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    INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY



    ENVIRONMENTAL HEALTH CRITERIA 159





    GLYPHOSATE








    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 H. Mensink and
    Dr. P. Janssen, National Institute of Public
    Health and Environmental Hygiene,
    Bilthoven, The Netherlands


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

    World Health Orgnization
    Geneva, 1994


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

    Glyphosate.

        (Environmental health criteria ; 159)

        1.Glycine - analogs and derivatives  2.Herbicides
        3.Environmental exposure        I.Series

        ISBN 92 4 157159 4        (NLM Classification: WA 240)
        ISSN 0250-863X

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    CONTENTS

    ENVIRONMENTAL HEALTH CRITERIA FOR GLYPHOSATE

    1. SUMMARY

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

    2. IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, AND ANALYTICAL
        METHODS

        2.1. Identity
        2.2. Physical and chemical properties
        2.3. Formulations
        2.4. Conversion factors
        2.5. Analytical methods
             2.5.1. Sample handling and preparation
             2.5.2. Analytical methods

    3. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE

        3.1. Anthropogenic sources
             3.1.1. Production levels and processes
             3.1.2. Uses
             3.1.3. Drinking-water

    4. ENVIRONMENTAL TRANSPORT, DISTRIBUTION AND TRANSFORMATION

        4.1. Transport and distribution between media
             4.1.1. Water
             4.1.2. Soil sorption
             4.1.3. Mobility in soils
             4.1.4. Dissipation from the soil in the field
             4.1.5. Uptake and dissipation from plants
             4.1.6. Ingestion by animals
        4.2. Abiotic degradation
             4.2.1. Hydrolytic cleavage
             4.2.2. Photodegradation

        4.3. Biodegradation
        4.4. Bioaccumulation
        4.5. Waste disposal

    5. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE

        5.1. Environmental levels
        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
        6.4. Elimination and excretion
        6.5. Retention and turnover

    7. EFFECTS ON LABORATORY ANIMALS AND IN VITRO TEST SYSTEMS

        7.1. Single exposure
        7.2. Short-term exposure
             7.2.1. Oral studies
             7.2.2. Dermal studies
             7.2.3. Inhalational studies
        7.3. Long-term toxicity and carcinogenicity
        7.4. Skin and eye irritation; sensitization
        7.5. Reproductive toxicity, embryotoxicity and
             teratogenicity
             7.5.1. Teratogenicity studies
             7.5.2. Reproduction studies
        7.6. Mutagenicity and related end-points

    8. EFFECTS ON HUMANS

        8.1. Cases of intentional and accidental exposure
        8.2. Occupational exposure
        8.3. Subpopulations at special risk

    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  Plants
                  9.1.2.2  Invertebrates
                  9.1.2.3  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  Plants
                  9.2.2.2  Invertebrates
                  9.2.2.3  Vertebrates
             9.2.3. Terrestrial organisms
                  9.2.3.1  Plants
                  9.2.3.2  Invertebrates
                  9.2.3.3  Vertebrates

    10. EVALUATION OF HUMAN HEALTH HAZARDS AND EFFECTS ON THE
         ENVIRONMENT

         10.1. Human health hazards
         10.2. Evaluation of effects on the environment
            10.2.1. Exposure levels and toxic effects
            10.2.2. Hazard evaluation for aquatic organisms
            10.2.3. Hazard evaluation for terrestrial organisms

    11. RECOMMENDATIONS FOR PROTECTION OF HUMAN HEALTH

    12. FURTHER RESEARCH

    REFERENCES

    RESUME

    RESUMEN
    

    WHO TASK GROUP ON ENVIRONMENTAL HEALTH CRITERIA FOR GLYPHOSATE

     Members

    Dr   S. Dobson, Institute of Terrestrial Ecology, Monks Wood
         Experimental Station, Huntingdon, United Kingdom
          (Chairman)

    Dr   A.H. El-Sebae, College of Agriculture, Alexandria University,
         El Shatby, Alexandria, Egypt

    Dr   P. Janssen, National Institute of Public Health and
         Environmental Hygiene, Bilthoven, The Netherlands

    Dr   H. Mensink, National Institute of Public Health and
         Environmental Hygiene, Bilthoven, The Netherlands

    Dr   M.S. Morrow, Health Effects Division, Office of Pesticide 
         Programs, US Environmental Protection Agency, Washington, DC,
         USA

    Professor R. Nilsson, Department of Scientific Documentation and
         Research, National Chemicals Inspectorate, Solna, Swedena

    Dr   R. Ye, National Environmental Protection Agency, Beijing, 
         People's Republic of China

     Observers

    Dr   C. Hastings, Agricultural Group, Monsanto, Missouri, St. 
         Louis, USA

     Secretariat

    Dr   M. Gilbert, International Programme on Chemical Safety, World
         Health Organization, Geneva, Switzerland  (Secretary)

    ___________

    a Invited but unable to attend.

    NOTE TO READERS OF THE CRITERIA MONOGRAPHS

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



                                  *   *   *


         A detailed data profile and a legal file can be obtained from
    the International Register of Potentially Toxic Chemicals, Case
    postale 356, 1219 Châtelaine, Geneva, Switzerland (Telephone No.
    9799111).



                                  *   *   *



         This publication was made possible by grant number 5 U01
    ES02617-14 from the National Institute of Environmental Health
    Sciences, National Institutes of Health, USA.

    ENVIRONMENTAL HEALTH CRITERIA FOR GLYPHOSATE

         A Task Group on Environmental Health Criteria for Glyphosate
    met at the Institute of Terrestrial Ecology, Monks Wood, United
    Kingdom, from 23 to 27 August 1993. Dr S. Dobson welcomed the
    participants on behalf of the host institution, and Dr M. Gilbert
    opened the Meeting on behalf of the three cooperating organizations
    of the IPCS (UNEP/ILO/WHO). The Task Group reviewed and revised the
    draft monograph and made an evaluation of the risks for human health
    and the environment from exposure to glyphosate.

         The first draft of this monograph was prepared by Dr H. Mensink
    and Dr P. Janssen, National Institute of Public Health and
    Environmental Hygiene, Bilthoven, The Netherlands.

         Dr M. Gilbert was responsible for the overall scientific
    content of the monograph and for the organization of the meeting,
    and Dr P.G. Jenkins, IPCS, for the technical editing of the
    monograph.

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

    ABBREVIATIONS

    a.i.           active ingredient

    ALAT           alanine aminotransferase

    AMPA           aminomethylphosphonic acid

    AP             alkaline phosphatase

    CHO            Chinese hamster ovary

    CNS            central nervous system

    HPLC           high-performance liquid chromatography

    i.p.           intraperitoneal

    IPA            isopropylamine

    MATC           maximum acceptable toxicant concentration

    NOAEL          no-observed-adverse-effect level

    NOEC           no-observed-effect concentration

    1.  SUMMARY

    1.1  Identity, physical and chemical properties, and analytical
         methods

         Glyphosate is a weak organic acid consisting of a glycine and a
    phosphonomethyl moiety. The empirical formula is C3H8NO5P.
    Glyphosate is usually formulated as a salt of the deprotonated acid
    of glyphosate and a cation, e.g., isopropylamine or
    trimethylsulfonium. The purity of technical grade glyphosate is
    generally above 90%. Technical grade glyphosate is an odourless
    white crystalline powder with a specific gravity of 1.704, a very
    low vapour pressure, and a high solubility in water. The
    octanol-water partition coefficient (log Kow) is -2.8. Glyphosate
    is amphoteric and may exist as different ionic species, dependent on
    the actual pH.

         Determination of glyphosate is in general laborious, complex,
    and costly. Derivatization with fluorogenic substances is the most
    common method and may be applied pre- or post-column. Determination
    is usually carried out with high performance liquid chromatography
    or gas liquid chromatography. Limits of determination for glyphosate
    in water, plants, soil and human urine, are 0.02-3.2 µg/litre,
    0.01-0.3 mg/kg, 0.05-1 mg/kg and 0.1 mg/litre, respectively.

    1.2  Sources of human and environmental exposure

         Glyphosate is a post-emergent, systemic and non-selective
    herbicide that is used in both agricultural and non-agricultural
    areas all over the world. Glyphosate is applied to many crops and in
    various commercial formulations. The major formulation is Roundup in
    which glyphosate is formulated as the isopropylamine salt.
    Recommended application rates do not exceed 5.8 kg a.i./ha and are
    dependent on the type of use. Environmental exposure may occur
    because of deposition due to drift and accidental releases.

    1.3  Environmental transport, distribution and transformation

         The most important processes of dissipation that may be
    involved after application of glyphosate are complexation in water
    with ions, e.g., Ca2+ and Mg2+, sorption to sediment, suspended
    particles in water, and soil, photodegradation in water, uptake by
    plants, and biodegradation.

         Glyphosate dissipates from the water with DT50 values
    (dissipation) ranging from a few days to more than 91 days. Sediment
    or suspended particles are shown to be the major sink.

         The adsorption coefficients (Ks/l) of glyphosate in
    laboratory experiments vary between 8 and 377 dm3/kg for various
    soils and clay minerals. No data on the sorption of
    aminomethylphosphonic acid (AMPA), the major metabolite, under
    laboratory conditions are available.

         Rf values of glyphosate do not exceed 0.2 in soil thin-layer
    chromatography experiments. Between less than 0.1% and 11% of the
    applied activity is recovered in the eluate of soil columns under
    leaching conditions simulating an extremely high rainfall. From
    field experiments it appears that AMPA is not likely to leach.

         Glyphosate dissipates in field experiments from the soil with
    DT50 values between 3 and 174 days, mainly depending on edaphic
    and climatic conditions. Up to 1.8% of the applied dose dissipated
    from the soil due to run-off in some field experiments.

         Under laboratory conditions, up to 45% of the applied activity
    may be absorbed by treated leaves, and this is followed by a
    substantial translocation.

         Hydrolysis of glyphosate in sterile buffers is very slow with
    DT50 values >> 35 days. Photodegradation in water under natural
    conditions occurs with DT50 values < 28 days. No substantial
    photodegradation in soil was recorded in a study lasting 31 days.

         The time needed for 50% biodegradation of glyphosate in the
    whole system of a test with water and sediment is > 14 days under
    aerobic conditions and 14-22 days under anaerobic conditions in the
    laboratory. The time needed for 50% biodegradation of glyphosate in
    the soil is 2-3 days under aerobic conditions.

         The major metabolite in soil and water is AMPA. Maximum amounts
    of AMPA in soils are approximately 20% of the applied activity under
    aerobic conditions and 0.5% under anaerobic conditions. Maximum
    amounts of AMPA in sediments are 25% under both aerobic and
    anaerobic conditions.

         Bioconcentration factors are low in laboratory tests with
    invertebrates and fish. Bluegill sunfish in a flow-through test
    showed a depuration half-life of 35 days, after being exposed for 35
    days. AMPA is recovered in bluegill sunfish up to 21 days after
    continuous exposure to glyphosate. Glyphosate has not been detected
    in fish living in directly sprayed water in field experiments. In
    one experiment, AMPA was detectable in carp up to 90 days after
    application. No biomagnification of glyphosate in litter by
    herbivorous and omnivorous small mammals in a forest brush ecosystem
    was indicated in a field experiment. Concentrations of up to 5 mg
    a.i./kg were measured in deermice immediately after spraying in this
    experiment.

         A range of bacterial strains can degrade glyphosate. Bacteria
    capable of using the compound as sole phosphorus, sole carbon or
    sole nitrogen source have been identified. Growth is slow compared
    to growth on inorganic sources of P, C and N. There is evidence from
    the field that bacterial populations adapted to metabolise
    glyphosate. The presence of inorganic phosphate inhibits degradation
    of glyphosate with some, but not all, bacteria. Biodegradation of
    glyphosate may involve co-metabolism with other energy sources.

    1.4  Environmental levels and human exposure

         Data on the occurrence of glyphosate in environmental biota and
    abiota as part of regular monitoring programmes are very scarce.
    Data from field experiments in which common agricultural practice is
    simulated are used to indicate maximum environmental concentrations:
    < 1-1700 µg/litre surface water, 0.07-40 mg/kg dry weight soil,
    < 0.05-19 mg/kg dry weight sediment, 261-1300 mg/kg foliage, 5 mg/kg
    the viscera of deermice, 1.6-19 mg/kg wild berries, and 45 mg/kg
    lichens. The corresponding maximum concentrations of AMPA are:
    < 1-35 µg/litre (surface water), 0.1-9 mg/kg dry weight (soil),
    < 0.05-1.8 mg/kg dry weight (sediment), 1.7-< 9 mg/kg (foliage),
    0.02-0.1 mg/kg (wild berries), and 2.1 mg/kg (lichens). The
    above-mentioned concentrations of glyphosate are generally found
    immediately after application. The concentration in lichens was
    found 270 days after application.

         Measurements of daily human intake of glyphosate via food and
    drinking-water (total diet studies) are not available. The few data
    on occupational exposure indicate that exposure levels for workers
    applying glyphosate as the herbicide formulation Roundup are low.

    1.5  Kinetics and metabolism in laboratory animals and humans

         Technical glyphosate is only partially absorbed from the
    gastrointestinal tract. In studies with 14C-labelled glyphosate,
    absorption percentages of 30-36% were found in several species.
    Dermal absorption is low. From the herbicide formulation Roundup,
    < 5.5% of the glyphosate present is absorbed through the skin
    (contact time about 24 h). In body tissues, the highest
    concentrations, approximately 1% of the oral dose, are found in
    bone. Following a single oral dose, 62-69% is eliminated in the
    faeces without absorption. Of the absorbed glyphosate, 14-29% is
    excreted in urine and 0.2% or less in expired air. Biliary excretion
    following intravenous application was only 5-8%. In lactating goats,
    excretion in milk was shown to occur to a minor extent only
    (concentration < 0.1 mg/kg whole milk at a dose level of
    120 mg/kg diet). Biotransformation of glyphosate occurs to a very
    low degree only. The only metabolite, AMPA, accounts for 0.3% of the
    dose or less; the rest is unchanged glyphosate. Whole body clearance
    (99% of an oral dose) occurs in approximately 168 h.

    1.6  Effects on laboratory mammals, and  in vitro test systems

         In experimental animals, technical glyphosate has very low
    acute toxicity by the oral and dermal administration routes; it is
    markedly more toxic by the intraperitoneal route than by other
    routes. Short-term feeding studies have been conducted in several
    species, but few effects were seen in most of these tests. In one
    13-week study in mice with technical glyphosate, increased weights
    of several organs and growth retardation were observed at
    50 000 mg/kg diet. In a 13-week study in rats no effect occurred
    (technical glyphosate dose levels up to 20 000 mg/kg diet). In
    another 13-week study, lesions of the salivary glands were found in
    rats and mice. In mice, the NOAEL was 3125 mg/kg diet; in rats, it
    was < 3125 mg/kg diet. These findings were not present in any other
    short-term or long-term studies conducted in different strains and
    species. The salivary lesions suggest that glyphosate may be acting
    as a weak adrenergic agonist.

         Long-term toxicity was studied in mice and rats. Few effects
    were observed and, in almost all cases, at relatively high dose
    levels only. In mice, technical glyphosate produced growth
    retardation, hepatocyte hypertrophy or necrosis and urinary bladder
    epithelial hyperplasia at 30 000 mg/kg. In rats, the same test
    compound produced decreased growth, increased liver weights,
    degenerative lens changes and gastric inflammation at 20 000 mg/kg
    diet.

         The available studies do not indicate that technical glyphosate
    is mutagenic, carcinogenic or teratogenic. Two multigeneration
    studies were carried out in rats. The main effects of technical
    glyphosate were decreased body weights of parent animals and pups
    and decreased litter size at 30 000 mg/kg diet. In one reproduction
    study, an increase in the incidence of unilateral renal tubular
    dilation in F3b male pups at 30 mg/kg body weight was reported.
    The absence of a renal effect in pups at a higher dose level in the
    other reproduction study indicates that the reproducibility of this
    lesion is uncertain.

    1.7  Effects on humans

         The available controlled studies are limited to three
    irritation/sensitization studies in human volunteers, the results of
    which indicated no effect. Several cases of (mostly intentional)
    intoxications with technical glyphosate herbicide formulation
    Roundup have been reported. In a study on health effects in workers
    applying Roundup herbicide formulation, no adverse effects were
    found. Available data on occupational exposure for workers applying
    Roundup indicate exposure levels far below the NOAELs from the
    relevant animal experiments.

    1.8  Effects on other organisms in the laboratory and field

         Technical grade glyphosate is moderately to slightly toxic to
    aquatic microorganisms, with EC50 (3-4 days) values of
    1.2-7.8 mg/litre, and 7-day NOEC values of 0.3-34 mg/litre.
    Formulations of glyphosate are slightly to highly toxic to aquatic
    microorganisms with 3-day EC50 values of 1.0 to > 55 mg product
    per litre. Cyanophyta (blue-green algae) are more sensitive to
    Roundup than true algae. Physiological processes that are affected
    include the greening process, respiration, photosynthesis, and the
    synthesis of aromatic amino acids.

         Soil bacteria in culture have shown effects of glyphosate on
    nitrogen fixation, denitrification and nitrification. However, field
    studies after application of formulations have not shown significant
    effects. Closely related species of bacteria have been shown capable
    of degrading glyphosate.

         Mycelial growth of ectomycorrhizal fungi in pure cultures is
    inhibited at concentrations of > 29 µg Roundup/litre. Sensitive
    genera are  Cenococcum, Hebeloma and Laccaria.

         Glyphosate is slightly toxic to aquatic macrophytes with a
    14-day NOEC value of 9 mg/litre, when dissolved in water. Roundup is
    also slightly toxic with 14-day NOEC values of 2.4-56 mg
    Roundup/litre, when dissolved in water. No data on acute toxicity
    are available. Phytotoxicity is much higher when sprayed deposits
    are not washed off.

         Technical grade glyphosate is slightly to very slightly toxic
    to aquatic invertebrates with 2- to 4-day LC50 or EC50 values of
    > 55 mg/litre, and a 21-day NOEC value of 100 mg/litre.
    Formulations of glyphosate are moderately to very slightly toxic to
    aquatic invertebrates with 2-day EC50 values of 5.3-5600 mg
    product/litre and 21-day MATC values of 1.4-4.9 mg product per
    litre. The higher toxicity of Roundup is mainly due to the presence
    of surfactants.

         Technical grade glyphosate is moderately to very slightly toxic
    to fish, with 4-day LC50 values of 10 to > 1000 mg/litre, a
    21-day NOEC value of 52 mg/litre, and an MATC value of >
    26 mg/litre. Formulations of glyphosate are also moderately to very
    slightly toxic to fish with 4-day LC50 values of 2.4 to > 1000 mg
    product per litre, and 21-day NOEC values of 0.8-2.4 mg
    product/litre. The most sensitive species is the carp, when exposed
    to the formulation Sting. No treatment-related effects of Roundup on
    fish have been found under field conditions, with the exception of
    stress immediately after application of a recommended rate and
    avoidance of concentrations of > 40 mg Roundup/litre.

         Nodulation of sub-clover inoculated with Rhizobium is inhibited
    in a dose-related way in soil-free systems with nutrient solutions
    at concentrations of > 2 mg a.i./litre. Seed germination of
    various forest species is not affected by glyphosate at the
    recommended application rates. The root length of red pine seedlings
    is decreased under laboratory conditions in a dose-related way at
    application rates of > 0.54 kg a.i./ha. This decrease was not
    confirmed in a comparable field experiment.

         Technical grade glyphosate and Roundup are slightly toxic to
    bees when applied either orally or topically. The 2-day LD50
    values are > 100 µg (a.i. or product) per bee. The oral 2-day
    LD50 of Sting to bees is > 100 µg/bee. Roundup and Roundup D-pak
    are slightly toxic to earthworms with 14-day NOEC values of 500 and
    158 mg product per kg dry weight, respectively. No adverse effects
    of Roundup were found on the fecundity and fertility of green
    lacewings, and there were no effects of Sting on the food uptake and
    mortality of the beetle  Poecilus.

         Technical grade glyphosate is slightly toxic to birds, with an
    LD50 of >3851 mg/kg body weight, an 8-day LC50 of >4640 mg/kg
    feed, and 112- to 119-day NOEC values of > 1000 mg/kg feed.
    Roundup and an unknown formulation are also slightly toxic to birds,
    with an LD50 of > 2686 mg product/kg body weight and an 8-day
    LC50 of > 5620 mg product/kg feed. Generally no treatment-related
    effects of technical grade glyphosate or Roundup on mammals are
    found under laboratory conditions, except at very high application
    rates. Treatment-related effects on birds and mammals under field
    conditions appear to be primarily due to habitat changes after
    treatment with Roundup.

    2.  IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, AND ANALYTICAL
        METHODS

    2.1  Identity

         Glyphosate is the primary name of a weak organic acid that
    consists of a glycine moiety and a phosphonomethyl moiety. The
    chemical name is  N-(phosphonomethyl)glycine according to IUPAC
    nomenclature. The CAS name is glycine,  N-(phosphonomethyl)-, and
    its CAS registry number is 1071-83-6. The empirical formula is
    C3H8NO5P, and the structural formula is as follows:

    CHEMICAL STRUCTURE

         The relative molecular mass of glyphosate is 169.07. Technical
    grade glyphosate has a purity of > 80%, but the purity generally
    exceeds 90%. Glyphosate usually is formulated as a salt of the
    deprotonated acid of glyphosate and a cation, e.g., isopropylamine.
    The CAS registry number of the salt of glyphosate and
    isopropyl-amine is 38641-94-0.

         Surfactants and inerts may be added to formulations of
    glyphosate. The type of surfactant and its concentration may differ
    per formulation. A common surfactant in the major formulation
    Roundup is polyoxyethylene amine. Other known surfactants are ortho
    X-77 (Mitchell et al., 1987), LI-700, R-11 and Widespread (Monsanto,
    1990a). Other additives in formulations may be sulfuric and
    phosphoric acids.

    2.2  Physical and chemical properties

         The physical and chemical properties of glyphosate are
    tabulated in Table 1. Glyphosate is an amphoteric compound of which
    the ionic species and their pKa values are presented in Fig. 1. Due
    to its high polarity glyphosate is practically insoluble in, for
    instance, ethanol, acetone and benzene.

    
    Table 1.  Physical and chemical properties of glyphosatea
                                                                                            

                                                               Remarks
                                                                                            

    Physical state                   crystalline powder

    Colour                           white

    Odour                            none

    Melting pointb                   184.5 °C                  decomposition at 187 °C

    Boiling point                    n.a.

    Specific gravity (density)c      1.704                     20 °C

    Vapour pressured                 < 1 x 10-5 Pa             25 °C

    Solubility in waterb,e           10 100 mg/litre           20 °C

    Henry's law constant             < 7 x 10-11

    Octanol-water partition
      coefficient (log Kow)d         -2.8

    Surface tensiond                 0.072 N/m                 0.5% (w/v) at approx. 25 °C

    pKa valuesd,f                    < 2, 2.6, 5.6, 10.6       Sprankle et al. (1975)

    Molar absorptivityc              0.086 litre/mol per cm    at 295 nm

    Flammabilityd                    not flammable

    Explosivenessd                   not explosive

    pHd                              2.5                       1% solution
                                                                                            

    a    data provided by Monsanto Ltd
    b    purity 96%
    c    purity 100%
    d    purity not reported
    e    pure glyphosate had been reported to have a water solubility of
         11 600 mg/litre at 25 °C
    f    free acid
    n.a. = not applicable
    
    2.3  Formulations

         Glyphosate can be applied in various formulations. A synopsis
    of these formulations, their concentrations of active ingredient,
    and the countries in which the use is permitted is presented in
    Table 2. This synopsis is not complete. Formulations may contain
    specific surfactants. The major formulation of glyphosate is Roundup
    containing 480 g/litre of the isopropylamine salt, which is
    equivalent to 360 g/litre of the free acid. Some other Roundup
    formulations that are characterized by other a.i. concentrations or
    other surfactants have been developed for specific applications.
    Other formulations that have been developed for special equipment
    are Roundup Ultrabax for CDA equipment, Glyphosate Nomix for Nomix
    equipment, and EZ-JECT for tree injections. In Canada, Roundup was
    re-labelled as Vision in 1987 for use in forestry.

    FIGURE 1

    
    Table 2.  Composition of various commercial formulations with glyphosatea
                                                                                               

    Name                 Synonyms             Concentration    Country
                                                a.i. (%)
                                                                                               

    Roundup              Spasor,              48.0 (w/v);      Most countries

    Sting                Vision,              41.0 (w/w)b

                         Swing,               21.7 (w/w)       Belgium, Cameroon, France,
                         Arcade,                               Holland, Kenya, Malawi,
                         Tomahawk                              Portugal, South Africa,
                                                               United Kingdom

    Armada               Frontier             16.6 (w/w)       Belgium, Cameroon, Ivory
                                                               Coast, Gabon, Greece, Zaire
    Dardo                Ricochetg,           12.2 (w/w)       Cameroon, Egypt, France,
                         Rival,                                Greece, Israel, Italy,
                         Ultrasonic                            Portugal, Spain, United
                                                               Kingdom

    Squadron                                  20.2 (w/w)       Argentina, Australia, Columbia
    Stirrup              Nomix, Expedite      18.3 (w/w)       France, United Kingdom
    Wallop                                    20.8 (w/w)       Malaysia
    Deploy Dryc                               94.0 (w/w)       USA
    Quotamakerd                               75.0 (w/w)       USA
    Landmaster IIe                            13.3 (w/w)       USA
    Landmaster BW,
      Campaignf                               12.9 (w/w)       USA
    Roundup D-Pak                             62.0 (w/w)       USA
    Rodeo                                     53.8 (w/w)       USA
    Ranger                                    28.6 (w/w)       USA
    Roundup Lawn and
      Garden Conc.                            18.0 (w/w)       USA
    Roundup-Ready-
      To-Use                                  0.96 (w/w)       USA
    Fusta                                     22.5 (w/w)       Spain
                                                                                               

    Table 2 (continued)
    a    all formulations produced by Monsanto Ltd; data provided by Monsanto Ltd
    b    based on the isopropylamine salt; equivalent to 36.0% (w/v) and 30.5% (w/w)
         of the free acid
    c    dry formulation of the monoammonium salt
    d    dry formulation of the sodium sesqui salt
    e    also contains 11.1% 2,4-D (isopropylamine salt)
    f    also contains 20.6% 2,4-D (isopropylamine salt)
    g    also contains simazine
    
         Formulations may contain other active ingredients, e.g.,
    simazine in Ricochet, 2,4-D in Landmaster, and MCPA in Fusta.

    2.4  Conversion factors

         1 ppm = 6.91 mg/m3 at 25 °C and 101.3 kPa

         1 mg/m3 = 0.145 ppm

    2.5  Analytical methods

    2.5.1  Sample handling and preparation

         The first preparative step before detection and measurement of
    glyphosate is generally extraction. As both glyphosate and its main
    metabolite aminomethylphosphonic acid (AMPA, see Fig. 2) show high
    polarity, and are therefore highly water soluble, they are difficult
    to extract with organic solvents. However, various methods have been
    developed. Some recently developed extraction methods for different
    media are summarized in Table 3.

    FIGURE 2


    
    Table 3.  Sampling, preparation, and analysis of glyphosate
                                                                                                                                              

    Medium            Sampling    Preparations           Derivatization       Analytical      Limit of           Recovery     Reference
                      volume or                          reagent              method          determinationa
                      weight
                                                                                                                                              

    Air               n.r.        collected onto an      trifluoroacetic      GC-MS and       approx. 0.3        94%          Jauhiainen
                                  absorption liquid;     anhydride and        GC-EC           µg/m3                           et al. (1991)
                                  evaporation to         trifluoroethanol
                                  dryness

    Cyano-bacteria    100 ml      dry, resuspend in      PITC                 HPLC with a     n.r.b              78%          Powell et al.
                                  methanol/sodium -                           radically                                       (1990)
                                  acetate/               column               compressed
                                  triethylamine

    Plants            5 g         extraction with        trifluoroacetic      GC-NPD          0.03 mg/kg         72-92%       Konar & Roy,
                                  water/chloroform;      anhydride and                                                        (1990)
                                  preconcentration       trifluoroethanol
                                  and clean-up on
                                  cation-exchange
                                  resin

    Plants            25-50 g     extraction with                             TLC with        0.01 mg/kg         n.r.         Bunyathyan &
                                  water/chloroform;                           ninhydrin                                       Gevorgyan
                                  preconcentration                            detection                                       (1984)
                                  and clean-up on
                                  anion-exchange 
                                  and cation-exchange
                                  resin; evaporation
                                  to dryness
                                                                                                                                              

    Table 3. cont'd (2)
                                                                                                                                              

    Medium      Sampling        Preparations             Derivatization      Analytical         Limit of           Recovery    Reference
                volume or                                reagent             method             determinationa
                weight
                                                                                                                                              

    Water       250 ml          extraction with          o-phthalaldehyde    LC                 3.2 µg/litre       89%         Wigfield &
                                dichloromethane;                                                                               Lanouette
                                adsorption on                                                                                  (1990)
                                anion-exchange
                                resin

    Water       25 ml           extraction with          FMOCCl              HPLC and TLC       0.02 µg/litre      80%         Gauch et al.
                                dichloromethane/                                                                               (1989)
                                2-propanol;
                                acidification
                                with H2SO4;
                                evaporation
                                to dryness

    Water       1-1.5 litre     no extraction;                               TLC with -         0.05 mg/litre      n.r.        Bunyathyan &
                                preconcentration                             ninhydrin                                         Gevorgyan
                                and clean-up                                 detectionc                                        (1984)
                                with anion-exchange
                                and cation-exchange
                                resin

    Soil        5 g             extraction with          trifluoroacetic     GC-NPD             0.05 mg/kg         75%         Roy & Konar
                                deionized water/         anhydride/                                                            (1989)
                                H3PO4; addition          trifluoroethanol
                                of Darco
                                charcoal
                                                                                                                                              

    Table 3. cont'd (3)
                                                                                                                                              

    Medium      Sampling        Preparations             Derivatization      Analytical         Limit of           Recovery    Reference
                volume or                                reagent             method             determinationa
                weight
                                                                                                                                              

    Soil        2 g (sandy      extraction with          FMOCCl              HPLC               1 mg/kg            80-119%     Miles & Moye,
                soil); 25 g     KH2PO4 (sandy soil),                                                                           (1988b)
                (clayish        KOH (clayish soil);
                soil)           no clean-up

    Soil,       5 g (soil);     extraction with NH4OH;   ninhydrin           LC                 0.05-0.1           73-79%      Thompson
    sediment,   20 g (sed);     adsorption on                                                   mg/kg (soil);      (soil)      et al. (1989)
    foliage     5 g (fol)       anion-exchange resin;                                           0.1 mg/kg          65-84% 
                                further clean-up with                                           (sed); 0.3         (sed)
                                Dowex cation-exchange                                           mg/kg (fol)d       81-84%
                                resin                                                                              (fol)

    Urine and   5-6 g           extraction with H2O                          HPLC (ion          n.r.               81-99%      Monsanto
    faeces of                   (only faeces); protein                       pair, strong                                      (1988a)
    the rat                     precipitation and                            anion and
                                lyophilization                               cation-exchange),
                                (only urine);                                LSC, 1H NMR, 31P
                                clean-up with C18                            NMR, GC/MS
                                column

    Urine       n.r.            adsorption on            trifluoroacetic     GC-MS and          0.1 mg/litre       n.r.        Jauhiainen
    (human                      anion-exchange resin     anhydride/          GC-EC                                             et al. (1991)
    male)                       (SAX); elution of the    trifluoroethanol
                                resin with HCl;
                                evaporation to
                                dryness
                                                                                                                                              

    Table 3. cont'd (4)
                                                                                                                                              

    Medium      Sampling        Preparations             Derivatization      Analytical         Limit of           Recovery    Reference
                volume or                                reagent             method             determinationa
                weight
                                                                                                                                              

    Serum       0.5 ml          extraction with           p-toluene           HPLC with UV       n.r.e              n.r         Tomita et
    (human)                     trichloroacetic          sulfonyl chloride   detection                                         al. (1991)
                                acid; adsorption
                                on anion-exchange
                                resin; elution
                                with HCl; evaporation
                                to dryness
                                                                                                                                              

    a    In no study with a non-liquid medium was it reported whether the limit of determination was based on dry or fresh weight,
         except in the study of Thompson et al. (1989).
    b    The order of magnitude was reported to be picomol.
    c    The use of TLC with ninhydrin, copper nitrate and rhodamine B detection is reported for glyphosate in distilled water in
         Ragab (1978).
    d    The limits of determination in soil, sediment, and foliage are expressed per kg dry weight.
    e    Only the limit of detection was reported: 0.3 mg/litre (approximately 75% recovery).

    PITC = phenylisothiocyanate; FMOCCl = 9-fluorenyl-methyl chloroformate; GC =gas chromatography;
    (HP)LC = (high-performance) liquid chromatography; TLC = thin layer chromatography; MS = mass spectroscopy;
    EC = electron capture detector; NPD = nitrogen-phosphorus detector; n.r. = not reported;
    LSC = liquid scintillation counting; NMR = nuclear magnetic resonance; sed = sediment; fol = foliage
    

         The second preparative step is the clean-up, which may include
    extraction, preconcentration by evaporation, ion-exchange
    chromatography or gel chromatography. Clean-up procedures may
    involve different combinations of chromatographic techniques. In a
    validation study in which plant tissues and water were analysed, a
    Chelex column was combined with anion-exchange clean-up (Cowell
    et al., 1986). No chromatography was included in the clean-up
    procedures for analysing glyphosate and AMPA in natural waters
    (Miles et al., 1986). In this procedure samples were successively
    filtrated, supplied with phosphate buffer, concentrated by
    evaporation, and filtrated, prior to derivatization.
    Samples with urine and faeces of the rat were subjected to clean-up
    with a C18 column (Monsanto, 1988a). Prior to this extraction,
    proteins were precipitated and the samples were lyophilized; samples
    of faeces were, however, only extracted with water.

         The third preparative step is derivatization. Derivatization
    with a fluorogenic reagent is common. Burns (1983), however,
    developed a preparation technique without derivatization.
    Derivatization prior to detection and measurement with HPLC can be
    pre-column (Miles et al., 1986; Lundgren, 1986; Miles & Moye,
    1988a) or post-column (Moye et al., 1983; Tuinstra & Kienhuis,
    1987). 9-Fluorenylmethyl chloroformate, phenylisothiocyanate and
    1-fluoro-2,4-dinitrobenzene may be used as pre-column reagents,
    whereas ortho-phthalaldehyde-mercaptoethanol and ninhydrin may be
    used as post-column fluorogenic reagents. With post-column
    techniques, derivatives can be formed on-line, but it requires more
    equipment and experience. On the other hand, pre-column techniques
    are often more rapid and require less equipment and experience. In
    general the facilities required for derivatization with fluorogenic
    substances are very specific, and therefore not available in many
    laboratories (Konar & Roy, 1990). These authors proposed
    derivatization with a mixture of trifluoroacetic anhydride and
    trifluoroethanol prior to analysis with gas chromatography as a
    simpler, less laborious and more economical method. This proposal
    referred to the determination of glyphosate and AMPA in plant
    tissues. This and other recently developed techniques of clean-up
    and derivatization are summarized in Table 2. These techniques are
    intended to simplify and improve preparative techniques, which in
    general used to be complex and costly (Marcotte et al., 1977;
    Guinivan et al., 1982; Roseboom & Berkhoff, 1982; Moye et al.,
    1983; Moye & Deyrup, 1984; Deyrup et al., 1985; Miles et al.,
    1986; Lundgren, 1986; Miles & Moye, 1988b).

         Sample preparation and derivatization, as developed by Powell
    et al. (1990) for cyanobacteria without deproteinization (see
    Table 3), should also be usable for plant and animal tissue. In this
    case, a simple maceration step prior to ethanol extraction should be
    included. Bunyathyan & Gevorgyan (1984) developed preparative
    techniques for different media prior to analysis with TLC. Only

    their procedures for plants and water are summarized in Table 3. The
    preparative technique for soil samples was comparable with that of
    Thompson et al. (1989), although samples of 25-50 g were required.
    Bunyathyan & Gevorgyan (1984) also developed a method for preparing
    20-litre air samples prior to TLC. They extracted the residues
    collected on a filter with water before clean-up on a
    cation-exchange resin.

    2.5.2  Analytical methods

         Various analytical methods for the determination of glyphosate
    have been described, including thin-layer chromatography (Young
    et al., 1977; Ragab, 1978; Bunyathyan & Gevorgyan, 1984),
    colorimetry (Glass, 1981), differential pulse polarography (Friestad
    & Bronstad, 1985), gas chromatography (Guinivan et al., 1982; Moye
    & Deyrup, 1984; Deyrup et al., 1985), high-performance liquid
    chromatography (Miles & Moye, 1988a; Powell et al., 1990), and
    31P NMR (Dickson et al., 1988). Some of these techniques, their
    analytical recoveries and limits of determination are listed in
    Table 3. The corresponding determination limits for AMPA, i.e.
    analysed with the same techniques, are listed in Table 4. Recoveries
    in the different media appear to be higher for glyphosate than for
    AMPA. This is probably due to optimization of the systems for
    glyphosate, as was done by Thompson et al.(1989).

        Table 4. Limits of determination of AMPA
                                                                                            

    Medium            Limit of              Recovery           Reference
                      determination
                                                                                            

    Plants            0.01 mg/kg             61-73%           Konar & Roy (1990)

    Water             1.2 µg/litre             86%            Wigfield & Lanouette (1990)

    Soil              0.01 mg/kg               66%            Roy & Konar (1989)

    Soil              0.03-0.05 mg/kg        58-68%           Thompson et al. (1989)

    Sediment          0.03 mg/kg             54-67%           Thompson et al. (1989)

    Foliage           0.008 mg/kg            55-70%           Thompson et al. (1989)

    Urine (human)     0.05 mg/litre           n.r.            Jauhiainen et al. (1991)

    Serum (human)     n.r.a                   n.r.            Tomita et al. (1991)
                                                                                            
    a  n.r. = not reported; only the limit of detection was reported: 0.2 mg/litre
       (approximately 88% recovery)
    
         TLC techniques are generally based on silica gel or cellulose
    plates; cellulose plates give a better separation (Dubelman, 1988).
    Ninhydrin and phosphate sensitive reagents may be used for
    detection, although interference from co-extractives may occur.
    According to Dubelman (1988), fluorogenic reagents may be preferable
    in case of interference.

         Fluorogenic derivatives can be determined in HPLC analysis with
    fluorescence detectors (Wigfield & Lanouette, 1990) and also with a
    spectrophotometer (Powell et al., 1990). In a GC analysis a
    nitrogen-phosphorus, electron capture or a flame photometric
    detector can be used.

    3.  SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE

    3.1  Anthropogenic sources

    3.1.1  Production levels and processes

         No data on the world production of glyphosate and its
    formulations are available. In addition, no data on losses to the
    environment during normal production and formulation or accidental
    losses have been reported.

         The first phase of the production of glyphosate consists of
    refluxing a mixture of glycine (50 parts), chloromethylphosphonic
    acid (92 parts), an aqueous solution with 50% sodium hydroxide (150
    parts), and water (100 parts) in a suitable reaction vessel. Another
    50 parts of an aqueous solution with 50% sodium hydroxide are added
    to maintain the pH between 10 and 12, whereafter the reaction
    mixture is refluxed for another 20 h. The mixture is then cooled to
    room temperature and filtered. After adding 160 parts of
    concentrated hydrochloric acid, this mixture is again filtered.
    Glyphosate will slowly precipitate in the filtrate (IRPTC, 1991).

    3.1.2  Uses

         Glyphosate is a post-emergent, systemic and non-selective
    herbicide intended for use against deep-rooted perennial species,
    and also biennial and annual broad-leaved, grass and sedge species
    (WSSA, 1983; Monsanto, personal communication to the IPCS, 1991).
    Glyphosate is used in both agriculture and forestry. Fields of
    agricultural use include grassland renovation, horticulture,
    fructiculture, arable cultivation, and rice cultivation. Use in
    forestry includes the killing of fast growing competitors in conifer
    plantations or conservation areas, and the treatment of tree stumps.
    Glyphosate may also be used for weed killing in non-agricultural
    areas such as water systems, including irrigation and temporarily
    drained waters, parks, road verges and gardens.

         The uses of glyphosate indicate that it can be applied in
    various crops for specific purposes. The major formulation Roundup
    may, for instance, be used in pre-plant treatments for seed bed
    preparations, and also against bracken infestations in forestry,
    against couchgrass  (Elytrigia repens) infestations on pastures, in
    direct treatments between rows of crops, or by direct wiping of the
    leaves of the weed, assuming the weeds are taller than the existing
    crop.

         Glyphosate is used worldwide. In 1987, 35 160 ha of the area in
    British Columbia where vegetation management activities took place
    had been treated with Roundup. This was 94% of the total area where
    there were such activities (Ackurst, 1989).

         The application rates of glyphosate are dependent on the
    formulation and type of use. In the Netherlands, recommended rates
    for the application of Roundup are 0.3-2.9 kg a.i./ha. In Canada the
    recommended application rates of Roundup are 1.1-1.7 kg a.i./ha for
    annual weeds and 1.2-5.8 kg a.i./ha for perennial weeds. The
    recommended application rates for Vision in Canadian forestry are
    1.1-2.1 kg a.i./ha (Task Force on Water Quality Guidelines, 1991).
    Glyphosate is generally applied as a 0.5-5% solution in water by
    spraying, and as a 10-50% solution in water by wiping with, for
    instance, a rope-wick (Monsanto, personal communication to the IPCS,
    1991).

         The timing of application is dependent on the use. Application
    in late summer or autumn is recommended for use in forestry in
    Canada (Hildebrand et al., 1982). Application in agriculture may
    be pre- or post-harvest. In the Netherlands, for instance,
    glyphosate may be applied to cereals, potatoes and asparagus
    immediately (up to 7 days) before harvest, but only when the
    ripening is complete. Treatment of immature crops would result in
    higher residue levels, early crop desiccation and reduced yields.

         Glyphosate may be applied in different ways. For large-scale
    treatments aerial application can be appropriate, small-scale
    treatments can be done with spraying equipment on the back or behind
    vehicles, or by wiping equipment.

         Aerial applications will lead to losses due to wind-drift.
    Exposure of flora and fauna due to off-target deposits may take
    place. These downwind deposits depend on the meteorological
    conditions, the plant canopy structure and the application method,
    including the release height (Payne et al., 1989; Feng  et al.,
    1990; Payne, 1992; Payne & Thompson, 1992). The non-volatile
    tank-mix fraction and the speed of the aircraft may influence the
    drop-size spectrum, and it can be expected that dispersal systems
    causing relatively small droplets and having a relatively low
    non-volatile fraction will cause the highest off-target deposits.
    Payne (1992) assumed that the large differences in deposits in two
    comparable experiments were due more to different aircraft airspeeds
    than to different wind speeds. In these experiments the maximum
    deposits at a downwind distance of 50 m were 19 and 3 mg a.i./m2
    at aircraft airspeeds of 45 and 11-20 m/second, respectively. The
    application rate in both experiments was 2.1 kg a.i./ha. In other
    experiments with the same application rate, Payne & Thompson (1992)
    found that the meteorological conditions had a considerable impact
    on the off-target deposition up to 400 m downwind when spraying at
    different wind speeds (2.2-5.7 m/second) and turbulences. The
    deposits at a downwind distance of 400 m varied between 0.001 and
    0.06 mg a.i./m2, whereas they varied between 0.6 and 4 mg
    a.i./m2 at a downwind distance of 50 m. Remarkably, the deposition
    was highest with an intermediate wind speed and intensity of

    turbulence. Payne et al.(1989) investigated the deposits for
    aerial applications of Roundup with different dispersal systems.
    When 2.1 kg a.i./ha was applied with a helicopter in a single
    crosswind swath over 100 ha, up to 13.4 mg a.i./m2 was deposited
    on a downwind distance of 50 m. This maximum deposition was caused
    by a D8-46 hydraulic nozzle, whereas the highest depositions with a
    Thru Valve Boom and a Microfoil Boom were 2 and 0.4 mg a.i./m2,
    respectively. These depositions were also found at a downwind
    distance of 50 m. At the time of application the windspeed 13 m
    above ground level was 0.4-0.5 m/second. Riley et al.(1991)
    modelled spray deposition of glyphosate using results from
    helicopter applications under semi-operational conditions. The study
    was designed to test the appropriateness of a New Brunswick "buffer
    zone" of 65 m to minimize the effects of spray drift. At a distance
    of 65 m, it was estimated that between 3.7% and 5.6% of the nominal
    spray rate was deposited.

    3.1.3  Drinking-water

     Appraisal

          The low mobility of glyphosate in soil would indicate a
     minimal potential for the contamination of drinking-water from
     groundwater aquifers. The only possible source of drinking-water
     contamination is, therefore, surface waters. There have been no
     reported incidences of drinking-water contamination with
     glyphosate.

          Conventional plants for processing of drinking-water would not
     remove glyphosate, but this could be achieved by coprecipitation
     after adding iron salts (AMA van der Linden, personal communication
     to the IPCS, 1991). Ozone, increasingly used as an alternative to
     chlorine in drinking-water treatment, does effectively remove
     glyphosate through the hydroxyl radical (HO. ) chain processes that
     occur in most ozonated waters (Yao & Haag, 1991; Haag & Yao, 1992).

    4.  ENVIRONMENTAL TRANSPORT, DISTRIBUTION AND TRANSFORMATION

    4.1  Transport and distribution between media

     Appraisal

          Following application, glyphosate selectively partitions to
     particulate matter suspended in surface water or to the soil
     substrate. This partitioning is usually rapid and occurred within 14
     days in reported studies. The mechanism of sorption to soil is only
     partially understood. Glyphosate can adsorb to soils through
     phosphate binding sites. Competition with inorganic phosphate has
     been demonstrated in the laboratory but not measured in the field.
     Specific ions (Fe2+,  Fe3+  and Al3+ ) complex glyphosate; metal
     complexes with humic acids in soil may be a main binding mechanism
     for glyphosate in soil. There is little reported information on
     desorption from soil; the data available suggest "strong" binding.
     This is supported by mobility studies which show little leaching of
     glyphosate below the upper few centimetres of the soil profile. The
     major metabolite, AMPA, is also retained in the upper soil layers.

          There is very little information on the bioavailability of
     sediment-bound glyphosate to either aquatic or terrestrial
     organisms. Bioaccumulation and ecotoxicity studies have not,
     generally, been performed with added sediment.

          Applied glyphosate can be translocated in plants. Glyphosate
     in plant foliage or leaf litter does not seem to represent a source
     of contamination of aquatic systems. Animals can ingest the
     herbicide residues in or on plants.

          Dissipation of glyphosate from soil has been widely studied
     with very variable results (DT50  between 3 and 174 days).
     Biodegradation appears to be the major source of dissipation.

          Run-off was minimal in experimental studies, but field results
     suggest that aquatic systems may be receiving glyphosate bound to
     soil particles following rainfall.

         In this chapter the terms biodegradation and dissipation are
    used to distinguish between the decrease of the concentration in,
    for instance, the soil that is due to microbes transforming the
    molecule to a smaller size (biodegradation) and the decrease of the
    concentration that might be due to microbial activity but also to
    other processes, e.g., sorption, leaching and run-off (dissipation).

    4.1.1  Water

         Glyphosate dissipates from the water with 50% dissipation times
    ranging from a few days to 2 weeks (Newton et al., 1984; Monsanto
    1990a; see also Table 5). These DT50 values were deduced from both
    laboratory and field experiments in which sediment or suspended
    particles were shown to be the major sink.

         In water with a near-neutral pH, the formation of an insoluble
    complex of Ca2+ with glyphosate was demonstrated in a laboratory
    experiment (Subramaniam & Hoggard, 1988). It was confirmed with
    X-ray powder diffraction and infrared spectra that this complex was
    not an ionic salt. At a near-neutral pH, the dianionic species of
    glyphosate is dominant. Insoluble complexes have also been found
    with Mg2+, Fe3+ and Cu2+.

         In a field experiment in a temperate coastal rainforest in
    British Columbia, Canada, the highest concentration of glyphosate in
    water was 162 µg/litre (Feng et al., 1990). This maximum was found
    in a directly sprayed tributary 2 h after an aerial application of
    Roundup at a rate of 2 kg a.i./ha. Concentrations in oversprayed
    tributaries without a high cover of overhanging riparian vegetation
    increased after the first rainfall. In oversprayed tributaries with
    a high cover of riparian vegetation almost no residues were found.
    Within 96 h after application the residues in all waters had
    declined below detection limits, indicating rapid dissipation. After
    rainstorms, peak concentrations of glyphosate were found in the
    sediments and on suspended particles of the oversprayed tributaries,
    with maximum concentrations of 7 mg a.i./kg dry weight and 0.06 µg
    a.i./litre unfiltered water, respectively. The amounts in the
    sediments of these waters were variable but declined over time. As
    0.1-2 mg residue/kg dry weight sediment was found between 196 and
    364 days after application, the residues appear to be persistent in
    sediments of oversprayed waters. Feng et al.(1990) concluded,
    therefore, that after rainstorms sediments appear to be the major
    sink.

         In another field experiment in the same forest ecosystem,
    glyphosate dissipated rapidly from a small perennial, very slow
    flowing stream, in a site of 8 ha aerially sprayed with Roundup at a
    rate of 3.3 kg a.i./ha (Newton et al., 1984). In water, 50% of the
    initial concentration had dissipated in 2 days. In sediment, maximum
    concentrations of approximately 0.6 mg a.i./kg were found 14 days
    after application. These were reduced to approximately 0.3 mg
    a.i./kg in 28 days, and to < 0.2 mg a.i./kg in 55 days after
    application. A comparable rapid dissipation from the water column
    was found for small forest ponds in a boreal forest in Manitoba,
    Canada, after applying Roundup at a rate of 0.9 kg a.i./ha
    (Goldsborough & Beck, 1989). The highest concentration in filtered
    water was 141 µg a.i./litre, within 6 h after application. The main

    mechanism of dissipation was probably sorption to the sediment. This
    was confirmed by additional experiments with polyethylene basins
    filled with unfiltered water and sediment that were placed in the
    spray zone. Without sediment, only a very small amount of the dose
    actually applied had dissipated after 30 days, whereas with sediment
    the initial concentrations in the water had decreased by 50%,
    approximately 6 days after application.

         Comparable dissipation patterns were found in a field
    experiment (Monsanto, 1990a) in which Accord (30.5% a.i. w/w) was
    applied at a rate of 4.2 kg Accord/ha on three forestry sites with
    non-flowing pond water and flowing water. Concentrations of up to
    1700 µg/litre filtered water were found in the pond water
    immediately after spraying. The initial concentrations in both pond
    and flowing water were reduced by 50% within 7 days. Concomitantly
    initial AMPA concentrations (maximally 35 µg/litre) were reduced by
    50% within the same period. In flowing water the dissipation of both
    glyphosate and AMPA was even more rapid. Concentrations of
    glyphosate increased up to 19 mg/kg dry weight in the sediment of
    one pond 28 days after application. Concentrations of up to 1 mg/kg
    of both compounds were measured in the sediments of non-flowing
    ponds up to 400 days after application.

         In field experiments in turbid Australian irrigation water,
    glyphosate adsorbed to suspended particles at different rates,
    apparently mainly depending on the application rate (Bowmer  et al.,
    1986). At an initial concentration of 5 mg a.i./litre, 10-16% of the
    load adsorbed to suspended matter, whereas at an initial
    concentration of 0.05 mg a.i./litre, 53-71% adsorbed. In more saline
    water the degree of sorption was less, probably due to rapid
    flocculation. Maximum adsorbed amounts were approximately 7000 mg
    a.i./kg in less saline supply water, and approximately 2500 mg
    a.i./kg in more saline drainage water. When a supply channel was
    emptied before spraying with 3.6 kg a.i./ha for control of aquatic
    weeds, and filled again with water 4 days after the treatment, the
    amount in the unfiltered water used for irrigation was 7% of the
    applied dose.

    4.1.2  Soil sorption

         Glyphosate is readily bound to many soils and clay minerals
    (Sprankle et al., 1975; Hance, 1976; Glass, 1987; Miles & Moye,
    1988b). In laboratory experiments in which glyphosate was added to
    aqueous soil suspensions, the adsorption coefficient Ks/l was
    18-377 dm3/kg in nine soils ranging from sandy loam to peat
    (Hance, 1976), and 33-76 dm3/kg in three soils ranging from sandy
    loam to clay loam (Glass, 1987). These Ks/l values indicate strong
    sorption. In both experiments the sorption could be described by the
    Freundlich equation. Glass (1987) found Ks/l values for the clay
    minerals montmorillonite, illite and kaolinite of 138, 115 and 8
    dm3/kg, respectively.


    
    Table 5.  Biodegradationa of technical grade glyphosate in water and sediments in the laboratory
                                                                                                                                              

    Water type        Sediment    Test    Sediment   Organic      Temperature    pH of      Experimental   Parameter   Time      Reference
                      type        type    (%)        matter in    (°C)           water      time                       (days)
                                                     sediment                               (days)
                                                     (%)
                                                                                                                                              

    Pond water        silty       A       17         0.9          23-25          5.9-7.0    30             DT50        14b       PTRL East
                      clay                                                                                                       Inc. (1990a)
                      loam

    Pond water        silty       An      16         0.9          20-27          5.7-6.5    365            DT50        14c,e     PTRL East
                      clay                                                                                                       Inc. (1990b)
                      loam

    Surface waterd    n.r.        A       9          n.r.         30             8.2-8.6    14             DT50        < 14      Monsanto
                                                                                                                                 (1972a)

    Lake water        sandy       An      33         1.4          30             6.6        42             DT50        22e       Monsanto
                      clay                                                                                                       (1978a)
                      loam
                                                                                                                                              

    a    Biodegradation in the whole system
    b    The biodegradation stopped after approximately 15 days
    c    The biodegradation stopped after approximately 150 days
    d    Three rivers and one lake in the USA
    e    Approximate value derived from data of the author(s)
         A = aerobic; An = anaerobic; n.r. = not reported.
    

         The mechanism of sorption of glyphosate to soil is only
    partially understood. Several factors may be involved. The
    phosphonic moiety adsorbs weakly to unoccupied phosphate binding
    sites and can be displaced by phosphate (Hance, 1976). In laboratory
    experiments with nine soils the author showed that sorption was
    positively correlated with the unoccupied phosphate sorption
    capacity, and not correlated with the total phosphate sorption
    capacity, organic matter, clay, iron or aluminium content. No data
    are available that confirm competition of glyphosate and phosphate
    under field conditions, e.g., after application of artificial
    fertiliser. Miles & Moye (1988b) suggested that the main mechanism
    was probably by H-bonding and ion-exchange, as the degree of
    sorption in their experiments was not correlated with cation
    exchange capacity (CEC) values or surface areas. Contrary to the
    results of Miles & Moye (1988b) and of Hance (1976), sorption
    appeared to be correlated with CEC values and clay content in a
    sorption study with clay loam, silt loam and sandy loam (Glass,
    1987).

         The binding is also influenced by the presence of specific
    cations. Hensley et al.(1978) demonstrated that Fe2+, Fe3+
    and Al3+ inactivated glyphosate much more than Ca2+, K+ and
    Na+. This was confirmed by Glass (1987) and Sprankle  et al.
    (1975). Glass (1987) suggested that glyphosate was complexed by
    cations, released from cation-saturated clays via a cation-exchange
    with solution protons.

         According to Heinonen-Tanski (1989), most of the soil-bound
    residues of glyphosate were recovered in the fulvic acid fraction
    (21-33%). Sorption of glyphosate to fulvic acids was also reported
    by Madhun et al.(1986), who added 14C-glyphosate to an aqueous
    soil extract (ASE) of peat. In this study sorption was mainly on ASE
    fractions with a relative molecular mass ¾ 1000. Piccolo  et al.
    (1992) studied the interaction of glyphosate with a pure iron-humic
    acid complex. Maximum adsorption values indicated that adsorption to
    the complex occurred to as great an extent as to whole soils. This
    suggested that humic acid complexes with polyvalent cations might
    represent a main binding substrate for glyphosate in soils. There
    was no desorption of bound residues of glyphosate following shaking
    with 0.01 mol CaCl2/litre solution for 48 h, the maximum shaking
    time for the adsorption studies.

         Desorption of glyphosate with ionized water from
    montmorillonite and illite needed three days before reaching an
    equilibrium in a study of Miles & Moye (1988b).

         It can be concluded that sorption of glyphosate can be expected
    in the presence of available phosphate binding sites, the presence
    of iron and aluminium (oxides or hydroxides), and appropriate
    combinations of clay and organic matter.

    4.1.3  Mobility in soils

         In view of its Ks/l, glyphosate can be expected to be
    immobile or slightly mobile in many soils. This was confirmed by
    several experiments, both in the laboratory and in the field. In
    thin-layer chromatography studies with sandy loam, clay loam and
    sandy clay loam, the Rf values of 14C-glyphosate were 0.14-0.20
    (Sprankle et al., 1975). In comparable studies with silt loam,
    silty clay loam, and sandy loam Rf values were < 0.2 (Monsanto,
    1972c). In a leaching study with columns of 30 cm and a high water
    flux of 51 cm over less than 2 days, < 0.1-6.6% of the applied
    activity was leached (Monsanto, 1978b). This experiment was
    performed with eight soils, ranging from sandy loam (organic matter
    content 0.7%) to volcanic ash (organic matter content 9.5%). More
    than 90% of the applied activity was recovered in the upper 0-14 cm
    layer.

         Only one leaching study under laboratory conditions with
    respect to the mobility of AMPA has been reported. In this
    experiment with 30-day-old residues, < 0.1-1.6% of the applied
    activity was leached over 45 days (Monsanto, 1978b). The columns
    were 30 cm and the water flux over 45 days was low (17 cm). The
    amount of AMPA that was recovered after 45 days in the upper 0-2 cm
    layer was low (0.5-12% of the applied activity), due to a high rate
    of mineralisation.

    4.1.4  Dissipation from the soil in the field

         Many field experiments on the dissipation of glyphosate from
    the soil have been performed. Some relevant studies are summarized
    in Table 6. They indicate DT50 values based on dissipation that
    range from 3 to 174 days depending on edaphic and climatic
    conditions. In a forest brush ecosystem in Oregon, USA, the DT50
    value in loam was 29 days with and 40 days without litter (Newton
    et al., 1984). In field experiments in Sweden, Roundup was sprayed
    over reforestated sites (Torstensson et al., 1989). In the soils
    of these sites the DT50 values were < 50 days, apparently
    depending on the soil respiration rate. The dissipation consisted of
    a fast first, and a much slower second phase, especially in sites in
    northern Sweden, which was possibly due to a longer frost period. In
    these sites 1-2% of the actually applied dose was recovered 1080
    days after application. A comparable dissipation pattern was found
    in a field experiment on Finnish agricultural soils (Heinonen-Tanski
    et al., 1985). In this experiment 25% of the concentration in a
    sandy loam 2 days after the treatment was recovered one year after
    application. The application rate was 1.4 kg a.i./ha.

         A study in a temperate coastal rain forest in British Columbia,
    Canada, showed that, 360 days after application, 6-18% of the
    initial levels was recovered (Feng et al., 1990). In this
    experiment Roundup was applied at a rate of 2 kg a.i./ha. The soils

    were alluvial sandy loam or sandy clay loam with highly organic
    surface horizons. Some of these soils were well drained, others were
    seasonally flooded. At each sampling time more than 90% of the
    recovered residues was in the upper 0-15 cm layer. Under all
    conditions the amount of glyphosate declined over time, whereas
    there was a transient increase of AMPA.

         In other field experiments on boreal forest soils, comparable
    dissipation patterns were found. Stark (1983) reported DT90 values
    of 30-720 days, and Roy et al.(1989b) found a DT50 value of
    approximately 20 days on a sandy soil planted with jackpines  (Pinus
     banksiana). In the field experiments of Roy et al.(1989b),
    glyphosate was detectable up to 335 days after application; almost
    all residues in the sandy soil were recovered in the organic top
    layer. In field experiments of Monsanto (1990a) in three forest
    locations in the USA, the concentration course of glyphosate
    appeared to be rather irregular, especially during the first four
    months. However, 50% of the initial concentrations in the soil had
    mostly dissipated within 120 days. One clear exception was a site in
    Corvallis in which glyphosate increased up to 0.15 mg/kg dry weight,
    180 days after application. On the same site AMPA increased up to
    0.32 mg/kg, 346 days after application. The application rate in
    these experiments was 4.2 kg Accord/ha.

         On a clay soil of a clear-cut boreal forest area, Roy  et al.
    (1989b) found no dissipation of glyphosate due to run-off on a 8°
    slope. In a field experiment on agricultural soils without
    conventional tillage, the dissipation of glyphosate due to run-off
    on 6-16° slopes was < 1% of the applied dose when 1.1-3.4 kg
    a.i./ha was applied (Edwards et al., 1980). However, when 9.0 kg
    a.i./ha was applied, 1.8% of the applied dose dissipated due to
    run-off, mainly because of a rainstorm shortly after application.

    4.1.5  Uptake and dissipation from plants

         Uptake of 14C-glyphosate by leaves of trembling aspen
    seedlings  (Populus tremuloides) was initially rapid, after which
    it slowed down (Sundaram, 1990). The seedlings were exposed to
    Roundup that was dripped with a micro-applicator on some central
    leaves. The application rate was 0.35 kg a.i./ha leaf surface area.
    Most activity was washable from the leaves (61-77%), and 22-28% was
    recovered in the treated leaves within 48 h. As only 1-10% was
    recovered in the other parts of the seedlings, this indicated a
    rather low translocation after absorption. A rapid uptake of
    14C-glyphosate within a few hours was indicated for sugar beets
     (Beta vulgaris), when applied to a mature leaf (Gougler & Geiger,
    1981). 14C-glyphosate probably entered the phloem in a
    non-facilitated way. The subsequent transport through the phloem


    
    Table 6.  Biodegradation and dissipation of glyphosate in soils
                                                                                                                                              

    Soil type          Compound   Test    Moisture   Temperature   pH        Organic    Experimental   DT50      Reference
                                  type    content    (°C)                    matter     duration       (days)
                                          (%)                                (%)        (days)
                                                                                                                                              

    Biodegradation

    Sandy loam         Tgg        L,A     14-16      25            7.3        2.8       360            2b        PTRL East Inc. (1991)
    Silt loam          Tgg        L,A     12-14      25            7.5        1.0       360            2b        PTRL East Inc. (1991)

    Dissipation

    Sandy loam         Tgg        G       11         32            5.7        1.0       112            130b      Monsanto (1972b); Rueppel
                                                                                                                 et al. (1977)
    Silt loam          Tgg        G       11         32            6.5        1.0       112            3b        Monsanto (1972b); Rueppel
                                                                                                                 et al. (1977)
    Silty clay loam    Tgg        G       11         32            7.0        6.0       112            25-27b    Monsanto (1972b); Rueppel
                                                                                                                 et al. (1977)
    Sand               Ru         F       n.r.       n.r.          3.5-3.7   40         762            approx    Roy et al. (1989b)
    (humoferric                                                                                        20a
    podsol)
    Sandy loam,        Ru         F       n.r.       n.r.          4.2-4.9   15-31      360            45-60b    Feng & Thompson (1990)
    sandy clay loam
    Loam               Ru         F       n.r.       n.r.          4.0-4.7   3.8-5.2    55             29-40b    Newton et al. (1984)
    Loamy sand         Ru         F       n.r.       n.r.          n.r.      0.8        370            3-4b      Monsanto (1983a)
    Sandy clay loam    Ru         F       n.r.       n.r.          n.r.      7.0        370            122-174b  Monsanto (1983a)
                                                                                                                                              

    a  Based on data of the author(s)             b  Data reported by the author(s)
    L = laboratory study; F = field study; G = greenhouse study; A = aerobic; An = anaerobic;
    Tgg = technical grade glyphosate; Ru = Roundup; n.r. = not reported
    

    appeared to be according to an "intermediate permeability
    mechanism". When exposed for a longer time, plants may show
    substantial translocation of absorbed 14C-glyphosate, as was shown
    for potatoes  (Solanum tuberosum) by Smid & Hiller (1981). In the
    treated leaves of the potatoes 45% of the absorbed activity was
    recovered, whereas the rest was mainly translocated to the apical
    meristem and the roots. Up to 5% was recovered in the mother tuber.
    The degree of translocation was age-dependent, as older plants
    showed less translocation than younger plants.

         The uptake of glyphosate by red raspberries  (Rubus strigosus)
    was 9% of the amount that was deposited on the leaves after spraying
    Roundup at a rate of 2 kg a.i./ha (Roy et al., 1989a). In the same
    field experiment the uptake was 14% by wild blueberries  (Vaccinium
     myrilloides). Most glyphosate was recovered in the washings, which
    was also found under laboratory conditions. The initial absorbed
    amounts were 0.92-2.0 mg a.i./kg dry weight. The absorbed and
    washable amounts together were reduced by 50% within 13 days in the
    raspberries and within 20 days in the blueberries. AMPA was
    detectable up to 33 days after application. Metabolism occurred to
    only a minor extent as AMPA concentrations were less than 1.5% of
    the concurrent concentrations of glyphosate (similar results were
    reported by FAO/WHO, 1986b). In a field experiment by Feng &
    Thompson (1990) in a temperate coastal rainforest in British
    Columbia, Canada, the main target species for treatment with Roundup
    were red alder  (Alnus rubra) and salmonberry  (Rubus spectabilis).
    Immediately after spraying, the concentrations in leaf tissue were
    up to 448 mg a.i./kg dry weight. Glyphosate dissipated rapidly from
    the leaf litter with a DT50 value of 8-9 days. The leaf litter
    included leaves directly exposed on the trees and existing leaf
    litter from natural defoliation before treatment with Roundup. The
    authors assumed that leaf litter of these major brush species is an
    insignificant source of glyphosate input into streams or onto forest
    floor, because of the fast dissipation. A rapid dissipation of
    glyphosate from fresh foliage was also found in a field study
    (Monsanto, 1990a) in which initial concentrations of up to 1300 mg
    a.i./kg and 2.6 mg AMPA/kg decreased rapidly. A transient
    accumulation of glyphosate and AMPA was found in the leaf litter on
    some sites, but these amounts were reduced by approximately 90%
    within 100 days.

         Glyphosate dissipated completely from wild berries  (Vaccinium
     vitis-idaea, Vaccinium myrtilus) within one year in a field
    experiment in Finland in which Roundup was applied at a rate of
    0.25-2.2 kg a.i./ha with a knapsack sprayer (Siltanen  et al.,
    1981). Contrary to this dissipation pattern was that of glyphosate
    in reindeer lichens  (Cladonia rangiferina) that were sampled in
    the same experimental plots. Around 270 days after application,
    dose-related concentrations of glyphosate and AMPA were recovered in

    lichens with maxima of 45 and 2.1 mg/kg for glyphosate and AMPA,
    respectively. Approximately 390 days after application of 0.8 kg
    a.i./ha, 6.4 and 0.3 mg/kg of glyphosate and AMPA were still
    detectable.

    4.1.6  Ingestion by animals

         As the concentration in the foliage may increase up to high
    amounts immediately after application, this implies the possibility
    of entry into the food chain through ingestion by herbivorous or
    omnivorous animals. This was confirmed by Sullivan & Sullivan (1979)
    who investigated the effects of glyphosate on the feed preference
    and daily chow consumption of black-tailed deer  (Odocoileus
     hemionus columbianus). These herbivores did not avoid eating
    browse of alder  (Alnus rubra) and alfalfa  (Medicago sativa) that
    was treated with glyphosate at a rate of 2.2 kg/ha. Sometimes the
    treated alder browse was even preferred. Reindeer may be exposed to
    glyphosate, since reindeer lichens, which are an important food
    source, can take up a substantial amount of glyphosate (see above).

    4.2  Abiotic degradation

    Appraisal

          Hydrolysis of glyphosate is very slow. Photodegradation in the
     field may occur.

    4.2.1  Hydrolytic cleavage

         Hydrolysis of glyphosate in sterile buffers is very slow. After
    32 days < 6.3% of the applied activity was recovered as AMPA,
    after applying 14C-glyphosate at rates of 25 and 250 mg/litre to
    aqueous buffer solutions of pH 3, 6 and 9 (Monsanto, 1978b). These
    tests were performed at both 5 and 35 °C.

    4.2.2  Photodegradation

         Photochemical degradation in water may occur under both
    laboratory and field conditions, mainly depending on the type of
    light source. In sterile aqueous buffers of pH 5, 7, and 9, less
    than 1% of the applied dose was degraded (photodecomposition of
    14C-phosphonomethyl-labelled glyphosate) during 29-31 days, when
    exposed to sunlight (PTRL Inc., 1990).

         Lund-Hoie & Friestad (1986) exposed Roundup to several light
    sources under different conditions. When exposed to UV light (lambda
    = 254 nm) under laboratory conditions, concentrations of 1 and
    2000 mg a.i./litre in deionized water showed DT50 values of 4 and 14
    days, respectively. When exposed to sunlight under field conditions
    1 mg a.i./litre in polluted water without sediment showed a much

    slower decomposition (DT50 > 63 days), possibly due to pollution
    preventing adequate UV penetration in the water. Polluted water with
    sediments showed a rapid dissipation from water, probably due to
    adsorption onto the sediments. In another field experiment 2 and
    100 mg a.i./litre in deionized or polluted water without sediment
    showed DT50 values of < 28 days, when exposed to sunlight. At
    the low concentration the dissipation in polluted water was more
    rapid than in deionized water. In the dark no dissipation occurred.

         In laboratory experiments 1 mg/litre of glyphosate in
    sterilized natural and deionized water showed DT50 values of 4 to >
    14 days when exposed to artificial light (350-450 nm) in
    photoreactors without sediment (Monsanto, 1978a). In these
    experiments Ca2+ acted as a photosensitizing agent.

         Photodegradation by sunlight of glyphosate applied to a soil
    appeared to be an insignificant route of dissipation (PTRL Inc.,
    1989). In this study, 14C-glyphosate mixed with unlabelled
    glyphosate was exposed for 31 days to natural sunlight, after
    application to a sandy loam at a rate of 4.5 kg a.i./ha.
    Extrapolated DT50 values that were based on first-order kinetics
    were 90 days in the sunlight and 96 days in the dark, indicating no
    substantial degradation due to photolysis. The temperature of the
    soil surface was 22-23 °C. Under unnatural light conditions
    glyphosate appeared not to be photodegraded substantially (Monsanto,
    1972c; Rueppel et al., 1977; Monsanto, 1978a).

    4.3  Biodegradation

    Appraisal

          Selected studies of the biodegradation of glyphosate have been
     considered; selection was on the basis of test conditions and modern
     methodologies. There is considerable variation in rate of breakdown
     in water, aquatic sediment and soil. Degradation occurs more rapidly
     in aerobic than anaerobic conditions. Half-times for biodegradation
     in the three media under laboratory conditions range between a few
     days and approximately 20 days. No data on biodegradation under
     anaerobic conditions are available.

          The main route of biodegradation of glyphosate appears to be
     by splitting the C-N bond to produce AMPA. However, a second route
     with splitting of the C-P bond can also occur.

          A range of bacterial strains can degrade glyphosate. Bacteria
     capable of using the compound as sole phosphorus, sole carbon or
     sole nitrogen source have been identified. Growth is slow compared
     to growth on inorganic sources of P, C or N. There is evidence from

     the field that bacterial populations adapt to the metabolism of
     glyphosate. Presence of inorganic phosphate inhibits degradation of
     glyphosate with some, but not all, bacteria. Biodegradation of
     glyphosate may involve co-metabolism.

         The most relevant laboratory experiments in which the
    biodegradation in systems with water and sediment have been studied
    are summarized in Table 5. These studies indicate that the rate of
    biodegradation may vary substantially, depending on experimental
    conditions, e.g., the availability of oxygen, temperature and type
    of sediment. The time needed for 50% biodegradation of glyphosate in
    the whole system of a test with water and sediment is < 14 days
    under aerobic and 14-22 days under anaerobic conditions in the
    laboratory.

         In the experiments of PTRL East Inc. (1990a,b), less then 10%
    of the applied activity was recovered in the pond water over a
    period of 30 days under aerobic condition and 365 days under
    anaerobic conditions. During all experiments more than 50% of the
    applied activity was recovered in the sediment.

         In experiments with water and their associated sediments the
    amount of a.i. declines over time with a generally transient
    increase of 14C-AMPA, an increase of 14CO2, and an increase of
    sediment-bound residues. An exception to this pattern of
    biodegradation can be observed in some aerobic and anaerobic
    experiments that were performed with pond water and a silty clay
    loam sediment (PTRL East Inc, 1990a,b). In this water/sediment
    system the biodegradation stopped after approximately 15 days under
    aerobic conditions and after approximately 150 days under anaerobic
    conditions. The glyphosate residues (a.i. plus AMPA) at both time
    points remained approximately 40% of the applied dose, which
    indicated substantial persistence in spite of the rapid initial
    degradation.

         AMPA is the main metabolite of glyphosate found in both the
    water column and the sediment. Maximum amounts of AMPA under both
    aerobic and anaerobic conditions in the sediment were 25% of the
    applied activity (PTRL East Inc., 1990a,b). These maxima were found
    at 7-20 days after application. In the same experiments maximum
    amounts of sediment-bound residue were 9% of the applied activity
    under aerobic conditions and 4% under anaerobic conditions. These
    maxima were found at the end of the experiments. The amounts of
    evolved 14CO2 in these studies gradually increased in most cases
    up to 24 and 35% of the applied activity after 30 days (aerobic),
    and 365 days (anaerobic), respectively. This indicates substantial
    differences in the mineralization rate. These differences are partly
    due to the availability of oxygen, since under anaerobic conditions
    the mineralization rate was slower than under aerobic conditions.

    This was also found by Monsanto (1972a, 1978a). In the aerobic
    experiments of Monsanto (1972a), four sediments that differed by up
    to two orders of magnitude in the total number of micro-organisms
    did not show substantial differences in mineralization rate.

         Biodegradation studies with glyphosate in the soil under
    conditions where unequivocal interpretation is justified are scarce.
    Table 6 summarizes some relevant studies, indicating that the
    biodegradation rate may differ substantially, depending on the
    experimental conditions. The laboratory and greenhouse experiments
    in Table 6 were performed with moisture contents (> 75% of the
    field capacity) that were adequate for optimal biodegradation.

         In most laboratory experiments the biodegradation rate of
    glyphosate in soils appears to be rapid (see Table 6). Mostly
    biodegradation can be described with linear first-order kinetics.

         Sometimes a non-linear first-order model taking into account
    spatial variability better describes the results observed (PTRL East
    Inc., 1991):

              C = C0 (1 + ßt)-alpha

         C in this equation is the concentration in the soil at time t,
    C0 the initial concentration, and alpha and ß are rate constants
    reflecting spatial variability.

         The main metabolite under aerobic conditions of glyphosate in
    soil is AMPA. In aerobic laboratory experiments the maximum amounts
    in sandy loam and silt loam were 27 and 29%, respectively, of the
    applied activity. These maxima were reached 14 days after
    application (PTRL East Inc., 1991). From the data of PTRL East Inc.
    (1991), DT50 values for AMPA of approximately 50 days in sandy and
    silty loam can be derived. That AMPA is more persistent than
    glyphosate was also shown in a laboratory experiment with sandy loam
    (Monsanto, 1972b). The amounts of AMPA after 111 days were 10-17% of
    the applied activity. In this study, the temperature (32 °C) was
    higher than in the other studies discussed above.

         Some minor unidentified metabolites were quantified in an
    aerobic laboratory experiment lasting 364 days with sandy loam and
    silt loam (PTRL East Inc., 1991). Two unknown metabolites did not
    exceed 3.5% of the applied activity, whereas some other unknown
    metabolites did not exceed 1.5% each. Rueppel et al.(1977)
    quantified some minor metabolites that did not exceed 1% of the
    applied activity. These metabolites were
     N-methylamino-methylphosphonic acid, glycine,
     N,N-dimethylaminomethylphos-phonic acid, hydroxymethylphosphonic
    acid, and two unknown metabolites.

         In aerobic laboratory experiments, the amounts of soil-bound
    residues immediately after application were 9-35% of the applied
    dose, after which they showed an irregular time-course during these
    experiments of approximately 112 days (Monsanto, 1972b). In general,
    the initial amounts were also the maximum amounts. In other
    laboratory experiments however, maximum amounts of soil-bound
    residues appeared to be reached after 14 days, whereafter they
    remained more or less constant or even decreased (PTRL East Inc.,
    1991). These maximum amounts were 7-9% of the applied activity, and
    were probably lower compared with other studies due to better
    extraction procedures.

         Mineralization in the soil occurs under both aerobic and
    anaerobic conditions in the laboratory, although the rates may
    differ greatly, apparently mainly depending on the soil respiration
    rate and the temperature. When 14C-phosphonomethyl-labelled
    glyphosate was applied to sandy loam and silt loam, 70-78% 14CO2
    evolved during an aerobic laboratory experiment of 360 days (PTRL
    East Inc., 1991). In this study the application rate was 4 mg
    a.i./kg dry weight. In an aerobic laboratory study with 15 Swedish
    forest soils, DT50 values based on 14CO2 evolution varied
    between 6 and 200 days. Mineralization was highly correlated with
    the soil respiration rate, but not with pH or organic matter content
    (Torstensson & Stark, 1981). This was confirmed by Torstensson &
    Stenström (1986) and Heinonen-Tanski (1989). Torstensson & Stenström
    (1986) reported that glyphosate was co-metabolized. In this case,
    co-metabolizing microorganisms are not supplied with energy by
    biodegrading glyphosate.

         Establishing the correlation between soil respiration and
    mineralization requires both a standardized measurement of the
    respiration rate and an accurate measurement of the actual dose that
    reaches the soil (Torstensson & Stenström, 1986). In a laboratory
    experiment simulating temperatures under arctic conditions in forest
    soils, 51-71% of the applied activity was recovered as 14CO2 217
    days after application of 14C-glyphosate. In this study the
    mineralization rate was reduced 10-15 times during a temperature
    decrease of 10 °C over the first part of the study. The rate
    increased only 3.7-4 times with a temperature increase of 10 °C
    during the second part (Heinonen-Tanski, 1989).

         Glyphosate in the soil appears to be degradable by
    micro-organisms in two ways (Jacob et al., 1988), as shown in
    Fig. 3. One route is via the formation of AMPA and a C2 fragment
    which might be glyoxylate. This scheme for degradation was proposed
    by many researchers (Monsanto, 1972b; PTRL East Inc., 1991). In this
    route the splitting of the C-N bond is the first step. There is,
    however, another route of biodegradation via sarcosine
     (N-methyl-glycine) and orthophosphate, after which sarcosine is

    degraded to glycine and a one-carbon unit that eventually might form
    CO2 via formaldehyde (Kishore & Jacob, 1987; Jacob et al., 1988).
    In this route the splitting of the C-P bond is the first step. In
    experiments with 14C-glyphosate, isolated cultures of  Pseudomonas
    sp. strain LBr were able to degrade glyphosate according to both
    routes (Jacob et al., 1988). Approximately 5% of the applied
    14C-glyphosate was not degraded via AMPA, but via sarcosine.

         The growth rate of bacteria isolated from a sandy loam garden
    soil that was sprayed with Tumbleweed (a garden product) was less
    inhibited by technical grade glyphosate than the growth rate of
    bacteria from an unsprayed control (Quin et al., 1988). This
    indicated adaptation of the bacterial populations of the sprayed
    site. As addition of aromatic amino acids prevented growth
    inhibition in the population of the unsprayed site to a greater
    extent than in the population of the sprayed site, different
    mechanisms of biochemical interference were indicated. The
    composition of the bacterial population on the unsprayed site was
    also different from the sprayed one.  Pseudomonas sp. and
    lactose-fermenting bacteria could be identified in an inoculum from
    the sprayed soil able to use glyphosate as a sole source of
    phosphorus (Quinn et al., 1988). A different regulatory mechanism
    for biodegradation in unsprayed and sprayed sites was assumed: in
    the latter the aromatic amino acid pathway might be regulated by
    direct control of 3-deoxy-D-arabino-heptulosonate-7-phosphate (DAHP)
    by the end-products, whereas in the unsprayed site DAHP synthase
    might be indirectly regulated by prephenate. Also in other
    experiments bacteria were shown to use glyphosate as a sole P source
    (Kishore & Jacob, 1987; Pipke & Amrhein, 1988; Weidhase et al.,
    1990), thereby primarily degrading glyphosate to orthophosphate and
    sarcosine, by splitting the C-P bond. In the study of Weidhase et
    al. (1990), 18.2% of the applied activity was recovered as sarcosine
    8 h after application of 14C-1-methyl-labelled glyphosate to a
    pure culture of  Pseudomonas sp. GS. This biodegradation route of
    glyphosate via sarcosine was also demonstrated by Kishore & Jacob
    (1987). In their experiments with glyphosate as sole P source for
     Pseudomonas sp. PG2982, one hour after application of
    14C-labelled glyphosate, glycine, phosphate, and a one-carbon
    unit, possibly formaldehyde, were identified as metabolites. After
    one hour, the 14CO2 evolution when the phosphonomethyl moiety
    was labelled was substantially higher, as compared with the 1- or
    2-glycine-labelled moieties. The authors suggested that the
    so-called phosphate-starvation-inducible proteins, as identified by
    others, might be responsible for cleaving the C-P bond. In an
    experiment with pure cultures of a mutant of  Arthrobacter sp.
    GLP-1 able to use glyphosate as a sole P source, 90% of the applied
    activity was released as orthophosphate at 240 h after application
    of 14C-1-methyl-labelled glyphosate (Pipke & Amrhein, 1988).
    Orthophosphate inhibited further biodegradation of glyphosate.
     Flavobacterium sp. was found by Balthazor & Hallas (1986) to be

    able to degrade glyphosate in spite of the presence of
    orthophosphate. Liu et al. (1991) showed that 12 strains of bacteria
    from the family  Rhizobiaceae could degrade glyphosate present in the
    medium as the sole phosphorus source; although growth of the
    bacteria was slower than with inorganic phosphate. Sarcosine was the
    intermediate breakdown product, indicating initial cleavage of the
    C-P bond, in  Rhizobium meliloti, the strain used for detailed
    metabolic studies.

    FIGURE 3

         Carlisle & Trevors (1986a) deduced from their experiments that
    nitrate-reducing bacteria are involved in metabolizing glyphosate.
    Involvement of nitrifying bacteria in the biodegradation of
    glyphosate was also demonstrated by Murthy et al. (1989), when they
    investigated the treatment of waste water from a Roundup formulating
    factory.

          Pseudomonas sp. may use glyphosate as a sole P or C source,
    as demonstrated by Weidhase et al. (1990). Only slight growth of the
    wild-type strain of the bacterium  Pseudomonas fluorescens was
    observed with glyphosate as sole carbon or nitrogen source. The
    herbicide was metabolized to aminomethylphosphonate (Zboinska
    et al., 1992). Murthy et al. (1989) isolated a denitrifying
    bacterial species that was also able to use glyphosate as a C
    source. This species was isolated from activated sludge in a
    waste-water treatment plant. A mutant of  Arthrobacter sp. strain
    GLP-1 was able to utilize glyphosate as a sole N source, whereas
    this was not possible for the normal strain (Pipke & Amrhrein,
    1988), probably due to the uptake of inorganic P released during
    biodegradation.

         As the Biological Oxygen Demand and the Chemical Oxygen Demand
    of glyphosate are < 2 mg/g and 0.53 g/g, respectively, glyphosate
    cannot be considered as readily biodegradable (LISEC, 1990a,b). In
    suitable systems, however, glyphosate is biodegradable, as shown by
    Murthy et al. (1989), who investigated the biodegradation of
    glyphosate in waste-water treatment plants under different
    conditions in sequencing batch reactors on a laboratory scale. These
    reactors were fed with waste water from a Roundup manufacturing
    facility. Glyphosate was degraded completely within one cycle of
    24 h, independent of whether there was an initial aerated or anoxic
    phase of 4 h. However, more glyphosate could be processed with an
    anoxic initial phase, probably due to better conditions for
    denitrification. Not only denitrifiers but also ammonifiers and
    nitrifiers appeared to be involved in the biodegradation of
    glyphosate. Only at the very high concentration of approximately
    5000 mg a.i./litre was biodegradation repressed by non-glyphosate
    COD and inhibited by excess ammonia production.

          Pseudomonas sp. strain LBr,  Flavobacterium sp. and a
    denitrifying bacterial species were isolated from activated sludge
    as species with the ability to use glyphosate as a P source
    (Balthazor & Hallas, 1986; Jacob et al., 1988; Murthy et al., 1989).
    The denitrifier was also able to use glyphosate as a sole C source.
     Flavobacterium sp. degraded glyphosate to AMPA in both the
    presence and absence of PO43- (Balthazor & Hallas, 1986). In
    this experiment the further degradation of AMPA appeared to be
    hampered in the presence of PO43-.

          Pseudomonas sp. strain LBr was capable of completely
    eliminating amounts of glyphosate up to 3212 mg/litre from a growth
    medium (Jacob et al., 1988).

         Continuous exposure of an activated sludge treatment system in
    a pilot plant increased the ability of the sludge to metabolize
    glyphosate to AMPA (Hallas et al., 1992). In this trial an influent
    concentration of 50 mg a.i./litre was reduced to less than 5 mg
    a.i./litre under continuous-flow conditions with an average
    residence time of 10 min. The sludge was inoculated with immobilized
    bacteria capable of degrading glyphosate. The effectiveness of the
    treatment was dependent on the presence of a nitrogen source and a
    non-glyphosate carbon source, and required a pH range of 6.0 to 8.0.

         No data are available on the amounts of glyphosate that can be
    eliminated in conventional waste-water treatment plants under
    practical conditions. In waste water from glyphosate-producing
    plants, 28-45% is reported to be eliminated through biological
    treatment (Task Force on Water Quality Guidelines, 1991).

         No data are available on the biodegradability of the
    surfactants in formulations. It is, however, probable that
    polyoxyethylene amine is biodegraded fairly rapidly in view of the
    biodegradability of structurally related compounds (Swisher, 1987).

    4.4  Bioaccumulation

    Appraisal

          Glyphosate is not expected to bioaccumulate in view of its
     high water solubility and its ionic character. This was confirmed by
     several laboratory experiments with fish, crustaceans and molluscs
     and by field experiments.

         In a static test, channel catfish  (Ictalurus punctatus) were
    exposed to 0.94-0.99 mg 14C-labelled a.i./litre (actual
    concentrations) for 10 days (ABC Inc, 1981d; Monsanto, 1981a). Of
    the absorbed amount, 76% was recovered in the viscera. More than 90%
    of the extractable residues in the viscera and the fillet was
    identified as glyphosate, whereas less than 2% was identified as
    AMPA. After 10 days of depuration 80% of the absorbed activity was
    eliminated. For exposed channel catfish the calculated
    bioconcentration factor based on the activity absorbed by the whole
    fish was 0.27. For depurated channel catfish the calculated
    bioconcentration factor was 0.052.

         The marsh clam  (Rangia cuneata) and crayfish  (Procambarus
     simulans) were exposed in static tests lasting 28 days to
    synthetic uncontaminated sea water and a sandy loam sediment that
    was incorporated with 3 mg 14C-labelled a.i./kg (ABC Inc.,
    1982d,e). These experiments were set up to assess the degree of
    bioconcentration of glyphosate when used in flooded rice levees and
    tidal water. The calculated bioconcentration factor for the edible

    parts of the clam increased during exposure up to 4.8, whereas for
    the whole crayfish it increased up to 12. The highest concentrations
    in the edible parts of the clam and the whole crayfish were 0.3 mg
    14C-labelled residues/kg for both. After 28 days of depuration 48%
    of the accumulated residues were eliminated from the edible parts of
    the clam. The concentration in these parts was then 0.16 mg
    14C-residues/kg. The crayfish finally had eliminated 91% after 14
    days of depuration. The concentration in the whole crayfish was then
    0.02 mg 14C-residues/kg. It must be stated that this test refers
    to the accumulation of 14C and not glyphosate.

         In a static test without sediment, in which rainbow trout
     (Salmo gairdnerii) were exposed to 2 mg a.i./litre (nominal
    concentration) for 12 h, the fillets of the fish contained 80 µg
    a.i./kg (in the original article the erroneous figure of 80 mg/kg
    was reported), and the eggs 60 µg a.i./kg (Folmar et al., 1979).
    This indicates a bioconcentration factor of 0.04 for the edible
    parts. Roundup was applied in this test.

         In a flow-through test in which bluegill sunfish  (Lepomis
     macrochirus) were exposed to 11-13 mg 14C-labelled a.i./litre
    (actual concentrations) for 35 days, calculated daily
    bioconcentration factors based on the whole fish increased from <
    0.1, 0.2 days after the start of the test, to 0.4-0.5 at the end
    (ABC Inc., 1989f). Maximum concentrations in the whole fish, viscera
    and fillet were 13, 7.6 and 4.8 mg 14C-residues/kg, respectively.
    The time required to reach 90% of the steady state and the uptake
    rate constant were calculated to be 120 days and 0.02 mg/kg fish x
    (mg/litre water)-1 x day-1, respectively. During 21 days of
    depuration, the half-life of depuration was calculated to be 35. A
    slow decrease in tissue concentration during depuration was
    indicated. After the period of depuration 2.2 mg 14C-residues/kg
    whole fish was still present. In an additional study to characterize
    the 14C-residues, 95-97% of the residues in the water was
    glyphosate, whereas in the whole fish and tissues 28-91% of the
    recovered activity was glyphosate (ABC Inc., 1989g). In a whole fish
    sample 21 days after starting the test, 49% of the recovered
    activity was found to be AMPA. By treating homogenates with
    proteinase K it was indicated that a substantial amount of the
    absorbed residues was tightly associated with, or incorporated into,
    protein.

         In a field experiment in a forest ecosystem in Oregon, USA,
    neither glyphosate nor AMPA were recovered in salmon fingerlings
     (Oncorhynchus kisutch) after aerial application of Roundup at a
    rate of 3.3 kg a.i./ha (Newton et al., 1984). The fingerlings were
    released at the downstream edge of the sprayed site and analysed up
    to 55 days after treatment. Glyphosate was not recovered in carp
     (Cyprinus carpio) in a field experiment in which ponds were
    sprayed with Roundup at rates of 1.3-1.4 kg a.i./ha (Monsanto,
    1980). In this experiment of approximately 90 days, AMPA was not

    recovered until 30 days after application. It then increased up to
    0.21 mg/kg whole fish, remained constant for another 30 days, and
    then decreased to around the limit of determination (0.1 mg/kg) at
    the end of the experiment.

         In a forest ecosystem in Oregon, USA, Roundup was aerially
    applied at a rate of 3.3 kg a.i./ha (Newton et al., 1984).
    Concentrations in mammals were of the same order of magnitude as the
    concentrations in litter and ground cover. The concentrations of
    glyphosate in the viscera of herbivorous small mammals decreased
    more slowly than in omnivorous and carnivorous small mammals, which
    was probably due to a higher ingestion of contaminated litter. The
    highest concentration was found in the viscera of omnivorous
    deermice  (Peromysces maniculatus) immediately after spraying: 5 mg
    a.i./kg. Only small traces of AMPA were found in mammalian viscera.

    4.5  Waste disposal

         Small amounts of glyphosate can be disposed of by mixing with
    alkali and soil prior to burial in a pit or trench, whereas large
    amounts should be incinerated in units equipped with effluent gas
    scrubbing (IRPTC, 1991).

    5.  ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE

    Appraisal

          The low toxicity, low volatility and low body absorption of
     glyphosate makes its application by backpack sprayer safe under
     field condition provided that the worker wears full protective
     clothing.

    5.1  Environmental levels

         A synopsis of concentrations of glyphosate is tabulated in
    Table 7. Measurements as part of regular monitoring programmes are
    very scarce; measurements in field experiments with recommended
    application rates simulating common agricultural practice are
    therefore included in Table 7. Only maximum amounts are tabulated as
    indicative values, since the rate at which they dissipate is not
    included here (see sections 4.1 and 4.3). Data on the occurrence of
    glyphosate and AMPA in sewage sludge are not available.

         In biota the highest concentrations of glyphosate and AMPA were
    found in fresh foliage and reindeer lichen  (Cladonia rangiferina).
    In abiota the highest concentrations of both compounds were found in
    the soil (see Table 7). The occurrence of glyphosate in the
    groundwater of Texas, USA, was reported by Hallberg (1989), but the
    measured concentration and the year of measurement were not
    specified.

         Use of glyphosate as a herbicide may result in the presence of
    residues in crops and animal tissues destined for human consumption.
    Application as a herbicide may also be responsible for the presence
    of glyphosate in drinking-water. Direct measurements of glyphosate
    in foodstuffs (as part of food surveillance), drinking-water or
    total diets have not been carried out. The only information
    available comes from controlled residue studies. With technical
    glyphosate formulated as the isopropylamine salt in aqueous
    solution, numerous residue studies have been carried out in
    vegetables, grasses, oil seeds, mammalian products, poultry products
    and primary feed commodities. The results are summarized in the
    various reports of the FAO/WHO Joint Meeting on Pesticide Residues
    (FAO/WHO, 1986a, 1987, 1988). For detailed information on these
    studies the reader is referred to these reports. The appraisals made
    by the JMPR included the following more general statements.
    Pre-harvest (5-14 days) application of glyphosate (isopropylamine
    salt) in the cultivation of cereals results in significant residues
    in the grain and plant materials. Studies are available to show the
    fate of glyphosate in milling, baking and brewing. Residue levels in
    white flour were approximately 10-20% of the levels in wheat, while
    the bran residue levels were 2 to 4 times as high as those in the
    wheat. Glyphosate residues were not lost during baking, but residue
    levels decreased when bread was made from flour because of dilution.

    Glyphosate residue levels in malt and beer derived from
    field-treated barley were, respectively, about 25% and 4% of the
    original level in the barley. Some glyphosate is lost during
    washing, but most of the decrease can be attributed to dilution. The
    levels in groats (processed oats) were about 50% of the levels in
    the pre-harvest-treated oats. In all these cases, AMPA contributed
    only a small proportion (average < 2.5%) of the total residues
    (FAO/WHO, 1986a, 1987).

         When administered to animals glyphosate is rapidly excreted
    without degradation. Residues in cattle, pig and poultry meat, eggs
    and milk were negligible after the animals were fed with a diet
    containing 100 mg/kg glyphosate and aminoglyphosate acid. The
    highest residues were in pig liver and kidney (up to 0.16 and
    0.91 mg/kg, respectively) and cattle kidney (up to 1.4 mg/kg).
    Residues in animal feeds arising from pre-harvest glyphosate
    applications to cereals will result in only low residues in meat,
    milk and eggs (FAO/WHO, 1986a).

         On the basis of these residue studies the JMPR has estimated
    the maximum residue levels that are likely to occur when glyphosate
    (as isopropylamine salt) is used in practice, and recommended these
    levels as Maximum Residue Limits (MRLs). These MRLs are presented in
    FAO/WHO (1986a, 1987, 1988).

    5.2  General population exposure

         Apart from the controlled residue studies mentioned above, no
    data are available.


    
    Table 7.  Maximum concentrations of glyphosate in environmental air, water, soil, sediment and biota
                                                                                                                                              

    Sample                   Compound       Location                Yeara      Concentration           Reference
                                                                                                                                              

    Netherlands

    Surface water            glyphosate     Drentsche Aa          1988-1989    0.5-1 µg/litreb         (Soppe, personal
    Surface water            AMPA           Drentsche Aa          1988-1989    6 µg/litreb             communication to the IPCS, 1991)

    Finland

    Loam soil (agric.)       glyphosate     Kettula                 1978       17 mg/kg d.w.           Müller et al. (1981)
    Loam soil (agric.)       AMPA           Kettula                 1978       3.2 mg/kg d.w.          Müller et al. (1981)
    Silt soil (agric.)       glyphosate     Kettula                 1978       3.8 mg/kg d.w.          Müller et al. (1981)
    Silt soil (agric.)       AMPA           Kettula                 1978       0.4 mg/kg d.w.          Müller et al. (1981)
    Wild berries             glyphosate     Laukaa, Konnevesi       1977       1.6-2.1 mg/kgc          Siltanen et al. (1981)
    Wild berries             AMPA           Laukaa, Konnevesi       1977       0.02-0.07 mg/kgc        Siltanen et al. (1981)
    Reindeer lichen          glyphosate     Laukaa, Konnevesi       1976       45 mg/kgc               Siltanen et al. (1981)
    Reindeer lichen          AMPA           Laukaa, Konnevesi       1976       2.1 mg/kgc              Siltanen et al. (1981)

    Canada

    Wild berries             glyphosate     Harker, Lamplugh        1985       8-19 mg/kg f.w.         Roy et al. (1989a)
    Wild berries             AMPA           Harker, Lamplugh        1985       0.06-0.1 mg/kg f.w.     Roy et al. (1989a)
    Surface waterd           glyphosate     Carnation Creek         1984       162 µg/litre            Feng et al. (1990)
    Surface watere           glyphosate     Carnation Creek         1984       < 1 µg/litre            Feng et al. (1990)
    Surface waterd           AMPA           Carnation Creek         1984       approx 3 µg/litre       Feng et al. (1990)
    Pond water               glyphosate     Manitoba                1986       141 µg/litre            Goldsborough & Beck (1989)
    Pond water               AMPA           Manitoba                1986       2.2 µg/litre            Goldsborough & Beck (1989)
    Sedimentd                glyphosate     Carnation Creek         1984       6.8 mg/kg d.w.          Feng et al. (1990)
    Suspended sediment       glyphosate     Carnation Creek         1984       0.06 µg/litre           Feng et al. (1990)
    Soil                     glyphosate     Carnation Creek         1984       40 mg/kg d.w.           Feng & Thompson (1990)
    Soil                     AMPA           Carnation Creek         1984       9 mg/kg d.w.            Feng & Thompson (1990)
                                                                                                                                              

    Table 7.  (cont'd)
                                                                                                                                              

    Sample                   Compound       Location                Yeara      Concentration           Reference
                                                                                                                                              

    Canada (cont'd)

    Foliage (fresh)          glyphosate     Carnation Creek         1984       261-448 mg/kg d.w.      Feng & Thompson (1990)
    Foliage (fresh)          AMPA           Carnation Creek         1984       < 9 mg/kg d.w.          Feng & Thompson (1990)
    Topcrown foliage         glyphosate     Oregon Coast Range      1978       489 mg/kgc              Newton et al. (1984)
    Topcrown foliage         AMPA           Oregon Coast Range      1978       2.1 mg/kgc              Newton et al. (1984)
    Deermice (viscera)       glyphosate     Oregon Coast Range      1978       5.1 mg/kgc              Newton et al. (1984)

    USA

    Pond water               glyphosate     Chassell, Corvallis,    1987       90-1700 µg/litre        Monsanto (1990a)
                                            Cuthbert
    Pond water               AMPA           Chassell, Corvallis,    1987       2-35 µg/litre           Monsanto (1990a)
                                            Cuthbert
    Stream water             glyphosate     Chassell, Corvallis,    1987       35-1237 µg/litre        Monsanto (1990a)
                                            Cuthbert
    Stream water             AMPA           Chassell, Corvallis,    1987       < 1.0-10 µg/litre       Monsanto (1990a)
                                            Cuthbert
    Pond sediment            glyphosate     Chassell, Corvallis,    1987       0.26-19 mg/kg d.w.      Monsanto (1990a)
                                            Cuthbert
    Pond sediment            AMPA           Chassell, Corvallis,    1987       0.13-1.8 mg/kg d.w.     Monsanto (1990a)
                                            Cuthbert
    Stream sediment          glyphosate     Chassell, Corvallis,    1987       0.11-0.69 mg/kg d.w.    Monsanto (1990a)
                                            Cuthbert
    Stream sediment          AMPA           Chassell, Corvallis,    1987       < 0.05-0.38 mg/kg d.w.  Monsanto (1990a)
                                            Cuthbert
    Soil (no litter on it)   glyphosate     Chassell, Corvallis,    1987       0.15-4.7 mg/kg d.w.     Monsanto (1990a)
                                            Cuthbert
                                                                                                                                              

    Table 7.  (cont'd)
                                                                                                                                              

    Sample                   Compound       Location                Yeara      Concentration           Reference
                                                                                                                                              

    USA (cont'd)

    Soil (no litter on it)   AMPA           Chassell, Corvallis,    1987       0.18-0.51 mg/kg d.w.    Monsanto (1990a)
                                            Cuthbert
    Soil (litter on it)      glyphosate     Chassell, Corvallis,    1987       0.07-1.4 mg/kg d.w.     Monsanto (1990a)
                                            Cuthbert
    Soil (litter on it)      AMPA           Chassell, Corvallis,    1987       0.14-0.68 mg/kg d.w.    Monsanto (1990a)
                                            Cuthbert
    Foliage (fresh)          glyphosate     Chassell, Corvallis,    1987       650-1300 mg/kgb         Monsanto (1990a)
                                            Cuthbert
    Foliage (fresh)          AMPA           Chassell, Corvallis,    1987       1.7-2.6 mg/kgb          Monsanto (1990a)
                                            Cuthbert
                                                                                                                                              

    a    Sampling invariably took place during the autumn period.
    b    Analytical procedure was unvalidated; the water was sampled at an inlet point of
         a drinking-water processing facility
    c    It was not reported whether values were based on dry or fresh weight
    d    Water unbuffered by vegetation
    e    Water buffered by vegetation
    

    5.3  Occupational exposure during manufacture, formulation or use

         In the study of Monsanto (1977), worker exposure to Roundup was
    measured during herbicide mixing and application operations and upon
    re-entry of treated fields. The formulation was applied on separate
    plots using three application devices, i.e. a broadcast boom
    sprayer, a handgun broadcast sprayer and a backpack/hand-gun
    sprayer. Exposure time during mixing was < 5 min; during
    application this was about 45 to 60 min. Inhalational exposure
    during mixing and spraying was determined using a sampling device
    placed close to the applicator's face; the total air volume sampled
    was 15-20 times the daily ventilation volume. Dermal exposure during
    mixing and spraying was monitored by determination of deposition on
    gauze pads placed outside or inside the operator's clothing on
    different parts of the body. Operator exposure (inhalation and skin,
    determination method as above) upon re-entry was determined at 1, 3
    and 7 days after application. In one treated plot, this was done by
    having operators walk through the plot for 28 to 44 min; in two
    other plots a dummy sampling device was used to determine exposure
    upon re-entry. Using the measured glyphosate residues total worker
    exposure was estimated. The results are presented in Table 8.

        Table 8. Applicator/worker exposure to glyphosatea
                                                                                            

    Operation                  Average dermal    Average inhalation    Total exposure
                                 exposure            exposure              (µg/h)
                                  (µg/h)             (µg/h)b
                                                                                            

    Tank filling                     805.1             17.9                823.0

    Boom spraying                    271.4              0.19               271.59

    Handgun spraying                7957.0              2.47              7959.47

    Backpack spraying               3619.0              0.92              3619.92

    Field re-entry:

    1 day after treatment           2046.6              4.58              2051.18

    3 days after treatment          2919.7              0.12              2919.82

    7 days after treatment            15.9              0.12                16.02
                                                                                            

    a    From: Monsanto (1977)
    b    Calculation based on an estimated breathing rate of 1.8 m3/h
    
         Monsanto (1990) conducted another collaborative study at three
    sites maintained by the USDA Forestry Service near Clayton, Georgia;
    the Savanah River Plant, South Carolina; and Edgefield, South
    Carolina. Glyphosate was being used to control vegetative growth
    around pine seedlings planted in clear-cut forest areas. The workers
    were biologically monitored by analysis of collected composite urine
    specimens. Additionally, dermal/clothing deposition and simulated
    inhalation exposure were monitored by passive dosimetry with cotton
    cloth patches, hand rinses and air filters. At the three sites,
    exposure of workers to glyphosate using backpack sprayers while
    performing their duties under normal use conditions was monitored.
    The analytical results indicated that the majority of urine
    composite samples had unmeasurable residues of glyphosate.
    Deposition on air filters, patches and hands from measurement of
    washes, however, indicated a small amount of body exposure. It was
    concluded that penetration of clothing did not exceed 3.84% and thus
    clothing produced 96.2% protection. Body burden, as shown in urine
    samples, was extremely low and in most cases below the detection
    limit.

         A new Monsanto study was conducted for the assessment of worker
    exposure to glyphosate during mist blower application of Roundup
    herbicide. Exposure was determined by a passive dosimetry technique
    while workers sprayed weeds around palm trees in a plantation in
    Malaysia. The workers were fitted with gauze patches at different
    locations on their clothing. Air sampling was performed in the
    breathing zone and the workers hands were washed at the end of the
    day. The passive dosimetry body dose estimates were calculated for a
    fully clothed worker with a long-sleeved shirt, long pants and
    rubber boots. The hand exposure would account for bare hands during
    the loading and spraying operations. Passive dosimetry estimates for
    the four replicates, corrected for clothing and dermal penetration,
    transport/storage/analytical recovery and normalized for body weight
    and amount of chemical handled, averaged 1.88 µg/kg body weight per
    kg a.i. This is little higher than the passive dosimetry estimates
    of forestry workers who applied Roundup with knapsack sprayers,
    which was 1.75 µg/kg body weight per kg a.i. Thus, it can be
    concluded that workers applying Roundup herbicide with mist blowers,
    experience some dermal exposure. In addition, inhalation exposure to
    glyphosate may be higher during mist blower application. However,
    Roundup demonstrated essentially no volatility. The only possible
    route would be via air-borne particles. The actual amount of
    glyphosate absorbed through inhalation would be much lower than the
    estimated values because the measurement includes particles too
    large to be inhaled (Monsanto, 1991).

         Jauhiainen et al. (1991) examined the exposure of workers to
    glyphosate during silvicultural clearing with brush saws equipped
    with herbicide sprayers. Measurements of air concentrations during
    spraying and urinary glyphosate levels both during and following
    spraying were carried out. Most of the air samples had glyphosate
    concentrations below 1.25 µg/m3; the highest value observed was
    15.7 µg/m3. All urine samples taken had glyphosate concentrations
    below the limit of detection of 0.1 mg/litre (Jauhiainen et al.,
    1991).

         Lavy et al. (1992) used two methods to monitor glyphosate
    exposure of workers planting conifer seedlings. Firstly, they
    estimated dermal exposure based on the rate of deposition on cotton
    gauze patches, surface area exposed, and a dermal penetration rate
    of 1.8%. This yielded dose estimates in the range of 0-875 or
    0-1.7 µg/kg body weight per h. Secondly, they attempted to measure
    urinary concentration levels but found no samples above the
    detection limit of 0.01 mg/litre. Based on the negative results with
    the urine samples, the authors concluded that the estimates based on
    patch deposition overestimated exposure by at least a factor of 11
    for the most highly exposed workers (Lavy et al., 1992).

         It should be noted that a dermal penetration rate of 1.8% was
    used in this calculation. In view of the discussion of the dermal
    penetration studies (section 6.1), a value of 5.5% is to be
    preferred. However, since the estimate based on 1.8% is already an
    overestimation of exposure, it is not considered necessary to adjust
    the estimate of Lavy et al. (1992) to 5.5%. In their final data
    assessment, the authors estimated exposure from the biological
    monitoring using postulated data, i.e. assuming a concentration in
    urine of half the lower limit of method validation. This yielded
    mean exposure values of 0.039 to 0.080 µg/kg body weight per h (Lavy
    et al., 1992).

    6.  KINETICS AND METABOLISM IN LABORATORY
        ANIMALS AND HUMANS

    Appraisal

          Absorption from the gastrointestinal tract after oral intake
     is limited to 30-36% of the dose or less in various species, i.e.
     rats, rabbits, laying hens and lactating goats. Percutaneous
     absorption in Rhesus monkeys amounts to 5.5% only, and glyphosate is
     very poorly absorbed through excised human abdominal skin.

          Radiolabelled glyphosate distributes widely in the body, but
     is primarily found in the bones where approximately 1% can be
     detected after oral administration.

          Glyphosate is essentially not metabolized. This validates
     kinetic studies performed with radiolabelled compound.

          After absorption, excretion of glyphosate occurs mainly in the
     urine. Biliary excretion is limited and elimination through exhaled
     air is very low. Total body clearance is 99% after 168 h.

    6.1  Absorption

         The absorption percentage in rats was reported to be 30-36%
    after single oral dosage at 10 and 1000 mg/kg body weight;
    calculations were based on excretion percentages in urine and
    faeces, and on the fact that biliary elimination is probably a minor
    route (Monsanto, 1988b; Brewster et al., 1991). From a similar study
    carried out in 1973, total absorption percentages of approximately
    20% (male rats) and 45% (female rats) can be derived (Monsanto,
    1973a). Comparable results were obtained in a recent single dose
    (5.6 or 56 mg/kg body weight) disposition study in F344/N rats (NTP,
    1992), which indicated that 30% of the dose was absorbed.

         In a 14-day oral study in rats with application of
    14C-glyphosate via the diet (dose levels 1, 10 and 100 mg/kg
    feed), the observed total excretion in urine was < 10% and in
    faeces approximately 80-90%. Given the minor importance of the
    biliary elimination route, these data indicate absorption levels of
    about 15% or less (Monsanto, 1973c). The results of oral studies
    with 14C-glyphosate in rabbits (Monsanto, 1973d), laying hens
    (Hazleton Lab. Inc., 1988a) and lactating goats (Hazleton Lab. Inc.,
    1988b) indicate gastrointestinal absorption percentages of
    approximately 30% or less.

         Percutaneous absorption has been studied in Rhesus monkeys and
    in human tissue  in vitro. After a single application of
    14C-glyphosate (isoproylamine salt) as the undiluted Roundup
    formulation to the shaven intact abdominal skin (contact time 24 h)
    of Rhesus monkeys, absorption amounted to only 1.8% of the dose. In
    this study, however, only 16% of the dose could be accounted for at
    the end of the study (Maibach, 1983). This low recovery strongly
    reduces the value of the study result (possibly skin absorption is
    seriously underestimated in this study). From an identical study
    with diluted Roundup formulation (1:29 with water), conducted by
    Wester et al. (1991), also in Rhesus monkeys (contact time 12 h),
    total absorption percentages of 3.7% (at low dose) or 5.5% (high
    dose) can be derived. Using both undiluted and diluted Roundup
    formulation, Wester et al. (1991) observed that percutaneous
    absorption of 14C-glyphosate through human skin  in vitro into
    human plasma as receptor fluid was < 2% (contact-time up to
    16 h). In another  in vitro study with human skin, absorption of
    14C-glyphosate from three undiluted formulations (i.e. MON 0139,
    Roundup and Roundup spray mix) was studied (contact time 24 h); very
    low absorption percentages of 0.028 to 0.152% were found (Franz,
    1983). With regard to these  in vitro results it should be pointed
    out that this technique has not yet been fully validated and
    therefore direct extrapolation to  in vivo human skin absorption
    should be undertaken cautiously.

         The absorption after inhalational intake has not been
    determined.

    6.2  Distribution

         Concentrations of 14C label in tissues were determined on day
    7 after administration of a single oral dose (10 or 1000 mg/kg body
    weight) of 14C-glyphosate to rats (Monsanto, 1988b). Although only
    a small proportion was absorbed, the isotope was widely distributed
    throughout the body, but was primarily found in bone. The principal
    results are presented in Table 9.

         In rats tissue concentrations of 14C label were determined on
    several occasions throughout a treatment period of 14 days and a
    post-dosing withdrawal period of 10 days (dietary administration of
    14C-glyphosate at 1, 10 and 100 mg/kg diet). Maximum tissue levels
    were reached after 10 days or less, with highest concentrations
    (maximum 0.85 mg/kg at the 100 mg/kg dose level) in kidneys
    (Monsanto, 1973c). It should be noted, however, that in this study
    concentrations in bone or bone marrow were not measured. An increase
    of 14C in excreta was observed during the withdrawal period after
    an initial rapid decrease; this indicated mobilization from storage
    in depot tissue (Monsanto, 1973c).

    
    Table 9. Concentrations of 14C label (as mg glyphosate-equivalents/kg
    fresh weight) in selected tissues of rats on day 7 after a single
    oral dose (rounded values) (Monsanto, 1988b)
                                                                                            

                      Dose: 10 mg/kg body weight          Dose: 1000 mg/kg body weight
                                                                                            

                      male          female                male          female

    Blood             0.0045        0.0027                 0.33           0.17

    Liver             0.030         0.014                  1.9            1.3

    Kidney            0.022         0.013                  1.9            1.4

    Spleen            0.012         0.0073                 2.6            3.0

    Lung              0.015         0.012                  1.5            1.1

    Thyroid           0.00080       0.00036                1.5            1.2

    Nasal mucosa      0.0050        0.023                  1.7            1.8

    Stomach           0.0080        0.0037                 2.4            2.4

    Small intestines  0.022         0.018                  1.9            1.6

    Colon             0.034         0.016                 11.0            9.2

    Bone              0.55          0.31                  30.6           19.7

    Bone marrow       0.029         0.0064                 4.1           12.5
                                                                                            
    

         In lactating goats concentrations of 14C label in milk were
    measured after giving capsules containing a 9:1 mixture of
    14C-glyphosate and 14C-aminomethylphosphonic acid (AMPA) to a
    dose level equivalent to 120 mg/kg diet (expressed as free acid) for
    5 days. Concentrations in milk (as mg equivalents glyphosate/kg
    whole milk) ranged from 0.019 to 0.086 mg/kg during the test period;
    at day 5 after the last dose the concentration was 0.006 mg/kg
    (Hazleton Lab. Inc., 1988b; Monsanto, 1988d).

         In a study on laying hens, carried out using a 9:1 mixture of
    14C-glyphosate and 14C-AMPA, concentrations of radiolabel were
    measured in eggs collected during a 7-day period of dietary
    administration at 120 or 400 mg/kg diet. At 400 mg/kg, residues in
    egg white were 0.010-0.032 mg/kg (expressed as
    glyphosate-equivalents) and in egg yolk 0.096-0.753 mg/kg; at
    120 mg/kg the corresponding concentration ranges were 0.003-0.017
    and 0.002-0.24 mg/kg, respectively. At 120 mg/kg diet, no 14C was
    detectable in egg white after 10 withdrawal days; in yolk
    0.019 mg/kg was present at that time (at 400 mg/kg no withdrawal
    test was conducted) (Hazleton Lab. Inc., 1988a).

    6.3  Metabolic transformation

         Biotransformation of glyphosate occurs to a very low degree
    only. In rats it was shown that all of the 14C in urine and
    faeces, after a single oral application of 14C-glyphosate, was
    present as unchanged parent compound (Monsanto, 1973b). Also in
    rats, > 97% of the 14C in excreta, after a single oral dose,
    was shown to be unchanged compound. AMPA was the only metabolite,
    covering only 0.2-0.3% of the applied 14C (Monsanto, 1988a). In
    laying hens also, AMPA was the only metabolite, accounting for only
    a minor part of the applied amount (Monsanto, 1988c).

    6.4  Elimination and excretion

         In the period of 0-5 days after a single oral application of
    14C-glyphosate (6.7 mg/kg body weight) to rats the total excretion
    in urine was 15% (males) and 35-43% (females) of the administered
    dose; total excretion in faeces was 85% (males) or 50-55% (females).
    Less than 1% of the radiolabel was expired as 14CO2 (Monsanto,
    1973a). In a more recent study (Monsanto, 1988b), the very low level
    of expiration as 14CO2 was confirmed but no significant
    inter-sex difference in the level of 14C in excreta was observed.
    The result of the latter study was that at both oral dose levels (10
    and 1000 mg/kg body weight) elimination in faeces was 62-70% and
    excretion in urine was 14-18% (1000 mg/kg body weight) or 22-29%
    (10 mg/kg body weight); less than 0.2% of the dose was expired as
    14CO2. After single intravenous application (dose 10 mg/kg body
    weight) 75-79% appeared in urine and only 5-8% in faeces, a finding
    that shows that biliary elimination occurs to a limited degree only
    (Monsanto, 1988b).

         Delayed excretion in rats during a 10-day post-dosing
    withdrawal period was observed after daily oral administration via
    the diet for 14 days (Monsanto, 1973c); this suggests that some
    storage in tissue(s) occurs when uptake is prolonged. Tissue
    equilibrium was attained by day 10 of the dosing period and
    excretion equalled intake by day 6 of the dosing period.

         In rabbits > 80% was eliminated in faeces (with additional
    14C present in the gut) and 7-11% in urine within 5 days after
    administration of a single oral dose (6.7 mg/kg body weight) of
    14C-glyphosate. Less than 1% of the dose was expired as 14CO2
    (Monsanto, 1973d). In one oral study in lactating goats lasting 5
    days, total excretion in urine varied from 20 to 24% and in faeces
    from 60 to 66% (Hazleton Lab. Inc., 1988b).

    6.5  Retention and turnover

         Total body clearance in the study of Monsanto (1973a) was
    94-98% (males) or 82-84% (females) over a 48-h period after giving a
    single oral dose of 14C-glyphosate; at 120 h post-dosing this was
    99% (both sexes). In Monsanto (1988b), the kinetics of whole body
    elimination were estimated using the radioactivity (14C) measured
    in urine and faeces after a single oral dose of 14C-glyphosate (10
    or 1000 mg/kg body weight). Because of the lack of biotransformation
    of glyphosate it is valid to base kinetics on total radioactivity.
    The elimination appeared to be biphasic. The half-life of the alpha
    elimination phase at 10 mg/kg body weight was 5.87 h (males) or
    6.22 h (females); at 1000 mg/kg body weight this was 5.26 h (males)
    or 6.44 h (females). The half-life of the beta phase at 10 mg/kg
    body weight was 79 h (males) or 106 h (females); at 1000 mg/kg body
    weight this was 181 h (males) or 337 h (females). Pretreatment with
    unlabelled compound for 14 days (carried out at the low dose level)
    did not have an effect on whole body elimination. Seven days after
    dosing, < 0.05% of the dose was present in organs and < 0.5% in
    the remaining carcass. Highest concentrations were present in bone.
    It was estimated that 0.2-0.6% of the oral dose was associated with
    this site; after intravenous dosing this was approximately 1%
    (Monsanto, 1988b). Brewster et al. (1991) reported that in rats
    nearly all of the absorbed material had been eliminated from the
    body 168 h after oral administration of 10 mg/kg body weight;
    approximately 1% of the dose was still associated with the bone.

         In a recent study on the disposition of glyphosate in F-344/N
    rats, 1% of a single oral dose (5.6 or 56 mg/kg) was found in the
    tissues 72 h after dosing; 20-30% of the administered radioactivity
    was eliminated via urine and 70-80% via the faeces (NTP, 1992).

    7.  EFFECTS ON LABORATORY ANIMALS AND  IN VITRO TEST SYSTEMS

    Appraisal

          Glyphosate, administered by oral and dermal routes, has a very
     low acute toxicity. Both glyphosate and its concentrated
     formulations produce moderate to severe eye irritation, but only
     slight dermal irritation. Neither glyphosate nor tested formulations
     induce sensitization.

          Short-term feeding studies have been conducted in several
     species. In CD-1 mice, increased liver, brain, heart and kidney
     weights, and growth retardation were reported at 50 000 mg/kg diet.
     At 10 000 mg/kg diet, an increase in relative liver weight was
     reported; however, there were no differences in absolute liver
     weights when this group was compared to controls. The relative
     increase represented only a 9% increase over the liver weight
     reported for controls and was not considered toxicologically
     significant. Additionally, there were no gross or histopathological
     changes in the liver at doses of 10 000 mg/kg or more. The Task
     Group considered the NOAEL to be 50 000 mg/kg diet.

          In a 13-week study conducted in Charles River CD
     (Sprague-Dawley) BR rats, no treatment-related effects were observed
     at doses up to 20 000 mg/kg diet. The NOAEL was greater than the
     highest dose tested.

          Two additional 13-week studies (one in rats and the other in
     mice) were conducted by NTP in which lesions of the salivary glands
     were observed in both species. The NOAEL in the rat study was 
     < 3125 mg/kg diet and that in the mouse study was 3125 mg/kg diet.
     Other short-term and long-term studies conducted in different
     strains and species did not reveal similar lesions. The lesions
     indicate that glyphosate may be acting as a weak adrenergic agonist.
     The toxicological significance of the salivary gland lesions
     observed in the NTP studies is unknown.

          In a 52-week study conducted in beagles, no compound-related
     effects were reported. The NOAEL was 500 mg/kg body weight per day.
     In a 7-day study with the Roundup formulation in female cattle, a
     NOAEL of 400 mg Roundup/kg body weight was reported. At higher dose
     levels, decreased feed intake and diarrhoea occurred.

          In long-term feeding studies in both rats and mice, few toxic
     effects were observed. These effects were present at relatively high
     dose levels only. In mice, technical glyphosate produced growth
     retardation, hepatocyte hypertrophy or necrosis at 30 000 mg/kg diet
     only. At 5000 and 30 000 mg/kg diet an increase in epithelial
     hyperplasia of the urinary bladder was reported. The increased
     incidence of this lesion did not follow a dose-related trend and in

     the highest dose tested the incidence was actually lower than that
     reported at the medium dose level, in spite of a 6-fold increase in
     glyphosate. The observation at the medium dose (5000 mg/kg) is not
     considered a compound-related effect and the NOAEL is, therefore,
     considered to be 5000 mg/kg diet (814 mg/kg body weight).

          Long-term feeding studies in rats resulted in decreased
     growth, increased liver weight and degenerative liver changes at
     20 000 mg/kg diet only. At 8000 and 20 000 mg/kg diet, there was an
     apparent increase in the incidence of inflammation of the gastric
     mucosa in both sexes. The only statistically significant increase
     was observed in the medium-dose females (15%). This value was also
     outside the historical control range of 0-13%. This finding was not
     considered to be a treatment-related effect. There was no
     dose-related trend across all groups of treated females and there
     was no statistically significant difference in any treated male
     groups. The NOAEL was therefore 8000 mg/kg diet (410 mg/kg body
     weight).

          Studies in rats and rabbits indicated that technical
     glyphosate is not teratogenic. Two multigeneration studies were
     conducted with technical glyphosate. In the first study, the only
     effect noted was an increased incidence of unilateral renal tubular
     dilation in F3b  male pups at 30 mg/kg body weight. In the second
     study, decreased body weights were reported for parents and pups and
     decreased litter size was associated with dose levels of
     30 000 mg/kg diet. Decreased body weights reported for parents and
     pups at 10 000 mg/kg diet were not toxicologically significant. In
     parents, the decrease was only 2 to 4% below controls and for pups
     the decrease was 5.6 to 6.6% lower than controls. The findings in
     pups were also transient and did not occur consistently in all
     litters. The NOAEL was 10 000 mg/kg diet. The absence of a renal
     effect in pups at a higher dose level (1500 mg/kg body weight),
     though not invalidating earlier findings of unilateral renal tubular
     dilation in male F3b  pups, indicates that the reproducibility of
     this lesion and its toxicological significance are uncertain. It
     should be noted that in no other toxicological study was an effect
     on kidneys found.

          Bioassays in mice and rats did not indicate that technical
     glyphosate was carcinogenic.

          Glyphosate has been shown to have no genotoxic potential in a
     range of in vitro and in vivo studies.

    7.1  Single exposure

         Numerous acute toxicity studies have been performed to
    determine LD50 values of glyphosate and of herbicide formulations
    containing glyphosate as active ingredient. The results of these
    studies are summarized in Tables 10 (results for glyphosate) and 11

    (results for formulations). These data show that glyphosate and its
    formulations have very low toxicity by the oral and dermal
    administration routes. By the intraperitoneal route glyphosate is
    markedly more toxic than by the other routes. General intoxication
    symptoms include breathing difficulties, ataxia and convulsions.

         The mechanism of the toxic action of glyphosate has been
    studied in rats. Olorunsaga et al. (1979) observed dose-related
    reduced respiratory control ratios and increased phosphatase
    activity in mitochondria isolated from rat livers 5 h after single
    intraperitoneal doses ranging from 15 to 120 mg/kg body weight. This
    effect was also seen in rat liver mitochondria  in vitro (Bababunmi
    et al., 1979; Olorunsaga, 1982a,b). The authors suggest that acute
    toxicity at lethal doses may occur as a result of the uncoupling of
    oxidative phosphorylation.

         The acute toxicity in rats of the surfactant
    polyoxyethyleneamine, with which glyphosate is commonly formulated
    in Roundup, was compared to that of glyphosate in a study by
    Martinez et al. (1990). Both compounds exhibited pulmonary toxicity
    following either oral or intratracheal administration. The toxicity
    of the herbicide formulation was greater than can be accounted for
    on the basis of the dose response data from either compound alone
    (Martinez et al., 1990; Martinez & Brown, 1991).

         A study was undertaken by Tai et al. (1990) to investigate the
    effects of glyphosate, surfactant, and their combination in Roundup
    on cardiovascular function in female beagles. They found that
    glyphosate alone at plasma levels ranging from 923 to 3450 mg/litre,
    which simulates the human ingestion situation, were shown to
    increase the myocardial contractility. The surfactant alone
    considerably reduced the cardiac output, the left ventricular stroke
    work index and the mean arterial pressure. The joint effect of both
    glyphosate and the surfactant in Roundup formulation resulted in
    cardiac depression, which was mostly due to the surfactant since
    glyphosate itself increased myocardial contractility. The authors
    indicated that the probable cause of the observed increases in
    pulmonary vascular resistance index and pulmonary artery pressure
    was a direct vasoactive effect of glyphosate on the pulmonary
    artery.

    
    Table 10. Acute toxicity of glyphosate to experimental animals
                                                                                            

    Species (sex)      Product tested            LD50/LC50a           Reference
                                                                                            

    Oral studies

    Rat (m,f)          glyphosate techn,         > 5000 mg/kg bw      FDRL (1988d)
                       purity 97.8%
    Rat (m,f)          glyphosate techn,         > 5000 mg/kg bw      Inveresk Research Int.
                       purity 96-99%                                  (1989a)
    Rat (m,f)          glyphosate techn,         > 5000 mg/kg bw      NOTOX (1988)
                       purity 96-99%
    Rat (m,f)          85.5% techn.              > 5000 mg/kg bw      Bio/Dynamics Inc.
                       glyphosate in                                  (1988c)
                       water
    Rat (m,f)          glyphosate, IPA salt,     > 5000 mg/kg bw      Monsanto (1981b)
                       65% in water
    Rat (m.f)          glyphosate, EO saltb      2047 mg/kg bw        Knapek et al. (1986)
    Goat (f)           glyphosate techn,         3500 mg/kg bw        USDA (1987c)
                       purity 98.7%
    Goat (f)           glyphosate, IPA salt,     5700 mg/kg bw        USDA (1987b)
                       65% in water

    Dermal studies

    Rat (m,f)          glyphosate techn,         > 2000 mg/kg bw      Inveresk Research Int.
                       purity 96-99%                                  (1989c)
    Rabbit (m,f)       glyphosate techn,         > 5000 mg/kg bw      FDRL (1988b)
                       purity 97.8%
    Rabbit (m,f)       85.5% techn.              > 5000 mg/kg bw      Bio/Dynamics Inc.
                       glyphosate in water                            (1988a)
    Rabbit (m,f)       glyphosate, IPA salt,     > 5000 mg/kg bw      Monsanto (1981c)
                       65% in water

    Intraperitoneal studies

    Mouse (m)          glyphosate (not           545 mg/kg bw
                       further specified)
    Mouse (f)          glyphosate (not           740 mg/kg bw         FAO/WHO (1986b)
                       further specified)
    Mouse (m,f)        glyphosate (not           134 mg/kg bw         FAO/WHO (1986b)
                       further specified)
    Rat (m)            glyphosate (not           281 mg/kg bw
                       further specified
                                                                                            

    Table 10. (cont'd)
                                                                                            

    Species (sex)      Product tested            LD50/LC50a           Reference
                                                                                            

    Rat (f)            glyphosate (not           467 mg/kg bw         FAO/WHO (1986b)
                       further specified)
    Rat (m,f)          glyphosate (not           238 mg/kg bw         FAO/WHO (1986b)
                       further specified)
                                                                                            

    a    All values expressed as mg of product tested (as presented in "product tested"
         column)

    b    EO is an abbreviation of 5-ethoxy-oleinamine salt.


    Table 11.  Acute toxicity of glyphosate formulations to experimental animals
                                                                                               

    Species (sex)     Product testeda              LD50/LC50b         Reference
                                                                                               

    Oral studies

    Mouse (m,f)       Roundup                      >5000 mg/kg bw     Mitsukaido Labs (1986)
    Rat (m,f)         Roundup                      5000 mg/kg bw      Bio/Dynamics Inc.
                                                                      (1988e)
    Rat (m,f)         "Compound No. 3607"          >5000 mg/kg bw     Inveresk Research Int.
                                                                      (1988a)
    Rat (m,f)         Roundup TX                   >5000 mg/kg bw     NOTOX (1987a,b)
    Rat (m,f)         Alphee                       >5000 mg/kg bw     Bio/Dynamics Inc.
                                                                      (1987a)
    Rat (m,f)         Sting TX                     >5000 mg/kg bw     NOTOX (1987f,g)
    Rat (m,f)         Sting                        2510 mg/kg bw      Younger Labs Inc.
                                                                      (1984)
    Rat (m,f)         Sting                        1950 mg/kg bw      Bio/Dynamics Inc.
                                                                      (1984b)
    Rat (m,f)         MON 8780                     >5000 mg/kg bw     Bio/Dynamics Inc.
                                                                      (1985a)
    Rat (m,f)         Agrichem Glyfosaat B         >5000 mg/kg bw     NOTOX (1990e)
    Rat (m,f)         "Glyfosaat 360 g/litre"      >2000 mg/kg bw     NOTOX (1989c)
    Rat (m,f)         Legend                       >2000 mg/kg bw     CIT (1991a)
    Goat (f)          Roundup                      4860 mg/kg bw      USDA (1983)
                                                                                               

    Table 11. (cont'd)
                                                                                               

    Species (sex)     Product testeda              LD50/LC50b         Reference
                                                                                               

    Dermal studies

    Rat (m,f)         "Compound No. 3607"          >2000 mg/kg bw     Inveresk Research Int.
                                                                      (1988b)
    Rat (m,f)         Roundup TX                   >4000 mg/kg bw     NOTOX (1987c)
    Rat (m,f)         Sting TX                     >4000 mg/kg bw     NOTOX (1987h)
    Rat (m,f)         MON 8780                     >5000 mg/kg bw     Bio/Dynamics Inc.
                                                                      (1985b)
    Rat (m,f)         Agrichem Glyfosaat           >4000 mg/kg bw     NOTOX (1990f)
                      B or 2
    Rat (m,f)         "Glyfosaat 360 g/litre"      >2000 mg/kg bw     NOTOX (1989b)
    Rat (m,f)         Legend                       >2000 mg/kg bw     CIT (1991b)
    Rabbit (m,f)      Roundup                      >5000 mg/kg bw     Bio/Dynamics Inc.
                                                                      (1988f)
    Rabbit (m,f)      Alphee                       >5000 mg/kg bw     Bio/Dynamics Inc.
                                                                      (1987b)
    Rabbit (m,f)      Sting                        >5000 mg/kg bw     Bio/Dynamics Inc.
                                                                      (1984a)

    Inhalation studies

    Rat (m,f)         Roundup (aerosol)            3180 mg/m3         Monsanto (1983d)
    Rat (m,f)         "Compound No. 3607"          >4860 mg/m3        Inveresk Research Int.
                                                                      (1989d)
    Rat (m,f)         Alphee                       >8900 mg/m3        Monsanto (1987b)
                                                                                               

    a    Composition of the formulations is given in Table 2, with the exceptions of
         Agrichem Glyphosaat B (or 2), "Glyfosaat 360 g/litre" and "Compound No. 3607",
         which all contain approximately 480 g/litre of the isopropylamine salt, and of MON
         8780 (32.8% isopropylamine salt), and Legend (40% isopropylamine salt).

    b    All values given as mg formulation.
    
    7.2  Short-term exposure

    7.2.1  Oral studies

         In CD-1 mice, a 13-week feeding study was conducted with
    technical glyphosate (purity 98.7%) using dose levels of 5000,
    10 000 and 50 000 mg/kg diet (equal to 940, 1890 and 9710 mg/kg body
    weight per day in males and 1530, 2730 and 14 860 mg/kg body weight
    per day in females). No effect on appearance or survival was
    observed. Growth retardation and increased weights of brain, heart

    and kidneys were observed at 50 000 mg/kg. Liver weights were
    increased at 50 000 and 10 000 mg/kg. Limited histopathology showed
    no adverse effects. The authors of the study concluded that the
    NOAEL was 10 000 mg/kg diet (1890 mg/kg body weight per day)
    (Bio/Dynamics Inc., 1979).

         In a 13-week feeding study with technical glyphosate,
    Sprague-Dawley rats received 1000, 5000 or 20 000 mg/kg diet (equal
    to 63, 317 and 1267 mg/kg body weight per day in males and 84, 404
    and 1623 mg/kg body weight per day in females). No effect on
    appearance, survival or growth occurred. Haematology, blood
    biochemistry and urinalysis, carried out at test end only, were also
    unaffected. Organ weights (determined for liver, kidneys and testes
    only) were not affected. Limited histopathology showed no adverse
    effect in any tissue. The NOAEL in this study was 20 000 mg/kg diet
    (1267 mg/kg body weight per day) (Monsanto, 1987a). Absence of
    toxicity was also found in another 13-week feeding study on rats
    using technical glyphosate and dose levels of 200 to 12 500 mg/kg
    diet (Tauchi, 1979 as cited by FAO/WHO, 1986b).

         Two further 13-week studies in rodents were conducted on behalf
    of the US National Toxicology Program (1992). Both rats (F-344/N)
    and mice (B6C3F1) were administered glyphosate (purity
    approximately 99%) in feed at levels of 3125, 6250, 12 500, 25 000
    and 50 000 mg/kg diet.

         In rats, reduced weight gains were observed at 25 000 mg/kg
    diet (males only) and at 50 000 mg/kg diet (males and females). No
    changes in feed consumption were found. Minor increases in the
    relative organ weight of liver, kidney and testes, and decreased
    thymus weight were observed in males only at several dose levels;
    these changes did not show a clear dose relation and therefore are
    not considered to be compound-related effects. Small increases in
    haematocrit and red cell blood counts were observed in male rats at
    > 12 500 mg/kg diet. Clinical chemistry showed increased alkaline
    phosphatase (AP) and alanine aminotransferase (ALAT) at >
    6250 mg/kg diet in males and at > 12 500 mg/kg diet in females.
    Bile acid levels in blood were increased at 25 000 mg/kg diet (males
    only) and at 50 000 mg/kg diet (males and females). Decreases in
    sperm count were observed in males at > 25 000 mg/kg diet. The
    only histopathological lesions found were cytoplasmic alterations of
    the parotid and submandibular salivary glands, consisting of
    basophilic changes and hypertrophy of acinar cells. The parotid
    gland was more severely affected. The magnitude of the effect was
    dose-dependent, with focal lesions in less severe cases to diffuse
    involvement at higher doses. Lesions of a similar nature and
    magnitude were observed in both sexes. The sublingual gland was not

    detectably altered. Effects on the salivary glands were observed
    already at the lowest dose level tested of 3125 mg/kg diet (equal to
    205 mg/kg body weight per day for males and 213 mg/kg body weight
    per day for females). Thus, this study did not yield a NOAEL (NOAEL
    < 3125 mg/kg diet) (NTP, 1992).

         In mice, reduced weight gains were observed at 50 000 mg/kg
    diet in both sexes. Increased organ weights were noted in heart,
    kidney, liver, thymus, lung and testis; these changes did not show a
    clear dose relation and therefore are not considered to be
    compound-related effects. Feed consumption levels were not changed
    significantly. Lesions in the parotid gland were observed, similar
    to those observed in rats. The sublingual gland and the
    submandibular salivary glands were not detectably altered. The
    effects on the parotid gland were observed in mice at 6250 mg/kg
    diet (equal to 1065 mg/kg body weight per day for males and
    1411 mg/kg body weight per day for females), but were not seen at
    the lowest dose level tested of 3125 mg/kg diet (equal to 507 mg/kg
    body weight per day for males and 753 mg/kg body weight per day for
    females). The NOAEL in this study was 3125 mg/kg diet (507 mg/kg
    body weight per day) (NTP, 1992).

         The salivary gland lesions could also be induced in rats by
    14-day exposure at feed levels of 50 000 mg/kg diet. The salivary
    glands lesions induced by glyphosate were similar to those which
    could be induced by exposure to high subcutaneous doses of the
    ß-adrenergic agonist isoproterenol and could be partially
    ameliorated with the ß-adrenergic antagonist propanolol. This
    indicates that glyphosate may induce the salivary gland lesions by
    acting as a weak adrenergic agonist (NTP, 1992).

         The short-term toxicity of technical glyphosate was also
    studied in dogs. Beagle dogs received technical glyphosate in
    gelatin capsules at dose levels of 0, 20, 100 or 500 mg/kg body
    weight per day for 52 weeks. No effect occurred with respect to
    clinical signs, body weight, feed consumption, ophthalmoscopy,
    haematology, blood biochemistry, urinalysis, gross pathology and
    histopathology. The only changes in treated groups relative to
    controls were increased pituitary weights (absolute and relative) in
    the medium- and high-dose males. Because there were no concomitant
    histological changes present in pituitaries and given the absence of
    an effect on this organ (and related organs) in all other studies,
    the toxicological significance of the increased pituitary weights is
    questionable. The NOAEL in this study was 500 mg/kg body weight per
    day (Monsanto, 1985).

         A 7-day oral study was carried out with Roundup in female
    cattle weighing 160 to 272 kg. Brahman-cross heifers received 400,
    500, 630 or 790 mg Roundup/kg body weight per day by nasogastric
    tube. At 790 mg/kg, 1/3 animals died before test end, showing

    laboured breathing and pneumonia caused by aspiration of rumen
    contents. Decreased feed intake was seen at 630 and 790 mg/kg;
    diarrhoea occurred at 500, 630 and 790 mg/kg body weight. Slight
    increases in a number of blood parameters, occurring at 790 mg/kg
    only, were probably due to extracellular fluid shifts and
    haemoconcentration secondary to diarrhoea. The NOAEL in this study
    was 400 mg Roundup/kg body weight per day (USDA, 1987a).

    7.2.2  Dermal studies

         Short-term dermal toxicity studies were carried out in rabbits
    with technical glyphosate and the formulation Roundup.

         Technical glyphosate, moistened with saline, was applied under
    occlusion at dose levels of 100, 1000 and 5000 mg/kg body weight per
    day to the shaven intact or abraded skin of rabbits for 6 h/day, 5
    days/week for 3 weeks. No effect on survival and growth occurred.
    Slight dermal irritation (barely perceptible erythema and oedema)
    was observed at 5000 mg/kg body weight only. No evidence of systemic
    toxicity was found (parameters: haematology and blood biochemistry
    in five animals of each sex per group, organ weights, gross
    pathology, limited histopathology) (IRDC, 1982). Absence of systemic
    effects was also found in the somewhat more limited 21-day study in
    rabbits (no haematology and blood biochemistry) with Roundup. Skin
    effects were more severe in this study: at both dose levels (76 and
    114 mg/kg body weight per day, undiluted formulation applied)
    erythema and oedema were seen and, in addition, exudate and
    fissuring occurred at the abraded skin sites. After a 4-week
    recovery period the skin effects were no longer present
    (Bio/Dynamics Inc., 1975).

    7.2.3  Inhalational studies

         A 4-week inhalation study was carried out on rats with a 1:3
    dilution of Roundup formulation. Test concentrations of 50, 160 and
    360 mg/m3 of the diluted formulation (equivalent to 17, 53 and
    120 mg Roundup/m3) were administered as an aerosol spray for
    6 h/day, 5 days/week. The mass median aerodynamic diameter of the
    test material ranged from 1.8 to 2.7 µm with geometric standard
    deviations between 1.7 and 2.0. An increased incidence of irritation
    of the nasal turbinates (subacute inflammation), trachea
    (mononuclear cell infiltration) and lungs (perivascular lymphoid
    infiltrates/aggregates) was observed among the high-dose females
    only. No signs of systemic toxicity were found (parameters:
    survival, growth, limited haematology and blood biochemistry, organ
    weights, limited histopathology) (Monsanto, 1983e).

    7.3  Long-term toxicity and carcinogenicity

         Only oral studies are available. Dietary studies using
    technical glyphosate were performed in mice and rats.

         In Charles River CD-1 mice technical glyphosate was fed in the
    diet at concentrations of 0, 1000, 5000 or 30 000 mg/kg diet for 24
    months (dose levels equal to 0, 157, 814 and 4841 mg/kg body weight
    per day for males and 0, 190, 955 and 5874 mg/kg body weight per day
    for females). No effect on survival or appearance was noted. Body
    weights were decreased in the males of the high-dose group.
    Haematology and organ weights showed no effects. Histopathology in
    liver revealed an increased incidence of central lobular hepatocyte
    hypertrophy among high-dose males (incidences: 9/49, 5/50, 3/50 and
    17/50 in control, low-dose, medium-dose and high-dose males,
    respectively) and an increased incidence of central lobular
    hepatocyte necrosis also among high-dose males (incidences: 0/49,
    2/50, 2/50 and 10/50). Increased incidences of epithelial
    hyperplasia in the urinary bladder were present in the medium-dose
    and high-dose males (incidences: 3/49, 3/50, 10/50 and 8/50). There
    were no statistically significant increases in the frequency of
    neoplastic lesions. The NOAEL in this study was 5000 mg/kg diet
    (814 mg/kg body weight per day) (Bio/Dynamics Inc., 1983a).

         Two chronic feeding studies on rats were conducted with
    technical glyphosate, one in 1979-1981 and the other in 1988-1990.
    The first study, carried out using Charles River CD (Sprague-Dawley)
    BR rats, had dose levels of 0, 60, 200 and 600 mg technical
    glyphosate/kg diet (equal to about 0, 3, 10 and 32 mg/kg body weight
    per day, respectively). Survival, appearance, haematology, blood
    biochemistry, urinalysis and organ weights were not changed. The
    NOAEL was > 600 mg/kg diet (> 32 mg/kg body weight per day).
    Slight growth retardation during part of the study was noted in the
    high-dose males. The incidence of interstitial cell tumours in
    testes showed a statistically significant increase (incidences:
    0/50, 3/50, 1/50 and 6/50; historical control range: 3-7%)
    (Bio/Dynamics Inc., 1981a). This finding, in itself constituting
    some evidence for a carcinogenic effect in rats, should be judged in
    the light of the absence of an effect at much higher dose levels in
    the more recent 2-year study in rats (see below). This is also valid
    for the slight growth retardation (i.e. no effect on growth at much
    higher dose levels in the more recent study, see below). In the
    recent 2-year study, rats (same strain) were fed 2000, 8000 or 20
    000 mg technical glyphosate/kg diet (equal to about 100, 410 and
    1060 mg/kg body weight per day) for 24 months. There was no effect
    on survival or appearance. Growth was retarded in the high-dose
    females. Haematology and blood biochemistry showed no effects. In
    the high-dose males, the urine specific gravity (after 6 months
    only) and urine pH were increased. A statistically significant
    increased incidence of degenerative lens changes (basophilic
    degeneration of the posterior subcapsular lens capsule or mature
    cataracts) was found among the high-dose males (incidences 3/60,
    4/60, 4/60 and 8/60 in the control, low-, medium- and high-dose
    groups, respectively. However this finding was within the historical
    control range of 0-33%. Liver weights were increased in the
    high-dose males only. Histopathology showed an increased incidence


    
    Table 12.  Skin irritation tests on rabbits with glyphosate and its formulations
                                                                                                                                              

    Product testeda                         Contact time (h)      Draize scoreb    Classificationc           Reference
                                                                                                                                              

    Glyposate

    Glyphosate techn. 85% in water                  4                  0.8         slightly irritating        Bio/Dynamics Inc. (1988b)
    Glyphosate techn., moistened powder             4                  0           not irritating             FDRL (1988a)
    Glyphosate techn., moistened powder             4                  0           not irritating             Inveresk Research Int. (1989e)
    Glyphosate, IPA salt 65% in water              24                  0.2         not irritating             Monsanto (1981d)

    Formulations

    Roundup, undiluted                              4                  1.9         slightly irritating        Bio/Dynamics Inc. (1988g)
    "Compound No. 3607", undiluted                  4                  1.2         slightly irritating        Inveresk Research Int. (1988c)
    Roundup TX, undiluted                           4                  0.7         slightly irritating        NOTOX (1987d)
    Alphee, undiluted                               4                  1.0         slightly irritating        Bio/Dynamics Inc. (1987c)
    Sting, undiluted                                4                  1.3         slightly irritating        Bio/Dynamics Inc. (1984c)
    Sting TX, undiluted                             4                  3.6         moderately irritating      NOTOX (1987i)
    Roundup L&G, undiluted                          4                  1.0         slightly irritating        Bio/Dynamics Inc. (1985c)
    "Glyfosaat 360 g/litre, undiluted               4                  0.3         not irritating             NOTOX (1989d)
    Agrichem Glyfosaat B, undiluted                 4                  1.7         slightly irritating        NOTOX (1990d)
    Agrichem Glyfosaat 2, undiluted                 4                  2.0         slightly irritating        NOTOX (1990b)
    Legend, undiluted                               4                  0.7         slightly irritating        CIT (1991c)
                                                                                                                                              

    a    For glyphosate content of the tested formulations, see footnote a in Table 11.
    b    This score is the mean score per animal and was calculated using the data from
         the original report. The standard Draize scoring system was used; in calculating
         the mean response per animal, the maximum response observed for an animal was used.
    c    The classification is based on the mean Draize score per animal. Specification:
         score 0-0.5 not irritating; 0.6-2.0 slightly irritating; 2.1-5.0 moderately
         irritating; 5.1-8.0 severely irritating.

    Table 13.  Results of eye irritation tests in rabbits for glyphosate and its formulations.
                                                                                                                                              

    Product testeda                                 Irritation indexb      Classificationc          Reference

                                                                                                                                              
    Glyphosate

    Glyphosate techn. 85% in water, undiluted             45               strongly irritating      Bio/Dynamics Inc. (1988d)
    Glyphosate, IPA salt, 65% in water                     0               not irritating           Monsanto (1981e)
    Glyphosate techn. (97.6%), undiluted                  54               strongly irritating      FDRL (1988c)
    Glyphosate techn. (96-99%), undiluted                 29               irritating               Inveresk Research Int. (1989f)

    Formulations

    Roundup, undiluted                                    54               strongly irritating      Bio/Dynamics Inc. (1990)
    "Compound No. 3607", undiluted                        13               irritating               Inveresk Research Int. (1989g)
    Roundup TX, undiluted                                 31               strongly irritating      NOTOX (1987e)
    Alphee, undiluted                                      6               slightly irritating      Bio/Dynamics Inc. (1987d)
    Sting, undiluted                                     104               extremely irritating     Bio/Dynamics Inc. (1984d)
    Sting TX, undiluted                                   19               irritating               NOTOX (1987j)
    Roundup L&G, undiluted                                19               irritating               Bio/Dynamics Inc. (1985d)
    "Glyfosaat 360 g/litre", undiluted                    33               strongly irritating      NOTOX (1989a)
    Agrichem Glyfosaat B, undiluted                       58               strongly irritating      NOTOX (1990c)
    Agrichem Glyfosaat 2, undiluted                       22               irritating               NOTOX (1990a)
    Legend, undiluted                                     21               irritating               CIT (1991d)
                                                                                                                                              

    Table 13 (continued)
    a    For glyphosate content of the tested formulations, see footnote a in Table 11.

    b    The irritation index represents a mean total score per animal for response in cornea, conjunctiva
         and iris, using the standard Draize scoring system. The irritation index was calculated using the data
         from the original report; in calculating the mean response per animal the maximum response observed
         for an animal was used. The calculation procedure is as follows:

              -    score conjunctiva:  A. chemosis: 0-4    Calculation partial
                                       B. discharge: 0-3   index for conjunctiva:
                                       C. erythema: 0-3    2 x (A + B + C) = 0-20
              -    score iris:         0-2                 Calculation partial index for iris: 5 x (0-2) = 0-10
              -    score cornea:       A. opacity: 0-4     Calculation partial index for cornea:
                                       B. area: 0-4        5 x A x B = 0-80

              The total irritation index is the sum of the partial indices (0-110).

    c         The classification is based on the following scheme: index 0-5 not irritating; 5.1-12 slightly
              irritating; 12.1-30 irritating; 30.1-60 strongly irritating; 60.1-80 severely irritating; 80.1-110
              extremely irritating.
    

    of inflammation of the gastric squamous mucosa in the medium- and
    high-dose groups (incidences in males: 2/58, 3/58, 5/59 and 7/59;
    females: 0/59, 3/60, 9/60, and 6/59; historical range: 0-13.3%). The
    incidence of pancreatic islet cell adenomas was increased
    (statistically significant) among low- and high-dose males
    (incidences: 1/58, 8/57, 5/60 and 7/59; historical control range of
    test laboratory 1.8-8.5%). The incidence in the control group was
    below the historical control range; the trend test for the observed
    increase was negative. No pancreatic carcinomas were found. The
    NOAEL in this study was 8000 mg/kg diet (410 mg/kg body weight per
    day) (Monsanto, 1990b).

         Glyphosate has been tested in the US National Toxicology
    Program; pre-chronic studies have been completed (NTP, 1992).

    7.4  Skin and eye irritation; sensitization

         Many studies have been carried out with rabbits to examine the
    potential of glyphosate and its formulations to produce skin and eye
    irritation. The results of these studies are briefly summarized in
    Tables 12 (skin irritation) and 13 (eye irritation). Glyphosate and
    its formulations produce only mild skin irritation after undiluted
    single application. The result of the 21-day dermal study in rabbits
    with the formulation Roundup (Bio/Dynamics Inc., 1975; see
    subsection 7.2.2) shows that repeated application of undiluted
    formulation to the skin does lead to irritation. The eye-irritating
    potential is considerable for undiluted glyphosate and, with a few
    exceptions, also for the formulations. Single application generally
    produces moderate to severe reactions.

         Sensitization studies have been carried out in guinea-pigs with
    glyphosate and its formulations. The results of these tests are
    summarized in Table 14. Neither glyphosate itself nor the tested
    formulations induced sensitization in any experiment.

         The results of two dermal irritation studies and one dermal
    sensitization study performed with human volunteers exposed to
    Roundup are presented in subsection 8.1.

    
    Table 14. Results of sensitization tests in guinea-pigs for glyphosate and its
              formulations
                                                                                       

    Product testeda          Method            Result           Reference
                                                                                       

    Glyphosate

    Glyphosate               Buehler           negative         Bio/Dynamics Inc.
    (purity 99.7%)                                              (1983c)

    Glyphosate technical     Magnusson &       negative         Safefarm Labs Inc.
                             Kligmanb                           (1991b)

    Glyphosate techn.        Magnusson &       negative         Inveresk Research
    (96-99%)                 Kligman                            Int. (1989b)

    Formulations

    Roundup                  Buehler           negative         Bio/Dynamics Inc.
                                                                (1983b)

    "Compound                Magnusson &       negative         Inveresk Research
    No. 3607"                Kligman           Int.             (1988d)
    Roundup L&G              Buehler           negative         Bio/Dynamics Inc.
                                                                (1987e)
    Legend                   Buehler           negative         Safefarm Labs Inc.
                                                                (1991a)
    Sting                    Buehler           negative         Bio/Dynamics Inc.
                                                                (1986)
                                                                                       

    a    For glyphosate content of the tested formulations, see footnote a
         in Table 11.
    b    Method also called the maximization test.
    
    7.5  Reproductive toxicity, embryotoxicity and teratogenicity

         Technical glyphosate has been tested for teratogenicity in rats
    and rabbits using the oral route. In addition, two oral
    multi-generation reproduction studies in rats have been reported.

    7.5.1  Teratogenicity studies

         Glyphosate (technical) was given to pregnant Charles River COBS
    CD rats by gavage at dose levels of 0, 300, 1000 and 3500 mg/kg body
    weight per day on days 6-19 of gestation. At 3500 mg/kg the
    following effects were observed: increased incidences of soft
    stools, diarrhoea, breathing rattles, red nasal discharge and
    reduced activity, increased mortality (6/25 dams dying before the
    end of the treatment period), growth retardation, increased
    incidence of early resorptions, decreases in total number of
    implantations and the number of viable fetuses, increased number of
    fetuses with reduced ossification of sternebrae. At the lower dose
    levels these effects were absent. The NOAEL in this study was
    1000 mg/kg body weight per day (IRDC, 1980b).

         In Dutch belted rabbits technical glyphosate was tested at dose
    levels of 0, 75, 175 and 350 mg/kg body weight per day
    (administration by gavage) from days 6-27 of gestation. In dams the
    incidences of diarrhoea and soft stools were increased in the
    high-dose group and, to a slight degree, also in the medium-dose
    group. The incidence of nasal discharge was increased in the
    high-dose group only. In the medium- and high-dose groups 2 and 10
    dams, respectively, died during the study from unknown causes. The
    NOAEL in this study was 175 mg/kg body weight per day (IRDC, 1980c).

    7.5.2  Reproduction studies

         A three-generation feeding study in Charles River CD
    (Sprague-Dawley) BR rats was conducted in 1980-1981 and a
    two-generation study, using higher dose levels, was completed in
    1990 with technical glyphosate. In the former study, dietary feeding
    levels were continuously adjusted to achieve dose levels of 0, 3, 10
    and 30 mg/kg body weight per day. The only effect noted was an
    increased incidence of unilateral renal tubular dilation in the male
    pups (randomly selected) of the F3b mating of the high-dose group
    (incidence 6/10 versus 0/10 in controls, not determined in
    intermediate groups, earlier litters not examined). The NOAEL in
    this study was < 30 mg/kg body weight (Bio/Dynamics Inc., 1981b).
    The more recent two-generation study had dose levels of 0, 2000,
    10 000 and 30 000 mg technical glyphosate/kg diet (equivalent to 0,
    100, 500 and 1500 mg/kg body weight per day). In the high-dose
    group, the following effects were observed: soft stools and
    decreased body weights in parent animals, slightly decreased litter
    size and decreased pup weights (the latter seen at day 14 or 21 of
    lactation). The decreased body weights of parents and pups were seen
    to a slight degree in the medium-dose group also. No histological
    effect on kidneys was present in the F2b male pups (15 and 23 pups
    examined in control and high-dose groups, respectively; first
    generation and F2a pups not examined). The NOAEL in this study was
    10 000 mg/kg diet (500 mg/kg body weight per day) (Monsanto, 1990c).

         With regard to these reproduction studies using technical
    glyphosate, it should be noted that in both studies the number of
    pups submitted to histopathological examination was limited. These
    limitations make evaluation of the renal effect in pups, seen (at
    30 mg/kg body weight) in the study of Bio/Dynamics Inc. (1981b),
    difficult.

    7.6  Mutagenicity and related end-points

         The results of several studies are summarized in Table 15. The
    results show that glyphosate is not mutagenic.

    
    Table 15.  Results of mutagenicity studies with glyphosate and its salts.
                                                                                               

    Test system                         Test compound and              Resultc     Reference
                                         concentrations
                                                                                               

    Ames test,  Salmonella               technical glyphosate             -         Monsanto
     typhimurium TA98, TA100,            (98.4%); 0.1-1000                          (1978c)
    TA1535 & TA1537, with and           µg/plate
    without metabolic activation

    Ames test,  S. typhimurium           technical glyphosate             -         IET (1978)
    TA98, TA100, TA1535 &               (98.4%); 10-5000
    TA1537, with and without            µg/plate
    activation

    Rec assay,  Bacillus subtilis,       technical glyphosate             -         IET (1978)
    strain H17 (rec+) & M45             (98.4%); 20-2000
    (rec-), without activation          µg/disc

    Reverse mutation assay in           technical glyphosate             -         IET (1978)
     Escherichia coli strain WP2hcr,     (98.4%); 10-5000
    with and without activation         µg/plate

    Forward mutation assay,             technical glyphosate             -         Monsanto
    CHO cells,  in vitro                 (98.7%); 0-20 mg/ml                        (1983b)
    (HGPRT-locus), with &               (- activation) or
    without activation                  5-25 mg/ml (+ activation)
                                                                                               

    Table 15.  (cont'd)
                                                                                               

    Test system                         Test compound and              Resultc     Reference
                                         concentrations
                                                                                               

    Cytogenetic study                   technical glyphosate             -         Monsanto
    (chromosome aberrations)            (98.7%); 200-1000 mg/kg                    (1983f)
    in rat bone marrow,                 body weight, i.p.b;
     in vivo                             sampling after 6, 12
                                        and 24 h

    Micronucleus test in                glyphosate (not                  -         Benova
    erythrocytes of mice,               specified); ´ LD50, oral;                  et al.
     in vivo                             sampling time not                          (1989)
                                        specified

    Dominant lethal test, mouse         technical glyphosate             -         IRDC
     in vivo                             (98.7%); 200-2000 mg/kg                    (1980a)
                                        body weight, oral

    Recessive sex-linked lethal         glyphosate (not                  -         Gopalan &
    test,  Drosophila melanogaster,      specified); dose not                       Njagi
     in vivo                             given                                      (1981)

    Unscheduled DNA repair assay        technical glyphosate             -         Monsanto
    rat hepatocytes,  in vitro           (98.7%); 0.0125-125                        (1983c)
                                        µg/ml
                                                                                               

    a    No higher concentrations tested because these would result in
         osmolalities much  higher than physiological levels; these high
         osmolalities can produce non-specific  chromosomal aberrations or
         sister chromatid exchange.

    b    In additional studies it was demonstrated that: (1) glyphosate
         produced no effect on viability and mitotic index of bone marrow
         cells of rats after i.p. doses of 200-1000 mg/kg body weight
         (Monsanto, 1983g); and (2) after giving 14C-labelled glyphosate
         i.p. significant concentrations of 14C reached the bone marrow
         (peak levels reached after 0.5 h remaining virtually constant up to
         10 h after dosing) (Monsanto, 1983h).

    c    - = negative result
    
    8.  EFFECTS ON HUMANS

    Appraisal

          The formulation Roundup containing glyphosate is acutely toxic
     to humans when ingested intentionally or accidentally.

          No controlled studies have been conducted, and therefore the
     human NOAEL level cannot not be derived.

          No data are available to show the impact on workers exposed
     during the manufacture or formulation of glyphosate. No
     compound-related effects were observed in a test group of five
     applicators prior to and after exposure for one week.

          The reported higher susceptibility of individuals older than
     40 years to ingested Roundup intoxication is important and requires
     further investigation.

    8.1  Cases of intentional and accidental exposure

         Many cases of acute intoxication with herbicides containing
    glyphosate and surfactant (Roundup) have been reported; most of
    these were suicide attempts. Talbot et al. (1991) reviewed 93 cases
    of exposure to Roundup (Chinese names: lan-da, hao-ni-chun,
    nian-nian-chun) in Taiwan. The classification of the severity of
    acute poisoning with Roundup as given by these authors is presented
    in Table 16. Severe effects occurred only in the cases of
    intentional ingestion (80 of the 93 reported). Accidental exposures
    led to only mild effects. The typical symptoms were erosion of the
    gastrointestinal tract (66% of the self-poisonings), seen as sore
    throat, dysphagia and gastrointestinal haemorrhage. Other organs
    were affected less often (nonspecific leucocytosis 65%, lungs 23%,
    liver 19%, cardiovascular system 18%, kidney 14% and CNS 12%). Death
    (in 7/80 cases) occurred within hours after ingestion. The amount of
    undiluted Roundup ingested (rough estimates) in the lethal cases
    varied from 85 to 200 ml (corresponding to roughly 30 to 70 g
    glyphosate acid); but much larger amounts (500 ml Roundup,
    corresponding to 180 g glyphosate acid) were reported to have been
    ingested by some patients with mild to moderate symptoms. Overall,
    moderate symptoms were associated with estimated intakes of 20 to
    500 ml, mild symptoms with 5 to 150 ml, no symptoms with 5 to 50 ml.
    The authors pointed out that the patient's estimates of the amount
    ingested, and the conversion ratio used in their paper may be
    inaccurate (Talbot et al., 1991). Other reviews of cases of
    intoxication with Roundup have reported similar findings (Sawada &
    Nagai, 1987; Tominack et al., 1991). The data of Tominack et al.
    (1991) suggested that people over 40 years of age who ingest amounts
    greater than 150 ml Roundup are at greatest risk of a fatal outcome.

    These authors also pointed out that the surfactant contained in
    Roundup may be responsible for the clinical syndrome (as suggested
    by Sawada & Nagai, 1987), but that the available evidence on this
    point is, as yet, inconclusive.

    
    Table 16. Classification of severity of acute poisoning with Roundupa
                                                                                            

    Classification                                Description
                                                                                            

    Asymptomatic      no complaints and no abnormalities on physical or laboratory
                      examination.

    Mild              mainly gastrointestinal tract(GIT) symptoms (nausea, vomiting,
                      diarrhoea, abdominal pain, mouth and throat pain) that resolved
                      within 24 h. Vital signs were stable, and there was no renal,
                      pulmonary or cardiovascular involvement.

    Moderate          GIT symptoms lasting longer than 24 h, GIT haemorrhages, endoscopically
                      verified oesophagitis or gastritis, oral ulceration, hypotension
                      responsive to intravenous fluids, pulmonary dysfunction not
                      requiring intubation, acid-base disturbance, evidence of transient
                      hepatic or renal damage, or temporary oliguria.

    Severe            pulmonary dysfunction requiring intubation, renal failure requiring
                      dialysis, hypotension requiring treatment with pressor amines, cardiac
                      arrest, coma, repeated seizures, or death.
                                                                                            

    a    From: Talbot et al. (1991)
    
         Further clinical experiences with patients exposed to Roundup
    either accidentally or through deliberate ingestion have been
    reported by Temple & Smith (1992). Symptoms resulting from dermal
    exposure incidental to the use of the product included periorbital
    oedema and chemosis of the eye, cardiovascular effects (tachycardia
    and elevated blood pressure), swelling and paraesthesia at the site
    of dermal contact and prolonged skin irritation. Deliberate
    ingestion resulted in more severe effects, including lethality from
    apparent respiratory and cardiac arrest (Temple & Smith, 1992).

         Two dermal irritation studies were carried out with volunteers.
    Application of 0.9 ml of a 9:1 dilution of Roundup formulation in
    water to the intact skin of the upper arm for 24 h produced no skin
    changes (Shelanski, 1973). Maibach (1986) tested undiluted Roundup
    (application of 0.1 ml to intact and abraded skin sites on the back
    for 24 h) and found erythema in only 1/24 subjects (23/24 no
    reaction) for the intact skin sites; for the abraded skin sites 4/24
    subjects showed an equivocal reaction and 10/24 showed erythema
    (10/24 no reaction). The same author reported very briefly the
    absence of effect in a photoirritation study in humans using
    undiluted Roundup as test compound (application to abraded skin of
    upper arm for 24 h with irradiation with UVA light for 45 min)
    (Maibach, 1986).

         A sensitization study was performed in 204 human volunteers
    with undiluted Roundup according to a modified Draize method. The
    summary report (no detailed report available) stated that there was
    no effect in any subject (Maibach, 1986). The same author reported
    absence of photosensitization by Roundup in volunteers.

    8.2  Occupational exposure

         The results of several studies focused primarily on the
    determination of the extent of exposure to glyphosate when the
    compound is used as herbicide are presented in section 5.3. The
    study of Jauhainen et al. (1991) included health examinations of a
    test group of five workers prior to and after an exposure period of
    1 week. These examinations included haematology, clinical chemistry,
    ECG, pulmonary function tests, an interview for a health
    questionnaire and a general clinical examination (including
    recording of blood pressure, pulse rate and pressure craft of
    hands). A control group consisted of five workers. No
    compound-related effects were observed (Jauhainen et al., 1991). The
    other studies described in section 5.3 did not include a health
    evaluation of workers.

    8.3  Subpopulations at special risk

         The only information available on this point is some suggestive
    evidence referring to oral intoxications with Roundup; Tominack
    et al. (1991) suggested that people older than 40 years are at
    greater-than-normal risk after ingestion of Roundup.

    9.  EFFECTS ON OTHER ORGANISMS IN THE LABORATORY AND
    FIELD

         In this chapter, concentrations or doses of formulations with
    glyphosate are always expressed as mg product per kg soil.
    Therefore, these figures may have been recalculated from the
    original data, in cases where the authors reported the data in, for
    instance, mg a.i./litre instead of mg Roundup per litre. For
    recalculation, the percentages of the active ingredient (free acid
    or the salt), given in Table 2, were used.

    9.1  Laboratory experiments

    9.1.1  Microorganisms

    9.1.1.1  Water

    Appraisal

          The prokaryotic cyanobacteria are generally more sensitive to
     the effects of glyphosate than the eukaryotic true algae. Similar
     enzyme systems are inhibited in microorganisms to those thought to
     be responsible for the herbicidal properties of glyphosate in higher
     plants. The single semi-field study suggests very variable results
     but with no significant effect on populations or community
     structure.

         The acute and chronic toxicities of technical grade glyphosate
    and its formulations to cyanobacteria, algae and diatoms are
    summarized in Table 17. Technical grade glyphosate is slightly toxic
    with 3- to 4-day EC50 values of 1.2-7.8 mg/litre, and 7-day NOEC
    values of 0.3-34 mg/litre. Formulations of glyphosate may be more
    toxic (3-day EC50 values of 1.0 to > 55 mg product/litre).

         Toxicity to cyanobacteria and algae is dependent on the species
    or strain tested. Wängberg & Blanck (1988) exposed 16 species in
    pure cultures to Roundup for 14 days. The concentration at which
    growth was inhibited completely was 16 mg Roundup/litre for the most
    sensitive species  (Raphidonema longiseta and  Anabaena sp.) and
    131 mg Roundup/litre for the least sensitive species  (Selenastrum
     capricornutum). The prokaryotic Cyanophyta were significantly more
    affected by Roundup than the eukaryotic  Chlorococcales.

         In  Pseudomonas chlororaphis Roundup severely inhibited
    respiration at concentrations of > 2623 mg/litre, whereas in
     Aeromonas hydrophila respiration was only slightly affected at
    these concentrations (Chan & Leung, 1986). The bacteria were exposed
    for 6 days.


    
    Table 17.  Aquatic microorganisms: acute and chronic toxicity of glyphosate and its formulations (EC50, NOEC)
                                                                                                                                              

    Organism            A    Test  Compound   Test     pH         Hardness     Temperature   Experimental  Parametera Concentration   Reference
                             type  water                          (mg CaCO3/   (°C)          duration                 (mg/litre)
                                                                  litre)                     (days)
                                                                                                                                              

    Cyanobacteria

     Anabaena            A    S     Tgg        a.m.     7.5        285          24            7             NOEC       9.7b,c          Malcolm
     flos-aquae                                                                                                                        Pirnie
                                                                                                                                      Inc.
                                                                                                                                      (1987a)

    Green algae

     Selenastrum         -    S     Tgg        a.m.     7.0        n.r.         26            3.5-4         EC50       7.8b,e          Bozeman
     capricornutum                                                                                                                     et al.
                                                                                                                                      (1989)
     S. capricornutum    A    S     Tgg        a.m.     7.5        285          24            7             NOEC       20b,c           Malcolm
                                                                                                                                      Pirnie
                                                                                                                                      Inc.
                                                                                                                                      (1987d)
     S. capricornutum    -    S     St         a.m.     7.7-9.0    26           23            3             EC50       1.0b            LISEC
                                                                                                                                      (1989b)
     S. capricornutum    -    S     St         a.m.     7.7-9.0    26           23            3             EC50       2.5d            LISEC
                                                                                                                                      (1989b)
     S. capricornutum    -    S     St         a.m.     7.7-9.0    26           23            3             NOEC       0.2b            LISEC
                                                                                                                                      (1989b)
     S. capricornutum    -    S     Ru         a.m.     7.7-10.0   26           24            3             EC50       2.1b            LISEC
                                                                                                                                      1989a)
     S. capricornutum    -    S     Ru         a.m.     7.7-10.0   26           24            3             EC50       8.0d            LISEC
                                                                                                                                      (1989a)
     S. capricornutum    -    S     Ru         a.m.     7.7-10.0   26           24            3             NOEC       0.7b            LISEC
                                                                                                                                      (1989a)
                                                                                                                                              

    Table 17 (contd).
                                                                                                                                              

    Organism            A    Test  Compound   Test     pH         Hardness     Temperature   Experimental  Parametera Concentration   Reference
                             type  water                          (mg CaCO3/   (°C)          duration                 (mg/litre)
                                                                  litre)                     (days)
                                                                                                                                              

     Chlorella           -    S     Ru         a.m.     7.5        n.r.         25            2.1-7         EC50       > 55b           Hernando
     pyrenoidosa                                                                                                                       et al.
                                                                                                                                      (1989)

     C. pyrenoidosa      -    S     Ru         a.m.     7.5        n.r.         25            2.1-7         NOEC       < 55b           Hernando
                                                                                                                                      et al.
                                                                                                                                      (1989)

    Diatoms

     Skeletonema         -    S     Tgg        a.m.h    8.2-8.5    n.r.         20            4             EC50       1.2g            E G &
     costatum                                                                                                                          Bionomics
                                                                                                                                      (1978a)
     S. costatum         -    S     Tgg        a.m.h    8.2-8.5    n.r.         20            4             EC50       1.3b            E G &
                                                                                                                                      Bionomics
                                                                                                                                      (1978a)
     S. costatum         -    S     Tgg        a.m.h    8.2-8.5    n.r.         20            4             NOEC       < 0.6f          E G &
                                                                                                                                      Bionomics
                                                                                                                                      (1978a)
     S. costatum         A    S     Tgg        a.m.h    7.5        285          20            7             NOEC       0.3b,c          Malcolm
                                                                                                                                      Pirnie
                                                                                                                                      Inc.
                                                                                                                                      (1987b)
     Navicula            A    S     Tgg        a.m.     7.5        285          20            7             NOEC       34b,c           Malcolm
     pelliculosa                                                                                                                       Pirnie
                                                                                                                                      Inc.
                                                                                                                                      (1987c)
                                                                                                                                              

    Table 17 (continued)

    a    concentration of a formulation is expressed as mg of the formulation per litre
    b    based on biomass decrease
    c    based on actual concentrations
    d    based on inhibition of growth rate
    e    NOEC value not reported
    f    based on biomass and chlorophyll a decrease
    g    based on chlorophyll a decrease
    h    salinity was 30%
         A = actual concentrations are measured; - = nominal concentrations; S = static system;
         Tgg = technical grade glyphosate; Ru = Roundup;
         St = Sting; a.m. = artificial medium; n.r. = not reported
    

         Chan & Leung (1986) found that the activity of
    5-enolpyruvyl-shikimic acid-3-phosphate synthase was inhibited
    strongly in aquatic bacteria at the lowest tested concentration of
    656 mg Roundup/litre. This enzyme takes part in the biosynthetic
    sequence to phenylalanine, tryptophan, and tyrosine via the
    conversion of shikimate to chorismate, and the conversion of
    chorismate to anthralinate. In an  in vitro experiment with
    cell-free extracts of  Aerobacter aerogenes, technical grade
    glyphosate inhibited the conversion of shikimate to chorismate at
    concentrations of > 0.2 mg a.i./litre (Amrhein et al., 1980).

         In  Chlorella pyrenoidosa Roundup affected the growth, the
    greening process and the photosynthetic metabolism at the lowest
    tested concentration of 55 mg/litre when exposed for 2.1 or 7 days
    (Hernando et al., 1989). Synthesis of chlorophyll a and b and
    carotenoids was then significantly inhibited. Roundup affected
    synthesis of chlorophyll a to a greater extent than that of the
    other pigments.  In vivo and  in vitro studies with isolated
    chloroplasts showed inhibition of the photosystems PS I and PS II at
    concentrations of > 55 mg Roundup/litre. Hernando et al. (1989)
    suggested that glyphosate acted as an electron transport inhibitor
    and that the stronger inhibition of PS II compared with PS I was due
    to the surfactant polyoxyethyleneamine.

         In periphytic algal communities that were collected in ponds of
    boreal forests, Roundup decreased the carbon fixation rate by 50% at
    concentrations of 243-479 mg/litre (Goldsborough & Brown, 1988). The
    algae were exposed for 4 h. Austin et al. (1991) cultured periphyton
    on glass plates suspended in artificial "stream-troughs" which were
    supplied with flowing water pumped from natural streams in British
    Columbia, Canada. The stream water was low in phosphorus and flowed
    out of an oligotrophic lake. Glyphosate was added to give nominal
    concentrations in the troughs of between 0.001 and 0.3 mg/litre. A
    further series of treatments added nutrients to troughs. The
    herbicide was not toxic to the periphyton. A transitory decrease in
    growth was followed by a stimulation of biomass in the
    glyphosate-treated troughs. Similar effects were seen with added
    nutrient. The authors considered the effect to be the result of
    algae using glyphosate as a phosphate source. Communities of
    periphyton were similar in all treatments.

         Various studies have indicated that glyphosate may affect
    aromatic amino acid synthesis in microorganisms, in addition to the
    greening process, respiration and photosynthesis (Amrhein et al.,
    1980; Chan & Leung, 1986; Pipke & Amrhein, 1988).

    9.1.1.2  Soil

    Appraisal


          Soil bacteria in culture have shown effects of glyphosate on
     nitrogen fixation, denitrification and nitrification. However, field
     studies after application of formulations have not shown significant
     effects. Closely related species of bacteria have been shown to be
     capable of degrading glyphosate (see chapter 4). The lack of
     information on the bioavailability of glyphosate in soil makes it
     difficult to relate the evaluation of effects in culture to the
     actual exposure in the field.

         Technical grade glyphosate inhibited the growth of bacteria
    isolated from a sprayed garden soil of sandy loam to a lesser extent
    than it did that of bacteria isolated from an unsprayed control
    (Quinn et al., 1988). In a mineral salt medium the growth rate of
    bacteria from the untreated site was reduced by 50% at 590 mg
    a.i./litre at 50 h after application.

         In various studies, glyphosate, applied at the recommended
    rates, caused neither inhibition of processes that are part of the
    nitrogen cycle nor inhibition of enzymes involved in microbial
    activity.

         Roundup did not affect dehydrogenase activity in sand and loamy
    sand when applied at rates of 4.9 and 24 kg a.i./ha (NATEC, 1990).
    In the same soils amended with lucerne  (Medicago sativa), the same
    application rates did not change significantly the amounts of
    nitrate nitrogen, although in treated loamy sand there was a slight
    increase. These experiments, with a duration of 28 days, indicated a
    slight stimulation of nitrification in loamy sand. A stronger
    stimulation of nitrification was found in silt loam, silty clay loam
    and sandy loam up to 84 days after application of technical grade
    glyphosate at rates of 5 and 25 mg a.i./kg (ABC Inc., 1978d). This
    stimulation was not dose-related. In this experiment no
    substance-related effects were found on nitrogen fixation and
    nitrite formation.

         Nitrogen fixation was not affected significantly at an
    application rate of 13 mg a.i./kg dry weight whether the product
    applied was technical grade glyphosate or Roundup, or whether
    aerobic or anaerobic conditions were used (Carlisle & Trevors,
    1986a). At concentrations > 127 mg a.i./kg dry weight, nitrogen
    fixation was inhibited under anaerobic conditions, whereas this
    could not be verified under aerobic conditions due to very low
    acetylene reduction rates. In the same agricultural sandy loam,
    denitrification under anaerobic conditions was stimulated at
    concentrations of > 13 mg a.i./kg dry weight, whether technical

    grade glyphosate or Roundup was applied. Denitrification was
    stimulated more strongly, when, in addition to the test compound,
    glucose was added. In the same soil, no substantial effects of
    either test compound on nitrification were found at 77 mg a.i./kg
    dry weight. Dose-related inhibition of nitrification was found at
    concentrations > 230 mg a.i./kg dry weight for glyphosate and
    Roundup with respect to nitrate and nitrite production,
    respectively. For both substances the inhibition was transient in
    these studies. The temperature used was not reported.


         No substantial effects on nitrification or denitrification were
    found in two agricultural soils from Southern Finland after
    application of an unknown formulation at a rate of 2.6 kg a.i./ha
    (Müller et al., 1981). A strong inhibition of nitrogenase activity
    was possibly an indirect effect due to a changed C/N ratio after the
    treatment. In this study lasting 28 days, pretreatment measurements
    were used as a control.

         Roundup did not inhibit nitrification in three agricultural
    soils after treatment at a recommended application rate of 2.9 kg
    a.i./ha in 25-day experiments (Stratton, 1990). In a sandy loam,
    nitrification was stimulated after the application of 145 kg
    a.i./ha, whereas nitrification rates were inhibited by 50% at rates
    of 194-435 kg a.i./ha. Mineralization, expressed as the time-course
    of mg N/g dry weight (N = ammonium or nitrate nitrogen), in two
    agricultural sandy loam soils was stimulated significantly due to
    the treatment with 100 mg a.i./kg dry weight during an experiment
    lasting 70 days (Marsh et al., 1977).

         Not only in agricultural soils but also in forest soils, the
    effects of Roundup on the nitrogen cycle appear not to be
    inhibitory. Preston & Trofymow (1989) found no significant effects
    on nitrification or the immobilization of urea nitrogen in an
    organic fir forest floor and its underlying mineral horizon due to
    the application of 10 and 50 mg a.i./kg dry weight. This experiment
    lasted for 40 days.

         In two agricultural sandy loam soils exposed to 100 mg a.i./kg
    dry weight for 210 days, transient slight effects on CO2 evolution
    were observed by Marsh et al. (1977). These effects were both
    stimulatory (first 20 days) and inhibitory (until approximately 130
    days) in both the soil from a permanent grassland and the soil from
    an arable field, but especially in the latter. Carlisle & Trevors
    (1986b) observed a dose-related increase in both O2 consumption
    and CO2 production in an agricultural sandy loam soil during a
    test of 10 days, with both technical grade glyphosate and Roundup.
    In general the increases were significant at concentrations greater
    than or equal to 127 mg a.i./kg. At the lowest dose (13 mg a.i./kg
    soil) no effects were observed except a significant increase in

    oxygen consumption due to Roundup. Oxygen consumption at doses
    greater than or equal to 127 mg a.i./kg was more strongly stimulated
    by Roundup than by technical grade glyphosate, possibly due to the
    isopropylamine or surfactant. Preston & Trofymow (1989) found no
    significant effect on CO2 production in an organic fir forest
    floor and its underlying mineral horizon after the application of 10
    and 50 mg a.i./kg dry weight. This experiment lasted 40 days.

         Aerobic H2 oxidation in an agricultural sandy loam was
    significantly inhibited by both technical grade glyphosate and
    Roundup, but only at the highest dose of 635 mg a.i./kg dry weight
    (Carlisle & Trevors, 1986b). Inhibition was stronger under anaerobic
    conditions, and was found at concentrations of > 127 mg a.i./kg
    dry weight to be significant and dose related. Anaerobic inhibition
    might have been due to increased H2 generation as a result of
    stimulated fermentation.

         No effects of glyphosate on the activities of 1,3-ß-glucanase
    and urease in a silt loam were found after application of Roundup at
    a rate of 12 mg a.i./kg dry weight (Lethbridge et al., 1981).
    Preston & Trofymow (1989) found no significant effect on urea
    hydrolysis in an organic fir forest floor and its underlying mineral
    horizon amended with 200 mg urea nitrogen per kg dry weight, due to
    treatment with Roundup at a rate of 50 mg a.i./kg dry weight.

         The degradation of cellulose, starch, protein, or leaf litter
    was not inhibited at concentrations of 5 and 25 mg a.i./kg soil in
    three agricultural soils, with the exception of litter in silt loam
    at the highest dose (ABC Inc., 1978e). In this experiment lasting 84
    days, technical grade glyphosate was applied.

         Bacterial growth of  Rhizobium trifolii in sterile solutions
    with Bergersen's broth was completely inhibited at solution
    concentrations of 10 and 20 mg a.i./litre, applied as an unknown
    formulation of glyphosate (Eberbach & Douglas, 1983). Only at 10 mg
    a.i./litre did the bacterial growth recover within 4 days. At lower
    concentrations no inhibition was found.

         Mycelial growth of ectomycorrhizal fungi in pure cultures on
    agar was inhibited by 50% or more at concentrations of > 1 mg
    a.i./litre for  Pisolithus tinctorius and of > 100 mg a.i./litre
    for  Cenococcum geophilum and  Hebeloma longicaudum (Estok et al.,
    1989). Ectomycorrhizal fungi commonly associated with pines  (Pinus
    sp) were significantly inhibited at concentrations of > 29 µg
    Roundup/litre (Chakravarty & Chatarpaul, 1990a). The most sensitive
    species were  Cenococcum graniforme, Hebeloma crustuliniforme and
     Laccaria laccata. Roundup increased the susceptibility of sandy
    soil for  Gaeumannomyces gramminis, a fungus causing "take-all
    disease" in wheat crops (Mekwatanakarn & Sivasithamparam, 1987).

    Application at a rate of 0.54 µg a.i./kg increased the survival and
    pathogenicity of the fungus significantly after 140 days of
    incubation at 25 °C. These authors concluded that Roundup affected
    microbial antagonists of the fungus.

    9.1.2  Aquatic organisms

    9.1.2.1  Plants

    Appraisal

          There is conflicting information on the effects of sediment on
     the phytotoxicity of glyphosate to aquatic plants; Lemna showed
     reduced effects whilst Carthamus did not. Generally, glyphosate is
     thought to be largely unavailable to plants when added to soil and
     only effective as a herbicide when applied to foliage.

         The chronic toxicity to aquatic macrophytes when exposed to
    technical grade glyphosate or Roundup dissolved in water is
    summarized in Tables 20 and 21. Glyphosate is slightly toxic with a
    14-day NOEC value of 9 mg/litre. Roundup is also slightly toxic with
    14-day NOEC values of 2.4-56 mg/litre. No data on acute toxicity for
    plants were available.

         When Roundup was sprayed at a rate of 0.8 kg a.i./ha, the
    phytotoxicity to floating plants of the common duckweed  (Lemna
     minor) was dependent on the extent of washed-off deposit (Lockhart
    et al., 1989). Phytotoxicity was highest when the sprayed deposits
    were not washed off within 6 h after application. Suspensions of
    50 mg/litre of inorganic bentonite clay reduced the phytotoxicity of
    Roundup to common duckweed significantly (Hartman & Martin, 1984).
    When exposed for 14 days, concentrations up to 24 mg Roundup/litre
    had no effect on plant growth when sediment was added, whereas the
    growth was reduced by 50% at 5 mg Roundup/litre when no sediment was
    added.

         Phytotoxicity of glyphosate to the safflower  (Carthamus
     tinctorius) was not significantly reduced when the test compound
    was added to drainage water with suspended particles instead of
    distilled water (Bowmer et al., 1986). In these bioassays the
    inhibition of root elongation was measured when the plants were
    exposed to concentrations in unfiltered water of approximately
    0.1-3 mg a.i./litre. The maximum concentration of adsorbed
    glyphosate was 2500 mg/kg.

         Although it had inhibitory effects at high concentrations,
    Roundup had a stimulatory effect on the growth of common duckweed
     (Lemna minor) and tubers of sago pondweed  (Potamogeton
     pectinatus) at lower concentrations (Hartman & Martin, 1985;
    Lockhart et al., 1989). Enhancement of growth of common duckweed and
    sago pondweed was found at 7-56 and 3 mg Roundup/litre,
    respectively. These stimulatory effects may refer to hormesis.

    9.1.2.2  Invertebrates

    Appraisal

          The data from laboratory toxicity tests show that formulations
     are often more toxic than technical glyphosate to aquatic
     invertebrates. The surprising result that addition of clay particles
     to Daphnia test systems increased the toxicity of glyphosate is
     probably due to ingestion of herbicide bounds to the particles. Few
     studies have been conducted in the presence of sediment; the
     reported toxicity of glyphosate is, therefore, difficult to relate
     to the field situation.

         The acute and chronic toxicity of technical grade glyphosate
    and its formulations to aquatic invertebrates are summarized in
    Tables 18-21. Technical grade glyphosate is slightly to very
    slightly toxic, with LC50 values of > 55 mg/litre and a 21-day
    NOEC value of 100 mg/litre. Formulations of glyphosate are
    moderately to very slightly toxic with 2-day EC50 values of
    5.3-5600 mg product/litre and 21-day MATC values of 1.4-4.9 mg
    product/litre. The higher toxicity of Roundup to crustaceans is
    mainly due to the presence of surfactants (Servizi et al., 1987).

         In a laboratory  in vitro test with the gills of  Mytilus
     californianus, a marine water mollusc, the active uptake of glycine
    was inhibited by 23 and 67%, respectively, when a mixture of
    14C-labelled and unlabelled glyphosate was applied at rates of
    0.2-1.7 mg/litre (Swinehart & Cheney, 1987). This inhibition might
    be an indication for Mg2+-moderated binding of glycine to the gill
    surface, whereas glyphosate can compete with glycine uptake by
    forming a metal complex.

         Water fleas  (Daphnia magna) were less sensitive to Roundup,
    when the water was aerated than when it was unaerated. The 2-day
    LC50 value decreased from 37 (with aeration) to 24 mg (without
    aeration) Roundup/litre (EG & G Bionomics, 1980f).

         Addition of 50 mg/litre of inorganic bentonite clay decreased
    the 2-day EC50 value for mature water fleas (Daphnia pulex) from
    16 to 7 mg Roundup/litre (Hartman & Martin, 1984). This higher
    toxicity might have been due to ingestion of particle-adsorbed
    glyphosate. Addition of bentonite also increased the toxicity in a
    chronic experiment with populations of  Daphnia pulex. Immature
    water fleas were more sensitive to glyphosate than the adults under
    all conditions. Populations in all treatments had recovered within
    14 days after the application (Hartman & Martin, 1984).

         No avoidance of glyphosate was found when mayfly nymphs
     (Ephemerella walkeri) were tested at concentrations of 0.2-2 mg
    Roundup/litre, whereas avoidance was demonstrated at 24 mg/litre
    (Folmar et al., 1979).

    9.1.2.3  Vertebrates

    Appraisal

          The toxicity tests for fish are generally performed without
     sediment. As the bioavailability of glyphosate itself will be
     reduced under most conditions due to sorption onto sediment, no
     toxic effects are expected. Toxic effects, however, can be expected
     due to surfactants in some formulations. To a lesser extent,
     life-stage, pH, water hardness, temperature, and the presence of
     feed all influence toxicity. No adverse effects on the
     osmoregulatory mechanism were found.

         The acute and chronic toxicity of technical grade glyphosate
    and its formulations to fish are summarized in Tables 18-21.
    Technical grade glyphosate is moderately toxic with 4-day LC50
    values of 10 to > 1000 mg/litre, a 21-day NOEC value of
    52 mg/litre, and an MATC value of > 26 mg/litre. Formulations of
    glyphosate have comparable toxicity with 4-day LC50 values of 2.4
    to > 1000 mg product/litre, and 21-day NOEC values of 0.8-2.4 mg
    product/litre. Toxicity may vary substantially, depending on the
    species, the test compound and test conditions. In general,
    technical grade glyphosate is less toxic than the formulations. This
    difference is mainly due to a higher toxicity of surfactants in the
    formulations (Folmar et al., 1979; Servizi et al., 1987; Mitchell
    et al., 1987; Wan et al., 1989).


    
    Table 18.  Aquatic organisms: acute toxicity of technical grade glyphosate in a static test system
                                                                                                                                              

    Organism                 A      Test     pH          Hardness     Temperature    Experimental   Parameter   Concentration   Reference
                                    water                (mg CaCO3/   (°C)           duration                   (mg/litre)
                                                         litre)                      (days)
                                                                                                                                              

    Molluscs

     Crassostrea              -      n.w.a    n.r.        n.r.         25             2              EC50        > 10b           Bionomics
     virginica, eggs                                                                                                             (1973a)

    Echinodermata

     Tripneustes              -      n.w.a    7.7-8.2     n.r.         20             4              EC50        > 1000c         E G & Bionomics
     esculentes                                                                                                                  (1978d)

    Crustaceans

     Daphnia magna,           -      w.w.     7.8-8.0     > 250        19             2              LC50        780             ABC Inc.
    first instar                                                                                                                (1978a)
     Uca pugilator            -      a.m.a    n.r.        n.r.         21             4              LC50        934             Bionomics
                                                                                                                                (1973b)
     Palaemonetes             -      a.m.a    n.r.        n.r.         21             4              LC50        281             Bionomics
     vulgaris                                                                                                                    (1973b)
     Mysidopsis bahia         -      n.w.a    6.4-8.3     n.r.         20             4              LC50        > 1000          E G &
                                                                                                                                      Bionomics
                                                                                                                                (1978c)

    Insects

     Chironomus plumosus,     -      r.w.     n.r.        40           22             2              EC50        55d             Folmar et al.
    fourth instar                                                                                                               (1979)
                                                                                                                                              

    Table 18 (contd).
                                                                                                                                              

    Organism                 A      Test     pH        Hardness      Temperature   Experimental   Parameter     Concentration   Reference
                                    water              (mg CaCO3/    (°C)          duration                     (mg/litre)
                                                       litre)                      (days)
                                                                                                                                              

    Fish

     Ictalurus                -      r.w.     n.r.      40            22            4              LC50          130             Folmar et al.
     punctatus                                                                                                                   (1979)
     Salmo gairdnerii,        -      divers   6.3-8.2   5.3-148       n.r.          4              LC50          10-197          Wan et al.
    0.4 cm, 0.5 g                                                                                                               (1989)
     Salmo gairdnerii,        -      r.w.     4.4-7.2   45            12            4              LC50          86              ABC Inc.
    0.4 cm, 0.6 g                                                                                                               (1978b)
     Oncorhynchus keta,       -      divers   6.3-8.2   5.3-148       n.r.          4              LC50          10-148          Wan et al.
    0.4 cm, 0.5 g                                                                                                               (1989)
     O. kisutch,              A      divers   6.3-8.2   5.3-148       n.r.          4              LC50          27-174          Wan et al.
    0.4 cm, 0.5 g                                                                                                               (1989)
     O. tshawytsha,           -      divers   6.3-8.2   5.3-148       n.r.          4              LC50          19-211          Wan et al.
    0.4 cm, 0.5 g                                                                                                               (1989)
     O. gorbusha,             -      divers   6.3-8.2   5.3-148       n.r.          4              LC50          14-190          Wan et al.
    0.4 cm, 0.5 g                                                                                                               (1989)
     Lepomis                  -      r.w.     n.r.      40            22            4              LC50          140             Folmar et al.
     macrochirus                                                                                                                 (1979)
     L. macrochirus,          -      r.w.     6.6-7.0   45            21            4              LC50          120             ABC Inc.
    0.3 cm, 1.0 g                                                                                                               (1978c)
     Pimephales               -      r.w.     n.r.      40            22            4              LC50          97              Folmar et al.
     promelas                                                                                                                    (1979)
     Rasbora                  -      r.w.     n.r.      25            21            4              LC50          168             HRC (1977)
     heteromorpha, 
    0.1-0.3 cm
     Cyprinodon               -      n.w.e    7.6-8.3   n.r.          20            4              LC50          > 1000          E G & Bionomics
     variegatus,                                                                                                                 (1978b)
    0.7-1.0 cm
                                                                                                                                              

    a    salinity 20-35%
    b    based on abnormal development of oyster larvae
    c    based on immobility, drooping spines, and retracted podia
    d    based on immobilisation
    e    salinity 18%
         A = actual concentrations are measured; - = nominal concentration; a.m. = artificial medium;
         r.w. = reconstituted water; w.w. = well water; n.w. = natural surface water;
         n.r. = not reported

    Table 19.  Aquatic organisms: acute toxicity of formulations with glyphosate
                                                                                                                                              

    Organism         A   Test    Compound  Test    pH        Hardness     Temperature   Experimental   Parameter   Concentration   Reference
                         type              water             (mg CaCO3/   (°C)          duration                   (mg/litre)a
                                                             litre)                     (days)
                                                                                                                                              

    Crustaceans

     Daphnia          -   S       Ru        r.w.   8.2-8.3       175          22         2              EC50          5.3b          E G & G
     magna, first                                                                                                                   Bionomics
    instar                                                                                                                         (1980e)
     D. magna,        -   S       Ru        r.w.   7.7-8.1       175          22         2              EC50         24-37b         E G & G
    first instar                                                                                                                   Bionomics
                                                                                                                                   (1980f)
     D. magna,        -   S       Ru        r.w.    n.r.         40           22         2              EC50          7.3b          Folmar
    first instar                                                                                                                   et al.
                                                                                                                                   (1979)
     D. magna,        -   S       St        w.w.   8.3-8.5     225-275        20         2              EC50           42b          ABC Inc
    first instar                                                                                                                   (1984c)
     D. magna,        -   S       RuD       w.w.   7.9-8.6       255          20         2              EC50          930b          ABC Inc
    first instar                                                                                                                   (1981a)
     D. pulex,        -   S       Ru        w.w.     7.6         282          15         2              EC50           19b          Hartman &
    mature                                                                                                                         Martin
                                                                                                                                   (1984)
     Gammarus         A   CF      Ru        w.w    7.9-8.3       255          17         2              EC50          42b,c         ABC Inc
     pseudolimnaeus                                                                                                                 (1982b)

    Insects

     Chironomus       -   S       Rod       r.w.   7.6-7.8      42-44         22         2              EC50          5600b         Buhl &
     riparius,                                                                                                                      Faerber
    fourth instar                                                                                                                  (1989)
                                                                                                                                              

    Table 19 (cont'd)(2)
                                                                                                                                              

    Organism         A   Test    Compound  Test    pH        Hardness     Temperature   Experimental   Parameter   Concentration   Reference
                         type              water             (mg CaCO3/   (°C)          duration                   (mg/litre)a
                                                             litre)                     (days)
                                                                                                                                              

     Chironomus       -   S       Ru        r.w.    n.r.         40           22         2              EC50           44b          Folmar 
     plumosus,                                                                                                                      et al.
    fourth instar                                                                                                                  (1979)

    Fish

     Ictalurus        -   S       Ru        r.w.    6.3-7.2    24-40         22          4              LC50           52           E G & G
     punctatus,                                                                                                                     Bionomics
    0.7 cm, 3 g                                                                                                                    (1980a)
     Ictalurus        -   S       Ru        r.w.     n.r.       40           22          4              LC50           32           Folmar
     punctatus                                                                                                                      et al.
                                                                                                                                   (1979)
     Salmo            -   S       St        r.w.    6.8-7.3    40-45         12          4              LC50           7.5          ABC Inc
     gairdnerii,                                                                                                                    (1984a)
    0.4 cm, 0.7 g
     Salmo            A   CF      Ru        w.w.    8.0-8.2     255          12          4              LC50          8.2c          ABC Inc
     gairdnerii,                                                                                                                    (1982c)
    0.5 cm, 2.4 g
     Salmo            -   S       Ru        divers  6.3-8.2   5.3-148       n.r.         4              LC50          14-33         Wan et al.
     gairdnerii,                                                                                                                    (1989)
    0.4 cm, 0.5 g
     Salmo            -   S       Ru        r.w.    6.6-7.6     40           12          4              LC50           36           E G & G
     gairdnerii,                                                                                                                    Bionomics
    0.3 cm, 0.3 g                                                                                                                  (1980c)
     Salmo            -   S       Ru        w.w     6.4-7.3    26-26         12          4              LC50           22           E G & G
     gairdnerii,                                                                                                                    Bionomics
    0.4 cm, 0.7 g                                                                                                                  (1980g)
                                                                                                                                              

    Table 19 (cont'd)(3)
                                                                                                                                              

    Organism         A   Test    Compound  Test    pH        Hardness     Temperature   Experimental   Parameter   Concentration   Reference
                         type              water             (mg CaCO3/   (°C)          duration                   (mg/litre)a
                                                             litre)                     (days)
                                                                                                                                              

     S. gairdnerii,   A   S       Ru        r.w     7.4-7.7     85           11          4              LC50           22           EVS
    0.4 g                                                                                                                          Consultants
                                                                                                                                   (1986a)
     S. gairdnerii,   A   S       Ru        n.w     7.4-7.8     81           11          4              LC50           15           EVS
    0.4 g                                                                                                                          Consultants
                                                                                                                                   (1986a)

     S. gairdnerii,   A   S       Ru        d.w    5.4-6.3      4.5           11         4              LC50           26           EVS
    0.4 g                                                                                                                          Consultants
                                                                                                                                   (1986a)

     S. gairdnerii,   -   S       Ru        r.w    approx.      40            12         4              LC50           3.2          Folmar et al.
    1.0 g                                           7.2                                                                            (1979)
     Salmo            -   S       RuD       w.w    4.6-7.1      45            12         4              LC50         > 1000         ABC Inc
     gairdnerii,                                                                                                                    (1981c)
    0.3 cm, 0.2 g
     S. gairdnerii,   -   S       Vis       n.r.     6.0        9.6          12.3        4              LC50           34           Morgan &
    9.5 cm                                                                                                                         Kiceniuk
                                                                                                                                   (1992)
     Lepomis          -   S       St        r.w.   6.8-7.5     40-45          22         4              LC50           4.5          ABC Inc
     macrochirus,                                                                                                                   (1984b)
    0.2 cm, 0.1 g
     Lepomis          -   S       RuD       w.w.   4.9-7.1      45            22         4              LC50         > 1000         ABC Inc
     macrochirus,                                                                                                                   (1981b)
    0.2 cm, 0.1 g
     Lepomis          A   CF      Ru        w.w.   8.0-8.2      255           22         4              LC50          5.8c          ABC Inc
     macrochirus,                                                                                                                   (1982a)
    0.2 cm, 0.2 g
                                                                                                                                              

    Table 19 (cont'd)(4)
                                                                                                                                              

    Organism         A   Test    Compound  Test    pH        Hardness     Temperature   Experimental   Parameter   Concentration   Reference
                         type              water             (mg CaCO3/   (°C)          duration                   (mg/litre)a
                                                             litre)                     (days)
                                                                                                                                              

     Lepomis          -   S       Ru        r.w.   6.4-7.5      40            22         4              LC50           46           E G & G
     macrochirus,                                                                                                                   Bionomics
    0.4 cm, 0.3 g                                                                                                                  (1980b)
     Pimephales       -   S       Ru        w.w.   6.7-7.7     39-44          22         4              LC50           31           E G & G
     promelas,                                                                                                                      Bionomics
    0.4 cm, 0.6 g                                                                                                                  (1980d)
     Pimephales       -   S       Ru        r.w.    n.r.        40            22         4              LC50           5.6          Folmar
     promelas                                                                                                                       et al.
                                                                                                                                   (1979)

     Cyprinus         A   S       St        w.w.    7.2-7.9     40-48        22-23       4               LC50          2.4          ABC Inc
     carpio,                                                                                                                        (1990)
    0.6 cm, 2.8 g

     Oncorhynchus     A   S       Ru        divers  6.3-8.2    5.3-148       n.r.        4              LC50          13-33         Wan et al.
     kisutch,                                                                                                                       (1989)
    0.4 cm, 0.5 g
     O. kisutch,      A   S       Ru        d.w.    5.5-6.4      4.5          11         4              LC50           22           EVS
    11.8 g                                                                                                                         Consultants
                                                                                                                                   (1986c)
     O. keta,         -   S       Ru        divers  6.3-8.2    5.3-148       n.r.        4              LC50          11-20         Wan et al.
    0.4 cm, 0.5 g                                                                                                                  (1989)
     O. tshawytsha,   -   S       Ru        divers  6.3-8.2    5.3-148       n.r.        4              LC50          17-33         Wan et al.
    0.4 cm, 0.5 g                                                                                                                  (1989)
     O. tshawytsha,   A   S       Ru        d.w.    5.8-6.7      4.5          11         4              LC50           20           EVS
    4.6 g                                                                                                                          Consultants
                                                                                                                                   (1986b)
                                                                                                                                              

    Table 19 (cont'd)(5)
                                                                                                                                              

    Organism         A   Test    Compound  Test    pH        Hardness     Temperature   Experimental   Parameter   Concentration   Reference
                         type              water             (mg CaCO3/   (°C)          duration                   (mg/litre)a
                                                             litre)                     (days)
                                                                                                                                              

     O. gorbusha,     -   S       Ru        divers  6.3-8.2    5.3-148       n.r.        4              LC50          14-33         Wan et al.
    0.4 cm, 0.5 g                                                                                                                  (1989)
     Oncorhynchus     A   S       Ru        n.w     7.7-8.0      84           4-5        4              LC50          27-29         Servizi
     nerka,                                                                                                                         et al.
    3-6.5 cm, 0.2-3.8 g                                                                                                            (1987)
                                                                                                                                              

    a    All concentrations are expressed as mg of the formulation per litre
    b    Based on immobilisation
    c    Based on actual concentrations
         A = actual concentrations are measured; - = nominal concentrations; CF = continous flow system;
         S = static system; Tgg = technical grade glyphosate; Ru = Roundup; Rod = Rodeo; St = Sting;
         RuD = Roundup D-Pak; d.w. = dechlorinated tap water; r.w. = reconstituted water;
         w.w. = well water; n.w. = natural surface water; a.m. = artificial medium

    Table 20.  Aquatic organisms: chronic toxicity of glyphosate (NOEC/MATC)
                                                                                                                                              

    Organism        A   Test  Test       Test    pH        Hardness    Temperature   Experimental   Parameter    Concentration   Reference
                        type  substance  water             (mg CaCO3/  (°C)          duration                    (mg/litre)
                                                           litre)                    (days)
                                                                                                                                              

    Macrophytes

     Lemna gibba     A   S     Tgg        a.m.      7.5     285            25            14          NOEC         9a,b            Malcolm
                                                                                                                                 Pirnie
                                                                                                                                 Inc.
                                                                                                                                 (1987e)
    Crustaceans

     Daphnia magna,  A   SS    Tgg        w.w.c   6.8-8.2   160-180        20            21          NOEC         100a,d          ABC Inc.
    first instar                                                                                                                 (1989c)

    Fish

     Salmo           A   CF    Tgg        w.w.    5.9-7.8   40-48         14-15          21          NOEC         52a,e           ABC Inc.
     gairdnerii,                                                                                                                  (1989e)
    0.5 cm, 1.3 g
     Pimephales      A   CF    Tgg        w.w.    6.5-7.6   32-42          25           255          MATC         > 26f           E G & G
     promelas,                                                                                                                    Bionomics
    1.5 g                                                                                                                        (1975)
                                                                                                                                              

    a    Based on actual concentrations          d    Based on survival and reproduction
    b    Based on biomass decrease               e    Based on survival, behaviour, and coloration
    c    Mixed with natural surface water        f    Based on survival, growth and reproduction
         A = actual concentrations are measured; CF = continous flow system; SS = semi-static system;
         S = static system; Tgg = technical grade glyphosate; w.w. = well water; a.m. = artificial medium

    Table 21.  Aquatic organisms: chronic toxicity of formulations with glyphosate
                                                                                                                                              

    Organism       A   Test   Test        Test    pH         Hardness     Temperature  Experimental   Parameter   Concentration   Reference
                       type   substance   water              (mg CaCO3/   (°C)                        duration    (mg/litre)f
                                                             litre)                                   (days)
                                                                                                                                              

    Macrophytes

     Lemna minor    -   S      Ru          a.m.      n.r.       n.r.          25            14         NOEC        56c,d           Lockhart
                                                                                                                                  et al.
                                                                                                                                  (1989)
     Lemna minor    -   S      Ru          a.m.      n.r.       n.r.          22            14         NOEC        2.4c,e          Hartman &
                                                                                                                                  Martin
                                                                                                                                  (1984)
     Potamogeton    -   S      Ru          a.m.      n.r.       n.r.          22            14         NOEC        33c,d           Hartman &
     pectinatus                                                                                                                    Martin
    (tubers)                                                                                                                      (1985)

    Crustaceans

     Daphnia        A   SS     St          w.w.b    7.2-8.2      174          20            21         MATC        1.4g,i          ABC Inc.
     magna,                                                                                                                        (1989a)
    first instar
     Daphnia magna, A   SS     Ru          w.w.b    7.6-8.3    160-180        20            21         MATC        4.9g,j          ABC Inc.
    first instar                                                                                                                  (1989b)

    Fish

     Salmo          A   CF     St          w.w.     7.3-7.8     40-48        14-16          21         NOEC        0.8a,h          ABC Inc.
     gairdnerii,                                                                                                                   (1989h)
    0.5 cm,
    1.3 g
                                                                                                                                              

    Table 21. (cont'd)
                                                                                                                                              

    Organism       A   Test   Test        Test    pH         Hardness     Temperature  Experimental   Parameter   Concentration   Reference
                       type   substance   water              (mg CaCO3/   (°C)                        duration    (mg/litre)f
                                                             litre)                                   (days)
                                                                                                                                              

     Salmo          A   CF     Ru          w.w.     7.1-7.8     24-48        14-16          21         NOEC        2.4a,h          ABC Inc.
     gairdnerii,                                                                                                                   (1989d)
    0.5 cm,
    1.8 g
     Salmo          A   CF     Vis         n.r.       6.0        9.6         12.3       approx. 60     NOEC        > 0.04a,k       Morgan &
     gairdnerii,                                                                                                                   Kiceniuk
    9.5 cm, 7.2 g                                                                                                                 (1992)
                                                                                                                                              

    a    Based on actual concentrations               g    Based on survival, reproduction and length of time to the first brood
    b    Mixed with natural surface water             h    Based on survival, behaviour and coloration
    c    Roundup dissolved in test water              i    NOEC is 1.0 mg Sting/litre
    d    Based on biomass decrease                    j    NOEC is 3.2 mg Roundup/litre (actual concentration)
    e    Based on reduction of frond formation        k    Based on mortality and growth
    f    All concentrations are expressed as mg of the formulation per litre

         A = actual concentrations are measured; - = nominal concentrations; CF = continous flow system;
         SS = semi-static system; Ru = Roundup; St = Sting; Vis = Vision; w.w. = well water;
         a.m. = artificial medium; n.r. = not reported.
    

         In laboratory experiments the major factors influencing the
    toxicity appear to be the tested species and its age, the presence
    of surfactants, the hardness, pH, temperature and the availability
    of ration. Wan et al. (1989) found that the toxicity of technical
    grade glyphosate to salmonids increased when hardness and pH
    decreased, whereas for Roundup and Accord CR the contrary was true,
    due to the presence of a 75% tallow amine surfactant in the
    formulations. This surfactant was most toxic in hard water with a
    relatively high pH. A higher toxicity of Roundup to salmonids at
    increasing hardness and pH was confirmed by Mitchell et al. (1987),
    but only partially by Servizi et al. (1987). It was also confirmed
    by Folmar et al. (1979), although they only investigated the effect
    of pH in reconstituted water with a moderate hardness. These authors
    demonstrated that the effects of a pH increase on the toxicity of
    Roundup and technical grade glyphosate were not only seen in rainbow
    trout  (Salmo gairdnerii) but also in bluegill sunfish  (Lepomis
     macrochirus). For both species Roundup became more toxic as the pH
    increased from 6.5 to 7.5, whereas technical grade glyphosate became
    less toxic as the pH increased from 6.5 to 9.5. At pH values higher
    than 7.5, the toxicity of Roundup remained constant. In the
    investigations of Servizi et al. (1987), an antagonistic effect of
    glyphosate on the toxic action of a surfactant was found.

         Increased toxicity of formulations due to surfactants was not
    only demonstrated for the tallow amine surfactant in Roundup but
    also for ortho X-77 in Rodeo (Mitchell et al., 1987). However, even
    in the presence of a surfactant, the acute toxicity of some
    formulations may be very slight, as was found by ABC Inc. (1980a,b).
    In these studies, 0.5% (v/v) of the surfactant X-77 was added to
    Roundup D-Pak, resulting in a 4-day LC50 value of this mixture for
    rainbow trout  (Salmo gairdnerii) and for bluegill sunfish
     (Lepomis macrochirus) of 240 and 830 mg/litre, respectively.

         Folmar et al. (1979) performed acute toxicity tests with
    various species in reconstituted water with a pH of 7.2 and a
    hardness of 40 mg CaCO3/litre. In these tests it was demonstrated
    for rainbow trout  (Salmo gairdnerii) and channel catfish
     (Ictalurus punctatus) that the sensitivity to Roundup increased in
    the following order: eyed eggs, 2-g fingerlings, sac fry, swim-up
    fry, and 1-g fingerlings. The 4-day LC50 values for the various
    life-stages of rainbow trout decreased from 39 mg Roundup/litre for
    eyed eggs to 3.2 mg/litre for small fingerlings. In an additional
    test, a 4-h exposure to concentrations of > 12 mg Roundup/litre
    affected the survival of sac fry and swim-up fry significantly.

         Holdway & Dixon (1988) also demonstrated that toxicity is
    dependent on the life-stage by applying technical grade glyphosate
    to larvae of flagfish  (Jordanella floridae). At concentrations up
    to 30 mg a.i./litre, no 2- or 4-day-old larvae died, whereas 20% of
    the 8-day-old larvae died at the top dose. The effect was even more

    drastic when the larvae were not fed. This treatment killed 20% of
    the oldest larvae even at 3 mg a.i./litre. According to the authors,
    the effect of age might fit the idea of saltatory ontogeny, implying
    critical periods for organs and tissues.

         While investigating the effect of temperature on toxicity,
    Folmar et al. (1979) demonstrated for rainbow trout  (Salmo
     gairdnerii) and bluegill sunfish  (Lepomis macrochirus) that
    toxicity increased with increasing temperatures. For the trout the
    4-day LC50 values decreased from 34 mg Roundup/litre at 7 °C to
    18 mg/litre at 17 °C. For the bluegill sunfish the 4-day LC50
    values decreased from 18 mg/litre at 17 °C to 9.8 mg/litre at 27 °C.

         Rainbow trout  (Salmo gairdnerii) showed the same sensitivity
    to Roundup, independent of whether the water was aerated or not
    (EG & G Bionomics, 1980g). In this experiment the 4-day LC50 under
    both conditions was 22 mg Roundup/litre.

         The potential of coho salmon smolt  (Oncorhynchus kisutch) to
    adapt to changes in water salinity encountered during migration was
    not influenced by the application of Roundup at actual
    concentrations up to 2.8 mg/litre (EVS Consultants, 1986d). The
    osmoregulatory mechanism, which is fully functional in smolts, was
    unaffected as indicated by plasma Na+ concentrations, haematocrit
    values and the condition of the fish. During the experiment in which
    the fish were exposed for 10 days in fresh water and subsequently
    allowed to recover in fresh or sea water, no abnormal behaviour was
    observed.

         No avoidance of glyphosate was found when rainbow trout were
    tested at concentrations up to 24 mg Roundup/litre (Folmar et al.,
    1979). Sublethal concentrations in acute toxicity tests with Roundup
    may cause loss of motility, complete loss of equilibrium, darkened
    pigmentation, or rapid respiration (EG & G Bionomics, 1980a,b,g; EVS
    Consultants, 1986a,b,c).

         Rainbow trout  (Salmo gairdnerii) were exposed to glyphosate
    (as Vision) at 0, 6.25, 25 and 100 µg Vision/litre in a continuous
    flow system. There were no effects on growth or foraging behaviour,
    and no histopathological liver effects. Two out of three types of
    aggressive behaviour were also unaffected; the third, a warning
    "wig-wag" increased in frequency at the top-dose 1 month after
    treatment. After two months the frequency was equal to that of the
    control (Morgan & Kiceniuk, 1992).

    9.1.3  Terrestrial organisms

    9.1.3.1  Plants

         Nodulation of sub-clover  (Trifolium subterraneum) was
    inhibited by an unknown formulation with glyphosate in a
    dose-related way at concentrations of 2-20 mg a.i./litre (Eberbach &
    Douglas, 1989). In this experiment, 3-day-old seedlings were
    inoculated with  Rhizobium trifolii. The seedlings were cultured
    for 56 days in soil-free systems with nutrient solutions. In an
    additional experiment in which Rhizobium was exposed to the same
    concentrations, repeated washing of the inoculi prior to nodule
    initiation did not reduce the inhibition of the nodulation of the
    sub-clover after inoculation. This indicated that the effect on
    nodulation might be the result of damage to the bacteria rather than
    to carry-over of glyphosate from the bacteria to the plant.

         Seed germination of various forest species was not affected by
    treatment with Roundup at concentrations up to 305 mg a.i. (free
    acid)/kg dry weight. Seed germination was affected at the highest
    tested dose of 1525 mg a.i. (free acid)/kg dry weight (Morash &
    Freedman, 1988). The effect on seed germination was confirmed by
    another experiment in which no differences were found between
    sprayed and unsprayed plots with respect to seedling composition and
    quantity. Morash & Freedman (1988) then incubated the soils of
    clear-cuts in a greenhouse. The application of Roundup in the field
    was at a rate of 2.3 kg a.i./ha.

         Red pine seedlings  (Pinus resinosa) were not affected by
    treatment with Roundup, with the exception of a dose-related
    decrease in root length (Chakravarty & Chatarpaul, 1990b).
    Non-affected growth parameters were shoot height, shoot weight, root
    weight, and mycorrhizal development. In this experiment lasting 60
    days, Roundup was applied at rates of 0.54 and 3.2 kg a.i./ha. The
    conifer seedlings were inoculated with an ectomycorrhizal fungus
     (Paxillus involutus), 14 days after germination.

         Glyphosate may affect various pathways of the secondary
    metabolism in the plant, although the actual targets in plants have
    not been located. The synthesis of aromatic amino acids, secondary
    hydroxyphenolic compounds, chlorophyll and delta-amino-levulinic
    acid were reported to be affected by glyphosate (Amrhein et al.,
    1980; Duke & Hoagland, 1981; Kitchen et al., 1981a,b). Aromatic
    amino acids are important for the synthesis of, for instance, some
    alkaloids, the phytohormone indole-3-acetic acid and phenolic
    compounds such as lignin and quinones.

         With respect to the synthesis of aromatic amino acids, there
    are indications that the actual target is not shikimate kinase or
    anthranilate synthase, but probably
    5-enolpyruvylshikimate-3-phosphate synthase or chorismate synthase
    (Amrhein et al., 1980). When applied to hypocotyls of buckwheat
     (Fagopyrum esculentum) and to cultured cells of smooth bedstraw
     (Galium mollugo), technical grade glyphosate inhibited the
    conversion of shikimate to chorismate. This inhibition  in vivo and
     in vitro was found at concentrations of > 10 mg a.i./litre. In
    the cultured cells of Galium, the accumulation of shikimate was
    concomitant with a decrease of anthraquinones (Amrhein et al.,
    1980). Possibly the non-transportable carbon pool, to which carbon
    was found to be diverted in sugar beets  (Beta vulgaris) due to
    glyphosate, was in fact shikimate (Gougler & Geiger, 1984). These
    effects on carbon metabolism were found at application rates
    equivalent to 5 kg a.i./ha leaves. Duke & Hoagland (1981) assumed
    that possibly chelation of divalent ions such as Ca2+ and Mg2+
    that are involved in many metabolic pathways was the main cause of
    damage.

    9.1.3.2  Invertebrates

    Appraisal

          Glyphosate has low toxicity for bees and earthworms.

         Oral 2-day LD50 values of technical grade glyphosate and
    Roundup for bees were 100 µg a.i./bee and > 100 µg Roundup/bee,
    respectively. Contact 2-day LD50 values for these two substances
    were likewise > 100 µg a.i./bee and > 100 µg Roundup/bee (HRC,
    1972). The oral 2-day LD50 of Sting was also > 100 µg Sting/bee
    (Altmann, 1984). In these experiments  Apis mellifera was tested. The
    LD50 values indicate a slightly acute toxicity of technical grade
    glyphosate, Roundup, and Sting to honey-bees. Roundup was also
    slightly toxic to green lacewings  (Chrysoperla carnea) when they
    were exposed by contact to 1 kg Roundup/ha (SFRSA, 1990). In this
    experiment the average number of eggs per female and the larval and
    pupal mortality were increased due to the treatment, resulting in an
    overall reduction in beneficial capacity of 41%. The beneficial
    capacity is a function of the larval and pupal mortality, and the
    average number of eggs per treated and untreated female. No effects
    on the food uptake and mortality of the beetle  Poecilus cupreus
    Bonelli were observed 15 days after application of 6 kg Sting/ha
    (IFU, 1990).

         When exposed to artificial soil contaminated with Roundup
    D-pak, earthworms  (Eisenia fetida) were soft and/or slack, in a
    dose-related way, at concentrations > 500 mg Roundup D-pak/kg dry
    weight (IBR, 1991a). No other adverse effects were found, indicating

    a 14-day NOEC of 158 mg/kg. A comparable experiment with Roundup
    also indicated slight toxicity for earthworms with a 14-day NOEC of
    500 mg Roundup/kg dry weight (IBR, 1991b). At higher concentrations
    thin, slack and lethargic worms with a dark skin were observed.

    9.1.3.3  Vertebrates

    Appraisal

          Glyphosate has low toxicity to birds after acute oral or
     short-term dietary exposure. Mammals tested showed effects (body
     weight loss) only after high levels of dosing. Herbicide-treated
     foliage was not avoided by deer in the single study reported.

         The acute, subacute and chronic toxicity of glyphosate and its
    formulations to birds is summarized in Table 22.

         Male marsupials  (Sminthopsis macroura) showed significant
    body weight loss after exposure to feed contaminated with
    concentrations of up to 5000 mg a.i/kg feed (Evans & Batty, 1986).
    No other treatment-related effects were found in the male
    marsupials. In female hopping-mice  (Notomys alexis and Notomys
     mitchelli) fed similar doses, no treatment-related effects were
    found.

         Glyphosate did not affect the daily chow consumption of
    black-tailed deer  (Odocoileus hemionus columbianus) when these
    herbivores were exposed to browse treated with glyphosate at a rate
    of 2.2 kg/ha (Sullivan & Sullivan, 1979). The deer did not avoid the
    contaminated browse, and sometimes even preferred it. Irrespective
    of whether they were given treated alfalfa  (Medicago sativa),
    treated alder  (Alnus rubra) or untreated feed, the deer had the
    same daily chow consumption.

    9.2  Field observations

    9.2.1  Microorganisms

    Appraisal

          Some effects on microorganisms have been reported in field
     studies following application of glyphosate formulations. However,
     these have been minor and reversible in most cases. It is not
     possible to separate the direct toxic effects of the herbicide from
     changes in the habitat caused by its intended herbicidal action.


    
    Table 22.  Birds: acute and chronic toxicity of glyphosate and its formulations
                                                                                                                                              
    Species                  Compound    Sex     Age        Route   Experimental   Parameter   Concentration          Reference
                             duration
                             (days)
                                                                                                                                              

     Colinus virginianus      Tgg         n.r.    n.r.       diet          8        LC50        > 4640 mg/kg feed      Hazleton Lab. Inc.
                                                                                                                      (1973a)
     Colinus virginianus      Tgg         n.r.    14 days    oral                   LD50        > 3851 mg/kg b.w.      Wildlife Int Ltd.
                                                                                                                      (1978c)
     Colinus virginianus      Tgg         M,F     5 months   diet        119        NOEC        > 1000 mg/kg feeda     Wildlife Int Ltd.
                                                                                                                      (1978b)
     Colinus virginianus      Ru          n.r.    10 days    diet          8        LC50        > 5620 mg/kg feedb     Wildlife Int Ltd.
                                                                                                                      (1990b)
     Anas platyrhynchos       Tgg         n.r.    n.r.       diet          8        LC50        > 4640 mg/kg feed      Hazleton Lab. Inc.
                                                                                                                      (1973b)
     Anas platyrhynchos       Tgg         M,F     6 months   diet        112        NOEC        > 1000 mg/kg feeda     Wildlife Int Ltd.
                                                                                                                      (1978a)
     Anas platyrhynchos       Ru          n.r.    10 days    diet          8        LC50        > 5620 mg/kg feedb     Wildlife Int Ltd.
                                                                                                                      (1990a)
     Poephilla guttata        Ru          M       mature     diet          7        LC50        < 16 393 mg/kg feedb   Evans & Batty (1986)

     Poephilla guttata        Ru          M       mature     diet          5        LC50        > 8197 mg/kg feedb     Evans & Batty (1986)
                                                                                                                                              

    a    Based on reproduction impairment of one generation
    b    Concentrations expressed as mg of the formulation per kg body weight or feed

         Tgg = technical grade glyphosate; Ru = Roundup; M = males; F = females; n.r. = not reported.
    

    9.2.1.1  Water

         In a pool located in Hong Kong, treatment with 656 mg
    Roundup/litre caused a substantial decrease in bacteria within 14
    days after treatment. The number of colony-forming units had
    returned to the control level 30 days after treatment (Chan & Leung,
    1986).

         Diatom populations in the water and sediments of a pond and a
    stream showed a significantly different density of some species when
    aerially treated with Roundup at a rate of 2.2 kg a.i./ha (Sullivan
    et al., 1981). The authors concluded that these differences were
    probably due to seasonal and habitat variation, rather than to
    treatment with the herbicide.

         Roundup may have affected the increase in ash-free dry weight
    and the chlorophyll a standing crop of periphyton in the first month
    after spraying with Roundup at a rate of 2.2 kg a.i./ha (Holtby &
    Baillie, 1989a). In this experiment lasting around 130 days, some
    tributaries in a watershed in British Columbia (Canada) were
    directly oversprayed.

    9.2.1.2  Soil

         When Roundup was applied to a sandy loam being prepared for
    conifer forestation, no significant changes in the soil respiration
    were found up to about 180 days after application of Roundup at a
    rate of 0.54 kg a.i./ha (Chakravarty & Chatarpaul, 1990a).
    Concomitantly the numbers of fungi and bacteria decreased
    significantly during the first 2 months, but after about 180 days
    the numbers had recovered. These results might indicate changes in
    microbial populations due to application of Roundup at recommended
    application rates.

         Preston & Trofymow (1989) found no significant effects on the
    number of bacteria, actinomycetes, and nitrogen fixers in
    ferro-humic podsols covered with alder trees  (Alnus rubra) in the
    Carnation Creek watershed, Canada, due to treatment with Roundup.
    The only consistent effect was a significant reduction of the number
    of fungi at one of the two treated sites. In this site the fungi
    appeared to have recovered at the end of the study. In this
    experiment of about 180 days, Roundup was hand-sprayed at a rate of
    2 kg a.i./ha. In an additional comparable intensive field trial of
    one month, microflora populations appeared to have recovered from
    treatment after one month, with the exception of the reduction of
    fungi in the litter of one of the treated sites. Actinomycetes and
    nitrogen fixers in the litter appeared to be reduced in numbers due
    to the treatment but they subsequently recovered. This reduction was
    not found in the underlying humus layer.

         Stratton & Stewart (1992) studied microbial activity in forest
    soil and litter following the application of Roundup at 4.7
    litres/ha (equivalent to 1.7 kg a.i./ha) to a coniferous forest
    previously clear-cut and replanted. Treated and untreated (covered
    with plastic sheeting during application) areas of forest were used
    to obtain soil and litter samples which were tested in situ over an
    8 months period following spraying. Glyphosate had a generally
    stimulatory effect on microbial biomass in litter (up to 80%
    increase) but no significant effect in soil. There were no
    significant effects on the numbers of bacteria, fungi or
    actinomycetes in either soil or litter. Glyphosate generally
    stimulated respiration in both soil and litter; the degree of
    stimulation was very variable throughout the sampling period,
    ranging from a few percent to 100% increases in CO2 evolution.

         No substance- or dose-related effects on aerobic bacteria were
    observed in a sandy soil in a semi-arid region of Argentina up to 96
    days after the application of Roundup (Gómez & Sagardoy, 1985).
    Doses of up to 2.8 kg a.i. (free acid)/ha were applied.

         No substance-related effects on the growth of an
    ectomycorrhizal fungus were found when Roundup was applied at
    concentrations of up to 3.2 kg a.i./ha (Chakravarty & Chatarpaul,
    1990b). In these experiments red pines  (Pinus resinosa) were
    inoculated with the fungus  Paxillus involutus. In the field 74-86%
    of the seedlings that were not inoculated were colonized by
    indigenous mycorrhizal fungi within two months.

    9.2.2  Aquatic organisms

    Appraisal

          Little effect has been reported on aquatic invertebrates or
     fish exposed to glyphosate formulations sprayed in the field. Minor
     mortality in a single study of young trout may reflect the greater
     sensitivity of early life-stages.

    9.2.2.1  Plants

         No field data on toxicity to aquatic macrophytes are available.

    9.2.2.2  Invertebrates

         An increased drift of midge larvae  (Chironomus plumosus) was
    found in artificial outdoor streams treated with 4.9 mg
    Roundup/litre (Folmar et al., 1979). No increased drift was found at
    0.5 mg Roundup/litre. In streams in a coastal rainforest in British

    Columbia, Canada, only the drift densities of an amphipod  (Gammarus
    sp) and mayflies  (Paraleptophlebia sp) increased after treatment
    with Roundup at a rate of 2 kg a.i./ha (Kreutzweiser et al., 1989).
    Density peaks partly coincided with periods immediately following
    rainfall, which might indicate an effect due to glyphosate run-off,
    or an effect due to increased discharges.

         During most streamflows, the abundance of benthic
    macro-invertebrates in a stream and at sites in tributary swamps was
    similar at untreated sites and at sites that had been treated with
    Roundup at a rate of 2.2 kg a.i./ha (Scrivener & Carruthers, 1989).
    However, after periods of frequent rainstorms leading to flooding,
    the abundance at the treated sites was 40-50% lower.

         Water-fleas  (Daphnia magna) did not show any mortality in a
    pond sprayed with Roundup at application rates of up to 220 kg
    a.i./ha (Hildebrand et al., 1980). In this experiment the
    water-fleas were exposed for 8 days in pens that were partly
    immersed in the water.

    9.2.2.3  Vertebrates

         Fingerlings (2.1 g, 5.8 cm) of rainbow trout  (Salmo
     gairdnerii) that were exposed for 14 days to Roundup in
    flow-through pens in shallow streams did not show any mortality or
    substance-related effects at application rates of up to 220 kg a.i.
    (free acid)/ha (Hildebrand et al., 1982). In this experiment Roundup
    was sprayed manually on moderately flowing forest streams in British
    Columbia, Canada. In the same area an aerial application of 2.2 kg
    a.i. (free acid)/ha did not cause mortality or obvious signs of
    stress in rainbow trout fingerlings in flow-through pens, which were
    also exposed to Roundup for 14 days (Hildebrand et al., 1982). A
    direct aerial application of 2.1 kg a.i./ha on a tributary of the
    Carnation Creek watershed (British Columbia, Canada), however,
    killed 2.6% of the 120 fingerlings of coho salmon  (Oncorhynchus
     kisutch) within 24 h after application, whereas no mortality
    occurred at the unexposed sites (Holtby & Baillie, 1989b). Also some
    stress of the caged fingerlings was indicated within the first 2 h
    after application. Up to 2 years after application no consistent
    effects of Roundup on over-winter mortality, probability of entering
    and leaving the tributary, timing of spring emigration, and growth
    rates were found.

         In artificial outdoor streams in Colorado, USA, rainbow trout
    (Salmo gairdnerii) were exposed to concentrations of up to 5 mg
    Roundup/litre for 12 h. No adverse effects on fecundity and
    gonadosomatic indices were found in this study by Folmar et al.
    (1979).

         Rainbow trout  (Salmo gairdnerii) in pens that were immersed
    in stream water avoided Roundup at concentrations of > 40 mg
    Roundup/litre (Hildebrand et al., 1982).

    9.2.3  Terrestrial organisms

    Appraisal

          Spray drift of herbicides will affect non-target plants.
     Adequate buffer zones have been defined for some application
     methods.

          Changes in species diversity and population size and structure
     have been reported for terrestrial invertebrates and vertebrates
     following applications of glyphosate formulation in the field.
     Modifications in available food plants, insect populations
     associated with vegetation killed by the herbicide, and ground cover
     following intended effects of the spray probably account for these
     changes.

    9.2.3.1  Plants

         Red pine seedlings  (Pinus resinosa) were not affected by
    treatment with Roundup in a field study. There was no dose-related
    decrease of the root length, as had been observed in a comparable
    laboratory experiment with the same doses (Chakravarty & Chatarpaul,
    1990b). In this experiment lasting 154 days Roundup was applied at
    rates of up to 3.2 kg a.i./ha. The conifer seedlings were inoculated
    with an ectomycorrhizal fungus  (Paxillus involutus) at 14 days
    after germination, and they were planted outdoor after being
    cultured in a greenhouse for up to 70 days after germination.

         No plants were found as indicator species for damage due to
    drift of a formulation of glyphosate when this was applied with
    hydraulic ground sprayers (Marrs et al., 1989). In this study,
    native British species commonly found in nature reserves were
    exposed to spray drift, at several distances downwind from a zone
    sprayed with 0.5 and 2.2 kg a.i./ha. The effect of windspeed was
    investigated by spraying at speeds of 2.5 and 3.5 m/second. Death
    and severe growth suppression occurred at a distance of 2-6 m from
    the sprayer. Sublethal damage also occurred, mostly near to the
    sprayer, although for  Prunella vulgaris damage occurred up to 20 m
    away. Epinasty was the most frequent symptom of damage. Most of the
    damaged plants recovered, however. Some of the species were
    consistently more sensitive, i.e.  Digitalis purpurea, Centaurea
     nigra, Prunella vulgaris and Lychnis flos-cuculi. Marrs et al.
    (1989) concluded that, when spraying with ground sprayers, buffer
    zones around nature reserves should be 5-10 m.

    9.2.3.2  Invertebrates

         No significant and consistent effects on the number of
    nematodes and springtails were found in the upper 3 cm of
    ferro-humic podsols due to treatment with Roundup (Preston &
    Trofymow, 1989). The soils were covered with alder trees  (Alnus
     rubra) and located in British Columbia, Canada. The only
    consistent effect was a significant reduction in the number of both
    oribatid and non-oribatid mites on one of the treated sites around
    20 days after application. On this site the number of mites appeared
    to have returned to normal by the end of the study. In this
    experiment of around 180 days Roundup was hand-sprayed at a rate of
    2 kg a.i./ha.

         No substance- or dose-related effects on mites and springtails
    were observed in a sandy soil in a semi-arid region of Argentina up
    to 96 days after application of Roundup (Gómez & Sagardoy, 1985).
    Applied doses were up to 2.8 kg a.i. (free acid)/ha.

         The numbers of herbivorous insects and ground invertebrates
    were significantly reduced up to 3 years after treatment with
    Roundup in a 4- to 5-year-old clear-cut planted with spruce
    seedlings  (Picea sp) (Santillo et al., 1989b). No effects were
    found on predatory insects. The clear-cut was located in Maine, USA,
    and sprayed with 1.7 kg a.i./ha. During this experiment lasting 3
    years, the vegetation did not recover completely, and, apparently,
    the effects on invertebrates were mainly due to habitat change.
    Unintentionally untreated areas in the sprayed site showed a much
    lesser reduction of invertebrates. These areas may therefore be
    considered as potential sources for recolonization.

    9.2.3.3  Vertebrates

         Treatment of 4- to 5-year old clear-cuts in Maine, USA, planted
    with seedlings of spruce  (Picea sp), with Roundup at a rate of
    1.7 kg a.i./ha affected breeding bird populations up to 3 years
    after treatment (Santillo et al., 1989a). Total bird densities
    decreased with 36% due to reduced habitat complexity, as expressed
    in regenerated hardwood, vegetation height and foliage height
    diversity. The most sensitive species were the insectivorous common
    yellowthroat  (Geothlypis trichas), Lincoln's sparrows  (Melospiza
     lincolnii) and alder flycatchers  (Empidonax alnorum). In less
    than 7-year-old clear-cuts in the Oregon coast range, Canada,
    planted with Douglas fir  (Pseudotsuga menziesii), some breeding
    bird populations were affected due to two closely spaced treatments
    with Roundup at a rate of 0.8 kg a.i./ha each (Morrison & Meslow,
    1984). Two years after the application, the densities of birds using
    shrubs for nesting and foraging had recovered, concomitant with the
    recovery of shrub vegetation. Sensitive species were the
    rufous-sided towhee  (Piplio erythrophthalmus) and MacGillyvray's

    warbler  (Oporornis tolmiei). On the other hand, the American
    goldfinch  (Carduelis tristis) increased one year after
    application, apparently due to the treatment. Morrison & Meslow
    (1986) stated that the effectiveness of the treatment, which was not
    maximal in their study, is crucial to the degree to which bird
    populations are affected.

         Small mammals may be affected by treatment with glyphosate,
    this depending mainly on the size of the treated area, the
    vegetation type and the extent to which the vegetation is damaged.
    Various experiments have been performed in and around clear-cuts
    planted with conifers in locations in the USA and Canada.
    Insectivorous shrews  (Blarina brivicauda, Sorex cinereus and Sorex
     hoyi) and herbivorous voles were less abundant due to a treatment
    with Roundup at a rate of 1.7 kg a.i./ha (Santillo et al., 1989b).
    This significant reduction of the number of shrews was maintained
    for 3 years after application, whereas the population of voles
    recovered. No effects on the omnivorous deer mice  (Peromyscus
     maniculatus) were observed. Deer mice also appeared to be
    unaffected in an experiment in which Roundup was applied at a rate
    of 3 kg a.i./ha (Sullivan, 1990). The population dynamics of both
    deer mice and the herbivorous Oregon voles  (Microtus oregoni)
    appeared not to be influenced, although some partially significant
    effects were observed that might have been due to the treatment. On
    the sprayed site there was an increase in the number of recruits of
    both species 3 years after application, and also an increased
    fecundity of deer mice and a higher survival of female voles 1 and 3
    years after application. Sullivan (1990) observed no physiological
    changes that might have been due to ingestion. In a clear-cut
    treated with Roundup at a rate of 1.1 kg a.i./ha, only the
    population density of Southern Redbacked voles  (Clethrionomys
     gapperi) was reduced by about 80% in a 1-year experiment (D'Anieri
    et al., 1987). In another clear-cut, no adverse effects on deer mice
    populations were evident after treatment with Roundup at a rate of
    2.2 kg a.i./ha (Sullivan & Sullivan, 1981). Contrary to the results
    of Sullivan (1990), D'Anieri et al. (1987), Sullivan & Sullivan
    (1981) and Santillo et al. (1989b), a significant reduction in the
    population density of deer mice was observed by Ritchie et al.
    (1987) on a sprayed clear-cut of 38 ha at around 11 months after
    spraying with Roundup at a rate of 1.1-1.2 kg a.i./ha. However, no
    adverse effects on fertility or fecundity were indicated. Probably
    the effect on abundance was due to habitat change with respect to
    feed provision and cover. Possibly the effects on deer mice observed
    by Ritchie et al. (1987) were less confounded by immigration as the
    sprayed area was larger than in the studies in which effects on deer
    mice were lacking. However, when a site of 36 ha was sprayed twice

    with Roundup at a rate of 0.8 kg a.i./ha on each occasion, deer mice
    were not affected, possibly due to a relatively low effectiveness of
    the treatment (Anthony & Morrison, 1985). In the treated area, even
    an increase of Oregon voles was found after 1 year, concomitant with
    an increase of grass and other plants. The results indicated that
    small mammal populations were able to recover within 2 years after
    application of glyphosate, dependent on the recovery of shrubs.

    10.  EVALUATION OF HUMAN HEALTH HAZARDS AND EFFECTS ON THE
         ENVIRONMENT

    10.1  Human health hazards

         Results of direct measurements of glyphosate concentrations in
    foodstuffs (as part of food surveillance), drinking-water or total
    diets are not available.

         Absorption from the gastrointestinal tract is limited to 36% or
    less and percutaneous absorption is 5.5% or less. Glyphosate is
    essentially not metabolized. Total body clearance is 99% in 7 days.
    Residues in livestock animals and their products (including milk)
    are minimal.

         Summarized information on short- and long-term studies is given
    in Table 23 and on teratogenicity and reproduction studies in Table
    24.

         In animals, glyphosate has very low acute toxicity by the oral
    and dermal administration routes. The formulation Roundup is acutely
    toxic to humans when ingested intentionally or accidentally. No
    controlled studies are available and therefore the human NOAEL
    cannot be derived.

         Animal studies show that glyphosate is not carcinogenic,
    mutagenic or teratogenic. Reproductive effects were only seen at
    dose levels producing maternal toxicity.

         In experimental animals (13-week studies in rats and mice), an
    effect was observed in the parotid salivary glands, indicating that
    glyphosate may be acting as a weak adrenergic agonist. In rats, this
    occurred at feeding levels of > 205 mg/kg body weight per day.
    The NOAEL in chronic feeding studies is > 410 mg/kg body weight
    per day. A NOAEL of 175 mg/kg body weight per day observed in a
    rabbit teratogenicity study was chosen as the appropriate basis for
    toxicological evaluations in humans. Through application of a
    suitable safety factor, safe levels for humans can be estimated for
    use in the toxicological evaluation of any actual exposures. For
    technical glyphosate a safety factor of 100 is considered
    appropriate given the elaborate data sets available.

         Glyphosate and its concentrated formulations produce moderate
    to severe eye irritation, but only slight skin irritation. Neither
    glyphosate nor tested formulations induce sensitization.

    
    Table 23. Short-term and long-term studies on glyphosate
                                                                                               

    Species   Test             Dose levels        Effects, dose level               NOAEL
              compound         (mg/kg-1 diet)     (mg/kg diet)                   [mg/kg diet]
                               unless otherwise                                mg kg-1 b.w. d-1
                               stated
                                                                                               

    Short-term studies

    Mouse     technical        5000, 10 000,      decreased growth and increased    [10 000]
              glyphosate       50 000             weights in brain, heart,           1890 m,
                                                  kidneys (50 000)                   2730 f

    Mouse     technical        3125, 6250,        reduced weight gain (50 000)       [3 125]
              glyphosate       12 500, 25 000,    lesions of salivary glands            507
                               50 000             (> 6250)

    Rat       technical        1000, 5000,        no adverse effects                [20 000]*
              glyphosate       20 000                                                  1267*

    Rat       technical        200 to 12 500      no adverse effects                [12 500]*
              glyphosate                                                                NG

    Rat       technical        3125, 6250,        increased AP and ALAT (> 6250)    [< 3 125]
              glyphosate       12 500, 25 000,    increased haematocrit and red      < 205 m
                               50 000             cell parameters (> 12 500),        < 213 f
                                                  increased bile acids,
                                                  decreased sperm counts
                                                  (> 25 000), histological
                                                  alterations in salivary glands
                                                  (> 3125), reduced weight
                                                  gain (> 25 000)

    Dogs      technical        20, 100, 500       no adverse effects                   [NG]
              glyphosate       mg kg-1 bw                                               500*

    Cattle    Roundup          400, 500, 630,     decreased feed intake (> 630)        [NG]
                               790                mg kg-1 bw d-1), diarrhoea (> 500)     400
                                                  increased blood parameters
                                                  (790)
                                                                                               

    Long-term studies

    Mouse     technical        1000, 5000,        decreased growth (30 000),        [5 000]
              glyphosate       30 000             increased incidence of               814
                                                  hepatocyte hypertrophy and
                                                  necrosis (30 000), increased
                                                  incidence of urinary bladder
                                                  epithelial hyperplasia
                                                  (30 000)

    Rat       technical        2000, 8000,        decreased growth (20 000),        [8 000]
              glyphosate       20 000             increased liver weights              410
                                                  (20 000), increased incidences
                                                  of degenerative lens changes
                                                  (20 000) and of gastric
                                                  inflammation (8000 and 20 000)

    Rat       technical        60, 200, 600       slightly decreased growth (600)   a
              glyphosate
                                                                                               

    m = males; f = females; * Highest dose tested; NG, not given;
    a    The slight effect at 600 mg/kg diet (32 mg/kg bw) is considered marginal in the light
         of the absence of an effect on growth at higher dose levels (2000 and 8000 mg/kg diet) 
         in a more recent 2-year study in rats.
    
         Available studies on exposures of workers involved in
    appli-cation of the herbicide show that exposure is low when
    protective clothing is worn. The following data illustrate this
    point.

    a)   The highest estimated exposure (dermal and inhalation) of about
         8000 µg/h, as reported in a study with spray applicators,
         corrected for incomplete absorption, equals about 50 µg/kg body
         weight per day (8-h working day for a 70-kg adult); between the
         latter level and the NOAEL of 175 mg/kg body weight per day,
         adjusted for incomplete absorption from the gastrointestinal
         tract (30-60%), i.e. 52-63 mg/kg body weight per day, there is
         a margin of safety of about 1100.

    b)   The highest exposure concentration found for forest brush saw
         workers was 15.7 µg/m3; between this level and the NOAEL
         expressed as glyphosate from the 4-week inhalation study with
         Roundup of 16 mg/m3 there is a margin of safety of 1000 (this
         is borne out by the absence of adverse findings in the workers'
         health examination in the study.

    
    Table 24.  Summary of teratogenicity and reproduction studies on glyphosate
                                                                                               

    Species   Test          Dose levels             Effects                             NOAELa
              compound      (mg/kg diet)                                                (mg/kg
                                                                                        body
                                                                                        weight)
                                                                                               

    Rat       technical     300, 1000, 3500         mortality, clinical signs and       1000
              glyphosate    mg/kg body              decreased growth in dams,
                            weight,                 early resorptions, decreased
                            gestation days 6-19     numbers of implantations and
                                                    visible fetuses, decreased
                                                    ossification of fetal
                                                    sternebrae (all at 3500 only);
                                                    no fetal malformations

    Rabbit    technical     75, 175, 350            diarrhoea and soft stools (350,     175
              glyphosate    mg/kg body              slight at 175), nasal discharge
                            weight,                 (350)
                            gestation days 6-27

    Rat       technical     3, 10, 30 mg/kg         increased incidence of renal        < 30b
              glyphosate    body weight             tubular dilation in F3b male
                            given in diet,          pups (30)
                            3 gens

    Rat       technical     2000, 10 000,           soft stools of parents (30 000),    100b
              glyphosate    30 000 mg/kg            decreased litter size (30 000),     (2000
                            diet, 2 gens            decreased body weights of           mg/kg
                                                    parents and pups (30 000            diet)
                                                    and 10 000)
                                                                                               

    a    Based on all observed effects (both in dams and offspring)
    b    There is some discrepancy in the results, and in the NOAELs, of the two
         reproduction studies carried out with technical glyphosate; the renal
         effects in the 3-generation study were not reproduced in the more recent
         2-generation study  with higher dose levels (for details, see section 7.5.2.).
    
    10.2  Evaluation of effects on the environment

         Following application, glyphosate will selectively partition to
    particulate matter suspended in surface water, aquatic sediment or
    to the soil substrate. This partitioning is usually rapid. The
    mechanism of sorption is only partially understood. There is little
    reported information on desorption from soil; the information
    available suggests "strong" binding. Mobility studies show little
    leaching of glyphosate below the upper few centimetres of the soil
    profile. The major metabolite, AMPA, also appears not to leach
    through soil.

         Given this environmental distribution, organisms living in
    aquatic sediment or soil would be expected to come into closest
    contact with residues of the herbicide.

         There is very little information on the bioavailability of
    sediment- or soil-bound glyphosate to either aquatic or terrestrial
    organisms. Few bioaccumulation or ecotoxicity studies have been
    performed with sediment present.

         Comparison of exposure concentrations and toxic effects is,
    therefore, difficult since the relevant organisms have not been
    tested and exposure of tested organisms is not by a realistic route.

    10.2.1  Exposure levels and toxic effects

         Exposure concentrations have been calculated from experimental
    application of Roundup in the field (see Table 7 in chapter 5). The
    methodology is presented in Fig. 4. No monitoring results of
    environmental concentrations following actual use in agriculture or
    forestry are available.

         The lowest LC(EC)50 and NOEC values for microorganisms,
    invertebrates and fish have been taken from the toxicity tests
    reported in chapter 9 (see Fig. 4). These all refer to organisms
    living in the open water and are, therefore, of questionable
    significance for a compound which partitions to sediment. There is
    no information on species living in aquatic sediment and little
    information available on soil-living organisms, with the exception
    of microorganisms.

    10.2.2  Hazard evaluation for aquatic organisms

         Tables 25 and 26 compare the estimated mean and maximum
    exposure concentrations, following aerial application of Roundup, to
    the lowest reported LC(EC)50 and NOEC concentrations for acute and
    chronic exposure of aquatic organisms. The ratio between exposure
    and effect concentrations has been calculated. These tables are

    meant as a guide to establishing possible hazard and are not
    intended to estimate the degree of effect likely to be seen in the
    field. The "possible hazard" classification is a simple one using
    different classification phrases for order of magnitude segments of
    the ratios.

         The toxicity of formulations to aquatic organisms is greater
    than for technical glyphosate in many cases. This increased toxicity
    is due to surfactants present in the product. No account has been
    taken of possible degradation of surfactants in the ratios
    presented, since no data are available. The ratios, therefore, may
    overestimate the toxicity of glyphosate. If the compound is bound to
    sediment in the environment, this could also reduce its toxic
    effect. Since no clear evidence is available to demonstrate this
    reduced toxicity, no account has been taken of partitioning to
    particulates. This will also tend to overestimate toxic effect of
    glyphosate.

         As can be seen from the tables, possible hazard for most
    aquatic organisms is small or negligible. Fish and aquatic
    invertebrates would not be affected by glyphosate use. Only
    microorganisms, with both acute and chronic exposure, appear to be
    susceptible to effects of the herbicide. The comparisons made in the
    table do not allow estimates of the degree of toxic effects likely
    to be seen in the field. From the field evidence available,
    populations and communities of algae are not likely to be affected
    after application of glyphosate formulation. Transitory changes in
    number and functioning of aquatic microorganisms are possible after
    use of the herbicide.

         Since data are not available, evaluation of the hazard of bound
    residues of glyphosate to sediment-living organisms is not possible.

         Minimal bioaccumulation of glyphosate has been reported in both
    laboratory experiments and in the field. The physicochemical
    properties of the compound are consistent with this conclusion.

    10.2.3  Hazard evaluation for terrestrial organisms

         Limited test data show low toxicity of glyphosate and its
    formulations to honey-bees, earthworms, birds and mammals. These
    data suggest low risk for these organisms from use of the herbicide.
    Field studies have been conducted and support the view that
    glyphosate does not affect soil microorganisms in the long term.

         Marked changes in populations of birds and small mammals have
    been seen in field studies following application of glyphosate.
    These seem to result from the changes in habitat, vegetation cover,
    food organisms, etc., resulting from the intended herbicidal effect
    of the compound.

    FIGURE 4


    
    Table 25.  Indications of environmental hazard for aquatic organisms by technical grade glyphosate
                                                                                                                                              

    Effect        Organisms           Estimated exposure   Toxicity data      End-point        Ratio of exposure    Possible
                                      concentration        (mg a.i./litre)                     to toxicity          hazard
                                      (mg a.i./litre)                                          concentrations
                                                                                                                                              

    Mean estimated exposure concentration

    Acute         microorganisms           0.1             EC50 = 1.2         mortality            0.082            small
    Acute         insects                  0.1             EC50 = 55          mortality            0.0018           negligible
    Acute         crustaceans              0.1             EC50 = 281         mortality            0.00035          negligible
    Acute         fish                     0.1             LC50 = 94          mortality            0.0010           negligible
    Chronic       algae                    0.05            NOEC = 0.3         growth               0.17             present
    Chronic       crustaceans              0.01            NOEC = 100         reproduction         0.00010          negligible
    Chronic       fish                     0.01            NOEC = 52          behaviour            0.00019          negligible

    Maximum estimated exposure concentration

    Acute         microorganisms           1.7             EC50 = 1.2         mortality            1.4              large
    Acute         insects                  1.7             EC50 = 55          mortality            0.031            small
    Acute         crustaceans              1.7             EC50 = 281         mortality            0.0060           negligible
    Acute         fish                     1.7             LC50 = 94          mortality            0.018            small
    Chronic       algae                    1.7             NOEC = 0.3         growth               5.7              large
    Chronic       crustaceans              0.17            NOEC = 100         reproduction         0.0017           negligible
    Chronic       fish                     0.17            NOEC = 52          behaviour            0.003            negligible
                                                                                                                                              

    Table 26.  Indications of environmental hazards for aquatic organisms by Roundup
                                                                                                                                              

    Effect            Organisms         Estimated exposure    Toxicity data          End-point     Ratio of exposure      Possible
                                        concentration         (mg Roundup/litre)                   to toxicity            hazard
                                        (mg Roundup/litre)                                         concentrations
                                                                                                                                              

    Mean estimated exposure concentration

    Acute             microorganisms         0.32             EC50 = 2.1             mortality          0.15              present
    Acute             crustaceans            0.32             EC50 = 10              mortality          0.032             small
    Acute             insects                0.32             EC50 = 44              mortality          0.0073            negligible
    Acute             fish                   0.32             LC50 = 13              mortality          0.025             small
    Chronic           microorganisms         0.16             NOEC = 0.7             growth             0.23              present
    Chronic           crustaceans            0.032            NOEC = 3.5             reproduction       0.0091            negligible
    Chronic           fish                   0.032            NOEC = 2.4             behaviour          0.013             negligible

    Maximum estimated exposure concentration

    Acute             microorganisms         5.6              EC50 = 2.1             mortality          2.7               large
    Acute             crustaceans            5.6              EC50 = 10              mortality          0.56              present
    Acute             insects                5.6              EC50 = 44              mortality          0.13              present
    Acute             fish                   5.6              LC50 = 13              mortality          0.43              present
    Chronic           microorganisms         5.6              NOEC = 0.7             growth             8.0               large
    Chronic           crustaceans            0.56             NOEC = 3.5             reproduction       0.16              small
    Chronic           fish                   0.56             NOEC = 2.4             behaviour          0.23              small
                                                                                                                                              
    

    11.  RECOMMENDATIONS FOR PROTECTION OF HUMAN HEALTH

    a)   Protective clothing is necessary to ensure the safety of
         herbicide applicators.

    b)   A market-basket survey would be useful to determine the
         possible exposure of the general population.

    12.  FURTHER RESEARCH

    a)   Further research is required to determine whether ß-adrenergic
         effects observed in rodents have any implications for human
         health.

    b)   The role of adjuvants in the toxicity of glyphosate
         formulations needs to be investigated further in laboratory
         mammals and organisms in the environment.

    c)   A controlled study on exposure of agricultural workers is
         needed.

    d)   The bioavailability of sediment- and soil-bound glyphosate in
         the environment should be studied.

    e)   Studies on the environmental behaviour and fate of adjuvants
         are required.

    f)   Further toxicity studies of sediment-living organisms are
         needed.

    g)   The effects of phosphate fertilizers on the binding of
         glyphosate to soils should be investigated.

    h)   Analytical techniques that are less costly but still adequate
         should be developed.

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    RESUME

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

         Le glyphosate, ou N-(phosphonométhyl)glycine, est un acide
    organique faible. Sa formule brute est C3H8NO5P. Il est
    généralement présenté sous la forme du sel de l'acide correspondant
    déprotoné et d'un cation comme l'isopropyl-ammonium ou le
    triméthylsulfonium. La pureté du glyphosate de qualité technique est
    généralement supérieure à 90%. Le glyphosate de qualité technique se
    présente sous la forme d'une poudre cristalline blanche inodore dont
    la densité est de 1,704. Sa tension de vapeur est très faible et il
    est très soluble dans l'eau. Le coefficient de partage octanol-eau
    (log Kow) est égal à -2,8. Le glyphosate est amphotère et il peut
    exister sous différentes formes ioniques, selon le pH du milieu.

         Le dosage du glyphosate est généralement une opération
    laborieuse, complexe et coûteuse. La méthode la plus courante
    consiste à en préparer un dérivé avec une substance fluorigène,
    avant ou après passage sur colonne. Le dosage s'effectue en général
    par chromatographie liquide à haute performance ou chromatographie
    gaz-liquide. Les limites de détection dans l'eau, les plantes, le
    sol et l'urine humaine sont respectivement de 0,02-3,2 µg/litre,
    0,01-0,3 mg/kg, 0,05-1 mg/kg et 0,1 mg/litre.

    2.  Sources d'exposition humaine et environnementale

         Le glyphosate est un herbicide non sélectif utilisé en
    traitement endothérapique après l'émergence; il est utilisé partout
    dans le monde sur des terrains agricoles ou non. On l'épand en
    plusieurs formulations commerciales sur de nombreuses récoltes. La
    plus courante est le Roundup qui consiste en un sel
    d'isopropylammonium. La dose d'emploi recommandée ne dépasse 5,8 kg
    de matière active par hectare et dépend de l'usage auquel on le
    destine. L'environnement peut être contaminé par suite du dépôt
    d'embruns ou de la libération accidentelle du produit.

    3.  Transport, distribution et transformation dans l'environnement

         Les principaux processus de dissipation qui interviennent après
    l'épandage de cet herbicide sont les suivants: formation de
    complexes avec certains ions présents dans l'eau comme Ca2+ et
    Mg2+, sorption aux sédiments ainsi qu'aux particules en suspension
    dans l'eau et le sol, photodécomposition dans l'eau, fixation par
    les plantes et biodégradation.

         Le glyphosate disparaît de l'eau avec des valeurs du TD50
    (temps de dissipation) qui vont de quelques jours à plus de 91
    jours. Le principal milieu récepteur est constitué par les sédiments
    ou les particules en suspension.

         En laboratoire, les coefficients d'adsorption (Ks/l) du
    glyphosate varient de 8 à 377 dm3/kg pour différents sols et
    substances argileuses. On ne dispose d'aucune donnée sur la sorption
    de l'acide aminométhylphosphonique (AMPA) qui en est le principal
    métabolite, dans les conditions du laboratoire.

         La valeur du Rf ne dépasse pas 0,2, selon certaines mesures par
    chromatographie sur couche mince de terre. Dans des conditions de
    lessivage reproduisant des précipitations extrêmement fortes, on
    récupère dans l'éluat d'une colonne de terre, entre 0,1 et 11% de
    l'activité appliquée initialement. L'expérimentation sur le terrain
    montre qu'il n'y a probablement pas lessivage de l'AMPA.

         L'expérimentation sur le terrain montre qu'en ce qui concerne
    la dissipation du glyphosate dans le sol, les valeurs du TD50
    varient de 3 à 174 jours, principalement en fonction des conditions
    édaphiques et climatiques. Certaines expériences sur le terrain ont
    montré que le ruissellement pouvait entraîner jusqu'à 1,8% de la
    dose appliquée sur la sol.

         Au laboratoire jusqu'à 45% de l'activité appliquée peut être
    absorbée par le feuillage après traitement, après quoi il y a une
    migration importante dans la plante.

         L'hydrolyse du glyphosate en tampon stérile est très lente, les
    valeurs du TD50 étant >> à 35 jours. En ce qui concerne la
    photodécomposition dans l'eau dans les conditions naturelles, les
    valeurs du TD50 sont < à 28 jours. Lors d'une étude qui s'est
    prolongée pendant 31 jours, on n'a pas enregistré de
    photodécomposition notable dans le sol.

         Le temps nécessaire à la biodégradation de 50% d'une quantité
    donnée de glyphosate dans l'ensemble d'un système d'épreuve en
    présence d'eau et de sédiments était < à 14 jours en aérobiose et
    compris entre 14 et 22 jours en anaérobiose. Dans le sol, le temps
    de demi-biodégradation du glyphosate est de 2 à 3 jours en
    aérobiose.

         Le principal métabolite qui se forme dans le sol et dans l'eau
    est l'AMPA. La quantité maximale d'AMPA présente dans le sol est
    d'environ 20% de l'activité appliquée en aérobiose et de 0,5% de
    cette activité en anaérobiose. Ce chiffre atteint 25% dans les
    sédiments dans les deux types de conditions.

         Les épreuves de laboratoire montrent que chez les invertébrés
    et les poissons, le facteur de bioconcentration est faible. Lors
    d'une épreuve en aquarium à écoulement continu, on a constaté que
    chez  Lépomis macrochirus le temps de demi-épuration était de 35
    jours après une exposition de même durée. Après exposition continue
    à du glyphosate, on retrouve de l'AMPA chez ce même poisson pendant
    des périodes allant jusqu'à 21 jours. Des mesures sur le terrain

    n'ont pas permis de déceler la présence de glyphosate chez les
    poissons vivant dans des eaux sur lesquelles cet herbicide avait été
    directement pulvérisé. Lors d'une expérience, on a décelé de l'AMPA
    chez les carpes jusqu'à 90 jours après l'épandage. Une autre
    expérience menée sur le terrain a montré qu'il n'y avait pas de
    bioamplification du glyphosate dans les portées de petits mammifères
    herbivores ou omnivores vivant en brousse. On a notamment mesuré des
    concentrations allant jusqu'à 5 mg de matière active par kg chez des
    souris du genre  Peromyscus, immédiatement après l'épandage.

         Plusieurs souches de bactéries peuvent décomposer le
    glyphosate. On a identifié des bactéries qui sont capables
    d'utiliser de composé comme seule source de phosphore, de carbone ou
    d'azote. La croissance est alors plus lente que lorsque elles
    utilisent des sources inorganiques de P, de C et de N. On est fondé
    à penser, d'après les observations effectuées sur le terrain, que
    certaines populations bactériennes se sont adaptées à la
    métabolisation du glyphosate. La présence de phosphate inorganique
    inhibe la décomposition du glyphosate par certaines bactéries mais
    pas toutes. La biodécomposition du glyphosate peut comporter un
    co-métabolisme avec d'autres sources d'énergie.

    4.  Concentrations dans l'environnement et exposition humaine

         Il n'existe que de très rares données provenant de programmes
    de surveillance systématique et concernant la présence de glyphosate
    dans la faune et la flore ainsi que dans le milieu abiotique. Pour
    avoir une idée des concentrations maximales dans l'environnement, on
    fait appel aux données fournies par des essais sur le terrain au
    cours desquels on simule des épandages à usage agricole; ces
    concentrations sont les suivantes: < 1-1700 µg/litre dans les eaux
    de surface, 0,07-40 mg/kg de poids sec dans le sol, < 0,05-19 mg/kg
    de poids sec dans les sédiments, 261-1300 mg/kg dans les feuilles,
    5 mg/kg dans les viscères des souris du genre  Peromyscus,
    1,6-19 mg/kg dans les baies sauvages et 45 mg/kg dans les lichens.
    Les concentrations maximales correspondantes d'AMPA sont les
    suivantes: < 1-35 µg/litre (eaux de surface), 0,1-9 mg/kg de poids
    sec (sol), < 0,05-1,8 mg/kg de poids sec (sédiments), 1,7-<
    9 mg/kg (feuilles), 0,02-0,1 mg/kg (baies sauvages) et 2,1 mg/kg
    (lichens). Les concentrations ci-dessus de glyphosate sont celles
    que l'on observe en général immédiatement après l'épandage. En ce
    qui concerne les lichens, la concentration mentionnée a été observée
    270 jours après l'épandage.

         On ne dispose pas de mesures de la dose journalière ingérée par
    l'homme avec les aliments et l'eau de boisson (études de rations
    totales). Les quelques données disponibles au sujet de l'exposition
    professionnelle indiquent que celle-ci est faible pour les ouvriers
    qui épandent du glyphosate comme désherbant sous forme de Roundup.

    5.  Cinétique et métabolisme chez les animaux de laboratoire et
        l'homme

         Le glyphosate technique n'est que partiellement résorbé au
    niveau des voies digestives. Lors d'études effectuées sur du
    glyphosate marqué au carbone-14, on a observé des pourcentages
    d'absorption de 30 à 36% chez plusieurs espèces. L'absorption par
    voie percutanée est faible. Dans le cas de l'herbicide Roundup, le
    glyphosate qu'il contient est absorbé dans une proportion < à
    5,5% à travers la peau (durée de contact environ 24 heures). En ce
    qui concerne les tissus de l'organisme, la concentration maximale,
    correspondant à environ 1% de la dose ingérée, se retrouve dans les
    eaux. Après administration d'une seule dose par voie orale, le
    produit est éliminé à hauteur de 62 à 69% dans les matières fécales
    sans absorption. Après absorption, 14 à 29% de la dose passe dans
    l'urine et 0,2% au maximum dans l'air expiré. Après administration
    par voie intraveineuse, le taux d'excrétion dans les voies biliaires
    n'a été que de 5 à 8%. Chez des chèvres en lactation, on a montré
    que le glyphosate n'était excrété dans le lait qu'en faible
    proportion (concentration < à 0,1 mg/kg de lait entier pour une
    dose ingérée de 120 mg/kg de nourriture). Le glyphosate n'est
    métabolisé que dans une très faible proportion. Son seul métabolite,
    l'AMPA, correspond à 0,3% de la dose ou même moins; le reste
    correspond au produit initial. Il faut environ 168 heures pour que
    le glyphosate soit éliminé en totalité de l'organisme (99% d'une
    dose orale).

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

         Chez l'animal de laboratoire, le glyphosate technique ne
    présente qu'une très faible toxicité aiguë lorsqu'il est administré
    par la voie orale ou percutanée; il est nettement plus toxique par
    la voie intrapéritonéale que par les autres voies d'administration.
    Des études d'alimentation de brève durée ont été effectuées sur
    plusieurs espèces, mais la plupart de ces épreuves n'ont guère
    révélé d'effets. Lors d'une épreuve de 13 semaines sur des souris au
    cours de laquelle on a utilisé du glyphosate technique, on a
    constaté une augmentation du poids de plusieurs organes ainsi qu'un
    retard de croissance à la dose de 50 000 mg/kg de nourriture. Lors
    d'une étude de même durée sur le rat, on n'a pas observé d'effet
    (les doses de glyphosate technique utilisées allaient jusqu'à
    20 000 mg/kg de nourriture). Lors d'une autre étude de 13 semaines,
    on a observé des lésions au niveau des glandes salivaires chez des
    rats et des souris. Chez les souris, la dose sans effet létal
    observable était de 3125 mg/kg de nourriture; chez le rat, elle
    était < à 3125 mg/kg de nourriture. Aucun de ces effets n'a été
    observé lors d'études à court ou à long terme effectuées sur
    diverses souches et espèces. Les lésions au niveau des glandes
    salivaires incitent à penser que le glyphosate pourrait se comporter
    comme un agoniste adrénérgique de faible activité.

         La toxicité à long terme a été étudiée sur des souris et des
    rats. Peu d'effets ont été observés et dans presque tous les cas,
    uniquement à des doses relativement élevées. Chez les souris, le
    glyphosate technique a produit un retard de croissance, une
    hypertrophie ou une nécrose des hépatocytes ainsi qu'une hyperplasie
    de l'épithélium vésical à la dose de 30 000 mg/kg. Chez les rats, le
    même composé a entraîné une réduction de la croissance, une
    augmentation du poids du foie, une dégénérescence du cristallin et
    une inflammation de la muqueuse gastrique à la dose de 20 000 mg/kg
    de nourriture.

         Les études dont on connaît les résultats ne concluent pas à
    l'existence d'un pouvoir mutagène, cancérogène ou tératogène du
    glyphosate technique. Deux études ont été effectuées sur plusieurs
    générations de rats. Les principaux effets du glyphosate technique
    consistaient en une réduction du poids corporel des géniteurs et des
    ratons ainsi qu'une diminution de la taille des portées, à la dose
    de 30 000 mg/kg de nourriture. Dans une étude portant sur la
    reproduction, on a constaté une augmentation dans l'incidence de la
    dilatation unilatérale des tubules rénaux chez les ratons mâles de
    la génération F3b, à la dose de 30 mg/kg de poids corporel.
    Toutefois la reproductibilité de cette lésion reste incertaine du
    fait qu'elle n'a pas été observée chez les ratons soumis à une dose
    plus élevée, dans la deuxième de ces études.

    7.  Effets sur l'homme

         Les études contrôlées dont on dispose se limitent à trois
    études sur l'irritation et la sensibilisation provoquées par le
    glyphosate chez des volontaires humains, et qui ont toutes donné des
    résultats négatifs. Plusieurs cas d'intoxication (la plupart du
    temps volontaires) avec un herbicide composé de glyphosate
    technique, le Roundup, ont été signalés. Une étude, consacrée à des
    travailleurs qui épandaient du Roundup, n'a pas révélé d'effets
    indésirables. Les données disponibles sur l'exposition
    professionnelle d'ouvriers appliquant du Roundup montrent que le
    niveau d'exposition est très inférieur à la dose sans effet létal
    observable qui ressort de l'expérimentation animale.

    8.  Effets sur les êtres vivants dans leur milieu naturel

         Le glyphosate de qualité technique est légèrement à modérément
    toxique pour les microorganismes aquatiques avec une CE50 (3 à 4
    jours) allant de 1,2 à 7,8 mg/litre et une concentration sans effets
    observables à 7 jours allant de 0,3 à 34 mg/litre. Sous ses
    différentes formulations, le glyphosate est légèrement à fortement
    toxique pour les microorganismes aquatiques avec des valeurs de la
    CE50 à 3 jours allant de 1,0 à plus de 55 mg de produit par litre.

    Les cyanophycées (algues bleues) sont plus sensibles au Roundup que
    les algues proprement dites. Les processus physiologiques affectés
    sont notamment le verdissement, la respiration, la photosynthèse et
    la synthèse des acides aminés aromatiques.

         Sur les bactéries terricoles en culture, le glyphosate agit au
    niveau de la fixation de l'azote, de la dénitrification et de la
    nitrification. Cependant des observations effectuées sur le terrain
    après épandage de diverses formulations de glyphosate n'ont pas
    révélé la présence d'effets sensibles. Des bactéries appartenant à
    des espèces étroitement apparentées aux bactéries précitées se sont
    révélées capables de dégrader le glyphosate.

         Chez les champignons ectomycorhiziens, la croissance du
    mycélium en culture pure est inhibée par des concentrations > à
    29 mg de Roundup par litre. Les genres sensibles à cette inhibition
    sont  Cenococcum, Hebeloma et  Laccaria.

         Le glyphosate est légèrement toxique pour les macrophytes
    aquatiques avec une valeur de la concentration sans effets
    observables à 14 jours de 9 mg/litre, en solution dans l'eau. le
    Roundup est également légèrement toxique avec, pour cette
    concentration, des valeurs allant de 2,4 à 56 mg/litre, également en
    solution dans l'eau. On ne dispose d'aucune donnée sur la toxicité
    aiguë. La phytotoxicité est beaucoup plus importante en l'absence de
    lessivage des dépôts d'herbicide.

         Le glyphosate de qualité technique est très légèrement à
    légèrement toxique pour les invertébrés aquatiques avec des valeurs
    de la CL50 ou de la CE50 à 2-4 jours > 55 mg/litre et une
    valeur de la concentration sans effets observables à 21 jours de
    100 mg/litre. Les diverses formulations de glyphosate sont très
    légèrement à modérément toxiques pour les invertébrés aquatiques
    pour des valeurs de la CE50 à 2 jours s'étageant entre 5,3 et
    5600 mg de produit par litre et des valeurs de la MATC à 21 jours
    allant de 1,4 à 4,9 mg de produit par litre. La toxicité plus forte
    du Roundup est essentiellement due à la présence d'agents
    tensioactifs.

         Le glyphosate de qualité technique est très légèrement à
    modérément toxique pour les poissons avec des valeurs de la CL50 à
    quatre jours allant de 10 à > 1000 mg/litre, une valeur de la
    concentration sans effets observables à 21 jours de 52 mg/litre et
    une valeur de la MATC de > 26 mg/litre. Les diverses formulations
    du glyphosate sont également très légèrement à modérément toxiques
    pour les poissons avec des valeurs de la CL50 à quatre jours de
    2,4 à > 1000 mg de produit par litre et des valeurs de la
    concentration sans effets observables à 21 jours allant de 0,8 à

    2,4 mg de produit par litre. C'est la carpe qui s'est révélée être
    l'espèce la plus sensible, après exposition à une formulation de
    glyphosate appelée Sting. Sur le terrain, on n'a pas constaté
    d'effets sur les poissons qui soient attribuables au traitement par
    le Roundup, à l'exception d'un stress constaté immédiatement après
    l'épandage du produit à la dose recommandée en évitant que celle-ci
    ne dépasse 40 mg de Roundup par litre.

         On constate que le glyphosate inhibe, dans une proportion qui
    dépend de la dose, la formation de nodosités par le trèfle
    souterrain inoculé par du Rhizobium, en culture hors-sol et en
    présence de solutions nutritives contenant une concentration de
    matière active > 2 mg/litre. La germination de diverses espèces
    forestières n'est pas affectée par la présence de glyphosate aux
    doses d'emploi recommandées. Avec des doses d'emploi > 0,54 kg de
    matière active par hectare, on constate au laboratoire qu'il y a
    réduction, proportionnée à la dose, de la longueur des racines des
    jeunes pousses de pins sylvestres. Cette diminution n'a pas été
    confirmée lors d'une étude du même genre sur le terrain.

         Le glyphosate de qualité technique et le Roundup sont
    légèrement toxiques pour les abeilles en application orale ou
    topique. Les valeurs de la DL50 à deux jours sont > 100 µg (de
    matière active ou de produit) par abeille. La DL50 par voie orale
    à deux jours du Sting pour les abeilles est > 100 µg/abeille. Le
    Roundup et le Roundup D-pack sont légèrement toxiques pour les
    lombrics avec des valeurs de la concentration sans effets
    observables à 14 jours respectivement égales à 500 et 158 mg de
    produit par kg de poids sec. Aucun effet nocif, attribuable au
    Roundup, n'a été observé sur la fécondité et la fertilité d'un
    certain nombre d'insectes appartenant au groupe des névroptères et
    le Sting n'a pas non plus produit d'effets sur la consommation de
    nourriture ou la mortalité des insectes du genre  Poecilus.

         Le glyphosate de qualité technique est légèrement toxique pour
    les oiseaux avec une DL50 > 3851 mg/kg de poids corporel, une
    CL50 à huit jours > 4640 mg/kg et des valeurs de la concentration
    sans effets observables à 112-119 jours, > 1000 mg/kg de
    nourriture. On a constaté que le Roundup et une autre formulation
    dont le nom n'est pas connu était également toxique pour les
    oiseaux, avec un DL50 > 2686 mg de produit par kg de poids
    corporel et une CL50 à huit jours > à 5620 mg de produit par kg
    de nourriture. Généralement, on ne constate, sur les mammifères de
    laboratoire, aucun effet qui soit attribuable au traitement par le
    glyphosate de qualité technique ou le Roundup. Les effets attribués
    au traitement par cet herbicide et constatés chez les oiseaux et les
    mammifères dans leur milieu naturel, semblent être dus
    principalement aux modifications du biotope consécutives au
    traitement herbicide.

    RESUMEN

    1.  Identidad, propiedades físicas y químicas y métodos analíticos

         El glifosato es un ácido orgánico débil formado por una
    molécula de glicina y otra de fosfonometilo. La fórmula empírica es
    C3H8NO5P. Normalmente se formula como una sal del ácido del
    glifosato en el que se ha sustituido un protón por un catión, por
    ejemplo la isopropilamina o el trimetilsulfonio. La pureza del
    glifosato de calidad técnica suele ser superior al 90%. Este es un
    polvo cristalino blanco e inodoro con un peso específico de 1,704,
    una presión de vapor muy baja y una solubilidad en agua alta. El
    coeficiente de reparto octanol/agua (log Kow) es -2,8. El
    glifosato es anfótero y se puede encontrar formando compuestos
    iónicos diversos, en función del Ph del medio.

         Su determinación es en general laboriosa, compleja y costosa.
    El método más habitual es la transformación con sustancias
    fluorogénicas en derivados más fácilmente detectables y se puede
    utilizar antes o después de la columna. La determinación se suele
    llevar a cabo mediante cromatografía líquida de alto rendimiento o
    cromatografía gas-líquido. Los límites de determinación del
    glifosato en el agua, las plantas, el suelo y la orina humana son de
    0,02-3,2 µg/litro, 0,01-0,3 mg/kg, 0,05-1 mg/kg y 0,1 mg/litro,
    respectivamente.

    2.  Fuentes de exposición humana y ambiental

         El glifosato es un herbicida que actúa después del brote de
    manera sistémica y no selectiva, y se utiliza en zonas agrícolas y
    no agrícolas de todo el mundo. Se aplica a numerosos cultivos con
    formulaciones comerciales diferentes. La más importante es el
    Roundup, en el que el glifosato aparece en forma de la sal de
    isopropilamina. Las dosis de aplicación recomendadas no superan los
    5,8 kg de a.i./ha y dependen del tipo de uso. Se puede producir
    exposición ambiental como consecuencia de la deposición debida a
    corrientes o escapes accidentales.

    3.  Transporte, distribución y transformación en el medio ambiente

         Las más importantes vías de desaparición del glifosato tras su
    aplicación son la formación en el agua de complejos con iones, por
    ejemplo con el Ca2+ y el Mg2+, la sorción al sedimento, las
    partículas suspendidas en el agua y el suelo, la fotodegradación en
    el agua, la fijación en las plantas y la biodegradación.

         El glifosato desaparece del agua con unos valores de TD50 que
    oscilan entre varios días y más de 91 días. Se ha comprobado que se
    deposita sobre todo en las partículas del sedimento o suspendidas.

         Los coeficientes de adsorción (Ks/l) del glifosato en
    experimentos de laboratorio varían entre 8 y 377 dm3/kg para
    diferentes suelos y minerales arcillosos. No se dispone de datos, en
    condiciones de laboratorio, sobre la sorción del ácido
    aminometilfosfónico (AAMF), su principal metabolito.

         En los experimentos de cromatografía en capa fina, los valores
    Rf del glifosato no son superiores a 0,2 en el suelo. En el eluato
    de columnas de suelo obtenido en condiciones de lixiviación
    simulando una precipitación muy intensa se recupera una cantidad que
    oscila entre menos del 0,1% y el 11% de la dosis aplicada. De los
    estudios sobre el terreno se desprende que no es probable la
    lixiviación del AAMF.

         En los experimentos sobre el terreno el glifosato desaparece
    del suelo con un TD50 que varía entre 3 y 174 días, principalmente
    en función de las condiciones edáficas o climáticas. En algunos
    experimentos sobre el terreno desaparecía del suelo, debido a la
    escorrentía, hasta el 1,8% de la dosis aplicada.

         En condiciones de laboratorio, las hojas tratadas podrían
    absorber hasta el 45% de la cantidad aplicada, produciéndose a
    continuación un importante desplazamiento.

         La hidrólisis del glifosato en tampones estériles es muy baja,
    con valores de TD50 >> 35 días. En condiciones naturales, la
    fotodegradación en agua se produce con valores de TD50 < 28 días.
    En el curso de un estudio de 31 días no se registró una
    fotodegradación importante en el suelo.

         El tiempo necesario para la biodegradación del 50% del
    glifosato en el sistema completo de una prueba con agua y sedimento
    es < 14 días en condiciones aerobias y de 14 a 22 días en
    condiciones anaerobias de laboratorio. El tiempo necesario para la
    biodegradación del 50% del glifosato en el suelo es de 2-3 días en
    condiciones aerobias.

         El metabolito principal en el suelo y el agua es el AAMF. Las
    cantidades máximas de AAMF en el suelo son de aproximadamente el 20%
    de la dosis aplicada en condiciones aerobias, y del 0,5% en
    condiciones anaerobias. Las cantidades máximas de AAMF en el
    sedimento son del 25%, tanto en condiciones aerobias como
    anaerobias.

         De las pruebas de laboratorio se desprende que los factores de
    bioconcentración en invertebrados y peces son bajos. Tras una
    exposición al glifosato de 35 días,  Lepomis macrochirus mostró en
    una prueba en corriente un periodo de semidepuración de 35 días. Se
    recuperó AAMF en  Lepomis macrochirus hasta 21 días después de una
    exposición continuada. No se detectó glifosato en peces que vivían

    en agua directamente rociada en experimentos sobre el terreno. En un
    experimento se detectó AAMF en carpas hasta 90 días después de la
    aplicación. En otro experimento sobre el terreno no se observó
    bioampliación del glifosato en el lecho de pequeños mamíferos
    herbívoros y omnívoros de un ecosistema de matorral boscoso. En este
    mismo experimento, inmediatamente después del rociado se
    determinaron concentraciones de hasta 5 mg de a.i./kg en ratones de
    pies blancos  (Peromyscus leucopus).

         Existen diversas bacterias que pueden degradar el glifosato. Se
    han identificado cepas capaces de utilizar este compuesto como única
    fuente de fósforo, de carbono o de nitrógeno. El crecimiento es
    lento si se compara con el obtenido de fuentes inorgánicas de P, C y
    N. Hay pruebas en el medio ambiente de la existencia de poblaciones
    bacterianas que se han adaptado para metabolizar el glifosato. La
    presencia de fosfato inorgánico inhibe la degradación de este
    compuesto por algunas bacterias, pero no por todas. La
    biodegradación del glifosato podría tener un metabolismo común con
    el de otras fuentes de energía.

    4.  Niveles ambientales y exposición humana

         Los datos sobre la presencia de glifosato en la biota y la
    abiota del medio ambiente como parte de programas de vigilancia
    regular son muy escasos. Se utilizan datos obtenidos en experimentos
    sobre el terreno en los que se simula la práctica agrícola normal
    para indicar las concentraciones máximas en el medio ambiente: <
    1-1700 µg/litro de agua superficial, 0,07-40 mg/kg de peso seco de
    suelo, < 0,05-19 mg/kg de peso seco de sedimento, 261-1300 mg/kg de
    follaje, 5 mg/kg de vísceras de ratón de pies blancos, 1,6-19 mg/kg
    de bayas silvestres y 45 mg/kg de líquenes. Las concentraciones
    máximas correspondientes de AAMF son las siguientes: <
    1-35 µg/litro (agua superficial), 0,1-9 mg/kg de peso seco (suelo),
    < 0,05-1,8 mg/kg de peso seco (sedimento), 1,7-< 9 mg/kg
    (follaje), 0,02-0,1 mg/kg (bayas silvestres) y 2,1 mg/kg (líquenes).
    Las concentraciones de glifosato mencionadas más arriba se suelen
    encontrar inmediatamente después de la aplicación. La concentración
    en los líquenes se determinó 270 días después de dicha aplicación.

         No se dispone de mediciones de la ingestión humana diaria de
    glifosato a través de los alimentos y el agua de bebida (estudios
    completos de alimentación). Los escasos datos disponibles sobre la
    exposición ocupacional ponen de manifiesto que los niveles de
    exposición para los trabajadores que aplican el glifosato en la
    formulación del herbicida Roundup son bajos.

    5.  Cinética y metabolismo en animales de laboratorio y en el ser
        humano

         La absorción del glifosato de calidad técnica en el tracto
    intestinal es sólo parcial. En estudios con glifosato marcado con
    14C, se encontró en varias especies un porcentaje de absorción del
    30% al 36%. La absorción cutánea es baja. De la formulación del
    herbicida Roundup, a través de la piel sólo se absorbe < 5,5% del
    glifosato presente (tiempo de contacto de unas 24 horas). En los
    tejidos del organismo, la concentración más alta, aproximadamente el
    1% de la dosis oral, se encuentran en los huesos. Tras una dosis
    oral única, se eliminó en las heces sin absorción el 62-69%. Del
    glifosato absorbido, un 14-29% se excretó en la orina y el 0,2% o
    menos en el aire expirado. La excreción biliar posterior a la
    administración intravenosa fue sólo del 5-8%. Se observó que la
    excreción en la leche de cabras lactantes se producía sólo en escasa
    proporción (concentración < 0,1 mg/kg de leche entera a un nivel
    de dosis de 120 mg/kg de alimentos). La biotransformación del
    glifosato se da únicamente en un grado muy bajo. El único
    metabolito, el AAMF, representa el 0,3% de la dosis o menos; el
    resto es glifosato inalterado. La eliminación de todo el organismo
    (99% de una dosis oral) se produce aproximadamente en 168 horas.

    6.  Efectos en animales de laboratorio y en sistemas de prueba
         in vitro

         El glifosato de calidad técnica administrado por vía oral y
    cutánea a animales de experimentación tiene una toxicidad aguda muy
    baja; por vía intraperitoneal es notablemente más tóxico que por
    cualquier otra. Aunque se han realizado estudios de alimentación de
    corta duración en varias especies, en la mayor parte de estas
    pruebas se han observado pocos efectos. En un estudio de 13 semanas
    realizado en ratones con glifosato de calidad técnica, a una
    concentración de 50 000 mg/kg de alimento, se observó aumento de
    peso de varios órganos y un retraso del crecimiento. En un estudio
    de 13 semanas en ratas no se advirtieron efectos (con dosis de
    glifosato de calidad técnica de hasta 20 000 mg/kg de alimento). En
    otro estudio de 13 semanas se detectaron lesiones en las glándulas
    salivales de ratas y ratones. En ratones, el NOAEL fue de 3125 mg/kg
    de alimento; en ratas fue < 3125 mg/kg de alimento. Estos
    resultados no se obtuvieron en ningún otro estudio de corta o larga
    duración realizado en diferentes razas y especies. Las lesiones de
    las glándulas salivales parecen indicar que el glifosato puede
    actuar como agonista adrenérgico débil.

         Se estudió la toxicidad a largo plazo en ratones y ratas. Se
    observaron escasos efectos y, en la mayor parte de los casos, sólo a
    dosis relativamente altas. Con dosis de 30 000 mg/kg de glifosato de
    calidad técnica se produjo en los ratones retraso del crecimiento,

    hipertrofia o necrosis de los hepatocitos e hiperplasia epitelial de
    la vejiga urinaria. La misma prueba en ratas con dosis de
    20 000 mg/kg de alimento provocó una disminución del crecimiento,
    aumento del peso del hígado, cambios degenerativos del cristalino e
    inflamación gástrica.

         De los estudios disponibles no se desprende que el glifosato de
    calidad técnica tenga actividad mutagénica, carcinogénica o
    teratogénica. Se realizaron con este compuesto dos estudios en
    varias generaciones de ratas. Los principales efectos del glifosato
    de calidad técnica con dosis de 30 000 mg/kg de alimento fueron una
    disminución del peso corporal de los padres y las crías y la
    reducción del tamaño de la camada. Se ha informado que en un estudio
    de reproducción con dosis de 30 mg/kg de peso corporal se produjo un
    aumento del número de casos de dilatación tubular renal unilateral
    en crías macho de la F3b. La ausencia de efectos renales en las
    crías con dosis más elevadas en el otro estudio de reproducción pone
    de manifiesto que la reproducibilidad de la lesión es incierta.

    7.  Efectos en el ser humano

         Sólo se dispone de tres estudios controlados sobre
    irritación/sensibilización en voluntarios, cuyos resultados indican
    la ausencia de efectos. Se ha informado de varios casos de
    intoxicación (la mayor parte intencionados) con la formulación
    Roundup de herbicida a base de glifosato de calidad técnica. No se
    detectaron efectos adversos tras realizar un estudio para determinar
    el estado de salud de los trabajadores que aplican la formulación
    del herbicida Roundup. Los datos disponibles sobre exposición en el
    trabajo de quienes aplican el Roundup indican que los niveles de
    exposición están muy por debajo del NOAEL obtenido en los
    experimentos correspondientes con animales.

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

         El glifosato de calidad técnica tiene una toxicidad de moderada
    a ligera para los microorganismos acuáticos, con valores de CE50
    (3-4 días) de 1,2-7,8 mg/litro y valores de NOEC (7 días) de
    0,3-34 mg/litro. Las formulaciones de glifosato son entre
    ligeramente tóxicas y muy tóxicas para los microorganismos
    acuáticos, con valores de CE50 en tres días de 1,0 a > 55 mg de
    producto por litro. Las cianofíceas (algas verdeazuladas) son más
    sensibles al Roundup que las algas verdaderas. Afecta a diversos
    procesos fisiológicos, entre ellos la formación del color verde, la
    respiración, la fotosíntesis y la síntesis de aminoácidos
    aromáticos.

         En cultivos de bacterias del suelo se ha comprobado la
    influencia del glifosato sobre la fijación del nitrógeno, la
    desnitrificación y la nitrificación. Sin embargo, en estudios sobre
    el terreno no se han observado efectos significativos tras la
    aplicación de varias formulaciones. Diversas bacterias estrechamente
    relacionadas han demostrado que son capaces de degradar el
    glifosato.

         A concentraciones > 29 µg de Roundup/litro se inhibe el
    crecimiento de los micelios de las ectomicorrizas en cultivos puros.
    Son géneros sensibles  Cenococcum, Hebeloma y  Laccaria.

         Cuando se disuelve en agua, el glifosato es ligeramente tóxico
    para las macrofitas acuáticas, con un valor de NOEC en 14 días de
    9 mg/litro. El Roundup disuelto en agua es también ligeramente
    tóxico, con valores de NOEC en 14 días de 2,4-56 mg de
    Roundup/litro. No se dispone de datos acerca de su toxicidad aguda.
    La fitotoxicidad es mucho más elevada cuando el agua no arrastra los
    depósitos del rociado.

         La toxicidad del glifosato de calidad técnica para los
    invertebrados acuáticos varía entre ligera y muy ligera, con unos
    valores de la CL50 o la CE50 en 2 a 4 días de > 55 mg/litro,
    y un valor de NOEC en 21 días de 100 mg/litro. Las formulaciones de
    glifosato tienen una toxicidad entre moderada y muy ligera para los
    invertebrados acuáticos, con valores de CE50 en 2 días de
    5,3-5600 mg de producto/litro y valores de MATC en 21 días de
    1,4-4,9 mg de producto por litro. La toxicidad más elevada del
    Roundup se debe fundamentalmente a la presencia de surfactantes.

         La toxicidad del glifosato de calidad técnica para los peces es
    entre moderada y muy ligera, con valores de CL50 en 4 días de 10 a
    > 1000 mg/litro, una NOEC en 21 días de 52 mg/litro, y un valor
    MATC de > 26 mg/litro. Las formulaciones del glifosato tienen
    también una toxicidad entre moderada y muy ligera para los peces,
    con valores de CL50 en 4 días de 2,4 a > 1000 mg de producto por
    litro y valores de NOEC en 21 días de 0,8-2,4 mg de producto/litro.
    La especie más sensible es la carpa, cuando se la expone a la
    formulación Sting. No se han observado en los peces efectos
    relacionados con el tratamiento de Roundup en el medio ambiente,
    salvo cierta tensión inmediatamente después de la aplicación de una
    dosis recomendada y evitando concentraciones > 40 mg de
    Roundup/litro.

         En sistemas de cultivo sin suelo con soluciones nutrientes en
    concentraciones > 2 mg de i.a./litro se produce una inhibición
    dependiente de la dosis de la nodulación del trébol subterráneo
    inoculado con Rhizobium. El glifosato en las dosis de aplicación
    recomendadas no afecta a la germinación de las semillas. La longitud

    de las raíces de los plantones de pino rojo disminuye en condiciones
    de laboratorio en función de la dosis con unas concentraciones de
    aplicación > 0,54 kg de i.a./ha. Esta reducción no se confirmó en
    un experimento comparable sobre el terreno.

         El glifosato de calidad técnica y el Roundup son ligeramente
    tóxicos para las abejas cuando se aplican por vía oral o tópica. Los
    valores de la DL50 en 2 días son > 100 µg (i.a. o producto) por
    abeja. La DL50 en 2 días por vía oral de Sting para las abejas es
    > 100 µg/abeja. Roundup y Roundup D-pak son ligeramente tóxicos
    para las lombrices de tierra, con valores NOEC en 14 días de 500 y
    158 mg de producto por kg de peso seco, respectivamente. No se
    observaron efectos adversos del Roundup sobre la fecundidad y
    fertilidad de especies de la familia Chrysopidae, y tampoco se
    detectaron efectos de Sting en la ingestión de alimentos y la
    mortalidad del escarabajo  Poecilus.

         El glifosato de calidad técnica es ligeramente tóxico para las
    aves, con una DL50 > 3851 mg/kg de peso corporal, una CL50 en 8
    días de > 4640 mg/kg de alimento, y valores NOEC en 112-119 días de
    > 1000 mg/kg de alimento. Roundup y una formulación desconocida
    son también ligeramente tóxicos para las aves, con una DL50 de >
    2686 mg de producto/kg de peso corporal y una CL50 en 8 días de >
    5620 mg de producto/kg de alimento. En condiciones de laboratorio no
    se han observado en general sobre los mamíferos efectos relacionados
    con el tratamiento de glifosato de calidad técnica o Roundup, salvo
    con dosis de aplicación muy elevadas. Los efectos relacionados con
    el tratamiento en las aves y mamíferos del medio ambiente parecen
    deberse fundamentalmente a cambios de hábitat después del
    tratamiento con Roundup.


    See Also:
       Toxicological Abbreviations
       Glyphosate (ICSC)
       Glyphosate (PDS)
       Glyphosate (Pesticide residues in food: 1986 evaluations Part II Toxicology)