This report contains the collective views of an international group of
    experts and does not necessarily represent the decisions or the stated
    policy of the United Nations Environment Programme, the International
    Labour Organisation, or the World Health Organization.

    Published under the joint sponsorship of
    the United Nations Environment Programme,
    the International Labour Organisation,
    and the World Health Organization

    First draft prepared by Dr P. Lundberg, Dr. J. Hogberg,
    and Dr P. Garberg, National Institute of Occupational
    Health, Sweden, Dr I. Lundberg, Karolinska
    Hospital, Sweden, and Dr S. Dobson and Mr. P. Howe,
    Institute of Terrestrial Ecology, United Kingdom

    World Health Orgnization
    Geneva, 1992

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    toxicology. Other activities carried out by the IPCS include the
    development of know-how for coping with chemical accidents,
    coordination of laboratory testing and epidemiological studies, and
    promotion of research on the mechanisms of the biological action of

    WHO Library Cataloguing in Publication Data

    Diethylhexyl phthalate.

        (Environmental health criteria ; 131)

        1.Diethylhexyl phthalate - adverse effects  2.Diethylhexyl
          phthalate - toxicity 3.Environmental exposure   I.Series

        ISBN 92 4 157131 4        (NLM Classification: QV 612)
        ISSN 0250-863X

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    1.   SUMMARY

         1.1   Identity, physical and chemical properties, and
               analytical methods
         1.2   Sources of human and environmental exposure
         1.3   Environmental transport, distribution, and
         1.4   Environmental levels and human exposure
         1.5   Kinetics and metabolism
         1.6   Effects on laboratory mammals and  in vitro test
         1.7   Effects on humans
         1.8   Effects on other organisms in the laboratory
               and field
         1.9   Evaluation


         2.1   Identity
         2.2   Physical and chemical properties
         2.3   Conversion factors
         2.4   Analytical methods


         3.1   Natural occurrence
         3.2   Anthropogenic sources
               3.2.1   Production levels
               3.2.2   Uses
               3.2.3   Disposal of plasticized products


         4.1   Environmental transport and distribution
               4.1.1   Transport in air
               4.1.2   Transport in soil and sediment
               4.1.3   Transport in water
               4.1.4   Transport between media
         4.2   Biotransformation
               4.2.1   Abiotic degradation
               4.2.2   Biodegradation
                Aerobic degradation
                Anaerobic degradation

               4.2.3   Bioaccumulation
                Model ecosystems
                Aquatic invertebrates


         5.1   Environmental levels
               5.1.1   Air
               5.1.2   Precipitation
               5.1.3   Water
               5.1.4   Sediment
               5.1.5   Soil
               5.1.6   Food
               5.1.7   Aquatic organisms
               5.1.8   Terrestrial organisms
         5.2   General population exposure
         5.3   Occupational exposure during manufacture,
               formulation or use


         6.1   Absorption
               6.1.1   Inhalation
               6.1.2   Dermal
               6.1.3   Oral
               6.1.4   Intraperitoneal
         6.2   Distribution
         6.3   Metabolism
         6.4   Elimination and excretion
         6.5   Retention and turnover
               6.5.1   Half-life and body burden
               6.5.2   Indicator media


         7.1   Single exposure
         7.2   Short-term exposure
         7.3   Long-term exposure
         7.4   Skin and eye irritation; sensitization
         7.5   Reproduction, embryotoxicity, and teratogenicity
               7.5.1   Reproduction
               7.5.2   Embryotoxicity and teratogenicity

         7.6   Mutagenicity and related end-points
               7.6.1   Mutation
                Mammalian cells
               7.6.2   DNA damage
               7.6.3   DNA binding
               7.6.4   Chromosomal effects
               7.6.5   Cell transformation
               7.6.6    In vivo effects
         7.7   Carcinogenicity
         7.8   Special studies
         7.9   Mechanisms of hepatotoxicity


         8.1   General population exposure
         8.2   Occupational exposure


         9.1   Toxicity to microorganisms
         9.2   Toxicity to aquatic organisms
               9.2.1   Invertebrates
               9.2.2   Fish
               9.2.3   Amphibians
         9.3   Toxicity to terrestrial organisms
               9.3.1   Plants
               9.3.2   Earthworms
               9.3.3   Insects
               9.3.4   Birds


         10.1  Evaluation of human health risks
               10.1.1  Exposure levels
               10.1.2  Toxic effects
               10.1.3  Conclusion
         10.2  Evaluation of effects on the environment
               10.2.1  Exposure levels
               10.2.2  Toxic effects
               10.2.3  Conclusion









    Dr D. Anderson, British Industrial Biological Research Association,
       Carshalton, Surrey, United Kingdom

    Dr R. Cattley, Department of Experimental Pathology and Toxicology,
       Chemical Industry Institute of Toxicology, Research Triangle Park,
       North Carolina, USA

    Dr U. Chantharaksri, Department of Pharmacology, Mahidol University,
       Bangkok, Thailand

    Dr S.D. Gangolli, British Industrial Biological Research Association,
       Carshalton, Surrey, United Kingdom

    Dr J. Högberg, Department of Toxicology, National Institute of
       Occupational Health, Solna, Sweden

    Mr P. Howe, Institute of Terrestrial Ecology, Monks Wood Experimental
       Station, Abbots Ripton, Huntingdon, United Kingdom

    Dr F. Matsumura, Toxic Substances Program, Department of Environmental
       Toxicology, University of California, Davis, California, USA

    Dr S. Oishi, Department of Toxicology, Metropolitan Research
       Laboratory of Public Health, Tokyo, Japan

    Dr C.-N. Ong, Department of Community, Occupational and Family
       Medicine, National University of Singapore, Singapore  (Joint

    Professor G. Pliss, Laboratory for Chemical Carcinogenic Agents, N.N.
       Petrov Research Institute of Oncology, Leningrad, USSR

    Professor Y.-L. Wang, Department of Occupational Health, School of
       Public Health, Shanghai Medical University, Shanghai, China

    Mr G. Welter, Federal Environmental Protection Agency, Berlin, Germany

     Representatives of other intergovernmental organizations

    Dr M. De Smedt, Commission of the European Communities, Luxembourg

     Representatives of non-governmental organizations

    Dr C. Elcombe, European Chemical Industry Ecology and Toxicology
       Centre, Brussels, Belgium

    Dr B. Lake, Conseil Européen des Fédérations de l'Industrie chimique
       (CEFIC), Brussels, Belgium


    Dr B.-H. Chen, International Programme on Chemical Safety, World
       Health Organization, Geneva, Switzerland  (Secretary)

    Dr P. Lundberg, Department of Toxicology, National Institute of
       Occupational Health, Solna, Sweden  (Joint Rapporteur)

    Dr D. McGregor, International Agency for Research on Cancer, World
       Health Organization, Lyon, France


       Every effort has been made to present information in the criteria
    monographs as accurately as possible without unduly delaying their
    publication.  In the interest of all users of the environmental health
    criteria monographs, readers are kindly requested to communicate any
    errors that may have occurred to the Manager of the International
    Programme on Chemical Safety, World Health Organization, Geneva,
    Switzerland, in order that they may be included in corrigenda.

                                    * * *

       A detailed data profile and a legal file can be obtained from the
    International Register of Potentially Toxic Chemicals, Palais des
    Nations, 1211 Geneva 10, Switzerland (Telephone No. 7988400 or


       A WHO Task Group on Environmental Health Criteria for Diethylhexyl
    Phthalate (DEHP) met at the British Industrial Biological Research
    Association (BIBRA), Carshalton, Surrey, United Kingdom, from 3 to 7
    June 1991.  Dr S.D. Gangolli opened the meeting on behalf of BIBRA.
    Dr B.-H. Chen, IPCS, welcomed the participants on behalf of the
    Manager, IPCS, and the three IPCS cooperating organizations
    (UNEP/ILO/WHO).  The Task Group reviewed and revised the draft
    criteria monograph and made an evaluation of the risks for human
    health and the environment from exposure to DEHP.

       The first draft of this monograph was prepared by Dr P. Lundberg,
    Dr J. Högberg, and Dr P. Garberg of the National Institute of
    Occupational Health, Sweden, Dr I. Lundberg of Karolinska Hospital,
    Sweden, and Dr S. Dobson and Mr P. Howe of the Institute of
    Terrestrial Ecology, Monks Wood Experimental Station, United Kingdom. 
    The second draft was prepared by Dr P. Lundberg incorporating comments
    received following the circulation of the first draft to the IPCS
    Contact Points for Environmental Health Criteria monographs.
    Particularly valuable comments on the draft were made by the European
    Chemical Industry Ecology and Toxicology Centre (ECETOC), the
    International Agency for Research on Cancer (IARC), the Toxicology
    Division, Exxon Biomedical Sciences, and the Conseil European des
    Federations de L'industrie Chimique (CEFIC).

       Dr B.-H. Chen and Dr P.G. Jenkins, both members of the IPCS Central
    Unit, were responsible for the overall scientific content and
    technical editing, respectively.

       The efforts of all who helped in the preparation and finalization
    of the document are gratefully acknowledged.

                                    * * *

       Financial support for this Task Group was provided by the United
    Kingdom Department of Health as part of its contributions to the IPCS.


    DEHP           diethylhexyl phthalate
    DBP            di- n-butyl phthalate
    DiBP           di-iso-butyl phthalate
    ECETOC         European Chemical Industry Ecology and Toxicology
    ECMO           extracorporeal membrane oxidation
    HPLC           high-performance liquid chromatography
    MEHP           monoethylhexyl phthalate
    NDMA            N-dimethylnitrosamine
    NOEL           no-observed-effect level
    PVC            polyvinyl chloride
    SHE            Syrian hamster embryo
    SPF            specific pathogen free
    UDS            unscheduled DNA synthesis
    US ATSDR       US Agency for Toxic Substances and Disease Registry
    US FDA         US Food and Drug Administration

    1.  SUMMARY

    1.1  Identity, physical and chemical properties, and analytical

         Di(2-ethylhexyl)phthalate (DEHP) is a benzenedicarboxylic acid
    ester which at room temperature is a colourless to yellow oily liquid.
    Its solubility in water is low (0.3-0.4 mg/litre), and is even lower
    in salt water. It is miscible with most common organic solvents. The
    volatility of DEHP is relatively low (8.6 x 10-4 Pa).

         Many sampling and analytical methods have been developed for the
    determination of DEHP in different media. Sensitive methods, such as
    gas chromatography, high-performance liquid chromatography, and mass
    spectrometry are being used increasingly. Analysis of low
    concentrations of DEHP is complicated by contamination from plastic
    equipment during sampling and analysis.

    1.2  Sources of human and environmental exposure

         Almost all the DEHP present in the environment arises from
    anthropogenic sources rather than from natural ones.

         The worldwide production of DEHP has been increasing during
    recent decades and at present amounts to about 1 x 106 tonnes per
    year. One third of the total production is in the USA and one third in

         DEHP is the most widely used plasticizer (comprising 50% of all
    phthalate ester plasticizers) that softens resins. It may account for
    40% (w/w) or more of the plastic. DEHP is used for making the
    polyvinyl chloride (PVC) utilized in building, construction and
    packaging, and for medical device components. Smaller amounts are used
    in industrial paints and as a dielectric fluid in condensers.

         Discarded plasticized products may be disposed of either by
    incineration or via dumping in a landfill site. During incineration at
    a low temperature, a large percentage of the DEHP may be lost to the
    atmosphere. The environmental fate of DEHP in landfill sites has not
    been well studied and no definite conclusions can be reached.

    1.3  Environmental transport, distribution, and transformation

         Transport in the air is the major route by which phthalates enter
    the environment. From the atmosphere DEHP either falls or is washed
    out via rainfall.

         DEHP has a high octanol-water partition coefficient, so the
    equilibrium between water and an organic-rich soil or sediment is in
    favour of the soil or sediment. It is readily adsorbed by organic soil

         Although the solubility of DEHP in water is low, the amount
    present in surface water may be higher due to adsorption onto organic
    particles and interaction with dissolved organic matter. It is
    adsorbed particularly by small particles, and adsorption is enhanced
    in salt water.

         Atmospheric photodegradation of DEHP is rapid, but its chemical
    hydrolysis in the environment is practically non-existent.

         Aerobic degradation has been found to be carried out by several
    soil microorganisms. However, the microbial degradation of DEHP in the
    environment has been reported to be slow. The biodegradation pathway
    begins with hydrolysis to the mono-ester, which is then converted to
    phthalic acid. The ring-opening degradation to pyruvate and succinate
    and then to CO2 and H2O is similar to the metabolic pathway of
    benzoic acid. The aerobic degradation is temperature dependent. Below
    10 °C little degradation takes place. At higher temperatures
    biodegradation proceeds in the upper layer of the soil, but it is
    virtually non-existent deeper down where conditions are anaerobic.
    Anaerobic degradation, if it exists, is very much slower than aerobic

         DEHP is highly lipophilic and moderately persistent. The degree
    of bioaccumulation depends on the capability of an organism to
    metabolize DEHP. It has been shown to accumulate to a high degree in
    a variety of aquatic invertebrates, fish, and amphibians.

         When DEHP was applied to plant leaves, there was little loss over
    a 15-day period. Uptake by plants from soil or sewage sludge was found
    to be low.

    1.4  Environmental levels and human exposure

         DEHP exists widely in the environment and is found in most
    samples, including air, precipitation, water, sediment, soil, and
    biota. Levels are generally highest in industrialized areas.

         DEHP concentrations of up to 300 ng/m3 have been found in urban
    and polluted air. Levels of between 0.5 and 5 ng/m3 have been
    reported in the air of oceanic areas, and the rainfall in these areas
    contained up to about 200 ng/litre. Precipitation samples from an area
    close to a plasticizer production plant indicated that the rate of dry
    deposition was 0.7 to 4.7 µg/m2 per day.

         In rivers and lakes the concentration of DEHP has been found to
    be up to 4 µg/litre, highest levels being associated with industrial
    effluent discharge points. The concentration in the sea is less than
    1 µg/litre, highest levels being in estuaries.

         Due to its hydrophobic character, DEHP is readily absorbed to
    soil, sediment, and particulate matter. River sediment levels of up to
    70 mg/kg (dry weight) have been reported, and these have reached 1480
    mg/kg (dry weight) near discharge points.

         The concentration of DEHP in biota varies from less than 1 to
    7000 µg/kg. It has been found in various types of food, such as fish,
    shellfish, eggs, and cheese. The estimated average exposure was around
    300 µg/person per day in the USA in 1974 and 20 µg/person per day in
    the United Kingdom in 1986.

         Blood transfusions and other medical treatment using plastic
    devices may lead to involuntary human exposure to DEHP. Levels from
    13.4 to 91.5 mg/kg (dry weight) in lung tissue have been detected in

         The few data available indicate that workplace concentrations of
    DEHP are usually below 1 mg/m3.

    1.5  Kinetics and metabolism

         Available data on oral administration indicate that DEHP is
    hydrolysed in the gut by pancreatic lipase. The metabolites formed,
    i.e. mono(2-ethylhexyl)phthalate (MEHP) and 2-ethyl-hexanol, are
    rapidly absorbed. When 14C-labelled DEHP (2.9 mg/kg) was given
    orally to rats, more than 50% was recovered in the urine or bile. The
    bioavailability of an oral dose of DEHP seems to be higher in young
    rats than in older ones.

         When administered orally, DEHP is extensively hydrolysed in the
    gut in certain animals, e.g., rats, and is mainly distributed as
    monoethylhexyl phthalate (MEHP). However, hydrolysis occurs to a much
    lesser extent in primates and humans. MEHP binds to plasma proteins.
    The liver seems to be the major organ for the metabolism of MEHP and
    2-ethylhexanol. Several further metabolites have been identified,
    omega- and omega-1-oxidation being the major metabolic pathways. One
    or several of the products of omega-oxidation may be further
    metabolized by ß-oxidation. Non-linear kinetics have been observed for
    the omega-oxidation. DEHP metabolism shows considerable species
    differences; e.g., the omega-oxidation pathway is less extensive in
    humans than in rats.

         Almost 100% of an oral dose of DEHP (2.9 mg/kg) was recovered in
    rat faeces and urine after a week. Bile and urine are the major
    excretory pathways. In a human study, 15-25% of an oral dose
    (0.45 mg/kg) of DEHP was excreted as MEHP, and oxidized metabolites
    constituted a major portion of the excretion products.

    1.6  Effects on laboratory mammals and in vitro test systems

         The oral LD50 for DEHP is about 25-34 g/kg, depending on the
    species, but the value for MEHP is lower. In feeding studies on rats
    and mice, DEHP dosages greater than 3 g/kg per day caused deaths
    within 90 days, and a level of 0.4 g/kg per day reduced weight gain
    within a few days. In other studies, 6.3-12.5 g/kg diet caused a body
    weight reduction.

         Hepatomegaly and increased relative kidney weights have been
    observed in treated animals in long-term studies. In one study, there
    were also hypertrophic cells in the anterior pituitary.

         Several studies have shown testicular atrophy, evident within a
    few days, related to DEHP administration (dietary levels of 10-20 g
    DEHP/kg). Younger rats seem to be more susceptible than older ones,
    and rats and mice seem to be more sensitive than marmosets and
    hamsters. Reversibility of the atrophy has been observed. MEHP has
    toxic effects on Sertoli cells  in vitro. DEHP, as well as MEHP,
    shows teratogenic properties. Malformations were observed at dietary
    levels of 0.5-2 g/kg in mice, and embryotoxic effects were observed at
    dietary levels greater than 10 g/kg.

         Tests for mutagenicity and related end-points have been negative
    in most studies. DEHP may induce cellular transformation, and it has
    been shown to be carcinogenic at doses of 6 and 12 g DEHP/kg diet in
    rats and 3 and 6 g/kg diet in mice. There was a dose-related increase
    in hepatocellular tumours in both sexes of both species. The induction
    of hepatic peroxisome proliferation and cell replication is strongly
    associated with the liver carci-nogenic effect of certain non-
    genotoxic carcinogens including DEHP. However, marked differences have
    been observed among animal species with respect to DEHP-induced
    peroxisome proliferation.

         In contrast to rat hepatocytes, DEHP metabolites do not produce
    peroxisome proliferation in cultured human hepatocytes.

    1.7  Effects on humans

         Only very limited information is available on the effects of DEHP
    on humans. Mild gastric disturbances, but no other deleterious
    effects, were reported for two subjects given 5 or 10 g DEHP.

    1.8  Effects on other organisms in the laboratory and field

         Most studies have yielded nominal LC50 values in acute toxicity
    tests that are in excess of 10 mg/litre, values which give a low
    toxicity rating for DEHP. However, these levels exceed the DEHP water
    solubility (0.3-0.4 mg/litre). One study suggested greater sensitivity
    of the water flea  Daphnia pulex, with a nominal 48-h LC50 of 0.133
    mg/litre. The only acute test with measured DEHP concentrations was on

    the fathead minnow and revealed a 96-h LC50 of > 0.33 mg/litre. In
    prolonged studies, the no-observed-effect level (NOEL) for  Daphnia
     magna was 72 µg/litre.  For adult fish a NOEL of > 62 µg/litre was
    determined. An exposure of 14 µg/litre, from 12 days prior to
    hatching, caused a significant increase in trout fry mortality. DEHP
    concentrations of between 3.7 and 11 µg/litre led to a reduction in
    the vertebral collagen of fish.

         The survival of zebra fish fry is adversely affected by DEHP
    concentrations of 50 mg/kg food. Sediment concentrations of 25 mg/kg
    (w/w) significantly reduced microbial activity and the number of
    tadpoles hatching.

         The acute toxicity of DEHP to algae, plants, earthworms, and
    birds is low.

    1.9  Evaluation

         DEHP causes reproductive and hepatocarcinogenic effects in rats
    and mice.

         Testicular atrophy is the main reproductive effect in rats and
    mice, and young animals are more susceptible than older ones to this
    effect. The induction of hepatic peroxisome proliferation and cell
    replication are strongly associated with the liver carcinogenic effect
    of certain non-genotoxic carcinogens including DEHP. However, marked
    differences have been observed among animal species with respect to
    DEHP-induced peroxisome proliferation. Currently there is not
    sufficient evidence to suggest that DEHP is a potential human

         There is no documented information that DEHP presents any hazard,
    based on acute exposure to fish and daphnids. However, a reduction of
    microbial activity in sediment at environmental levels of DEHP was
    reported. A comparison between environmental levels and the
    concentrations that produce effects in prolonged studies, especially
    early life-stage tests on fish and amphibians, indicates that a hazard
    for the environment, particularly via water and sediment, cannot be
    excluded. Adverse effects on organisms are likely in areas with highly
    contaminated water and sediments which are near to point emission

         Although few relevant studies have been reported, the acute
    toxicity of DEHP to algae, plants, earthworms, and birds appears to be


    2.1  Identity

    Common name:             di(2-ethylhexyl) phthalate

    Structural formula:      CHEMICAL STRUCTURE 1

    Empirical formula:       C24H38O4

    Abbreviation:            DEHP

    Relative molecular
     mass:                   390.57

    Common synonyms:

         1,2-benzenedicarboxylic acid bis(2-ethylhexyl) ester (CAS name);
         phthalic acid bis(2-ethylhexyl) ester (IUPAC name); BEHP;
         1,2-benzenedicarboxylic acid bis(ethylhexyl) ester;
         bis(2-ethylhexyl) 1,2-benzenedicarboxylate; bis(2-ethyl-hexyl)
         ester of phthalic acid; bis(2-ethylhexyl) phthalate;
         di(2-ethylhexyl)  ortho-phthalate; di(ethylhexyl) phthalate;
         dioctyl phthalate; DOP; ethylhexyl phthalate; 2-ethylhexyl
         phthalate; octyl phthalate; di- sec-octyl phthalate; phthalic
         acid dioctyl ester

    Common trade names:

         Bisoflex 81; Bisoflex DOP; Compound 889; DAF 68; Ergoplast FDO;
         Eviplast 80; Eviplast 81; Fleximel; Flexol DOP; Goodrite GP 264;
         Hatcol DOP; Kodaflex DOP; Mollan O; Nuoplaz DOP; Octoil;
         Palatinol AH; Platinol DOP; Pittsburgh; PX-138; Reomol DOP;
         Reomol D 79P; Sicol 150; Staflex DOP; Truflex DOP; Vestinol AH;
         Vinicizer 80; Witcizer 312 (IARC, 1982; NIOSH, 1985b)

    CAS registry
     number:                 117-81-7

    RTECS number:            TI 035000

         Di(2-ethylhexyl) phthalate (DEHP) is available in a variety of
    technical grades. In the USA typical product specifications are:
    minimal ester content, 99.0-99.6%; maximal moisture content, 0.1%;
    acidity (as acetic acid or phthalic acid), 0.007-0.01%; specific
    gravity, 0.980-0.985 (25 °C/25 °C); refractive index, 1.4850-1.4870
    (23 °C); and minimal flash-point, 216 °C (IARC, 1982).

         In western Europe, DEHP is available with the following
    specifications: maximal acid value, 0.06; maximal weight loss on
    heating at 140 °C for 3 h, 1%; and saponification value, 284-290 mg
    KOH/g (IARC, 1982).

         In Japan, DEHP must fulfill the following specifications: maximal
    acid value, 0.05; maximal weight loss on heating at 125 °C for 3 h,
    0.1%; and specific gravity, 0.983-0.989 (20 °C/20 °C) (IARC, 1982).

    2.2  Physical and chemical properties

         DEHP is a colourless to yellow, oily liquid at room temperature
    and normal atmospheric pressure. The melting point is -46 °C (pour
    point) and the boiling point is 370 °C (at atmospheric pressure, 101.3
    kPa; 236 °C at 1.33 kPa, and 231 °C at 0.67 kPa). The flash point is
    425 °C (open cup) (Clayton & Clayton, 1981; IARC, 1982; SAX, 1984). At
    20 °C, the density is 0.98 g/ml (Fishbein & Albro, 1972) and the
    vapour pressure 8.6 x 10-4 Pa (Howard et. al., 1985). The log  n-
    octanol-water partition coefficient is 3-5.

         The solubility of uncolloidal DEHP in water is low (45 µg/litre
    at 20 °C) (Leyder & Boulanger, 1983). However, DEHP may form colloidal
    dispersions which lead to higher values for solubility (Klöpfer et
    al., 1982). Values of 285 µg/litre (Hollifield, 1979), 340 µg/litre
    (Howard et al., 1985), and 360 µg/litre (Defoe et al., 1990) have been
    determined at 20-25 °C. These higher values are probably more
    realistic in the environment. Howard et al. (1985) determined a value
    of 160 µg/litre at 25 °C in salt water.

         DEHP is miscible with most common organic solvents and is more
    soluble in blood than water. It is lipophilic and the distribution
    ratio in dichloromethane-Krebs bicarbonate buffer has been measured to
    be 1130 (Krauskopf, 1973; Clayton & Clayton, 1981; IARC, 1982; Sax,
    1984; Weast et al., 1984; Sittig, 1985).

    2.3  Conversion factors

              1 ppm = 15.87 mg/m3
              1 mg/m3 = 0.063 ppm

    2.4  Analytical methods

         Methods used for the analysis of di(2-ethylhexyl) phthalate in
    many types of samples are summarized in Table 1.

         Analysis of samples with low concentrations of DEHP is
    complicated by the risk of contamination from plastic equipment.

        Table 1.  Methods for the analysis of di(2-ethylhexyl) phthalate
    Sample           Sample                              Assay             Limit of
    matrix           preparation                         procedure         detection       Reference
    Air              collect on cellulose                GC/FID            range:          NIOSH (1977)
                     ester membrane filter;                                2.03-10.9       NIOSH (1985a)
                     extract disulfide)                                    mg/m3 for
                                                                           a 32-litre
                                                                           sample at
                                                                           23 °C

    Air              collect with impinger               GC/ECD            not given       Thomas (1973)
                     (ethylene glycol);                  GC/MS
                     extract (hexane)

    Marine air       trap on glass-fibre filters         GC/ECD            0.5 ng/m3       Giam et al.
                     with foam plugs; Soxhlet                                              (1980)
                     extract (petroleum ether);
                     concentrate extracts; clean-up
                     on deactivated Florisil

    Air-borne        Soxhlet extract (methanol);         GC/MS             not given       Karasek et al.
    particulate      concentrate; centrifuge                                               (1978)

    River water      extract (hexane); filter            HPLC/UV           2 ng at         Mori (1976)
                                                         (normal and       224 nm
                                                         and gel

    Table 1 (contd).
    Sample           Sample                              Assay             Limit of
    matrix           preparation                         procedure         detection       Reference
    River water      extract (chloroform);               thin-layer        50 µg/litre     Kataeva (1988)
                     concentrate extract; dry by         chromatography
                     sodium sulfate treatment;
                     evaporate; dissolve residue

    Industrial       add hydrochloride acid;             GC/MC (EI         not given       Sheldon &
    and municipal    extract (dichloromethane);          and CI modes);                    Hites (1979)
    waste water      clean-up by liquid                  with SIM
                     chromatographic fractionation

    River            freeze-dry; homogenize;             HPLC/UV           10 ng           Schwartz et al.
    sediment         extract (hexane; acetone;           (233 nm)                          (1979)
                     methanol); evaporate;
                     dissolve (hexane); filter

    Human serum      centrifuge; extract (chloroform:    GC/FID            50 µg/litre     Lewis et al.
                     methanol); evaporate; dissolve      GC/MS                             (1977)
                     (ethyl acetate); treat with
                     alumina; decant; rinse;
                     filter; evaporate; dissolve
                     residue (hexane-containing
                     butyl benzyl phthalate as an
                     internal standard)

    Human plasma     separation on Celite 545;           GC/FID            50 ng           Piechocki &
                     extract (diethyl ether);                                              Purdy (1973)
                     evaporate; dissolve (carbon

    Table 1 (contd).
    Sample           Sample                              Assay             Limit of
    matrix           preparation                         procedure         detection       Reference
    Stored blood;    lyophilize; suspend; filter;        GC/FID            not given       Contreras et
    whole blood      wash residue; mix with distilled                                      al. (1974)
                     water; centrifuge; add silicic
                     acid to chloroform phase;
                     mix; centrifuge; decant;
                     evaporate; dissolve; centrifuge

    Human and        grind wet tissue samples            GC/FID            0.3 µg/g        Chen et al.
    animal           in saline; extract homogenate                         (wet tissue)    (1979b)
    tissue and       or urine; dilute with                                 15 ng/ml        Chen et al.
    urine            chloroform:methanol                                   (urine)         (1979a)

    Intravenous      add hydrochloric acid;              GC/ECD            4 µg/litre      Arbin &
    solutions        extract (dichloromethane);                                            Östelius
                     redissolve                                                            (1980)

    Organic          evaporate; dissolve                 GC/FID            not given       Ishida et al.
    solvents         (diethyl ether)                                                       (1980)

    Solid            immerse in chloroform:              GC/FID            not given       Ishida et al.
    reagents         methanol; filter; rinse;                                              (1980)

    Aluminium        cut into small pieces;              GC/FID            not given       Ishida et al.
    foil; rubber     immerse in chloroform:                                                (1980)
    tubing, etc.     methanol; extract
    Abbreviations:  GC = gas chromatography; FID = flame-ionization detection; ECD =
                    electron capture detection; MS = mass spectrometry; HPLC = high-
                    performance liquid chromatography; UV = ultraviolet spectroscopy;
                    EI = electron impact; CI = chemical ionization; SIM = selected ion

         It seems likely that by far the major part of the phthalate
    esters present in the environment arises from human activity and not
    from natural sources. Some dialkyl esters may be found in coals, crude
    oil, and shales while others may have plant origins, but most
    originate either directly or indirectly from industrial processes.
    Phthalic acid has been reported to be formed during the bacterial
    metabolism of phenanthrene.

    3.1  Natural occurrence

         Phthalates have been reported in a wide variety of substances
    (oil, soil, plants, and animals) and over a wide geographical area.
    Most occurrences have anthropogenic origins but some could be of
    natural origin. The nature of the origin is further complicated by the
    fact that sampling techniques often lead to contamination of samples
    via contamination from plastic bags or bottles. Mathur (1974a)
    critically reviewed this question and concluded that the possibility
    of the phthalic acid esters found in biological and geochemical
    samples being of biosynthetic origin cannot be ruled out. Both Mathur
    (1974a) and Peakall (1975) cite reports where phthalates were detected
    yet no anthropogenic source could be found. Studies by Manandhar et
    al. (1979) and Pare et al. (1981) also revealed residues of phthalates
    in biological samples where the source seemed to be natural. Peterson
    & Freeman (1984) suggested that some of the phthalates found in older
    samples (from the 1920s and 1930s) of sediment cores from Chesapeake
    bay, USA, may have been of natural origin.

         An ECETOC (European Chemical Industry Ecology and Toxicology
    Centre) task force concluded that, although knowledge of naturally
    produced phthalates is limited or uncertain, it is unlikely that this
    contribution is of significance except, possibly, in very localized
    areas (ECETOC, 1985).

    3.2  Anthropogenic sources

    3.2.1  Production levels

         About 2.7 x 106 tonnes of total phthalates are produced
    annually, of which the non-plasticizer (dimethyl and diethyl)
    phthalates represent a very small fraction. Of the plasticizer
    phthalates, DEHP accounts for well over 50% of the tonnage, the
    contribution of the remaining compounds ranging from about 1% to 10%
    each (ECETOC, 1985).

         The production of DEHP has been increasing since it was first
    used commercially in 1949. During the period 1950-1954, the production
    in the USA was 106 x 103 tonnes, and by the period 1965-1969 the
    level had risen to 650 x 103 tonnes (Peakall, 1975).

         The estimated world consumption of DEHP in 1984 was 1.09 x 106
    tonnes (SRI, 1985).

    3.2.2  Uses

         Phthalate acid esters are the most widely used plasticizers for
    the production of polyvinyl chloride (PVC) products (with DEHP as the
    plasticizer). Phthalates are used for the insulation of wires and
    cables, in floor tiles, weatherstripping, upholstery, garden hose,
    swimming pool liners, footwear and clothing. They are also used in
    food wrapping and containers, although in some countries this use is
    prohibited by law. They also have non-plasticizer uses, e.g., as
    pesticide carriers.

         DEHP has been widely used since 1949. An important property is
    that it softens resins without reacting with them chemically. This has
    led to about 95% of DEHP production being directed towards plasticizer
    use, particularly in PVC products such as tubing and medical device
    components. It is also used as a plasticizer in cellulose ester
    plastics and synthetic elastomers. The DEHP content of these products
    generally ranges from 20 to 40%, but for some uses it is up to 55%.
    The most important non-plasticizer use of DEHP is as a dielectric
    fluid in capacitors.

    3.2.3  Disposal of plasticized products

         Most discarded plasticized products are disposed of either by
    incineration or via dumping in a tip/landfill site. When incinerated
    at high temperature the combustion of phthalates is nearly complete.
    However, if combustion is uncontrolled and occurs at a low
    temperature, a large percentage of the phthalates may be lost to the
    atmosphere. After dumping in a landfill site, phthalates may leach
    into the aquatic environment, but because of their high affinity for
    organic soil particles and their low water solubility this is not
    likely to be a major route into the environment. Indiscriminate
    dumping is more likely to lead to volatilization of phthalates to the
    atmosphere rather than leaching to the aquatic environment (ECETOC,


    4.1  Environmental transport and distribution

         The release of phthalates to the environment may occur as

         a)   during production and distribution;
         b)   during the manufacture of plasticized products;
         c)   during the use of plasticized product;
         d)   after disposal.

         It was concluded by ECETOC (1985) that most of the phthalates
    entering the environment are likely to do so by volatilization to the
    atmosphere, only a minor part (perhaps 10%) entering the aquatic
    environment by leaching.

         The estimated worldwide emissions of DEHP are given in Table 2.
    However, as reported by ECETOC (1985), the loss of DEHP from modern
    production plants is negligible.

    4.1.1  Transport in air

         ECETOC (1985) suggested that transport in air is the major route
    by which phthalates enter the environment. DEHP is volatilised to the
    atmosphere and then either falls as dry deposition or is "washed out"
    via rainfall.

         DEHP has been measured in air samples from remote sites at
    Enewetak Atoll in the North Pacific Ocean (Atlas & Giam, 1981), the
    North Atlantic (Giam et al., 1978), the Gulf of Mexico (Giam et al.,
    1978, 1980), and Sweden (Thurén & Larsson, 1990).

    4.1.2  Transport in soil and sediment

         DEHP has a high  n-octanol-water partition coefficient, so the
    equilibrium between water and an organic-rich soil or sediment will be
    very much in favour of the soil or sediment. Absorbance of DEHP by
    soil or sediment is also enhanced by van de Waals type bonding with
    natural soil minerals, promoted by the presence of benzene rings and
    carbonyl groups, and also by the low solubility of DEHP (ECETOC,

         From results with other organic substances, Wams (1987) estimated
    that 90% of DEHP is readily adsorbed by organic soil particles.

         As can be seen from section 5.1.4, the sediment or hydrosoil
    tends to act as a sink for DEHP.

        Table 2.  Estimated worldwide emission of DEHP, based on an estimated
              total annual production of 4 x 106 tonnesa
    Phase                    Emission (tonnes/year)      Route
    Production                   up to 40 000         waste water
    Distribution                     2000             sewage systems
    Production of PVC               32 000            air and water
    During use of plastics          14 000            air
                                     6000             water

    After disposal:

      to landfill sites          up to 200 000        percolating water
      to waste incinerators            ?              air
      uncontrolled burning             ?              air

    a    Adapted from Wams (1987). The values are higher than those from
         other sources (ECETOC, 1985)
    4.1.3  Transport in water

         The solubility of DEHP in water is low (0.3 mg/litre at 25 °C). 
    However, the amount present in surface water may be higher than the
    actual solubility as a result of adsorption onto organic particles
    (Taylor et al., 1981) and interactions with dissolved organic matter
    of high relative molecular mass, such as humic and fulvic acid
    (Matsuda & Schnitzer, 1971).

         DEHP has been found to adsorb to suspended particulate matter
    fairly rapidly, in less than 2 to 3 h, especially to small particles
    (Al-Omran & Preston, 1987). This adsorption was more rapid in salt
    water than in fresh water. Taylor et al. (1981) reported that between
    one-half and two-thirds of the DEHP in Mississippi River water is
    associated with particulate matter.

         By extrapolating laboratory data on the volatilization of DEHP
    from water under defined conditions, Klopffer et al. (1982) obtained
    a half-life in water of 146 days, although on purely theoretical
    grounds a value of only 25 days was calculated.

         Using the Exposure Analysis Modeling System (EXAMS), Wolfe et al.
    (1980a) calculated that at equilibrium the loss of DEHP via
    volatilization from a model river, a pond, an eutrophic lake, and an
    oligotrophic lake would be 0%, 2.8%, 2.2% and 2.3%, respectively.

    4.1.4  Transport between media

         Eisenreich et al. (1981) estimated that the total annual
    deposition of DEHP from air into the Great Lakes, North America,
    varied from 3.7 tonnes (Lake Ontario) to 16 tonnes (Lake Superior).

         DEHP has a high  n-octanol/water partition coefficient. This
    means that biota living in phthalate-containing water would be
    expected to have a higher phthalate level than the water itself (see
    section 4.2.3). However, many organisms are able to metabolize DEHP,
    and the concentrations found may be lower than those expected on the
    sole basis of partition coefficient.

         During the 33-day period of a model ecosystem study, the
    concentration of 14C in the aquatic phase reached a peak of 31
    µg/litre at the fifth day after treatment and had declined to 7.7
    µg/litre by the end of the experiment. This decline was stated to be
    the result of the uptake of DEHP and its degradation products by the
    organisms in the model ecosystem (Metcalf et al., 1973).

         Lokke & Bro-Rasmussen (1981) treated the leaves of  Sinapsis alba
    with a mixture of di-iso-butyl phthalate (DiBP), di- n-butyl
    phthalate (DBP), and DEHP at a rate of 2.5 µg/cm2. Only very small
    amounts of DEHP evaporated from the leaves during the 15-day
    experiment, compared with DiBP (71%) and DBP (43%).

    4.2  Biotransformation

    4.2.1  Abiotic degradation

         As a result of atmospheric photodegradation, the atmospheric
    half-life of DEHP is less than one day (ECETOC, 1985).

         Chemical hydrolysis of DEHP is practically non-existent, the
    half-life being > 100 years in water at pH 8 and 30 °C (Wolfe et al.,

    4.2.2  Biodegradation

         Aerobic degradation has been found from several micro-organisms
    in soil, sludge, sediment, and water. Anaerobic degradation is very
    much slower, or possibly even non-existent.

         The first step in the metabolic pathway for the biodegradation of
    DEHP is the hydrolysis of the diester to the monoester by esterases
    with low substrate specificity (Kurane et al., 1980; Taylor et al.,
    1981). The monoester is then converted into phthalic acid (Engelhardt
    et al., 1975). The ring-opening degradation to pyruvate and succinate
    and then to CO2 and H2O is similar to the  metabolism of benzoic
    acid. According to Kurane et al. (1984), this is probably why the
    biodegradation of phthalic esters is so widespread. It appears that
    mixed populations of microorganisms are the most successful at
    completely degrading DEHP (Engelhardt et al., 1975; Kurane et al.,
    1979). When pure cultures of bacteria, selectively isolated in the
    laboratory, are used for the biodegra-dation of phthalates,
    accumulation of the breakdown products tends to occur (Keyser et al.,
    1976).  Aerobic degradation

         Aerobic degradation of DEHP has been found with several
    microorganisms, including bacteria and fungi. Overall, it appears that
    phthalates with short alkyl chains undergo rapid degradation, whereas
    those with longer chains, such as DEHP, are only 40-90% degraded after
    10-35 days (ECETOC, 1985).

         Graham (1973) reported that laboratory-scale activated sludge
    processes degraded 91% of introduced DEHP within 38 h. Saeger & Tucker
    (1973) demonstrated that all phthalates tested underwent complete
    aerobic degradation in activated sludge and river water.

         Aerobic degradation of DEHP depends on temperature. Mathur
    (1974b) incubated a loam soil with DEHP at 4, 10, 22-25, and 32 °C,
    and soil respiration rates were measured after 14 weeks.  Increased
    rates of respiration, showing that microbial degradation was taking
    place, were found at all temperatures. However, at 4 and 10 °C results
    indicated that only marginal degradation was taking place.

         Johnson & Lulves (1975) incubated freshwater hydrosoil containing
    14C-DEHP (1 mg/litre) under aerobic conditions, and after 14 days,
    50% of the DEHP had been degraded. This was a much slower rate of
    degradation than that found with DBP, where 98% was degraded within 5
    days. Johnson et al. (1984) studied the biodegradation of phthalic
    acid esters in freshwater sediment and  found that the length and
    configuration of the alkyl phthalate diester significantly affected
    the primary biodegradation rate.  After a 14-day incubation in aerobic
    sediment at 22 °C, less than 2% of the branched-chain alkyl phthalate
    DEHP had been degraded whereas over the same period the linear alkyl
    DBP showed 85% degradation. DEHP degradation was significantly greater
    at very high concentrations (10 mg/litre) and at temperatures above 22
    °C. Neither inorganic nitrogen nor phosphorus influenced the
    degradation of DEHP. Engelhardt et al. (1977) found that the fungus
     Penicillium lilacinum degraded approximately half of the initial
    amount of DEHP within 30 days, yielding the corresponding monoester,
    a second metabolite, which is hydroxylated in the alcohol moiety, and
    at least four minor metabolites. The bacterium  Pseudomonas
     acidovorans completely degraded DEHP at a medium concentration of
    5000 mg/kg within 72 h (Kurane et al., 1977).

         Saeger & Tucker (1976) found that 60% of the DEHP had undergone
    primary biodegradation within 5 weeks in Mississippi River water.
    Rapid primary degradation was found when DEHP was added to activated
    sludge at the rate of 5 mg/24 h. Depending on the source of the
    sludge, between 70% and 78% was degraded.  To monitor whether complete
    biodegradation was being achieved, the authors measured CO2
    evolution. Within the 14 days of incubation DEHP had essentially been
    completely degraded to CO2 and water under the conditions of this
    test. Taylor et al. (1981) showed the presence of significant
    populations of taxonomically distinct bacteria that grew on a range of
    phthalic acid esters, including DEHP, in the water and sediments of
    the Mississippi River region.

         Sugatt et al. (1984) used an acclimated shake-flask
    CO2-evolution test to study the biodegradation of DEHP and reported
    an initial breakdown of the parent compound of > 99% within the 28
    days. The authors calculated a half-life of 5.25 days for the primary
    biodegradation of DEHP.

         In surface waters, DEHP is strongly adsorbed to organic particles
    (Taylor et al., 1981), which tends to reduce degradation (Baughman et
    al., 1980).

         In the upper layer of soil, biodegradation of phthalates proceeds
    as in surface water, but deeper down, where conditions are anaerobic,
    it is virtually nonexistent (Engelhardt & Wallnofer, 1978). Shanker et
    al. (1985) incubated garden soil containing DEHP at a concentration of
    500 mg/kg. Within 20 days, 75% of the DEHP had been degraded and,
    after 30 days, more than 90%.  Again the rate of degradation was much

    slower than that found for either di- n-methyl or di- n-butyl
    phthalate. No degradation was detectable when sterilized soil was
    used.  Anaerobic degradation

         Johnson & Lulves (1975) found DEHP to be completely resistant to
    microbial attack under anaerobic conditions. After 30 days, there was
    no significant loss of 14C-DEHP activity in freshwater hydrosoils
    overlaid with nitrogen.

         Shanker et al. (1985) reported that degradation of DEHP was much
    slower in anaerobic soil, flooded with sterile water to reduce the
    oxygen tension. After a 30-day incubation, 33% of the DEHP had been
    degraded, compared with more than 90% in the case of aerobic soil.

         O'Connor et al. (1989) studied the biodegradation of DEHP under
    anaerobic conditions in a medium containing municipal digester sludge
    over a period of 140 days. DEHP, which was the only carbon source, was
    added at a rate of 20, 100, and 200 mg/litre, and 100%, 69%, and 54%
    of the DEHP was degraded at the three respective concentrations.
    However, complete biodegradation to carbon dioxide and methane was

         Ziogou et al. (1989) studied the behaviour of DEHP (0.5, 1, and
    10 mg/litre) during batch anaerobic digestion of sludge over a 32-day
    period. No degradation of DEHP was observed during this period.

    4.2.3  Bioaccumulation

         DEHP is highly lipophilic, the log  n-octanol-water partition
    coefficient being 3 to 5, and it is moderately persistent. The
    accumulation of DEHP is also influenced by the capability of an
    organism to metabolize it. Melancon (1979) reviewed the metabolism of
    phthalates in aquatic organisms. Bioconcentration factors for DEHP in
    a variety of aquatic organisms are given in Table 3.

        Table 3.  Bioaccumulation of DEHP in aquatic organisms
    Organism                    Stat/     Exposure   Exposure            Bioconcentration     Reference
                                flowa     period     concentration           factorc
    Freshwater organisms

    Canadian pondweed            stat       24 h          10                274.8d            Metcalf et al. (1973)
      (Elodea canadensis)        stat       12 h        10 000              133.8d            Metcalf et al. (1973)

    Snail                        stat       48 h          10                857.5d            Metcalf et al. (1973)
     ( Physa sp)                 stat       6 h         10 000e               402d            Metcalf et al. (1973)

    Scud                         flowb      7 day         0.1               13 600            Sanders et al. (1973)
      (Gammarus pseudolimnaeus)  flow       7 day         0.1                3900             Mayer & Sanders (1973)

    Water flea                   flowb      7 day         0.3                5200d            Sanders et al. (1973)
      (Daphnia magna)            flow       7 day         0.3                 420             Mayer & Sanders (1973)
                                 stat       1 h           10                 421d             Metcalf et al. (1973)
                                 stat       12 h        10 000e             133.8d            Metcalf et al. (1973)

    Sowbug                       flowb      21 day       62.3                 250d            Sanders et al. (1973)
      (Asellus brevicaudus)

    Mosquito (larvae)            stat       12 h          10               1320.2d            Metcalf et al. (1973)
      (Culex pipiens             stat       24 h        10 000e            1187.3d            Metcalf et al. (1973)
      quinquefasciatus)          stat       24 h          10                 20.3d            Metcalf et al. (1973)
                                 stat       48 h        10 000e             434.6d            Metcalf et al. (1973)

    Midge larvae (3rd instar)    flowb       7 day        0.2                 408             Streufert et al. (1980)
      (Chironomus plumosus)      flowb       7 day        0.3                3100d            Sanders et al. (1973)
                                 flow        7 day        0.3                 350d            Mayer & Sanders (1973)

    Mayfly                       flowb       7 day        0.1                2300d            Sanders et al. (1973)
      (Hexagenia bilineata)      flow        7 day        0.1                 575d            Mayer & Sanders (1973)

    Table 3 (contd).
    Organism                    Stat/     Exposure   Exposure            Bioconcentration     Reference
                                flowa     period     concentration           factorc

    Mosquito fish                stat       48 h         100                  265.3d        Metcalf et al. (1973)
      (Gambusia affinis)          stat      12 h        10 000e               129.4d        Metcalf et al. (1973)

    Fathead minnow               flow      14 day         1.9                  458d         Mayer & Sanders (1973)
      (Pimephales promelas)      flow      56 day         1.9                  886d         Mehrle & Mayer (1976)

    Marine organisms

    Eastern oyster (muscle)      stat       24 h          100                  11.2         Wofford et al. (1981)
      (Crassostrea virginica)    stat       24 h          500                   6.9         Wofford et al. (1981)

    Brown shrimp                 stat       24 h          100                  10.2         Wofford et al. (1981)
      (Penaeus aztecus)          stat       24 h          500                  16.6         Wofford et al. (1981)

    Sheepshead minnow            stat       24 h          100                  10.7         Wofford et al. (1981)
      (Cyprinodon variegatus)    stat       24 h          500                  13.5         Wofford et al. (1981)

    a  Stat = static conditions (water unchanged for the duration of the test); flow = flow-through conditions
       (DEHP concentration in water continuously maintained, unless stated otherwise)
    b  Intermittent flow-through conditions
    c  Bioconcentration factor = concentration of DEHP in organism divided by concentration of DEHP in water
    d  Bioconcentration factor calculated using a radioactive isotope (values represent parent compound plus
       radiolabelled products)
    e  DEHP applied directly to water  Model ecosystems

         Metcalf et al. (1973) studied the uptake of 14C-labelled DEHP
    from water by aquatic organisms in a model ecosystem containing algae
     (Oedogonium), snails ( Physa sp.), mosquito larvae  (Culex pipiens
     quinquefasciatus), and fish ( Gambusia sp). The mosquito larvae
    showed the highest concentration factor and the fish the lowest.
    Labelled DEHP was added to  Sorghum plants and at the end of the 33-
    day experiment the water contained 0.34 µg DEHP per litre, the algae
    18.32 mg/kg (53 890 x), the snails 7.3 mg/kg (21 480 x), the mosquito
    larvae 36.61 mg/kg (10 7670 x), and the fish 0.044 mg/kg (130 x).

         Sodergren (1982) exposed fish, aquatic invertebrates, and plants
    to 14C-labelled DEHP at a concentration of 1.4 mg/litre for 27 days
    under static conditions. After 5 days, 1/50 of the added amount of
    DEHP was still present in the water, and at the end of the experiment
    62% was recovered from the various surfaces (glass walls, sediment and
    surface microlayer). All organisms accumulated DEHP. The amphipod
     Gammarus pulex, larvae of trichopterans, and the snail  Planorbis
     corneus accumulated the DEHP to the highest degree, the
    concentration factors ranging from 17 000 to 24 000. The submerged
    plants,  Mentha aquatica and  Chara chara, also showed uptake and
    storage of large amounts (concentration factor of 18 000). However,
    the fish (stickleback,  Pungitius pungitius, and minnow,  Phoxinus
     phoxinus) did not accumulate 14C-DEHP to any great extent
    (concentration factors of 300 or less). Large accumulations of DEHP
    occurred in organisms living and/or feeding at interfaces.  Aquatic invertebrates

         Brown & Thompson (1982a) exposed Daphnia magna to nominal
    14C-labelled DEHP concentrations of 3.2, 10, 32, and 100 µg/litre
    for 21 days and obtained bioconcentration factors of 166, 140, 261,
    and 268 at the four respective concentrations.

         When Brown & Thompson (1982b) exposed mussels  (Mytilus edulis)
    to labelled DEHP at concentrations of 4.1 and 42.1 µg per litre, in
    both cases equilibrium was reached within 14 days with a concentration
    factor of 2500. Exposure ceased on day 28 but the mussels were
    monitored for a further 14 days. The half-life for loss of DEHP over
    this period was calculated to be 3.5 days.

         Laughlin et al. (1978) exposed grass shrimp, during larval
    development, to DEHP concentrations of up to 1 mg/litre for 28 days.
    DEHP was not detectable in shrimp tissues at or above a level of 2

         When Streufert et al. (1980) exposed midge larvae  (Chironomus
     plumosus) to a radioactively labelled DEHP concentration of 0.2
    µg/litre, the larvae accumulated DEHP to 292 times the concentration
    in water within 2 days. DEHP levels in the midge larvae reached a

    plateau after 7 days at a bioconcentration factor of 408. Some of the
    larvae were transferred to clean water after 4 days, by which time
    they had accumulated 56 µg DEHP/kg, and the half-life for loss was
    calculated to be 3.4 days.

         After 9 weeks of exposure to sediment containing approximately
    600 mg/kg, dragonfly larvae had taken up 14.7 mg DEHP per kg. This was
    significantly more than control larvae, which contained 2.9 mg/kg
    (Woin & Larsson 1987).

         Hobson et al. (1984) fed penaeid shrimps on a diet containing
    between 40 and 50 000 mg DEHP/kg for 14 days at a rate of 40 g/kg body
    weight per day. Whole body residues ranged from 0.249 to 18.3 mg/kg in
    a dose-related manner.  Fish

         Macek et al. (1979) exposed bluegill sunfish (Lepomis
    macrochirus) to 14C-DEHP, both via food at a concentration of 2.8
    mg/kg and via water at 5.6 µg/litre, for up to 35 days. The steady-
    state body burden of 14C-DEHP after exposure via food and water was
    0.73 mg/kg and via water alone was 0.64 mg/kg. The authors concluded
    that the uptake of DEHP via the aquatic food chain was statistically
    indistinguishable from that due to aqueous exposure. The time required
    for the fish to eliminate 50% of the residue burden during depuration
    in uncontaminated water was < 3 days.

         In a study by Karara & Hayton (1989), sheepshead minnows
     (Cyprinodon variegatus) were exposed to a 14C-DEHP concentrations
    of 60 ng/litre at temperatures ranging from 10 °C to 35 °C for a
    period of between 72 h and 160 h. The amount of DEHP accumulated after
    72 h was 6 times greater at 35 °C than at 10 °C, and the
    bioconcentration factors increased exponentially with temperature from
    45 at 10 °C to 6510 at 35 °C. Metabolic clearance also increased as a
    function of temperature, the maximum value being reached at a
    temperature of between 29 °C and 35 °C.

         Tarr et al. (1990) exposed three sizes (2.9 g, 61 g, and 440 g)
    of rainbow trout  (Oncorhynchus mykiss) to 14C-DEHP at 20 ng/ml
    under static conditions for up to 96 h at 12 °C. The body-weight-
    associated changes in the pharmacokinetic parameters caused the
    bioconcentration factor to decline from 51.5 to 1.6 as body weight

         When Mehrle & Mayer (1976) exposed rainbow trout  (Salmo
     gairdneri) eggs (12 days prior to hatching to 24 days post-hatching)
    to 14C-labelled DEHP at concentrations of 5, 14, and 54 µg/litre,
    the bioconcentration factors were 78, 113, and 42, respectively.

         Mayer (1976) exposed fathead minnows  (Pimephales promelas) to
    DEHP concentrations ranging from 1.9 to 62 µg/litre for 56 days under
    flow-through conditions. As the exposure concentration increased,
    concentration factors, measured after 56 days, decreased from 569 to
    91. Equilibrium was attained after 28 days at the lowest dose and
    after 56 days at the highest dose. After exposure the fish were placed
    in uncontaminated water for 28 days, and the half-lives for loss
    ranged from 6.2 days (at 2.5 µg/litre) to 18.3 days (at 62 µg/litre).  Amphibians

         Larsson & Thuren (1987) exposed moorfrog eggs to sediment DEHP
    concentrations ranging from 10 to 800 mg/kg (fresh weight of
    sediment). The eggs hatched after about 3 weeks and the tadpoles were
    analysed after 60 days. The DEHP was released from the sediment to the
    overlying water, and the losses to the water increased linearly with
    increasing levels in the sediment (from 0.89 to 187.4 µg/litre). DEHP
    accumulated in the tadpoles at concentrations ranging from 0.28 to
    246.8 mg/kg wet weight, and the accumulation increased with increasing
    DEHP concentration, both in sediment and water.  Plants

         Lokke & Bro-Rasmussen (1981) applied DEHP as a mixture that also
    contained DiBP and DBP at a concentration of 2.5 µg/cm2 to the
    leaves of  Sinapis alba. The residue level of DEHP on the leaves
    immediately after application was 2.7 µg/cm2. After 15 days, DEHP
    levels had decreased to 0.8 µg/cm2, but when the growth of the plant
    was taken into account, no significant loss of DEHP over this period
    was found. Lokke & Rasmussen (1983) also found little loss of DEHP
    over a 15-day period when they applied it (as a mixture with DBP) to
     Achillea at a concentration of 3.5 µg/cm2 or to Sinapis at 3.1
    µg/cm2. Residues ranged from 120 to 155 µg/plant, and approximately
    80% of the DEHP accumulated was on the surface of the leaf.

         When Schmitzer et al. (1988) grew barley from seed in soil
    containing 1 or 3.33 mg 14C-DEHP/kg dry soil for 7 days, only 0.61%
    and 1.25%, respectively, of the applied 14C was taken up by the
    plants. Aranda et al. (1989) also found low accumulation of DEHP by
    plants grown in sewage sludge containing DEHP.  Lettuce (Lactuca
    sativa), carrot  (Daucus carota), chile pepper  (Capiscum annuum),
    and tall fescue  (Festuca arundinaca) were all grown in sludge
    containing 14C-DEHP levels of between 2.57 and 14.07 mg/kg.
    Bioconcentration factors ranged from 0.06 to 0.53 (based on initial
    soil concentration and plant dry weight).  Birds

         Belisle et al. (1975) fed mallard ducks  (Anas platyrhynchos) on
    a diet containing 10 mg DEHP/kg for a period of 5 months. No DEHP was

    detected in fat tissue, but 0.1 and 0.15 mg/kg (wet weight) were found
    in breast muscle and lung tissue, respectively.

         O'Shea & Stafford (1980) exposed starlings  (Sturnus vulgaris)
    to a dietary DEHP concentration of 25 or 250 mg/kg for 30 days.  One
    of eight birds fed 25 mg/kg contained detectable residues (1.6 mg/kg)
    after 30 days exposure, and five of eight birds fed 250 mg/kg
    contained an average of 1.8 mg/kg. The same proportion of birds fed
    the higher dose still had detectable residues (an average of 1.3
    mg/kg) 14 days after dosing had finished.

         When Ishida et al. (1982) fed hens on a diet containing 5 or 10
    g/kg for up to 230 days, DEHP was detected in all tissue monitored
    except the brain. Residues ranged from non-detectable to 42.5 mg/kg
    for most tissues. However, adipose tissue (192.7 to 899.6 mg/kg) and
    feathers (513.1 to 1165.2 mg/kg) accumulated the highest
    concentrations. A similar pattern of uptake was observed in hens fed
    2 g/kg for 25 days, although the amount of DEHP accumulated was much
    lower. At this dose level no accumulation had occurred in tissues
    other than liver and feather within 5 days.


    5.1  Environmental levels

         DEHP exists widely in the environment and is found in most
    samples, including air, precipitation, water, sediment, soil, and
    biota. Residues have also been detected in food and in humans.

         In many cases it is not clear whether the phthalate measured in
    samples is naturally occurring or is exogenous. However, there seem to
    be clear indications that high levels of DEHP are anthropogenic in

    5.1.1  Air

         The levels of DEHP in air have been monitored in the North
    Atlantic, the Gulf of Mexico, and on Enewetak Atoll in the North
    Pacific and found to range from not detectable to 4.1 ng/m3 (Giam et
    al., 1978; Giam et al., 1980; Atlas & Giam 1981). Similar levels
    (between 0.5 and 5 ng/m3) have been found in the Great Lakes
    ecosystem (Eisenreich et al., 1981) and in the Swedish atmosphere
    (Thurén & Larsson, 1990). The DEHP content of these samples was at
    least an order of magnitude lower than those found in urban areas such
    as New York city, where levels of up to 28.6 ng/m3 have been found
    (Bove et al., 1978). Based on the analysis of snow samples, Lokke &
    Rasmussen (1983) calculated DEHP concentrations in air of 22 ng/m3
    at Lyngby, Denmark. Levels of 29-132 ng/m3 have been found in
    Antwerp, Belgium (Cautreels et al., 1977), 126 ng/m3 in polluted air
    in Belgium (Cautreels & Van Cauwenberghe, 1978), 300 ng/m3 in
    polluted air in Canada (Thomas, 1973), and 38-790 ng/m3 in Japan in
    1985 (Environment Agency of Japan, 1989).

    5.1.2  Precipitation

         Atlas & Giam (1981) measured levels of DEHP in rainfall at
    Enewetak Atoll, North Pacific, of 5.3-213 ng/litre (mean, 55
    ng/litre). Eisenreich et al. (1981) reported between 4 and 10 ng/litre
    in precipitation falling on the Great Lakes ecosystem and  Thurén &
    Larsson (1990) a level of 55 ng/litre in Sweden. Goto (1979) found a
    range of mean rainwater concentrations of 0.65 to 3.16 µg/litre in
    various Japanese cities.

         Lokke & Rasmussen (1983) analysed snow sampled near a plasticizer
    production plant 14 days after a snowfall. Levels of DEHP ranged from
    0.7 to 4.7 µg/m2 per day over this period, the highest levels being
    within 150 m of the plant and the lowest levels at least 600 m away.

    5.1.3  Water

         Levels of DEHP found in water are summarized in Table 4.

        Table 4.  Concentrations of DEHP in water
    Location              Country         Year   Concentration         Reference

    Northern Atlantic                             0.0001-0.006    Giam et al. (1978)
    Gulf of Mexico                                 0.006-0.316    Giam et al. (1978)
    Estuaries            Germany                     ND-0.3       Weber & Ernst (1983)
    Nueces Estuary,
       Texas             USA              1980      0.2-0.77      Ray et al. (1983b)
    Estuaries            United Kingdom   1981     0.058-0.078    Waldock (1983)


    Various rivers       Japan                       ND-3.1       Kodama et al. (1975)
    Various cities       Japan            1974      0.1-2.19      Goto (1979)
    River Meuse          Netherlands      1983      < 0.1-3.5     Wams (1987)
    River Rhine          Netherlands      1983       ND-1.2       Wams (1987)
    River Rhine          Netherlands      1982       ND-4.0       Wams (1987)
    a  ND = not detectable
         Thuren (1986) analysed water samples from the Rivers Ronnebyan
    and Svartan, Sweden, and found DEHP concentrations ranging from 0.32
    to 3.1 µg/litre and from 0.39 to 1.98 µg/litre, respectively. The
    highest concentrations were associated with industrial effluent
    discharge points.

         Few samples of ground water have been analysed for DEHP. Wams
    (1987) reported that contaminated ground water in the Netherlands
    contained between 20 and 45 µg/litre, while Rao et al. (1985) found
    DEHP levels of up to 170 µg/litre in New York state ground water.

         In a non-industrialised estuary in the United Kingdom, Waldock
    (1983) measured DEHP levels of 58 to 78 ng/litre. Ray et al. (1983b)
    found levels of DEHP ranging from 210 to 770 ng/litre in a Nueces
    estuary in Texas, USA, while Weber & Ernst (1983) found DEHP levels of
    up to 300 ng/litre in German estuaries. DEHP levels of up to 316
    ng/litre have been found in the Gulf of Mexico, but levels in the
    North Atlantic were much lower (Giam et al., 1978). In Japan, river
    and marine levels in 1982 ranged from 0.1 to 0.8 µg/litre (Environment
    Agency of Japan, 1989).

         Ritsema et al. (1989) analysed samples from Lake Yssel,
    Netherlands, and found DEHP levels of < 0.1 to 0.3 µg/litre in water
    and levels of 12-25 mg/kg in suspended particulate matter. The authors
    concluded that the probable source of DEHP was the River Yssel.

         Preston & Al-Omran (1986) sampled water and suspended
    particulates from the Mersey estuary, United Kingdom, in 1985, and
    reported DEHP concentrations ranging from 83 to 335 ng/litre in water
    and from 182 to 1700 µg/kg in particulate matter.  However, in 1986,
    levels were 125-693 ng/litre in water and 280-640 µg/kg in particulate
    matter (Preston & Al-Omran, 1989).

    5.1.4  Sediment

         DEHP levels in sediment are summarized in Table 5.

         Being lipophilic DEHP tends to be adsorbed onto sediment, which
    acts as a sink. Sediment samples from various Dutch rivers have been
    found to contain between 1 and 70 mg/kg (Schwartz et al., 1979; Wams,
    1987). Taylor et al. (1981) analysed sediment samples from the
    Mississippi River and found similar levels.  Sediment levels of DEHP
    in the Chester River Maryland, USA,  were found to be less than 45
    µg/kg dry weight, but, in a tributary of this river, sediment levels
    were up to 4.8 mg/kg about 2 km downstream from a phthalate ester
    plant outfall. The Chester River flows into Chesapeake Bay, which
    contained sediment DEHP levels of 110 µg/kg (Peterson & Freeman 1984).
    In Sweden, Thuren (1986) found sediment DEHP levels ranging from 1.2
    to 628 mg/kg (dry weight) in the River Ronnebyan and 0.15 to 1480
    mg/kg in the River Svarten. As was found with water samples (see
    section 5.1.3), in both rivers the highest residues of DEHP were near
    to industrial effluent discharge points.

         Giam et al. (1978) analysed sediment from the Mississippi delta
    and reported mean DEHP levels of 69 µg/kg. Sediment samples from
    Nueces Estuary, Texas, USA, unlike the water samples, reflected local
    inputs of pollutants. The highest levels (up to 16 mg/kg) were
    associated with industrial areas, whereas other areas of the estuary
    contained levels ranging from 40 to 330 µg/kg.  Much lower levels of
    DEHP were found on the Gulf of Mexico coast and in the open sea (mean
    levels of 6.6 and 2 µg/kg, respectively) (Giam et al., 1978).

        Table 5.  Concentrations of DEHP in sediment
    Location                      Country             Year       Concentration           Reference
    Gulf of Mexico                                              < 0.1-248 ns         Giam et al. (1978)
    Nueces Estuary, Texas         USA                 1980        40-16 000 dw       Ray et al. (1983b)
    Portland, Maine               USA                 1980         60-7800 dw        Ray et al. (1983a)
    Chester River, Maryland       USA                 1978         20-4800 dw        Peterson & Freeman (1984)
    River Mississippi             USA                 1981          140 dw           Taylor et al. (1981)
    River Meuse                   Netherlands         1977       1000-17 000 dw      Schwartz et al. (1979)
    River Ijssel                  Netherlands         1977       2500-52 500 dw      Schwartz et al. (1979)
    River Rhine                   Netherlands         1978       4000-36 000 ns      Wams (1987)
    River Rhine                   Netherlands         1977       6500-70 500 dw      Schwartz et al. (1979)
    Various cities                Japan               1974         80-1360 dw        Goto (1979)
    Crouch Estuary, Essex         United Kingdom      1981        11.2-26.2 ww       Waldock (1983)
    River Usk                     United Kingdom      1974         30 000 dw         Eglinton et al. (1975)
    a  dw = dry weight; ww = wet weight; ns = not stated whether concentrations refer to dry or wet weight
         In Japan, the levels of sediments in rivers and seas in 1982
    ranged from 9 to 35 000 µg/kg dry weight (Environment Agency of Japan,

    5.1.5  Soil

         Contaminated soil analysed in the Netherlands was found to
    contain up to 1.5 mg DEHP/kg (Wams 1987), while residues of DEHP in
    soil collected in the vicinity of a DEHP manufacturing plant contained
    up to 0.5 mg/kg (Persson et al., 1978). 

         Fatoki & Vernon (1990) reported a DEHP concentration of 1.9
    µg/litre in treated sewage effluent from the Manchester area, United
    Kingdom, and stated that such a level was consistent with the
    industrial activities of the city.

    5.1.6  Food

         DEHP has been found in many samples of fish and shellfish (see
    Table 6). It has also been detected in milk (Cerbulis & Ard, 1967),
    bovine pineal gland (Taborsky, 1967), bovine heart muscle (Nazir et
    al., 1971), and chicken eggs (Ishida et al., 1982). Perkins (1967)
    isolated a substance similar to DEHP from corn oil. The US ATSDR
    (1988) quoted an US FDA survey of various foods in 1974 which showed
    that DEHP levels in most foods were less than 1 mg/kg; the foods
    surveyed included margarine, cheese, meat, cereal, eggs, milk, white
    bread, canned corn, corn meal, and baked beans.

         Ishida et al. (1981) collected and analysed chicken eggs
    available in Japanese markets. DEHP levels in egg white ranged from
    0.05 to 0.4 mg/kg, but no DEHP was detected in the egg yolk.

         In a study by Antonyuk (1975), DEHP migration from PVC materials
    into foodstuffs was noted following 7 days of contact.  Levels of 4-16
    mg DEHP/kg were detected in cheese, sausage, meat, flour, and rice
    while after 30 days levels of 30-150 mg/kg were found in sunflower
    oil. The permissible level of DEHP migration into foodstuffs was
    considered to be 2.0 mg/kg. Zitko (1972) detected DEHP concentrations
    in hatchery-reared juvenile Atlantic salmon of 13 to 16 mg/kg lipid;
    fish food contained 8 to 9 mg/kg lipid. When Williams (1973) analysed
    fish available to the Canadian consumer, only 6 out of 21 samples
    contained measurable amounts of DEHP. Levels of 0.1 and up to 0.16
    mg/kg were found in unprocessed eels and in processed canned tuna
    fish, respectively.

        Table 6.  Concentrations of DEHP in biota
    Organisms                            Location                Country           Concentration           Reference
    Mollusc (digestive gland)        Crouch Estuary, Essex     United Kingdom       9.2-214 ww        Waldock (1983)
    Dragonfly naiads                 Iowa (industrial)         USA                      200           Mayer et al. (1972)
    Commercial fish food                                       North America         2000-7000        Mayer et al. (1972)
    Channel catfish                  Mississippi & Arkansas    USA                     3200           Mayer et al. (1972)
    Channel catfish                  Iowa (industrial)         USA                      400           Mayer et al. (1972)
    Various fish species                                       Japan                  70-450          Kodama & Takai (1974)
    Various fish species                                       Japan               < 50-1800 ww       Kamata et al. (1978)
    Various fish species             various cities            Japan                  50-720          Goto (1979)
    Mainly fish                      Gulf of Mexico                                   < 1-135         Giam et al. (1978)
    Various fish species (liver)     Tees Bay                  United Kingdom       43-85.9 ww        Waldock (1983)
    Various fish species (muscle)    Tees Bay                  United Kingdom       13-51.3 ww        Waldock (1983)
    Walleye                          Lake Superior, Ontario    Canada                   800           Mayer et al. (1972)
    Tadpoles                         Iowa (industrial)         USA                      300           Mayer et al. (1972)
    Common seal pup (blubber)                                                        10 600 lw        Zitko (1972)
    a  ww = wet weight; lw = lipid weight
    5.1.7  Aquatic organisms

         Ray et al. (1983a) found DEHP levels of up to 490 µg/kg in
    sandworms  (Neanthes virens) and up to 170 µg/kg in clams collected
    from Portland, Maine, USA, but these levels did not seem to reflect
    the sediment levels and local pollutant sources.

         Musial & Uthe (1980) collected fish of various species from the
    Gulf of St Lawrence, Canada, and analysed them for DEHP in lipid
    extracts. They reported levels of up to 6.5 mg/kg on a wet weight
    basis (51.3 mg/kg on fat weight basis) in mackerel muscle and up to
    7.2 mg/kg (47.1 mg/kg fat weight) in herring muscle.  Lower levels of
    0.37 mg/kg (wet weight) were found in eels, and in both plaice and
    redfish concentrations were less than 0.001 mg/kg.

         Persson et al. (1978) collected aquatic organisms from the
    vicinity of a DEHP factory in Finland. Invertebrates contained up to
    0.1 mg DEHP/kg, and levels of 1.1 mg/kg in roach muscle and 2.3 mg/kg
    in pike liver were measured. Thuren (1986) analysed biota near to an
    industrial discharge point and reported levels of up to 5.3 mg/kg
    (fresh weight) in  Odonata sp and up to 14.4 mg/kg in  Asellus

         In Japan, DEHP levels in various species of fish in rivers and
    seas ranged from 0.01 to 19 mg/kg wet weight in 1974 (Environment
    Agency of Japan, 1989).

    5.1.8  Terrestrial organisms

         Persson et al. (1978) analysed soil arthropods collected near a
    DEHP factory in Finland and found residues of 2.8 mg/kg.

    5.2  General population exposure

         Few data are available on general population exposure.

         Based on an analysis of DEHP levels in various foods, the average
    exposure in the USA has been estimated to be around 0.3 mg/person per
    day and the maximum 2 mg/person per day (see section 5.1.6).

         In a survey of plasticizer levels in food-contact material and
    food, the United Kingdom Ministry of Agriculture, Fisheries and Food
    (MAFF, 1987) stated that DEHP has very limited use in food-contact
    material, and the maximum daily intake from food sources has been
    estimated to be less than 20 µg/person per day.

         DEHP found in human tissues may be derived from medical devices,
    since it was recognised by Trimble et al. (1966) and by Guess &
    Haberman (1968) that DEHP is leached from certain medical devices.
    Samples of soft PVC fluid bags containing normal saline and glucose
    (50 mg/ml) were shaken for 24 h and analysed for plastic additives 

    (Smistad et al., 1989). The PVC plastic materials contained DEHP,
    epoxidized vegetable oils, and stearates as the main additives. The
    same components were found in the solutions. MEHP was also detected,
    but only in the solutions.

         Marcel & Noel (1970) reported the presence of phthalate esters in
    human plasma that had been stored in plastic blood bags.

         Jaeger & Rubin (1972) found that DEHP was extracted from PVC
    plastic blood bags by human blood at the rate of 2.5 mg/litre per day
    at 4 °C. The DEHP was found in both lipid-containing and lipid-free
    fractions of plasma, whereas the red cells contained only minor
    amounts. Seven out of twelve samples of lung tissue, taken at autopsy
    from patients who had received transfusions of stored blood, contained
    DEHP at concentrations of 13.4-91.5 mg/kg (dry weight). Rubin & Nair
    (1972) reported that DEHP had also been found in the tissues and urine
    of patients who had not received blood transfusions, but no details
    were given. Mes et al. (1974) analysed human adipose tissue in Canada
    and found the levels of DEHP in most samples to range from 0.3 to 1.0

         Schneider et al. (1989) estimated that the highest exposure to
    DEHP associated with medical devices resulted from extracorporeal
    membrane oxygenation (ECMO), which could result in an exposure of 14
    mg/kg per day. In an infant receiving ECMO for 14 and 24 days, the
    DEHP serum levels were 26.8 and 33.5 mg/litre, respectively. In
    another infant, ECMO for 6 days resulted in liver, heart, and
    testicular concentrations of 3.5, 1.0, and 0.4 mg/kg, respectively.

         Newborn infants given exchange transfusions may have plasma
    levels of about 10 mg/litre (Sjöberg et al., 1985b).

    5.3  Occupational exposure during manufacture, formulation or use

         Few data on occupational exposure to DEHP have been reported.

         In a phthalate manufacturing plant in the USA producing DEHP from
    phthalic anhydride and alcohols, Liss et al. (1985) measured, in the
    case of six heavily exposed workers, 8-h TWA workplace air
    concentrations of DEHP ranging from 0.02 to 4.1 mg/m3. The exposure
    level for 44 other workers in the same plant was below the detection
    limit (10 µg/sample).

         In an Italian factory producing n-butyl phthalate, isobutyl
    phthalate, and DEHP, Gilioli et al. (1978) measured total phthalate
    exposure concentrations of between 1 and 60 mg/m3, the average being
    5 mg/m3.

         Nielsen et al. (1985) measured total phthalic acid esters in air
    in a PVC-processing plant in Sweden where diisodecyl phthalate, DEHP,
    and some butylbenzylphthalate were used. Concentrations of between

    0.01 and 2.0 mg/m3 were recorded in 96 2-h personal samples from 54

         Total phthalates concentrations in air of between 1.7 and 66
    mg/m3 were recorded in a PVC-processing plant in the USSR using
    mainly dibutyl phthalate and higher alkyl phthalates but also some
    DEHP and other phthalates (Milkov et al., 1973). Stankevich & Zarembo
    (1978) measured DEHP levels of 1.5 to 40 mg/litre in the blood of
    workers manufacturing PVC in the USSR.

         DEHP concentrations in air of between 0.09 and 0.16 mg/m3 were
    recorded in a German factory for phthalate production (Thiess et al.,
    1978a). In 9 PVC-processing plants in Finland, the mean concentration
    of DEHP ranged from less than 0.02 mg/m3 to 0.5 mg/m3 and the
    highest single value was 1.1 mg/m3 (Vainiotalo & Pfäffli, 1990).


    6.1  Absorption

    6.1.1  Inhalation

         There is no quantitative information available on the pulmonary
    route of absorption, although aerosols of DEHP are readily formed
    (Albro & Lavenhar, 1989).

    6.1.2  Dermal

         After a single application of 14C-labelled DEHP (61.5 mg/kg) to
    the back of F-344 rats, clipped one hour before treatment, urine and
    faeces were collected every 24 h for 7 days. The amount of 14C
    excreted was taken as an index of the percutaneous absorption, and
    about 5% of the dose given was excreted (Elsisi et al., 1989).

    6.1.3  Oral

         Studies with 14C-labelled DEHP indicated that at least 50% of
    the radioactivity of a single dose (2.9 mg/kg) was absorbed in the rat
    intestine since 42% and 14% were excreted in urine and bile,
    respectively, after 7 days (Daniel & Bratt, 1974). The same authors
    also found that DEHP was rapidly hydrolysed by pancreatic lipase,
    suggesting that DEHP is hydrolysed in the gut before absorption.  This
    was supported by the fact that no unmetabolized DEHP was found in
    liver after the administration of low oral doses (< 0.4 g/kg),
    although at higher doses (> 0.5 g/kg) DEHP was detected (Albro et
    al., 1982). At an oral dose level of 2 g/kg, the bioavailability of
    DEHP in rats, as measured in blood by HPLC, was 14%, whereas at an
    intraperitoneal dose level of 4 g/kg only 5% was recovered, again
    indicating a role for hydrolysis of DEHP in the gut (Pollack et al.,
    1985b). Using an inhibitor of mucosal esterases ( S,S,S-tributyl
    phosphorothionate), White et al. (1980) observed a marked inhibition
    in the uptake of DEHP by the gut.  Studies involving oral
    administration of MEHP indicated that this metabolite is well
    absorbed. After radiolabelled MEHP or DEHP was given to rats, the
    radioactivity recovered in plasma from MEHP was 16 times more than
    that from DEHP (Teirlynck & Belpaire, 1985).

         Oral administration of DEHP (1 g/kg) to young rats leads to a
    larger area under the plasma concentration-time curve (measured with
    gas chromatography) for MEHP (twice that of DEHP) than in older rats
    (Sjöberg et al., 1985a). This indicates either a more rapid hydrolysis
    of DEHP or a more efficient absorption of MEHP in young rats.

         Cynomolgus monkeys hydrolyse DEHP in the gut less efficiently
    than rats or mice (Astill, 1989). Rhodes et al. (1986) also noted that
    there is less absorption from the gastrointestinal tract in marmosets
    than in rodents.

    6.1.4  Intraperitoneal

         The systemic availability of DEHP was only 5% when a dose of 4
    g/kg was given intraperitoneally to rats. Relatively small amounts of
    MEHP were recovered in the blood in this study (Pollack et al.,

    6.2  Distribution

         Intravenously administered DEHP is rapidly eliminated from blood.
    This was demonstrated in experiments where radioactive DEHP was
    injected into male CFN rats and blood levels were determined by thin-
    layer chromatography (Schulz & Rubin, 1973).  At a low dose level (0.1
    mg/kg), there was an initial phase with a half-time of 4.5 min and a
    second phase with a half-time of 22 min. At a higher dose level (200
    mg/kg) the initial phase had a half-time of 9 min. This indicated that
    DEHP was taken up in a tissue compartment by a saturable process
    (Schulz & Rubin, 1973).  Radioactivity from DEHP was rapidly
    distributed to the liver, lungs, and spleen when administered
    intravenously (Schulz & Rubin, 1973; Daniel & Bratt, 1974).

         Orally administered DEHP is mainly distributed as MEHP in rats
    (Pollack et al., 1985b; Teirlynck & Belpaire, 1985).  Unmetabolized
    DEHP was recovered in the liver only after large oral doses
    (> 0.5 g/kg) were given, indicating a threshold phenomenon in the
    absorption and distribution (Albro et al., 1982; Agarwal, 1986). The
    distribution kinetics of MEHP have been analysed by Pollack et al.
    (1985b) and by Teirlynck & Belpaire (1985). Pollack et al. (1985b)
    found that the peak concentration of MEHP in blood was reached 15 min
    after oral or intraperitoneal administration of MEHP. The half-time of
    MEHP in blood or plasma in the rat is shorter than that of DEHP
    (Pollack et al., 1985b; Teirlynck & Belpaire, 1985). The  in vitro
    plasma protein binding of MEHP in the rat reaches approximately 98%
    (Sjöberg et al., 1985a).

         A phenomenon known as "shock-lung" has been reported to occur
    following intravenous administration of DEHP to rats and other
    species. Two hours after an emulsion of DEHP was given, between 13%
    and 48.6% of DEHP-radiolabelled material was found in the lungs of
    rats, as compared to 26.3-38.2% in the liver (Daniel & Bratt, 1974).
    This phenomenon may be relevant to human exposure via intravenous
    administration from bags and tubing containing DEHP.

         The DEHP plasma level in newborn infants given exchange
    transfusions may reach about 10 mg/litre (Sjöberg et al., 1985b). 
    This level is about twice as high as those found in leukaemia patients
    receiving platelet concentrations and about five times as high as
    levels found in haemodialysed patients. After treatment, this level
    falls rapidly to about 3 mg/litre within 2 h, and then there is a
    further drop with a half-time of about 10-12 h (Sjöberg et al.,

    6.3  Metabolism

         DEHP is hydrolysed  in vitro by pancreatic lipase to MEHP
    (Daniel & Bratt, 1974), indicating that this metabolism would occur
    mainly in the gut lumen. In rats about 80% of an oral dose of DEHP
    undergoes mono-deesterification (Pollack et al., 1985b), while intra-
    arterially administered DEHP is only slowly converted to MEHP (Pollack
    et al., 1985b). Studies on the hydrolysis of DEHP in homogenates from
    different organs (Table 7) indicate a very high activity in pancreatic
    juice and a comparatively low activity in liver (Albro & Thomas, 1973;
    Daniel & Bratt, 1974).

         At relatively high doses of DEHP (2 g/kg body weight per day),
    administered by gavage for up to 14 days, approximately 25-40% of the
    dose in rats and 50-75% of the dose in marmosets was excreted in the
    faeces. This implies that incomplete intestinal hydrolysis and
    absorption may occur (Rhodes et al., 1986).

         MEHP may in turn be metabolized in the gut wall (Pollack et al.,
    1985b) or in other organs. Rat liver cell cultures have been shown to
    convert MEHP to several metabolites, as occurs in the intact rat
    (Albro et al., 1973; Lhuguenot et al., 1985). Cultures of testicular
    cells were, however, apparently not able to metabolize MEHP beyond
    slight hydrolysis to phthalic acid within 18-24 h (Albro et al.,

         The metabolic pathways for MEHP are shown in Fig. 1. The omega-
    and omega-1-carbon oxidation products constitute more than 85% of the
    metabolites (Albro et al., 1973; Mitchell et al., 1985a; Lhuguenot et
    al., 1985). The ethyl side chain may also be oxidized (Lhuguenot et
    al., 1985). It has been suggested that omega-oxidation leads to a
    metabolite that is further degraded by ß-oxidation in the peroxisomes
    (Albro et al., 1973; Lhuguenot et al., 1985). Non-linear dose-
    dependency has been reported for these pathways in the rat; the
    predominance of omega-oxidation over omega-1-oxidation was increased
    by high doses of MEHP (Lhuguenot et al., 1985).

         The administration of 2-ethyl-(1-14C)-hexyl-labelled DEHP led
    to a low level of radioactivity being recovered from purified rat
    liver DNA (Albro et al., 1982). In a more recent study (Lutz, 1986),
    the administration of 14C-carboxylate-labelled DEHP resulted in no
    measurable radioactivity in DNA, whereas radioactivity was clearly
    measurable after the administration of DEHP that was 14C- or
    3H-labelled in the alcohol moiety.

        Table 7.  DEHP hydrolase activity of various tissue lipasesa
                                                          DEHP hydrolase activity
    Enzyme preparation                                                               
                                                     units/mg protein   units/g tissue
    Liver acid lipase, homogenate                           0.34               101
    Liver alkaline lipase, homogenate                       0.14                43
    Liver "lysosomal" concentrate (acacia)b                 8.80                 -
    Liver microsome + supernatant fraction, pH 8.2          1.22                 -
    Kidney acid lipase, homogenate                          0.15                45
    Lung homogenate (cholate)b                              0.072               15
    Lung homogenate (acacia)b                               0.10                21
    Mucosal homogenate, pH 7.4                              0.86                83
    Muscosal homogenate, pH 9.0                             0.43                41
    Pancreas homogenate                                    54.9             34 400
    Adipose "monoglyceride lipase"                          0.021             0.53
    Adipose "hormone-sensitive lipase"                      0.14              0.82
    Purified cholesteryl esterase                           0                    -
    a  From: Albro & Thomas (1973)
    b  14C-labelled DEHP supplied as dispersion in either sodium cholate or
       gum acacia
    FIGURE 1

         There are marked species differences in the metabolism of DEHP.
    Thus omega-oxidation seems to play a dominante role in the rat and
    guinea-pig (Albro et al., 1982; Lhuguenot & Elcombe, 1984; Lhuguenot
    et al., 1985), but to be a minor pathway in the mouse, hamster, green
    monkey, cynomolgus monkey, and marmoset (Albro et al., 1982; Lhguenot
    & Elcombe, 1984). In guinea-pigs there are few omega-1 metabolites of
    MEHP (Albro et al., 1982). This may have toxicological significance
    because certain omega-1-oxidation metabolites have been identified as
    active agents in peroxisomal proliferation in rat hepatocytes
    (Mitchell et al., 1985a).

         No conjugated metabolites were detected in the urine of
    DEHP-treated rats, but a minor portion was conjugated in the urine of
    hamsters (Albro et al., 1982; Lhuguenot & Elcombe, 1984). A major
    portion of glucuronide conjugates was found in the urine of the
    marmoset, mouse, guinea-pig, and green monkey, and in human urine
    (Albro et al., 1982). Albro (1986) reported that glucuronidation of
    DEHP metabolites was insignificant in rats. Studies in primates,
    including the African green monkey (Albro et al., 1981), marmoset
    (Rhodes et al., 1986), cynomolgus monkey (Short et al., 1987; Astill,
    1989), and man (Schmid & Schlatter, 1985), demonstrated that
    conjugation of DEHP can occur at the carboxylic acid moiety following
    a single ester hydrolysis.

         The level of unmetabolized MEHP excreted in urine also varies
    considerably between species; it is low in the rat and hamster, but
    high in the mouse, guinea-pig, green monkey, and man (Albro et al.,

         Repeated oral administration of DEHP or MEHP at high doses (500
    mg/kg) to rats leads to a change in the metabolic profile; there is an
    increase in omega-oxidized metabolites and a decrease in
    omega-1-oxidized metabolites (Lhuguenot et al., 1985). In rats given
    2% DEHP in the diet for one week, a 4-fold increase in peroxisomal ß-
    oxidation was found. ß-Oxidation of fatty acids induced by DEHP
    appears to occur via mitochondrial and peroxisomal pathways that are
    similar to normal pathways (Ganning et al., 1989). Drug-metabolizing
    enzyme activities have been studied after DEHP administration, and in
    some cases changes were observed (Walseth et al., 1982; Agarwal et
    al., 1982; Gollamudi et al., 1985; Pollack et al., 1989).

         The same metabolites as those found in rat urine can be detected
    in human urine. One study on intravenously injected DEHP (Albro et
    al., 1982) and one on orally administered DEHP (Schmid & Schlatter,
    1985) indicated that humans metabolize DEHP by omega- and
    omega-1-oxidation as well as by oxidation of the ethyl side chain.
    However, the omega-oxidation-pathway seems to be a minor pathway in
    man (Albro et al., 1982; Schmid & Schlatter, 1985). More than half of
    the metabolites recovered in human urine are conjugated metabolites
    (Albro et al., 1982; Schmid & Schlatter, 1985).

         Time-averaged concentrations of DEHP, MEHP, and phthalic acid in
    the blood of patients undergoing maintenance haemodialysis were 1.9,
    1.3, and 5.2 mg/litre, respectively (Pollack et al., 1985a). Such
    patients are considered to be at risk of potential DEHP toxicity
    through prolonged contact with medical plastic products that contain
    DEHP. The relatively high circulating level of phthalic acid may
    indicate an altered metabolism of DEHP in uraemic patients (Pollack et
    al., 1985a).

         The levels of DEHP and MEHP in plasma have been studied in
    newborn infants given blood exchange transfusions. In one case the
    MEHP half-life was the same as for DEHP (about 12 h), indicating that
    the hydrolysis of DEHP was the rate-limiting metabolic step. However,
    in other children the half-time of MEHP was longer than that of DEHP
    (Sjöberg et al., 1985b).

    6.4  Elimination and excretion

         Radioactivity from intravenously injected 14C-labelled DEHP is
    mainly recovered in urine and faeces after 24 h (Schulz & Rubin,
    1973), indicating that urine and bile are major excretory pathways.
    When a low dose level (0.1 mg/kg) was given to rats, 50-60% of
    injected radioactivity was recovered in urine and faeces after 24 h,
    whereas at a high dose level (200 mg/kg) less than 50% was recovered
    (Schulz & Rubin, 1973). Seven days after an oral dose (2.9 mg/kg) of
    DEHP was given to rats, 42% of the radioactivity was recovered in the
    urine and 57% in the faeces (Daniel & Bratt, 1974). Biliary excretion
    was also measured in these experiments, and it was found that 14% of
    the radioactivity was recovered in bile after 4 days (Daniel & Bratt,
    1974). The almost 100% recovery reported by Daniel & Bratt (1974) has
    been confirmed by Teirlynck & Belpaire (1985). Oral administration of
    MEHP (50-500 mg/kg) gave a higher urinary recovery than orally
    administered DEHP (50-500 mg/kg) as measured after 24 h (Lhuguenot et
    al., 1985).

         After the oral administration of non-radioactive DEHP (0.45
    mg/kg) to human volunteers, it was found that 15-25% was excreted in
    urine as MEHP or oxidized metabolites within 2-3 days (Schmid &
    Schlatter, 1985).

         In the rat no unmetabolized DEHP is excreted in the urine, but
    small amounts are found in mouse or green monkey urine (Albro et al.,
    1982). Major amounts of MEHP are excreted in mouse, guinea-pig, green
    monkey, and human urine (Albro et al., 1982). However, oxidized
    metabolites, either free or conjugated, constitute a major portion of
    excretion products in rat, mouse, hamster, green monkey, and human
    urine (Albro et al., 1982).

         Changes in excretion pathways have been observed after prolonged
    administration of DEHP. After oral dosing of rats without
    pre-treatment with DEHP, the faecal excretion pathway dominated, while

    in rats fed with DEHP for 7 days the urinary pathway dominated (Daniel
    & Bratt, 1974).

    6.5  Retention and turnover

    6.5.1  Half-life and body burden

         After the intravenous administration of radiolabelled DEHP, at
    least two elimination phases of radioactivity, with short half-lives
    (4.5-9 and 22 min, respectively), were observed in rat blood (Schulz
    & Rubin, 1973). After 7 weeks of oral administration, the elimination
    phase in the liver was considerably slower, the half-life being 3-5
    days (Daniel & Bratt, 1974). No accumulation of DEHP or MEHP was
    observed when the dosage was 2.8 g/kg per day for 7 days (Teirlynck &
    Belpaire, 1985), nor was there any in a long-term (5-7 weeks) feeding
    study at a dose level of 1 or 5 g/kg diet (corresponding to a daily
    dose of about 50 and 250 mg/kg body weight) (Daniel & Bratt, 1974).

    6.5.2  Indicator media

         Analysis of the total amount of urinary metabolites, measured as
    derivatized phthalic acid, indicate a weak positive correlation
    between occupational exposure to phthalate and the presence of
    metabolites in the urine (Nielsen et al., 1985; Liss et al., 1985). In
    the study by Nielsen et al. (1985), workers were exposed mainly to
    DEHP and diisodecyl phthalate, and the urinary level of phthalate
    ester metabolites rose from the background level (17 µmol/litre) to
    23-25 µmol/litre. In the study by Liss et al. (1985), workers were
    exposed to DEHP and phthalic anhydride. Urinary phthalate
    concentrations in exposed workers more than doubled after a workshift
    and levels up to 44 µmol/litre were recorded. The authors concluded
    that phthalic anhydride influenced the urinary level more than DEHP.
    Phthalic acid is not a specific marker for DEHP exposure.


    7.1  Single exposure

         Numerous LD50 values have been reported for DEHP. Oral LD50
    values generally exceed 25 g/kg in rats and 30 g/kg in mice
    (Stankevich et al., 1984; NIOSH, 1985b; Woodward et al., 1986); in the
    rabbit it is 33.9 g/kg (Shaffer et al., 1945) and in the guinea-pig
    26.3 g/kg (Krauskopf, 1973). Dermal LD50 values for guinea-pigs and
    rabbits of 10 g/kg and 25 g/kg, respectively, have been reported
    (NIOSH, 1985b).

         The LD50 values after intraperitoneal administration were
    30.7 g/kg in rats (Shaffer et al., 1945) and 14-75 g/kg in mice
    (Lawrence et al., 1975; Woodward et al., 1986). LD50 values in the
    range of 200-250 mg/kg were reported for the rat after intravenous
    administration of DEHP solubilized with a nonionic detergent (Schmidt
    et al., 1975; Rubin & Chang, 1978).

         The main symptom of DEHP toxicity after single oral or
    intraperitoneal dosing is diarrhoea (Hodge, 1943). An intraperitoneal
    dose of 500 mg/kg in rats decreased spontaneous running activity,
    thereby indicating behavioural changes (Rubin & Jaeger, 1973). After
    intravenous dosing, lung lesions including oedema, haemorrhage, and
    infiltrations of polymorphonuclear leucocytes were observed in rats at
    doses as low as 50 mg/kg (Schulz et al., 1975). The etiology of the
    lung lesions is unknown.  It has, however, been suggested that some
    changes could be due to the release of lysosomal enzymes from alveolar
    macrophages, which was found to occur  in vitro in rabbit alveolar
    macrophages cultured with DEHP (Bally et al., 1980).

         Rabbits treated intravenously with 350 mg DEHP/kg showed a
    decrease in blood pressure and an increase in breathing rate. No
    deaths occurred after doses up to 650 mg/kg were administered (Calley
    et al., 1966).

         The monoester, MEHP, may be more toxic than the diester but data
    are very limited. In a short note by Villeneuve et al. (1978), the
    oral LD50 of MEHP was reported to be 1.34 g/kg in female rats and
    1.8 g/kg in males.

    7.2  Short-term exposure

         Doses of 3.4 g/kg body weight per day given by gavage (in olive
    oil) for periods of up to 90 days caused the death of 15 out 20 rats
    (Nikonorow et al., 1973). However, no deaths were reported among rats
    fed 3% DEHP in the diet (1.9 g/kg body weight) for 90 days (Shaffer et
    al., 1945) or in a rat study of the US National Toxicology Program
    after dietary dosing (< 50 g/kg) for 14 days (NTP, 1982).

         Oral administration of DEHP at a rate of > 0.4 g/kg body
    weight per day resulted in a weight gain decrease in rats within a few
    days (Nikonorow et al., 1973). In a 17-week feeding study where rats
    were given 2, 10 or 20 g DEHP/kg diet, a decreased body weight was
    observed (Gray et al., 1977). Reduction in body weight was also
    observed in rats given dietary levels of 12.5 or 25 g/kg for 13 weeks.
    Dosages of 1.6-6.3 g/kg resulted in either slight elevations of body
    weight or no effect (NTP, 1982).

         MEHP given at a level of 6.4 g/kg diet caused reduction in the
    body weight gain of rats (Chu et al., 1981). No effects on body weight
    occurred at dietary levels of 0.625 g/kg given for 3 months, but a
    significant decrease in blood glucose was observed.

         Reductions in haemoglobin, packed cell volume and erythrocyte
    numbers were observed in rats given 10 or 20 g DEHP/kg diet for 17
    weeks, but not when they were given 2 g/kg for the same period (Gray
    et al., 1977).

         Cystic kidneys and centrilobular necrosis were noted in one
    strain of mice (ddY) fed 2.5 or 25 g DEHP/kg for 2 weeks, but not in
    another strain (B6C3F1), even with a higher exposure level and a
    longer exposure period (Woodward et al., 1986).

         DEHP administered intravenously at a rate of 25-500 mg/kg per day
    for 2-4 weeks to beagle dogs resulted in pulmonary haemorrhage and
    inflammatory response similar in appearance to the "shock-lung" effect
    (Woodward et al., 1986).

         In an inhalation study, Wistar rats were exposed in a head-nose
    inhalation system to DEHP aerosols of respirable particle size. 
    Exposure duration was 6 h/day, 5 days/week for 4 weeks at target
    concentrations of 0, 0.01, 0.05, and 1.0 mg/litre. A statistically
    significant increase in relative lung weights was found in the males
    given the highest dosage, and this was accompanied by foam cell
    proliferation and thickening of the alveolar septa (Klimisch et al.,

         Another inhalation study has been reported, but this is
    inadequate for assessment (Timofievskaja et al., 1980).

         A discussion of the effects of DEHP on the liver is given in
    section 7.9.

         When rats were treated with DEHP by intraperitoneal injection
    (Walseth et al., 1982) or by repeated oral dosing (Agarwal et al.,
    1982), an increase in cytochrome P-450 levels was observed.  The
    increase in hepatic microsomal oxidation appears to be primarily due
    to the deesterification products of DEHP, i.e. MEHP and
    2-ethoxyhexanol, in long-term exposure, whereas DEHP and its two
    metabolites may inhibit microsomal oxidation after acute exposure to

    DEHP (Pollack et al., 1989). However,  in vitro rat liver microsomal
    cytochrome P-450 levels were not affected by DEHP (Gollamudi et al.,

         Liver mitochondrial enzymes and mitochondrial morphology have
    been reported to be influenced by DEHP administration (Ohyama, 1977;
    Shindo et al., 1978). Recent results suggest that the  in vitro
    effects of DEHP (> 20 µmol/litre) on mitochondrial functions are
    mainly related to the action on membrane lipids surrounding the
    adenine nucleotide translocator, which reduces the rate of adenine
    nucleotide exchange across the mitochondrial membrane (Kora et al.,

         In male rats given DEHP in the diet, the urinary excretion of
    zinc was enhanced and the testicular level of zinc decreased (Gray et
    al., 1982; Oishi, 1985) (section 7.5). These changes in zinc
    homeostasis could be due to altered levels of metallothionein. In mice
    fed 6 or 12 g DEHP/kg diet for 24 weeks, hepatic levels of
    metallothionein were increased up to 11-fold (Waalkes & Ward, 1989).

         Several studies on rats have shown that DEHP given in the diet
    (5-20 g/kg) decreases plasma triglyceride and cholesterol levels
    (Yanagita et al., 1978; Sakurai et al., 1978; Bell et al., 1978a; Bell
    et al., 1978b; Bell et al., 1979; Yanagita et al., 1979; Curstedt &
    Sjövall, 1983). DEHP inhibits the biosynthesis of cholesterol, an
    effect which is accompanied by phospholipidosis, and the same effects
    have been observed with MEHP (Oishi & Hiraga, 1982).

    7.3  Long-term exposure

         In a 24-month study by Harris et al. (1956), three groups of
    Wistar rats each comprising 43 males and 43 females were fed diets
    containing 0, 1, and 5 g DEHP/kg, interim kills being made at 3, 6,
    and 12 months. At the end of the study, only two control, four
    low-dose, and seven high-dose animals were alive. During the first
    year the body weights of the high-dose group were slightly reduced,
    but by the second year the body weights of all groups were similar.
    During the first 6 months an increase in relative liver and kidney
    weights was seen in DEHP-treated animals but later they were similar
    in control and treated animals. After 3 months of treatment, one out
    of eight rats in the low-dose group was found to have mild renal
    tubular atrophy. After 6, 12, and 24 months of treatment, no
    compound-related pathological changes were evident. Because of high
    mortality due to disease, this study is difficult to validate.

         In another 24-month study (Carpenter et al., 1953), groups of
    Sherman rats consisting of 32 males and 32 females were given diets
    containing 0, 0.4, 1.3 or 4 g DEHP/kg. Owing to reduced life
    expectancy due to disease and the small numbers of animals used, the
    study was inadequate for assessing the chronic toxicity of DEHP.

         In a 12-month study by Nikonorow et al. (1973), a group of 20
    male and 20 female Wistar rats was given a diet containing 3.5 g
    DEHP/kg, and a control group received the diet without DEHP.  The only
    gross or micropathological change noted in exposed animals at necropsy
    was hepatomegaly. During the study, however, about 30% of the animals
    died due to congestion of the small intestine and loss of the gastric
    and/or intestinal mucosa, which was complicated by purulent pneumonia
    and endometritis.

         Crocker et al. (1988) described the renal effects of DEHP given
    by gavage to young male rats at a dosage of 2.14 mg/kg body weight
    three times per week for up to 12 months. A 50% reduction in
    creatinine clearance and an increase in the severity of renal cyst
    formation was observed. This lesion was consistent with spontaneous
    nephropathy commonly observed in old rats; exposure may cause an onset
    in younger rats. Furthermore, DEHP fed at 6 and 12 g/kg diet for two
    years did not produce renal lesions in male and female F-344 rats
    (NTP, 1982).

         In a 2-year study (NTP, 1982; Kluwe et al., 1982), groups of
    F-344 rats were given dietary levels of 0, 6, and 12 g DEHP/kg.
    Decreased body weight in exposed groups was noted from week 30 until
    the cessation of exposure. In addition to neoplastic effects (section
    7.7) and testicular atrophy (section 7.5), a compound-related
    hypertrophy of cells in the male anterior pituitary was noted in the
    high-dose group. In both exposed groups an increased incidence of
    clear changes in liver cells was observed.  This study also
    investigated B6C3F1 mice exposed to 3 or 6 g DEHP/kg diet. A
    dose-related decrease of body weight was observed in female mice.
    There was no increased incidence of non-neoplastic lesions except for
    seminiferous tubular degeneration in the testes of male mice (section

         In a study on groups of male and female guinea-pigs fed diets
    containing 0, 0.4 or 1.3 g DEHP/kg for 12 months (Carpenter et al.,
    1953), the body weight of the low-dose animals was statistically
    higher than that of controls, and liver weight relative to body weight
    of dosed females slightly increased. No other exposure-related lesions
    were found.

         A study on groups of male ferrets fed diets containing 0 and 10
    g DEHP/kg for 14 months revealed a reduction in body weight and an
    increase in relative liver weight, but no evidence of peroxisome
    proliferation (Lake et al., 1976).

    7.4  Skin and eye irritation; sensitization

         DEHP has been shown to be a weak irritant to mammalian skin when
    administered topically or intradermally (0.2 ml of an emulsion of
    100 g/litre) (Calley et al., 1966; Woodward et al., 1986).

         In a study by Lawrence et al. (1975), no irritation occurred when
    undiluted DEHP was instilled into the eye of rabbits.

         No data are available on the potential for DEHP to induce skin
    sensitization in animals.

    7.5  Reproduction, embryotoxicity, and teratogenicity

    7.5.1  Reproduction

         The effects of DEHP on male reproductive organs have been studied
    extensively. The majority of the studies have been carried out on rats
    or mice given DEHP in the diet.

         Seminiferous tubular atrophy, comprising a loss of spermatids and
    spermatocytes, occurred when 4-week-old Wistar rats were given 2800 mg
    DEHP/kg by oral intubation for 10 days (Gray & Butterworth, 1980). In
    similarly treated 10-week-old rats, about 50% of the tubules were
    atrophic and the remainder unaffected.  However, no testicular damage
    was detected in treated 15-week-old rats. When 20 g DEHP/kg was given
    in the diet (approximately 1200 mg DEHP/kg per day) to 4-week-old
    rats, the lesions produced were reversible whether treatment stopped
    before or continued until after the control rats had reached sexual

         In rats given 10 or 20 g DEHP/kg diet, the testis atrophy was
    dose dependent after approximately 2 weeks of feeding. This atrophy
    was accompanied by pituitary changes, i.e. enlargement and
    vacuolization of the basophils of the pars distalis, corresponding to
    the formation of the so-called castration cells seen after gonadectomy
    (Gray et al., 1977). In a subsequent study, there was a reduction in
    testicular and prostatic zinc levels concomitant with increased
    urinary excretion of zinc (Gray et al., 1982).

         In a rat study by Oishi & Hiraga (1980a), the serum testoster-one
    levels in rats fed 20 g DEHP/kg diet were reduced by approximately
    50%. The total amount per testis decreased, but the concentration rose
    to 150% of the original value because of the testicular atrophy.
    Simultaneous administration of testosterone or zinc had no protective
    effect on the atrophy but did prevent the weight reduction of the sex
    organs such as the epididymis (Gray & Butterworth, 1980; Oishi &
    Hiraga, 1983). In a further study, it was found that a low-zinc diet
    aggravated the DEHP-induced testicular atrophy (Agarwal et al.,

         DEHP given to 7 young (5 weeks old) Wistar rats in the diet
    (20 g/kg) for one week decreased the testicular weight significantly
    (P < 0.05), compared to controls, and also the testicular zinc
    concentration (Oishi, 1984). In a study by Oishi (1985), groups of 20
    male rats were given DEHP (2.0 g/kg per day) by gavage for 14 days.
    Ten rats were then killed and the remaining ten were kept for 45 days

    on a DEHP-free diet. The histopathological changes of the testes seen
    on day 15 were marked shrinkage of the seminiferous tubules, a
    germinal epithelium consisting only of Sertoli cells, and very few
    spermatogonia. After 45 days the percentage of spermatogenic tubules
    had increased from 0 (at day 15) to 12.8%, indicating a limited
    reversibility of the testicular atrophy.

         Similar degeneration of the seminiferous tubules was observed
    when 13-week-old Wistar rats were orally given 2 g DEHP/kg body weight
    for 7 consecutive days (Saxena et al., 1985) .

         In a dietary study, DEHP was administered to F-344 rats at 6 and
    12 g/kg for 103 weeks (NTP, 1982). Degeneration of the seminiferous
    tubules was observed only at the higher dose.

         The testicular changes induced by oral DEHP administration appear
    to be age dependent. A daily dosage of 2.8 g DEHP/kg body weight given
    for 10 days caused seminiferous tubular atrophy in 4-week-old Wistar
    rats, but this effect was less severe in 10-week-old rats and absent
    in 15-week-old rats (Gray & Butterworth, 1980). Sjöberg et al. (1986)
    showed that DEHP fed at a level of 1.7 g/kg diet caused reductions in
    testis weight to 20%, 55%, and 92% of the control values in
    Sprague-Dawley rats at 25, 40, and 60 days of age, respectively. This
    was apparently not due to differences in pharmacokinetic parameters,
    since the plasma levels of the metabolite MEHP were identical in the
    25- and 40-day-old rats (Sjöberg et al., 1982).

         Rats given DEHP intraperitoneally at a dosage of 100 mg/kg per
    day for 5 days did not develop testicular atrophy (Curto et al.,
    1982). However, this lack of response was probably a consequence of
    the dose used, rather than the dose route, and there was a 30%
    reduction in testicular zinc. Intraperitoneal administration of higher
    doses (1-25 g/kg per day for 5 days) to Wistar rats decreased serum
    testosterone levels (Oishi & Hiraga, 1979).

         Some degenerated primary spermatocytes and altered Sertoli cells
    were observed in Sprague-Dawley rats given 3-h intravenous infusions
    of an emulsion at a rate of 1 ml/h, which corresponded to a daily dose
    of 500 mg DEHP/kg (Sjöberg et al., 1985c). The infusions were given
    every other day on six occasions. The emulsion contained DEHP,
    fractionated egg yolk phosphatides, glycerol, and water. No effects
    were observed when emulsions corresponding to 0, 5 or 50 mg DEHP/kg
    were given.

         In a study by Curto & Thomas (1982), groups of sexually mature
    Swiss-Webster mice were given intraperitoneal injections of 50 or 100
    mg DEHP/kg either daily for 5 days or alternate daily for 20 days (10
    injections). The animals were killed 24 h after the last injection. No
    significant alterations in testicular weight or zinc levels occurred.

         Young rats were not more susceptible to testicular damage than
    older ones following intravenous infusion of DEHP. It has been
    suggested that the age-related difference observed in some studies may
    be due to the fact that the rate of gastrointestinal absorption of the
    DEHP-derived metabolite MEHP is higher in younger animals than in
    older ones (Sjöberg et al., 1986).

         In a NTP-sponsored study (Melnick et al., 1987), CD-1 mice given
    3 g DEHP/kg diet showed significantly diminished testis and epididymis
    weights compared to controls. In addition, the sperm concentration in
    the cauda epididymis was reduced and the percentage of abnormal sperm
    in the cauda was significantly higher in the treated mice than in the

         A high oral dose (4.2 g/kg) of DEHP gave minimal tubular atrophy
    in hamsters but did not produce any effects on urinary zinc excretion,
    testicular zinc levels or testicular weights (Gray et al., 1982).

         The effects of the monoester MEHP have not been as well studied
    as those of DEHP. An oral dosage of 1 g MEHP/kg per day for 5 days
    produced a significant decrease in rat testis weight and extensive
    testicular atrophy (Gray et al., 1982). On the other hand, rats given
    intraperitoneal doses of up to 100 mg MEHP/kg daily for 5 days showed
    no abnormal histology (Curto et al., 1982) and an intraperitoneal dose
    of 50 mg/kg given on alternate days for 20 days produced only a
    reduction in prostatic zinc levels (Curto & Thomas, 1982).

         Mice fed diets containing 20 g MEHP/kg for 1 week revealed
    markedly reduced testicular zinc and testosterone levels, but there
    were no reductions in testicular weight (Oishi & Hiraga, 1980b). 
    Hamsters given MEHP (1 g/kg per day) for 9 days showed more severe
    testicular effects than those given DEHP at a level of 4.2 g/kg per
    day for the same period (Gray et al., 1982). More recent studies
    (Albro et al., 1989) indicate that testicular atrophy resulting from
    DEHP exposure is most probably due to the formation of MEHP.

         These studies indicate that the rat is the species most
    susceptible to DEHP-induced testicular atrophy. The mechanism of
    phthalate-induced testicular damage is not fully understood. 
    Testicular zinc depletion has been suggested to be a primary event
    (Foster et al., 1982, 1983). However, Gray et al. (1982) showed that
    an effect on zinc level is not always associated with testicular
    atrophy in all species tested. Zinc is essential for normal testicular
    function and its depletion is known to lead to testicular atrophy
    (Barney et al., 1969). Inhibition of dehydrogenase enzymes, e.g.,
    those controlling the biosynthesis of testosterone, leads to reduced
    testosterone levels. DEHP administration has been shown to reduce the
    level of serum testosterone in the rat (Oishi & Hiraga, 1979; Oishi &
    Hiraga, 1980a) as well as in the mouse (Gray et al., 1982), although
    in the mouse no testicular atrophy was observed.  Administration of
    testosterone or zinc did not prevent the testicular damage induced by

    DEHP (Gray & Butterworth, 1980; Oishi & Hiraga, 1983).
    Co-administration of DEHP and testosterone, on the other hand,
    apparently enhanced the testicular damage caused by DEHP (Oishi,
    1989a). In a similar study (Oishi, 1989b), luteinizing
    hormone-releasing hormone (LRH) significantly decreased testis weight,
    sulfhydryl content, and lactate dehydrogenase when given together with
    DEHP, whereas LRH or DEHP alone had no effects.

         Similar effects were observed with exogenously added follicle
    stimulating hormone (FSH) in an  in vitro study. Primary testicular
    cell cultures pretreated with MEHP showed a dose-related reduction in
    FSH-stimulated cyclic adenosine monophosphate (cAMP) production (Lloyd
    & Foster, 1988). These results suggest that MEHP produces a
    perturbation at the level of the FSH membrane receptor, causing an
    inhibition of FSH action.

          In vitro studies have indicated that the Sertoli cell is the
    target cell. Mixed cultures of Sertoli and germ cells prepared from
    rat testes were exposed to DEHP or MEHP (10-7-10-4 mol/litre)
    (Gray & Beamand, 1984; Gray & Gangolli, 1986). DEHP had no effect, but
    MEHP caused a dose-dependent increase in the rate of germ cell
    detachment from Sertoli cells, accompanied by changes in Sertoli cell
    morphology. More recent data indicate that Sertoli cell mitochondria
    are a target for MEHP (Chapin et al., 1988).  However, it has also
    been shown that testicular mitochondrial respiratory functions are
    decreased in rats given 2 g DEHP/kg body weight by gavage (Oishi,

         In a study by Melnick et al. (1987), CD-1 mice were given 0, 0.1,
    1 or 3 g DEHP/kg diet during a 7-day pre-mating period and a
    subsequent 98-day cohabitation period. There was complete suppression
    of fertility in the 3-g/kg group and a significant reduction in
    fertility in the 1-g/kg group, compared to controls, but no effect on
    fertility at 0.1 g/kg.

         In a fertility study designed to investigate earlier significant
    results (Singh et al., 1972), groups of male ICR mice were dosed
    subcutaneously with undiluted DEHP at levels of 1, 2, 5, and 10 ml/kg
    (equivalent to 0.99, 1.97, 4.93, and 9.86 g/kg) on days 1, 5, and 10
    of the study (Agarwal et al., 1985a). They were subsequently mated
    with virgin females on days 2, 6, 11, 16 and 21, and then at weekly
    intervals until week 8. There were reductions in the incidence of
    pregnancies in several groups, but these were dose related only at the
    first mating (i.e. on day 2 of the experiment). Examination of
    pregnant mice on day 13 of gestation showed that there were
    pre-implantation losses corresponding to a DEHP dose level of 10 ml/kg
    and a mating on day 6. There were consistent increases in early fetal
    deaths relating to matings at day 21, week 4, and week 5. In a similar
    study, male and female mice were treated subcutaneously with 1-100 ml
    DEHP/kg, and this led to a reduction in testicular but not ovarian
    weight. The ovaries exhibited histological injury at lower doses of

    DEHP than the testes. Unlike the situation in the testes, there was no
    significant dose-related increase in histopathological changes in the
    ovaries (Agarwal et al., 1989).

         In an investigation of the effects of phthalates on female
    reproductive organs, three doses of 4.93 g DEHP/kg were given
    intraperitoneally at 5-day intervals to female rats (Seth et al.,
    1976). No histopathological changes in the ovaries were seen 22 days
    after the first injection, but reductions in the activities of some
    enzymes were noted.

         The administration of DEHP (1000 mg/kg intraperitoneal or 2000
    mg/kg oral) to marmosets daily for 14 days did not lead to testicular
    atrophy (Rhodes et al., 1986).

         Hence, it appears that rats and guinea-pigs are sensitive to
    DEHP-induced testicular atrophy, while mice are fairly resistant and
    hamsters and marmosets are highly resistant. The fact that at least
    some of the effects associated with this atrophy can be produced
     in vitro argues against a hormonally mediated indirect effect. The
    earliest effects are seen in Sertoli cells and are described as
    vacuolation (Albro, 1987).

    7.5.2  Embryotoxicity and teratogenicity

         In a study by Nikonorow et al. (1973) on female Wistar rats given
    0.34 or 1.7 g DEHP/kg by gavage during the first 21 days of gestation,
    the only untoward effect was a reduction in fetal body weight. When
    DEHP (0, 5, 10, 15 or 20 g/kg) was administered orally to Fischer-344
    rats on gestational days 0 to 20, maternal toxicity and reduced fetal
    body weight per litter were observed at the three highest dose levels.
    The number of fetuses per litter was unaffected by the treatment (Tyl
    et al., 1988).

         Intraperitoneal injections of 4.93 or 9.86 g DEHP/kg on days 5,
    10, and 15 of gestation resulted in an increase in the number of
    resorptions and reduced fetal weight in Sprague-Dawley rats (Singh et
    al., 1972). In the highest-dose group, gross abnormalities, such as
    twisted hind legs and anophthalmia, were noted but no skeletal defects
    were observed.

         Rat plasma soluble extracts of two PVC plastics containing DEHP
    were administered intravenously to groups of pregnant Sprague-Dawley
    rats daily from the 6th to the 15th day of gestation, and the animals
    were killed at day 20 (Lewandowski et al., 1980). The daily doses of
    DEHP were equivalent to 1.3 mg/kg and 5.2 mg/kg. No significant
    teratogenic or embryotoxic effects were noted.

         Groups of ICR mice were given DEHP in the diet at levels of 0.5
    to 10 g/kg for the first 18 days of gestation and were then sacrificed
    (Shiota et al., 1980; Shiota & Nishimura, 1982). The food intake was

    an average of 7 g/day. At the 4 g/kg and 10 g/kg dose levels, no live
    fetuses were found. At 2 g/kg, 40% of the fetuses had malformations
    including exencephaly, spina bifida, and malformed tail. Delayed
    ossification was seen in about 15% of the fetuses at 1 g/kg and
    2 g/kg.

         In a study by Tyl et al. (1988), DEHP was administered in the
    diet to CD-1 mice on the first 17 days of gestation at levels of 0,
    0.25, 0.5, 1, and 1.5 g/kg. At the two highest dose levels maternal
    toxicity, increased resorptions and late fetal deaths, decreased
    numbers of live fetuses, and reduced fetal body weight per litter were
    observed. The number and percentage of malformed fetuses per litter
    were elevated at the three highest dose levels.

         A single oral administration of DEHP (0.1 ml/kg) on day 7 of
    gestation to ddY-strain SPF (specific pathogen free) mice decreased
    the number and the body weights of live fetuses (Tomita et al.,
    1982b). In a study by Yagi et al. (1980), DEHP was given orally to SPF
    mice on days 6, 7, 8, 9 or 10 of gestation. When 5.0 or 10.0 ml/kg was
    given on day 7 there were no live fetuses, whereas 2.5 ml/kg
    administered on the same day resulted in 14% live embryos and 1.0
    ml/kg gave 40% live embryos. The percentages of live embryos when 10.0
    ml/kg was given on day 8, 9 or 10 of gestation were 18, 92, and 95%,
    respectively. Gross and skeletal abnormalities occurred in fetuses
    given 2.5 and 7.5 ml/kg on day 7 or 8. The abnormalities included
    exencephaly, open eyelid, and club-foot.

         In a study by Shiota & Mima (1985), groups of ICR mice were given
    DEHP by stomach intubation on days 7, 8, and 9 of gestation. The DEHP
    doses (in olive oil) were 250, 500, 1000, and 2000 mg/kg, and the mice
    were sacrificed on day 18. In the two highest dose groups, the numbers
    of resorptions and malformed fetuses were significantly increased.
    Fetal weights were significantly depressed. The most frequent
    malformations were anencephaly and exencephaly. When doses of up to
    8000 mg/kg were given by intraperitoneal injections on days 7,8, and
    9 of gestation, no effects were noted. DEHP is highly embryotoxic and
    terato-genic in mice when given orally but not when given

         The monoester MEHP, at oral doses of 225, 450, and 900 mg/kg per
    day, produced significant signs of maternal toxicity when given to
    pregnant Wistar rats on days 6-15 of gestation (Ruddick et al., 1981).
    In the highest dose group a 73% mortality was observed in the dams.
    There was a dose-related decrease in the number of litters and in the
    litter weights of live pups.

         Oral dosing with 0.1 or 1.0 g MEHP/kg on day 7 of gestation led
    to increased incidence of early embryonic deaths in SPF mice (ddY
    strain), but dosing on day 8 or 9 had less effect (Yagi et al., 1980;
    Tomita et al., 1986). The fetuses had reduced body weight, and there
    was a higher incidence of gross abnormalities, compared to controls,

    when the higher dose level was given on day 8 or 9.  The mice dosed on
    day 8 produced fetuses with a high incidence of skeletal effects.

         Intravenous injections of MEHP (11.38 mg/kg) to rabbits on days
    6-17 of gestation gave a high incidence of resorptions (Thomas et al.,
    1979). The incidence of fetal anomalies was similar to that in

         When Nikonorow et al. (1973) administered DEHP (0.34 or 1.7 g/kg
    per day) to female Wistar rats by gavage for 3 months prior to mating,
    there was an increase in the number of resorptions but no effects on
    fetal weights or the incidence of skeletal anomalies. 

          In utero administration of DEHP (1000 mg/kg body weight) to
    rats daily from days 6 to 15 of gestation resulted in retardation of
    fetal growth and an increase in fetal liver weight. There were
    significant quantities of DEHP and decreased mitochondrial enzyme
    activities in the fetal liver. These results indicate an effect on
    fetal liver cell bioenergetics (Srivastava et al., 1989).

         The mechanism of DEHP or MEHP teratogenicity is not known.
    Teratogenic activity could result from zinc deficiency, which is known
    to produce such effects (Swenerton & Hurley, 1971).

    7.6  Mutagenicity and related end-points

         The possible genotoxic effect of DEHP has been thoroughly
    investigated in several different short-term tests. The effects of the
    major metabolites of DEHP, i.e. MEHP and 2-ethylhexanol, as well as
    phthalic acid and phthalic anhydride, have also been studied.

    7.6.1  Mutation

         Studies on the possible mutagenic effect of DEHP have been
    performed in bacteria, fungi, and in cultured mammalian cells.
     Drosophila melanogaster has also been used and results from a few
     in vivo studies on mice have been reported.  Bacteria

         Many studies have been performed using a variety of strains of
     Salmonella typhimurium and DEHP doses of up to 10 mg/plate. 
    Incubations both with and without exogenous activation systems have
    been performed. S9 mix from rats induced by Aroclor 1254 has
    frequently been used, but other species and other inducers have also
    been used to produce metabolic activation systems. With one exception
    (Tomita et al., 1982a) these test results have all been negative
    (Kirby et al., 1983; Yoshikawa et al., 1983; Zeiger et al., 1985;
    Agarwal et al., 1985b), and in a IPCS collaborative study (Ashby et
    al., 1985) all five laboratories reported negative results.  Bacteria

    other than  S. typhimurium have also been used; negative results were
    obtained with E. coli WP2 at doses of up to 2 mg per plate (Yoshikawa
    et al., 1983).

         The major metabolites of DEHP have also been tested for mutagenic
    activity in bacteria. Concentrations of up to 3.333 mg per plate for
    MEHP and phthalic anhydride (Zeiger et al., 1985) and 2 mg/plate for
    2-ethylhexanol and phthalic acid (Agarwal et al., 1985b) have yielded
    negative results in strains of Salmonella (see also Kirby et al.,
    1983; Yoshikawa et al., 1983). However, Tomita et al. (1982a) reported
    a significant increase in TA100 revertants following exposure to
    either DEHP or MEHP (both with and without S9). It should be noted,
    however, that MEHP mutagenicity is only demonstrable within a narrow
    range of concentration, because it has a sterilizing effect at high
    concentration and shows no mutagenic activity at low concentration.
    Tomita et al. (1982a) also detected dose-dependent (0.4 and 0.5 mg
    MEHP/plate) DNA damage in a  B. subtilis Rec assay, while DEHP,
    phthalic acid, and 2-ethylhexanol were all negative.  In this study
    MEHP also gave a positive result in  E. coli WP2 b/r.

         Negative results were obtained when pooled urine from rats,
    treated with 2000 mg DEHP/kg per day for 15 days, was tested for
    genotoxic activity. A direct plating procedure was used with
     S. typhimurium strains TA98, TA100, TA1535, TA1537, and TA1538, both
    with and without S9 and ß-glucuronidase/aryl sulfatase as the
    activation system. When 2-ethylhexanol was tested according to the
    same protocol, the result was also negative (Divincenzo et al., 1985).  Fungi

         The induction of mutations by DEHP has been studied in various
    fungal species. In the IPCS collaborative study on  in vitro assay
    systems (Ashby et al., 1985), DEHP was considered to be negative in
    six out of seven assays. Positive results were obtained with
     Saccharomyces cerevisiae, both with and without S9 activation, at
    lowest effective concentrations of 1541 mg/litre and 3081 mg/litre,
    respectively. However, other laboratories using different strains of
     S. cerevisiae or  Schizosaccharomyces pombe reported negative
    results at a maximum tested concentration of 5000 mg/litre.  Mammalian cells

         Mouse lymphoma cells (L5178Y), Chinese hamster V79 cells, and
    human lymphoblasts have been used to study the mutagenic effect of
    DEHP in cultured mammalian cells. Several investigators obtained
    negative results, but a few positive results have also been reported.

         In the IPCS collaborative study (Ashby et al., 1985), only one
    out of ten investigators reported a positive response. Mouse lymphoma
    cells were exposed to DEHP without S9, and two concentrations (7.5 and
    20 mg/litre) gave positive results. In a separate study (Kirby et al.,

    1983), where MEHP, 2-ethylhexanol, and DEHP were tested in the mouse
    lymphoma cell assay, all three substances were found to be
    non-mutagenic. The concentrations used were 0.016-1.0 ml/litre
    (without S9) and 0.067-5.0 ml/litre (with S9) for DEHP, and 0.013-1.0
    ml/litre for MEHP and 2-ethylhexanol.  Drosophila

         DEHP has also been tested for mutagenicity in  Drosophila
     melanogaster using the sex-linked recessive lethal (SLRL) test and
    various somatic recombination and mutation assays. SLRLs were not
    induced by DEHP (20 µg/g) administered by injection (Yoon et al.,
    1985). Negative results for DEHP were also reported in the SLRL
    mutation assay on  Drosophila melanogaster larvae (Zimmering et al.,

         In the IPCS study (Ashby et al., 1985), DEHP gave a positive
    response in the unstable eye mosaic test at a dose of 6.1 g/litre, in
    two separate experiments, but neither lower nor higher concentrations
    produced any response. No activity was seen in the wing spot test with
    a single dose of 6.1 g/litre. It was concluded that DEHP exhibits
    marginally positive mutagenicity (Ashby et al., 1985).

    7.6.2  DNA damage

         Various end-points, such as unscheduled DNA synthesis (UDS) and
    single strand breaks, have been used to detect DNA damage induced by
    DEHP in a variety of mammalian test systems.  In the IPCS study (Ashby
    et al., 1985), negative results were obtained when single strand
    breaks were measured, either by alkaline elution in hepatocytes (up to
    3.907 g/litre) or alkaline sucrose sedimentation in Chinese hamster
    ovary (CHO) cells (up to 39 g/litre). UDS in either isolated
    hepatocytes or cultured HeLa cells was investigated by four different
    laboratories. One investigator detected a positive response using
    isolated hepatocytes, but, since this result was only statistically
    significant at one dose and not dose related, the consensus was that
    DEHP does not cause UDS.

         In a study by Butterworth et al. (1984), DEHP did not induce DNA
    repair in primary rat hepatocytes. Similarly, neither DEHP nor MEHP
    induced any DNA repair in primary human hepatocytes from three
    different subjects. In this study, concentrations as high as 0.01 mol
    DEHP/litre and 5 x 10-4 mol MEHP/litre were used and exposure
    continued for 18 h. No induction of DNA repair or increased alkaline
    elution of DNA was seen in hepatocytes from either female or male
    F-344 rats treated with DEHP  in vivo. UDS was not induced in male
    rats treated with 500 mg DEHP/kg by gavage 2, 12, 24 or 48 h before
    sacrifice.  Furthermore, treatment of male rats with 150 mg/kg per day
    by gavage for 14 days and treatment of female rats with dietary DEHP
    (12 g/kg) for 30 days resulted in peroxisome proliferation, but no UDS
    was induced.

         Similar  in vivo/in vitro results in studies of UDS in
    hepatocytes were reported by Kornbrust et al. (1984). No UDS was
    observed in primary rat hepatocytes exposed  in vitro to 10-5 to
    10-2 mol DEHP/litre or  in vivo by a single gavage dose of 5 g/kg
    2, 15 or 24 h prior to the isolation of hepatocytes. A dietary
    concentration of 20 g DEHP/kg led to a marked proliferation of
    peroxisomes after 4 weeks. Neither this treatment nor the additional
    administration of a single gavage dose of 5 g/kg (15 h before
    sacrifice) to animals fed the 20-g/kg diet for either 8 weeks or for
    4 weeks with or without pretreatment with 3-amino-1,2,4-triazole (to
    inhibit endogenous catalase activity) induced any detectable DNA
    repair in hepatocytes. Indeed, the administration of DEHP and some
    other proliferators has been shown to increase the levels of
    8-hydroxy-deoxyguanosine in rat liver DNA (Takagi et al., 1990).

    7.6.3  DNA binding

         In an  in vivo study by Albro et al. (1982), radioactivity from
    carbonyl-labelled DEHP did not associate with purified protein, RNA or
    DNA from rat liver. Label from 2-ethyl-(1-14C)-hexyl-labelled DEHP
    or MEHP appeared to associate strongly with purified DNA, but this was
    not the case with label from free 14C-labelled 2-ethylhexanol.
    According to Albro (1987), although there is incorporation into normal
    nucleosides, there is no evidence of alkylation.

         In a similar study (von Däniken et al., 1984; Lutz, 1986), DEHP
    radiolabelled in different positions was administered orally to female
    F-344 rats with or without pretreatment with unlabelled DEHP (10 g/kg
    diet) for 4 weeks. The administration of 14C-carboxylate-labelled
    DEHP resulted in no measurable DNA radioactivity, whereas
    radioactivity was clearly measurable after the administration of DEHP,
    14C- or 3H-labelled in the alcohol moiety, or of
    2-ethyl(1-14C)hexanol. HPLC analysis showed that the normal
    nucleosides had incorporated radiolabel, but fractions expected to
    contain carcinogen-modified nucleoside adducts did not contain any

         DNA isolated from the livers of male F-344 rats administered 2000
    mg DEHP/kg daily by gavage for 3 days was analysed for possible
    carcinogen-DNA adducts by the 32P-postlabelling technique (Gupta et
    al., 1985). No adducts were detected in the DNA, which also was the
    case when DNA from hepatocytes exposed to 10-3 mol DEHP/litre
     in vitro for 4 h was analysed.

    7.6.4  Chromosomal effects

         Chromosomal effects of DEHP have mainly been studied  in vitro,
    although some studies on the induction of micronuclei in the
    peripheral blood erythrocytes of mice have been published.

         DEHP did not induce any increase in the level of sister-chromatid
    exchange (SCE) in Chinese hamster ovary (CHO) cells, treated for 1 h,
    either with or without S9, at levels of up to 0.01 mol/litre (Douglas
    et al., 1986). On the other hand, MEHP has been reported to induce SCE
    in V79 cells treated with 25 or 50 mg/litre for 24 h or 1500 mg/litre
    for 3 h (Tomita et al., 1982a). This metabolite also induced
    chromosomal aberrations in CHO cells and RL4 cells (from rat liver),
    but only at cytotoxic concentrations in the CHO cells (1.0 and 1.3
    mmol/litre, with or without S9). MEHP was less toxic to RL4 cells
    and concentrations between 2.0 and 5.0 mmol/litre gave a dose-related
    increase in chromosomal aberrations (Phillips et al., 1986). According
    to the authors, observations on changes in CHO cell structure and
    permeability and on the haemolytic effects of phthalate monoesters
    suggest that MEHP cytotoxicity may be due primarily to an action on
    cell membranes.

         The induction of aneuploidy by DEHP was investigated both in
    mammalian cells and in fungi in the IPCS study (Ashby et al., 1985).
    The mammalian assays gave positive responses at 50 mg/litre in a
    fibroblast cell line from Chinese hamster liver and at levels of
    between 25 and 50 mg/litre in Chinese hamster primary liver cells. Two
    out of four studies using fungi were also positive and the consensus
    was that DEHP is capable of inducing aneuploidy  in vitro in both
    fungi and mammalian cells.

         In a study by Ahmed et al. (1990), male Wistar rats were fed for
    alternate 7-day periods with a diet containing 20 g DEHP/kg and a
    control diet. The rats were examined after 3 days on the DEHP diet or
    after 7 days on the control diet. An analysis of nuclear size gave
    results consistent with an increase in tetraploid hepatocytes after
    treatment with DEHP, which was reversed when the rats returned to the
    control diet.

    7.6.5  Cell transformation

         DEHP-induced cellular transformation has been studied in several
    different experimental systems. In a test programme (Astill et al.,
    1986), the BALB/3T3 cell transformation assay was used both with and
    without rat primary hepatocytes. Both DEHP (0.875-1 µl/litre) and the
    two metabolites MEHP and 2-ethylhexanol gave negative results. On the
    other hand, the majority of transformation tests in the IPCS study
    (Ashby et al., 1985) were positive for DEHP. Negative results were
    obtained with BALB/c-3T3 cells, while a study measuring the
    enhancement of viral transformation of Syrian hamster embryo (SHE)
    cells was considered to be inconclusive. Positive responses were
    obtained by four other investigators using SHE cells at 1-300 mg/litre
    (two different laboratories), embryonic mouse fibroblasts at 1000
    mg/litre with S9 and 10 mg/litre without S9, or retrovirus-infected
    Fischer rat embryo cells at 2000 mg/litre (the highest dose tested).
    In a separate study (Tomita et al., 1982a), both DEHP (7.5 and 15
    g/kg) and MEHP (375 and 750 mg/kg) induced morphological

    transformation, as well as chromosomal aberrations, in SHE cells after
    transplacental administration.

         In a study by Diwan et al. (1985), anchorage-independent growth
    of JB6 mouse epidermal cells was enhanced by DEHP at concentrations of
    500 to 20 000 ppm/ml (the Task Group noted that the unit used in
    Diwan's paper is ppm/ml, but it should read ppm or mg/litre). Ward et
    al. (1986) used the same model system and reported that both DEHP
    (1.3-51 x 10-6 mol/litre) and MEHP (2-5 x 10-8 mol/litre) were
    effective, while 2-ethylhexanol (4-77 x 10-7 mol/litre) was without
    effect. The morphological transformation in the SHE cell system
    induced by DEHP, MEHP, and other hepatic peroxisome proliferators
    seemed not to be correlated with increased peroxisomal ß-oxidation,
    increased production of oxidative radicals or peroxisome proliferation
    (Mikalsen et al., 1990a; Mikalsen et al., 1990b; Mikalsen et al.,

         A few studies on DEHP-induced inhibition of metabolic
    cooperation, which may be indicative of the promoting potential of a
    substance, have been reported. Metabolic cooperation in Chinese
    hamster V79 cells was not inhibited by DEHP at non-cytotoxic
    concentrations, i.e. 3 x 10-4 mol/litre (0.12 mg/litre) or less
    (Kornbrust et al., 1984). In the IPCS study (Ashby et al., 1985), one
    investigator reported inhibition in V79 cells at non-cytotoxic
    concentrations of DEHP (25-200 mg/litre in two separate experiments
    and 5-25 mg/litre in another), while another investigator, using V79
    cells in a microassay method, detected a slight (but non-significant)
    increased inhibition at doses of between 10-5 and 2 x 10-4
    mol/litre (3.9-78 mg/litre).

         In a study by Malcolm & Mills (1989) on V79 cells, DEHP inhibited
    intercellular communication (gap junctions) at non-cytotoxic
    concentrations (10-30 mg/litre).

    7.6.6  In vivo effects

         Putman et al. (1982) dosed male Fischer-344 rats orally for 5
    days with DEHP (5.0, 1.7, and 0.5 g/kg per day), MEHP (0.14, 0.05, and
    0.01 g/kg per day), and 2-ethylhexyl (0.21, 0.07, and 0.02 g/kg per
    day), and bone marrow metaphase cells were examined. No significant
    increases were observed in gap breaks or structural rearrangements. In
    addition, the mitotic index was unaffected by treatment.

         A micronucleus assay on mouse (B6C3F1) peripheral blood
    erythrocytes (intraperitoneal doses of 0.6, 3.0, and 6.0 g/kg per day
    for 5 days), sampled at 0, 2, and 4 weeks after the last treatment,
    yielded negative results (Douglas et al., 1986). A sperm morphology
    assay in B6C3F1 mice and Sprague-Dawley rats at the same dose levels
    also gave negative results (Douglas et al., 1986).  However, Agarwal
    et al. (1986b) reported increases in sperm morphology changes in adult
    F-344 rats given 20 g DEHP/kg for 60 consecutive days.

         Two studies have yielded negative results in dominant lethal
    tests (Hamano et al., 1979; Rushbrook et al., 1982). Hamano et al.
    (1979) administered MEHP and DEHP orally, and the observation period
    was six weeks. Rushbrook et al. (1982) dosed ICR/SRM mice orally for
    5 days with DEHP (2465, 4930, and 9860 mg/kg per day), MEHP (50, 100,
    and 200 mg/kg per day) or 2-ethylhexanol (250, 500, and 1000 mg/kg per
    day). Each male was mated weekly with virgin females for 8 consecutive
    weeks, and females were evaluated for pregnancy, live fetuses, and
    early and late fetal deaths. All data were within the normal ranges.

         DEHP and its major metabolites have also been tested for their
    potential to induce micronuclei. Exposure to 0.6, 3.0 or 6.0 g DEHP/kg
    per day for 5 days over a 4-week period did not induce micronuclei in
    the peripheral blood erythrocytes of B6C3F1 male mice (Douglas et
    al., 1986). Negative results were also obtained in another mouse
    micronucleus test after either a single or daily doses of 5 g DEHP/kg.
    In this study MEHP and 2-ethylhexanol also gave negative results
    (Astill et al., 1986).

    7.7  Carcinogenicity

         In a carcinogenicity study (NTP, 1982; Kluwe et al., 1982),
    groups of 50 male and 50 female Fischer-344 rats were fed diets
    containing 6 or 12 g DEHP/kg diet, and 50 male and 50 female B6C3F1
    mice were fed diets containing 3 or 6 g DEHP/kg for 103 consecutive
    weeks. Concurrent controls (50 of each sex and species) were fed a
    diet without the addition of DEHP. Food and water were supplied
     ad libitum. All animals were given the control diet for 1-2 weeks
    after 103 weeks of treatment and were then killed and examined both
    grossly and microscopically. The administered concentrations of DEHP
    were estimated to be equal to, or one half of, the maximum tolerated
    doses. Under the conditions studied, DEHP caused an increased
    incidence of hepatocellular carcinomas in female rats and male and
    female mice, and an increased incidence of hepatocellular carcinomas
    and neoplastic nodules in female rats (Table 8). Twenty of the 57
    hepatocellular carcinomas in the DEHP-treated mice (sexes and doses
    combined) had metastasized to the lung. The figure of nine
    hepatocellular carcinomas in control male mice is considered to be
    within the normal range (NTP, 1982).

         The reported decreased incidence of tumours of the thyroid,
    pituitary, and testis could be related to an increased endocrine
    activity of the pituitary gland (NTP, 1982).

         The carcinogenicity of DEHP in rats has been confirmed in two
    further studies. Rao et al. (1990) found a 78.5% incidence in a group
    of 14 male Fischer-344 rats fed a diet containing 20 g DEHP/kg for up
    to 108 weeks, whereas the incidence in the 10 controls was 10%. Popp
    et al. (1987) found either hepatocellular carcinomas or neoplastic
    nodules in 6 out of 20 animals after female Fischer-344 rats were
    exposed for 2 years to a diet containing 12 g/kg.

        Table 8.  Carcinogenic effects of DEHP on the livera
                              Control   Low dose   High dose   P valueb
    Hepatocellular carcinoma

    Male rats                  1/50        1/49      5/49      < 0.05
    Female rats                0/50        2/49      8/50      < 0.005
    Male mice                  9/50       14/48     19/50      < 0.05
    Female mice                0/50        7/50     17/50      < 0.0001

    Neoplastic nodules

    Male rats                  2/50        5/49      7/49      n.s.
    Female rats                0/50        4/49      5/50      < 0.05

    Hepatocellular adenoma

    Male mice                  6/50       11/48     10/50      n.s
    Female mice                1/50        5/50      1/50      n.s
    a  From: NTP (1982)
    b  Probability level in Codiran-Armitage test for linear trend when
         P < 0.05; otherwise not significant (n.s.)
         Two other long-term studies on DEHP have been performed by
    Carpenter et al. (1953) and Harris et al. (1956), but, due to the
    small numbers of animals used, the studies are inadequate to assess
    the carcinogenic potential.

         DEHP has been investigated in two life-time studies on Syrian
    hamsters (Schmezer et al., 1988). In one study, groups of 25 male and
    25 female 6-week-old hamsters were assigned to each of six groups:
    untreated control; 3 g DEHP/kg body weight given intraperitoneally
    once every week for 18 weeks; the same dose once every two weeks for
    18 weeks; the same dose once every four weeks for 32 weeks; the same
    dose once every four weeks for 32 weeks plus  N-dimethylnitrosamine
    (NDMA) at 1.67 mg/kg body weight given orally once per week; and the
    same dose of NDMA without DEHP treatment. NDMA increased the tumour
    rate for malignant liver tumours, mainly haemangioendotheliomas.
    Co-administration of DEHP with NDMA neither increased nor decreased
    the tumour rates. No significant differences of tumour rates were
    observed in groups treated with DEHP alone compared with controls.

         Schmezer et al. (1988) also investigated the life-time, whole-
    body exposure of Syrian hamsters to DEHP vapour alone in air or in
    combination with orally administered NDMA. The low doses of DEHP
    (15 µg/m3, resulting in a total exposure of about 7.5 µg/kg body
    weight) in this study render it inadequate for the assessment of the
    long-term effects of DEHP alone. In combination with NDMA, this low

    level of DEHP was associated with highly significant (P < 0.001)
    decreases in hepatic haemangioendothe-lioma and fibrosarcomas combined
    in males and females combined. This unexpected result should be
    further investigated to evaluate its significance.

    7.8  Special studies

         Since DEHP lacks genotoxic activity in most test systems, it has
    been suggested that the carcinogenic effect is exerted during the
    promotion phase of hepatocarcinogenicity. DEHP has therefore been
    tested in several initiation/promotion experiments in rats and mice
    where the end-point has been the number and/or volume of altered liver
    cell foci. As expected, DEHP lacked initiating activity in these
    studies (Ward et al., 1986; Popp et al., 1987).

         DEHP appears to be a tumour promotor in mouse liver. In male
    B6C3F1 mice given an intraperitoneal initiating dose of
    diethylnitrosamine (80 mg/kg), DEHP at levels of 6 and 12 g/kg diet
    caused accelerated growth of foci and increased incidence of
    hepatocellular adenomas (Ward et al., 1983; Ward et al., 1986). 
    However, in a 6-month study, DEHP at a level of 12 g/kg diet did not
    appear to promote altered foci in male F-344 rats given an
    intraperitoneal initiating dose of diethylnitrosamine of 150 mg/kg
    (Popp et al., 1987). In other promotion studies DEHP appeared to
    accelerate the regression or inhibit the appearance of some kinds of
    foci in rats (DeAngelo & Garrett, 1983; DeAngelo et al., 1984). 
    However, other studies have shown that peroxisome proliferators can
    promote the development of altered foci and tumours in rat liver.
    These results are somewhat different from those of other liver tumour
    promotors with respect to the histological phenotype of the foci and
    the relatively low frequency of the foci (Cattley & Popp, 1989). Thus,
    further studies are required to demonstrate that DEHP can promote
    altered foci in rat liver.

    7.9  Mechanisms of hepatotoxicity

         Together with a number of structurally diverse chemicals,
    including certain hypolipidaemic drugs, herbicides, and chlorinated
    solvents, DEHP has been shown to produce hepatic peroxisome
    proliferation in rats and mice (Reddy & Lalwani, 1983; Lock et al.,
    1989). This proliferation is accompanied by liver enlargement,
    stimulation of replicative DNA synthesis and cell division, and the
    induction of peroxisomal and microsomal fatty acid oxidizing enzyme
    activities (Reddy & Lalwani, 1983; Marsman et al., 1988; Lock et al.,
    1989). The increase in microsomal fatty acid oxidation is due to the
    induction of a cytochrome P-450 IVA1 (Lock et al., 1989). Peroxisome
    proliferation and induction of peroxisomal and microsomal fatty acid
    oxidizing enzyme activities can be observed in primary hepatocyte
     in vitro cultures, as well as in the intact animal (Elcombe &
    Mitchell, 1986; Lake et al., 1986). Both  in vivo and  in vitro
    studies with rat hepatocytes have demonstrated wide compound potency

    differences (Reddy et al., 1986; Barber et al., 1987; Lake et al.,
    1988). Compared to some other compounds, DEHP is not considered to be
    a particularly potent peroxisome proliferator in rodent liver (Reddy
    et al., 1986; Barber et al., 1987). Possible mechanisms of peroxisome
    proliferation include a perturbation of hepatic lipid metabolism
    and/or the presence of a receptor protein (Lalwani et al., 1987; Lock
    et al., 1989; Issemann & Green, 1990).

         Certain peroxisome proliferators, including DEHP, have been shown
    to increase the incidence of liver tumours in rats and mice (Reddy &
    Lalwani, 1983; Reddy et al., 1986; Butterworth et al., 1987).
    Generally, there is a good correlation between the potency of a
    compound to produce peroxisome proliferation in rat and mouse liver
    and its hepatocarcinogenic properties (Reddy et al., 1986). However,
    the magnitude of peroxisome induction does not necessarily always
    correlate with the carcinogenic response when different compounds are
    compared (Marsman et al., 1988). Since the peroxisome proliferators
    known to date are non-genotoxic carcinogens, Reddy and coworkers
    (Reddy & Lalwani, 1983; Rao & Reddy, 1987; Reddy & Rao, 1989) have
    suggested that liver tumour formation arises from a sustained
    oxidative stress to the hepatocytes due to an imbalance in the
    production and degradation of hydrogen peroxide (H2O2). DEHP
    administration results in increased peroxisomal production of H2O2
    in hepatocytes (Tomaszewski et al., 1986). There is an imbalance in
    H2O2 production and degradation due to the fact that catalase is
    induced to a much lesser extent than peroxisomal ß-oxidation enzymes.
    In addition, the level of cytosolic glutathione peroxidase is reduced
    on prolonged exposure to DEHP (Lake et al., 1987; Marsman et al.,
    1988; Conway et al., 1989b; Tamura et al., 1990). Treatment with
    peroxisome proliferators also results in a reduction of superoxide
    dismutase and glutathione  S-transferase activities (Ciriolo et al.,
    1982; Goel et al., 1986; Lake et al., 1987). It has been suggested
    that the increased level of H2O2 in hepatocytes, either directly or
    via other reactive oxygen species (e.g., hydroxyl radical), damages
    intracellular membranes and/or DNA (Reddy & Rao, 1989). Indeed chronic
    administration of DEHP and other peroxisome proliferators results in
    increased lipid peroxidation and lipofuscin deposition in rat
    hepatocytes (Mitchell et al., 1985b; Goel et al., 1986; Cattley et
    al., 1987; Lake et al., 1987; Reddy & Rao, 1989; Conway et al.,
    1989b). Oxygen radicals may damage DNA (Bridges, 1985), leading to
    increased levels of 8-hydroxydeoxy-guanosine and other modified bases
    in DNA (Reddy & Rao, 1989).  Indeed, the administration of DEHP and
    some other peroxisome proliferators has been shown to increase levels
    of 8-hydroxyde-oxyguanosine in rat liver DNA (Kasai et al., 1989;
    Takagi et al., 1989, 1990, 1991). Apart from a role in peroxisome
    proliferation, leading to oxidative stress, recent studies on DEHP
    have demonstrated a role in the stimulation of replicative DNA
    synthesis in the hepatocarcinogenicity of peroxisome proliferators
    (Butterworth et al., 1987; Smith-Oliver & Butterworth, 1987; Marsman
    et al., 1988). Increased cellular division may result in spontaneous
    mutational events or promotional effects (Bridges, 1985; Butterworth

    et al., 1987), including the promotion of spontaneously initiated
    cells found in the livers of aging rodents (Schulte-Hermann et al.,
    1989). A comparative study of the hepatic effects of Wy-14643
    ([4-chloro-6 (2,3-xylidino) 2-pyrimidinylthio]acetic acid), a potent
    peroxisome proliferator, and DEHP indicated that, although both
    compounds produced a similar induction of peroxisome proliferation at
    the dose levels administered, only Wy-14643 produced liver tumours
    within one year (Marsman et al., 1988; Conway et al., 1989b). However,
    analysis of these data demonstrates that Wy-14643 produces both a more
    marked increase in lipofuscin deposition (presumably reflecting
    oxidative stress) and a sustained stimulation of replicative DNA
    synthesis. These data suggest a role for both oxidative stress and
    increased cell replication in the hepatocarcinogenicity of DEHP and
    other peroxisome proliferators.

         The mechanism of hepatocarcinogenicity induced by peroxisome
    proliferators could include both enhanced cell replication and DNA
    damage induced by oxygen radicals. Thus, at dose levels that do not
    produce either significant peroxisome proliferation or cell
    replication, it is unlikely that DEHP or other peroxisome
    proliferators would elicit hepatic tumour formation.  In this context
    data on threshold values for liver effects in sensitive species (e.g.,
    the rat and mouse), together with an assessment of species differences
    in response (see below), should be considered.  The value for the
    subchronic liver effects of DEHP (e.g., peroxisome proliferation and
    DNA synthesis) in the rat appears to be around 50 mg/kg per day (Lake
    et al., 1984, 1991; Mitchell et al., 1985b; Tomaszewski et al., 1986).
    Furthermore, a two-year study of DEHP in female rats demonstrated no
    effects at 0.3 g/kg diet, a value which corresponds well to that noted
    above (Cattley et al., 1987; Popp et al., 1987). 

         Many studies have shown dramatic species differences in response
    to peroxisome proliferators including DEHP (ECETOC, 1992). For
    example, DEHP threshold values for peroxisome proliferation of 25 and
    250 mg/kg, respectively, were found in rats and hamsters after 14 days
    of daily gastric intubation (Lake et al., 1984). Several other studies
    showing species differences in the response to DEHP have been
    reported. One experiment demonstrated a large increase in hepatic
    peroxisomes after 14 days of oral DEHP administration (2000 mg/kg per
    day) to rats, but the same treatment of marmosets resulted in no
    peroxisome proliferation (Rhodes et al., 1986). A limited study in
    cynomolgus monkeys also revealed no evidence of peroxisome
    proliferation due to DEHP (Short et al., 1987). In addition, DEHP did
    not elicit peroxisome proliferation in guinea-pigs (Osumi & Hashimoto,
    1978; Mitchell et al., 1985a). Some studies have indicated that
    intrinsic species differences in hepatocellular sensitivity exist. For
    example, MEHP and MEHP metabolite VI (mono(2-ethyl-5-oxohexyl)
    phthalate), which is a proximate peroxisome proliferator in the rat,
    were good peroxisome proliferators in cultured rat hepatocytes, but
    had little or no effect in guinea-pig, marmoset or human hepatocytes
    (Mitchell et al., 1985a; Elcombe & Mitchell, 1986). More recently,

    Bichet et al. (1990) and Butterworth et al. (1989) have also
    demonstrated the inability of MEHP to elicit peroxisome proliferation
    in human hepatocyte cultures. Studies using a variety of other
    peroxisome proliferators (e.g., clofibric acid, beclobric acid,
    methylclofenapate, trichloroacetic acid, ciprofibrate, and
    benzbromarone) have also failed to demonstrate peroxisome
    proliferation in cultured human hepatocytes, in spite of a good
    response in rat hepatocytes (Elcombe, 1985; Allen et al., 1987;
    Elcombe & Styles, 1989; Butterworth et al., 1989; Bichet et al., 1990;
    Blaauboer et al., 1990).

         Species differences in cytochrome P-450 IVA1 induction and
    stimulation of replicative DNA synthesis have been less well studied.
    However, MEHP and metabolite VI did not induce P-450 IVA1-mediated
    lauric acid hydroxylation  in vivo in marmosets (Rhodes et al., 1986)
    or in cultured marmoset or human hepato-cytes (Elcombe & Mitchell,

         In conclusion, it appears that the livers of rats and mice are
    exquisitely sensitive to peroxisome proliferators, including DEHP,
    while those of guinea-pigs, monkeys, and humans show minimal or no
    response (ECETOC, 1992).


    8.1  General population exposure

         Two adults who swallowed 5 or 10 g DEHP experienced no untoward
    effects apart from mild gastric disturbances and moderate diarrhoea
    with 10 g DEHP (Shaffer et al., 1945). Three cases of nonspecific
    hepatitis were described among 27 haemodialysis patients with terminal
    renal failure. The PVC blood tubing used released DEHP at a
    concentration of 10-20 mg/litre of perfusate. The symptoms and signs
    of hepatitis disappeared rapidly when the use of tubing that did not
    contain DEHP was resumed (Neergaard et al., 1971).

         In three pre-term infants artificially ventilated with PVC
    respiratory tubes, unusual lung disorders (opacification of the lung)
    were observed during the fourth week of life. It was assumed by the
    authors that these lung disorders were causually related to the
    exposure to DEHP released from the respiratory tubes (Roth et al.,

    8.2  Occupational exposure

         There are very few data on the effects of occupational exposure
    specifically to DEHP.

         Two studies reported symptoms and signs of polyneuropathy among
    47 out of 147 workers at a PVC-processing plant in the USSR and 12 out
    of 23 workers at a plant for phthalate production in Italy. The
    workers were exposed to a mixture of phthalates and DEHP was a minor
    constituent, at least in the USSR plant.  Furthermore, tricresyl
    phosphate (a neurotoxin) was a component of the incombustible
    materials produced in 10-20% of machines assigned to various workers
    (Milkov et al., 1973). In the Italian study there was no corresponding
    unexposed control group and the authors concluded that no definite
    conclusion could be drawn from the study because of the limited number
    of workers examined (Gilioli et al., 1978). The total phthalate air
    concentrations recorded varied between 1.7 and 66 mg/m3 in the USSR
    and 1 and 60 mg/m3 in Italy (Milkov et al., 1973; Gilioli et al.,

         In a study involving a Swedish PVC-processing factory, peripheral
    nervous system symptoms and signs were investigated among 54 male
    workers exposed mainly to DEHP, diisodecylphthalate, and some
    butylbenzylphthalate. The workers were divided into three groups of
    approximately equal size and with mean phthalate exposures of 0.1,
    0.2, and 0.7 mg/m3. Some workers displayed various peripheral
    nervous system symptoms and signs, but these were not related to the
    level of exposure.  None of the workers reported symptoms indicating
    work-related obstructive lung disease. Conventional lung function
    tests also showed no association with exposure (Nielsen et al., 1985). 
    However, several biochemical parameters showed significant

    associations with exposure. There was a slight decrease in the
    haemoglobin level with longevity of employment as well as with
    exposure in the last year. The serum ý-1-antitrypsin level increased
    slightly with length of employment and the serum immunoglobulin A
    level rose with rising exposure during the last year (Nielsen et al.,

         One case of occupational asthma due to DEHP was reported in
    worker at a PVC-processing plant (Brunetti & Moscato, 1984). 
    Diagnosis was made by exposing the patient to DEHP vapour in an
    inhalation chamber; this evoked a dual asthmatic reaction. The action
    was inhibited by prior administration of sodium chromoglycate.

         A study of blood lipids, serum activities of liver enzymes, and
    routine haematological tests was carried out among workers at a German
    plant for DEHP production. The results were negative and uninformative
    due to very low exposure levels (below 0.16 mg/m3) and the lack of
    a control group (Thiess et al., 1978b).  Thiess & Flieg (1978)
    investigated the frequency of chromosome aberrations in 10 workers who
    were employed from 10 to 30 years in this DEHP production plant. The
    exposure levels were very low (0.09-0.16 mg/m3), and there was no
    increase in chromosome aberrations compared to the control group. A
    mortality study of 221 workers exposed to DEHP in the same plant was
    also conducted. Eight deaths occurred in the cohort compared with
    expected values of 15.9 and 17.0 from the city and county data,
    respectively (Thiess et al., 1978a).


    9.1  Toxicity to microorganisms

         Bringmann & Kühn (1980) calculated toxic thresholds from cell
    multiplication inhibition tests and found thresholds of > 400
    mg/litre for the bacterium  Pseudomonas putida, 10 mg/litre for the
    alga  Scenedesmus quadricaudaia, and 19 mg/litre for the protozoan
     Entosiphon sulcatum.

         Mathur (1974b) reported that DEHP added to soil (3 g/kg)
    inhibited respiration, whereas addition of the same amount of DEHP to
    soil preincubated for 14 weeks with DEHP or dioctyl phthalate had no
    effect on respiration rate. The experiments were conducted at 22 °C
    since no degradation of DEHP occurred at 4 °C or 10 °C. In an
    intermittent flow-through hydrosoil microcosm, DEHP (at 1 and 100
    mg/litre) produced no significant effect on the numbers or
    physiological activity of the microflora monitored, nor on any of the
    overall activities that were studied (Mutz & Jones 1977).

         Larsson et al. (1986) studied the effect of DEHP on the microbial
    activity in sediments. Sediment cores were taken, together with the
    overlying water, from a eutrophic lake and the sediment was spiked
    with between 25 and 400 mg DEHP/kg at a level of 5 mm below the
    surface. Uncontaminated sediment samples had a significantly higher
    oxygen uptake than sediment containing DEHP. Microbial activity, as
    estimated from decreased oxygen saturation in the soil column,
    correlated inversely with increasing levels of DEHP in the sediment.

    9.2  Toxicity to aquatic organisms

         Many of the toxicity values given in this section are above the
    water solubility of DEHP, which ranges from 0.3 to 0.4 mg/litre (see
    section 2.2). DEHP adsorbs strongly to sediment, food, dissolved
    organic carbon, and even the testing vessels. Many of the toxicity
    tests are also based on nominal concentrations. Therefore, the actual
    exposure of the organisms is very difficult to determine and care must
    be taken when interpreting the results.

    9.2.1  Invertebrates

         Acute toxicity to aquatic invertebrates is summarized in Table 9.

        Table 9.  Toxicity of DEHP to aquatic invertebratesa

    Organism                     Age     Temperature    Hardnessb       pH      Duration     LC50             Reference
                                             (° C)                                  (h)   (mg/litre)d
    Water flea                 < 24 h       17                                     48        0.133      Passino & Smith (1987)
      (Daphnia pulex)

    Water flea                 < 24 h       21-23         173        7.4-9.4       24         > 68      Leblanc (1980)
      (Daphnia magna)          < 24 h       21-23         173        7.4-9.4       48           11      Leblanc (1980)

    Crayfish                                                                       96         > 10      Mayer & Sanders (1973)
      (Orconectes nais)

    Harpacticoid               adult        20-22         7c           7.8         96        > 300      Linden et al. (1979)
      (Nitocra spinipes)

    Scud                                                                           96         > 32      Sanders et al. (1973)
      (Gammarus pseudolimnaeus)

    Midge                      larvae       21-23         270          7.4         48         > 18      Streufert et al. (1980)
      (Chironomus plumosus)
    a    All experiments were performed using static conditions (water unchanged for duration of test)
    b    Hardness is expressed as mg calcium carbonate per litre.  The tests were performed in fresh water
         except where stated otherwise
    c    Salinity (%)
    d    Nominal values
         Brown & Thompson (1982a) found no mortality of  Daphnia magna
    over an exposure period of 48 h and at DEHP dose levels up to 320
    µg/litre. However, at levels of 180 µg/litre or more, the daphnids
    were seen to float in the surface layer. The authors suggested that
    this observation was connected with the solubility of DEHP (which
    tends to precipitate out at > 180 µg/litre, possibly onto the
    daphnids, thereby causing them to float). Stephenson (1983) found no
    mortality in  Gammarus pulex exposed for up to 120 h to a DEHP
    concentration (400 µg/litre) in excess of its solubility.

         Laughlin et al. (1978) exposed grass shrimps  (Palaemonetes
     pugio) to DEHP concentrations of up to 1 mg/litre (well above the
    solubility limit of the ester) and found no significant difference in
    mortality between the control and treated shrimps during the 28-day
    exposure period. DEHP had no significant effect on the development
    rate of larvae from hatching to moult. No apparent effects were noted
    when mussels  (Mytilus edulis) were exposed to 50 µg/litre for 28
    days (Brown & Thompson, 1982b).

         Hobson et al. (1984) found no significant mortality of penaeid
    shrimps  (Penaeus vannamei) fed a diet containing up to 50 g DEHP/kg
    for 14 days, this representing 4% of the body weight per day.
    Histological examination revealed no changes and no significant dose-
    related effects were observed on moulting.

         Sanders et al. (1973) and Mayer & Sanders (1973) exposed  Daphnia
     magna for a complete life cycle (21 days) to 3, 10 or 30 µg
    DEHP/litre in an intermittent-flow system. Reproduction was
    significantly inhibited at all concentrations (60, 70, and 80%
    inhibition at the three dose levels, respectively). In contrast, Brown
    & Thompson (1982a) found no effect on the reproduction of  Daphnia
     magna at concentrations of up to 100 µg/litre and suggested that the
    difference in the numbers of young born to the controls in the two
    studies, i.e. 11 offspring per parent (Mayer & Sanders, 1973) compared
    with 170 per parent (Brown & Thompson, 1982a), could account for these
    conflicting results.

         When Knowles et al. (1987) exposed  Daphnia magna to DEHP
    concentrations of between 12 and 811 µg/litre for up to 21 days under
    flow-through conditions (the reproduction rate was approximately 200
    offspring per adult), survival was not affected at levels of < 158
    µg/litre. At 811 µg/litre, however, survival was significantly reduced
    after both 7 and 21 days. The mean number of young per surviving adult
    was also reduced at this dose level. The levels of major biochemical
    components such as protein, RNA, DNA, and glycogen were all
    significantly reduced after 7 days at 811 µg/litre, but all except
    glycogen were unaffected after 21 days. The authors concluded that the
    maximum acceptable toxicant concentration (MATC), based on survival
    and reproduction, was between 158 and 811 µg/litre. A no-observed-
    effect level (NOEL) of 72 µg/litre was identified both by DNA content
    and RNA/DNA ratio at day 7 and by surfacing of  Daphnia at day 0.

         Thuren & Woin (1991) exposed the freshwater amphipod  Gammarus
     pulex to DEHP at concentrations of 100 or 500 µg/litre for 10 days
    under flow-through conditions. There was a 5 day pre- and post-
    exposure period. The overall locomotor activity of G. pulex was
    significantly decreased at the higher exposure level, and the effect
    persisted throughout the post-exposure period. No significant effects
    were observed at the lower dose level.

         Flow-through chronic toxicity tests in both sand and hydrosoil
    showed that DEHP concentrations as high as 360 µg/litre (sand) and 240
    µg/litre (hydrosoil) had no significant effects on the growth or
    development of midge larvae  (Chironomus plumosus) over a 35-day
    period. The continuous exposure of first-generation midge eggs in sand
    substrate to mean DEHP concentrations of between 140 and 360 µg/litre
    had no significant effect on hatchability (Streufert et al., 1980).

         Woin & Larsson (1987) found that dragonfly larvae ( Aeshna sp.)
    caught significantly fewer prey  (Chaborus larvae) per attempt when
    exposed to sediment DEHP concentrations of approximately 600 mg/kg for
    between 3 and 9 weeks.

    9.2.2  Fish

         The acute toxicity to fish is summarized in Table 10 and the
    toxicity to the embryo-larval stages of fish is shown in Table 11.

         DEHP caused no deaths among one-year-old rainbow trout  (Salmo
     gairdneri) exposed to between 1 and 1000 mg/litre for 48 h (Silvo,
    1974) or juvenile Atlantic salmon  (Salmo salar) exposed to 100
    mg/litre for 96 h (Zitko, 1972).

         Mehrle & Mayer (1976) reported no effects on growth or survival
    of adult fathead minnows  (Pimephales promelas) exposed to between 1
    and 62 µg/litre for 56 days. They also exposed rainbow trout eggs to
    DEHP levels of 5, 14, and 54 µg/litre for 12 days prior to hatching,
    but there was no significant effect on hatchability. The resulting fry
    were continuously exposed to DEHP for a further 90 days. The two
    highest concentrations caused a significant, but not dose related,
    increase in the mortality of sac fry within 5 days of hatching. After
    yolk absorption (at 24 days), DEHP caused no significant mortality or
    effects on growth and development for the remainder of the exposure

        Table 10.  Toxicity (96-h LC50 values) of DEHP to fish

    Organism                      Size         Stat/    Temperature     Hardnessb      pH        LC50            Reference
                                               flowa        (° C)                             (mg/litre)c
    Bluegill sunfish                                                                             > 10       Mayer & Sanders (1973)
      (Lepomis macrochirus)     0.32-1.2 g     stat        21-23          32-48      6.7-7.8     > 770      Buccafusco et al. (1981)

    Fathead minnow                                                                               > 10       Mayer & Sanders (1973)
      (Pimephales promelas)     0.055-0.25 g               24-26          44-46      7-8         > 0.33d    Defoe et al. (1990)

    Channel catfish                                                                              > 10       Mayer & Sanders (1973)
      (Ictalurus punctatus)

    Rainbow trout               fingerling     stat        14-16                                  540       Hrudey et al. (1976)
      (Oncorhynchus mykiss)
                                                                                                 > 10       Mayer & Sanders (1973)
    a    Stat = static conditions (water unchanged for duration of test)
    b    Hardness is expressed as mg calcium carbonate per litre.  The tests were performed in fresh water
         except where stated otherwise
    c    Nominal values unless stated otherwise
    d    Measured value

    Table 11.  Toxicity of DEHP to the embryo-larval stages of fisha

    Organism                      Stat/flowb     Hardnessc       pH       LC50 (mg/litre)   95% confidence limits

    Channel catfish                 stat          90-115       7.6-8.1         0.69               0.55-0.86
      (Ictalurus punctatus)

    Redear sunfish                  stat          90-115       7.6-8.1         6.18               4.65-8.04
      (Lepomis microlophus)

    Largemouth bass                 flow           45-55       7.5-8.0        42.1                28.8-77.8
      (Micropterus salmoides)
                                    flow          190-225      7.5-8.0        32.9                19.8-58.9

    Rainbow trout                   flow           45-55       7.5-8.0       139.5                123.2-165.2
      (Oncorhynchus mykiss)
                                    flow          190-225      7.5-8.0       149.2                125.8-203.8
    a    From: Birge et al. (1978).  Exposure was initiated 2 to 6 h after spawning (except in the case of rainbow trout where exposure
         was initiated 15 min after fertilisation) and continued until 4 days after hatching.  The hatching times were 22 days for rainbow
         trout (13.5-14.3 °C), 3 days for catfish (29-31 °C), and 3-4 days for bass and sunfish (20-24 °C).
    b    Stat = static conditions, but water renewed every 12 h; flow = flow-through conditions (DEHP concentration in water continuously
    c    Hardness is expressed as mg calcium carbonate per litre.  The tests were performed in fresh water except where stated otherwise.
         DeFoe et al. (1990) exposed rainbow trout  (Oncorhynchus mykiss)
    and Japanese medaka  (Oryzias latipes) to DEHP using concentrations
    of up to 0.502 mg/litre for a 90-day trout embryo-larval test and
    0.554 mg/litre for a 168-day larval test on medaka. No significant
    adverse effects were observed on trout hatchability, survival or
    growth, but there was a significant reduction in the growth of
    Japanese medaka.

         Mayer & Sanders (1973) studied the effects of DEHP on the
    reproduction of zebra fish  (Brachydanio rerio) and guppies
     (Lebistes reticulatus). During the 90-day dietary exposure, zebra
    fish were fed on 50 or 100 mg DEHP/kg food, and guppies were fed 100
    mg/kg. Although the treated zebra fish spawned more frequently than
    the control fish, the latter produced more eggs per spawn. Fry
    survival was significantly reduced by DEHP. Treated guppies produced
    fewer fry per adult and had an 8% incidence of abortions, whereas
    there were no abortions in the control group (no statistics were

         In a study of fish exposed to DEHP, Mayer et al. (1977) found
    reduced vertebral collagen levels after 150 days at a DEHP
    concentration of 3.7 µg/litre in adult brook trout  (Salvelinus
     fontinalis), after 127 days at 11 µg/litre in fathead minnow, and
    after 90 days at 5 µg/litre in rainbow trout. However, there was no
    effect on fish growth. Pfuderer & Francis (1975) found that DEHP had
    no effect on the heart rate of goldfish when the fish were exposed to
    200 mg/litre for 10 min.

    9.2.3  Amphibians

         The acute toxicity of DEHP to tadpoles of Fowler's toad and the
    leopard frog is given in Table 12.

         When Larsson & Thuren (1987) exposed eggs of the moorfrog  (Rana
     arvalis) to sediment concentrations of between 10 and 800 mg DEHP/kg
    (fresh weight), the number of tadpoles hatching decreased with
    increasing exposure concentration. The percentage hatch was 90% for
    the controls eggs, 50% at 150 mg DEHP/kg, and < 30% at > 400 mg/kg.
    The lowest DEHP concentration that caused a significant decrease in
    the number of tadpoles hatching was 25 mg/kg. After hatching, the
    survival of tadpoles was unaffected. There were no delays in hatching
    and no abnormalities in the developing tadpoles exposed to the various
    DEHP concentrations.

    Table 12.  Toxicity of DEHP to frog and toad tadpolesa
    Organism            LC50 (mg/litre)     95% confidence limits
    Fowler's toad            3.88                  3.08-4.84
     (Bufo fowleri)

    Leopard frog             4.44                  3.65-5.37
     (Rana pipiens)
    a  From: Birge et al. (1978). Exposure occurred under static
       conditions, but water was renewed every 12 h. It was initiated 2
       to 6 h after spawning and continued until 4 days after hatching.
       The hatching times varied between 3 and 4 days, and so the
       exposure period varied between 7 and 8 days. The temperature was
       20-24 °C, hardness 90-115 mg CaCO3/litre, and pH 7.6-8.1.

         Wams (1987) quoted an unpublished Dutch report which stated that
    an exposure of 2 mg DEHP/litre caused a reduction in the growth rate
    of clawed toad  (Xenopus laevis) larvae.

    9.3  Toxicity to terrestrial organisms

    9.3.1  Plants

         When Herring & Bering (1988) grew spinach and pea plants from
    seed for 14 to 16 days in soil containing 100 mg DEHP/kg, there was no
    effect on plant growth, as measured by height. The effect of DEHP on
    seed germination was observed by placing seeds in petri dishes
    containing a solution of 100 mg/litre. A reduction of 40% to 50% in
    the number of seeds germinating was found.

         Lokke & Rasmussen (1983) reported that DEHP had no visible effect
    on  Sinapis alba, Brassica napusor or  Achillea millefolium when
    sprayed in the field at concentrations of up to 87.5 kg/ha. According
    to Stanley & Tapp (1982), DEHP at a level of 1000 mg/kg soil had no
    effect on the growth of rape  (Brassica rapa) and only a slight
    effect on that of oats  (Avena sativa).

    9.3.2  Earthworms

         In a series of contact toxicity tests, red earthworms  (Eisenia
     foetida) were exposed to DEHP via filter paper in glass vials. An
    LD50 could not be calculated because DEHP was not toxic even at the
    highest dose of 25 mg/cm2 (Neuhauser et al., 1986).

    9.3.3  Insects

         Al-Badry & Knowles (1980) found DEHP to be non-toxic to female
    houseflies  (Musca domestica) when applied topically or by injection
    at a concentration of 20 µg/fly (equivalent to 1000 mg/kg). When it

    was topically applied simultaneously with various organophosphates, an
    antagonistic interaction was apparent; mortality of 85% to 95% was
    reduced to less than 10% by the DEHP. However, when DEHP was topically
    applied 30 min before exposure to an organophosphate concentration
    that caused 10% to 30% mortality, the resulting interaction was
    synergistic with mortality rising to over 60% and in most cases to
    over 80%.

    9.3.4  Birds

         Hill et al. (1975) found no mortality among 10-day-old ring-
    necked pheasants or mallard fed up to 5000 mg DEHP/kg for 5 days
    followed by 3 days on a normal diet.

         When Wood & Bitman (1984) fed broiler hens a diet containing 2000
    mg DEHP/kg (226 mg DEHP/hen per day) for 4 weeks, egg production and
    body weights were significantly decreased. Ishida et al. (1982)
    reported the cessation of egg production and an abnormality of the
    ovaries in laying hens fed 5 or 10 g DEHP/kg diet for up to 230 days.

         In a study by O'Shea & Stafford (1980), starlings  (Sturnus
     vulgaris) were fed on a diet containing 25 or 250 mg DEHP/kg for a
    30-day period. At both dose levels, the treated birds gained
    significantly more body weight than controls during the exposure
    period, but at the lower dose level food consumption was significantly
    reduced compared to controls.

         When Peakall (1974) fed ring doves  (Streptopelia risoria) on a
    diet containing 10 mg/kg, there were no effects on the eggshell
    thickness, ashed egg weight, rate of water loss, surface area or
    permeability of the eggs laid.


    10.1  Evaluation of human health risks

    10.1.1  Exposure levels

         DEHP concentrations of up to 300 ng/m3 have been measured in
    urban air, but usually the levels in ambient air are well below 100
    ng/m3. Data on occupational exposure levels are limited.
    Concentrations of up to 4.1 mg/m3 have been reported but they are
    usually below 1 mg/m3. Total phthalate exposure levels have been
    reported to be between 0.1 and 60 mg/m3. Clinical treatment,
    including transfusion, haemodialysis, extracorporal circulation, and
    artificial respiration, may lead to DEHP exposure.

         The exposure to DEHP from drinking-water and food is low.

    10.1.2  Toxic effects

         Dose-dependent kinetics of DEHP or its metabolites and species
    differences in the metabolism have been confirmed in several studies.

         In animals some inhalation studies have been performed. However,
    no consistent findings are reported.

         The oral and intraperitoneal LD50 values exceed 25 g/kg,
    indicating that DEHP has low acute toxicity.

         In rats and mice, DEHP administration produces hepatic
    hyperplasia and hypertrophy, which is characterized by peroxisome
    proliferation. The threshold dose level for hepatic peroxisome
    proliferation in the rat is approximately 50 mg/kg per day. High DEHP
    dose levels (12 g DEHP/kg diet in rats; 6 g DEHP/kg diet in mice) in
    a feeding study resulted in an increased incidence of hepatic tumours
    in rats and mice.

         Marked species differences in DEHP-induced hepatomegaly and
    peroxisome proliferation exist. Rats and mice are very responsive,
    Syrian hamsters less responsive, and guinea-pigs, marmosets, and
    cynomolgus monkeys non-responsive. Studies with primary hepatocyte
    cultures have shown an excellent  in vitro/in vivo correlation
    concerning responsiveness to DEHP/DEHP metabolites. Several studies
    have demonstrated the non-responsive nature of cultured human
    hepatocytes to DEHP metabolites and other peroxisome proliferators.
    The induction of peroxisome proliferation and cell replication is
    strongly associated with the development of hepatic tumours in rats
    and mice. Thus, the available data suggest that species which do not
    respond to the hepatic effects of DEHP are unlikely to be susceptible
    to the development of hepatic tumours.

         Testicular atrophy is one of the most consistent effects of DEHP
    in  in vivo studies on a variety of experimental animal species. Such
    effects have been observed in rats fed 6 g DEHP/kg diet and mice fed
    3 g/kg diet, and they are more pronounced in young animals. Hamsters
    and marmosets appear to be more resistant to the testicular effects of

         When both male and female CD-1 mice were treated with 1 g DEHP/kg
    diet, a significant reduction in fertility was observed.

         A dietary level of 20 g DEHP/kg throughout gestation produced an
    increased incidence of resorptions in rats but no malformations. In
    the mouse, however, 1 g/kg throughout pregnancy increased the
    incidence of embryolethality and abnormalities. Sensitivity was
    greatest on days 7-9 of gestation. The dose level (0.5 g/kg) that
    induced fetotoxicity in mice did not induce maternal toxicity. Results
    from several different genotoxicity tests indicate that DEHP and its
    major metabolites do not exhibit any direct genotoxic effect in either
    bacteria or  in vitro mammalian cells. This has been confirmed in
     in vivo binding studies, which indicated that DEHP and its
    metabolites do not interact covalently with DNA. However, it has been
    established that DEHP has the potential of inducing aneuploidy in
    fungi as well as in  in vitro mammalian cells. There are no
    consistent results from dominant lethal studies.

         Few data on the human health effects of DEHP exposure have been
    reported. There have been a few studies on workers exposed to
    phthalate mixtures, but no consistent health effects that could be
    directly related to DEHP have been reported.

    10.1.3  Conclusion

         DEHP causes reproductive and hepatocarcinogenic effects in rats
    and mice.

         Testicular atrophy is the main reproductive effect in rats and
    mice, and young animals are more susceptible than older ones to this
    effect. The induction of hepatic peroxisome proliferation and cell
    replication are strongly associated with the liver carcinogenic effect
    of certain non-genotoxic carcinogens including DEHP. However, marked
    differences have been observed among animal species with respect to
    DEHP-induced peroxisome proliferation. Currently there is not
    sufficient evidence to suggest that DEHP is a potential human

    10.2  Evaluation of effects on the environment

    10.2.1  Exposure levels

         DEHP exists widely in the environment, being released during
    production, processing, usage, and disposal. Transport in the air is

    the major route by which it enters the environment. It can be
    deposited by dry or wet deposition and has been found to leach into
    ground water.

         DEHP is readily photodegraded in the atmosphere. In the
    laboratory, aerobic biodegradation has been shown to occur under
    certain circumstances. However, in the environment aerobic
    biodegradation seems to be slow and anaerobic degradation is even

         Based on a measured log Pow of about 5 and measured
    bioaccumulation factors for biota and sediment, it can be seen that
    DEHP is highly bioaccumulative.

         In rivers and lakes, DEHP concentrations of up to 4 µg/litre have
    been measured.

         Due to its hydrophobic nature, DEHP adsorbs readily to soil,
    sediment, and particulate matter. Levels of up to 70 mg/kg dry weight
    have been found in river sediments, but near to a discharge point
    levels of up to 1480 mg/kg dry weight have been reported.

    10.2.2  Toxic effects

         Most acute toxicity studies on aquatic organisms show DEHP to be
    of low toxicity. However, one study suggested a greater sensitivity
    for the water flea  (Daphnia pulex), the nominal 48-h LC50 being
    133 µg/litre.

         In a 21-day study on  Daphnia magna, the NOEL for biochemical
    and behavioural effects was 72 µg/litre. A significant increase in
    mortality was recorded in trout fry exposed to 14 µg DEHP/litre from
    12 days prior to hatching. At concentrations of 3.7 to 11 µg/litre,
    reductions in vertebral collagen were reported in three species of
    fish exposed for 150 days.

         Zebra fish fry survival and guppy reproduction were adversely
    affected at DEHP concentrations in food of 50 and 100 mg/kg,
    respectively, during a 90-day exposure period.

         In two separate studies, DEHP concentrations of 25 mg/kg
    (wet weight) of sediment significantly reduced microbial activity and
    the number of frog tadpoles hatching.

         Available studies indicate that the acute toxicity of DEHP to
    algae, plants, earthworms, and birds is low. There are no data
    relating to effects upon wild mammals.

    10.2.3  Conclusion

         There is no documented information that DEHP presents any hazard,
    based on acute exposure to fish and daphnids. However, a reduction of
    microbial activity in sediment at environmental levels of DEHP was
    reported. A comparison between environmental levels and the
    concentrations that produce effects in prolonged studies, especially
    early life-stage tests on fish and amphibians, indicates that a hazard
    for the environment, particularly via water and sediment, cannot be
    excluded. Adverse effects on organisms are likely in areas with highly
    contaminated water and sediments which are near to point emission

         Although few relevant studies have been reported, the acute
    toxicity of DEHP to algae, plants, earthworms, and birds appears to be


    a)   Current disposal practises should be improved.

    b)   Measures must be undertaken to reduce the release of DEHP to the

    c)   Medical devices and products that contribute to the body burden
         of DEHP must be scrutinized to reduce exposure to DEHP via the
         intravenous route.


         More research is needed on the effects of DEHP on the ecosystem
    of sediments and on the subject of DEHP leaching, particularly from
    landfill to ground water.

         Since DEHP occurs in the air, including workplace air, further
    inhalation studies are needed.

         Epidemiological surveys on exposed populations should be
    encouraged. Further studies are needed on the mechanism of peroxisome-
    proliferator-induced hepatocarcinogenicity, with emphasis on human
    health effects.


         DEHP was one of the substances discussed by a working group of
    the International Agency for Research on Cancer in 1981 (IARC, 1982).
    The evaluation by the group was that there was sufficient evidence for
    the carcinogenicity of DEHP in mice and rats, but that no adequate
    epidemiological study was available. The working group concluded that
    DEHP is possibly carcinogenic to humans and placed it in group 2B
    (IARC, 1987).

         DEHP was evaluated at the 28th meeting of the Joint FAO/WHO
    Expert Committee on Food Additives. The recommendation was to reduce
    human exposure to DEHP from food-contact materials to the lowest level
    technologically attainable (WHO/FAO, 1984a,b). During the 33rd meeting
    of the Joint FAO/WHO Expert Committee on Food Additives, DEHP was
    again evaluated and the recommendation was the same: DEHP in food
    should be reduced to the lowest level attainable. The Committee
    considered that this might be achieved by using alternative
    plasticizers or alternatives to plastic material containing DEHP
    (WHO/FAO, 1989a,b).


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    1.  Identité, propriétés physiques et chimiques et méthodes d'analyse

         Le phtalate de di(2-éthylhexyle) est un ester de l'acide
    benzènedicarboxylique qui se présente, à la température ambiante, sous
    la forme d'un liquide huileux, incolore à jaune. Il est peu soluble
    dans l'eau (0,3-0,4 mg/litre) et encore moins dans l'eau salée. Il est
    miscible à la plupart des solvants organiques courants. Sa volatilité
    est relativement faible (tension de vapeur: 8,6 x 10-4Pa).

         On a mis au point, pour le dosage du DEHP dans différents
    milieux, un grand nombre de méthodes de prélèvement et d'analyse. On
    fait de plus en plus appel à des méthodes sensibles telles que la
    chromatographie en phase gazeuse, la chromatographie en phase liquide
    à haute performance et la spectrométrie de masse. La contamination due
    au matériel en plastique utilisé lors des prélèvements et de l'analyse
    complique le dosage du DEHP à faibles concentrations.

    2.  Sources d'exposition humaine et environnementale

         Le DEHP présent dans l'environnement est, dans sa presque
    totalité, d'origine artificielle.

         La production mondiale de DEHP augmente depuis quelques décennies
    et s'élève actuellement à environ un million de tonnes par an. Les
    Etats-Unis d'Amérique et l'Europe entrent chacun pour un tiers dans
    cette production.

         Le DEHP est le plastifiant le plus largement utilisé pour
    augmenter la fluidité des résines puisqu'il représente 50% de
    l'ensemble des esters phtaliques utilisés comme plastifiants. Il peut
    représenter jusqu'à 40% en poids ou davantage de la matière plastique.
    Le DEHP est utilisé dans la fabrication du chlorure de polyvinyle
    (PVC) utilisé dans l'industrie du bâtiment et de l'emballage ainsi que
    pour la fabrication d'accessoires à usage médical. On l'utilise
    également en plus petites quantités pour la préparation de peintures
    industrielles ainsi que comme diélectrique dans les condensateurs.

         Les produits contenant du plastique qui sont mis au rebut peuvent
    être éliminés soit par incinération soit par enfouissement dans une
    décharge. Lors de l'incinération à basse température, une proportion
    importante du produit peut se dissiper dans l'atmosphère. On n'a pas
    très bien étudié ce qu'il advient du DEHP déposé dans les décharges et
    il n'est pas possible à ce stade de tirer des conclusions définitives.

    3.  Transport, distribution, et transformation dans l'environnement

         C'est principalement en étant transportés dans l'atmosphère que
    les phtalates pénètrent dans l'environnement. A partir de là, le DEHP
    tombe sur le sol à moins d'être éliminé par les précipitations.

         Le coefficient de partage du DEHP entre l'octanol et l'eau est
    élevé, de sorte que l'équilibre entre l'eau et un sol ou un sédiment
    riche en substances organiques s'établit aux dépens de la phase
    aqueuse. Le DEHP s'adsorbe facilement sur les particules organiques et
    du sol.

         Bien que peu soluble dans l'eau, le DEHP peut se rencontrer en
    quantités relativement élevées dans les eaux de surface en raison de
    son adsorption sur les particules organiques et de son interaction
    avec les substances organiques en solution. L'adsorption s'effectue
    plus particulièrement sur les particules de petite taille et elle est
    favorisée par la présence d'eau salée.

         Dans l'atmosphère, le DEHP subit une photodégradation rapide mais
    son hydrolyse dans l'environnement est pratiquement inexistante.

         Plusieurs microorganismes terricoles effectuent la dégradation
    aérobie du DEHP. Toutefois dans l'environnement, cette dégradation
    microbienne s'effectue avec lenteur. La biodégradation commence par
    l'hydrolyse du mono-ester qui est transformé en acide phtalique. Il y
    a ensuite dégradation en pyruvate et succinate par ouverture du cycle
    puis en CO2 et H2O, comme dans la dégradation métabolique de
    l'acide benzoïque. La dégradation aérobie dépend de la température. Au
    dessous de 10 °C la dégradation est peu importante. A température
    supérieure, la biodégradation se produit dans la partie supérieure du
    sol mais, à plus grande profondeur elle est pratiquement inexistante
    du fait des conditions d'anaérobiose. La dégradation anaérobie, pour
    autant qu'elle existe, est beaucoup plus lente que la dégradation

         Le DEHP est très lipophile et modérément persistant. Le degré de
    bioaccumulation dépend de la capacité de l'organisme qui l'absorbe à
    métaboliser la substance. On a montré qu'il s'accumulait en forte
    proportion chez divers invertébrés aquatiques, les poissons et les

         Après application de DEHP sur des feuilles de végétaux, on n'a
    constaté qu'une faible perte de matière sur une période de 15 jours.
    Le DEHP présent dans le sol ou dans les boues d'égout n'est que peu
    fixé par les plantes.

    4.  Concentrations dans l'environnement et exposition humaine

         Le DEHP est très répandu dans l'environnement et on le retrouve
    dans la plupart des échantillons prélevés dans l'air, dans les
    précipitations, l'eau, les sédiments, le sol et les biotes. C'est
    généralement dans les zones industrielles que les teneurs sont les
    plus fortes.

         On a relevé des concentrations allant jusqu'à 300 ng/m3 dans
    l'air des villes et en atmosphère polluée. Dans l'atmosphère
    surmontant les océans, on a fait état de concentrations allant de 0,5
    à 5 ng/m3 et les précipitations de ces régions en contenaient
    jusqu'à 200 ng/litre. L'étude d'échantillons de précipitations
    prélevés dans un secteur situé à proximité d'une usine produisant des
    plastifiants, a montré que la vitesse de dépot à sec était de 0,7 à
    4,7 µg/m2 et par jour.

         Dans les rivières et les lacs, on a trouvé des concentrations de
    DEHP allant jusqu'à 4 µg/litre, les teneurs les plus élevées étant
    relevées au point de décharge d'effluents industriels. Dans la mer, la
    concentration est inférieure à 1 µg/litre, les concentrations les plus
    fortes étant observées dans les estuaires.

         En raison de son caractère hydrophobe, le DEHP est facilement
    absorbé par le sol, les sédiments et les particules. Dans les
    sédiments de cours d'eau, des concentrations allant jusqu'à 70 mg/kg
    (de poids à sec) ont été signalées, concentrations qui peuvent monter
    jusqu'à 1480 mg/kg (de poids à sec) au voisinage des points de

         Dans les biotes, la concentration du DEHP varie de moins de 1 à
    7000 µg/kg. On le rencontre dans divers types d'aliments: poisson,
    coquillages, oeufs et fromages. L'exposition moyenne a été estimée à
    environ 300 µg par personne et par jour aux Etats-Unis d'Amérique en
    1974 et à 20 µg par personne et par jour au Royaume-Uni en 1986.

         Les transfusions de sang et autres traitements utilisant du
    matériel en plastique peuvent entraîner l'exposition involontaire des
    malades au DEHP. C'est ainsi qu'on a relevé dans le poumon de malades
    ainsi traités, des teneurs allant de 13,4 à 91,5 mg/kg de poids à sec.

         Les quelques données disponibles indiquent que sur les lieux de
    travail, la concentration de DEHP est généralement inférieure à 1

    5.  Cinétique et métabolisme

         Des données disponibles au sujet de l'administration par voie
    orale indiquent que le DEHP est hydrolysé dans l'intestin par la
    lipase pancréatique. Les métabolites qui se forment, c'est-à-dire le
    phtalate de mono-2-éthylhexyle (MEHP) et le 2-éthylhexanol sont
    rapidement absorbés. Après administration de DEHP marqué au 14C
    (2,9 mg/kg) par voie orale à des rats, on a récupéré 50% de la dose
    initiale dans l'urine et la bile de ces animaux. Il semble que la
    biodisponibilité d'une dose orale de DEHP soit plus élevée chez les
    jeunes animaux.

         Après administration par voie orale, le DEHP est largement
    hydrolysé dans l'intestin de certains animaux, par exemple les rats,
    et se répartit dans l'organisme, principalement sous forme de phtalate
    de monoéthylhexyle (MEHP). Toutefois chez les primates et l'homme,
    l'hydrolyse est beaucoup moins importante. Il semble que ce soit
    principalement au niveau du foie que s'effectue la métabolisation du
    MEHP et du 2-éthylhexanol. On a identifié plusieurs autres
    métabolites, l'omega et omega-1-oxydation étant les principales voies
    métaboliques. Un ou plusieurs des produits résultant de l'omega-
    oxydation peuvent encore être métabolisés par ß-oxydation. On a
    constaté que l'omega-oxydation s'effectuait selon une cinétique non
    linéaire. Le métabolisme du DEHP offre des différences considérables
    selon les espèces; par exemple, l'omega-oxydation est moins importante
    chez l'homme que chez le rat.

         Une dose de DEHP de 2,9 mg/kg, administrée par voie orale à des
    rats, a été récupérée une semaine plus tard presque intégralement dans
    les matières fécales et l'urine des animaux. La bile et l'urine
    constituent les principales voies d'excrétion. Lors d'une étude sur
    l'homme, 15 à 25% d'une dose orale (0,45 mg/kg de DEHP) ont été
    excrétés sous forme de MEHP et la majeure partie des produits
    d'excrétion étaient constitués de métabolites oxydés.

    6.  Effets sur les mammifères de laboratoire et les systèmes
        d'épreuve in vitro

         La DL50 par voie orale du DEHP est d'environ 25 à 34 g/kg,
    selon l'espèce, mais cette valeur est plus faible dans le cas du MEHP.
    Lors d'études d'alimentation sur des rats et des souris, on a constaté
    que des doses quotidiennes de DEHP supérieures à 3 g/kg entraînaient
    la mort dans les 90 jours et qu'une dose de 0,4 g/kg réduisait le gain
    de poids en l'espace de quelques jours. Dans d'autres études, c'est
    une dose de 6,3-12,5 g/kg de nourriture qui a entraîné une réduction
    du poids corporel.

         Lors d'études à long terme, on a observé chez les animaux traités
    une hépatomégalie accompagnée d'une augmentation du poids relatif des
    reins. Dans l'une de ces études, on observait également une
    hypertrophie cellulaire au niveau du lobe antérieur de l'hypophyse.

         Un certain nombre d'études ont permis de mettre en évidence une
    atrophie testiculaire, apparaîssant au bout de quelques jours, et liée
    à l'administration de DEHP (doses dans l'alimentation allant de
    10-20 g/kg). Les jeunes rats semblent plus vulnérables et les rats et
    les souris plus sensibles que les hamsters et les ouistitis. On a
    constaté que cette atrophie était réversible. Le MEHP exerce des
    effets toxiques  in vitro sur les cellules de Sertoli. Le DEHP de
    même que le MEHP ont des propriétés tératogènes. Des malformations ont
    été observées à des doses de 0,5-2 g/kg de nourriture chez la souris
    et, à des doses supérieures à 10 g/kg, on a observé des effets

         Les tests de mutagénicité et autres manifestations du même genre
    ont donné des résultats négatifs dans la plupart des études. Le DEHP
    peut provoquer la transformation des cellules et on a montré qu'il
    était cancérogène à des doses respectives de 6 et 12 g/kg de
    nourriture chez le rat et de 3 et 6 g/kg de nourriture chez la souris.
    On a constaté une augmentation, liée à la dose, des tumeurs
    hépatocellulaires chez les deux sexes de l'une et l'autre espèce. La
    prolifération des peroxysomes hépatiques et la réplication cellulaire
    sont fortement liées aux effets cancérogènes sur le foie de certains
    produits non génotoxiques comme le DEHP. Toutefois on a observé des
    différences importantes entre les espèces animales en ce qui concerne
    la prolifération des peroxysomes induite par le DEHP.

         Contrairement à ce qui se passe dans le cas des hépatocytes de
    rat, les métabolites du DEHP ne provoquent pas de prolifération des
    peroxysomes dans les cultures d'hépatocytes humains.

    7.  Effets sur l'homme

         On ne dispose que de données très limitées sur les effets que le
    DEHP peut exercer sur l'homme. On a fait état chez deux sujets qui
    avaient reçu 5 ou 10 g de DEHP respectivement, d'effets qui se
    limitaient à de légers troubles gastriques.

    8.  Effets sur les autres êtres vivants en laboratoire et dans
        leur milieu naturel

         Dans la plupart des études, les valeurs nominales de la CL50
    obtenues lors des tests de toxicité aiguë dépassent 10 mg/litre, ce
    qui indique que le DEHP est peu toxique. Toutefois ces valeurs sont
    supérieures à la solubilité du DEHP dans l'eau (0,3-0,4 mg/litre).
    D'après une étude, une daphnie,  Daphnia pulex, serait
    particulièrement sensible, la CL50 à 48 heures étant dans son cas de
    0,133 mg/litre. La seule épreuve de toxicité aiguë où l'on ait mesuré
    des concentrations de DEHP a été effectuée sur un poisson,  Pimephales
     promelas, et elle a donné une CL50 à 96 heures supérieure à 0,33
    mg/litre. Lors d'études de longue durée, on a obtenu une dose sans
    effet nocif observable de 72 µg/litre pour  Daphnia magna. Pour les
    poissons adultes, cette dose était supérieure à 72 µg/litre. Une
    exposition à une concentration de 14 µg/litre, pendant 12 jours avant
    l'éclosion, a provoqué une importante augmentation de la mortalité
    parmi des alevins de truites. Des concentrations comprises en 3,7 et
    11 µg/litre ont entraîné une réduction de la teneur en collagène
    vertébral chez les poissons.

         La présence de DEHP dans la nourriture à raison de 50 mg/kg a eu
    des effets nocifs sur la survie des alevins de  Melambaphes zebra. A
    la concentration de 25 mg/kg en poids dans les sédiments, on a
    constaté une réduction de l'activité microbienne et du nombre d'
    éclosions chez les tétards.

         Le DEHP est peu toxique pour les algues, les végétaux, les
    lombrics et les oiseaux.

    9.  Evaluation

         Le DEHP exerce des effets cancérogènes sur le foie et affecte le
    système reproducteur chez le rat et la souris.

         Le principal effet au niveau des gonades consiste, chez le rat et
    la souris, en une atrophie des testicules qui affecte davantage les
    jeunes animaux. La prolifération des peroxysomes hépatiques et la
    réplication cellulaire sont fortement liées à l'effet cancérogène
    qu'exercent sur le foie certains produits non génotoxiques au nombre
    desquels figure le DEHP. Toutefois, on a observé de fortes différences
    selon les espèces animales pour ce qui est de la prolifération des
    peroxysomes induite par le DEHP. On ne possède pas actuellement de
    preuves suffisantes permettant de conclure que le DEHP est
    potentiellement cancérogène pour l'homme.

         Il n'existe pas de données attestées selon lesquelles le DEHP
    serait dangereux pour la faune, si l'on s'en tient aux résultats
    obtenus lors de l'exposition de poissons et de daphnies. Toutefois, on
    a observé une réduction de l'activité microbienne dans les sédiments
    aux doses auxquelles le DEHP est présent dans l'environnement. Si l'on
    compare les doses de DEHP présentes dans l'environnement aux
    concentrations qui sont susceptibles de produire des effets lors
    d'études de longue durée, notamment sur les larves de poissons et
    d'amphibiens, on ne peut exclure l'existence d'un risque pour
    l'environnement, les effets s'exerçant notamment par l'intermédiaire
    de l'eau et des sédiments. Des effets nocifs sur les êtres vivants
    sont possibles dans les secteurs où l'eau et les sédiments sont
    fortement pollués en raison de leur proximité des points de décharge.

         Bien que peu d'études valables aient été effectuées, il semble
    que le DEHP présente une faible toxicité aiguë pour les algues, les
    végétaux, les lombrics et les oiseaux.


    1.  Identidad, propiedades físicas y químicas y métodos analíticos

         El di(2-etilhexil) ftalato (DEHF) es un éster del ácido
    bencenodicarboxílico que a temperatura ambiente es un líquido oleoso
    entre incoloro y amarillo. Su solubilidad en agua es baja (0,3-0,4
    mg/litro) y aún más baja en agua salada. Es miscible con la mayor
    parte de los disolventes orgánicos normales. La volatilidad del DEHF
    es relativamente baja (8,6 x10-4 Pa).

         Se han perfeccionado numerosos métodos de muestreo y análisis
    para la determinación del DEHF en diferentes medios. Cada vez se
    utilizan más métodos de gran sensibilidad, como la cromatografía de
    gases, la cromatografía líquida de alto rendimiento y la
    espectrometría de masas. El análisis de concentraciones bajas de DEHF
    se complica por la contaminación debida al material de plástico
    utilizado en el muestreo y el análisis.

    2.  Fuentes de exposición humana y ambiental

         Casi todo el DEHF presente en el medio ambiente procede de
    fuentes antropogénicas.

         La producción mundial de DEHF ha ido aumentando durante los
    últimos decenios y en la actualidad es de alrededor de 1 x 106
    toneladas al año. Un tercio del total se produce en los Estados Unidos
    y otro tercio en Europa.

         El DEHF es el plastificante más utilizado (representa el 50% de
    todos los plastificantes a base de ésteres de ftalato) para ablandar
    las resinas. A él corresponde el 40% (p/p) o más del total de
    plásticos. El DEHF se emplea en la fabricación de cloruro de
    polivinilo (PVC) utilizado en los edificios, en la construcción, en
    embalajes y en componentes de aparatos médicos. Se utiliza en
    cantidades más pequeñas en las pinturas industriales y como fluido
    dieléctrico en los condensadores.

         Los productos plastificados de desecho se pueden eliminar por
    incineración o terraplenado. Durante la incineración a baja
    temperatura, se puede liberar un porcentaje elevado del DEHF a la
    atmósfera. No se ha estudiado bien el destino medioambiental del DEHF
    depositado en terraplenes, por lo que no se pueden sacar conclusiones

    3.  Transporte, distribución y transformación en el medio ambiente

         El transporte en el aire es la vía más importante de
    incorporación de los ftalatos al medio ambiente. De la atmósfera, el
    DEHF precipita o es lavado por la lluvia.

         El DEHF tiene un coeficiente de reparto octanol-agua elevado, de
    manera que el equilibrio entre el agua y un suelo o sedimento rico en
    materia orgánica está desplazado a favor de este último. Las
    partículas orgánicas del suelo lo adsorben fácilmente.

         Aunque la solubilidad del DEHF en agua es baja, la cantidad
    presente en las aguas de superficie puede ser mayor debido a la
    adsorción a las partículas orgánicas y a la interacción con la materia
    orgánica disuelta. Las pequeñas partículas lo adsorben más fácilmente,
    y el proceso se potencia en el agua salada.

         La fotodegradación atmosférica del DEHF es rápida; en cambio, su
    hidrólisis química en el medio ambiente es prácticamente inexistente.

         Se ha observado que varios microorganismos del suelo llevan a
    cabo la degradación aerobia. Sin embargo, se ha informado que la
    degradación microbiana del DEHF en el medio ambiente es lenta. El
    proceso de biodegradación comienza con la hidrólisis para formar el
    monoéster, que se transforma luego en ácido ftálico. La degradación
    con ruptura del anillo para dar primero piruvato y succinato y después
    CO2 y H2O es análoga a la vía metabólica del ácido benzoico. La
    degradación aerobia es dependiente de la temperatura. Por debajo de
    los 10 °C es escasa. A temperaturas más altas se produce
    biodegradación en la capa más superficial del suelo, pero es
    prácticamente inexistente en las capas más profundas, donde las
    condiciones son de anaerobiosis. La degradación anaerobia, si existe,
    es mucho más lenta que la aerobia.

         El DEHF es muy lipófilo y moderadamente persistente. El grado de
    bioacumulación depende de la capacidad de los organismos para
    metabolizarlo. Se ha observado un grado elevado de acumulación en
    diversos invertebrados acuáticos, peces y anfibios.

         Cuando se aplicó DEHF a hojas de plantas, la pérdida fue pequeña
    durante un período de 15 días. Se ha observado que su absorción por
    las plantas a partir del suelo o de los fangos cloacales es muy baja.

    4.  Niveles medioambientales y exposición humana

         El DEHF está muy ampliamente distribuido en la naturaleza y se
    encuentra en la mayor parte de las muestras, por ejemplo de aire,
    precipitación, sedimento, suelo y biota. Las concentraciones más altas
    aparecen generalmente en las zonas industrializadas.

         En el aire urbano y contaminado se han encontrado concentraciones
    de hasta 300 ng/m3. Se han comunicado de niveles comprendidos entre
    0,5 y 5 ng/m3 en el aire de zonas oceánicas, y la lluvia de esas
    zonas contenía hasta alrededor de 200 ng/litro. En muestras de
    precipitación procedentes de una zona cercana a una fábrica de
    plastificantes se observó que la tasa sedimentación seca era de 0,7 a
    4,7 µg/m2 al día.

         Se han encontrado concentraciones de DEHF en ríos y lagos de
    hasta 4 µg/litro; los niveles más altos están asociados a los puntos
    de descarga de efluentes industriales. La concentración en el mar es
    inferior a 1 µg/litro, y los niveles más altos se detectan en los

         Debido a su carácter hidrófobo, el DEHF se adsorbe muy fácilmente
    al suelo, los sedimentos y la materia particulada. En los sedimentos
    fluviales se han comunicado niveles de hasta 70 mg/kg (peso seco), y
    cerca de los puntos de descarga ha llegado a ser de 1480 mg/kg (peso

         La concentración de DEHF en la biota oscila entre menos de 1 y
    7000 µg/kg. Se ha detectado en diversos tipos de alimentos, como
    pescado, marisco, huevos y queso. En 1974 se estimó en los Estados
    Unidos una exposición media de 300 µg/persona al día y en el Reino
    Unido en 1986 de 20 µg/persona al día.

         Las transfusiones de sangre y otros tipos de tratamiento médico
    en los que se utilizan aparatos de plástico pueden dar lugar a una
    exposición humana involuntaria al DEHF. En el tejido pulmonar de
    algunos pacientes se han detectado concentraciones comprendidas entre
    13,4 y 91,5 µg/kg (peso seco).

         Los escasos datos disponibles indican que las concentraciones de
    DEHF en el lugar de trabajo suelen estar por debajo de los 1 mg/m3.

    5.  Cinética y metabolismo

         Los datos disponibles sobre la administración oral indican que la
    lipasa pancreática hidroliza el DEHF en el intestino. Los metabolitos
    formados, que son el mono (2-etilhexil) ftalato (MEHF) y el
    2-etilhexanol, se absorben rápidamente. Cuando se administró por vía
    oral a ratas DEHF marcado con 14C (2,9 mg/kg), se recuperó en la
    orina o la bilis más del 50%. La biodisponibilidad de una dosis oral
    de DEHF parece ser más elevada en las ratas jóvenes que en las más

         El DEHF administrado por vía oral se hidroliza en el intestino de
    ciertos animales, por ejemplo las ratas, y se distribuye sobre todo
    como monoetilhexil ftalato (MEHF). Sin embargo, la hidrólisis es mucho
    menor en los primates y el ser humano. El MEHF se liga a las proteínas
    del plasma. El hígado parece ser el principal órgano metabolizador del
    MEHF y del 2-etilhexanol. Se han identificado algunos otros
    metabolitos, siendo las principales vías metabólicas la omega-y la
    omega-1-oxidación. Uno o varios productos de la omega-oxidación pueden
    metabolizarse ulteriormente por ß-oxidación. En la omega-oxidación se
    han observado cinéticas no lineales. En el metabolismo del DEHF se
    registran considerables diferencias entre las especies; por ejemplo,
    la omega-oxidación es menor en el hombre que en las ratas.

         Una semana después de administrar una dosis oral de DEHF
    (2,9 mg/kg) se recuperó casi el 100% en las heces y la orina de ratas.
    La bilis y la orina son las principales vías de excreción. En un
    estudio humano, del 15 al 25% de la dosis oral (0,45 mg/kg) de DEHF se
    excretó en forma de MEHF; los metabolitos oxidados constituían la
    parte más importante de los productos de excreción.

    6.  Efectos en los mamíferos de laboratorio y en sistemas de
        ensayo in vitro

         La DL50 del DEHF por vía oral es de alrededor de 25-34 g/kg,
    según las especies, pero el valor para el MEHF es más bajo. En
    estudios de alimentación en ratas y ratones, las dosis de DEHF
    superiores a 3 g/kg al día causaron muertes en un plazo de 90 días, y
    con una concentración de 0,4 g/kg al día se redujo el aumento de peso
    a los pocos días. En otros estudios, con dosis de 6,3-12,5 g/kg de
    alimentos se produjo una reducción del peso corporal.

         En los animales tratados en estudios de larga duración se ha
    observado hepatomegalia y un aumento del peso relativo de los riñones.
    En un estudio se encontraron también células hipertróficas en el
    lóbulo anterior de la hipófisis.

         En varios estudios se ha detectado atrofia testicular, evidente
    al cabo de pocos días, relacionada con la administración de DEHF
    (concentraciones de 10-20 g de DEHF/kg de alimentos). Las ratas más
    jóvenes parecen ser más susceptibles que las viejas, y las ratas y los
    ratones parecen ser más sensibles que los titís y los hámsters. Se ha
    observado que la atrofia es reversible. El MEHF tiene efecto tóxico
     in vitro en las células de Sertoli. Tanto el DEHF como el MEHF
    muestran propiedades teratogénicas. Con concentraciones de 0,5 a 2
    g/kg de alimentos se observaron malformaciones en ratones, y con
    niveles superiores a los 10 g/kg se apreciaron efectos embriotóxicos.

         Las pruebas de mutagenicidad y puntos finales asociados han dado
    resultados negativos en la mayor parte de los estudios. El DEHF puede
    inducir la transformación celular y se ha demostrado que con dosis de
    6 y 12 g/kg de alimentos en ratas y de 3 y 6 g/kg de alimentos en
    ratones tiene efectos carcinógenos. En ambos sexos de las dos especies
    se produjo un aumento de tumores hepatocelulares dependiente de la
    dosis. La inducción de la proliferación de peroxisomas hepáticos y de
    la replicación celular está fuertemente vinculada al efecto
    carcinógeno en el hígado de determinados compuestos carcinógenos no
    genotóxicos, entre los que figura el DEHF. Sin embargo, se han
    observado notables diferencias entre distintas especies animales con
    respecto a la proliferación de peroxisomas inducida por el DEHF.

         A diferencia de lo que ocurre con los hepatocitos de rata, los
    metabolitos del DEHF no inducen proliferación de peroxisomas en
    cultivos de hepatocitos humanos.

    7.  Efectos en el ser humano

         La información disponible sobre los efectos del DEHF en el ser
    humano es muy escasa. Se han comunicado dos casos de trastornos
    gástricos leves, con dosis de 5 y 10 g de DEHF, pero sin ningún otro
    efecto nocivo.

    8.  Efectos en otros organismos en el laboratorio y en el
        medio ambiente

         En la mayoría de los estudios de toxicidad aguda se han obtenido
    unos valores nominales para la CL50 que se sitúan por encima de los
    10 mg/litro, por lo que el índice de toxicidad del DEHF es bajo. Sin
    embargo, esos niveles son superiores a los correspondientes a su
    solubilidad en agua (0,3-0,4 mg/litro). En un estudio se detectó en la
    pulga de agua  Daphnia pulex una sensibilidad mayor, con un valor
    nominal de la CL50 a las 48 h de 0,133 mg/litro. La única prueba de
    la toxicidad aguda realizada con concentraciones medidas de DEHF se
    hizo en el pez  Pimephales promelas y puso de manifiesto una CL50
    > 0,33 mg/litro a las 96 h. En estudios prolongados, el nivel sin
    observación de efectos (NOEL) en  Daphnia magna fue de 72 µg/litro.
    En peces adultos se determinó un NOEL > 62 µg/litro. Una exposición
    a 14 µg/litro desde 12 días antes de la eclosión produjo un
    significativo aumento de la mortalidad de los alevines de trucha. Las
    concentraciones entre 3,7 y 11 µg/litro provocaron una reducción del
    colágeno vertebral en peces.

         Con concentraciones de 50 mg/kg de alimentos, el DEHF tiene
    efectos adversos sobre la supervivencia de los alevines de
     Brachydanio rerio. Concentraciones de 25 mg/kg (p/p) en el sedimento
    redujeron de manera considerable la actividad microbiana y el número
    de renacuajos que nacían.

         La toxicidad aguda del DEHF en algas, plantas, lombrices de
    tierra y aves es baja.

    9.  Evaluación

         El DEHF tiene efectos reproductivos y hepatocarcinogénicos en
    ratas y ratones.

         El principal efecto sobre la reproducción en ratas y ratones es
    la atrofia testicular; los animales jóvenes son más susceptibles que
    los viejos. La inducción de la proliferación de peroxisomas hepáticos
    y de la replicación celular está estrechamente relacionada con el
    efecto carcinogénico en el hígado de determinados compuestos
    carcinógenos no genotóxicos, incluido el DEHF. Sin embargo, se han
    observado notables diferencias entre las distintas especies de
    animales con respecto a la proliferación de peroxisomas inducida por
    el DEHF. En la actualidad no hay pruebas suficientes que indiquen que
    el DEHF sea un posible carcinógeno para el ser humano.

         No hay información documentada de que el DEHF constituya un
    riesgo, tomando como base la exposición aguda de peces y dáfnidos. Sin
    embargo, se ha informado de una reducción de la actividad microbiana
    en los sedimentos con los niveles de DEHF presentes en el medio
    ambiente. La comparación entre los niveles medioambientales y las
    concentraciones que producen efectos en estudios prolongados,
    especialmente las pruebas en las fases vitales tempranas de peces y
    anfibios, indica que no se puede descartar un cierto peligro para el
    medio ambiente, sobre todo por conducto del agua y los sedimentos. Es
    probable que se produzcan efectos adversos en los organismos en zonas
    con agua y sedimentos muy contaminados que están próximos a las
    fuentes de emisión.

         Aunque son pocos los estudios que se conocen al respecto, la
    toxicidad aguda del DEHF en algas, plantas, lombrices de tierra y aves
    parece ser baja.

    See Also:
       Toxicological Abbreviations