samples of alcoholics by headspace SPME-GC/MS of the chloroform/methanol

extracts.

Project 112 2004. Project SHAD Glossary. DeploymentLink [DoD].

[http://deploymentlink.osd.mil/current_issues/shad/shad_glossary.shtml]

Project 65-17 2004. Fearless Johnny. DeploymentLink [DoD].

[http://deploymentlink.osd.mil/pdfs/fearless_johnny.pdf]

RTECS [Registry of Toxic Effects of Chemical Substances] 2004. Diethyl Phthalate.

http://www.cdc.gov/niosh/rtecs/ti100590.html.

Saarma, et al. 2003. Heat shock protein synthesis is induced by diethyl phthalate but not

by di(2-ethylhexyl) phthalate in radish (Raphanus sativus). J.Plant Physiol. 160(9): 1001-

1010.

The toxicity and effects on protein synthesis of the phthalate esters diethyl phthalate

(DEP) and di(2-ethylhexyl) phthalate (DEHP) was studied in radish seedings (Raphanus

sativus cv. Koopenhaminan tori). Phthalate esters are a class of commercially important

compounds used mainly as plasticizers in high molecular-weight polymers such as many

plastics. They can enter soil through various routes and can affect plant growth and

development. First the effect of DEP and DEHP on the growth of radish seedings was

determined in an aqueous medium. It was found that DEP, but not DEHP, caused

retardation of growth in radish. A further investigation on protein synthesis during DEP-



39

stress was executed by in vivo protein labeling combined with two-dimensional gel

electrophoresis (2D-PAGE). For comparisons with known stress-induced proteins a

similar experiment was done with heat shock, and the induced heat shock proteins (HSPs)

were compared with those of DEP-stress. The results showed that certain HSPs can be

used as an indicator of DEP-stress, although the synthesis of most HSPs was not affected

by DEP. DEP also elicited the synthesis of numerous proteins found only in DEP-treated

roots. The toxic effect of phthalate esters and the roles of the induced proteins are

discussed.

Schulsinger, et al. 1980. Polyvinyl chloride dermatitis not caused by phthalates. Contact

Dermatitis. 6(7): 477-480.

Seven cases of contact dermatitis in children due to identification bracelets made of

polyvinyl chloride plastic are reported. Patch tests with the bracelets were negative in the

five cases tested. It is concluded that the reactions were irritant due to some unknown

chemical in the bracelets. The most widely used plasticizers in PVC, phthalates, must

have very low sensitizing properties, as only one positive patch test was found in 1532

patch tests with phthalate mix, performed as a joint study by the International Contact

Dermatitis Research Group.

Scott, et al. 1987. In vitro absorption of some o-phthalate diesters through human and rat

skin. Environ.Health Perspect. 74: 223-227.

The absorption of undiluted phthalate diesters [dimethyl phthalate (DMP),

diethylphthalate (DEP), dibutyl phthalate (DBP) and di-(2-ethylhexyl)phthalate (DEHP)]

has been measured in vitro through human and rat epidermal membranes. Epidermal

membranes were set up in glass diffusion cells and their permeability to tritiated water

measured to establish the integrity of the skin before the phthalate esters were applied to

the epidermal surface. Absorption rates for each phthalate ester were determined and a

second tritiated water permeability assessment made to quantify any irreversible

alterations in barrier function due to contact with the esters. Rat skin was consistently

more permeable to phthalate esters than the human skin. As the esters became more

lipophilic and less hydrophilic, the rate of absorption was reduced. Contact with the

esters caused little change in the barrier properties of human skin, but caused marked

increases in the permeability to water of rat skin. Although differences were noted

between species, the absolute rates of absorption measured indicate that the phthalate

esters are slowly absorbed through both human and rat skin.

Seed. 1982. Mutagenic activity of phthalate esters in bacterial liquid suspension assays.

Environ.Health Perspect. 45: 111-114.

The mutagenic activities of several phthalate esters have been evaluated in an 8-



40

azaguanine resistance assay in Salmonella typhimurium. Three phthalate esters were

found to be mutagenic: dimethyl phthalate, diethyl phthalate and di-n-butyl phthalate. A

number of other phthalate esters were not found to be mutagenic, including di(2-

ethylhexyl) phthalate, di-n-octyl phthalate, diallyl phthalate, diisobutyl phthalate and

diisodecyl phthalate. A metabolite of di(2-ethylhexyl) phthalate, 2-ethylhexanol, was also

noted to be mutagenic. The mutagenic activity of this agent and others in this series was

dose dependent but weak. No dose-response curve exceeded more than 3.5 times

background at maximally testable concentrations. A liquid suspension histidine reversion

assay of dimethyl phthalate showed levels of mutagenic activity similar to that observed

in the azaguanine resistance assay. The data suggest a need for further investigation of

the mutagenic potential of these agents in other assay systems.

Silva, et al. 2004. Urinary levels of seven phthalate metabolites in the U.S. population

from the National Health and Nutrition Examination Survey (NHANES) 1999-2000.

Environ.Health Perspect. 112(3): 331-338.

We measured the urinary monoester metabolites of seven commonly used phthalates in

approximately 2,540 samples collected from participants of the National Health and

Nutrition Examination Survey (NHANES), 1999-2000, who were greater than or equal to

6 years of age. We found detectable levels of metabolites monoethyl phthalate (MEP),

monobutyl phthalate (MBP), monobenzyl phthalate (MBzP), and mono-(2-ethylhexyl)

phthalate (MEHP) in > 75% of the samples, suggesting widespread exposure in the

United States to diethyl phthalate, dibutyl phthalate or diisobutylphthalate, benzylbutyl

phthalate, and di-(2-ethylhexyl) phthalate, respectively. We infrequently detected

monoisononyl phthalate, mono-cyclohexyl phthalate, and mono-n-octyl phthalate,

suggesting that human exposures to di-isononyl phthalate, dioctylphthalate, and

dicyclohexyl phthalate, respectively, are lower than those listed above, or the pathways,

routes of exposure, or pharmacokinetic factors such as absorption, distribution,

metabolism, and elimination are different. Non-Hispanic blacks had significantly higher

concentrations of MEP than did Mexican Americans and non-Hispanic whites. Compared

with adolescents and adults, children had significantly higher levels of MBP, MBzP, and

MEHP but had significantly lower concentrations of MEP. Females had significantly

higher concentrations of MEP and MBzP than did males, but similar MEHP levels. Of

particular interest, females of all ages had significantly higher concentrations of the

reproductive toxicant MBP than did males of all ages; however, women of reproductive

age (i.e., 20-39 years of age) had concentrations similar to adolescent girls and women 40

years of age. These population data on exposure to phthalates will serve an important role

in public health by helping to set research priorities and by establishing a nationally

representative baseline of exposure with which population levels can be compared.



41

Singh, et al. 1975. Maternal-fetal transfer of 14C-di-2-ethylhexyl phthalate and 14C-

diethyl phthalate in rats. J.Pharm.Sci. 64(8): 1347-1350.

14C-Di-2-ethylhexyl and 14C-diethyl phthalates were administered intraperitoneally to

pregnant rats on either Day 5 or 10 of gestation. Rats were sacrificed at 24-hr intervals

starting on Days 8 and 11, respectively; maternal blood, fetal tissue, amniotic fluid, and

placentas (whenever possible) were obtained. The 14C-activity of each sample was

determined by scintillation counting. It was found that both diesters and/or their

metabolic products were present in each of these compartments throughout the gestation

period, thus suggesting that the embryo-fetal toxicity and teratogenesis reported

previously could be the results of a direct effect of the compound (or its metabolites)

upon developing embryonic tissue. Additionally, the reduction in concentration of 14C

from these tissues as a function of time was found to fit a first-order excretion curve.

From this model curve, the half-life for both compounds was calculated; the average was

about 2.33 days for di-2-ethylhexyl phthalate and 2.22 days for diethyl phthalate.

Singh et al 1972. Teratogenicity of phthalate esters in rats. J Pharm Sci. 61(1):51-5.

Smirnov, et al. 1983. [Experience with the diagnosis and treatment of scabies]. Voen

Med.Zh. (4)(4): 60-61.

Sonde, et al. 2000. Simultaneous administration of diethylphthalate and ethyl alcohol and

its toxicity in male Sprague-Dawley rats. Toxicology. 147(1): 23-31.

Phthalate esters have been implicated as xenoestrogens. One among them is di-

ethylphthalate (DEP), which is used as plasticizer, detergent base, and binder in incense

sticks and after-shave lotions. DEP is one of the contaminants of freshwater and marine

ecosystems. Incense stick workers are occupationally exposed to DEP and some workers

are chronic alcoholics. Therefore, a study was undertaken to evaluate the interactive

toxicity of DEP with ethyl alcohol (EtOH) in young male Sprague-Dawley rats. The rats

were given 50 ppm DEP (w/v), 5% EtOH (v/v) and a combined dose of 50 ppm DEP

(w/v)+EtOH (5% v/v) in water ad libitum for a period of 120 days and were maintained

on normal diet. Control animals received normal diet and plain water. During the

treatment rats were weighed every week and water consumption per day was measured.

After the completion of treatment, liver weight/body weight, liver weight, body weight,

serum enzymes and other biochemical parameters were assessed. It was found that there

was no significant change observed in body weight, liver weight, liver weight/body

weight and water consumption. It was observed that there was a significant decrease in

liver aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels in



42

EtOH, DEP and EtOH+DEP treated rats in the order of EtOH>DEP>EtOH+DEP

as compared with control. Serum AST, ALT, acid phosphatase (ACP), alkaline

phosphatase (ALP), succinate dehydrogenase (SDH) and liver ACP showed significant

increase in DEP and EtOH+DEP treated rats in the order of DEP>EtOH+DEP as

compared with control and EtOH treated rats. On the contrary, there was no significant

change in liver ALP levels in treated rats. There was significant increase in liver SDH,

glycogen, total triglyceride, total cholesterol and lipid peroxidation in DEP and

EtOH+DEP treated rats, but no significant changes in the serum SDH, glucose and total

triglyceride levels. Serum total cholesterol levels in DEP and EtOH+DEP treated rats

were significantly high as compared to control and EtOH treated rats. These results show

that there is no interaction of DEP with EtOH but DEP alone leads to severe impairment

of lipid metabolism coupled with toxic injury to the liver as evident from significantly

altered lipid and enzyme levels in the liver and serum. Long term simultaneous exposure

to DEP and EtOH may have severe implications for humans who are occupationally

exposed to these two xenobiotics.

Sugamori, et al. 1989. Microencapsulation of pancreatic islets in a water insoluble

polyacrylate. ASAIO Trans. 35(4): 791-799.

Rat pancreatic islets were encapsulated in a water insoluble polyacrylate (Eudragit RL), a

model polymer, by coaxial extrusion and interfacial precipitation. Despite exposure to

organic solvents and nonsolvents (diethyl phthalate, corn oil, and mineral oil) and to

shear, the islets survived encapsulation. They continued to secrete insulin into the tissue

culture medium and responded to glucose in both static glucose challenges and perifusion

assays as well and as long as control islets which were not encapsulated, but were

maintained in tissue culture alongside the encapsulated islets. Unfortunately, there was a

great deal of variability in the performance of all islets studied, making unequivocal

conclusions difficult. Some encapsulated islets survived more than 140 days in vitro and

histologically appeared healthy. However, there appeared to be a general deterioration in

insulin secretion capacity following prolonged culture in all islets, with corresponding

changes (e.g., central necrosis) visible by microscopy. Although Eudragit RL is not

practical as an encapsulation polymer, this study was useful in demonstrating that islets

may be encapsulated in materials other than alginate-polylysine, and ultimately in

materials that may have a more optimum blend of the desired properties:

biocompatibility, permselectivity, and mechanical durability.

Sung, et al. 2003. Effects and toxicity of phthalate esters to hemocytes of giant

freshwater prawn, Macrobrachium rosenbergii. Aquat.Toxicol. 64(1): 25-37.

Phthalate esters (PAEs) have been considered as environmental pollutants and have been

subject to control in the United States of America and Japan. The aim of this study was to



43

investigate the effects and toxicity of eight PAEs to hemocytes and the defense functions

of giant freshwater prawn (Macrobrachium rosenbergii), including hemocytic adhesion,

pseudopodia formation, phenoloxidase (PO) activity, and superoxide anion (O(2)(-))

production, by means of in vitro exposure experiments. After hemocytes were treated

separately with eight PAEs at concentrations of 100 microg/ml, the results showed that

two PAEs (dipropyl phthalate, DPrP and diethyl phthalate, DEP) increased cells with

pseudopodia formation, but decreased adhesive cells; reduction in the percentages of both

pseudopodia formation and adhesive cells were detected in the dihexyl phthalate (DHP)

and diphenyl phthalate (DPP) experiment groups; and di-(2-ethyl hexyl) phthalate

(DEHP) decreased pseudopodia formation, but did not affect the adhesion. In addition,

both PO activity and O(2)(-) production were decreased after hemocytes were treated

with five PAEs (benzyl butyl phthalate (BBP), di-n-butyl phthalate (DBP), DEP, DHP

and DPrP), respectively. At the same time, microscopy showed that both DPrP and DHP

altered morphology of the cell nucleus and led to the presence of vacuoles in cytosol of

hemocytes. Using the annexin assay, and after analysis of DNA fragmentation and

transmission electron microscopy (TEM), it was found that hemocytes exposed to DHP

and DPrP for more than 10 min would primarily die via apoptosis, the fatality correlates

with increasing treatment time; and hemocytes treated with either BBP, dicyclohexyl

phthalate (DCP), DEP or DPP would primarily die via necrosis. According to these

results, we suggest that all eight PAEs examined could damage hemocytes and further

influence the defense mechanism of prawns. This study reveals an important precaution

for prawn cultivation.

Teghtsoonian, et al. 1978. Invariance of odor strength with sniff vigor: an olfactory

analogue to size constancy. J.Exp.Psychol.Hum.Percept.Perform. 4(1): 144-152.

Previous evidence has shown that detection threshold in humans and olfactory neural

discharge rate in animal preparations both depend on flow rate of odorous vapor. But no

data have been reported that show the effects of flow rate in humans on perceived odor

strength at suprathreshold intensities. Subjects learned to inspire at two flow rates, one

twice as great as the other, by adjusting (on a cathode ray tube) the transduced trace of a

sniff-produced pressure change to match either of two target contours. They then made

magnitude estimations of odor strength, while producing either weak or strong sniffs, for

odorants presented over a wide range of concentrations via a specially designed sniff-

bottle system. The odorant, diluted in diethyl phthalate, was n-butanol in two experiments

and n-amyl acetate in two others. Subject-controlled flow rate had no effect on odor

strength for either odorant. There was an apparent contradiction between these data and

those on neural discharge rate that may, however, be resolved by adopting an odor

constancy model: When sniff intensity varies during the olfactory exploration of an odor

source, information about the rate at which odorant molecules are established at the



44

receptor site is combined with information about sniff vigor so that the resulting percept

is of invariant odor strength.

Teranishi, et al. 1980. The effects of phthalate esters on fibroblasts in primary culture.

Toxicol.Lett. 6(1): 11-15.

The toxicity of butylbenzyl phthalate(BLP), di-n-heptyl phthalate (DNHP) and n-butyl

lauryl phthalate (BLP) to fibroblasts from newborn rat cerebellum in primary culture was

significant at concentrations of 7.0, 2.7, and 5.0 x 10(-4) M, respectively. The toxicity of

di-methoxyethyl phthalate(DMEP), butyl phthalyl butyl glycolate(BPBG), di-n-octyl

phthalate(DNOP), and di-(2-ethylhexyl) phthalate(DEHP) was not significant. Phthalic

acid and potassium hydrogen phthalate (K-phthalate) were the least toxic to fibroblasts.

Comparison of the toxicity to fibroblasts of five phthalate esters of normal series showed

that dimethyl phthalate(DMP) < diethyl phthalate(DEP) < di-n-butyl

phthalate(DNBP) > DNHP > DNOP.

US EPA. 2004 Integrated Risk Information System (IRIS) [

http://www.epa.gov/iris/subst/0226.htm]

WHO [World Health Organization] 2003 Concise International Chemical Assessment

Document 52 DIETHYL PHTHALATE .

[//www.inchem.org/documents/cicads/cicads/cicad52.htm]

Zaitsev, et al. 1990. [Health-related regulation of diethyl phthalate, di-n-hexyl phthalate

and dialkyl phthalate 810 in water]. Gig.Sanit. (9)(9): 26-28.

On the basis of studies of hygienic regulation of diethylphthalate (DEP), di-n-

hexylphthalate (DHP) and dialkylphthalate-810 (DAP-810) in the water medium) it has

been found out that the compounds are highly persistent in the water medium, are of low

toxicity (LD50 from 10.3 up to 33 g/kg and more for white rats), belong to the third and

fourth (DHP and DAP-810) classes of danger. The threshold concentrations of DEP,

DHP, DAP-810 according to the organoleptic water properties and sanitary regimen of

water reservoirs were determined on the level of 1, 0.46, 0.3 and 0.1, 1, and 1 mg/l

respectively. DEP has moderately expressed cumulative properties while in DHP and

DAP-810 they are clearly expressed. No specific effect was observed in the compounds.

MACs for DEP, DHP and DAP-810 in the water medium are recommended on the level

of 0.1, 0.5 and 0.3 mg/l according to the general toxic and organoleptic indices of

harmfulness.

Zou, et al. 1997. Effects of estrogenic xenobiotics on molting of the water flea, Daphnia

magna. Ecotoxicol.Environ.Saf. 38(3): 281-285.



45

The effects of five xenobiotics, 2,4,5-trichloribiphenyl (PCB29), the polychlorinated

biphenyl (PCB) Aroclor 1242, diethyl phthalate, lindane, and 4-octylphenol, on molting

of Daphnia magna were investigated. All except PCB29 are known to have unexpected

estrogenicity in vertebrates. Daphnids exposed to PCB29, Aroclor 1242, and diethyl

phthalate took significantly more time to complete four molts than did the controls. The

inhibitory effects of these ortho-chlorinated PCBs suggest that certain structural features,

most probably including ortho-chlorination, are related to the ability of a PCB to affect

molting. Agents with multicyclic structures, such as PCBs, are more effective in

inhibiting molting than are single-ringed xenobiotics, such as diethyl phthalate, which

suggests that hydrophobicity may be a requirement for binding to the ecdysteroid

receptor. These molt-inhibiting agents with multiple rings appear to bear more structural

resemblance to the steroidal molting hormones of arthropods, the ecdysteroids, than do

the single-ringed ones. While the possibility of alternative mechanisms, such as

impairment of ecdysteroidogenesis exists, the results obtained herein support the

hypothesis that some xenobiotics which disrupt endocrine processes in vertebrates can

also interfere with the hormonally regulated molting process in arthropods through acting

as antagonists of endogenous ecdysteroids by binding to and thereby blocking the

ecdysteroid receptor.

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