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    UNITED NATIONS ENVIRONMENT PROGRAMME
    INTERNATIONAL LABOUR ORGANISATION
    WORLD HEALTH ORGANIZATION


    INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY



    ENVIRONMENTAL HEALTH CRITERIA 184





    Diflubenzuron





    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.


    First draft prepared by Dr M. Tasheva, Sofia, Bulgaria


    Published under the joint sponsorship of the United Nations
    Environment Programme, the International Labour Organisation, and the
    World Health Organization, and produced within the framework of the
    Inter-Organization Programme for the Sound Management of Chemicals.


    World Health Organization
    Geneva, 1996

         The International Programme on Chemical Safety (IPCS) is a joint
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    WHO Library Cataloguing in Publication Data

    Diflubenzuron.

    (Environmental health criteria ; 184)

    1. Diflubenzuron - adverse effects   2. Diflubenzuron - toxicity
    3. Insecticides - adverse effects    4. Insecticides - toxicity
    5. Environmental exposure  I. Series

    ISBN 92 4 157184 1                 (NLM Classification: WA 240)
    ISSN 0250-863X

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    CONTENTS

    ENVIRONMENTAL HEALTH CRITERIA FOR DIFLUBENZURON

    Preamble

    1. SUMMARY AND EVALUATION; CONCLUSIONS AND RECOMMENDATIONS

         1.1. Summary

              1.1.1. Identity, physical and chemical properties, and
                       analytical methods
              1.1.2. Sources of human and environmental exposure
              1.1.3. Environmental transport, distribution and
                       transformation
              1.1.4. Environmental levels and human exposure
              1.1.5. Kinetics and metabolism in laboratory animals
              1.1.6. Effects on laboratory mammals and  in vitro test
                       systems
              1.1.7. Effects on humans
              1.1.8. Effects on other organisms in the laboratory and
                       field
         1.2. Evaluation
              1.2.1. Evaluation of human health risks
              1.2.2. Evaluation of effects on the environment
              1.2.3. Toxicological criteria for setting guidance values
         1.3. Conclusions and recommendations

    2. IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, ANALYTICAL METHODS

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

    3. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE

         3.1. Natural occurrence
         3.2. Anthropogenic sources
              3.2.1. Production levels and processes
              3.2.2. Formulations
              3.2.3. Uses

    4. ENVIRONMENTAL TRANSPORT, DISTRIBUTION, TRANSFORMATION AND FATE

         4.1. Appraisal
         4.2. Transport and distribution between media
              4.2.1. Soil mobility
              4.2.2. Dissipation
              4.2.3. Evaporation
              4.2.4. Crop residue data

         4.3. Transformation
              4.3.1. Abiotic degradation
                       4.3.1.1   Photolysis
                       4.3.1.2   Hydrolysis
              4.3.2. Biodegradation
                       4.3.2.1   Water
                       4.3.2.2   Soil
         4.4. Bioaccumulation and biomagnification
         4.5. Interaction with other physical, chemical or
              biological factors
         4.6. Ultimate fate following use

    5. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE

         5.1. Environmental levels
              5.1.1. Air
              5.1.2. Water
              5.1.3. Food and feed
              5.1.4. Forest plants and litter
              5.1.5. Aquatic organisms
         5.2. General population exposure
         5.3. Occupational exposure during manufacture, formulation or use

    6. KINETICS AND METABOLISM IN LABORATORY ANIMALS AND HUMANS

         6.1. Absorption
         6.2. Distribution
         6.3. Metabolic transformation
              6.3.1. Metabolites - distribution, excretion, retention
                       and turnover
         6.4. Elimination and excretion
         6.5. Retention and turnover
              6.5.1. Biological half-life

    7. EFFECTS ON LABORATORY MAMMALS AND  IN VITRO TEST SYSTEMS

         7.1. Single exposure
         7.2. Short-term exposure
         7.3. Long-term exposure
         7.4. Skin and eye irritation; sensitization
         7.5. Reproductive toxicity, embryotoxicity and teratogenicity
         7.6. Mutagenicity and related end-points
         7.7. Carcinogenicity
         7.8. Other special studies
              7.8.1. Special studies on met- and sulfhaemoglobin
                       formation
         7.9. Toxicity of metabolites
              7.9.1. Carcinogenicity studies with 4-chloroaniline

    8. EFFECTS ON HUMANS

    9. EFFECTS ON OTHER ORGANISMS IN THE LABORATORY AND FIELD

         9.1. Laboratory experiments
              9.1.1. Microorganisms
                       9.1.1.1   Water
                       9.1.1.2   Soil
              9.1.2. Aquatic organisms
                       9.1.2.1   Microorganisms
                       9.1.2.2   Plants
                       9.1.2.3   Invertebrates
                       9.1.2.4   Vertebrates
              9.1.3. Terrestrial organisms
                       9.1.3.1   Plants
                       9.1.3.2   Invertebrates
                       9.1.3.3   Vertebrates
         9.2. Field observations
              9.2.1. Microorganisms
                       9.2.1.1   Water
                       9.2.1.2   Soil
              9.2.2. Aquatic organisms
                       9.2.2.1   Plant
                       9.2.2.2   Invertebrates
                       9.2.2.3   Vertebrates
              9.2.3. Terrestrial organisms
                       9.2.3.1   Invertebrates
                       9.2.3.2   Vertebrates

    10. PREVIOUS EVALUATIONS BY INTERNATIONAL BODIES

    REFERENCES

    RESUME

    RESUMEN
    

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         This publication was made possible by grant number 5 U01
    ES02617-15 from the National Institute of Environmental Health
    Sciences, National Institutes of Health, USA, and by financial support
    from the European Commission.



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    WHO TASK GROUP ON ENVIRONMENTAL HEALTH CRITERIA FOR DIFLUBENZURON

     Members

    Dr T. Bailey, US Environmental Protection Agency, Washington DC, USA

    Dr A.L. Black, Department of Human Services and Health, Canberra,
         Australia

    Mr   D.J. Clegg, Carp, Ontario, Canada

    Dr S. Dobson, Institute of Terrestrial Ecology, Monks Wood, Abbots
         Ripton, Huntingdon, Cambridgeshire, United Kingdom
          (Vice-Chairman)

    Dr P.E.T. Douben, Her Majesty's Inspectorate of Pollution, London,
         United Kingdom  (EHC Joint Rapporteur)

    Dr P. Fenner-Crisp, US Environmental Protection Agency, Washington DC,
         USA

    Dr R. Hailey, National Institute of Environmental Health Sciences,
         National Institutes of Health, Research Triangle Park, USA

    Ms   K. Hughes, Environmental Health Directorate, Health Canada,
         Ottawa, Ontario, Canada  (EHC Joint Rapporteur)

    Dr D. Kanungo, Central Insecticides Laboratory, Government of India,
         Ministry of Agriculture & Cooperation, Directorate of Plant
         Protection, Quarantine & Storage, Faridabad, Haryana, India

    Dr L. Landner, MFG, European Environmental Research Group Ltd,
         Stockholm, Sweden

    Dr M.H. Litchfield, Melrose Consultancy, Denmans Lane, Fontwell,
         Arundel, West Sussex, United Kingdom  (CAG Joint Rapporteur)

    Professor M. Lotti, Institute of Occupational Medicine, University of
         Padua, Padua, Italy  (Chairman)

    Professor D.R. Mattison, University of Pittsburgh, Graduate School of
         Public Health, Pittsburgh, Pennsylvania, USA

    Dr J. Sekizawa, National Institute of Health Sciences, Tokyo, Japan

    Dr P. Sinhaseni, Chulalongkorn University, Bangkok, Thailand

    Dr S.A. Soliman, King Saud University, Bureidah, Saudi Arabia

    Dr M. Tasheva, National Centre of Hygiene, Medical Ecology and
         Nutrition, Sofia, Bulgaria  (CAG Joint Rapporteur)

    Mr J.R. Taylor, Pesticides Safety Directorate, Ministry of 
         Agriculture, Fisheries and Food, York, United Kingdom

    Dr H.M. Temmink, Wageningen Agricultural University, Wageningen, The
         Netherlands

    Dr M.I. Willems, TNO Nutrition and Food Research Institute, Zeist,
         The Netherlands

     Representatives of GIFAPa (Groupement International des
    Associations Nationales de Fabricants de Produits Agrochimiques)

    Dr M. Bliss, Jr., ISK Biosciences Corporation, Mentor, Ohio, USA

    Dr A.C. Dykstra, Registration Department BPID, Solvay-Duphar BV, CP
         Weesp, The Netherlands

    Dr H. Frazier, ISK Biosciences Corporation, Mentor, Ohio, USA

    Dr R. Gardiner, GIFAP, Brussels, Belgium

    Dr B. Julin, Regulatory Affairs, Du Pont de Nemours (Belgium),
         Agricultural Products Department, Mercure Centre, Brussels,
         Belgium

    Dr S.M. Kennedy (Environmental Science), Du Pont de Nemours (Belgium),
         Agricultural Products Department, Mercure Centre, Brussels,
         Belgium

    Dr J. Killeen, ISK Biosciences Corporation, Mentor, Ohio, USA

    Dr Th. S.M. Koopman, Toxicology Department, Solvay-Duphar BV, CP
         Weesp, The Netherlands

    Dr R.L. Mull, Du Pont Agricultural Products, Wilmington, Delaware, USA

    Dr J.L.G. Thus, Environmental Research Department, Solvay-Duphar BV,
         CP Weesp, The Netherlands

     Secretariat

    Ms A. Sundén Byléhn, International Register of Potentially 
         Toxic Chemicals, United Nations Environment Programme,
         Châtelaine, Switzerland

    Dr P. Chamberlain, International Programme on Chemical Safety, World
         Health Organization, Geneva, Switzerland

               

    a  Participated as required for exchange of information.

    Dr J. Herrman, International Programme on Chemical Safety, World
         Health Organization, Geneva, Switzerland

    Dr K. Jager, International Programme on Chemical Safety, World Health
         Organization, Geneva, Switzerland

    Dr P. Jenkins, International Programme on Chemical Safety, World
         Health Organization, Geneva, Switzerland

    Dr W. Kreisel, World Health Organization, Geneva, Switzerland

    Dr M. Mercier, International Programme on Chemical Safety, World
         Health Organization, Geneva, Switzerland

    Dr M.I. Mikheev, Occupational Health, World Health Organization,
         Geneva, Switzerland

    Dr G. Moy, Food Safety, World Health Organization, Geneva, Switzerland

    Mr I. Obadia, International Labour Organisation, Geneva, Switzerland

    Dr R. Pleœtina, International Programme on Chemical Safety, World
         Health Organization, Geneva, Switzerland

    Dr E. Smith, International Programme on Chemical Safety, World Health
         Organization, Geneva, Switzerland  (EHC Secretary)

    Mr J. Wilbourn, International Agency for Research on Cancer, Lyon,
         France

    ENVIRONMENTAL HEALTH CRITERIA FOR DIFLUBENZURON

         The Core Assessment Group (CAG) of the Joint Meeting on Pesticide
    Residues met in Geneva from 25 October to 3 November 1994.
    Dr W. Kreisel of the WHO welcomed the participants on behalf of WHO,
    and Dr M. Mercier, Director, IPCS, on behalf of the IPCS and its
    cooperating organizations (UNEP/ILO/WHO).  The Group reviewed and
    revised the draft monograph and made an evaluation of the risks for
    human health and the environment from exposure to diflubenzuron.

         The first draft of the monograph was prepared by Dr M. Tasheva,
    Sofia, Bulgaria.  The second draft, incorporating comments received
    following circulation of the first draft to the IPCS contact points
    for Environmental Health Criteria monographs, was prepared by the IPCS
    Secretariat.

         Dr K.W. Jager and Dr P.G. Jenkins, both members of the IPCS
    Central Unit, were responsible for the overall scientific content and
    technical editing, respectively.

         The fact that Solvay-Duphar, BV, made available to the IPCS its
    proprietary toxicological information on diflubenzuron is gratefully
    acknowledged.  This allowed the CAG to make its evaluation on a more
    complete database.

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

    ABBREVIATIONS

    ADI       acceptable daily intake

    a.i.      active ingredient

    AP        alkaline phosphatase

    bw        body weight

    4-CPU     4-chlorophenylurea

    DFB       diflubenzuron

    2,6-DFBA  2,6-difluorobenzoic acid

    ECD       electron capture detection

    G         granular formulation

    GC        gas chromatography

    GLC       gas-liquid chromatography

    Hb        haemoglobin

    HPLC      high performance liquid chromatography

    MATC      maximum acceptable toxicant concentration

    MCH       mean cell haemoglobin

    MCHC      mean cell haemoglobin concentration

    MCV       mean cell volume

    NOAEC     no-observed-adverse-effect concentration

    NOEL      no-observed-effect level

    NPD       nitrogen-phosphorus detector

    PCA        para-chloroaniline (4-chloroaniline)

    PCV       packed cell volume

    SAP       serum alkaline phosphatase

    SGOT      serum glutamic-oxaloacetic transaminase (aspartate
              aminotransferase)

    SGPT      serum glutamic-pyruvic transaminase (alanine
              aminotransferase)

    TLC       thin-layer chromatography

    WP        wettable powder

    1.  SUMMARY AND EVALUATION; CONCLUSIONS AND RECOMMENDATIONS

    1.1  Summary

    1.1.1  Identity, physical and chemical properties, and analytical
           methods

         Diflubenzuron is a member of the benzoylphenylurea group of
    insecticides.  Its insecticidal action is due to interaction with
    chitin synthesis and/or deposition.  It forms odourless white crystals
    with a melting point of 230-232°C.  It is sparingly soluble in water
    (0.2 mg/litre at 20°C) and is virtually non-volatile.  It is
    relatively stable in acidic and neutral media but it hydrolyses in
    alkaline conditions.

         Diflubenzuron is produced by the reaction of 2,6-difluoro-
    benzamide with 4-chlorophenylisocyanate.

         Diflubenzuron residues may be measured in water, biological
    samples and soils by HPLC with UV detection or by GC with ECD for
    analysis of the intact molecule or following derivatization of the
    liberated 4-chloroaniline with trifluoroacetic anhydride.

    1.1.2  Sources of human and environmental exposure

         Diflubenzuron is a synthetic compound used in agriculture,
    forestry and public health programmes to control insect pests and
    vectors.  Different formulations of diflubenzuron are available for
    these uses.  There is no relevant information on human exposure to
    diflubenzuron.

    1.1.3  Environmental transport, distribution and transformation

         Diflubenzuron is usually applied directly to plants and water. 
    Uptake of diflubenzuron through plant leaves does not occur.

         The adsorption of diflubenzuron to soil is rapid.  It is
    immobilized in the top 10 cm layer of soil to which it is applied.  It
    is unlikely to leach.  Diflubenzuron is degraded in soils of various
    types and origin under aerobic or anaerobic conditions with a half-
    life of a few days.  The rate of degradation depends greatly on the
    diflubenzuron particle size.  The main metabolic pathway (over 90%) is
    hydrolysis leading to 2,6-difluorobenzoic acid and 4-chlorophenylurea;
    these are degraded with half-lives of about 4 and 6 weeks,
    respectively. Free 4-chloroaniline has not been detected in soils.

         Diflubenzuron degrades rapidly in neutral or alkaline waters. 
    Studies of application of diflubenzuron to water show rapid partition
    to sediment; the parent compound and 4-chlorophenylurea may persist on
    sediment for more than 30 days.

         Diflubenzuron does not bioaccumulate in fish.

    1.1.4  Environmental levels and human exposure

         Exposure of the general population to diflubenzuron via water or
    food as a result of its use in agriculture, against forest insects or
    in mosquito control is negligible.

    1.1.5  Kinetics and metabolism in laboratory animals

         In experimental animals, diflubenzuron is absorbed from the
    digestive tract and to a lesser extent through the skin.  There is a
    saturable absorption mechanism in the rat gastrointestinal tract. 
    Consequently a large proportion of orally administered diflubenzuron
    is found in the faeces.  Diflubenzuron has widespread distribution in
    the tissues, but it does not accumulate.

         The metabolic fate of diflubenzuron has been studied in various
    species.  The major route of metabolism in mammals is via
    hydroxylation.  Hydrolysis of diflubenzuron may occur at any of the
    three carbon-nitrogen bonds.  In pigs and chickens the major route of
    hydrolysis is at the ureido bridge.  In rats and cows the major
    metabolic pathway is hydroxylation.  The major metabolites in sheep,
    swine and chickens are 2,6-difluorobenzoic acid and 4-chloro-
    phenylurea; minor metabolites are 2,6-difluorobenzamide and
    4-chloroaniline.  In rats and cattle 80% of the metabolites are
    2,6-difluoro-3-hydroxydiflubenzuron, 4-chloro-2-hydroxy-diflubenzuron
    and 4-chloro-3-hydroxydiflubenzuron.  The metabolic studies indicate
    that little or no 4-chloroaniline is formed in rats or cattle.

         The major route of elimination is via the faeces, ranging from 70
    to 85% in cats, pigs and cattle.  In sheep elimination is roughly
    equally distributed between the urine and faeces.  Urinary excretion
    in rats and mice decreases proportionally with increasing dosage
    level.  Less than 1% of an oral dose is recovered in exhaled air. 
    Only trace residues are found in milk.

         No human studies on the kinetics and metabolism of diflubenzuron,
    including the extent of biotransformation to 4-chloroaniline, are
    available.

    1.1.6  Effects on laboratory mammals and in vitro test systems

         Diflubenzuron has low acute toxicity by any route of exposure. 
    It has been classified by WHO as a "product unlikely to present an
    acute hazard in normal use", based on an acute oral LD50 of more than
    4640 mg/kg body weight in rats.  The acute dermal LD50 in rats is
    greater than 10 000 mg/kg body weight while the acute inhalational
    LC50 for rats is greater than 2.49 mg/litre.  No signs of
    intoxication have been observed during the 14-day period following
    single administration of diflubenzuron by various routes to a variety
    of animal species.

         Diflubenzuron is not a skin irritant (in rabbits) and not a skin
    sensitizer (in guinea-pigs).  It is marginally irritating to the eyes
    of rabbits.

         Diflubenzuron causes methaemoglobinaemia and sulfhaemo-
    globinaemia.  Dose-related methaemoglobinaemia has been demonstrated
    after oral, dermal or inhalatory exposure to diflubenzuron in various
    species.  This effect is the most sensitive toxicological end-point in
    experimental animals.  The NOEL based on methaemoglobin formation is
    2 mg/kg body weight per day in rats and dogs and 2.4 mg/kg body weight
    per day in mice.  In long-term toxicity studies with mice and rats,
    treatment-related changes were principally associated with oxidation
    of haemoglobin or with hepatocyte changes.

         In carcinogenicity studies in mice and rats at dietary levels up
    to 10 000 mg/kg in the diet, there were no treatment-related effects
    on tumour incidence.  Specifically, there were no mesenchymal
    neoplasms of the spleen or liver as observed in carcinogenicity
    studies with 4-chloroaniline.

         In several reproductive toxicity studies on rats, mice, rabbits
    and three avian species, no effects were seen on reproduction and
    there was no embryotoxicity.  Teratogenicity studies in rats and
    rabbits demonstrated no teratogenic effects.

         Diflubenzuron and its main metabolites have been examined in a
    variety of  in vitro and  in vivo mutagenicity tests.  Neither
    diflubenzuron nor its major metabolites have a mutagenic effect.

         The minor metabolite, 4-chloroaniline, was shown to be positive
    in several  in vitro mutagenicity assays using various end-points. 
    It is carcinogenic in rats and mice.  The neoplastic lesions related
    to administration of 4-chloroaniline were benign and malignant
    mesenchymal tumours in the spleens of male rats and haemangiomas and
    haemangiosarcomas, primarily in the spleen and liver of male mice.

    1.1.7  Effects on humans

         The diflubenzuron metabolite, 4-chloroaniline, has been reported
    to cause methaemoglobinaemia in exposed workers and in neonates
    inadvertently exposed.  Some individuals who are deficient in
    NADH-methaemoglobin reductase may be particularly sensitive to
    4-chloroaniline and hence to diflubenzuron exposure.

    1.1.8  Effects on other organisms in the laboratory and field

         All chitin-synthesizing organisms show susceptibility to
    diflubenzuron.

         Bacteria were not affected by diflubenzuron at concentrations of
    500 mg/kg soil; some stimulation of nitrogen fixation was seen.

    Diflubenzuron acetone solutions were degraded; the acetone was used 
    as carbon source.  Algal biomass increased at a diflubenzuron
    concentration of 1 µg/litre.  There were no adverse effects at
    concentration above the limit of diflubenzuron solubility.  Fungi were
    temporarily affected at 0.1 µg/litre in laboratory streams.

         Aquatic invertebrates show variable responses to diflubenzuron. 
    Molluscs are insensitive, the LC50 being greater than 200 mg/litre. 
    LC50 values for other invertebrates ranged from 1 to > 1000
    µg/litre, reflecting the effects of the compound on juvenile,
    moulting stages.  A MATC for  Daphnia has been estimated at > 40 and
    < 93 ng/litre; as expected, larval mayflies and other aquatic insect
    juveniles are highly susceptible.  Overspray of water bodies would be
    expected to kill some aquatic invertebrates.

         In ecosystems and field experiments where diflubenzuron was
    applied directly to the water, the effects on most taxa were less
    severe than predictions from acute laboratory tests.  No effects on
    aquatic organisms have been found after aerial applications to
    forests.

         The LC50 values for fish are > 150 mg/litre.  In field
    experiments, fish kills have never been recorded.

         The oral and contact LD50 for honey-bees is greater than
    30 µg/bee.  Honey-bee colonies were not affected after aerial
    application of 350 g diflubenzuron/ha.

         A 5-day dietary study on the mallard duck and bobwhite quail with
    levels of up to 4640 mg/kg feed revealed no observable signs of
    toxicity.  Small songbirds in the forest ecosystem were not affected
    after aerial application of diflubenzuron at 350 g/ha.

         Small mammal species showed no reductions in numbers after
    application of diflubenzuron at 67 g/ha to a forest.

    1.2  Evaluation

    1.2.1  Evaluation of human health risks

         The primary manifestation of diflubenzuron toxicity is
    methaemoglobin induction.  This toxicity occurs in a range of test
    animal species. It is attributable to the metabolite, 4-chloroaniline,
    which is known to induce methaemoglobin formation in several animal
    species and in humans.

         Diflubenzuron does not cause other toxicities on chronic dietary
    administration.  It is not mutagenic or carcinogenic in mice or rats. 
    However, its metabolite, 4-chloroaniline, is mutagenic  in vitro and
    is carcinogenic in mice and male rats.  Although 4-chloroaniline is a

    minor urinary metabolite of diflubenzuron in rats, the extent to which
    it is formed  in vivo in various animal species remains unknown. 
    Similarly, the comparative degree of absorption of its parent compound
    in various species is unknown.

         The sensitivity of human haemoglobin to methaemoglobin formation
    by 4-chloroaniline  in vivo is not known.  However, since induction
    of methaemoglobinaemia is consistently the most sensitive measure of
    diflubenzuron toxicity in the various animal species tested, it may be
    used as the basis to estimate the levels causing no toxicological
    effect.

    1.2.2  Evaluation of effects on the environment

         Diflubenzuron adsorbs readily to soil with little subsequent
    desorption.  Its mobility in soil is very low, practically all
    residues remaining within 15 cm of the top, even in sandy loam soils;
    diflubenzuron does not leach.  It is only partly removed from foliage
    by heavy rainfall.  Nevertheless, some diflubenzuron may be present in
    surface water shortly after application, due to flooding of treatment
    areas or agricultural run-off.

         Dissipation of diflubenzuron from water is rapid. Adsorption to
    sediment occurs within 4 days; both parent compound and 4-chloro-
    phenylurea metabolite may persist on sediment for at least 30 days.

         Uptake of diflubenzuron by plants through the leaves after aerial
    application does not occur.  Some uptake of soil residues does occur
    in plants and this may be translocated.  At the highest application
    rate (1 kg a.i./ha), following 1 month ageing of residues, up to
    1 mg/kg residue may be found in various crops.

         Photolysis of diflubenzuron is slow with a calculated half-life
    of 40 days.  Under environmental conditions abiotic degradation in
    water and soil represents a minimum route of break-down.  Aerobic
    degradation in water is a microbial process with a half-life of a few
    days under both laboratory and field conditions.  In the field,
    degradation of diflubenzuron applied at practical rates is influenced
    by pH, temperature, formulation, organic matter content and depth of
    the water.

         Degradation in soil through microbial hydrolysis is a rapid
    process, with a half-life of a few days, depending on diflubenzuron
    particle size.  The major break-down products are 2,6-difluorobenzoic
    acid and 4-chlorophenylurea; a minor metabolite is parachloroaniline. 
    All these are irreversibly bound to soil and/or further metabolized.

         The half-life of diflubenzuron residues on citrus fruits is
    significantly decreased by high temperature and humidity.

         Anaerobic degradation in water and sediment is slower than
    aerobic.

         Fish bioconcentrate diflubenzuron and some bioaccumulation takes
    place during extended exposure up to a plateau, depending on the water
    concentration, owing to fast degradation of diflubenzuron and
    excretion of metabolites; the depuration half-life is less than one
    day.  The 4-chloroaniline metabolite has not been detected in fish.

         Fish are not sensitive to diflubenzuron, the LC50 values being
    > 150 mg/litre.  Metabolites of diflubenzuron are also of low
    toxicity to fish.  Chronic exposure has shown no effects on fish at
    recommended application rates; the compound does not persist in water
    and no chronic exposure is expected.

         Diflubenzuron is not phytotoxic to duckweed at the diflubenzuron
    solubility limit concentration.

         Honey-bees were not affected by topical applications of
    > 30 µg/bee or dietary concentrations of up to 1000 mg/kg diet.
    Brood in hives was reduced when bees were fed syrup at 59 mg
    diflubenzuron/kg.  Brood was also reduced following exposure of
    flying colonies.

         Earthworms were not affected at a concentration of 780 mg/kg
    soil, which is at least three orders of magnitude above reported soil
    residues.

         Diflubenzuron has low acute toxicity to birds, the oral and
    dietary LD(LC)50 values being greater than 3000 mg/kg diet. 
    Following recommended application rates diflubenzuron is not expected
    to pose a hazard to birds.

         Extensive field studies have shown minimal or reversible effects
    on most aquatic invertebrates; daphnids were most seriously affected,
    with short-term reductions in populations of up to 75% following a
    single application of diflubenzuron.  Fish were not affected by water
    overspraying.  Neither bird nor mammal populations were adversely
    affected following forest spraying with diflubenzuron.

         A summary of risk quotients for birds, fish and aquatic
    invertebrates is given in Table 1.

    1.2.3  Toxicological criteria for setting guidance values

         The toxicological studies on diflubenzuron of relevance for
    setting guidance values are shown in Table 2.

        Table 1.  Toxicity/exposure ratios for birds, fish and aquatic invertebrates based on
              application rates of 2.5 kg a.i./ha of diflubenzuron to soybeans (worst case)
                                                                                           

    Risk category              LC50 (mg/litre      Estimated exposure    Toxicity/exposure
                               or mg/kg diet)      (mg/litre or          ratio (TER)c
                                                   mg/kg diet)a,b
                                                                                           

    Acute bird                      3762             73.7-535.7             51.0-7.0

    Acute fish (stream)              150                 0.0007              214 300

    Acute fish (pond)                150                   0.01               15 000

    Acute aquatic
     invertebrate (stream)         0.005                 0.0007                  7.1

    Acute aquatic
     invertebrate (pond)           0.005                   0.01                  0.5
                                                                                           

    a  Estimated environmental concentration in the terrestrial environment (for bird
       exposure) is based on the stated application rate and the assumption of
       deposition on short grass using the US EPA nomogram.

    b  Aquatic exposure concentrations were taken from the STREAM model based on a
       single application and estimated runoff into water; no direct overspray is
       included.

    c  TER is the toxicity (as LC50) divided by the exposure; values at or below
       1.0 indicate likely exposure to toxic concentrations by organisms in the
       different risk categories.

    Table 2.  Toxicological criteria for estimating guidance values for diflubenzuron
                                                                                 

    Exposure scenario   Relevant route/effect/        Result/remarks
    (technical          species
    diflubenzuron)
                                                                                 

    Short-term          dermal, irritation, rabbit    non-irritant
    (1-7 days)
                        ocular, irritation, rabbit    marginal, high dose

                        dermal, sensitization,        non-sensitizing
                        guinea-pig

                        inhalational, toxicity, rat   LC50 > 2.49 mg/litre
                                                      (single exposure)
    Mid-term
    (1-26 weeks)

    3 weeks; 5 days     dermal, irritation, rabbit    NOEL = 70 mg/kg body
     per week                                         weight per day

    3 weeks; 5 days     inhalational, methaemoglobin  NOAEL = < 0.12 mg/litre
     per week           formation, rat

    Long-term           dietary, methaemoglobin       NOEL = 2 mg/kg body weight
                        formation, rat                per day

                        dietary, methaemoglobin       NOEL = 2.4 mg/kg body weight
                        formation, mouse              per day

                        dietary, methaemoglobin       NOEL = 2 mg/kg body weight
                        formation, dog                per day
                                                                                 
    
    1.3  Conclusions and recommendations

         Considering the toxicological characteristics of diflubenzuron,
    both qualitatively and quantitatively, it was concluded, on the basis
    of the NOEL of 2 mg/kg body weight per day derived in long-term
    toxicity studies on mice, rats and dogs and applying a 100-fold
    uncertainty factor, that 0.02 mg/kg body weight per day will probably
    not cause adverse effects in humans whatever the route of exposure.

         Biomonitoring of 4-chloroaniline during occupational exposures
    needs to be carried out.

    2.  IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, ANALYTICAL METHODS

    2.1  Identity

    Molecular structure

    CHEMICAL STRUCTURE 1

    Empirical formula        C14H9ClF2N2O2

    Common name              Diflubenzuron

    Common trade names       Dimilin; Micromite; Vigilante

    Common abbreviation      DFB

    IUPAC name               1-(4-chlorophenyl)-3-(2,6-difluorobenzoyl)-
                             urea

    CAS chemical name        N-[[(4-chlorophenyl) amino] carbonyl]-
                             2,6-difluorobenzamide

    CAS registry number      35367-38-5

    RTECS registry number    YS6200000

         Technical diflubenzuron contains > 95% pure compound.

    2.2  Physical and chemical properties

         Diflubenzuron is an odourless white crystalline solid. It is
    almost insoluble in water and poorly soluble in apolar organic
    solvents.  In polar to very polar solvents, the solubility is moderate
    to good, e.g., in acetone it is 6.5 g/litre at 20°C.  Diflubenzuron is
    highly soluble in  N-methylpyrolidone (200 g/litre), dimethyl-
    sulfoxide and dimethylformamide (both 120 g/litre).

         Some physical and chemical properties of diflubenzuron are given
    in Table 3.

        Table 3.  Physical and chemical properties of diflubenzuron
                                                                                      

    Relative molecular mass                    310.7

    Melting point technical > 95%              210-230°C
                            > 99% pure         230-232°C

    Vapour pressure at 25°C                    0.00012 mPa

    Volatility
      solid material                           < 4%
      from water pH 5.6                        < 2% (virtually non-volatile)

    Specific gravity                           1.56

     n-Octanol/water partition coefficient
     (log Kow)                                 5000

    Solubility in water (at 25°C and pH 5.6)   8 × 10-5 g/litre

    Stability in water (0.0001 g/litre         4% decomposition after 3 weeks at pH 5
                       in the dark)            8% decomposition after 3 weeks at pH 7
                                               26% decomposition after 3 weeks at pH 91
                                                                                      
    
    2.3  Conversion factor

         1 ppm = 12.7 mg/m3 at 25°C
         1 mg/m3 = 0.079 ppm at 25°C

    2.4  Analytical methods

         Analytical methods for determining diflubenzuron in crops, soil,
    water and biological samples are summarized in Table 4.

         A review of the analytical methods has been presented by Rabenort
    et al. (1978).  Two general types of assay procedures for
    diflubenzuron are available: high performance liquid chromatography
    (HPLC) and gas chromatography (GC).

        Table 4.  Methods for the determination of diflubenzuron residues
                                                                                                                                           

    Sample type              Extraction/clean-up         Analytical  Limit of      Comments                        Reference
                                                         method      detection
                                                                                                                                           

    Crops, soil, water       dichloromethane; clean-up   HPLC        0.03 mg/kg                                    Rabenort et al. (1978)
                             on a Florisil column

    Milk                     ethyl acetate               HPLC        0.1 mg/kg                                     Corley et al. (1974)

    Crops                    acetone (n-hexane)          HPLC        0.01 mg/kg                                    Nakayama (1977a)

    Apples                   acetonitrile                HPLC        0.008 mg/kg                                   Goto (1977a)

    Tea                      acetone/dichloromethane     HPLC        0.1 mg/kg                                     Nakayama (1977b)

    Tea                      acetone or water            HPLC        0.2 mg/kg                                     Goto (1977b)

    Crops, soil, sediment;   acetonitrile                HPLC        0.05 mg/kg    the procedures involve Celite   Di Prima et al. (1978)
    aquatic and forest                                                             liquid-liquid partition, and
    foliage; fish and                                                              Florisil-aluminasilica gel
    shellfish; animal                                                              column chromatography;
    tissues                                                                        20 g sample

    Crops                    acetone-hexane              GLC-ECD     0.20 mg/kg                                    Lawrence & Sundaram
                             (1+4)                                                                                 (1976); Di Prima (1976)

    Soybean                  acetonitrile for process    GC-ECD      0.05 mg/kg    after hydrolysis and            Lawrence & Sundaram
                             fractions, hulls and meal;                            derivatization                  (1976); Di Prima (1976)
                             hexane-acetonitrile for
                             oil

    Water                    dichloromethane             TLC         0.1 mg/kg                                     Singh & Kaira (1989)
                                                                                                                                           

    Table 4 (Con't)
                                                                                                                                           

    Sample type              Extraction/clean-up         Analytical  Limit of      Comments                        Reference
                                                         method      detection
                                                                                                                                           

    Water & soil             hexane/ethyl acetate;       GC/ECD      0.05 ng       100 ml sample of water          Smith et al. (1983)
                             evaporate to dryness;                                 or 10 g sample of soil
                             dissolve residue in
                             benzene; derivatize with
                             trifluoroacetic anhydride
                             (with trimethylamine as
                             catalyst); LC on Florisil/
                             hexane: ethylether
                             (9:1 v/v)

    Water                    ethyl acetate, KCl;         GC/ECD      20 µg/litre   % DEGS-LAC 728 on               Cooke & Ober (1980)
                             derivatize with                                       Chromosorb W-AW at 165°C
                             trifluoroacetic anhydride;
                             LC on Florisil

    Exposure pads            methylene chloride or       HPLC/UV     3 ng          103.2 cm2 pads                  Bogus et al. (1985)
                             other solvents; clean-up    (254 nm)
                             on SEPPAC C18; elute with
                             methanol
                                                                                                                                           
             The HPLC method is recommended by CIPAC as a method of choice
    (van Rossum et al., 1984).  An alternative method for analysis of
    residues in crops, soil, mud and  water using Celite column
    chromatography has been described by Di Prima et al. (1978).  A gas
    chromatographic method used on the acetylated derivative of
    diflubenzuron was described by Worobey & Webster (1977) but has not
    been applied to crop samples.  The formation of 4-chloroaniline from
    diflubenzuron under acidic conditions provides the basis for the GC
    method.

         Most of the recommended extraction procedures use acetonitrile or
    acetone followed by  n-hexane or dichloromethane.

         Wie & Hammock (1982, 1984) developed three enzyme-linked
    immunosorbent assays (ELISA) for diflubenzuron. All three assays were
    based on antibodies raised against an  N-carboxypropyl hapten of
    diflubenzuron, while a diflubenzuron phenylacetic acid derivative
    coupled to a carrier other than the immunizing antigen was used as the
    coating antigen. None of these assays demonstrated significant cross-
    reactivity with benzamide, urea, phenylurea or aniline components of
    diflubenzuron. Each of the three assays was shown to be as sensitive
    as the recommended HPLC methodology for the analysis of diflubenzuron
    in water.  Using ELISA, DFB was detected in milk at a level of
    1-2 µg/litre without any sample extraction procedure.

         Wimmer et al. (1991) developed a gas chromatography/mass
    spectrometry (GC/MS) method using deuterated diflubenzuron as internal
    standard and claimed high sensitivity.

         The Joint FAO/WHO Codex Alimentarius Commission has given
    recommendations for the methods of analysis to be used in determining
    diflubenzuron residues (FAO/WHO, 1989).

    3.  SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE

    3.1  Natural occurrence

         Diflubenzuron does not occur naturally in the environment.

    3.2  Anthropogenic sources

    3.2.1  Production levels and processes

         Diflubenzuron was first commercialized by Philips-Duphar BV, The
    Netherlands (now Solvay Duphar BV).  Solvay Duphar BV produces
    diflubenzuron under the trade name Dimilin, but production figures are
    not available.

         Diflubenzuron is synthesized by the reaction of 2,6-difluoro-
    benzamide with  p-chlorophenyl isocyanate.

    3.2.2  Formulations

         Technical diflubenzuron is made into diflubenzuron 90%
    concentrate by air-milling with a grinding aid and sufficient kaolin
    to attain 90% active material.  This is the product from which all
    other formulations are made; these are listed below.

    Dry products

    *    Dimilin 25W: a 25% wettable powder (more or less the standard
         product)

    *    Dimilin 5W: a local Italian formulation containing 5% active
         ingredient

    *    Various granular formulations used locally in specific
         situations; these products are expected to be removed from the
         market within 2 or 3 years

    Water-based products

    *    Dimilin SC-48: a suspension concentrate containing 48% active
         ingredient

    *    Dimilin SC-15: a suspension concentrate containing 15% active
         ingredient for the French market

    *    Dimilin 4L, a suspension concentrate (0.4 kg/litre) containing
         48% active ingredient for the USA market

    Oil-based products

    *    Dimilin ODC-45: an oil-based dispersible concentrate containing
         45% active ingredient to be diluted with mineral or vegetable oil
         for spraying operations; this formulation may not be mixed with
         water

    *    Dimilin OF-6: a dispersion in oil ready for direct spraying,
         containing 6% active ingredient; this product must not be diluted
         or mixed with water

    *    Dimilin 2F: an oil-based suspension concentrate containing 24%
         active ingredient; it must not be diluted with water for spraying
         and is a local formulation development for the USA market

         The all-round formulations are Dimilin 25W, Dimilin 5W, Dimilin
    SC-48, Dimilin SC-15 and Dimilin 4L.  Dimilin ODC-45 was developed
    specially for aerial spraying operations on non-food crops and
    forestry.  Dimilin OF-6 was developed for broadcast aerial spraying
    operations to control locusts and grasshoppers.  Dimilin 2F was
    developed for those purposes where oil must be added to improve spray
    deposit tenacity on crops such as cotton.

    3.2.3  Uses

         Diflubenzuron was the first benzoylphenylurea to be discovered. 
    Its insecticidal properties were first described by van Daalen et al.
    (1972).

         Diflubenzuron is effective as a stomach and contact insecticide,
    acting by inhibiting chitin synthesis and so interfering with the
    formation of the cuticle.  Thus, all stages of insects that form new
    cuticles should be susceptible to diflubenzuron exposure.  It has no
    systemic activity and does not penetrate plant tissue.  Consequently,
    plant sucking insects are generally unaffected, and this forms the
    basis of its selectivity.

         The recommended application rates for diflubenzuron are given in
    Table 5.

         Diflubenzuron is effective at a concentration of 15-300 mg
    a.i./litre of water against leaf-feeding larvae and leaf miners in
    forestry  (Lymantria dispar, Thaumethopoea pityocampa), top fruit
    ( Cydia pomonella, Psylla spp), citrus  (Phyllocoptruta oleivora),
    field crops including cotton and soybeans  (Anthonomus grandis,
     Anticarsia gemmatalis), and horticultural crops  (Pieris
     brassicae).  It is also effective against the larvae of  Sciaridae
    and  Phoridae in mushrooms (1 g/m2 casing at case mixing or
    as a drench in 2.5 litre of water to the finished casing), against
    mosquito larvae (20-45 g/ha water surface) and against fly

    larvae  (Stomoxys calcitrans, Musca domestica) as a surface
    application in animal housings (0.5-1.0 g/m2 surface) (Worthing &
    Walker, 1987).


    Table 5.  Recommended application rates for diflubenzuron on
              different cropsa
                                                                        

    Pest                 Crop                Rate/concentration

    Apple rust mite      apples/pears        0.01-0.015% a.i.
    Codling moth         apples/pears        0.01-0.015% a.i.
    Leaf miners          apples/pears        0.01-0.015% a.i.
    Leaf rollers                             0.01-0.02% a.i.
    Pear suckers                             0.01 (+0.3% crop oil)% a.i.
                                             0.02-0.03% (without oil) a.i.
    Winter moth                              0.02% a.i.
    Plum fruit moth      plum                0.02% a.i.
    Olive moth           plum                0.01-0.02% a.i.
    Citrus rust mite     citrus fruit        0.0075-0.0125% a.i.
    Citrus weevil        citrus fruit        0.015-0.03% a.i.
    Cotton ball weevil   cotton              70 g/ha a.i.
    Army worms           cotton              150-300 g/ha a.i.
    Army worms           maize and Sorghum   70-150 g/ha a.i.
    Cotton leaf worms                        75-150 g/ha a.i.
    Beet army worms      peanuts             150-300 g/ha a.i.
    Rice water weevil    rice                75-150 g/ha a.i.
    Fall army worms      rice                70-100 g/ha a.i.
    Mosquitoes                               up to 100 g/ha a.i.
    Rice leaf rollers    rice                75-250 g/ha a.i.
    Various pests        peanuts             up to 75 g/ha a.i.
    Various pests        oil palm            50-150 g/ha a.i.
    Various pests        soybean             20-150 g/ha
                                                                        

    a  Solvay Duphar (1994)

    4.  ENVIRONMENTAL TRANSPORT, DISTRIBUTION, TRANSFORMATION AND FATE

    4.1  Appraisal

         Diflubenzuron is hydrolysed and photolysed slowly (see section
    2.2).  Residues in the aquatic environment may decrease rapidly, due
    to adsorption by organic and inorganic matter.  This process greatly
    reduces the availability of diflubenzuron to aquatic organisms.

    4.2  Transport and distribution between media

         Diflubenzuron is generally applied either directly on plants or
    on water for mosquito control.

    4.2.1  Soil mobility

         Diflubenzuron and its two formulations, Dimilin WP-25 and Dimilin
    SC-48, were applied separately at 17.23, 51.69 and 155.07 µg a.i.
    (corresponding to 70, 210 and 630 g a.i./ha) to the top layers of
    columns (30 × 5.6 cm internal diameter) packed with either sandy or
    clay loam forest soils.  Water (1.25 litre) equivalent to 50.8 cm of
    precipitation (representing an average annual rainfall) was allowed to
    leach through each column.  After leaching, the columns were divided
    into five segments from bottom to top as follows: two 10-cm
    increments, one 5-cm increment and two 2.5 cm increments. 
    Diflubenzuron residues in soils were extracted and analysed by HPLC. 
    Diflubenzuron mobility was low and did not increase with dosage.  At a
    deposit rate equivalent to 70 g a.i./ha, nearly all the residues were
    found within the top 2.5 cm of the column.  Even at 630 g a.i./ha,
    only about 9% of the technical diflubenzuron, 7% of Dimilin SC-48 and
    4% of Dimilin WP-25 moved below the 2.5 cm level in sandy loam.  The
    mobility of diflubenzuron in clay loam was lower than in sandy loam. 
    No residues were found below the 10 cm level or in the leachates in
    either soil type at any dosage levels.  The mobility of diflubenzuron
    was also influenced by the additives present in the formulation, the
    mobilities being in the following order: technical diflubenzuron
    > Dimilin SC-48 > Dimilin WP-25 (Sundaram & Nott, 1989).

         Helling (1985) investigated the movement of 14C-labelled
    diflubenzuron in five soils and classified it as immobile in all of
    them.  After six treatments of cotton fields with 14C-labelled
    diflubenzuron, most radioactivity was detected in the top 10 cm layer
    of soil (Bull & Ivie, 1978).  Diflubenzuron was found to adsorb very
    rapidly to eight soil types (greater than 87% of the initial amount),
    and there was only limited desorption (Booth et al., 1987).

         Fourteen days after a single foliar application of 14C-labelled
    diflubenzuron to field-grown cotton, only just over 10% of the dose
    was absorbed into the plants.  After 21 days and following a heavy
    rainfall, approximately 23% of the applied diflubenzuron remained on
    the treated leaf surfaces (Bull & Ivie, 1978).

         No leaching occurred when 14C-labelled diflubenzuron was applied
    to soil at the rate of 134.52 g/ha in an area with a normal rainfall
    of 32 cm (Danhaus et al., 1976).

    4.2.2  Dissipation

         Diflubenzuron might enter an estuary either as a result of
    flooding of treated supra-tidal mosquito breeding lagoons during
    spring tides or from agricultural run-off after significant rainfall
    (Cunningham & Myers, 1986).

         Following aerial application at 67.26 g/ha to a watershed,
    diflubenzuron was found to reach the stream channel.  It was also
    washed from the foliage as a result of several subsequent rainfalls
    (Jones & Konchenderfer, 1988).  However, these discharges were very
    short-lived.

         No residues were found in sediments from a lake treated with
    diflubenzuron, suggesting rapid dissipation before or upon reaching
    the bottom sediment (Apperson et al., 1978).

         Pritchard & Bourquin (1981) demonstrated some affinity of
    diflubenzuron for sediments, i.e. a partition coefficient of 380 in
    simulated estuarine conditions.  According to Cunningham & Myers
    (1986), sediment appeared to be a major site for diflubenzuron
    adsorption in a supra-tidal salt marsh.  Carringer et al. (1975) found
    that the organic content of soil was the most important factor in
    determining adsorption and dissipation of diflubenzuron, and that
    adsorption was inversely related to the water solubility of
    diflubenzuron.

    4.2.3  Evaporation

         When diflubenzuron was applied as Dimilin WP 80 at a
    concentration of 75 g/ha a.i. to bare soil (less than 1.5% organic
    matter) and red kidney bean leaves, no significant evaporation was
    measured under the following simulated climatological conditions: wind
    speed 1-2 m/s; temperature 20-21°C; relative humidity 25-45% (van der
    Laan-Straathof & Thus, 1994).

    4.2.4  Crop residue data

         When soybean and maize (corn) seedlings and potato tubers were
    planted into soil treated with 3H- or 14C-labelled diflubenzuron,
    only small amounts of radioactivity were taken up (Nimmo & de Wilde,
    1976a).  When 3H- or 14C-labelled diflubenzuron was applied to soil
    in which the seedlings of wheat and rice were already present, the
    14C residues in rice and wheat leaves were between 0.1 and 0.5 µg/kg. 
    The residues consisted mainly of 4-chlorophenylurea and polar
    conjugates.  The 14C residues in the wheat seeds were 0.02-0.04 mg/kg
    and 3H residues were lower (Nimmo & de Wilde, 1976b).

         The fate of diflubenzuron was studied following application to
    soybeans both in greenhouse and field conditions.  It was found that
    75 to 100% of the total residues in soybean plants consisted of
    unaltered diflubenzuron.  There was no significant absorption or
    translocation of residues.  Less than 0.05 mg/kg of the total residues
    was found in harvested soybean seed (Gustafson & Wargo, 1976).

         The diflubenzuron spray residue on aerial parts of plants is
    essentially stable.  Leaf permeation does not occur and the compound
    is not translocated to other parts of the plant.  It has been
    demonstrated that there is virtually no absorption, translocation or
    metabolism of foliar-applied diflubenzuron on greenhouse cotton plants
    (Nimmo & de Wilde, 1974; Nimmo, 1976a,b; Mansager et al., 1979).

         Plant metabolism studies in corn, soybean, cabbage and apples
    have demonstrated that no degradation products are found in plant
    tissues.  The only residue component present was the parent compound
    diflubenzuron.  Similar results were reported for cotton.  Studies on
    citrus fruits, apples and soybeans have confirmed that the only
    residue component is the parent compound diflubenzuron.  It can be
    concluded that plants do not metabolize diflubenzuron (Nimmo & de
    Wilde, 1974; Nimmo et al., 1978; Bull & Ivie, 1978; Nigg, 1989;
    Joustra et al., 1989; Serra & Joustra, 1990; van Kampen & Joustra,
    1991; Thus & van der Laan, 1993).

    4.3  Transformation

    4.3.1  Abiotic degradation

         Under environmental conditions abiotic degradation of
    diflubenzuron represents a very minor route of breakdown, owing to
    the stability of the substance.

    4.3.1.1  Photolysis

         On the basis of results from a 15-day photolysis experiment, a
    photolytic half-life of 40 days was calculated for diflubenzuron by
    regression analysis (Boelhouwers et al., 1988a,b).  After one week of
    storage at 50°C or after one day at 100°C, there was no significant
    decomposition (< 2%).  The solid is stable to sunlight.

    4.3.1.2  Hydrolysis

         Abiotic hydrolysis of diflubenzuron in solution does not occur at
    normal pH values.  At pH 9 the hydrolytic half-life is 32.5 days,
    4-chlorophenyl urea (4-CPU) and 2,6-difluorobenzoic acid (2,6-DFBA)
    being the degradation products (Boelhouwers et al., 1988a).

         High temperature (121°C) increases the degradation of
    diflubenzuron in aqueous media at levels greatly above its solubility
    in water and result in its rapid degradation to as many as seven

    identified products: 4-CPU, 2,6-DFBA, 2,6-difluorobenzamide,
    4-chloroaniline,  N,N'-bis (4-chlorophenyl) urea, 1-(4-chlorophenyl)-
    5-fluoro-2,4 (1H,3H)-quinazolinedione and 2-[(4-chlorophenyl) amino]-
    6-fluorobenzoic acid.  4-Chloroaniline,  N,N'-bis (4-chlorophenyl)
    urea and 2[(4-chlorophenyl) amino]-6-fluorobenzoic acid were not
    detected at lower temperatures (0.1 mg [14C]-diflubenzuron/litre
    water or buffer at 36°C).  4-Chloroaniline was a major degradation
    product of diflubenzuron in heat-treated samples, but it was not seen
    at lower temperatures (Ivie et al., 1980).

         The heat-induced degradation of diflubenzuron increased with
    increasing pH (Schaefer & Dupras, 1976).  Nigg et al. (1986) found
    that high temperature and humidity significantly decreased the half-
    life of diflubenzuron residues on citrus fruit.

    4.3.2  Biodegradation

    4.3.2.1  Water

    a)  Laboratory studies

         Degradation in water can also occur through microbial action,
    since in sterile water no breakdown or hydrolysis occurs (Boelhouwers
    et al., 1988a).  In freshly sampled ditch water,  Nimmo & De Wilde
    (1975a) demonstrated 50% degradation in 1-4 weeks.  The breakdown
    products were the same as the primary soil metabolites (4-CPU and
    2,6-DFBA).  Ivie et al. (1980) reported the same metabolites.  Anton
    et al. (1993) calculated the half-life of diflubenzuron in aerated and
    unaerated tap water to be less than half a day and less than one day,
    respectively.

         When diflubenzuron (1.3 mg/litre) was added to an anaerobic silt
    loam/water system, disappearance from the water phase showed a half-
    life of 18 days and  from the total system a half-life of 34 days. 
    The metabolites were 4-CPU and 2,6-DFBA, and almost no bound residue
    was formed (Thus et al., 1991).  After 90 days less than 2% of added
    diflubenzuron remained in the system (Thus & van Dyk, 1991).

         In another study, van der Laan-Straathof & Thus (1993) calculated
    the half-life of diflubenzuron in water to be 2.5 days.  Of the two
    degradation products, 4-CPU underwent no further degradation but
    2,6-DFBA was mineralized.

    b)  Outdoor models

         Schaefer et al. (1980) reported that, in pasture water with a pH
    of 8.2 and afternoon temperatures as high as 38-40°C, there was a
    decline from an initial nominal concentration of 30 µg/litre to a
    one-hour measured concentration of 20.3 µg/litre and subsequently to
    21.6, 13.6, 4.4, and 2.4 µg/litre on days 1, 2, 3, and 4 respectively.

         Schaefer & Dupras (1976) applied two formulations of
    diflubenzuron (a wettable powder and a flowable formulation) to
    artificial ponds of 1 m2 surface area containing 318 litres of pond
    water.  An initial concentration of 80 µg/litre decreased to 50%
    within about 2 days.  The diflubenzuron residue level after one week
    was 2-3 µg/litre.

         The half-life of diflubenzuron (1 µg/litre) in the aqueous
    fraction of sludge experiments was 4-15 h (Booth et al., 1987), and
    the half-life in sea water was reported to be less than 4 days 
    (Schimmel et al., 1983).  Cunningham & Myers (1986) estimated a half-
    life of less than 1 day for residues of diflubenzuron in water
    following three applications of 0.4% granules and three applications
    of 25% WP at a rate of 45 g a.i./ha to a supra-tidal salt marsh.

         Madder & Lockhart (1980) studied model ponds (20 m2) to which
    Dimilin WP-25 was applied at 56 g/ha (equivalent to 11.2 µg/litre). 
    For an unexplained reason, the measured concentration reached a
    maximum value of about 17.5 µg/litre, 4 days after treatment.  It
    decreased by around 50% during the next 5 days. A residue of
    2 µg/litre remained 2 weeks after application.  On the basis of a
    bioassay, a diflubenzuron half-life of about 3 days was calculated.

         Collwell & Schaefer (1980) applied diflubenzuron to five
    experimental ponds (each 100 m2) at a mean concentration of
    13 µg/litre.  The residue levels in water declined to an average of
    7.2 µg/litre after 24 h.

         In a study by Sarkar (1982), a 3 × 1 × 0.3 m open tank containing
    water was sprayed with a dispersion of Dimilin WP-25.  Three
    subsequent applications were made, giving diflubenzuron concentrations
    of 25, 35 and 50 µg/litre, respectively.  These concentrations
    decreased to 50% in about 3-4 days.

         Pritchard & Bourquin (1981) studied the environmental fate of
    diflubenzuron under simulated estuarine conditions in a laboratory
    continuous-flow estuarine system and a  static test system.  The
    hydrolytic half-life of diflubenzuron was 17 days in the static test
    system, whereas the biological half-life was 5 days.  4-Chloroaniline
    was not detected in either of the systems.

         Thus & van der Laan-Straathof (1994) studied the fate of
    diflubenzuron in two model ditch systems.  Diflubenzuron was added at
    a concentration of 0.94 mg/kg to two sediments (sandy loam and silt
    loam), both of which were covered with aerated surface water. It
    disappeared rapidly from the water phase through degradation and
    adsorption to the sediment, the half-lives being 1.9 and 1.1 days,

    respectively. Dissipation of diflubenzuron from the complete sandy
    loam and silt loam systems occurred with half-lives of 25 and 10 days,
    respectively.  The metabolites (> 1% of the added diflubenzuron)
    consisted of CO2, 4-CPU and 2,6-DFBA.

    c)  Field studies

         Apperson et al. (1978) described the treatment of three farm
    ponds with  diflubenzuron levels of 2.5, 5 and 10 µg/litre, and a lake
    with 5 µg/litre.  Shortly after the  application, a rapid decline in
    diflubenzuron residues occurred, resulting in half-life values of only
    a few days.  In the lake no residues were found in the sediment
    samples, suggesting that diflubenzuron was rapidly dissipated before,
    or upon reaching, the bottom sediment.

         Hester (1982) applied diflubenzuron at 0.045 kg a.i./ha to
    specially constructed estuarine ponds.  The water residue levels
    decreased rapidly from 7.5 to 2 µg/litre in 2-3 days (study II) and
    from 3.3 to 0.6 µg/litre in 7 days (study I).

    d)  Discussion and appraisal

         The rate of decrease in diflubenzuron concentration after
    application of the formulated product to natural waters depends on the
    combined action of many environmental factors.  Factors affecting the
    degradation rate of diflubenzuron include the acidity (pH), the
    relative local abundance of soil and organic debris, and the water
    depth.

         Half-life values vary from less than 4 days to 4 weeks in
    laboratory experiments.

         The use of artificial ponds or basins, preferably outdoors,
    yields more relevant data and fairly consistent results.  Dissipation
    half-life values vary from 1-5 days after diflubenzuron has been
    applied at recommended rates.

         The dissipation half-life of diflubenzuron in the aquatic
    environment is between one day and one week in most cases, depending
    on the properties of the applied formulation and on the
    characteristics of the application site.  The presence of organic
    sediments (hydrosoil, plant debris) and a relatively high local
    temperature are factors that particularly accelerate the disappearance
    of diflubenzuron.

    4.3.2.2  Soil

    a)  Mobility in soil

         Diflubenzuron is immobile in soil, as demonstrated by Helling
    (1985) in column leaching experiments and Booth et al. (1987) in
    adsorption-desorption studies with eight soil types.

         The work of Carringer et al. (1975) suggests that soil organic
    matter is an important parameter in soil adsorption.  Due to its
    immobility in soil, diflubenzuron is not likely to contaminate
    groundwater by vertical movement in soil or to contaminate open water
    by lateral movement in groundwater.

         This has been confirmed in studies carried out in field soils
    with growth of citrus fruits (Verhey, 1991a; Kramer, 1991), apple
    (Kramer, 1990, Verhey, 1991b), soybean (Kramer, 1992b) and cotton
    (Kramer, 1992a).  After three applications of diflubenzuron (Dimilin
    25W) at normal rates, most residue was found in the top 15 cm of soil
    and no residue was encountered below 30 cm.

    b)  Degradation in soil

         The rates of disappearance of technical diflubenzuron applied at
    10 mg/kg on quartz sand to natural sandy loam and muck soils were
    significantly greater than for the corresponding sterilized soils
    (e.g., 2-12% and 80-87% diflubenzuron, respectively, remaining at
    12 weeks), demonstrating that soil microorganisms play a major role in
    their degradation (Chapman et al., 1985).

         Diflubenzuron is very rapidly hydrolysed in soil.  The half-life
    time is 2 days to one week.  The primary metabolites are 2,6-DFBA and
    4-CPU. The process is microbial, since in sterilized soil no breakdown
    occurs.  The rate of breakdown is strongly dependent on the particle
    size of diflubenzuron (see Fig. 1) (Nimmo et al., 1984, 1986).

         The half-life in water in alkaline pastures is 1 day and in
    neutral lake water it is from 10 to 15 days (Nimmo & de Wilde, 1975a).

         Metabolic routes other than 4-CPU and 2,6-DFBA are virtually
    irrelevant.  Both primary metabolites are further metabolized,
    2,6-DFBA with a half-life of about 4 weeks and 4-CPU with a
    dissipation time of 1 to 3 months.  Radiolabelling of both primary
    metabolites and of a carbon atom in the ureido bridge shows carbon
    dioxide development from mineralization.  However, both the benzoic
    acid ring carbon and the ureido bridge carbon are mineralized
    much faster than the aniline moiety carbon, suggesting that
     para-chloroaniline (PCA) is a major secondary metabolite that is
    virtually irreversibly bound to soil (Bollag et al., 1978; Mansager et
    al., 1979; Nimmo et al., 1984, 1986, 1990).

    FIGURE 2

         Even as a bound residue PCA is metabolized.  Apparently, the
    breakdown of 4-CPU in soil is a complex process in which PCA is a
    transient metabolite or intermediate.  The breakdown process leads to
    products beyond the aniline structure.  If PCA is applied to soil,
    6 weeks incubation at 25°C yields 60% breakdown products of a
    different nature (Bollag et al., 1978).  The aniline itself is firmly
    bound to soil and immobilized (Hsu & Bartha, 1974; Moreale & van
    Bladel, 1976; Bollag et al., 1978; Simmons et al., 1989).

         Fig. 2 shows metabolic pathways in soil.

         The main metabolic pathway (over 90%) is hydrolysis, leading to
    2,6-DFBA and 4-CPU.  The second site of cleavage occurs at CœN bonds 2
    and 3.  Both reactions lead to the formation of 2,6-difluorobenzamide
    (DFBAM), which readily hydrolyses to 2,6-DFBA (Verloop & Ferrell,
    1977; Nimmo et al., 1984).

         The major metabolite in an activated sludge system is 4-CPU. 
    This is the major metabolite reported in most soil metabolism
    experiments (Booth et al., 1987).  4-CPU was found to be converted
    into bound residues with a half-life of 5-10 weeks.  In the bound
    residues, 4-CPU and PCA were present in roughly equal amounts after
    2 months (Verloop & Ferrell, 1977).  Free PCA was not found in soil
    (Nimmo et al., 1986).  The soil type and characteristics appear to
    have no influence on the rate of degradation (Nimmo et al., 1984).

         Metcalf et al. (1975) found no significant degradation of
    diflubenzuron in a silty clay loam after incubation at 26.7°C for
    periods of 1, 2 and 4 weeks.  However, the authors did not take into
    account the particle size of the soil, and used techniques that have a
    negative influence on breakdown.

         The rate of degradation of 14C- or 3H-diflubenzuron applied to
    a mushroom growth medium (dose 2 g/m2) was between 30-50% in one
    month.  The main degradation products, 4-CPU and 2,6-DFBA, were
    absorbed from the growth medium by the  mushrooms, resulting in
    residue levels of 0.1-0.6 mg/kg and 1-3 mg/kg, respectively (Nimmo &
    de Wilde, 1977a).  Free PCA or its further possible degradation
    products were not present in the extractable residues (Nimmo & de
    Wilde, 1975a; Verloop & Ferrell, 1977).  Organic matter in soil
    significantly contributed to the adsorption of chloroaniline compounds
    and their immobilization (Hsu & Bartha, 1974; Moreale & van Bladel,
    1976; Bollag et al., 1978).

         Nimmo & de Wilde (1975a) found a degradation half-life of
    0.5-1 week at a diflubenzuron concentration of 1 mg/kg (corresponding
    to an application dose of approximately 300 g/ha).  2,6-DFBA was
    degraded with a half-life of approximately 4 weeks, and 4-CPU with a
    half-life of 2-3 months.

    FIGURE 3

         Walstra & Joustra (1990) applied 0.69 mg diflubenzuron/kg to
    sandy loam.  When incubated in the dark at 24°C, they obtained a half-
    life for diflubenzuron of 50 h.

         Diflubenzuron was found to be rapidly degraded by four soil fungi
    ( Fusarium sp.,  Cephalosporium sp.,  Penicillium sp. and  Rhodotorula
    sp.), the half-lives being 7, 13, 14 and 18 days, respectively
    (Seuferer et al., 1979).

         Several degradation studies on diflubenzuron (Dimilin 25 W) in
    field soils have been conducted (Kramer, 1990, 1991, 1992a,b; Verhey,
    1991a,b).  Most of the degradation half-lives were between one and two
    weeks, except in the case of the two Verhey studies, which yielded
    half-lives of more than two months.  In all studies, the metabolites
    were 4-CPU and 2,6-DFBA.

         No degradation of diflubenzuron by the soil microorganism
     Pseudomonas putida was observed (Booth & Ferrell, 1977).

    4.4  Bioaccumulation and biomagnification

         Metcalf et al. (1975) studied the fate of 14C-diflubenzuron in a
    laboratory model ecosystem. Diflubenzuron was clearly persistent in
    some organisms, such as algae  (Oedogonium cardiacum), snails
    ( Physa sp.), caterpillars  (Estigmene acrea) and mosquito larvae
     (Culex pipiens quinquefasciatus).  The fish  Gambusia affinis was
    able to degrade diflubenzuron more efficiently. Diflubenzuron did not
    biomagnify in the fish through food chain transfer.  The biomagnifi-
    cation was about 40-fold greater in mosquito larvae than in  Gambusia
     affinis.

         When the bluegill sunfish  (Lepomis macrochirus) was exposed to
    10 µg diflubenzuron/litre for 24 h the tissues contained an average of
    264 µg/kg.  After 24 to 48 h of exposure, fish degraded and eliminated
    the diflubenzuron. The excretory products were neither the parent
    compound nor 4-CPU. The amount of diflubenzuron remaining in fish
    tissues at various times was dependant on the reduction of residue
    concentration in water.  However, the potential for degradation and
    elimination was very great (Schaefer et al., 1979).

         A dynamic 42-day study was conducted by Burgess (1989) in order
    to evaluate the bioconcentration of 14C-diflubenzuron by bluegill
    sunfish  (Lepomis macrochirus).  A flow-through proportional diluter
    system was used for a 28-day exposure period.  Radioanalysis of
    fillet, whole fish and visceral portions was performed throughout the
    exposure period. Daily bioconcentration factors ranged from 34 to 200,
    78 to 360, and 100 to 550 for fillet, whole fish and viscera,
    respectively.  Uptake tissue concentrations of 14C-diflubenzuron
    ranged from 0.25 to 1.7 mg/kg for fillet, 0.58 to 3.3 mg/kg for whole
    fish, and 0.75 to 4.7 mg/kg for viscera.  To measure the elimination

    of 14C-diflubenzuron, the test fish were placed in clean water for 14
    days.  Radioanalysis throughout the depuration period indicated 99%
    depuration for each of fillet, whole fish and viscera.  The fillet
    concentration of 14C-diflubenzuron decreased from 1.6 mg/kg on day 28
    of exposure to 0.012 mg/kg by day 14 of the depuration period.  Whole
    fish levels decreased from 3.3 mg/kg on day 28 of exposure to
    0.038 mg/kg by the end of the study; whereas, viscera concentrations
    dropped from 4.4 mg/kg on day 28 of exposure to 0.056 mg/kg by day 14
    of depuration.  BIOFAC modelling estimated the uptake rate constant
    (K1) to be 370 (± 57) mg/kg fish per mg/litre water per day, the
    depuration rate constant (K2) 1.2 (± 0.18) day-1, the time for 50%
    depuration 0.60 (± 0.09) days, the bioconcentration factor (BCF) 320
    (± 70), and the time to reach 90% or steady state 2.0 (± 0.31) days.
    The BIOFAC-calculated BCF value was the same as the observed mean
    whole fish BCF of 320 for days 3, 7, 14, 21 and 28.  Fig. 3 shows the
    accumulation, plateauing and depuration in this study.

         During the study, no mortality or abnormal behaviour was observed
    in the test fish.  This appeared to indicate that the test fish were
    in good health and would provide acceptable data for defining the
    uptake/depuration potential of 14C-diflubenzuron.  Analysis of fish
    revealed parent compound (80%), 2,6-difluorobenzamide (10-13%) and
    three other minor metabolites (one of which probably was 4-CPU).  PCA
    was demonstrated to be absent (sensitivity limit 0.01 mg/kg).

         White crappies  (Pomoxis annularis) contained residues from
    355.1 to 62.2 µg/kg at 4 and 21 days, respectively, following
    treatment of a lake with 5 µg diflubenzuron/litre (Apperson et al.,
    1978).

         Channel catfish  (Ictalurus punctatus) did not bioaccumulate
    diflubenzuron residues (less than 0.05 mg/kg) from treated soil in a
    simulated lake ecosystem (Booth & Ferrell, 1977).

         Assuming a biomagnification of 50-160, and that fish are capable
    of rapidly depleting residues from the body, the likelihood of fish
    accumulating significant residues of diflubenzuron is low (Apperson et
    al., 1978; Schaefer et al., 1980).

    4.5  Interaction with other physical, chemical or biological factors

         Schaefer & Dupras (1976) reported that application of the
    technical grade compound in an ethanol carrier or as a flowable liquid
    formulation resulted in higher concentrations in the upper water
    levels of mosquito ponds for a period of 3 days following spray
    treatment than in the case of spray treatment with wettable powder
    formulation (the actual formulation used for mosquito control
    spraying).

    FIGURE 4

         Seuferer et al. (1979) reported that the soil microorganisms
     Rhodotorula sp.,  Penicillium sp. and  Cephalosporium sp. cannot
    utilize diflubenzuron as a sole carbon and energy source.  However,
    accelerated breakdown of diflubenzuron occurred in the presence of
    these organisms.

    4.6  Ultimate fate following use

         It appears that after direct spraying diflubenzuron is persistent
    on foliage, it remains almost completely at the site of application on
    the surface, and it does not penetrate the plants.

         Diflubenzuron is readily degraded in soils of various types and
    origin under aerobic or anaerobic conditions with a half-life in the
    range of 0.5 to 1 week.  It is metabolized by microorganisms
    principally to 4-CPU and 2,6-DFBA.  The latter is unstable with a
    half-life of 3-5 days (Nimmo et al., 1984) to 4 weeks (Verloop &
    Ferrell, 1977).  The half-life of 4-CPU is about 6 weeks (Nimmo et
    al., 1984).  Free PCA has not been detected in soil.

         In spite of rapid degradation in soil, small amounts of residue
    (up to 1 mg/kg, depending on ageing time and growth stage of plants)
    may be taken up by crops in treated soil (Thus et al., 1994).

         Field applications of diflubenzuron produce soil residues which
    might possibly lead to residues in rotational crops by re-uptake from
    soil.

         Studies with direct applications to field water show a moderate
    persistence of diflubenzuron in water.  Half-life values average one
    week or less.  This rapid rate of loss may be more dependent on
    adsorption to organic matter than on microbial degradation.

    5.  ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE

    5.1  Environmental levels

    5.1.1  Air

         No information is available on air concentrations of
    diflubenzuron.

    5.1.2  Water

         A total of 1160 ha of insect-infested forest in Finland was
    sprayed with diflubenzuron (25% WP) from a fixed-winged aircraft at an
    application rate of 75 g a.i. in 50 litres water per ha.  The residues
    in "run-off" water (gathered in specially dug pits adjacent to the
    sprayed area) decreased from 5 µg/litre one day after spraying to
    0.1 µg/litre after 2 months.  The concentration in water in open pits
    was 0.1 µg/litre 1 and 7 days after application and 0.2 µg/litre
    1 month after application.  After 2 months no residues were detected. 
    All water samples taken from outside the treated area contained less
    than 0.1 µg/litre (the limit of sensitivity) (Mutanen et al., 1988).

         Diflubenzuron was found in the water of the Fraser River, Canada,
    up to 71 days following application with diflubenzuron (1% granular
    formulation) at a rate of 4.5 kg/ha (45 g a.i./ha).  The peak value
    was 1.8 µg/litre 8 days after treatment (Wan & Wilson, 1977).

         After aerial application of diflubenzuron (25% WP formulation) to
    two forest ponds in Canada, the maximum residue levels in water,
    sediment, aquatic plants and fish were 13.82 µg/litre (at 1 h),
    0.24 mg/kg (at 1 day), 0.36 mg/kg (at 1 day) and 0.11 mg/kg
    (at 1 day), respectively.  The rate of dissipation was rapid, non-
    detectable levels being reached in 20 days for water, 5 days in
    aquatic plants and 3 days in fish (Kingsbury et al., 1987).

         A pond in Salt Lake County, Utah, USA, was treated with three
    applications of diflubenzuron at a rate of 280.25 g a.i./ha. 
    Diflubenzuron was found at less than 0.05 mg/litre 4 days following
    treatment (Booth et at., 1987).

         Residues in three farm ponds in California treated with
    diflubenzuron (2.5, 5 and 10 µg/litre) averaged 1.9, 4.6 and
    9.8 µg/litre, respectively, 1-4 h after the applications.  They
    declined steadily averaging 0.5, 0.3 and 0.2 µg/litre, respectively,
    2 weeks later.  Residues in a small lake treated at 5 µg/litre
    averaged 3.3 µg/litre following treatment and 0.4 µg/litre after
    35 days.  No residues were found in sediment samples taken post-
    treatment (Apperson et al., 1978).

         One hour after a single application of 45 g diflubenzuron/ha to
    brackish water pools the residues in water and in sediment were
    3.6 µg/litre and 80 µg/kg, respectively.  The concentration in
    sediment increased to 520 µg/kg after 1 day and reached its maximum of
    780 µg/kg 4 days following application (Hester et al., 1986).  After
    6 applications of diflubenzuron at a rate of 145.73 g/ha to Utah Lake,
    USA, the residues in sediments were less than 0.05 mg/kg (Booth et
    al., 1987).

         Other field studies with similar results have been reported by
    Anon (1980), Smith & Edmunds (1985), Van Den Berg (1986), Huber &
    Collins (1987), Jones & Kochenderfer (1988),  Huber & Manchester
    (1988), Downey (1990) and Sundaram et al. (1991).  It is clear that a
    variety of application scenarios will result in measurable residues of
    diflubenzuron in water (Table 6).

         The overall conclusion is that diflubenzuron residues in stagnant
    water dissipate rapidly within days.  In flowing water, e.g., in
    wooded areas, diflubenzuron residues may peak shortly after rainfall
    but such peak concentrations are very transient in nature.

    5.1.3  Food and feed

         Data on residues in food resulting from treatment with
    diflubenzuron have been summarized by FAO/WHO (1982a,b, 1985a,b,
    1986a,b).

         Residue data obtained from various countries showed residues in
    apples below 1.0 mg diflubenzuron/kg at 2 weeks after the last
    application at recommended rates.  Residues in whole citrus fruit were
    below 0.5 mg/kg 1 week after the last application at the recommended
    rate.  Residues in soybean seed and cottonseed were generally below
    the limit of determination (0.05 mg/kg).

         Mushrooms have a residue pattern different from other plant
    material.  In mushrooms growing on diflubenzuron-treated soil, high
    levels of the metabolite 2,6-DFBA are taken up from the soil.
    Diflubenzuron was found at a level of 0.1 mg/kg, while the 2,6-DFBA
    level was around 1 mg/kg (see chapter 4).

         Residues in wild mushrooms after aerial application to forests in
    Finland were on average 0.07 mg/kg 1 week after spraying with 75 g
    diflubenzuron in 50 litre water per ha.  In bilberries the residues
    decreased on average from 0.2 mg/kg 1 day after spraying to 0.09 mg/kg
    after 1 month (Mutanen et al., 1988).

         Diflubenzuron applied as a wettable powder spray to growing
    alfalfa at 20-100 g/ha showed initial residue levels of 1.8-8.5 mg/kg. 
    Residues of 0.3-1.5 mg/kg remained 22 days after applications (Lauren
    et al., 1984).

        Table 6.  Summary and comparison of experimental parameters among key studies designed to measure environmental concentrations of
              diflubenzuron in water
                                                                                                                                              

    Medium       Formulation  a.i.%  Method of      Application     Maximum           Time for       Minimum          Time for       References
    treated                          application    rate a.i.       concentration     maximum        concentrationa   minimum
                                                                                      concentration                   concentration
                                                                                                                                              

    Farm ponds      25 WP     25     hand sprayer   2.5-10 µg/litre  1.9-9.8 µg/litre    1-4 h       0.5-0.2 µg/litre    14 days      Apperson
    (0.06-0.2 ha)                    from boat                                                                                        et al.
                                                                                                                                      (1978)

    Small lake      25 WP     25     hand sprayer   5 µg/litre       3.3 µg/litre        4 h         0.4 µg/litre        35 days      Apperson
    (18.6 ha)                        from boat                                                                                        et al.
                                                                                                                                      (1978)

    Pond            W-25      25     hand-operated  0.28 kg/ha       56 µg/litre         96 h        < 0.01 µg/litre     40 days      Booth
                                     spray                                                                                            et al.
                                     applicator                                                                                       (1987)

    Brackish        25 WP     25     clothes        0.045 kg/ha      7.5 µg/litre        48-72 h     < 0.3 µg/litre      25-30 days   Hester
    pools                            sprinkler                                                                                        (1986)

    Forest ponds    25 WP     25     aircraft       0.07 kg/ha       13.82 µg/litre      1 h         < DL                20 days      Kingsbury
    (25 ha)                          (four                                                                                            et al.
                                     atomizers)                                                                                       (1987)

    Field plot      25 WP     25     fixed-wing     0.075 kg in 50   5.0 µg/litre        24 h        < DL                60 days      Mutanen
    (1160 ha)                        aircraft       litre water/ha                                                                    et al.
                                                                                                                                      (1988)

    Fixed plots     granular  1.0    aircraft       0.023 kg/ha,     1.8 µg/litre        192 h       < DL                60 days      Wan &
    (3-40 ha)                                       0.46 kg/ha                                                                        Wilson
                                                                                                                                      (1977)
                                                                                                                                              
    a  DL = determination limit
             After two soil applications of 67.26 g/ha, the residues of
    diflubenzuron in the rotational crops (wheat, cabbage and onions) were
    less than 0.01 mg/kg (Danhaus & Sieck, 1976).

         Mian & Mulla (1983) studied the persistence of diflubenzuron in
    stored wheat after applications of 1, 5 and 10 mg/kg.  The residue
    levels were 0.59, 2.75 and 5.00 mg/kg, respectively, 23 months after
    treatment.

    5.1.4  Forest plants and litter

         The level of diflubenzuron residues in pine needles was on
    average 3.0 mg/kg 1 day after application to the forest in Finland at
    a rate of 75 g diflubenzuron in 50 litres water per ha.  The level had
    decreased to 0.2-0.3 mg/kg or was not detectable 2 months later
    (Mutanen et al., 1988).

         Booth et al. (1987) found diflubenzuron residues of less than
    0.05 mg/kg in the forest litter 1, 4, 10 and 21 days after treatment
    with 0.28 kg a.i./ha.

         Sundaram (1986) studied the residues in a forest in Canada after
    simulated aerial spraying of diflubenzuron in acetone and in fuel oil:
    Arotex 3470 mixture, each at 90 g a.i. in 18 litre/ha.  The residue
    levels 1 h after application varied, respectively, from 23.8 to
    30.6 µg/g in foliage and from 3.08 to 4.60 µg/g in litter.  Forty-five
    days after spraying the residue levels in foliage were 0.80 and
    3.9 µg/g, respectively, for the above-mentioned formulations.

         Spray deposit patterns and persistence of diflubenzuron in white
    pine ( Pinus strobus L.) and sugar maple ( Acer saccharum Marsh.)
    canopies, forest litter and soil were studied after aerial application
    of a 250 g/kg wettable powder formulation (Dimilin WP-25) at
    70 g a.i./ha, using three volume rates (2.5, 5 and 10 litres/ha), over
    three blocks in a mixed forest near Kaladar, Ontario, Canada, during
    1986 (Sundaram, 1991).  In the spray block that received 10 litres/ha,
    diflubenzuron persisted in foliage as long as 120 days after
    treatment, but it lasted for only about a week in forest litter and
    soil samples.  At 2.5 and 5 litres/ha, diflubenzuron failed to persist
    in foliage as long, and residues in litter and soil, which were barely
    above the quantification limit, persisted only for a few days.

    5.1.5  Aquatic organisms

         Residues in fish are given in section 4.4.

    5.2  General population exposure

         Exposure of the general population to diflubenzuron via food and
    drinking-water may occur.

         Twelve volunteers with whole body dosimeters were exposed for 4 h
    to Dimilin 25 W after simulated indoor treatment of carpets at
    0.16 g/m2.  Average deposition was 5.3 ± 2.3 µg diflubenzuron/cm2
    carpet.  Total dermal exposure ranged from 0.053 to 0.25 mg/kg body
    weight per day to (average 0.15 ± 0.066 mg/kg body weight per day). 
    Assuming a dermal absorption of 0.2%, the total exposure via the
    dermal route was calculated to be 0.0003 mg/kg body weight per day. 
    Air concentrations ranged from 10.2 to 32.4 µg/m3 during the first
    4 h and were < 1 µg/m3 at 12-16 h.  The total respiratory exposure
    was calculated to be 0.0011 mg/kg body weight per day.  The total
    exposure, via the dermal and respiratory route, was calculated to be
    0.0014 mg/kg body weight (Honeycutt, 1993).

    5.3  Occupational exposure during manufacture, formulation or use

         In a US Department of Agriculture report, human exposure via a
    variety of exposure scenarios was estimated using standardized methods
    and assumptions. The exposure scenarios included mixing and loading by
    workers, via aircraft or truck spillage, and general public exposure
    via the diet or resulting from occupational aerial spraying.  Dermal
    absorption of diflubenzuron was assumed to be 10%.  Estimated
    realistic doses for humans were < 0.003 mg/kg body weight per day
    except where aircraft or truck spillages occurred, in which case
    exposures were significantly higher.  Estimated worst-case doses for
    humans were < 0.01 mg/kg body weight per day, except where aircraft
    or truck spillages occurred (USDA, 1985).

    6.  KINETICS AND METABOLISM IN LABORATORY ANIMALS AND HUMANS

    6.1  Absorption

         Diflubenzuron is absorbed from the digestive tract but only
    poorly absorbed through the skin.  Willems et al. (1980) found that in
    rats the relative intestinal absorption diminished greatly with
    increasing dose.  Following a dose of 4 mg/kg body weight 42.5% was
    absorbed, but only 3.7% of a 900 mg/kg body weight dose was absorbed.

         Dermal absorption of 14C-diflubenzuron was only 0.2% when it was
    applied to the shaved skin of rabbits as an aqueous micro-suspension
    of 150 mg/kg (De Lange, 1979).

         When applied dermally to cattle 14C-diflubenzuron was not
    absorbed or degraded through the skin to any detectable degree (Ivie,
    1978).

    6.2  Distribution

         Body tissues show little tendency to retain diflubenzuron. 
    Analysis of tissues for radiocarbon residues, 4 days (for sheep) or
    7 days (for cows) after a single oral dose of 10 mg/kg body weight,
    indicated that only the liver contained appreciable levels of
    radioactivity, ranging from 2 to 4 mg/kg diflubenzuron equivalents
    (Ivie, 1977).

         More than one third of an oral diflubenzuron dose appeared in the
    bile of a cannulated sheep (Ivie, 1977).

         The highest 14C-diflubenzuron residue present in pig tissues
    after a single oral dose of 5 mg/kg body weight was 0.43 mg/kg in the
    gall bladder.  All other tissue residue levels were found to be less
    than 0.30 mg/kg (Opdycke et al., 1982a).

         Twenty-two dairy cows were fed 14C-diflubenzuron (labelled in
    both phenyl moieties) in a diet at dose levels of 0.05, 0.5, 5, 25 and
    250 mg/kg feed for 28 days. Residues in blood, fat and muscle were
    below the detection limit (0.0067-0.04 mg/kg) at all dose levels. They
    were only detected following a dose of 250 mg/kg in the liver and
    kidney where residues were 6.040 and 1.038 mg/kg, respectively. 
    Residues in milk were found at dose levels of 5 and 250 mg/kg, where
    the highest levels of diflubenzuron were 0.009 and 0.20 mg/kg,
    respectively (Smith & Merricks, 1976a).

         In a study by Miller et al. (1976a), two dairy cows were fed
    diflubenzuron at 0.25 or 1 mg/kg body weight per day for 4 months.  A
    third cow received an increased dosage of 8 to 16 mg/kg body weight

    per day, the highest value being maintained for three months.  In the
    fat, liver and milk of the third cow, residues were 0.2, 0.13 and
    0.02 mg/kg, respectively.

         When dairy bull calves (four treated and four controls) received
    diflubenzuron at 1.0 to 2.8 mg/kg body weight, residues were detected
    only in the tissue samples of one bull (0.02 mg/kg in liver and
    kidney, 0.04 mg/kg in the subcutaneous fat, and 0.08 mg/kg in the
    renal and omental fat (Miller et al., 1979).

         The maximum total residue in eggs 3 days after a single dose of
    5 mg/kg 14C-diflubenzuron to hens was 0.248 mg/kg (Opdycke, 1976).

         When laying hens were administered 14C-diflubenzuron at dose
    levels 0.05, 0.5, and 5.0 mg/kg feed for 28 days, dose-related
    residues ranging from 0.007 mg/kg at the lowest to 1.2 mg/kg at the
    highest dose level were found in kidney, liver and fat.  After 7 days
    of withdrawal, residues in all tissues and eggs were below the
    detection limit (0.0006-0.032 mg/kg) for all dose levels (Smith &
    Merricks, 1976b).

         When diflubenzuron was fed to white leghorn and black sex-linked
    cross hens at a level of 10 mg/kg feed for 15 weeks, detectable
    residues were found in eggs, liver and visceral fat.   Residues were
    significantly higher in eggs from white leghorn hens than in eggs from
    black sex-linked cross hens, the average levels being 0.55 and
    0.38 mg/kg, respectively (Miller et al., 1976b).

    6.3  Metabolic transformation

         The metabolic fate of diflubenzuron has been studied in various
    species.  Metabolic pathways of diflubenzuron are shown in Fig. 4

         In rats and cows the major metabolic pathway involves
    hydroxylation of the phenyl moieties of the compound. About 80% of the
    metabolites in rat urine were identified as 2,6-difluoro-3-
    hydroxydiflubenzuron and 4-chloro-2-hydroxy- and 4-chloro-3-
    hydroxydiflubenzuron.  About 20% underwent scission of the benzoyl
    ureido bridge.  The major part was excreted as 2,6-DFBA and
    constituted more than half of the urinary metabolites.  4-CPU was not
    detected in bile or urine in a significant quantity (De Lange et al.,
    1975; Willems et al., 1980).

         The major metabolite in cow urine was 2,6-difluoro-3-hydroxy-
    diflubenzuron (45%). Relatively small quantities of 4-chloro-2-
    hydroxy- (1.6%) and 4-chloro-3-hydroxydiflubenzuron (3.7%) and the
    scission products 4-CPU (0.6%), 2,6-DFBA (6.0%) and 2,6-difluoro-
    hippuric acid (6.9%) were present (Ivie, 1978).

    FIGURE 5

         The major metabolites (approximately 50%) in sheep urine were
    2,6-DFBA and 2,6-difluorohippuric acid (Ivie, 1978).

         14C-Diflubenzuron uniformly radiolabelled in both rings was
    administered to a pig as an oral dose of 5 mg/kg body weight.  Of the
    administered dose, 82% was eliminated in faeces as parent compound and
    5% was recovered in urine.  Identification of the metabolic products
    in urine revealed 2,6-DFBA (0.28% of the dose), 4-CPU (0.82%), PCA
    (1.03%) and 2,6-difluorobenzamide (0.83%).  Cleavage of the urea
    moiety between the benzoyl carbon and urea nitrogen was shown to be
    the primary degradation pathway in pigs (Opdycke et al., 1982a).

         In chickens only small quantities of the metabolites 2,6-DFBA,
    4-CPU and PCA were found in excreta and tissues (Opdycke, 1976).
    Neither induction nor inhibition of mixed-function oxidase activity
    altered diflubenzuron metabolism in chickens (Opdycke et al., 1982b).

         After 4 days daily doses of 7.8 g diflubenzuron/kg body weight,
    De Bree et al. (1977) found PCA at a level of 30 ng/ml in rat plasma
    and 323 ng/g in erythrocytes.  PCA, estimated by the concentration in
    the urine, represented at most 0.01% of the dose actually absorbed.

    6.3.1  Metabolites - distribution, excretion, retention and turnover

         When 14C-PCA was administered orally as single doses of 0.3,
    3.0 or 30.0 mg/kg to male Fischer-344 rats, approximately 75% of the
    administered radioactivity was excreted in the urine within 24 h,
    while approximately 10% appeared in the faeces.  Excretion was
    virtually complete (92-97%) 7 days after dosing.  The highest tissue
    levels of radioactivity following a single intravenous dose of
    3.0 mg/kg were found in the liver, fat, muscle and skin.  Tissue
    levels peaked within 5-60 min after dosing. By 3 days, concentrations
    in all tissues except the blood had declined to < 0.3% of the dose
    (Sipes & Carter, 1988).  At this time, the only tissue containing more
    than 1% of the dose was the cellular compartment of blood, which
    contained 1-2% of the dose.  The decline of PCA concentration in all
    tissues, except for urine, faeces and intestinal contents, was
    biexponential. The t alpha 1/2 for fat, muscle and skin was about
    1.5 h, while the tß1/2 was approx. 43-59 h. The t alpha 1/2 for
    liver was 3.5 h.  Levels of unchanged PCA in all tissues peaked after
    5 min following intravenous administration.  The highest amount of
    unchanged PCA was attained in muscle (15% of radioactivity in the
    tissue) followed by skin (6%), fat (3%) and liver (2%). The decline of
    PCA in all tissues, except for the liver, followed biexponential kinetics
    with an estimated t alpha 1/2 of 8 min and a tß1/2 of 3 to 5 h.
    PCA is rapidly metabolized to  p-chloroacetanilide (PCAA) as the
    initial step in the metabolism and excretion of PCA.  The decline of PCAA
    was monoexponential, the appearance half-life being approx. 6 min in the
    testes and 15 min in the brain.  The elimination half-life in the
    brain, kidney, testes, muscle, skin and fat was around 1.0 to 2.0 h. 
    The elimination of PCA does not depend on either the dose or route of

    administration.  Approximately 4% of the urinary radioactivity in the
    0-24 h urine sample was unchanged PCA; less than 1% was found in the
    faeces.  PCAA was not detected in either urine or faeces over a 3-day
    period (Sipes & Carter, 1988).

         After a single intravenous dose of 14C-PCA (3 mg/kg), maximal
    tissue levels were reached within 15 min in most tissues.  At this
    time, most of the radioactivity was located in muscle (34%), fat
    (14%), skin (12%), liver (8%) and blood (7%).  Elimination half-lives
    from tissues ranged between 1.5 and 4 h.  By 8 h, approximately 90% of
    the administered dose had been eliminated into urine and faeces.  By
    3 days, concentrations in all tissues, except blood, had declined to
    < 0.3% of the dose (US NTP, 1989).

    6.4  Elimination and excretion

         After oral administration to rats of 5 mg diflubenzuron labelled
    with 3H in the benzoyl and with 14C in the aniline moiety, 95% of
    the 3H and 70-75% of the 14C radioactivity were retrieved in urine
    and faeces.  2,6-DFBA was shown to constitute more than half of the
    urinary metabolites (De Lange et al., 1977).  Up to 1% of an oral dose
    of 5 mg 14C-diflubenzuron labelled at the benzoyl moiety was
    recovered in the expired air of rats (De Lange et al., 1974; Willems
    et al., 1980).

         When 14C-diflubenzuron, labelled in the aniline moiety, was
    administered by gavage (4, 16, 48, 128, 900 and 1000 mg/kg body
    weight) to rats, the urinary excretion was complete after 48-72 h. 
    Urinary excretion after single oral administration of diflubenzuron
    relatively decreased with increasing dose level, being 27.6% of the
    dose at 4 mg/kg and 1% at 1000 mg/kg (De Lange et al., 1977).

         When 14C-diflubenzuron was administered at single oral doses of
    12.5, 63.5, 202.5 and 925 mg/kg body weight to Swiss mice, the
    excretion was almost completed within 48 h.  The cumulative percentage
    of the dose excreted in the urine decreased from 15% at the dose level
    of 12.5 mg/kg to approximately 2% at 925 mg/kg (De Lange & Post,
    1978).

         Hawkins et al. (1980) studied the excretion of radioactivity in
    urine and faeces after oral administration of 3H/14C-diflubenzuron
    (7 mg/kg) to male cats.  The radioactive dose was given on day 10 of a
    15-day dosing regime of non-radioactive diflubenzuron (days 1-9 and
    days 11-15).  The excretion of radioactivity in urine accounted for
    9.5 and 9.6% of the 14C and 3H doses, respectively, during 6 days
    after dosing.  The elimination of radioactivity in faeces accounted
    for 77.3 and 71.6% of the 14C and 3H doses, respectively, during 6
    days after dosing.

         After an oral administration of 14C-diflubenzuron (5 mg/kg) to
    female pigs, 82% of the dose was eliminated via faeces and 5% via
    urine in 11 days (Opdycke, 1976).

         About 85% of a single oral dose of 14C-diflubenzuron (10 mg/kg
    body weight) administered to a cow was recovered in the faeces during
    the first 4 days after treatment.  About 15% was recovered in urine
    and only about 0.2% was secreted in the milk (Ivie, 1977, 1978).

         Sheep excreted 41% of the dose (10 mg/kg) in the urine and 43% in
    the faeces during the 4 days after treatment.  Bile-cannulated sheep
    eliminated 24% of the dose in the urine, 32% in the faeces and 36% in
    the bile.  Sheep treated with 500 mg 14C-diflubenzuron/kg as a single
    oral dose eliminated a much smaller proportion of the 14C in urine
    and bile.  This was probably due to reduced absorption from the
    gastrointestinal tract when the sheep were given an exaggerated dose
    (Ivie, 1977).

         An oral dose of 5 mg 14C-diflubenzuron/kg administered to white
    leghorn hens and Rhode Island red-barred Plymouth Rock buff cross hens
    was rapidly excreted unaltered within the first 8 h.  Up to 91 and
    82%, respectively, were excreted within 13 days (Opdycke, 1976).

    6.5  Retention and turnover

    6.5.1  Biological half-life

         From the studies of Willems et al. (1980) and Ivie (1978), the
    half-life of diflubenzuron appears to be 12 h in rat and sheep and
    18-20 h in the cow.

         Diflubenzuron has been shown to pass intact through the
    intestinal tract and remained active in the manure (Nimmo & de Wilde,
    1977b).

    7.  EFFECTS ON LABORATORY MAMMALS AND IN VITRO TEST SYSTEMS

    7.1  Single exposure

         The acute toxicity of diflubenzuron and its formulations to
    different species is summarized in Table 7.  No signs of intoxication
    were observed during the 14-day period following a single
    administration of diflubenzuron.

         Van Eldik (1974) reported an intraperitoneal LD50 of
    > 2150 mg/kg in rats and mice.

    7.2  Short-term exposure

         Rats (5 of each sex per group) were fed on a diet containing
    diflubenzuron at concentrations of 0, 800, 4000, 20 000 and
    100 000 mg/kg feed for 4 weeks.  Behaviour, body weight, food and
    water consumption were not affected by the treatment.  There was a
    dose-related increase in the met- and sulfhaemoglobin content of the
    blood in all treated groups except for the methaemoglobin value for
    females in the 800-mg/kg dose group.  Lower erythrocyte, packed cell
    volume (PCV) and haemoglobin values were observed in both sexes of the
    100 000-mg/kg dose group.  There was a dose-related increase in spleen
    and liver weights.  Only the dose level of 800 mg/kg did not affect
    the liver weight (Palmer et al., 1977).

         Five male Swiss-albino rats were given 96.7 mg diflubenzuron/kg
    body weight per day, dissolved in corn oil, in their diet for 48 days. 
    The total dose was 4640 mg/kg body weight.  Five controls were given
    corn oil only in their diet.  At the end of the study the treated
    groups showed a significantly lower mean haemoglobin concentration
    than the controls, and decreases in MCH and MCHC values (Berberian &
    Enan, 1989).

         Diflubenzuron was administered to Sprague-Dawley rats of the CD
    strain (20 of each sex per group) at dietary levels of 10 000 and
    100 000 mg/kg feed for 9 weeks, followed by a 4-week withdrawal
    period.  Lower values for red blood cell parameters were recorded at
    both dose levels. An increase in reticulocyte count and a pallor of
    the extremities and eyes were observed.  The formation of
    methaemoglobin occurred in both males and females, with approximately
    5-8% of the available haemoglobin being transformed to methaemoglobin. 
    After a withdrawal period of 4 weeks methaemoglobin comprised less
    than 2% of the available haemoglobin in tested animals compared with
    approximately 0.6% in controls.  Higher values for SGPT were recorded
    as well as heavier liver, spleen and adrenal weights.   Minor
    enlargement of centrilobular hepatocytes in the highest dose group was
    observed, but this finding disappeared after the withdrawal period
    (Hunter et al., 1979).

        Table 7.  Acute toxicity of diflubenzuron and its formulations
                                                                                                                                              

                     Acute oral LD50      Acute dermal LD50     Acute inhalation     Primary eye         Primary skin        Dermal
                                                                LC50                 irritation          irritation          sensitization
                                                                                                                                              

    Diflubenzuron    mouse: > 4640        rat: > 10 000         rabbit: > 30 mg/     rabbit: 40 mg       rabbit: non-        guinea-pig: non-
    technical        mg/kg body weight    mg/kg body weight     litre nominal        instilled in eye;   irritant (Taylor,   sensitizing
                                          (Koopman, 1977c)                           marginal irritant   1973a)              (Prinsen, 1992)
                                                                                     (Davies &
                     rat: > 4640 mg/kg    rabbit: > 4 ml/kg     > 3.75 mg/litre      Ligget, 1973)
                     body weight          body weight of 50%    actual (Berczy et
                     (van Eldik, 1973a;   in gum tragacanth     al., 1975a)
                     Koopman, 1977a)      (Davies &
                                          Halliday, 1974)

    Diflubenzuron    mouse: > 5000        rat: > 2000 mg/kg     rat: > 13.8 mg/      rabbit: 100 mg      rabbit: 500 mg      guinea-pig: 25%
    90% concentrate  mg/kg body weight    body weight           litre nominal        instilled in eye;   non-irritant        w/w in paraffin
                     (Koopman & Pot,      (Koopman, 1984a)                           very slight         (OECD Guideline     (Grade 1); weak
                     1986)                                      > 2.49 mg/litre      irritant (Koopman,  404) (Koopman,      sensitizer (OECD
                                                                actual (Greenough    1984c)              1984d)              Guideline 406)
                     rat: > 5000 mg/kg                          & McDonald,                                                  (Kynoch & Smith,
                     body weight                                1986)                                                        1986)
                     (Koopman, 1984b)

    Dimilin WP 25%   rat: > 40 000        rat: > 20 000         rat: > 150 mg/       rabbit: 100 mg      rabbit: 500 mg      guinea-pig: non-
                     mg/kg body weight    mg/kg body weight     litre nominal        instilled in eye;   non-irritant to     sensitizing
                                          (Janssen & Pot,                            slight to           only minimally      (Kynoch & Elliott,
                     mice: > 40 000       1987e)                > 3.5 mg/litre       moderate,           irritant            1978,a,b)
                     mg/kg body weight                          actual (Arts,        transientsient eye  (Taylor, 1973b;
                     (van Eldik, 1973b;                         1991)                irritation (Snoeij  Chandran, 1981;
                     Koopman, 1977b)                                                 & Busé-Pot, 1991)   Snoeij &
                                                                                                         Busé-Pot, 1990)
                                                                                                                                              

    Table 7.  (Con't)
                                                                                                                                              

                     Acute oral LD50      Acute dermal LD50     Acute inhalation     Primary eye         Primary skin        Dermal
                                                                LC50                 irritation          irritation          sensitization
                                                                                                                                              

    Dimilin SC-48    rat: > 5000 mg/kg    rat: > 2000 mg/kg                          rabbit: 0.1 ml      rabbit: non-        guinea-pig: weak
                     body weight          body weight                 -              instilled in eye;   irritant (Janssen   sensitizer
                     (Janssen & Pot,      (Janssen & Pot,                            slight irritant     & Pot, 1987b)       (Kynoch &
                     1987c)               1987d)                                     (Janssen & Pot,                         Parcell, 1987)
                                                                                     1987a)

    Dimilin SC-15                                                                    albino rats: 0.1    rabbit: minimally
                            -                    -                    -              ml instilled in     irritant                  -
                                                                                     eye; non-irritant   (Prinsen, 1989a)
                                                                                     (Prinsen, 1989b)

    Dimilin 4F       rat: > 5000 mg/kg    rat: > 2000 mg/kg     rat: > 1.9 mg/litre  rabbit: 0.1 ml      rabbit: 0.5 ml      guinea-pig: non-
                     body weight          body weight           actual (Jackson      instilled in eye;   minimally irritant  sensitizing
                     (Spanjers, 1988a)    (Spanjers, 1988b)     et al., 1990)        non-irritant        (Prinsen, 1988a)    (Prinsen, 1989c)
                                                                                     (Prinsen, 1988b)

    Dimilin 2F       rat: > 5000 mg/kg    rat: > 2000 mg/kg     rat: > 4.4 mg/litre  rabbit: 0.1 ml      rabbit: 0.5 ml      guinea-pig:
                     body weight          body weight           actual very          instilled in eye;   moderate irritant   moderate (Grade
                     (Koopman, 1985d)     (Koopman, 1985c)      slightly irritant    moderate irritant   OECD Guideline      III) (Kynoch &
                                                                (Zwart, 1985)        OECD Guideline      404 (Koopman,       Parcell, 1987)
                                                                                     405 (Koopman,       1985a)
                                                                                     1985b)

    Dimilin ODC 45   rat: > 37 300 mg/kg  rat: > 37 300 mg/kg                        rabbit: 0.1 ml      rabbit: 0.5 ml
                     body weight          body weight                                instilled in eye;   moderate irritant
                     (Koopman &           (Koopman, 1980b)                           marginally          (Koopman,
                     Jongeling, 1979)                                                irritant (Koopman,  1980c)
                                                                                     1980a)
                                                                                                                                              

    Table 7.  (Con't)
                                                                                                                                              

                     Acute oral LD50      Acute dermal LD50     Acute inhalation     Primary eye         Primary skin        Dermal
                                                                LC50                 irritation          irritation          sensitization
                                                                                                                                              

    Dimilin OF 6     rat: > 5000 mg/kg    rat: > 2000 mg/kg     rat: > 95.7 mg/      rabbit: 0.1 ml      rabbit: 0.5 ml      guinea-pig: non-
                     body weight          body weight           litre nominal:       instilled in eye;   topical; mild       sensitizing
                     (Besten et al.,      (Besten et al.,       > 2.17 mg/litre      non-irritant        irritant            (Prinsen, 1993)
                     1993a)               1993b)                actual (Janssen &    (Janssen & van      (Janssen & van
                                                                van Doorn, 1993a)    Doorn, 1993c)       Doorn, 1993b)
                                                                                                                                              
             Wistar rats (10 of each sex per group) were fed diflubenzuron in
    the diet for 13 weeks at concentrations of 0, 3.125, 12.5, 50 or
    200 mg/kg feed.  Behaviour, growth and food intake were unaffected by
    the treatment.  At the highest dose level the PCV value, the
    haemoglobin concentration and the number of erythrocytes were
    decreased.  There was an increase in the SGPT and SGOT activities in
    the males of the highest dose group at the end of the experiment.  A
    slight increase in the number of normally occurring scattered small
    foci of necrotic parenchymal cells was observed, accompanied by
    mononuclear inflammatory cell infiltration and proliferation of
    reticuloendothelial system cells in the liver of both males end
    females of the 50 and 200 mg/kg groups (Kemp et al., 1973a,b).

         Absence of toxic effects in chronic/oncogenicity studies at low
    dose levels necessitated re-evaluation of liver histopathology in the
    90-day feeding study; "piece meal" liver cell necrosis was reported at
    the two highest dose levels, i.e. 50 and 200 mg/kg feed.  The original
    slides of the study were re-evaluated by four histopathologists in
    four different laboratories.  They independently and unanimously
    agreed that the lesions in the livers of treated rats were found to
    the same extent in the livers of untreated rats.  This showed that the
    high dose levels in the study did not demonstrate a treatment-related
    necrotic effect in the liver (Offringa, 1977).

         Technical grade diflubenzuron was administered in the diet to
    male and female 21- to 28-day-old Sprague-Dawley rats (40 of each sex
    per group) at dose levels of 0, 160, 400, 2000, 10 000 and
    50 000 mg/kg feed for 13 weeks.  No apparent treatment-related effects
    were noted on mortality, clinical observations, body weight gain, food
    consumption, clinical chemistry or urinalysis.  A treatment-related
    significant increase in methaemoglobin concentration was noted in all
    treated groups. Sulfhaemoglobin values showed increases at dose levels
    of 2000 mg/kg or more.  A significant treatment-related decrease in
    haemoglobin, PCV and erythrocyte count was observed in males and
    females at all dose levels by the end of the study.  An increase was
    noted in the reticulocyte count at all dose levels except 160 mg/kg,
    and the number of the Heinz bodies was higher in the 10 000 and
    50 000 mg/kg groups. After 7 weeks, spleen weights were increased in
    the females at all dose levels, but after 13 weeks no effect was found
    at 160 mg/kg.  With the exception of the lowest dose level, all
    treated groups showed a higher liver weight.  The administration of
    diflubenzuron resulted in a dose-related increase in the incidence of
    chronic hepatitis and haemosiderosis of the liver. It was also
    associated at all dose levels with haemosiderosis and congestion of
    the spleen and mild erythroid hyperplasia of the bone marrow.  The
    severity of the lesions tended to increase with the dose.  Liver
    lesions were more severe in males than in females and were more severe
    at 13 weeks than at 7 weeks.  A no-observed-effect level (NOEL) was
    not established (Burdock et al., 1980b; Goodman, 1980b).

         Diflubenzuron was given to CFLP mice for 6 weeks at levels of 16
    and 50 mg/kg feed.  There were no clinical signs and no effects on
    food consumption, body weight, blood chemistry or macroscopic
    pathology.  In three of the eight animals given 50 mg/kg, foci of
    liver cell necrosis, with or without inflammatory cell filtration,
    were noted.  Other organs were not examined microscopically (Hunter et
    al., 1974).

         Diflubenzuron was administered to Swiss Webster mice in a 30-day
    oral intubation study.  Groups of five mice each received either no
    treatment, vehicle control (Polyethylene glycol 400) or diflubenzuron
    suspensions at dose levels of 125, 500 or 2000 mg/kg body weight. 
    Hepatic glutathione- S-transferase activity and morphological
    characteristics were studied.  Diflubenzuron was shown to elicit
    hepatocellular changes at all dose levels.  The activities of three
    glutathione- S-transferases ( S-aryl,  S-aralkyl and  S-epoxide)
    were irregularly altered in a non-dose-related manner.  Light
    microscopy revealed radial arrays of hepatocellular vacuolization
    between the portal and central vein areas.  There was evidence of an
    increase in the amount of endoplasmic reticulum (Young et al., 1986).

         Male and female mice of the B6C3F1 strain (40 of each sex per
    group) received diflubenzuron (97.2% a.i.) in the diet at dose levels
    of 0, 16, 50, 400, 2000, 10 000 or 50 000 mg/kg feed for 13 weeks.  An
    additional group of 100 of each sex served as a control.   No
    compound-related effects were apparent with respect to clinical signs,
    survival, growth rates, total food consumption or gross pathology. 
    Significant treatment-related increases in met- and sulfhaemoglobin
    concentrations were noted in all treated groups, except in the group
    fed 16 mg/kg.  At the higher dose levels, there was a decrease in
    haematocrit and erythrocyte counts and an increase in reticulocyte,
    platelet and Heinz body counts.  Significantly higher alkaline
    phosphatase activity was noted in the 10 000 and 50 000 mg/kg groups. 
    Compound-related effects on the weights of liver and spleen were
    noted.  In the females, adrenal weight was decreased (but not in a
    dose-related fashion) at all dose levels after 7 weeks and increased
    after 13 weeks in higher dose levels.  Higher adrenal weight was
    observed in treated males than in the controls.  Histopathological
    examination revealed treatment-related centrilobular hypertrophy of
    hepatocytes, with or without cell necrosis, haemosiderosis of the
    liver and spleen, extramedullary haematopoiesis and mild chronic
    hepatitis in treated animals of both sexes, some of which effects were
    observed at the lowest dose level (16 mg/kg).  The liver lesions were
    more severe in males than in females, being most severe in the high-
    dose males.  The NOEL for methaemoglobin formation was 16 mg/kg feed
    (Burdock et al., 1980a; Goodman, 1980a).

         HC/CFLP mice (40 of each sex per group) were fed diflubenzuron
    (97.2% purity) for 14 weeks at levels of 0, 80, 400, 2000, 10 000 and
    50 000 mg/kg feed.  On the second day of treatment, the majority of

    mice treated with 10 000 or 50 000 mg/kg showed dark eyes and/or
    prominent caudal blood vessels.  On day 5, blue/grey discoloration of
    the extremities was noted for the majority of mice treated with
    50 000 mg/kg.  Mice in the lowest dose group exhibited no clinical
    signs.  Mortality, food consumption, water consumption and body weight
    changes were not significantly affected by the treatment.  Lower PCV
    and red blood cell counts were found at all dose levels except
    80 mg/kg.  The total white blood cell count, lymphocyte count,
    haemoglobin concentration, incidence of Heinz bodies and red blood
    cell count were increased in all treated groups.  At week 7, there was
    an increase in the number of reticulocytes in treated mice,
    particularly in males treated at 10 000 or 50 000 mg/kg.  At week 14,
    the reticulocyte counts were similar to those of the controls.  A
    treatment-related increase in both met- and sulfhaemoglobin was
    recorded in all treated groups at weeks 7 and 14 of the investigation. 
    Plasma glutamic-pyruvic transaminase values were increased at all dose
    levels, with the exception of 80 mg/kg feed.  Lower blood cholesterol
    levels were noted in the 2000, 10 000 and 50 000 mg/kg groups. 
    Macroscopic examination showed dark discoloration and/or enlargement
    of the spleen and pale subcapsular areas of the liver in all dose
    groups after both 7 and 14 weeks.  Histopathological examination of
    the spleen revealed increased haemosiderosis at all dose levels except
    80 mg/kg.  In the liver, areas of focal necrosis and/or fibrosis in
    the parenchyma, with or without associated inflammatory cells,
    fibroblasts or pigment-laden macrophages, were observed.  At higher
    dose levels necrotic and fatty hepatocytes and brown pigment-laden
    Kupffer cells were found.  A NOEL was not established (Colley et al.,
    1981a,b).

         Diflubenzuron was fed to groups of three male and three female
    beagle dogs for 13 weeks at concentrations of 0, 10, 20, 40 and
    160 mg/kg diet.  No effect of the treatment on behaviour, body weight
    or food and water consumption was observed.  Elevated SAP and SGPT
    values were recorded for some dogs receiving 40 or 160 mg
    diflubenzuron/kg feed.  After 6 weeks, methaemoglobin and another
    abnormal pigment, probably sulfhaemoglobin, were demonstrated in dogs
    given 160 mg/kg.  After 12 weeks of administration, some recovery was
    observed.  Organ weights and gross and microscopic evaluation did not
    reveal any treatment-related effects.  The NOEL for methaemoglobin
    formation was 40 mg/kg feed (Chesterman et al., 1974).

         Diflubenzuron was administered daily in gelatin capsules to male
    and female beagle dogs (6 of each sex per group) at dose levels of 2,
    10, 50 or 250 mg/kg body weight per day for 52 weeks. There were no
    treatment-related effects on mortality, food consumption or body
    weight gain.  Dose-related marginal increases in methaemoglobin and
    sulfhaemoglobin were recorded from 10 mg/kg upwards.  At 50 and
    250 mg/kg, haemoglobin concentration and MCHC were decreased whereas
    reticulocyte and platelet counts were increased.  Heinz bodies were

    also detected in several animals receiving 50 and 250 mg/kg.  Dose-
    related increases in liver and spleen weights were found in the 50 and
    250 mg/kg males.  Histopathological evaluation of the liver revealed
    an increase in the incidence of pigmented macrophages and Kupffer cell
    siderosis at 50 and 250 mg/kg in both males and females.  The NOEL
    based on the increase in met- and sulfhaemoglobin was 2 mg/kg body
    weight (Greenough et al., 1985).

         In a study by Berczy et al. (1975c), rats were exposed daily for
    a one-hour period to technical diflubenzuron dust at nominal
    concentrations of 0, 0.5, 5.0 and 50 mg/litre air, respectively (the
    actual concentrations were 0, 0.12, 0.87 and 1.85 mg/litre,
    respectively).  Exposures were repeated over a period of 3 weeks,
    5 days per week.  The methaemoglobin levels in male rats at the two
    lower concentrations and female rats of all test groups were
    significantly higher than those of controls.

         Rabbits were exposed daily for one-hour periods to technical
    diflubenzuron dust at concentrations of 0.5, 5.0 and 25 mg/litre air
    (the measured concentrations were 0.15, 0.75 and 1.99 mg/litre,
    respectively).  Exposures were repeated over a period of 3 weeks,
    5 days each week.  There were no signs of irritation in animals
    exposed to 0.5 mg/litre, but mild transient respiratory irritation was
    seen at the two higher concentrations.  Haematological examination,
    biochemistry tests and macroscopic inspection revealed no treatment-
    related abnormalities (Berczy et al.,  1975b).

         Diflubenzuron, at levels of 4.64, 10 and 21.5% weight/volume was
    applied daily to the intact and abraded skin of rabbits at a dosage
    level of 1.5 ml/kg body weight, 5 days a week, for 3 consecutive weeks
    (equivalent to 69.6, 150 and 322.5 mg diflubenzuron/kg body weight per
    day).  The sulfhaemoglobin level was increased in one rabbit out of 20
    in the 10% group, and in 5 rabbits out of 20 at the high treatment
    level.  The 10% treatment level was considered to be the NOEL based on
    sulfhaemoglobin formation (Davies et al., 1975).

    7.3  Long-term exposure

         Diflubenzuron was administered to Sprague-Dawley rats (60 of each
    sex per group) in their diet at levels 0, 10, 20, 40 and 160 mg/kg
    feed for 104 weeks.  There were no treatment-related effects on body
    weight gain, food intake, renal function or on macroscopic and
    microscopic pathology.  In rats treated with 160 mg/kg, significantly
    higher methaemoglobin levels were recorded.  The tumour profile of
    treated rats was similar to that of the controls.  The NOEL based on
    methaemoglobin was 40 mg/kg, equivalent to a mean intake of 1.43 and
    1.73 mg/kg per day for males and females, respectively (Hunter et
    al.,1976).

         When diflubenzuron was administered in the diet to male and
    female Sprague-Dawley rats (50 of each sex per group) at dosage levels
    of 0, 156, 625, 2500 and 10 000 mg/kg feed for 104 weeks, there was no
    evidence of an effect on mortality or treatment-related clinical
    signs.  Significantly increased absolute methaemoglobin and
    sulfhaemoglobin values were observed in all male treatment groups. 
    However, increases in relative methaemoglobin  values (% of total
    haemoglobin) were only noted in the 156, 2500 and 10 000 mg/kg groups,
    while sulfhaemoglobin increases were noted in the 156, 625 and
    10 000 mg/kg females.  There was a significant increase in absolute
    and relative spleen weights in the two highest dose groups of both
    sexes, together with haemosiderosis in spleen and liver.  There was no
    evidence of carcinogenicity after 2 years of feeding diflubenzuron
    (Burdock et al., 1984).

         In a study by Hunter et al. (1975), diflubenzuron was
    administered to CFLP mice for 80 weeks at dietary levels of 0, 4, 8,
    16 and 50 mg/kg feed.  There were no overt signs of reaction to
    treatment.  Behaviour, mortality, food and water consumption and body
    weight were unaffected by the treatment.  The macroscopic changes
    observed were those commonly seen in mice of this strain and age.  No
    histopathological changes were seen that were considered to be related
    to the administration of diflubenzuron.  The incidence of liver cell
    tumours was higher in this study than it was in other studies
    performed in these laboratories using this strain of mouse.  The
    increase was seen in both treated and control groups of either sex and
    showed no evidence of a treatment-related effect.  There was no
    evidence of a treatment-related effect on tumour incidence in the CFLP
    mouse.

         Male and female HC/CFLP mice (88 of each sex per group) were fed
    diflubenzuron at dietary levels of 0, 16, 80, 400, 2000 and
    10 000 mg/kg feed for 91 weeks.  There was no indication of a
    treatment-related effect on survival, food consumption or body weight
    gain.  Treatment-related elevations of MCH and MCHC values were
    recorded from week 26 onwards in mice given 10 000 mg/kg.  The
    incidence of Heinz bodies increased in a dose-related manner among
    mice given 400, 2000 or 10 000 mg/kg from week 52 onwards.  Dose-
    related increases in methaemoglobin levels were found from week 26
    onwards and in sulfhaemoglobin levels from week 52 onwards in the mice
    fed 80 mg/kg or more.  Elevated alkaline phosphatase (AP) activities
    were seen at weeks 24, 76 and 89 among male mice receiving 2000 and
    10 000 mg/kg and at week 84 among mice of the 400 mg/kg dose group. 
    An increased incidence of splenic and/or hepatic enlargement was seen
    among mice treated with 10 000 mg/kg.  Cyanosis of the skin was noted
    among mice at 400, 2000 and 10 000 mg/kg.  Increased liver and spleen
    weights were found among mice given 2000 and 10 000 mg/kg.  An
    increased incidence of hepatocyte enlargement and increased
    extramedullary haematopoiesis in the liver and spleen were seen in
    mice at high dose levels.  In the 400, 2000 and 10 000 mg/kg groups,

    there was an increased incidence of siderocytosis in the spleen and of
    pigmented Kupffer cells in the liver.  There was no treatment-related
    effect on tumour incidence (Colley et al., 1984).

    7.4  Skin and eye irritation; sensitization

         Relevant data are given in Table 7.

         Administration of technical diflubenzuron to the intact and
    abraded skin of albino rabbits did not produce irritation of the skin
    after exposure times of 24 and 72 h (Taylor, 1973a,b).

         Diflubenzuron was found to be a moderate irritant to the rabbit
    skin after application of 0.5 ml of 45% oil dispersible concentrate
    for 24 h (Koopman, 1980a; Prinsen, 1990).

         Diflubenzuron (both technical and 45% oil dispersible
    concentrate) was considered to be marginally irritant to the rabbit
    eye (Davies & Ligget, 1973; Koopman, 1980b).

         Diflubenzuron (48% water-based paste) was not found to be a
    dermal sensitizer in guinea-pigs (Kynoch & Parcell, 1987).

         Technical diflubenzuron was studied for skin sensitization in a
    maximization test on guinea-pigs and was found to be non-sensitizing
    (Prinsen, 1992).  However, some formulations are mild sensitizers
    (Table 7).

    7.5  Reproductive toxicity, embryotoxicity and teratogenicity

         Diflubenzuron was fed to pathogen-free rats of the CFY strain
    (20 of each sex per group) at dietary levels of 0, 1000 and
    100 000 mg/kg for one generation and one litter.  The animals were
    maintained on their respective diets for 9 weeks prior to mating. 
    There were no clear effects on mating performance, pregnancy rate,
    duration of gestation, litter size, offspring mortality, litter weight
    or the type and distribution of abnormalities.  Dose-related effects
    of diflubenzuron were demonstrated at 17 weeks in adults and consisted
    of reduced values for PCV, haemoglobin, total red cells count and
    MCHC, increased values for methaemoglobin, MCV, spleen weight and
    siderocyte incidence in the spleen, and the occurrence of iron-
    pigment-containing Kupffer cells in the liver.  A dose-related effect
    on the liver was also shown by increased weight and SGPT activity and
    centrilobular hepatocyte enlargement.  Reduced blood glucose
    concentrations were recorded in both treated groups.  At the highest
    dosed level, the offspring showed increased liver and spleen weights
    for both sexes (Palmer et al., 1978).

         In a three-generation reproductive study with rats fed
    diflubenzuron at concentrations of 10, 20, 40 and 160 mg/kg feed, no
    adverse treatment-related effects on mating performance, pregnancy
    rate, duration of gestation or litter parameters (total loss, size,
    mean pup weight, mortality, abnormalities) were found (Palmer & Hill,
    1975a).

         After gavage administration of diflubenzuron at doses of 1, 2 and
    4 mg/kg body weight per day to pregnant rats during days 6-15 of
    gestation, no effects were observed on embryonic or fetal development
    (Palmer & Hill, 1975b).

         Treatment of pregnant New Zealand white rabbits at oral doses of
    1, 2 and 4 mg/kg body weight per day during days 6-18 of gestation did
    not affect embryonic or fetal development as assessed by the incidence
    of major malformations, minor anomalies and skeletal variants (Palmer
    & Hill, 1975c).

         In a study by Booth (1977), pregnant Swiss mice were fed a diet
    containing 50 mg/kg of diflubenzuron (partly 14C) for a period of
    17 days.  Some of these mice were killed at day 17 after conception,
    while the others were allowed to give birth.  The results of this
    study showed that mucopolysaccharide synthesis in embryonic mouse-limb
    cartilage was normal.  Diflubenzuron did not pass through embryonic
    membranes nor was it passed from mother to suckling young mice. 
    Analysis of 226 embryos showed no gross teratogenic effects.

         Two groups of 24 timed-mated female rats of the Crl:CD (SD) BR
    strain were dosed once daily by the oral route between days 6 and 15
    of pregnancy.  One group was dosed with diflubenzuron at 1000 mg/kg
    per day, while the other group was given the vehicle (1.0% gum
    tragacanth) only.  Clinical signs, body weights and food consumption
    were recorded. The females were killed on day 20 of pregnancy and a
    necropsy was performed.  The fetuses were subjected to detailed
    external, visceral and skeletal examinations.  There were no maternal
    deaths or treatment-related changes in clinical condition.  Treatment
    did not affect maternal growth or food consumption.  There were no
    maternal abnormalities at necropsy that were considered to be related
    to treatment.  The mean numbers of corpora lutea, implantations and
    live fetuses were similar in all groups.  Both pre- and post-
    implantation losses were unaffected by treatment.  Fetal weights and
    sex ratio were unaffected by diflubenzuron treatment.  The incidences
    of major external/visceral and skeletal abnormalities, and minor
    external/visceral abnormalities were not affected by diflubenzuron
    treatment.  The incidence of fetuses with minor skeletal abnormalities
    was slightly higher in the treated group, but was within the usual
    background range.  There were intergroup differences in the
    proportions of fetuses with specific minor abnormalities and variants
    of skeletal ossification. For some bones, the differences achieved

    statistical significance.  However, on balance, treated group fetuses
    were considered to be similarly ossified to the control fetuses.  Oral
    administration of diflubenzuron at a dose level of 1000 mg/kg per day
    did not elicit maternal toxicity or any evidence of embryotoxicity
    (Kavanagh, 1988a).

         Two groups of 16 timed-mated female New Zealand White rabbits
    were dosed once daily by the oral route from days 7 to 19 of
    pregnancy, inclusive. One group was dosed with diflubenzuron at
    1000 mg/kg per day and the other group with the vehicle (1.0% gum
    tragacanth) only. Clinical signs, body weights and food consumption
    were recorded. The females were killed on day 28 of pregnancy and a
    necropsy was performed. The fetuses were subjected to detailed
    external, visceral and skeletal examinations. There were no maternal
    deaths, changes in clinical condition or abnormalities at maternal
    necropsy considered to be related to diflubenzuron treatment. Two
    animals were killed prematurely during the study, one from the control
    group and one from the treated group. The changes in clinical
    condition and abnormalities observed at necropsy in these animals were
    considered to be unrelated to treatment. Treatment did not affect
    maternal growth or food consumption. The mean numbers of corpora
    lutea, implantations and live fetuses were similar in all groups. Both
    pre- and post-implantation losses were unaffected by treatment. Fetal
    weights and sex ratio were unaffected by diflubenzuron treatment. The
    incidence of both major and minor external/visceral and skeletal
    abnormalities, as well as the numbers of skeletal variants, was
    unaffected by diflubenzuron treatment. Oral administration of
    diflubenzuron at a dose level of 1000 mg/kg per day did not elicit
    maternal toxicity or any evidence of embryotoxicity (Kavanagh, 1988b).

         In a study by Kubena (1982), diflubenzuron was fed at levels of
    0, 2.5, 25 and 250 mg/kg feed to male and female layer-breed chickens
    from 1 day of age through a laying cycle. Characteristics measured
    were egg production, egg weight, eggshell weight, fertility,
    hatchability and effects on the progeny. Feeding diflubenzuron at
    levels up to 250 mg/kg feed did not affect these characteristics.

         Groups of chicken eggs were injected near the embryonic coelom
    with a suspension of 10 mg diflubenzuron in 0.1 ml of peanut oil. 
    Diflubenzuron did not cause significant malformations in the embryos
    (Seegmiller & Booth, 1976).

    7.6  Mutagenicity and related end-points

         Diflubenzuron was examined by  in vitro and  in vivo 
    mutagenicity tests.  The results are summarized in Table 8. 
    Mutagenicity tests have also been carried out with the major
    metabolites of diflubenzuron (Table 9).

        Table 8.  Mutagenicity tests with diflubenzuron
                                                                                                                                              

    End-point             Organisms,                      Dose level,                   Metabolic activationa      Results     References
                          cells, strains                  concentration                 (presence/absence = +/-)
                                                                                                                                              

                          Microorganisms

    Reverse               Salmonella typhimurium
    mutation              TA98, TA100,                    10-1000 µg/plate                       +/-               negative    Bryant (1976)
                          TA98, TA100, TA1537, TA1978     1000 µg/spot                           +/-               negative    Bryant (1976)

    Reverse               S. typhimurium                  0.1-500 µg/plate                       +/-               negative    Brusick & Weir
    mutation              TA98, TA100, TA1535,                                                                                 (1977a)
                          TA1537, TA1538

    Reverse               S. typhimurium                  10, 100 or 1000 µg/plate;              +/-               negative    McGregor et al.
    mutation              TA98, TA100,                    19, 186, 1860 µg DFB/plate                                           (1979)
                          TA1535, TA1537                  (as Dimilin W-25)

    Reverse               S. typhimurium                  up to 5000 µg/plate                    +/-b              negative    Moriya et al.
    mutation              TA98, TA100, TA1535,                                                                                 (1983)   
                          TA1537, TA1538
                          Escherichia coli. WP2 hcr

    Reverse               S. typhimurium                  up to 1000 µg/plate                    +/-               negative    Koorn (1990)
    mutation              TA98, TA100, TA1535,
                          TA1537, TA1538
                                                                                                                                              

    Table 8.  (Con't)
                                                                                                                                              

    End-point             Organisms,                      Dose level,                   Metabolic activationa      Results     References
                          cells, strains                  concentration                 (presence/absence = +/-)
                                                                                                                                              

                          Mammalian cells in vitro

    Forward               L5178Y mouse lymphoma           1.17-300 µg/ml                         +/-               negative    McGregor et al.
    mutation              cells                                                                                                (1979) 

    Chromosomal           Chinese hamster ovary           up to 200 µg/ml                        +/-               negative    Taalman & Hoorn
    aberrations           cells                                                                                                (1986)

    Unscheduled           human diploid WI-38 cells       50-1000 µg/ml                          +/-               negative    Brusick & Weir
    DNA synthesis         (blocked in the G1 phase)                                                                            (1977c)

    DNA repair            rat hepatocytes                 up to 333 µg/ml                                          negative    Enninga (1990)
    (unscheduled
    DNA synthesis)

    Cell transformation   BALB-3T3 cells,                 0.02-0.312 µg/ml                       -                 negative    Brusick & Weir
                          in vitro                                                                                             (1977b)

    Cell transformation   pregnant hamster, ip            10, 200 and 500 mg/kg                                    negative    Quarles et al.
    (transplacental       injection on 10th day of        body weight                                                          (1980)
    transformation)       gestation, fetal cell culture
                          3 days after injection
                                                                                                                                              

    Table 8.  (Con't)
                                                                                                                                              

    End-point             Organisms,                      Dose level,                   Metabolic activationa      Results     References
                          cells, strains                  concentration                 (presence/absence = +/-)
                                                                                                                                              

                          Mammals

    Micronucleus          mouse bone marrow               15, 150, 1500 mg/kg body                                 negative    McGregor et al.
                                                          weight 30 and 6 h before                                             (1979)
                                                          necropsy

    Dominant              male mice mated to              1000 and 2000 mg/kg body                                 negative    Arnold et al.
    lethal                3 females weekly for 6 weeks    weight intraperitoneal                                               (1974)
                                                                                                                                              

    a  Unless indicated otherwise, S9 was obtained from livers of rats treated with Arochlor-1254
    b  Source of S9 not indicated

    Table 9.  Mutagenicity tests with diflubenzuron metabolites
                                                                                                                                              

    End-point           Test system                   Dose level            Metabolic activationa        Results      References
                                                                            (presence/absence = +/-)
                                                                                                                                              

    A.  4-chlorophenylurea

                        Microorganisms

    Reverse mutation    Salmonella typhimurium
    and differential    TA98, TA100, TA1535,          1000 µg/spot                  +/-                 negativeb     Dorough (1977)
    killing             TA1537, TA1538, TA1978

    Reverse mutation    TA98, TA100                   10, 100, 500,                                     negative
                                                      1000 µg/plate

    Reverse mutation    S. typhimurium                0.1-500 µg/plate              +/-                 negative      Jagannath & Brusick
                        TA98, TA100, TA1535,                                                                          (1977a)
                        TA1537, TA1538

                        Mammalian cells

    Unscheduled         human WI-38 cells             6.25-50 µg/ml                 +/-                 negative      Matheson & Brusick
    DNA synthesis       (blocked in the G1 phase)                                                                     (1978b)

    Cell                BALB-3T3 cells,               0.019-0.312 mg/ml             not stated          weak          Matheson & Brusick
    transformation      in vitro                                                                        positive at   (1977a)
                                                                                                        0.312 mg/ml
                                                                                                                                              

    Table 9. (Con't)
                                                                                                                                              

    End-point           Test system                   Dose level            Metabolic activationa        Results      References
                                                                            (presence/absence = +/-)
                                                                                                                                              

    B.  2,6-Difluorobenzoic acid

                        Microorganisms

    Reverse mutation    S. typhimurium                1000 µg/spot                  +/-                 negative      Dorough (1977)
    and differential    TA98, TA100, TA1535,
    killing             TA1537, TA1538, TA1978

    Reverse mutation    S. typhimurium                10, 100, 500,
                        TA98, TA100                   1000 µg/plate

    Reverse mutation    S. typhimurium                0.1-500 µg/plate              +/-                 negative      Jagannath & Brusick
                        TA98, TA100, TA1535,                                                                          (1977b)
                        TA1537, TA1538

                        Mammalian cells

    Unscheduled         human WI-38 cells             75-500 µg/ml                  +/-                 positive      Matheson & Brusick
    DNA synthesis       (blocked in the G1 phase)                                                       with          (1978a)
                                                                                                        activation

    Cell                BALB-3T3 cells,               0.156-2.5 mg/ml               not stated          weak          Matheson & Brusick
    transformation      in vitro                                                                        positive at   (1977b)
                                                                                                        2.5 mg/ml

    C.  4-Chloroaniline (PCA)

                        Microorganisms

    Reverse mutation    S. typhimurium                1000 µg/spot                  +/-                 negativeb     Dorough (1977)
    and differential    TA98, TA100, TA1535,
    killing             TA1537, TA1538, TA1978
                                                                                                                                              

    Table 9. (Con't)
                                                                                                                                              

    End-point           Test system                   Dose level            Metabolic activationa        Results      References
                                                                            (presence/absence = +/-)
                                                                                                                                              

    Reverse mutation    TA98, TA100                   10, 100, 500,                                     positive in
                                                      1000 µg/plate                                     TA98 at 500
                                                                                                        and 1000 µg
                                                                                                        with 
                                                                                                        activation

    Reverse mutation    S. typhimurium                0.1-500 µg/plate              +/-                 negative      Jagannath & Brusick
                        TA98, TA100, TA1535,                                                                          (1977c)
                        TA1537, TA1538

    Reverse mutation    S. typhimurium
                        TA98, TA100, TA1530,          0-1500 µg/plate               +/-                 negative      Gilbert et al. (1980)
                        TA1535, TA1537, TA1538

    Reverse             S. typhimurium C3076,         1000 µg/plate                 +/-                 negative      Thompson et al.
    mutation            D3052, G46, TA98, TA100,                                                                      (1983)
                        TA1535, TA1537, TA1538,
                        Escherichia coli WP2,
                        WP2 uvrA

    Reverse             S. typhimurium                3333 µg/plate                 +/-                 positivec     Dunkel et al. (1985)
    mutation            TA98, TA100, TA1535,
                        TA1537, TA1538,
                        Escherichia coli

    Reverse             S. typhimurium                1666 µg/plate                 +/-                 positived     Mortelmans et al.
    mutation            TA97, TA98, TA100, TA1535                                                       negative      (1986)

    DNA damage          Escherichia coli polA+/polA-  250 µg/plate                  -                   positive      Rosenkranz & Poirier
                                                                                                                      (1979)
                                                                                                                                              

    Table 9. (Con't)
                                                                                                                                              

    End-point           Test system                   Dose level            Metabolic activationa        Results      References
                                                                            (presence/absence = +/-)
                                                                                                                                              

    Mutation            Aspergillus nidulans          200 µg/ml                     -                   positive      Prasad (1970)

                        Mammalian cells

    Forward mutation    L5178Y mouse lymphoma                                       +/-                 positived     Caspary et al. (1988)
                        tk+/- cells

    Unscheduled         human WI-38 cells             250-1000 µg/ml                +/-                 negative      Matheson & Brusick
    DNA synthesis       (blocked in the G1 phase)                                                                     (1978b)

    Unscheduled         rat primary hepatocytes       5-50 µg/ml                    -                   positive      Williams et al.
    DNA synthesis                                                                                                     (1982)

    Unscheduled         rat primary hepatocytes       50 nmol/ml                    -                   negative      Thompson et al.
    DNA synthesis                                                                                                     (1983)

    Sister chromatid    Chinese hamster ovary         1600 µg/ml                    +/-                 positived     US NTP (1989)
    exchange            cells

    Chromosomal         Chinese hamster ovary cells   1000 µg/ml                    +/-                 positived     US NTP (1989)
    aberrations                                                                                         and negative

    Cell                BALB-3T3 cells,               0.039-0.625 mg/ml             not stated          negative      Matheson & Brusick
    transformation      in vitro                                                                                      (1978c)
                                                                                                                                              

    a  Unless indicated otherwise, S9 was obtained from livers of rats treated with Arochlor-1254
    b  No increase in revertants, strains TA1538/TA1978 positive for differential killing
    c  Tested in three independent laboratories
    d  Tested in two independent laboratories
             According to the findings presented, neither diflubenzuron nor
    its major metabolites may be considered to have mutagenic effect.
    Several positive effects were, however, obtained with PCA.

    7.7  Carcinogenicity

         Long-term carcinogenicity studies were described in section 7.3. 
    In both mouse and rat oncogenicity studies, diflubenzuron at dose
    levels up to 10 000 mg/kg feed caused no change in tumour profile or
    onset of tumours.  In the rat oncogenicity study, the incidence of
    sarcoma in the spleen and phaeochromocytomas was not increased.  In
    the mouse oncogenicity study, the incidence of hepatocellular
    neoplasms or haemangiosarcomas in spleen and liver was not increased. 
    Therefore, diflubenzuron, in combination with its metabolites as
    generated in the animal metabolic system, is not oncogenic. 
    Significantly, there were no non-neoplastic or neoplastic lesions of
    the vasculature, including that of liver and spleen, in B6C3F1 male
    mice treated with diflubenzuron. Similarly, there were no fibrotic or
    carcinomatous lesions in the spleen of treated male F-344/N rats.

    7.8  Other special studies

         Diflubenzuron has been studied in mice for its growth-inhibiting
    activity in serially transplanted B16 malignant melanoma and CA1025
    skin carcinoma.  A single 800 mg/kg intraperitoneal injection of
    diflubenzuron induced rapid (24 h) decreases in tumour volume in 78%
    and 66% of the tumours, respectively, while in control mice 85% of the
    melanomas and 91% of the skin carcinomas increased in volume over the
    same time period (Jenkins et al., 1984).  These observations were
    later confirmed, and it was suggested that the activity was due to
    derivatives of a hydroxylated metabolite (Jenkins et al., 1986). 
    Studies of nucleoside uptake by Harding Passey melanoma cells
     in vitro indicated a rapid (< 5 min) inhibition of the uptake of
    uridine, adenosine and cytidine, but not of thymidine, by
    diflubenzuron; this could not be reversed by washing.   De novo 
    nucleic acid synthesis was not impaired and  in vitro cell growth was
    unaffected (Mayer et al., 1984).

         El-Sebae et al. (1988) tested the effect of diflubenzuron on
    protein and RNA biosynthesis by rabbit liver and muscle tissues kept
    in an incubation medium.  The synthesis of protein and RNA was
    significantly stimulated in the liver and inhibited in the muscle by
    graded doses.  The maximum effect on both tissues was reached at 5 µg
    diflubenzuron/ml for protein synthesis and at 0.2 µg/ml for RNA
    synthesis, the effect on protein synthesis being more pronounced than
    that on RNA synthesis in both tissues.

    7.8.1  Special studies on met- and sulfhaemoglobin formation

         The ability of diflubenzuron to induce methaemoglobin and
    sulfhaemoglobin formation has been recognized since the initial
    toxicity studies on the compound.  Methaemoglobinaemia has been
    demonstrated after oral, dermal and inhalatory exposure to
    diflubenzuron in various species (see section 7.2 and 7.3).  It is the
    most sensitive parameter in this case.

         Fifteen male Wistar rats received diflubenzuron (technical) by
    gastric intubation at a dose level of 5000 mg/kg body weight per day
    for 8 days.  No effect was observed on Heinz body formation, whereas
    met- and sulfhaemoglobin levels were significantly increased when
    compared with the control group.  The increase in the methaemoglobin
    level was about 6% (Keet, 1977a).

         When diflubenzuron was administered to male Wistar rats for 28
    days at oral doses of 100 and 500 mg/kg body weight, it induced
    elevation of methaemoglobin concentration and reticulocytes count in
    both of the treated groups.  However, there was no dose-response
    relationship at the dose levels investigated  (Tasheva & Hristeva,
    1991, 1993).

         Technical diflubenzuron was administered by gastric intubation to
    male Swiss mice daily for a period of 14 days at dose levels of 0, 8,
    40, 200, 1000 and 5000 mg/kg body weight.  Body weight measurement and
    macroscopic evaluation did not reveal any effect of the treatment.  At
    dose levels of 1000 and 5000 mg/kg the percentages of methaemoglobin
    and erythrocytes containing Heinz bodies were increased.  The
    sulfhaemoglobin level was statistically significantly increased at
    200, 1000 and 5000 mg/kg in comparison to the control group.  The NOEL
    was considered to be 40 mg/kg body weight based on sulfhaemoglobin
    (Keet, 1977b).

         When female mice were fed 0, 50, 200, 400, 1000 and 2000 mg
    diflubenzuron/kg feed for 30 days, sulfhaemoglobin was demonstrated in
    the blood from 200 mg/kg onwards (being 13% of total haemoglobin at
    2000 mg/kg).  Mice fed 1000 and 2000 mg/kg showed signs of cyanosis
    after 3 weeks.  Recovery was completed after a 3-week withdrawal
    period (Bentley et al., 1979).

         When 15 male New Zealand White rabbits were fed technical or
    analytically pure  diflubenzuron (640 mg/kg feed) for 21 or 18 days,
    respectively, the methaemoglobin and sulfhaemoglobin levels were
    significantly increased (Keet, 1977c).

         Male and female cats (24 of each sex per group) received
    diflubenzuron orally for 21 days at dose levels of 0, 30, 70, 100, 300
    and 1000 mg/kg body weight and were observed for the subsequent 14-day
    period.  A dose-related elevation of methaemoglobin level was observed

    with ceiling values at 300 and 1000 mg/kg.  For male cats the NOEL was
    30 mg/kg, but no NOEL was achieved for females.  Increased
    sulfhaemoglobin and Heinz bodies were observed in all treated groups. 
    NOEL values for sulfhaemoglobin were not achieved.  The haemoglobin
    concentration, reticulocyte number and organ weights were not affected
    by the treatment (Schwartz & Borzelleca, 1981).

         After dermal application of technical diflubenzuron at a dose
    level of 1.5 ml/kg body weight for 18 days to rabbits, the
    methaemoglobin level was increased (Keet, 1977c).

         The available data demonstrate that dose-response relationship
    for production of methaemoglobin exists.  This is considered to be the
    most sensitive end-point after repeated exposure in experimental
    animals.

         Table 10 summarizes the effects on methaemoglobinaemia as
    determined in various studies.

    7.9  Toxicity of metabolites

         In rat metabolic studies it was shown that about 20% of absorbed
    diflubenzuron is metabolized to 2,6-DFBA and its counterpart 4-CPU. 
    Only a small fraction of the 4-CPU is metabolized to PCA (see
    section 6).

         The acute oral toxicity of the major metabolites of diflubenzuron
    is summarized in Table 11.

         Loss of activity, catatony, paralysis and severe bradypnoea were
    observed in rats treated with the metabolite 4-CPU.  The minimum
    symptomatic dose level was 100 mg/kg body weight.  At autopsy, the
    animals showed congested blood vessels and haemorrhage in the
    gastrointestinal tract (Koelman-Klaus, 1978a).

         Rats dosed with 2,6-DFBA showed slight increase in startle
    response, activity, abdominal and limb tone, slight decrease in
    grooming activity, slightly abnormal gait and body posture, mild
    restlessness, irritation and aggressivity, pilo-erection and increased
    alertness. The minimum symptomatic dose level was 464 mg/kg body
    weight (Koelman-Klaus, 1978b).

         Mutagenicity tests have been carried out with 2,6-DFBA, 4-CPU and
    PCA (see section 7.6 and Table 9).

        Table 10.  Summary of the effects on methaemoglobinaemia in various species
                                                                                                                                     

    Species   Route               Duration       No-observed-effect level                               Reference
                                                                                                                                     

    Rat       diet                4 weeks        male: not achieved; female: 800 mg/kg feed             Palmer et al. (1977)
                                                 (equivalent to 45 mg/kg per day)

    Rat       diet                13 weeks       not established - lowest dose tested 160 mg/kg feed    Burdock et al. (1980b)

    Rat       diet                104 weeks      40 mg/kg feed (equivalent to 2 mg/kg                   Hunter et al. (1976)
                                                 per day body weight)

    Mouse     oral                2 weeks        200 mg/kg body weight                                  Keet (1977b)

    Mouse     diet                13 weeks       male/female: 16 mg/kg feed (equivalent to              Burdock et al. (1980a)
                                                 2.4 mg/kg body weight per day)

    Mouse     diet                14 weeks       not established - lowest dose tested 80 mg/kg feed     Colley et al. (1981a,b)

    Mouse     diet                91 weeks       male/female: 16 mg/kg feed (equivalent to              Colley et al. (1984)
                                                 2.4 mg/kg body weight per day)

    Cat       oral                3 weeks        male: 30 mg/kg body weight; female: not achieved       Schwartz & Borzelleca (1981)

    Dog       diet                13 weeks       male/female: 40 mg/kg feed                             Chesterman et al. (1974)

    Dog       oral (capsules)     52 weeks       male/female: 2 mg/kg body weight                       Greenough et al. (1985)
                                                                                                                                     
            Table 11.  Acute toxicity of diflubenzuron metabolites
                                                                                  

    Metabolite              Species   Sex       Route     LD50       Reference
                                                          (mg/kg)
                                                                                  

    4-Chlorophenylurea      rat       male      oral      1080       Koelman-Klaus
                                      female    oral      1210       (1978a)

    2,6-Difluorobenzoic     rat       male,     oral      4640       Koelman-Klaus
     acid                             female                         (1978b)
                                                                                  
    
    7.9.1  Carcinogenicity studies with 4-chloroaniline

         The diflubenzuron metabolite, 4-chloroaniline (PCA), has been
    assayed for carcinogenicity by the US NCI (1979) and by the US NTP
    (1989) using Fischer-344 rats and B6C3F1 mice on both occasions.  In
    the earlier of these studies, technical-grade PCA was administered at
    dietary concentrations of 250 and 500 mg/kg to rats and 2500 and
    5000 mg/kg to mice.  Groups of 50 male and 50 female animals of each
    species were randomized to the treatment groups at approximately six
    weeks of age.  The control groups consisted of 20 animals of each sex
    and species.  All animals which survived were treated for 78 weeks and
    observed, untreated, for a further 24 weeks (rats) or 13 weeks (mice). 
    Survival in all groups was good and it was judged that there were
    adequate numbers at risk for late-developing tumours.  In rats, the
    most significant findings were treatment-related proliferative splenic
    capsular and parenchymal lesions in males and females and, in male
    rats of the high-dose group, the occurrence of several types of
    unusual splenic neoplasms (i.e. fibroma, fibrosarcoma, sarcoma NOS,
    haemangiosarcoma and osteosarcoma) which appeared to arise from areas
    of capsular or parenchymal fibrosis.  These neoplasms were combined
    for analysis because it was considered that the fibromas were a benign
    form of sarcoma and that the neoplasms had a common cellular origin. 
    The combined incidences were 0/20 control, 0/49 low-dose and 10/49
    high-dose rats.  This result indicated a carcinogenic effect of
    treatment in male rats. There was no similar effect in the spleen of
    females.  In mice of each sex, there was increased incidence of
    haemangiomas and haemangiosarcomas in various organs.  The combined
    incidences were 2/20 in control males, 10/50 in low-dose males and
    14/50 in high-dose males, and 0/18 in control females, 3/49 in low-
    dose females and 8/42 in high-dose females.  In addition, there was,
    in female mice only, a non-significant increase in hepatocellular

    carcinomas and adenomas combined (0/18 control, 1/49 low-dose and 6/41
    high-dose).  The smaller number of control group animals in these
    experiments significantly reduced the statistical power.  It was
    recognized that PCA is unstable in feed and so the animals in the
    early experiments may have received lower doses than intended. 
    Consequently, PCA was administered by gavage with an aqueous vehicle
    containing hydrochloric acid in the later experiments (US NTP, 1989).

         Groups of 49 or 50 rats and mice of each sex (7 to 8 weeks old)
    were administered PCA at dose levels of 2, 6 or 18 mg/kg (rats) or 3,
    10, or 30 mg/kg (mice) on 5 days per week for 103 weeks.  Vehicle
    control groups of 50 males and 50 females received deionized water by
    gavage.  Survival was adequate for analysis, although variable. It was
    lower, for example, in the vehicle control groups of rats, which was
    attributable to the higher incidence of mononuclear cell leukaemia in
    these groups.  Significant, non-neoplastic findings in rats included
    treatment-related increases in the incidence of splenic fibrosis
    (males: 3/49 control, 11/50 low-dose, 12/50 mid-dose, 41/50 high-dose;
    females: 1/50 control, 2/50 low-dose, 3/50 mid-dose, 42/50 high-dose),
    lipocytic infiltration of the spleen (males: 0/49, 0/50, 0/50, 24/50;
    females: 0/50, 0/50, 0/50, 11/50) and adrenal medullary hyperplasia 
    in female rats (4/50, 4/50, 7/50, 24/50).  In addition, the 
    methaemoglobin level was consistently increased in the mid- and high-
    dose groups of male rats at 6, 12, 18 and 24 months.  There was no
    NOEL for this parameter in male rats.  Bone-marrow hyperplasia was
    also observed.  The combined incidences of uncommon splenic sarcomas
    (fibrosarcomas, osteosarcomas or haemangiosarcomas) were increased in
    male rats but not in females (males: 0/49, 1/50, 3/50, 38/50; females:
    0/50, 0/50, 1/50, 1/50).  There was also a small increase in
    phaeochromocytomas or malignant phaeochromocytomas in male rats
    (13/49, 14/48, 15/48, 26/49).  The incidences of mononuclear cell
    leukaemia were reduced in male and female rats of the treatment groups
    (males: 21/49, 3/50, 2/50, 3/50; females: 10/50, 2/50, 1/50, 1/50). 
    This reduction may be related to splenic toxicity, since splenectomy
    of Fischer rats at one to two months of age markedly reduces the
    incidence of mononuclear cell leukaemia later in life (Boorman et al.,
    1990).  However, similar reductions in the incidence of this neoplasm
    are not seen with several other aniline-like chemicals that also cause
    splenic toxicity.  In male mice, but not in females, the incidence of
    haemangiosarcomas of the liver and spleen was slightly greater in the
    high-dose group than in the controls (males: 4/50, 4/49, 1/50, 10/50)
    and the incidences of hepatocellular carcinomas or adenomas (combined)
    were also increased in treated male mice (males: 11/50, 21/49, 20/50,
    21/50).  The incidences of malignant lymphomas were slightly reduced
    in treated male and female mice (males: 10/50, 3/49, 9/50, 3/50;
    females: 19/50, 12/50, 5/50, 10/50).

         There are various points of similarity between the two studies
    described above: (a) in male and female rats there was splenic
    toxicity; (b) in male rats, there was a treatment-related increase in
    uncommon splenic sarcomas; and (c) in male mice, there was a
    treatment-related increase in haemangiosarcomas and haemangiomas
    combined.  The reductions in the incidence of mononuclear cell
    leukaemia in rats and malignant lymphomas in mice seen in the latter
    study were not observable in the earlier study because their incidence
    in the control groups of that study was already low.  It is concluded
    that PCA is carcinogenic in both mice and rats.

    8.  EFFECTS ON HUMANS

         No data concerning the effects of diflubenzuron on human health
    are available.

    9.  EFFECTS ON OTHER ORGANISMS IN THE LABORATORY AND FIELD

    9.1  Laboratory experiments

    9.1.1  Microorganisms

         Diflubenzuron was tested for its effects on morphogenesis in
     Streptomyces spp. Exposure to 400 mg diflubenzuron/litre resulted in
    reduced dominance of spore hairs and reduced width of the outer wall,
    and prevented formation of the inner spore wall in  S. babergiensis. 
    In  S. coelicolor, 1600 mg diflubenzuron/litre altered the structure
    of the fibrillar pattern of spore envelopes.  Exposure to diflubenzuron
    resulted in small increases in exported protein and in an approximately
    20% increase in chitinase in both  Streptomyces spp (Smucker &
    Simon, 1986).

    9.1.1.1  Water

         Aquatic bacterial biomass and density were not affected by
    diflubenzuron (1.0 µg/litre), although after three months of
    continuous exposure some decline in species diversity was found. 
    Owing to detrimental effects on aquatic arthropods, e.g., filter
    feeders that may use bacteria as a food source, such effects may very
    well be secondary in nature and not a direct effect of diflubenzuron
    (Hansen & Garton, 1982b).

         In a test on the alga  Selenastrum capricornutum, a nominal
    concentration of 0.20 mg diflubenzuron/litre did not reduce the
    biomass and can be considered a no-observed-effect concentration
    (NOEC) (Berends & Thus, 1992b).

    9.1.1.2  Soil

         Soil microorganisms were able to use diflubenzuron as carbon
    source when added as an acetone solution (Seuferer et al., 1979; see
    section 4.3.2.2).

         The effect of diflubenzuron (100, 200, 300, 400 and 500 mg/kg
    soil) was studied in non-sterile soil incubated under aerobic
    conditions and in sterilized soil inoculated with  Azotobacter
     vinelandii.  The presence of diflubenzuron had a stimulatory effect
    on nitrogen fixation in both non-sterile and sterile soil (Martinez-
    Toledo et al., 1988a).  At similar concentrations, diflubenzuron did
    not affect the growth of  Azotobacter vinelandii in culture media,
    either with or without a nitrogen source (Martinez-Toledo et al.,
    1988b).

    9.1.2  Aquatic organisms

    9.1.2.1  Microorganisms

         Cyanobacteria (the blue-green alga  Plectonema boryanum) grew
    rapidly in the presence of diflubenzuron (initial concentration
    0.1 mg/litre) with no visible signs of inhibited growth (Booth &
    Ferrell, 1977).  Concentrations of 1, 10, 50 and 100 mg/litre did not
    affect the growth of six species of fungi:  Rizopus arrhizus,
     Aspergillus niger, Fusarium oxysporum, Mycorrhizae-Rhizopogan
     vinicolor, Pythum debaranum and  Trichoderma viride (Booth et al.,
    1987).  However, these authors autoclaved the medium containing
    diflubenzuron, which led to extensive breakdown of the pesticide
    (Willems et al., 1977).

         Five-day lethality tests on the algae  Selenastrum capricornutum
    and  Anabaena flos-aquae resulted in NOAEC values above the exposure
    concentrations of 300 and 330 µg/litre, respectively (Thompson &
    Swigert, 1993b,c).  Similarly, tests on the diatoms  Navicula
     pelliculosa and  Skeletonema costatum led to NOAEC values of 380
    and 270 µg/litre, respectively (Thompson & Swigert, 1993d,e).

         Algae were affected at 1.0 µg/litre by technical diflubenzuron in
    dimethylformamide added to laboratory stream channels: the alga
    biomass increased and chlorophyll and phaeocitin levels were elevated. 
    Fungi were only affected temporarily by 0.1 µg/litre under the same
    circumstances.  Changes in species diversity had disappeared after
    2 months of continuous dosing (Hansen & Garton, 1982 b).

    9.1.2.2  Plants

         In a 14-day toxicity test on diflubenzuron in duckweed
     (Lemna gibba), the NOAEC was higher than the tested concentration
    (190 µg/litre) (Thompson & Swigert, 1993a).

    9.1.2.3  Invertebrates

         The acute toxicity of diflubenzuron to a number of non-target
    aquatic invertebrates is presented in Table 12.  Comprehensive eviews
    on the subject have been published recently, e.g., by Fischer & Hall
    (1992) and by Cunningham (1986) on the effects of diflubenzuron on
    estuarine crustaceans. The acute LC50 for insects ranges from
    1 µg/litre  (Diptera) to 250 µg/litre  (Coleoptera), and mayflies
    have an LC90 of 1 to 10 µg/litre. Other aquatic arthropods such as
    water fleas, scuds and sow bugs have LC50 values of 5 to 15 µg/litre.

        Table 12.  Acute toxicity of diflubenzuron for non-target aquatic invertebrates
                                                                                                                                              

    Species                Size/age    Stat/flowa     Temperature   Hardness     pH      Parameter      Concentration        Reference
                                                      (°C)          (mg/litre)                          (µg/litre)
                                                                                                                                              

    Daphnia magna          1st instar     stat            22            40       7.2     48-h EC50         15          Julin & Sanders (1978)
    (Water flea)

    Daphnia magna          24 h           stat            20            50               48-h LC50         4.55        Hansen & Garton (1982a)
    (Water flea)                          stat            20            100              48-h LC50         6.89        Hansen & Garton (1982a)
                                          stat            20            200              48-h LC50         4.42        Hansen & Garton (1982a)

    Daphnia magna          0-24 h                         20            294      8.1     48-h EC50         7.1         Kuijpers (1988)
    (Water flea)                                          20            294      8.1     24-h EC50         68          Kuijpers (1988)

    Gammarus pulex         mature         stat            12            40       7.2     96-h LC50         30          Julin & Sanders (1978)
    (Scud)

    Hyallela azteca        2-4 mm         flow            20            25               96-h LC50         1.84        Hansen & Garton (1982a)
    (Amphipod)

    Chironomus plumosus    4th instar     stat            22            40       7.2     48-h EC50         560         Julin & Sanders (1978)
    (Midge)

    Cricotopus sp.         4th instar     flow            20            25               EC50 (moulting    1.72        Hansen & Garton (1982a)
    (Midge)                                                                              success)

    Tanytarsus dissimilis  2nd instar     flow            20            25               EC50 (moulting    1.02        Hansen & Garton (1982a)
    (Midge)                                                                              success)

    Acartia tonsa          adult          constant        20            10b              5-day LC50        > 1000      Tester & Costlow
    (Copepod)                             daily                                                                        (1981)   
                                          replenishment
                                                                                                                                              

    Table 12 (Con't)
                                                                                                                                              

    Species                Size/age    Stat/flowa     Temperature   Hardness     pH      Parameter      Concentration        Reference
                                                      (°C)          (mg/litre)                          (µg/litre)
                                                                                                                                              

    Mysidopsis bahia       adult          intermittent    24-25         24-27b           96-h LC50         2.06        Nimmo et al. (1980)
    (Mysid shrimp)                        flow

                           adult          continuous      24-26         23-29b           21-day LC50       1.24        Nimmo et al. (1980)
                                          flow

    Palaemonetes pugio     larvae         stat            22            20b              96-h LC50         1.44        Wilson & Costlow
    (Grass Shrimp)                                                                                                     (1987)

                           post larval    renewal         22            20               96-h LC50         1.62        Wilson & Costlow (1987)
                                                                                                                                              

    a  Stat = static conditions (water unchanged for duration of test); flow = flow-through conditions (diflubenzuron concentration in water
       continuously maintained)
    b  Salinity (expressed as parts per thousand)
             Concentrations of diflubenzuron causing significant mortality of
    several freshwater invertebrates are presented in Table 13.

    Table 13.  Concentrations of diflubenzuron causing significant
               mortality of freshwater invertebratesa
                                                           

    Organisms                               Concentration
                                            (µg/litre)
                                                           

    Water flea (Daphnia magna)              2.0
    Amphipod (Hyalella azteca)              2.0
    Snail (Juga plicifera)                  > 36
    Snail (Physa spp.)                      > 36
    Caddis fly (Clisforonia magnifica)      0.1
    Midge (Tanytarsus dissimilis)           4.9
    Midge (Cricotopus spp.)                 1.6
                                                           

    a  From: Nebeker et al. (1983)


         The 96-h LC50 for the snails  Juga plicifera and  Physa sp.
    was > 45 µg/litre (Hansen & Garton, 1982a).

         The 48-h EC50 values of diflubenzuron metabolites for midge
    larvae were > 100 mg/litre for 4-CPU and 2,6-DFBA and 43 mg/litre for
    PCA (Julin & Sanders, 1978).

         Diflubenzuron suspended in water at a concentration of
    200 mg/litre was not directly toxic to the freshwater clam  Anodonta
     cygnea during 3 months of treatment.  Diflubenzuron produced
    disturbances in the calcification process in the lamellar layer of the
    shell.  Positive PAS reaction of the secretory cells on the outer
    mantle epithelium has been observed (Machado et al., 1990).

          Daphnia magna was continuously exposed to 14C-diflubenzuron at
    concentrations of 5.6, 14, 23, 40 and 93 ng/litre.  After 21 days of
    exposure daphnid survival at the highest concentration (93 ng/litre)
    was 50%.  In the remaining concentrations it ranged from 93 to 98%,
    which was comparable to the survival (99%) of the control organisms. 
    Reproduction and body length were affected only at the highest
    concentration.  The maximum acceptable toxicant concentration (MATC)
    of 14C-diflubenzuron for  Daphnia magna was > 40 and < 93 ng/litre
    (Surprenant, 1988).

         Technical diflubenzuron in dimethylformamide at 0.1, 1, 10 and
    50 µg/litre was added continuously to complex laboratory stream
    channels supplied periodically with field-collected microorganisms for
    5 months.  LC50 values ranging from 1.0 to 1.8 µg/litre were
    determined for four insect and crustacean species ( Tanytarsus
     dissimilis, Cricotopus sp. and  Hyalella azteca).  A chronic effect
    level was obtained only for  Daphnia magna (0.06 µg/litre).  Mayflies
    and stoneflies were the most sensitive.  They were severely affected
    at 1 µg/litre within one month (the sampling interval) and numbers
    were two to three orders of magnitude lower, almost leading to their
    elimination.  The survival of chironomids was reduced by 10 µg/litre
    (Hansen & Garton, 1982a,b) (see also section 9.1.1.1).

         Benthic communities in outdoor experimental streams were exposed
    to 1 or 10 mg/litre of diflubenzuron for 30 min.  The effect was
    assessed daily by examining drifting pupal exuviae over a period of
    one month following the treatment.  No drift of macrobenthos was
    induced at the time of application.  However, diflubenzuron affected
    the emergence of all species examined.  High larval mortality for a
    species of chironomid was observed directly in the stream treated with
    diflubenzuron, where numbers of mayfly nymphs and caddisfly larvae
    were also decreased (Yasuno & Satake, 1990).

         Technical diflubenzuron in acetone was applied at 5 µg/litre to
    two aquaria in a simulated field test outdoors.  Daphnid numbers were
    markedly reduced three days after treatment, but recovered slowly. 
    Copepods numbers were moderately reduced, exceeding the control
    numbers after 18 days.  The seed shrimp population showed no harmful
    effect (Miura & Takahashi, 1974a,b).

         In a study by Collwell & Schaefer (1980), diflubenzuron was
    applied to experimental ponds (mean concentration of 13.2 µg/litre) in
    California.  An hour after treatment, cladoceran ( Ceriodaphnia sp.,
     Diaphanosoma sp.,  Chydorus sp.,  Bosmina sp. and  Daphnia sp.)
    numbers were strongly reduced and did not return to pretreatment
    levels until more than 5 weeks after treatment. Copepod ( Diaptomus 
    sp. and  Cyclops sp.) numbers were also reduced but to a lesser
    extent and for a shorter period than for the cladocerans.  The
    rotifers increased in abundance in both the control and treated ponds
    during the first 8 days following treatment.

         Using laboratory tests to study mortality, Miura & Takahashi
    (1974a) found that crustaceans, especially the tadpole shrimp
     (T. longicaudatus), clam shrimp ( Eulimnadia spp.) and water fleas
    ( Daphnia and  Moina spp.), were highly susceptible to diflubenzuron
    at levels below 0.01 mg/litre. Copepods,  Cyclops and  Diaptomus 
    spp. showed some tolerance, whereas seed shrimp ( Cypricerus and
     Cypridopsis spp.) tolerated as much as 0.5 mg/litre.  Among aquatic

    insects tested, mayfly nymphs ( Callibaetis spp.) were most
    susceptible.  Aquatic midge larvae  (G. holoprasinus) also
    showed susceptibility.  However, dytiscid ( T. bassillaris and
     Laccorhilus  spp.) and hydrophilid beetles ( H. triangularis and
     T. lateralis  (mosquito predators)) demonstrated a strong tolerance. 
    Mosquito fish  (Gambusia affinis) showed no effect at a relatively
    high dose of
    1 mg/litre.

         When larvae of the crab  Rhithropanopeus harrisii were exposed
    to sublethal concentrations of diflubenzuron (0.05, 0.1, 0.3 and
    0.5 µg/litre), swimming speed increased in stage I, II and III zoeae,
    0.3 µg/litre being the lowest effective concentration.  Phototaxis was
    altered in stage IV only at concentrations as low as 0.1 µg/litre 
    (Forward & Costlow, 1978).

         Christiansen et al. (1978) showed that nearly 100% of
     Rhithropanopeus harrisii larvae at each of the four zoeal stages
    died when moulting to the succeeding stage after only 3 days of
    exposure to 10 µg diflubenzuron/litre.  This concentration was also
    lethal for larvae of  Sesarma reticulatum (Say).

         Christiansen & Costlow (1980) exposed larvae of the estuarine
    crab  Rhithropanopeus harrisii in laboratory conditions to 10 µg
    diflubenzuron/litre as an indicator of persistence of
    diflubenzuron in brackish water.  Disturbance of endocuticle
    deposition seemed to occur as soon as newly hatched larvae (less than
    12 h old) were exposed to diflubenzuron.  Diflubenzuron not only
    affected endocuticle deposition in the larvae, but also exocuticle
    deposition.  The only part of the crab larval exoskeleton that did not
    seem to be affected by diflubenzuron was the epicuticle (Cristiansen
    et al., 1978; Cristiansen & Costlow, 1982).

         Diflubenzuron, at concentrations of 0.02 and 0.2 mg/litre,
    enhanced mortality during moulting of the crab  Carcinus 
     mediterraneus (Czerniavsky) (Cardinal et al., 1979).

         Cirripede crustaceans, (barnacles,  Balanus eburneus) exposed to
    concentrations of 1 to 1000 µg/litre over a 28-day period showed a
    dose-dependent mortality.  Heavy mortality occurred during the second
    week of exposure.  Lethal and sublethal effects were observed at
    concentrations as low as 50 µg/litre (Gulka et al., 1980).  Disruption
    of the exoskeleton of  B. eburneus caused by diflubenzuron was
    similar to that observed in insects.  Development of barnacles exposed
    to diflubenzuron for 10 days or more at 750 and 1000 µg/litre was
    delayed in the premoult phase of cuticle secretion (Gulka et al.,
    1982).

         Wilson & Costlow (1986) found that diflubenzuron concentrations
    of 2.5 and 5 µg/litre were lethal to larvae of the grass shrimp
     (Palaemonetes pugio), causing 100% mortality on days 14 and 6,
    respectively.  Wilson et al. (1985) studied the effects of
    diflubenzuron upon phototaxis of larvae of  P. pugio.  The depression
    in positive phototaxis and elevation in negative phototaxis were most
    pronounced at 0.5 µg/litre, and the lowest test concentration to
    affect phototaxis was 0.3 µg/litre.  The alterations in photo
    responses varied with the embryonic stage at which exposure to
    diflubenzuron commenced.  This study was carried out at optimal
    salinity and temperature, but in the estuary these conditions
    fluctuate daily, which may amplify the observed effects.

         Nimmo et al. (1980) observed that chronic exposure of the mysid
    shrimp  Mysidopsis bahia to 0.075 µg diflubenzuron/litre reduced the
    reproductive success of both parents and progeny even after they were
    transferred to uncontaminated sea water.

         Diflubenzuron reduced the reproductive life span of adult brine
    shrimps  (Artemia salina) at levels of 2-10 µg/litre and caused death
    of immature shrimps within 3 days at concentrations above 10 µg/litre
    (Cunningham, 1976).

         Fiddler crabs  (Uca pugilator) were exposed to diflubenzuron
    (Dimilin WP-25%) at 0.5, 5 and 50 µg/litre for 1, 2, 3 and 4 weeks
    after multiple autonomy of one chela and five walking legs.  Exposure
    to diflubenzuron retarded the rate of limb regeneration in a dose-
    dependent fashion.  The effects of diflubenzuron were seen even in
    crabs exposed for only one week.  Significant retardation was evident
    by day 21 at 5.0 µg/litre  but was not statistically significant at
    0.5 µg/litre.  At 5 and 50 µg/litre moult-associated mortality was
    seen.  The number of setae on regenerated limbs was less than the
    number on the intact limbs.  The effects were reduced in experiments
    in which sediment was present (Weis et al., 1987).

         The burrowing activity of  Uca pugilator in sand under
    laboratory conditions was not altered when the sand was contaminated
    with 1 mg diflubenzuron/litre, indicating a lack of avoidance of
    diflubenzuron-contaminated sand.  However, exposure for 1 week to 0.5,
    5.0 or 50 µg/litre led to a decrease in the amount of burrowing
    activity.  The behavioral response was not changed after exposure for
    1, 2 or 3 weeks and was not concentration-related (Weis & Perlmutter,
    1987).

         Survival, moulting and behaviour of juvenile fiddler crabs were
    significantly affected by exposure to diflubenzuron (0.2, 2, 20 and
    200 µg/litre) for 24 h weekly during 10 weeks.  All crabs in the 200
    and 20 µg/litre groups died after 8 and 23 weeks, respectively.  The

    no-observed-effect concentrations (NOEC) for moulting (time to the
    first moult), survival (time until death), and behaviour (ability to
    escape from the test container) were 20, 2 and 0.2 µg/litre,
    respectively (Cunningham & Myers, 1987).

         Larvae of the stone crab  (Menippe mercenaria) were exposed to
    0.5, 1.0, 3.0, and 6.0 µg diflubenzuron/litre in combination with
    different temperature and salinity.  All of these concentrations were
    lethal to the larvae.  Evidence for synergistic effects of
    diflubenzuron and temperature or salinity was observed. Tolerance of
    the megalopa of the blue crab  Callinectes sapidus to diflubenzuron
    at concentrations of 0.5, 1.0, 3.0 and 6.0 µg/litre was slightly
    higher than for  Menippe mercenaria but was also dependent upon
    temperature and salinity.  At 20°C the percentage survival at a
    concentration of 1 µg/litre was similar to that observed for the
    controls (Costlow, 1979).

         Tester & Costlow (1981) reported that the marine copepod
     Acartia tonsa exposed to 1 and 10 µg diflubenzuron/litre for 36 h
    failed to produce viable nauplii even after they had been placed in
    clean sea water.  No viable nauplii were produced by these females for
    at least 30 h after treatment ended.

         Adult crustaceans were more resistant to exposure than their
    larvae at high concentrations (100-200 µg/litre) of technical grade
    diflubenzuron. Adults also exhibited significant mortality associated
    with moulting (Cunningham, 1976; Cardinal et al., 1979; Gulka et al.,
    1980).

         When larvae of horseshoe crabs  (Limulus polyphemus) were
    exposed to 5 and 50 µg diflubenzuron/litre, the crabs in the
    50 µg/litre group exhibited severe mortality immediately after
    ecdysis.  The larval stages were shown to be quite resistant to
    diflubenzuron, compared with other crustacean larvae (Weis & Ma,
    1987).

    9.1.2.4  Vertebrates

         Data on the acute toxicity of diflubenzuron and its metabolites
    for fish are presented in Tables 14 and 15, respectively.

         A 96-h acute toxicity test on juvenile sheepshead minnow
     (Cyprinodon variegatus) in a flow-through system resulted in no
    mortality at the nominal exposure concentration of 130 µg
    diflubenzuron/litre (Graves & Swigert, 1993).  A similar test under
    semi-static conditions on zebra fish  (Brachydanio rerio) and rainbow
    trout  (Oncorhynchus mykiss) at a nominal diflubenzuron concentration
    of 200 µg/litre also showed no mortality, nor changes in the appearance

        Table 14.  Acute toxity of diflubenzuron to fish
                                                                                                                                              

    Organism                 Size/age      Stat/flowa     Temperature   Hardness      pH        Parameter    Concentration   Reference
                                                          (°C)          (mg/litre)                           (mg/litre)
                                                                                                                                              

    Coho salmon              1 g           stat           11            4.55          6.5       96-h LC50    > 150           McKague &
    (Oncorhynchus kisutch)                                                                                                   Pridmore (1978)

    Rainbow trout            1 g           stat           11            4.55          6.5       96-h LC50    > 150           McKague &
    (Oncorhynchus mykiss)                                                                                                    Pridmore (1978)

    Rainbow trout            1.2 g         stat           12            40            7.2       96-h LC50    240             Julin & Sanders
    (Oncorhynchus mykiss)                                                                                                    (1978)

    Rainbow trout            not           flow-through                                         96-h LC50    140             Marshall & Hieb
    (Oncorhynchus mykiss)    reported                                                                                        (1973)

    Rainbow troutb           not           flow-through                                         96-h LC50    195             Marshall & Hieb
    (Oncorhynchus mykiss)    reported                                                                                        (1973)

    Fathead minnow           0.87 g        stat           22            40            7.2       96-h LC50    430             Julin & Sanders
    (Pimephales promelas)                                                                                                    (1978)

    Channel catfish          2.2 g         stat           22            40            7.2       96-h LC50    370             Julin & Sanders
    (Ictalurus punctatus)                                                                                                    (1978)

    Bluegill                 0.5 g         stat           22            40            7.2       96-h LC50    660             Julin & Sanders
    (Lepomis macrochirus)                                                                                                    (1978)

    Bluegill                 not           flow-through                                         96-h LC50    135             Marshall & Hieb
    (Lepomis macrochirus)    reported                                                                                        (1973)

    Bluegillb                not           flow-through                                         96-h LC50    230             Marshall & Hieb
    (Lepomis macrochirus)    reported                                                                                        (1973)
                                                                                                                                              

    Table 14.  (Con't)
                                                                                                                                              

    Organism                 Size/age      Stat/flowa     Temperature   Hardness      pH        Parameter    Concentration   Reference
                                                          (°C)          (mg/litre)                           (mg/litre)
                                                                                                                                              

    Mummichog                adult 2.7 g   stat           24            22 ppt        8.0       96-h LC50    32 990          Lee & Scott
    (Fundulus heteroclitus)                renewal                      salinity                                             (1989)

    Mummichogb               not           flow-through                                         96-h LC50    255             Marshall & Hieb
    (Fundulus heteroclitus)  reported                                                                                        (1973)
                                                                                                                                              

    a  Stat = static conditions (water unchanged for duration of test); flow = flow-through conditions (diflubenzuron concentration in water
       continuously maintained)
    b  Formulated product 25 WP

    Table 15.  Acute toxicity of metabolites of diflubenzuron to fish (from: Julin & Sanders, 1978)
                                                                                                                         

                                                                               Concentrations (mg/litre)
    Organism                  Water         Effect                                                                       
                              temperature   measured
                              (°C)                         4-Chlorophenyl      2,6-Difluorobenzoic      4-Chloroaniline
                                                           urea                acid
                                                                                                                         

    Rainbow trout             12            96-h LC50      72 (57-90)          > 100                    14 (11-16)
    (Oncorhynchus mykiss)

    Channel catfish           22            96-h LC50      > 100               > 100                    23 (18-29)
    (Ictalurus punctatus)

    Fathead minnow            22            96-h LC50      > 100               69 (55-87)               12 (7-18)
    (Pimephales promelas)

    Bluegill                  22            96-h LC50      > 100               > 100                    2.4 (1.8-3.2)
    (Lepomis macrochirus)
                                                                                                                         
        or behaviour of the fish (Berends & van der Laan-Straathof, 1994a,b).
    All these test concentrations were above the diflubenzuron solubility
    level of 80 µg/litre.

         Nebeker et al. (1983) found no significant reduction in
    survival of fathead minnow  (Pimephales promelas) or guppy
     (Poecilia reticulata) as a result of exposure to diflubenzuron at
    concentrations below 36 µg/litre during acute (96-h) and chronic
    tests.

         No acute response resulted from exposure of fish to a
    diflubenzuron concentration of 45 µg/litre, and no chronic effects
    were observed at this concentration, the highest one tested (Hansen &
    Garton, 1982a,b).

         Diflubenzuron was not toxic to either rainbow trout or coho
    salmon exposed at concentrations up to 150 mg/litre for a 96-h period. 
    A 15-min exposure to 1 g/litre did not result in any fish mortality
    (McKague & Pridmore, 1978).

         Madder & Lockhart (1978) found a dose-related decrease of
    glutamic-oxaloacetic transaminase activity in rainbow trout
     (Oncorhynchus mykiss) exposed to diflubenzuron at concentrations of
    0.625, 1.25, 2.5, 5 and 10 mg/litre.

         At a concentration of 0.01 mg/litre, diflubenzuron had a
    repellent effect on precocious male Atlantic salmon parr (Granett et
    al., 1978).

    9.1.3  Terrestrial organisms

    9.1.3.1  Plants

         Photosynthesis, respiration and leaf ultrastructure of soybeans
    were unaffected by diflubenzuron at doses up to a level of
    0.269 kg a.i./ha (Hatzios & Penner, 1978).

         Diflubenzuron is used as an insecticide in forestry, agriculture
    and horticulture.  No phytotoxicity was reported in the field studies
    cited in section 9.2.

    9.1.3.2  Invertebrates

         The oral and contact LD50 values of diflubenzuron for honey-bees
    are greater than 30 µg/bee (Stevenson, 1978).

         Diflubenzuron did not show any toxicity to bees at concentrations
    up to 1000 mg/kg in the diet (Yu et al., 1984).

         Barker & Taber (1977) found that diflubenzuron reduced brood
    production when fed for 10 days to honey-bees at 59 mg/kg in sugar
    syrup, but not at 5.9 or 0.59 mg/kg.

         Barker & Waller (1978) confirmed that 60 mg/kg in sugar syrup or
    100 mg/litre in water caused colonies to produce less brood.

         Stoner & Wilson (1982) found that diflubenzuron fed to flying
    colonies at 1 or 10 mg/kg for a year significantly reduced the amount
    of sealed brood.

         Nation et al. (1986) reported that diflubenzuron did not cause
    reduction in pollen consumption or brood production when fed for 10
    weeks to caged colonies of honey-bees at 10 mg/kg, but it caused more
    than 50% reduction in the amount of syrup stored.

         Gordon & Cornect (1986) showed that, at concentrations that are
    effective in suppressing egg hatching and larva development of the
    cabbage maggot  Delia radicum, diflubenzuron did not adversely
    affect eggs, first-instar larvae or adults of the rove beetle
     Aleochara bilineata, an important predator and parasitoid of the
    cabbage maggot.

         When the acute toxicity of diflubenzuron to the earthworm
     Eisenia fetida was tested in artificial soil to which diflubenzuron
    had been added, 780 mg/kg dry soil was the no-observed-effect
    concentration (NOEC) for the 14-day test period (Berends et al.,
    1992).  For the WP-25 formulation, the NOEC was 1.0 g/kg (Berends &
    Thus, 1992a).

    9.1.3.3  Vertebrates

    a)  Birds

         The acute oral LD50 of technical diflubenzuron for red-winged
    blackbirds  (Agelaius phoeniceus) is 3762 mg/kg body weight.  In an
    8-day dietary LD50 study on mallard duck and bobwhite quail using
    technical diflubenzuron, levels up to 4640 mg/kg in the feed gave no
    observable signs of toxicity (Maas et al., 1980).

         When diflubenzuron was fed to mature White leghorn hens at
    dietary levels of 10, 50, 100, and 500 mg/kg for 8 weeks, there were
    no adverse effects on feed consumption, body weight, egg production,
    egg weight, eggshell thickness, fertility, hatchability or progeny
    performance.  The highest dose used was 50 times higher than the
    efficacy level for fly control (Cecil et al., 1981).

         In chronic toxicity tests, groups consisting of 10 male and 10
    female one-day-old chicks of barred Plymouth rocks and white leghorn
    hens, Nicholas white turkeys, mallard ducks and ring-necked pheasants
    were given diets containing diflubenzuron levels of 0.25, 1.25, 25 and
    250 mg/kg for 91 days after hatching.  No differences between control
    and treatment groups were observed in mortality, food consumption,
    body weight, comb and wattle development, weight of inner organs,
    serum hormone levels or general behaviour (Maas et al., 1980).

         There was no effect of diflubenzuron on the content of hyaluronic
    acid in the skin of Hubbard broiler chickens when they were fed at
    levels of 2.5 and 250 mg/kg in the diet for 98 days after hatching
    (Deul & Jong, 1977). In an experiment with the same dose levels and
    duration, diflubenzuron had no effect on hyaluronic acid synthesis or
    comb deposition in either growing broilers or layers (Crookshank et
    al., 1978).

         White leghorn and black sex-linked cross hens were fed
    diflubenzuron at a level of 10 mg/kg in the ration for 15 weeks. 
    Diflubenzuron had no effect on body weight gain, egg production,
    fertility or hatchability (Miller et al., 1976b).

         In a feeding study with 2.5, 25 and 250 mg/kg, diflubenzuron did
    not affect bobwhite quail reproduction (Booth et al., 1987). 
    Diflubenzuron did not show significant teratogenic activity on chick
    embryos over time when injected at 10 mg/egg (Booth et al., 1987).

         A one-generation bobwhite quail  (Colinus virginianus)
    reproduction study was conducted in which a diflubenzuron-containing
    diet was administered  ad libitum to young adults (24 weeks old at
    test initiation) approaching their first breeding season.  Dietary
    concentrations of 250, 500 or 1000 mg/kg did not result in treatment-
    related mortality, overt signs of toxicity or effects upon adult body
    weight or feed consumption during the 21-week exposure period.  There
    were no apparent treatment-related effects upon reproductive
    parameters at 250 or 500 mg/kg.  There may have been a slight reduction
    in the number of eggs laid, although this was not statistically
    significant or dose-related at 1000 mg/kg.  On the basis of a possible
    effect on egg production at 1000 mg/kg, the NOEC for diflubenzuron in
    this study was above 500 mg/kg (Beavers et al., 1990a).

         In a one-generation reproduction study on the mallard duck
     (Anas platyrhynchos), diets containing diflubenzuron were
    administered  ad libitum to young adults (27 weeks old at test
    initiation) approaching their first breeding season.  Dietary
    diflubenzuron concentrations of  250, 500 and 1000 mg/kg did not
    result in treatment-related mortality, overt signs of toxicity  or
    effects upon adult body weight or feed consumption during the 20-week
    exposure period.  There were no apparent treatment-related effects

    upon reproductive performance at any of the concentration tested.  At
    1000 mg/kg there was a slight, but statistically significant, 
    reduction in mean eggshell thickness.  On the basis of the effect upon
    eggshell thickness at 1000 mg/kg, the NOEC for diflubenzuron in this
    study was 500 mg/kg (Beavers et al., 1990b).

    b)  Mammals

         In studies by Ross et al. (1977a,b), diflubenzuron was
    administered in the feed to sheep (3 of each sex per group) as a model
    for ruminant wildlife at concentrations of 500, 2500 and 10 000 mg/kg
    feed for 13 weeks.  No treatment-related effects were observed on food
    consumption, body weight gain, haematological parameters or
    urinalysis.  Increase in met- and sulfhaemoglobin levels were observed
    at 13 weeks and there was a reduction in the weight of the thyroid. 
    No histopathological abnormalities were observed.  Both the plasma and
    the erythrocyte cholinesterase activities were unaffected by the
    treatment after 6 weeks.

    9.2  Field observations

    9.2.1  Microorganisms

    9.2.1.1  Water

         The laboratory data available (see section 9.1.1.1) make it
    unlikely that detrimental effects will occur.

         Rotifers were unaffected by diflubenzuron (28 and 56 g a.i./ha)
    in both experimental and naturally treated ponds (Ali & Lord, 1980).

    9.2.1.2  Soil

         One aerial application of diflubenzuron at 67.26 g/ha had no
    adverse effects upon populations of bacteria, actynomycetes or fungi
    in leaf litter and forest soil (Wang, 1975; Kurczewski et al., 1975).

    9.2.2  Aquatic organisms

    9.2.2.1  Plant

         The laboratory data available (see section 9.1.2.2) make it
    unlikely that detrimental effects will occur.

    9.2.2.2  Invertebrates

         Recent field studies have demonstrated that effects on aquatic
    fauna are limited and  transient, and that recovery is evident after

    3 months (Huber & Collins, 1987; Kingsbury et al., 1987; Huber &
    Manchester, 1988; Ali et al., 1988; Ali & Kok-Yokomi, 1989; Sundaram
    et al., 1991).

         Diflubenzuron (25% WP) applied at 33.63 and 134.52 g a.i./ha,
    4 times at 2-week intervals, had no adverse effect on freshwater clams
    10 days after final treatment (Jackson, 1976).

         When diflubenzuron (25% WP) was applied at 1.121-280.25 g a.i./ha
    to flooded rice fields in Louisiana, USA, significant reduction of
     Tropisternus spp. and  Libellulidae was found 80 days after
    treatment.  Significantly more chironomid and baetid immatures
    occurred due to reduction in the number of predators (Steelman et al.,
    1975).

         Mulla et al. (1975) found that at an application rate of 28 and
    56 g a.i./ha diflubenzuron reduced slightly, and only for a short
    time, the number of mayfly ( Beatis sp.) in the treated ponds.  The
    numbers, however, appeared to be within natural fluctuation limits
    equal to the check ponds or the pretreatment levels during all other
    sampling periods.  At these application rates, diflubenzuron caused a
    short-term reduction in copepod populations, but they started to
    increase on the 11th or 15th day of post-treatment sampling.  There
    was little or no effect on the ostracod population.

         In a forestry spraying programme at 0.0672 kg a.i./ha, aquatic
    arthropods were studied in a water shed (White Deer Creek).  Sampling
    at three test stations and one control station in the watershed
    revealed a rich fauna, dominated numerically by Ephemeroptera,
    Chironomida, Trichoptera and Plecoptera in descending order of
    abundance.  Significant differences in larval abundance in the surber
    samples were found to occur from one sampling day to the next, or
    between pre-spray and post-spray periods, for most of the organisms
    examined.  In all cases, control and test stations showed similar
    patterns of variation, so treatment effects were discussed as a
    probable cause.  The number of organisms tended to be higher at the
    upstream stations.  Chironomida and Trichoptera pupal abundance in the
    surber samples was examined for decreases after spraying but numbers
    were too low to permit statistical analysis.  Pupal abundance showed
    some direct relationship to larval abundances.  Nymphal drift rates
    were examined for possible increases after spraying, but most of the
    significant changes were decreases during the post-spray period. 
    These decreases also occurred at the control station.  The drift rates
    of nymphal and pupal exuviae also did not indicate a treatment effect. 
    Individual species abundances in the surber samples were examined for
    possible selective effects of diflubenzuron.  Comparisons were made
    between pre-spray and post-spray periods at both test and control
    stations, but no evidence of a treatment-related decrease in the
    abundance of any of these organisms was found.  The net effect was
    usually an increase in the density of these organisms during the post-
    spray period.  No organisms showing abnormal ecdysis or pupation were

    found in any of the samples.  It was concluded that spraying with
    diflubenzuron had no adverse effect on the macrobenthic community in
    White Deer Creek (White, 1975).

         In a study by Booth (1975), diflubenzuron (25% WP) was applied at
    44.84 g a.i./ha to small ponds in Utah.  Biosamples taken 30 and 80
    days later showed that, although larva and immature aquatic insect
    populations were decreased at 30 days, the total number of adults was
    not significantly different from the control number at 80 days.

         Booth & Ferrell (1977) examined the effect of diflubenzuron on
    over 20 different species (corixidae and collembolans) after multiple
    pond applications at 45 g a.i./ha.  Only immature corixidae (water
    boatmen) and collembolans (springtails) were significantly affected.

         Segmented worms  (Oligochaeta) and midges  (Chironomidae) 
    were unaffected by six applications of diflubenzuron at a rate of
    145.73 g a.i./ha to Utah Lake (Booth et al., 1987).

         Diflubenzuron (28 and 56 g a.i./ha) reduced numbers of
     Chaoborus sp. and  Baetis sp. in both experimental and natural
    treated ponds.   Chaoborus sp. recovered within 1-3 weeks (Ali &
    Lord, 1980).

         The application of granular diflubenzuron at 0.11 kg a.i./ha
    (about 3.7 µg/litre) to residential-recreational lakes caused a 62-75%
    reduction of  Daphnia pulex and  Daphnia galeata during 7 days
    following treatment.  The populations recovered in the second week
    after treatment.  A 30% reduction in  Diaptomus spp.  was noted 2
    days after the treatment.  Hyalella azteca was affected markedly,
    with a maximum reduction of 97% after 3 weeks of treatment.  No
    detectable effects on  Cyprinotus sp.,  Cyclops sp. or  Bosmina
     longirostris were observed (Ali & Mulla, 1978a).  Ali & Mulla
    (1978b) also studied the impact of diflubenzuron on invertebrates in a
    residential-recreational lake after two treatments with 25% WP
    formulation at 156 g a.i./ha (about 12 µg/litre).  Diflubenzuron
    concentrations in water were not measured but were based on nominal
    initial concentrations.  The results from this study are summarized in
    Table 16.

         Diflubenzuron (25% WP) at rates of 0.02, 0.025, 0.03, 0.035,
    0.04, 0.045 and 0.05 lb a.i./acre was applied to 19 irrigated pastures
    and spring ponds (in some cases up to 4 times) at 2- to 3-week
    intervals in California.  Cladoceran and nymphal mayfly populations
    were temporarily and slightly reduced, but recovered within a short
    period of time.  Even repeated treatment of the same pastures did not
    eliminate the populations.  The impact on copepod populations was
    inconclusive.  Adult beetles demonstrated high tolerance to

        Table 16.  Effect of diflubenzuron on invertebrates in Vallage Grove Lake, Californiaa
                                                                                                                                     

    Species                        First application (April)                       Second application (August)
                                                                                                                                     

                             Effect                   Recovery                 Effect                   Recovery
                                                                                                                                     

    Daphnia leavis           elimination within       no recovery 6 months
    Birge                    1 week                   after treatment

    Ceriodaphnia sp.         elimination within       no recovery 6 months
                             1 week                   after treatment

    Bosmina longirostris     elimination within       recovery after           elimination after        reappearance in small numbers
    (O.F. Muller)            1 week                   11 weeks                 1 week                   8-9 weeks after treatment

    Cyclops spp.             elimination within       recovery within          elimination within       recovery after 4 weeks
                             1 week                   6-7 weeks                1-2 weeks

    Diaptomus sp.            elimination within       recovery after           absent prior to          reappearance in small numbers
                             1 week                   4 months                 treatment                1-2 months later

    Hyalella azteca          elimination within       no recovery 6 months
    (Saussure)               4 weeks                  after treatment

    Caenis sp.               elimination within       recovery within          elimination within       recovery in 4-5 weeks
                             3 weeks                  6-7 weeks                2-3 weeks

    Physa sp.                no adverse effects       no adverse effects

    Cypridopsis sp.          no adverse effects       no adverse effects
                                                                                                                                     

    a  From:  Ali & Mulla (1978b)
        diflubenzuron, but few dead larvae ( Laccophilus spp.,  Hydrophilus
     triangularis and  Tropisternus lateralis) were observed.  Spiders
    ( Pardosa spp. and  Lycosa spp.) showed no apparent effects.  There
    were no deleterious effects on planarian, rotifer, seed shrimp and
    fresh water flagellate populations (Miura, 1974).

         Diflubenzuron (25 WP formulation) was applied 3 times at 2.5-week
    intervals to man-made pools in Manitoba, Canada at 56 g/a.i. ha (this
    produced initial concentrations of 0.02 mg/litre).  The treatment
    produced no detectable effect on chironomid larvae in terms of numbers
    or composition, although dead larvae were noted after treatment. 
    Daphnid populations were significantly reduced on most sampling dates. 
    Repetitive treatments prevented the recovery of cladocerans.  Despite
    these reductions, daphnids were not annihilated.  At a higher rate
    (0.22 kg a.i./ha, about 7.4 µg/litre), both species of  Daphnia were
    eliminated for 3 months after treatment.  Populations of  Diaptomus 
    spp. declined to zero 7 days after treatment, but recovered during the
    second week following treatment.  Diflubenzuron caused reductions in
    the number of  H. azteca (32-100%) during the 2´ months following
    treatment.  Seed shrimp populations were stressed for only 2 weeks,
    and there were no observable effects on oligochaetes at a treatment
    level of 0.22 kg a.i./ha (about 7.4 µg/litre).  Copepods only
    occasionally showed significant reduction, and recovered within 10
    days after treatment.  Of the ten other non-target invertebrates (i.e.
     Chaoboridae, Tipulidae, Ephemeroptera, Corixidae, Nonectidae,
     Gerridae, Dystiscidae, Hydrophilidae, Hydrarina and  Gastropoda),
    only mayfly nymphs  (Ephemeroptera) were significantly reduced
    (Madder, 1977).

         When diflubenzuron at a rate of 34.2 g/ha was applied aerially to
    a forested area in the USA, there were no significant effects on the
    population structure of the benthic macro-invertebrate community. 
    Some decreases in the number of mayflies  Heptagenia sp. and
     Rhithrogenia sp. were noted in treatment streams, but the total taxa
    richness values remained high. An increase in abundance of the
    caddisfly  Lepidostoma sp. was found.   Allonarcys sp. was not
    affected by the treatment (Huber & Collins, 1987; Huber & Manchester,
    1988).

         After aerial application of diflubenzuron to two forest ponds in
    Canada the greatest effect was on crustacean zooplankton, especially
    cladocerans, with limited effect on pond benthos.  Recovery of
    population of even the most severely affected organism such as
    Daphnids was well established by 3 months after treatment.  The
    maximum residue found in water was 13.82 µg/litre 1 h after
    application (Kingsbury et al., 1987).

         Diflubenzuron WP-25 was applied at a rate of 70 g active
    ingredient in 10, 5 or  2.5 litre/ha to three spray blocks in a mixed
    boreal forest near Kaladar, Ontario, Canada.  Water, sediment and
    aquatic plants were collected from two ponds and a stream at intervals
    up to 30 days after treatment for analysis of diflubenzuron residues. 
    The duration of detectable residues was different for each substrate,
    but in all cases it was less than 2 weeks.  Zooplankton and benthic
    invertebrate populations were monitored for up to 110 days after
    spraying in two ponds in the high-volume rate block and in control
    ponds. Significant mortality occurred in two groups of caged
    macroinvertebrates (amphipoda and immature corixidae) 1 to 6 days
    after the ponds were treated with diflubenzuron.  Three taxa of
    littoral insects ( Caenis, Celithemis and  Coenagrion) were
    significantly reduced in abundance in the treated ponds 21 to 34 days
    after treatment but recovered to pre-treatment levels by the end of
    the season.  Of the six remaining groups studied, only one (immature
    corixidae) may have been slightly affected by treatment.  Zooplankton
    (cladocera and copepoda) population numbers were reduced 3 days after
    treatment and remained suppressed for 2-3 months (Sundaram et al.,
    1991).

         Blumberg (1986) found no effect on population numbers in a
    forested ecosystem after aerial application of diflubenzuron (25 WP)
    at rate 140 g a.i./ha.

         When diflubenzuron (33.63 and 134.52 g a.i./ha) was applied 4
    times at 2-week intervals to small ponds in Virginia, numbers of
    daphnids were markedly reduced at both treatment levels.  There was no
    appreciable effect on two other invertebrate species ( Chironomidae 
    or  Chaoborus) (Birdsong, 1977).

         In a study by Rodrigues (1982), diflubenzuron (25% WP) was
    applied at a rate of 1 mg/litre per 30 min to three forest streams in
    New York.  The reduction of chironomid larvae 15 days after treatment
    ranged from 35 to 91% at different sites in the three streams.  Of the
    two stonefly families present in the three streams,  Nemouridae were
    affected more than  Leuctridae, with decreases of 75-94% and 70%,
    respectively.   Ephemeroptera were reduced, but  Trichoptera and
     Coleoptera were unaffected.

         Following aerial application at 67.26 g a.i./ha to a watershed,
    diflubenzuron reached the stream channel (also as a result of wash-off
    from foliage following several subsequent rainfalls), but the
    invertebrate populations were not affected (Jones & Konchenderfer,
    1988).

         After two applications of 44.8, 112 and 224 g a.i./ha, grass
    shrimp showed mortality of 86.6, 100, and 100%, respectively.  At the
    two lowest application rates, fiddler crabs ( Uca spp.) showed
    mortalities of 53.3 and 66.6%, respectively (McAlonan, 1976).

         Diflubenzuron (25% WP) at 33.63 and 134.52 g/ha was applied 4
    times at 2-week intervals to man-made ponds in North Carolina,
    Arkansas and Texas, USA.  In North Carolina no apparent effect on
    phytoplankton was found.  There was a marked decrease in the numbers
    of crustaceans ( Cladocera and  Copepoda), as well as of certain
    species from benthic communities such as  Hexagenia and  Chaoborus 
    in Arkansas.  A relative decrease in copepod numbers was observed, but
    the authors attributed this partly to natural mortality during winter. 
    In Texas there was a marked decrease in the numbers of all crustaceans
    and a corresponding increase in rotifer populations, particularly
     Asplanchina (Aquatic Environmental sciences, 1976a,b,c).

         When diflubenzuron (25% WP) at 33.63 and 134.52 g/ha was applied
    4 times at 2-week intervals to man-made ponds in Alabama, USA, daphnid
    populations were reduced to 50% of the pre-treatment level at a rate
    of 134.52 g/ha but increased at 33.63 g/ha to a level of 15% of that
    in a control pond.  Gastropod numbers decreased in the benthic samples
    but increased in the periphyton samples from the suspended plate
    samplers in the treated ponds (Jackson, 1976).

         Diflubenzuron at 33.63 and 134.52 g/ha was applied 4 times at
    2-week intervals to a tidally influenced salt marsh in Virginia, which
    was sampled biologically 14 times over a 70-day period.  Three species
    of arthropods,  Cyathura polita, Ceratopogonidae sp., and  Psychodidae
    sp., showed significant reductions in numbers.  Oligochaetes,  Hanynkia
     speciosa and the molluscs were not significantly affected (Matta,
    1976).

         Six applications of diflubenzuron (28 g a.i./ha) over an 18-month
    period caused statistically significant differences in the population
    density of aquatic organisms in a Louisiana coastal marsh. None of the
    organisms affected were completely eliminated from the ecosystem. 
    Populations of five taxa (nymphs of  Trichocorixa louisianae  Jaczewski
    and  Buenoa spp., Coenagrionidae naiad spp.,  Berosus  infuscatus Le Conte
    adults, and  Hyalella azteca Saussure) were significantly reduced.
    Population of 15 taxa, i.e.  Physa sp.,  Ceanis sp. and  Callibaetis sp.
    naiads,  Noteridae larvae,  Hydrovatus cuspidatus, Kunze adults,
     Hydrovatus sp. larvae, Dytiscidae larvae,  Mesovelia mulsanti Jaczewski
    adults,  Trichocorixa louisiana adults, larvae of Chironomidae, Ephudridae,
    Dolichopodiae and Tabanidae, and the fish  Gambusia affinis (Baird and
    Griard) and  Jordanella floridae (Goode and Bean), showed significant
    increases after exposure to diflubenzuron.  The 27 remaining aquatic
    organisms (members of the Hemiptera, Coleoptera, Mysidacea, Decapoda,
    Diptera and Odonata) showed no statistically significant differences
    when compared with untreated populations (Farlow et al., 1978).

         After treatment with diflubenzuron (as 1% granular formulation)
    at rate of 4.5 g a.i./ha to a marsh habitat on the Fraser River,
    Canada, the population of cladocerans appeared to be depressed for
    about 5 days but they recovered 2 weeks after treatment.  There was no
    effect on copepods or ostracods.  Significant reduction in the numbers
    of water beetles and zooplankton occurred (Wan & Wilson, 1977).

         In a field study by Hester et al. (1986), WP-25 formulation was
    evaluated in natural salt water pools to determine its toxicity to
    juvenile stages of three estuarine crustaceans exposed to a single
    application of 45 g diflubenzuron/ha.  One hour after application,
    water residues of diflubenzuron averaged 3.6 µg/litre and mortality
    was 46.5, 60.7 and 40.6% for  Callinectes sp.,  Palaemonetes pugio 
    and  Uca pugilator, respectively, over the 10-day observation period. 
    When these species were introduced into the pools 7 days after
    treatment, mortality was not significantly greater in the treated
    group than in the controls over the 21-day observation period.  The
    maximum concentration after organisms were introduced was
    0.69 µg/litre, and mortalities of 22.2, 1.5 and 4.2% for  Callinectes
    sp.,  Palaemonetes pugio and  Uca pugilator, respectively, were
    reported.

         In a study by Hester (1982), diflubenzuron (25% WP) at the rate
    of 0.045 kg a.i./ha was applied to tidal ponds.  Residue analysis
    showed 2 to 5 µg/litre after 1 h.  The mortality of three
    estuarine decapods: blue crabs ( Callinectes sp.), grass shrimp
     (Palaemonetes pugio) and fiddler crabs  (Uca pugilator) was 44,
    61 and 41%, respectively.  When these decapods were introduced to the
    ponds 7 days after application, the residues in water were
    0.4-0.7 µg/litre and mortality was 22, 1.5 and 4%, respectively.

    9.2.2.3  Vertebrates

         During a forestry spraying programme at 0.067 kg diflu-
    benzuron/ha, a survey on fish was performed in the watershed.  
    Observations made on caged brown trout in the stream from day -7 to
    day +6 revealed no immediate mortality or erratic behaviour
    attributable to treatment.  Barrier seines placed at the upstream and
    downstream boundaries of the spray area did not collect any dead or
    dying fish during the 1-week period they were in the stream.  Visual
    observations made by walking along the stream during this same period
    also revealed no dead or dying fish.  Post-spray population
    estimations revealed increases in both test and control areas. 
    However, statistical analysis indicated no significant differences
    pre- and post-spray between stations.  The treatment had no observable
    effect on the development of fish larvae.  Immediate mortality was not
    observed in caged fish or stream fish collected by barrier seines. 
    Delayed mortality attributable to spraying was not shown by population
    estimations taken 6 weeks following spray, nor were any dead or dying

    fish seen during this period.  There appeared to be no adverse effect
    of diflubenzuron sprayed at 0.067 kg a.i./ha on the fish species
    observed in this study (White, 1975).

         No effect on fish populations ( Micropterus salmoides, Lepomis
     macrochirus and  Gambusia affinis) was observed after four
    applications of diflubenzuron (33.69 and 134.52 g a.i./ha) at 2-week
    intervals to small ponds in Virginia (Birdsong, 1977).

         Killifish  (Fundulus heteroclitus) showed no effect after three
    applications of diflubenzuron at rates from 11.12 to 224.2 g a.i./ha
    to replicated semi-natural pools (McAlonan, 1976). Diflubenzuron
    revealed no toxic effect on bullheads or sunfish after aerial
    application of 35 g/ha in Canada (Buckner et al., 1975).

         Growth of bluegill sunfish ( Lepomis macrochirus Rafinesque) was
    not affected by diflubenzuron applications at rates of 2.5-10 µg/litre
    to ponds and lakes in California (Apperson et al., 1978).  Ten days
    after the fourth treatment to man-made ponds with diflubenzuron at
    33.63 and 134.52 g a.i./ha no apparent adverse effect on largemouth
    bass was observed (Jackson, 1976).

         No death of fish ( Pomoxis nigromaculatus and  Ictalurus
     nebulosis) occurred after application of diflubenzuron (mean
    concentration of 13.2 µg/litre one hour after treatment) to
    experimental ponds in California.  For 1 month following treatment,
    stomach content analyses showed alteration in the diet of the fish
    (Collwell & Schaefer, 1980).

    9.2.3  Terrestrial organisms

    9.2.3.1  Invertebrates

         Honey-bee colonies remained normal after an aerial application of
    350 g diflubenzuron/ha in Canada (Buckner et al., 1975).

         A comparative study of the effect of diflubenzuron on carabid
    fauna in oak woods in Westphalia in 1987-1988 was reported.  The total
    number of captured beetles in treated areas was lower in 1989. 
    Evaluation of the 12 beetle species most frequently caught indicated
    lower numbers than in control areas.  The decrease coincided with the
    diflubenzuron treatment in the middle of May.  No higher active
    density of species reproducing in late summer or autumn was observed
    in the untreated areas (Klenner, 1990).

         In a forestry spraying programme (0.0672 kg diflubenzuron per
    ha), a survey of microarthropods collected from leaf litter and soil
    in control and sprayed areas of the Bald Eagle State Forest,
    Pennsylvania, USA, showed little or no effect of diflubenzuron on the
    spray plot fauna.  Increases and decreases in both the control and
    spray population sizes during the 6-week study period could seemingly

    be explained on the basis of seasonal population cycles, increased
    soil moisture condition and differential extraction efficiencies
    (White, 1975).

    9.2.3.2  Vertebrates

         Small song birds in a forest ecosystem were unaffected after an
    aerial application of 350 g diflubenzuron/ha in Canada (Buckner et
    al., 1975).

         In a forestry spraying programme (0.0672 kg diflubenzuron/ha)
    surveys were made on birds and small mammals.  Effects on birds were
    assessed using single male mapping surveys.  The survey methodology
    was designed to minimize bias due to differences between observers,
    plots, times of day and abundance of birds.  Vegetation and
    defoliation types were identified and mapped and defoliation was
    sampled at 136 sites.  Four hundred and sixteen surveys were conducted
    on treated plots, defoliated plots, and undisturbed plots.  No
    differences were observed in the number of individuals singing, the
    amount of time each individual spent singing or the song repetition
    rate during song bouts.  The results show that diflubenzuron did not
    lead to the death or emigration of singing birds and probably had no
    direct adverse physiological effects on birds since their vocal
    behaviour changed little, if at all.  The singing male surveys did not
    detect a presumed food shortage among canopy feeders caused by the
    defoliation.  Small mammals studied in this project were censused as
    thoroughly as possible.  The short pre-spray sampling period for the
    treated plot was a handicap in evaluating the pre-spray population
    levels; however, post-spray sampling compensated for this to some
    degree.   Peromyscus, Clethrionomys and  Sorex consume arthropods to
    varying degrees and may be exposed to diflubenzuron present as
    residues.  Diflubenzuron had no demonstrable effects on any of these
    small mammals sampled in this study.  The application of diflubenzuron
    at the rate of 0.067 kg/ha provided good foliage protection within the
    spray sites and apparently made the dying gypsy moth larvae
    unpalatable.  Other contact and stomach pesticides generally do not
    have this effect on caterpillars and are consequently consumed by
    insectivorous mammals.  The reduced exposure by non-ingestion is no
    doubt a beneficial aspect.  None of the species of small mammals
    showed reductions in numbers from pre-spray to post-spray periods that
    could be attributed to the compound.  Stomach analysis did not
    indicate any change in the feeding habits of any of the small mammals
    from either plot in the study.  Changes in reproduction could not be
    detected.  However, of the  Peromyscus and  Clethrionomys animals
    that were dissected and examined, 7 out of 12 females were in a
    healthy condition or had gravid reproductive organs.  Others were
    either virgins or placental scars could not easily be seen, as in
     Sorex.  The body weight of all the small mammals in the study was
    normal for each species.  Juveniles were easily distinguished by
    weight and gonad development.  A regression comparing gonad weights

    and body weights of  Clethrionomys indicated a strong correlation. 
    Since the body weight of small mammals increases with age, this was to
    be expected (White, 1975).

         Diflubenzuron sprayed at 0.14 and 0.28 kg/ha over forests in
    north-eastern Oregon, USA, did not have any detectable adverse effect
    on bird population numbers, nesting or bird behaviour (Richmond et
    al., 1979).

    10.  PREVIOUS EVALUATIONS BY INTERNATIONAL BODIES

         Diflubenzuron was evaluated by the Joint FAO/WHO Meeting on
    Pesticide Residues (JMPR) in 1981, 1984, and 1985 (FAO/WHO, 1982a,b;
    1985a,b; 1986a,b).  In 1985, JMPR established an Acceptable Daily
    Intake (ADI) for human beings of 0-0.02 mg/kg body weight per day,
    based on the fact that the following levels produced no toxicological
    effects:

    rat:    2 mg/kg body weight (40 mg/kg diet)
    mouse:  2.4 mg/kg body weight (16 mg/kg diet)
    dog:    2 mg/kg body weight per day

         Diflubenzuron has been classified as "a product unlikely to
    present an acute hazard in normal use", on the basis of an acute oral
    LD50 for the rat that is greater than 4640 mg/kg body weight (WHO,
    1994).

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    Davies RE, Elliot PH, Street AE, Heywood R, & Prentice DE (1975)
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    De Bree H, De Lange N, Overmars H, & Post LC (1977) Diflubenzuron:
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    De Lange N, Overmans H, Willems AGM, & Post LS (1975) Diflubenzuron
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    De Lange N, Fronik GM, & Post LC (1977) Diflubenzuron: intestinal
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    Deul DH & Jong BJ de (1977) The possible influence of DU 112307 on the
     in vivo synthesis of hyaluronic acid in chicken skin. Weesp, The
    Netherlands, Philips-Duphar BV (Unpublished proprietary report
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    Di Prima SJ (1976) Determination of diflubenzuron residues in soybean
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    Di Prima SJ, Cannizaro RD, Roger JC, & Ferrell CD (1978) Analysis of
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    Solvay Duphar BV, Weesp, The Netherlands).

    Goodman DG (1980b) Histopathologic evaluation of rats administered
    diflubenzuron in the diet. Washington, DC, Clement Associates
    (Unpublished proprietary report submitted to WHO by Solvay Duphar BV,
    Weesp, The Netherlands).

    Gordon R & Cornect M (1986) Toxicity of the insect growth regulator
    diflubenzuron to the rove beetle  Aleochara bilineata a parasitoid
    and predator of the cabbage maggot  Delia radicum Entomol Exp Appl,
    42: 179-185.

    Goto M (1977a) Crop residue analysis of Dimilin. Japan, Institute
    of Pesticide Residues (Unpublished proprietary report
    No. K/Resid./004/1977, submitted to WHO by Solvay Duphar BV, Weesp,
    The Netherlands).

    Goto M (1977b) Crop residue analysis of Dimilin. Japan, Institute of
    Pesticide Residues (Unpublished proprietary report No.
    K/Resid./007/1977, submitted to WHO by Solvay Duphar BV, Weesp, The
    Netherlands).

    Granett J, Morang S, & Hatch R (1978) Reduced movement of precocious
    male Atlantic salmon parr into sublethal Dimilin-G1 and carrier
    concentrations. Bull Environ Contam Toxicol, 19(4): 462-464.

    Graves WC & Swigert JP (1993) Diflubenzuron: A 96-h flow through acute
    toxicity test with the sheepshead minnow ( Cyprinodon variegatus.
    Final report. Wildlife International Ltd. USA (Unpublished proprietary
    report No. 56835/13/93, submitted to WHO by Solvay Duphar BV, Weesp,
    The Netherlands).

    Greenough RJ, Goburdhun R, Hundson R, & MacNaughtan F (1985)
    Diflubenzuron. 52 week oral toxicity study in dogs: Volumes 1 and 2
    (Project No. 630146,2728). Musselburgh, Scotland, Inveresk Research
    International (Unpublished proprietary report No. 56645/32/1985,
    submitted to WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Greenough RJ & McDonald P (1986) Diflubenzuron VC 90 acute inhalation
    toxicity study in rats (limit test). Musselburgh, Scotland, Inveresk
    Research International (Proprietary report No. 56645/41/1986,
    submitted to WHO by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Gulka G, Doscher CM, & Watabe N (1980) Toxicity and molt-accelerating
    effects of diflubenzuron on the barnacle,  Balanus eburneus Bull
    Environ Contam Toxicol, 25: 477-481.

    Gulka G, Gulka CM, & Watabe N (1982) Histopathological effects of
    diflubenzuron on the cirripede crustacean,  Balanus eburneus Arch
    Environ Contam Toxicol, 11: 11-16.

    Gustafson DE & Wargo JP (1976) Fate of Dimilin following application
    to soybeans (ADC Project No 222). Colorado Springs, Colorado,
    Analytical Development Corporation (Unpublished proprietary report
    submitted to WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Hansen SR & Garton RR (1982a) Ability of standard toxicity test to
    predict the effects of the insecticide diflubenzuron on laboratory
    stream communities. Can J Fish Aquat Sci, 39: 1273-1288.

    Hansen SR & Garton RR (1982b) The effects of diflubenzuron on a
    complex laboratory stream community. Arch Environ Contam Toxicol,
    11: 1-10.

    Hatzios KK & Penner D (1978) The effect of diflubenzuron on soybean
    photosynthesis, respiration and leaf ultrastructure. Pestic Biochem
    Physiol, 9: 65-69.

    Hawkins DR, Jackson AJS, & Roberts NL (1980) Excretion of radio-
    activity after oral administration of 3H/14C-diflubenzuron to
    cats (PDR/302/80443). Huntingdon, England, Huntingdon Research Centre
    (Unpublished proprietary report No. 56654/14/1980, submitted to WHO by
    Solvay Duphar BV, Weesp, The Netherlands.

    Helling CS (1985) Soil mobility of three Thompson-Hayward pesticides.
    Interim report. Beltsville, Maryland, US Department of Agriculture,
    Agricultural Research Centre (Unpublished proprietary report No. E-34,
    submitted to WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Hester PG (1982) Efficacy of diflubenzuron on three estuarine decapods
    ( Callinectes sp.,  Palaemonetes pugio and  Uca sp.) (Unpublished
    document submitted to WHO by Solvay Duphar BV, 'sGraveland, The
    Netherlands).

    Hester PG, Olson MA, & Floore TG (1986) Effects of diflubenzuron on
    three estuarine decapods,  Callinectes sp.,  Palaemonetes pugio and
     Uca pugilator J Fla Anti-Mosq Assoc, 57: 8-14.

    Honeycutt R (1993) Indoor occupant exposure study with Dimilin 25 W.
    Colorado Springs, Colorado, Analytical Research and Development
    Corporation (Report No. 56635/56/1992, submitted to WHO by Solvay
    Duphar BV, Weesp, The Netherlands).

    Hsu TS & Bartha R (1974) Interaction of pesticide-derived chloroaniline
    residues with soil organic matter. Soil Sci, 1974: 444-452.

    Huber CM & Collins DL (1987) Final report on the 1987 spray project to
    eradicate Gypsy moth from the Tusquitee ranger district on the
    Nantahala national forest. Asheville, North Carolina, Field Office,
    Forest Pest Management (Unpublished report No. 88-1-4, submitted to
    WHO by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Huber CM & Manchester EH (1988)  Final report on the eradication of
    the Gypsy moth from the Tusquitee ranger district on the Nantahala
    national forest. Forest Pest Management (Unpublished report
    No. 89-1-7, submitted to WHO by Solvay Duphar BV, 'sGraveland, The
    Netherlands).

    Hunter B, Batham P, Street AE, & Cherry CP (1974) DU 112307
    preliminary assessment of the toxicity to male mice in dietary
    administration for 6 weeks. Huntingdon, England, Huntingdon Research
    Centre (Unpublished proprietary report No. PDR/174/74199, submitted to
    WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Hunter B, Batham P, Offer JM, & Prentice DE (1975) Tumorigenicity
    study of DU 112307 to mice. Dietary administration for 80 weeks. Final
    report. Huntingdon, England, Huntingdon Research Centre (Unpublished
    proprietary report No. PDR/170/75685, submitted to WHO by Solvay
    Duphar BV, Weesp, The Netherlands).

    Hunter B, Colley J, Street AE, Heywood R, Prentice DE, & Offer J
    (1976) Effects of DU 112307 in dietary administration to rats for 104
    weeks. Huntingdon, England, Huntingdon Research Centre (Unpublished
    proprietary report No. PDR/171/75945, submitted to WHO by Solvay
    Duphar BV, Weesp, The Netherlands).

    Hunter B, Jordan J, Heywod R, Street AE, Prentice DE, Wight DG, &
    Gibson WA (1979) DU 112307 toxicity to rats in dietary administration
    for nine weeks followed by a four week withdrawal period (final
    report). Huntingdon, England, Huntingdon Research Centre (Unpublished
    report No. PDR/248/77883).

    Ivie GW (1977) Metabolism of insect growth regulators in animals.
    In: Ivie GW & Dorough HW ed. Fate of pesticides in large animals. New
    York, London, Academic Press, pp 111-125.

    Ivie GW (1978) Fate of diflubenzuron in cattle and sheep. J Agric Food
    Chem, 26(1): 81-89.

    Ivie GW, Bull DL, & Veech JA (1980) Fate of diflubenzuron in water.
    J Agric Food Chem, 28(2): 330-337.

    Jackson GA (1976) Dimilin (TH 6040): Biological impact on pond
    organisms. Marion, Alabama, Southeastern Fish Cultural Laboratory
    (Unpublished proprietary report NTP-42, submitted to WHO by Solvay
    Duphar BV, Weesp, The Netherlands).

    Jackson GC, Hardy CJ, Simms F, Mullins PA, & Gopinath Ch (1990)
    Dimilin 4F acute inhalation toxicity study in rats 4-hour exposure.
    Huntingdon, England, Huntingdon Research Centre (Proprietary report
    No. 56645/68/1990, submitted to WHO by Solvay Duphar BV, 'sGraveland,
    The Netherlands).

    Jagannath DR & Brusick DJ (1977a) Mutagenicity evaluation of
    4-chlorophenyl urea, final report. Kensington, Maryland, Litton
    Bionetics Inc. (Unpublished proprietary report No. 56645/42/1977,
    submitted to WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Jagannath DR & Brusick DJ (1977b) Mutagenicity evaluation of
    2,6-difluorobenzoic acid. Final report. Kensington, Maryland, Litton
    Bionetics Inc. (Unpublished proprietary report No. 56645/40/1977,
    submitted to WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Jagannath DR & Brusick DJ (1977c) Mutagenicity evaluation of
    4-chloroanilin. Final report. Kensington, Maryland, Litton Bionetics
    Inc. (Unpublished proprietary report No. 56645/41/1977, submitted to
    WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Janssen PJM & Pot TE (1987a) Primary irritation study of dimilin SC-48
    to the eye of the male rabbit. Weesp, The Netherlands, Solvay Duphar
    BV, Department of Toxicology (Proprietary report No. 56645/46/1987).

    Janssen PJM & Pot TE (1987b) Primary irritation study of dimilin SC-48
    to the skin of the male rabbit. Weesp, The Netherlands, Solvay Duphar
    BV, Department of Toxicology (Proprietary report No. 56645/45/1987).

    Janssen PJM & Pot TE (1987c) Acute oral toxicity study with dimilin
    SC-48 in rats. Weesp, The Netherlands, Solvay Duphar BV, Department of
    Toxicology (Proprietary report No. 56645/44/1987).

    Janssen PJM & Pot TE (1987d) Acute dermal toxicity study with dimilin
    SC-48 in rats. Weesp, The Netherlands, Solvay Duphar BV, Department of
    Toxicology (Proprietary report No. 56645/43/1987).

    Janssen PJM & Pot TE (1987e) Acute dermal toxicity study with dimilin
    WP-25 in rats. Weesp, The Netherlands, Solvay Duphar BV, Department of
    Toxicology (Proprietary report No. 56645/58/1987).

    Janssen PJM & van Doorn WM (1993a) Acute inhalation toxicity study
    with dimilin OF-6 in male and female rats. Weesp, The Netherlands,
    Solvay Duphar BV, Department of Toxicology (Proprietary report
    No. 56345/01/1993).

    Janssen PJM & van Doorn WM (1993b) Primary irritation study of dimilin
    OF-6 to the skin of female rabbits. Weesp, The Netherlands,
    Solvay Duphar BV, Department of Toxicology (Proprietary report
    No. 56345/30/1993).

    Janssen PJM & van Doorn WM (1993c) Primary irritation study of dimilin
    OF-6 to the eye of female rabbits. Weesp, The Netherlands, Solvay
    Duphar BV, Department of Toxicology (Proprietary report
    No. 56345/31/1993).

    Jenkins VK, Mayer RT, & Perry RR (1984) Effects of diflubenzuron on
    growth of malignant melanoma and skin carcinoma tumors in mice. Invest
    New Drugs, 2: 19-27.

    Jenkins VK, Perry RR, Ahmer AE, & Ives K (1986) Role of metabolism in
    effects of diflubenzuron on growth of B16 melanomas in mice. Invest
    New Drugs, 4: 325-335.

    Jones AS & Konchenderfer JN (1988) Persistence of diflubenzuron
    (Dimilin) in a small eastern watershed and its impact on invertebrates
    in a headwater stream. Research Triangle Park, North Carolina,
    Southeastern Forest Experiment Station (Unpublished proprietary report
    No. 56637/26/1988, submitted to WHO by Solvay Duphar BV, Weesp, The
    Netherlands).

    Joustra KD, van Kampen WG, & Walstra P (1989) Metabolism of [14C]
    diflubenzuron in citrus fruit (Analytical report). Weesp, The
    Netherlands, Duphar BV, Analytical Development Department (Proprietary
    report No. 56630/86/1989).

    Julin AM & Sanders HO (1978) Toxicity of IGR, diflubenzuron, to
    freshwater intervertebrates and fishes. Mosq News, 38: 256-259.

    Kavanagh P (1988a) Diflubenzuron oral (gavage) rat teratology limit
    study. Ledbury, England, Toxicol Laboratories Ltd. (PHD/11/87)
    (Proprietary report No. 56645/68/1987, submitted to WHO by Solvay
    Duphar BV, 'sGraveland, The Netherlands).

    Kavanagh P (1988b) Diflubenzuron oral (gavage) rabbit teratology limit
    study. Ledbury, England, Toxicol Laboratories Ltd. (PHD/12/87)
    (Proprietary report No. 56645/79/1987, submitted to WHO by Solvay
    Duphar BV, 'sGraveland, The Netherlands).

    Keet CMJF (1977a) The methaemoglobin, sulphaemoglobin and Heinz body
    forming properties of DU 112307 after oral administration to male rats
    during 8 days. Weesp, The Netherlands, Duphar BV (Unpublished
    proprietary report No. 56645/15/1977).

    Keet CMJF (1977b) The effect of DU 112307 (tech) in male mice after
    daily oral administration (14-days) on body weight, methaemoglobin,
    sulphaemoglobin and Heinz body formation. Weesp, The Netherlands,
    Duphar BV (Unpublished proprietary report No. 56645/33/1977).

    Keet CMJF (1977c) The methaemoglobin and sulphaemoglobin forming
    properties of DU 112307 in male rabbits after prolonged dietary and
    dermal administration. Weesp, The Netherlands, Duphar BV (Unpublished
    proprietary report No. 56645/02/1977).

    Kemp A, van der Heijden CA, & van Eldik A (1973a) Dietary
    administration of DU 112307 to male and female rats for three months.
    Weesp, The Netherlands, Philips-Duphar BV (Unpublished proprietary
    report No. 56645/13A/1973).

    Kemp A, van der Heijden CA, & van Eldik A (1973b) Appendix III to
    report No. 56645/13A/1973 individual data: dietary administration of
    DU 112307 to male and female rats for 3 months. Weesp, The
    Netherlands, Philips-Duphar BV (Unpublished proprietary report
    No. 56645/13B/1973).

    Kingsbury P, Sundaram KMS, Holmers KMS, Nott R, & Kreutzweiser D
    (1987) Aquatic fate and impact studies with Dimilin. Ottawa, Ontario,
    Forest Pest Management Institute (Unpublished report No. 78, submitted
    to WHO by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Klenner MF (1990) [A comparative study of the carabid fauna in the
    Dimilin-treated and untreated oak stands in Westphalia.] Mitt BBA
    (Berlin-Dahlem), 266: 279 (in German).

    Koelman-Klaus HJS (1978a) Acute oral toxicity study of
    4-chlorophenylurea in male and female rats. Weesp, The Netherlands,
    Duphar BV (Unpublished proprietary report No. 56645/09/1978).

    Koelman-Klaus HJS (1978b) Acute oral toxicity study of
    2,6-difluorobenzoic acid in male and female rats. Weesp, The
    Netherlands, Duphar BV (Unpublished proprietary report
    No. 56645/25/1978).

    Koopman TSM (1977a) Acute oral toxicity study with DU 112307 technical
    in mice. 'sGraveland, The Netherlands, Philips-Duphar BV, Department
    of Toxicology (Unpublished proprietary report No. 56645/04/1977).

    Koopman TSM (1977b) Acute oral toxicity study with DU 112307 WP 25%
    in mice and rats. 'sGraveland, The Netherlands, Philips-Duphar
    BV, Department of Toxicology (Unpublished proprietary report
    No. 56645/03/1977).

    Koopman TSM (1977c) Acute dermal toxicity study with DU 112307
    technical in rats. 'sGraveland, The Netherlands, Philips-Duphar
    BV, Department of Toxicology (Unpublished proprietary report
    No. 56645/07/1977).

    Koopman TSM (1980a) Primary irritation of Dimilin ODC-45 to the rabbit
    eye. 'sGraveland, The Netherlands, Duphar BV, Department of Toxicology
    (Unpublished proprietary report No. 56645/02/1980).

    Koopman TSM (1980b) Acute dermal toxicity study of dimilin ODC-45 in
    rats. 'sGraveland, The Netherlands, Duphar BV, Department of
    Toxicology (Unpublished proprietary report No. 56645/04/1980).

    Koopman TSM (1980c) Primary irritation of Dimilin ODC-45 to the rabbit
    skin. 'sGraveland, The Netherlands, Duphar BV, Department of
    Toxicology (Unpublished proprietary report No. 56645/03/1980).

    Koopman TSM (1984a) Acute dermal toxicity study with diflubenzuron
    VC-90 in rats. 'sGraveland, The Netherlands, Duphar BV, Department of
    Toxicology (Unpublished proprietary report No. 56645/31/1984).

    Koopman TSM (1984b) Acute oral toxicity study with diflubenzuron VC-90
    in rats. 'sGraveland, The Netherlands, Duphar BV, Department of
    Toxicology (Unpublished proprietary report No. 56645/30/1984).

    Koopman TSM (1984c) Primary irritation study of diflubenzuron VC-90 to
    the rabbit eye. 'sGraveland, The Netherlands, Duphar BV, Department of
    Toxicology (Unpublished proprietary report No. 56645/29/1984).

    Koopman TSM (1984d) Primary irritation study of diflubenzuron VC-90 to
    the rabbit skin. 'sGraveland, The Netherlands, Duphar BV, Department
    of Toxicology (Unpublished proprietary report No. 56645/44/1984).

    Koopman TSM (1985a) Primary irritation study of dimilin 2F to the
    skin of the male rabbit. 'sGraveland, The Netherlands, Duphar BV,
    Department of Toxicology (Unpublished proprietary report
    No. 56645/93/1985).

    Koopman TSM (1985b) Primary irritation study of dimilin 2F to
    the eye of the male rabbit. 'sGraveland, The Netherlands,
    Duphar BV, Department of Toxicology (Unpublished proprietary report
    No. 56645/91/1985).

    Koopman TSM (1985c) Acute dermal toxicity study with dimilin 2F in
    rats. 'sGraveland, The Netherlands, Duphar BV, Department of
    Toxicology (Unpublished proprietary report No. 56645/95/1985).

    Koopman TSM (1985d) Acute oral toxicity study with dimilin 2F in rats.
    'sGraveland, The Netherlands, Duphar BV, Department of Toxicology
    (Unpublished proprietary report No. 56645/94/1985).

    Koopman TSM & Jongeling AJ (1979) Acute oral toxicity study with
    Dimilin ODC 45% in rats. 'sGraveland, The Netherlands, Duphar BV,
    Department of Toxicology (Unpublished proprietary report No.
    56645/38/1979).

    Koopman TSM & Pot TE (1986) Acute oral toxicity study with
    diflubenzuron VC-90% in mice. 'sGraveland, The Netherlands, Duphar
    BV, Department of Toxicology (Unpublished proprietary report
    No. 56645/39/1986).

    Koorn JC (1990) Study to examine the possible mutagenic activity of
    diflubenzuron in the Ames  Salmonella microsome assay. 'sGraveland,
    The Netherlands, Duphar BV, Department of Toxicology (Proprietary
    report No. 56645/74/1990).

    Kramer HT (1990) Determination of diflubenzuron and the metabolites
    4-chlorophenyl urea and 2,6-difluorobenzoic acid in soil from a
    dissipation trial in an apple orchard in Phelps (New York). Madison,
    Wisconsin, Hazleton Laboratories (Proprietary report No. HLA-6012-271,
    submitted to WHO by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Kramer HT (1991) Determination of diflubenzuron and the metabolites
    4-chlorophenyl urea and 2,6-difluorobenzoic acid in soil from a
    dissipation trial in a citrus orchard in Oviedo (Florida). Madison,
    Wisconsin, Hazleton Laboratories (Proprietary report No. HLA-6012-270,
    submitted to WHO by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Kramer HT (1992a) Field dissipation study with diflubenzuron
    insecticide applied for control of insects on cotton (a bare soil
    application in Arkansas). Madison, Wisconsin, Hazleton Laboratories
    (Proprietary report No. HLA-6248-110, submitted to WHO by Solvay
    Duphar BV, 'sGraveland, The Netherlands).

    Kramer HT (1992b) Field dissipation study with diflubenzuron
    insecticide applied for control of insects on soybeans (a bare soil
    application in Louisiana). Madison, Wisconsin, Hazleton Laboratories
    (Proprietary report No. HLA-6248-116, submitted to WHO by Solvay
    Duphar BV, 'sGraveland, The Netherlands).

    Kubena LF (1982) The influence of diflubenzuron on several
    reproductive characteristics in male and female layer-breed chickens.
    Poult Sci, 61: 268-271.

    Kuijpers LAM (1988) The acute toxicity of diflubenzuron to  Daphnia
     magna Weesp, The Netherlands, Duphar BV (Unpublished proprietary
    report No. 56635/26/1988).

    Kurczewski F, Wang C, Grimble D, Smith R, & Brezner J (1975)
    Environmental impact of dimilin. A final report: Effects of dimilin
    upon microorganisms in leaf litter and forest soil. Syracuse, New
    York, State University of New York, pp 28-43.

    Kynoch SR & Elliot PH (1978a) Screening test for delayed contact
    hypersensitivity with Dimilin WP 25%. Huntingdon, England, Huntingdon
    Research Centre (Proprietary report No. 0911/D258/78, submitted to WHO
    by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Kynoch SR & Elliot PH (1978b) Screening test for delayed contact
    hypersensitivity with dummy formulation for dimilin WP 25% in the
    albino guinea-pig. Huntingdon, England, Huntingdon Research Centre
    (Proprietary report No. 9112/D258/78, submitted to WHO by Solvay
    Duphar BV, 'sGraveland, The Netherlands).

    Kynoch SR & Parcell BI (1987) Delayed contact hypersensitivity in the
    guinea-pig with Dimilin 2F. Huntingdon, England, Huntingdon Research
    Centre (Proprietary report No. 56645/13/1987, submitted to WHO by
    Solvay Duphar BV, 'sGraveland, The Netherlands).

    Kynoch SR & Parcell BI (1987) Delayed contact hypersensitivity in the
    guinea-pig with Dimilin SC-48. Huntingdon, England, Huntingdon
    Research Centre (Proprietary report No. 56645/72/1987, submitted to
    WHO by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Kynoch SR & Smith PA (1986) Delayed contact hypersensitivity in the
    guinea-pig with diflubenzuron VC-90. Huntingdon, England, Huntingdon
    Research Centre (Proprietary report No. 86558D/PDR 432SS, submitted to
    WHO by Solvay Duphar BV, 'sGraveland, The Netherlands).

    Lauren DR, Agnew MP, & Henzell RF (1984) Field life of the insect
    growth regulator diflubenzuron on lucerne. N Z J Agric Res,
    27: 425-429.

    Lawrence JF & Sundaram KMS (1976) Gas-liquid chromatographic analysis
    of N-(4-chlorophenyl-Nœ-2,6-difluorobenzoyl) urea insecticide after
    chemical derivatization. J AOAC, 59(4): 938-941.

    Lee BM & Scott GI (1989) Acute toxicity of temephos, fenoxycarb,
    diflubenzuron and methoprene and  Bacillus thuringiensis var.
    Israelensis to the mummichug ( Fundulus heteroclitus. Bull Environ
    Contam Toxicol, 43: 827-832.

    Maas W, Van Hes R, Grosscurt AC, & Deul DH (1980) [Benzoylphenylurea
    insecticides.] In: Wegler R ed. [Chemistry of plant protection
    chemicals and pesticides.] Berlin, Heidelberg, Springer Verlag, vol 6,
    pp 432-470 (in German).

    McAlonan WG (1976) Effects of two insect growth regulators on some
    selected salt-marsh non-target organisms. University of Delaware, USA
    (Thesis) (NTP-53).

    Machado J, Coimbra J, Castilho F, & Sa C (1990) Effects of
    diflubenzuron on shell formation of the freshwater clam,  Anodonta
     cygnea Arch Environ Contam Toxicol, 19: 35-39.

    McGregor JT, Gould DH, Mitchell AD, & Sterling GP (1979) Mutagenicity
    tests of diflubenzuron in the micronucleus test in mice, the L5178Y
    mouse lymphoma forward mutation assay, and the Ames  Salmonella
    reverse mutation test. Mutat Res, 66: 45-53.

    McKague AB & Pridmore RB (1978) Toxicity of Altosid and Dimilin to
    juvenile rainbow trout and coho salmon. Bull Environ Contam Toxicol,
    20: 167-169.

    Madder DJ (1977) The disappearance from, efficacy in and effect on
    non-target organisms of diflubenzuron, methoprene and chlorpyrifos in
    a lentic ecosystem. Canada, University of Manitoba, Faculty of
    Graduate Studies, 139 pp (Thesis) (NTP-71).

    Madder DJ & Lockhart WL (1978) A preliminary study of the effects of
    diflubenzuron and methoprene on rainbow trout ( Salmo gairdneri. Bull
    Environ Contam Toxicol, 20: 66-70.

    Madder DJ & Lockhart WL (1980) Studies on the dissipation of
    diflubenzuron and methoprene from shallow prairie pools. Can Entomol,
    112: 173-177.

    Mansager ER, Still GC, & Frear DS (1979) Fate of 14C diflubenzuron on
    cotton and in soil. Pestic Biochem Physiol, 12: 172-182.

    Marshall BF & Hieb BF (1973) 96-h LC50  Salmo gairdneri (n.m.
     Oncorhynchus mykiss, Lepomis machrochirus and  Fundulus heteroclitus
    Marine Research Institute (Unpublished study submitted to WHO by Solvay
    Duphar BV, Weesp, The Netherlands).

    Martinez-Toledo MV, Rubia T, De La Moreno J, & Gonzalez-Lopez J
    (1988a) Effect of diflubenzuron on azotobacter nitrogen fixation in
    soil. Chemosphere, 17(4): 829-834.

    Martinez-Toledo MV, Gonzalez-Lopez J, Rubia T, De La Moreno J, &
    Ramos-Cormenzana A (1988b) Diflubenzuron and the acetylene-reduction
    activity of  Azotobacter vinelandii Soil Biol Biochem,
    20(2): 255-256.

    Matheson DW & Brusick DJ (1977a) Evaluation of 4-chlorophenylurea
     in vitromalignant transformation in BALB/3T3 cells. Kensington,
    Maryland, Litton Bionetics Inc. (Unpublished proprietary report
    submitted to WHO by Solvay Duphar BV, Weesp, The Netherlands).

    Matheson DW & Brucisk DJ (1977b) Evaluation of 2,6-diflurobenzoic acid
     in vitromalignant transformation in BALB/3T3 cells. Final report.
    Kensington, Maryland, Litton Bionetics Inc. (Unpublished proprietary
    report No. 56645/10/1978, submitted to WHO by Solvay Duphar BV, Weesp,
    The Netherlands).

    Matheson DW & Brusick DJ (1978a) Mutagenicity evaluation of
    2,6-difluorobenzoic acid in unscheduled DNA synthesis in the human WI-
    38 cells assay. Kensington, Maryland, Litton Bionetics Inc.
    (Unpublished proprietary report No. 56645/16/1978, submitted to WHO by
    Solvay Duphar BV, Weesp, The Netherlands).

    Matheson DW & Brusick DJ (1978b) Mutagenicity evaluation of
    4-chlorophenylurea in unscheduled DNA synthesis in the human WI-38
    cells assay. Final report. Kensington, Maryland, Litton Bionetics Inc.
    (Unpublished proprietary report No. 56645/24/1978, submitted to WHO by
    Solvay Duphar BV, Weesp, The Netherlands).

    Matheson DW & Brusick DJ (1978c) Mutagenicity evaluation of
    4-chloroanilin in the  in vitro transformation of BALB/3T3 cells
    assay. Final report. Kensington, Maryland, Litton Bionetics Inc.
    (Unpublished proprietary report No. 56645/23/1978, submitted to WHO by
    Solvay Duphar BV, Weesp, The Netherlands).

    Matta JF (1976) The effect of dimilin on non-target organisms in a
    marsh community. Norfolk, Virginia, Environmental Consultants (Report
    prepared for Thompson-Hayward Chemical Company) (Unpublished
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    RESUME

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

         Le diflubenzuron appartient au groupe des insecticides dérivés de
    la benzoylphénylurée.  Son activité  insecticide est due à une
    interaction avec la synthèse et le dépôt de la chitine.  Il forme des
    cristaux blancs inodores dont le point de fusion est de 230-232°C.
    Il est légèrement soluble dans l'eau (0,2 mg/litre à 20°C)
    et pratiquement non volatil.  Il est relativement stable en milieu
    acide ou neutre mais s'hydrolyse en milieu alcalin.

         Le diflubenzuron s'obtient par réaction du 2,6-difluoro-
    benzamide sur l'isocyanate de 4-chlorophényle.

         Le dosage des résidus de diflubenzuron présents dans l'eau, les
    échantillons biologiques,et le sol peut s'effectuer par
    chromatographie liquide à haute performance avec détection par UV ou
    encore par chromatographie en phase gazeuse avec détection par capture
    d'électrons, soit directement sur la molécule initiale, soit sur un
    dérivé (libération de 4-chloroaniline et action de l'anhydride
    trifluoracétique).

    2.  Sources d'exposition humaine et environnementale

         Le diflubenzuron est un produit de synthèse utilisé en
    agriculture, en foresterie et dans les programmes de santé publique
    pour détruire les ravageurs et les vecteurs de maladies.  Différentes
    formulations existent à cet usage.  On ne possède pas de
    renseignements au sujet de cas d'exposition humaine au diflubenzuron
    qui auraient pu se produire.

    3.  Transport, distribution et transformation dans l'environnement

         En général, le diflubenzuron est appliqué directement sur les
    végétaux et les eaux à traiter.  Le feuillage ne constitue pas une
    porte d'entrée dans la plante.

         Le diflubenzuron est rapidement adsorbé sur les particules du
    sol.  Il reste fixé dans les 10 premiers cm du sol sur lequel il est
    épandu.  Il est peu probable qu'il subisse un lessivage.  Dans divers
    types de sol, il subit une décomposition aérobie ou anaérobie avec une
    demi-vie de quelques jours.  La vitesse de décomposition dépend
    largement de la taille des particules de diflubenzuron.  La principale
    voie métabolique (plus de 90%) esst l'hydrolyse qui conduit à la
    formation d'acide 2,6-difluorobenzoïque et de 4-chlorophénylurée; ces
    deux composés sont dégradés à leur tour, respectivement en 4 et 6
    semaines.  On n'a pas décelé la présence de 4-chloroaniline libre
    dans le sol.

         Le diflubenzuron se décompose rapidement dans les eaux neutres ou
    alcalines.  On constate qu'une fois épandu sur l'eau, il se répartit
    rapidement entre celle-ci et les sédiments.Le composé initial et la 
    4-chlorophénylurée peuvent persister plus de 30 jours dans les
    sédiments.

         Le diflubenzuron ne s'accumule pas chez les poissons.

    4.  Concentrations dans l'environnement et exposition humaine

         L'utilisation en agriculture, en foresterie ou pour la
    démoustication n'entraîne qu'une exposition négligeable de la
    population générale par l'intermédiaire de la nourriture ou de l'eau.

    5.  Cinétique et métabolisme chez les animaux de laboratoire

         Chez l'animal de laboratoire, le diflubenzuron est absorbé au
    niveau des voies digestives et, à un moindre degré, au niveau cutané. 
    Chez le rat, il existe un mécanisme d'absorption saturable.  Dans ces
    conditions, une forte proportion du diflubenzuron administré par voie
    orale se retrouve dans les matières fécales.  Le diflubenzuron se
    répartit largement dans les tissus, mais il ne s'y accumule pas.

         Le métabolisme du diflubenzuron a été étudié chez diverses
    espèces animales. Chez les mammifères, la principale voie métabolique
    comporte une hydroxylation.  L'hydrolyse peut se produire au niveau de
    l'une quelconque des trois liaisons carbone-azote.  Chez le porc et le
    poulet, l'hydrolyse s'effectue principalement au niveau du pont
    uréido.  Chez le rat et la vache, l'hydroxylation constitue la
    principale voie métabolique.  Chez le mouton, le porc et le poulet,
    les principaux métabolites sont le 2,6-difluorobenzamide et la 
    4-chlorophénylurée; on trouve aussi, en moindre proportion, du
    2,6-difluorobenzamide et de la 4-chloroaniline.  Chez le rat et les
    bovins, 80% des métabolites sont constitués de 2,6-difluoro-3-
    hydroxydiflubenzuron, de 4-chloro-2-hydroxydifluorobenzuron et de 
    4-chloro-3-hydroxydiflubenzuron.  Les études de métabolisme indiquent
    qu'il ne se forme pratiquement pas de 4-chloroaniline chez le rat et
    les bovins.

          Chez les chats, les porcs et les bovins, l'élimination
    s'effectue principalement par la voie fécale, à hauteur de 70 à 85%. 
    Chez les ovins, la voie urinaire et la voie fécale ont à peu près la
    même importance de ce point de vue.  Chez  le rat et la souris,
    l'excrétion urinaire décroît proportionnellement à l'augmentation de
    la dose.  Moins de 1% de la dose administrée par la voie orale se
    retrouve dans l'air expiré.  Le diflubenzuron n'est présent qu'à
    l'état de résidus dans le lait.

         Il n'existe pas d'étude sur la cinétique et le métabolisme du
    diflubenzuron chez l'homme et notamment, sur son degré de
    biotransformation en 4-chloroaniline.

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

         Quel que soit le mode d'exposition, le diflubenzuron présente une
    faible toxicité aiguë.  En se basant sur le fait que sa DL50 aiguë
    par voie orale est supérieure à 4640 mg/kg de poids corporel chez le
    rat, l'OMS estime qu'il s'agit d'un produit qui ne présente
    vraisemblablement pas de risque d'intoxication aiguë en utilisation
    normale.  Chez ce même animal, la DL50 aiguë par voie percutanée est
    supérieure à 10 000 mg/kg de poids corporel et la CL50 dépasse 2,49
    mg/litre par la voie respiratoire.  Au cours d'une période de deux
    semaines pendant laquelle diverses espèces animales avaient reçu du
    diflubenzuron en une seule prise et selon divers modes
    d'administration, on n'a constaté aucun signe d'intoxication.

         Le diflubenzuron n'est pas irritant pour la peau (chez le lapin)
    et ne provoque pas non plus de sensibilisation cutanée (chez le
    cobaye).  Il est légèrement irritant pour la muqueuse oculaire chez le
    lapin.

         Le diflubenzuron provoque une méthémoglobinémie et une
    sulfhémoglobinémie.  Une méthémoglobinémie liée à la dose a été mise
    en évidence après exposition d'animaux de diverses espèces au
    diflubenzuron par la voie orale, percutanée ou respiratoire.  Cet
    effet constitue le point d'aboutissement toxicologique le plus
    sensible chez les animaux de laboratoire.  En prenant comme critère la
    méthémoglobinémie, la dose sans effet observable est de 2 mg/kg de
    poids corporel par jour chez les rats et les chiens et de
    2,4 mg/kg de poids corporel par jour chez les souris.  Les études de
    toxicité à long terme effectuées sur des souris et des rats
    ont montré que les modifications imputables au traitement
    correspondaient principalement à l'oxydation de l'hémoglobine et à une
    altération des hépatocytes.

         Des études de cancérogénicité effectuées sur des rats et des
    souris à des doses allant jusqu'à 10 000 mg/kg de nourriture, n'ont
    pas révélé de modification de l'incidence tumorale qui soit imputable
    au traitement.  En particulier, on n'a pas observé de néoplasmes au
    niveau du mésenchyme splénique ou hépatique lors d'études de
    cancérogénicité utilisant de la 4-chloroaniline.

         Plusieurs études toxicologiques portant sur la fonction de
    reproduction ont été menées sur des rats, des souris, des lapins et
    trois espèces d'oiseaux; elles n'ont pas mis en évidence d'effets
    pathogènes et le produit ne s'est pas révélé embryotoxique.  Les
    études de tératogénicité effectuées sur des rats et des lapins se sont
    également révélées négatives.

          Le diflubenzuron et ses principaux métabolites ont également été
    soumis à diverses épreuves de mutagénicité  in vivo e  in vitro 
    Ni le diflubenzuron ni ses principaux métabolites n'ont donné de
    résultat positif dans ces épreuves.

         Le métabolite secondaire, c'est-à-dire la 4-chloroaniline,
    a donné des résultats positifs dans plusieurs épreuves de mutagénicité
     in vitro portant sur divers points d'aboutissement toxicologiques.
    Il est cancérogène pour le rat et la souris.  Les tumeurs imputables à
    l'administration de 4-chloroaniline se sont révélées bénignes; quant
    aux tumeurs malignes observées, il s'agissait de tumeurs du mésenchyme
    splénique chez les rats mâles ainsi que d'hémangiomes et
    d'hémangiosarcomes spléniques ou hépatiques chez les souris mâles.

    7.  Effets sur l'homme

         On a signalé des cas de méthémoglobinémie chez des travailleurs
    exposés de par leur profession et des nouveau-nés exposés par
    inadvertance à de la 4-chloroaniline, un métabolite secondaire du
    diflubenzuron.  Les sujets qui présentent un déficit en NADH-
    méthémoglobine-réductase peuvent être particulièrement sensibles à la
    4-chloroaniline et par conséquent à une exposition au diflubenzuron.

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

         Tous les organismes qui synthétisent la chitine sont sensibles au
    diflubenzuron.

         A la concentration de 500 mg/kg de terre, les bactéries n'ont pas
    souffert d'une exposition au diflubenzuron.  Il y a eu une certaine
    stimulation de la fixation d'azote.  Les bactéries décomposent les
    solutions de diflubenzuron dans l'acétone, solvant qu'elles utilisent
    comme source de carbone.  Une concentration de diflubenzuron de 1
    µg/litre a provoqué un accroissement de la biomasse algaire.  Aucun
    effet nocif n'a été observé à des concentration supérieures à la
    limite de solubilité du diflubenzuron.  Des champignons placés dans un
    courant créé en laboratoire ont été temporairement affectés à la
    concentration de 0,1 µg/litre.

         Les invertébrés aquatiques présentent des réactions variées au
    diflubenzuron.  Les mollusques n'y sont pas sensibles, avec une CL50
    supérieure à 200 mg/litre.  Chez les autres invertébrés, la CL50 peut
    aller de 1 à > 1000 µg/litre, ce qui peut refléter la sensibilité de
    ces organismes au moment de la mue.  On estime que pour la daphnie, la
    concentration tolérable  maximale en produit toxique est supérieure à
    40 ng/litre et inférieure à 93 ng/litre.  Comme prévu, les larves
    d'éphémères et autres formes pré-imaginales d'insectes divers, sont
    très sesnsibles au diflubenzuron.  Le traitement des eaux de surface
    par le diflubenzuron est donc probablement susceptible de causer une
    certaine mortalité parmi les insectes aquatiques.

         Lors de traitements expérimentaux effectués sur le terrain et
    dans divers écosystèmes, on a constaté que la plupart des organismes
    avaient moins souffert que ne le laissaient prévoir les études
    toxicologiques en laboratoire.  Aucun effet n'a été constaté sur les
    organismes aquatiques après traitement de forêts par voie aérienne.

         Pour les poissons, la CL50 est supérieure à 150 mg/litre.  Les
    essais effectués sur le terrain n'ont enregistré aucune mortalité chez
    les poissons.

         La DL50 par voie orale et par contact est supérieure à 30 µg par
    insecte chez l'abeille mellifique.  Après épandage de diflubenzuron
    par voie aérienne à raison de 350 g/ha, on n'a pas constaté de dommage
    parmi les colonies d'abeilles des alentours.

         Une étude alimentaire de 5 jours sur des colverts et des
    gallinacés du genre colin avec des doses allant jusqu'à 4640 mg/kg de
    nourriture, n'a pas révélé de signes de toxicité.  Après épandage de
    diflubenzuron par voie aérienne sur des forêts à raison de 350 g/ha,
    on n'a pas constaté de dommages parmi les oiseaux chanteurs de
    l'écosystème forestier.

         Après épandage de diflubenzuron à raison de 67 g/ha sur une
    forêt, on n'a pas observé de réduction dans l'effectif des populations
    de petits mammifères.

    RESUMEN

    1.  Identidad, propiedades físicas y químicas y métodos analíticos

         El diflubenzurón pertenece al grupo de los insecticidas
    derivados de la benzoilfenilurea.  Su acción insecticida se debe
    a la interacción con la síntesis y/o deposición de quitina.  Forma
    cristales blancos inodoros con un punto de fusión de 230-232°C.  Es
    bastante soluble en agua (0,2 mg/litro a 20°C) y prácticamente
    involátil.  Es relativamente estable en medios ácidos y neutros, pero
    se hidroliza en condiciones alcalinas.

         El diflubenzurón se produce haciendo reaccionar 2,6-difluoro-
    benzamida con 4-clorofenilisocianato.

         Los residuos de diflubenzurón pueden medirse en el agua, en
    muestras biológicas y en el suelo mediante cromatografía líquida de
    alta resolución con detección de radiación ultravioleta o mediante
    cromatografía de gases con detector de captura de electrones para el
    análisis de la molécula intacta o tras la derivatización de la
    4-cloroanilina liberada con anhídrido trifluoracético.

    2.  Fuentes de exposición humana y ambiental

         El diflubenzurón es un compuesto sintético utilizado en la
    agricultura, en la silvicultura y en programas de salud pública para
    combatir plagas de insectos y vectores.  Para esos usos existen
    diferentes formulaciones de diflubenzurón.  No se dispone de
    información pertinente sobre la exposición humana a este producto.

    3.  Transporte, distribución y transformación en el medio ambiente

         El diflubenzurón suele aplicarse directamente a las plantas y al
    agua.  No se produce absorción a través de las hojas.

         La adsorción del diflubenzurón en el suelo es rápida.  El
    producto se inmoviliza en la capa superior de 10 cm del suelo al que
    se aplica, y las probabilidades de lixiviación son escasas.  El
    diflubenzurón se degrada en suelos de diversos tipos y orígenes en
    condiciones aerobias o anaerobias, con una semivida de pocos días.  La
    velocidad de degradación depende en gran medida del tamaño de las
    partículas de diflubenzurón.  La principal ruta metabólica (más del
    90%) es la hidrólisis, que produce 2,6-ácido difluorobenzoico y
    4-clorofenilurea; estos productos se degradan con semividas del orden
    de cuatro y seis semanas, respectivamente.  No se ha detectado
    4-cloroanilina libre en los suelos.

         El diflubenzurón se degrada rápidamente en aguas neutras o
    alcalinas.  Estudios de aplicación al agua revelan que el
    diflubenzurón se concentra rápidamente en el sedimento; el compuesto
    de origen y la 4-clorofenilurea pueden persistir en el sedimento por
    más de 30 días.

         El diflubenzurón no es objeto de bioacumulación en los peces.

    4.  Niveles ambientales y exposición humana

         La exposición de la población general al diflubenzurón por medio
    del agua o los alimentos de resultas de su utilización en la
    agricultura, contra insectos forestales o en la lucha contra los
    mosquitos, es insignificante.

    5.  Cinética y metabolismo en animales de laboratorio

         En animales de experimentación, el diflubenzurón se absorbe en el
    tubo digestivo y, en menor medida, a través de la piel.  En el tubo
    digestivo de la rata existe un mecanismo de absorción saturable, por
    lo que una gran proporción del diflubenzurón administrado oralmente
    aparece en las heces.  El diflubenzurón tiene una amplia distribución
    en los tejidos, pero no se acumula.

         El destino metabólico del diflubenzurón ha sido estudiado en
    diversas especies.  La principal vía metabólica en los mamíferos es la
    hidroxilación.  La hidrólisis del diflubenzurón puede producirse en
    cualquiera de los tres enlaces carbono-nitrógeno.  En los cerdos y los
    pollos, la principal ruta de hidrólisis es el puente ureido.  En las
    ratas y las vacas, la principal vía metabólica es la hidroxilación. 
    En las ovejas, los cerdos y los pollos, los metabolitos más
    importantes son el 2,6-ácido difluorobenzoico y la 4-clorofenilurea;
    los metabolitos secundarios son la 2,6-difluorobenzamida y la
    4-cloroanilina.  En las ratas y el ganado vacuno, el 80% de los
    metabolitos está constituido por 2,6-difluoro-3-hidroxidiflubenzurón,
    4-cloro-2-hidroxi-diflubenzurón y 4-cloro-3-hidroxidiflubenzurón.  Los
    estudios metabólicos indican que en las ratas o el ganado vacuno se
    forman cantidades mínimas o nulas de 4-cloroanilina.

         La principal ruta de eliminación es a través de las heces, con
    porcentajes de entre el 70 y el 85% en los gatos, los cerdos y el
    ganado vacuno.  En el ganado ovino, la eliminación se distribuye
    aproximadamente por igual entre la orina y las heces.  En las ratas y
    ratones, la excreción urinaria disminuye proporcionalmente al aumento
    de la dosis.  Menos del 1% de una dosis oral se recupera en el aire
    exhalado.  En la leche sólo se han hallado residuos ínfimos.

         No se dispone de ningún estudio humano de la cinética y el
    metabolismo del diflubenzurón, incluido el alcance de la
    biotransformación en 4-cloroanilina.

    6.  Efectos en mamíferos de laboratorio y en sistemas de pruebas
        in vitro

         El diflubenzurón tiene una toxicidad aguda baja por cualquier vía
    de exposición.  La OMS lo ha clasificado como producto con pocas
    probabilidades de presentar un riesgo agudo en el uso normal, sobre la
    base de una DL50 aguda por vía oral de más de 4640 mg/kg de peso
    corporal en las ratas.  La DL50 aguda por vía cutánea en las ratas es
    superior a 10 000 mg/kg de peso corporal, mientras que la CL50 aguda
    por inhalación en las ratas excede de 2,49 mg/litro.  No se han
    observado signos de intoxicación en los 14 días siguientes a una
    administración única de diflubenzurón por diversas rutas a una
    variedad de especies animales.

         El diflubenzurón no provoca irritación cutánea (en el conejo) ni
    sensibilización de la piel (en el cobayo).  Produce una ligera
    irritación a los ojos en el conejo.

         El diflubenzurón causa metahemoglobinemia y sulfohemoglobinemia. 
    Tras la exposición oral, cutánea o por inhalación de diversas especies
    al diflubenzurón se ha demostrado la presencia de metahemoglobinemia
    dosisdependiente.  Este efecto es la variable de evaluación
    toxicológica más sensible en los animales de experimentación.  El NOEL
    basado en la formación de metahemoglobina es de 2 mg/kg de peso
    corporal por día en las ratas y los perros, y de 2,4 mg/kg de peso
    corporal por día en los ratones.  En estudios de toxicidad a largo
    plazo realizados con ratones y ratas, los cambios relacionados con el
    tratamiento se han asociado principalmente a la oxidación de la
    hemoglobina o a alteraciones de los hepatocitos.

         En estudios de la carcinogenicidad en ratones y ratas con niveles
    de hasta 10 000 mg/kg en la alimentación, no se observaron efectos
    relacionados con el tratamiento en la incidencia de tumores. 
    Específicamente, no se registraron neoplasias mesenquimatosas del bazo
    o el hígado como las observadas en los estudios de carcinogenicidad
    con 4-cloroanilina.

         En varios estudios de la toxicidad reproductiva en ratas,
    ratones, conejos y tres especies aviarias no se observó ningún efecto
    en la reproducción, ni tampoco embriotoxicidad.  Los estudios de
    teratogenicidad en ratas y conejos no revelaron ningún efecto
    teratogénico.

         El diflubenzurón y sus principales metabolitos han sido sometidos
    a una serie de ensayos de mutagenicidad  in vitro e  in vivo Ni el
    diflubenzurón ni sus principales metabolitos tienen efecto mutagénico.

         El metabolito secundario 4-cloroanilina ha dado un resultado
    positivo en varios ensayos de mutagenicidad  in vitro con diversas
    variables de valoración.  Es carcinógeno en las ratas y los ratones. 
    Las lesiones neoplásicas relacionadas con la administración de

    4-cloroanilina fueron tumores mesenquimatosos benignos y malignos en
    el bazo de ratas macho, y hemangiomas y hemangiosarcomas,
    principalmente en el bazo y el hígado de ratones macho.

    7.  Efectos en el ser humano

         Se ha notificado que el metabolito 4-cloroanilina del
    diflubenzurón ha causado metahemoglobinemia en trabajadores sometidos
    a exposición y en neonatos expuestos por inadvertencia.  Algunas
    personas con carencia de NADH metahemoglobina reductasa pueden ser
    particularmente sensibles a la 4-cloroanilina y, por lo tanto, a la
    exposición al diflubenzurón.

    8.  Efectos sobre otros organismos en el laboratorio y en el terreno

         Todos los organismos que sintetizan quitina presentan sensibilidad
    al diflubenzurón.

         Las bacterias no resultaron afectadas por el diflubenzurón a
    concentraciones de 500 mg/kg de suelo; se observó cierta estimulación
    de la fijación de nitrógeno.  Las soluciones de diflubenzurón-acetona
    se degradaron; la acetona se utilizó como fuente de carbono.  La
    biomasa de algas aumentó a una concentración de diflubenzurón de
    1 µg/litro.  No se observaron efectos adversos a concentraciones
    superiores al límite de solubilidad del diflubenzurón.  Los hongos
    resultaron temporalmente afectados a 0,1 µg/litro en condiciones de
    laboratorio.

         Los invertebrados acuáticos presentan respuestas variables al
    diflubenzurón.  Los moluscos son insensibles, con una CL50 superior a
    200 mg/litro.  Los valores de la CL50 de otros invertebrados van
    desde 1 hasta más de 1000 µg/litro, en función de los efectos del
    compuesto en las fases juvenil y de muda.  Para  Daphnia se ha
    estimado una MATC de > 40 y < 93 ng/litro; como era de prever, las
    larvas de mosca de mayo y otros insectos acuáticos juveniles son
    sumamente sensibles.  El rociamiento de masas de agua matará
    probablemente algunos invertebrados acuáticos.

         En los ecosistemas y experimentos de campo en que se aplicó
    diflubenzurón directamente al agua, los efectos en la mayoría de los
    grupos taxonómicos fueron menos graves de lo previsto a partir de las
    pruebas de laboratorio sobre efectos agudos.  No se han observado
    efectos en los organismos acuáticos después de aplicaciones aéreas a
    los bosques.

         Los valores de la CL50 para los peces son de > 150 mg/litro. 
    En los experimentos prácticos no se ha registrado nunca la muerte de
    peces.

         La DL50 oral y por contacto en la abeja de miel es superior a
    30 µg/individuo.  Las colonias de abejas no resultaron afectadas tras
    la aplicación aérea de 350 g de diflubenzurón/hectárea.

         Un estudio de alimentación de cinco días de duración en patos
    silvestres y codornices con niveles de hasta 4640 mg/kg de pienso no
    reveló ningún signo observable de toxicidad.  Las pequeñas aves
    canoras del ecosistema forestal no resultaron afectadas por la
    aplicación aérea de diflubenzurón a razón de 350 g/hectárea.

         Las especies mamíferas pequeñas no sufrieron mermas de las
    poblaciones tras la aplicación de diflubenzurón en un bosque a razón
    de 67 g/hectárea.
    


    See Also:
       Toxicological Abbreviations
       Diflubenzuron (HSG 99, 1995)
       Diflubenzuron (PDS)
       Diflubenzuron (Pesticide residues in food: 1981 evaluations)
       Diflubenzuron (Pesticide residues in food: 1983 evaluations)
       Diflubenzuron (Pesticide residues in food: 1984 evaluations)
       Diflubenzuron (Pesticide residues in food: 1985 evaluations Part II Toxicology)
       Diflubenzuron (JMPR Evaluations 2001 Part II Toxicological)