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    FAO/PL:1968/M/9/1

    WHO/FOOD ADD./69.35

    1968 EVALUATIONS OF SOME PESTICIDE RESIDUES IN FOOD

    THE MONOGRAPHS

    Issued jointly by FAO and WHO

    The content of this document is the result of the deliberations of the
    Joint Meeting of the FAO Working Party of Experts and the WHO Expert
    Committee on Pesticide Residues, which met in Geneva, 9-16 December,
    1968.

    FOOD AND AGRICULTURE ORGANIZATION OF THE UNITED NATIONS

    WORLD HEALTH ORGANIZATION

    Geneva, 1969

    ETHION

    IDENTITY

    Chemical name

         OOO'O'-tetraethyl SS'-methylene di(phosphorothiolothionate) (IUPAC)

    Formula

    CHEMICAL STRUCTURE 

    Other information on identity and properties

    Liquid, practically non-volatile at ordinary temperatures, solidifying
    at -12 to -15C. Insoluble in water, somewhat soluble in kerosene and
    petroleum oils, soluble in most organic solvents. Slowly oxidized in
    air to a more reactive material.

    EVALUATION FOR ACCEPTABLE DAILY INTAKE

    Biochemical aspects

    Studies in vitro showed that the rate of reaction of ethion with
    beef erythrocyte cholinesterase is relatively slow, the velocity
    constant being about 3.5  103 litres per mole per minute. Liver
    fortified with diphosphopyridine nucleotide (DPN) converts ethion into
    a more potent cholinesterase inhibitor. Non-fortified liver did not
    enhance the activity (Hazleton Laboratories, Inc., 1958a).

    Acute toxicity

                                                                                
                         LD50 mg/kg
    Animal      Route    body-weight          References
                                                                            

    Mouse       oral         69        May and Baker. Ltd., 1960

                s.c.        630        May and Baker, Ltd., 1960

    Rat (M)     oral         65        May and Baker, Ltd., 1960

        (F)     oral         63        May and Baker, Ltd., 1960

    (continued)
                                                                            
                         LD50 mg/kg
    Animal      Route    body-weight          References
                                                                            
        (M)     s.c.        380        May and Baker, Ltd., 1960

        (F)     s.c.        360        May and Baker, Ltd., 1960

    Rat (M)     oral         97        Hazleton Laboratories, Inc., 1961

    Rat         oral        161        Industrial Bio-Test Laboratories, 1965
                                                                            
    
    Short-term studies

    In two separate studies, groups comprising equal numbers of male and
    female rats were fed dietary levels of 0, 3, 10, 30 and 100 ppm of
    ethion for periods up to 13 weeks. Five male and five female animals
    from each group were sacrificed after 30, 63 and 93 days for
    cholinesterase determination. All animals exhibited normal physical
    appearance, behaviour, growth and food consumption during the studies.
    There appeared to be little difference in the response of male and
    female rats with respect to cholinesterase activity at these levels.
    In the 100 ppm group significant inhibition of plasm, red blood cell
    and brain cholinesterase activity was evident after the 30, 63 and 93
    day test periods. Similar findings were noted in the 30 ppm group but
    to a lesser degree. In the 10 ppm group slight depression in plasma
    and red blood cell cholinesterase was noted after 30 days but not
    after 63 and 93 days, and there was never any effect observed in brain
    cholinesterase. At 3 ppm cholinesterase was normal. At the 100 ppm
    level plasma cholinesterase activity returned to normal 14 days after
    withdrawal of ethion from the diet, but red blood cell and brain
    cholinesterase remained slightly depressed. No histological changes
    were noted at any level (Hazleton Laboratories, Inc., 1958b).

    Five groups, each containing 10 male and 10 female rats, were fed
    ethion at dietary levels of 0, 300, 600, 1000 and 1500 ppm for 13
    weeks. Growth suppression was observed in all the test groups, except
    for the male rats at the 300 and 600 ppm levels. Signs of
    intoxication, with increasing frequency and severity in proportion to
    the dietary levels, were observed in all test animals, except for the
    males fed 300 ppm. All the animals in the 1500 ppm group died during
    the first two weeks. Six male and nine female rats died in the 1000
    ppm group. Survival in the other groups was comparable to controls.
    Complete inhibition of plasma and red cell cholinesterase activity
    occurred in all test groups, except in the males fed 300 ppm, where
    inhibition was marked but not complete. Almost total inhibition of
    brain cholinesterase activity was observed in the female test animals.
    The inhibition noted in the males was less marked, but in proportion
    to increasing dietary levels. No gross or histological changes, which

    could be attributed to ethion, were observed in any of the surviving
    rats (Hazleton Laboratories Inc., 1959).

    Dog

    Groups, each containing two male and two female dogs, received ethion
    orally in daily doses of 0.0125, 0.025, 0.075 and 0.25 mg/kg body
    weight for 90 days. A control value relative to cholinesterase
    activity was established for each dog during a two- week pre-dosage
    period.Physical appearance, behaviour, appetite, body weight and
    survival were unaffected. Autopsy at the conclusion of the experiment
    revealed no pathological changes. Plasma cholinesterase activity was
    significantly inhibited in one male dog and one female dog fed 0.25
    mg/kg (Hazleton Laboratories Inc., 1958c).

    Ethion was administered to groups, each containing three male and
    three female dogs, for six days a week during a 90-day period. Dosage
    levels were 0, 0.05, 0.075, 0.125, 0.25, 1.25 and 2.5 mg/kg body
    weight. No significant cholinesterase depression was observed at
    levels below 0.25 mg/kg. Depression of erythrocyte cholinesterase
    occurred only in the 2.5 mg/kg group. This depression was observed
    early in the test and persisted throughout its duration, but recovery
    was complete 24 days after withdrawal. Plasma cholinesterase was
    depressed more readily, occurring early in the test at the 2.5 mg/kg
    level but was not evident until the eleventh week of the test at the
    0.25 mg/kg level. Recovery was achieved 24 days after withdrawal in
    the case of the higher dose level and within nine days in the case of
    the 0.25 mg/kg level (Industrial Bio-Test Laboratories Inc., 1961a,
    1961b).

    Special studies

    (a) Reproduction

    Groups of male and female rats were fed diets containing 0, 2 and 30
    ppm of ethion through three generations. The second litter from each
    generation (F1b and F2b) was selected as parental animals to
    establish the next generation. No adverse effects were noted among
    either the parental animals or their progeny as regards growth,
    mortality, reactions, pathology, reproduction performance and survival
    indices of the young. Cholinesterase determinations were not conducted
    in this study (Industrial Bio-Test Laboratories Inc., 1965b).

    (b) Potentiation

    When ethion was administered to rats in combination with nine other
    organo-phosphorus insecticides, potentiation was observed only with
    malathion, where the ratio of observed to theoretical acute LD50 was
    2.9:1 (Hazleton Laboratories Inc., 1958d).

    Comments

    Ninety-day toxicity studies have been made in both rats and dogs
    together with a three-generation reproduction study in rats. The
    levels causing no toxic effects have been determined mainly on the
    results of the extensive cholinesterase inhibition studies, and were
    based on the absence of red cell cholinesterase depression. However,
    it would have been desirable to have data on cholinesterase depression
    at more frequent intervals.

    TOXICOLOGICAL EVALUATION

    Level causing no significant toxicological effect

         Rat: 3 ppm in the diet, equivalent to 0.15 mg/kg per day;
         Dog: 0.125 mg/kg per day.

    Estimate of acceptable daily intake for man

         0-0.00125 mg/kg body weight.

    RESIDUES IN FOOD AND THEIR EVALUATION

    Use pattern

    Pre-harvest treatments

    Ethion is used as a pre-harvest topical application to a variety of
    food crops especially citrus and deciduous fruit for the control of
    many species of aphids, scale insects, mites, leaf miners and leaf
    hoppers. It is also used in a cattle dip for the control of ticks and
    as a "back-line" treatment for the control of buffalo fly.

    Post-harvest treatments

    No known post-harvest uses.

    Residues resulting from supervised trials

    The typical data presented below have been extracted from Internal
    Reports of Niagara Chemical Division, FMC, except for that on grapes
    (Taschenberg et al., 1963) and strawberries (Fakey et al., 1962).

                                                                                           
                            Rate of            No. of       Pre-harvest     Residue
    Crop              application (kg/ha)    treatments   interval (days)    (ppm)
                                                                                       

    Almonds                   1.9                              0-28         < 0.1

    Apples                                       1               45           1

    (continued)
                                                                                       
                            Rate of            No. of       Pre-harvest     Residue
    Crop              application (kg/ha)    treatments   interval (days)    (ppm)
                                                                                       

    Beans                     1.9                1                7           0.27-0.43

    Beef               0.075% emulsion           3                3           0.01

    Beef, fat                 "                  3                3           0.86

    Citrus (juice)            1.25               2               15           0

      "    (peel)             1.25               1               30           1

    Cucumbers                 2.5                6               11           0.04

    Grapes                    2.5                1               28           2

    Melons                    1.25               1                7           0.07

    Raisins                   9                  3                9           1.25-10.4

    Strawberries              0.6                3                8           0.1

    Tea, leaves               0.9                1                7           1.71-7.06

    Tomatoes                  0.6                2                7           0.17-0.34
                                                                                       
    
    Fate of residues

    General comments

    Ethion shows some systemic activity. It is taken into the plant and
    translocated to other parts where it kills mites and certain insects.
    This effect diminishes rather rapidly, and correlates with the
    relative instability of the oxidized material by comparison with
    ethion itself. The residues are reduced by oxidation and subsequent
    hydrolysis on the surface or in the plant.

    In soils. It is usually used as a spray and no data on its
    degradation in soil have been made available.

    In plants. A surface residue on cotton foliage of 86 ppm was reduced
    to less than 1 ppm After eight days. Simultaneous analysis of
    cottonseed showed no detectable residue. Onions planted in 20-feet
    rows treated with 10 lb of three per cent dust per 20-feet rows
    showed insignificant residues at harvest, 123 days later (Niagara,
    1958).

    Tea, at harvest, sometimes shows residues as high as 7 ppm, as the
    result of "spot" treatment of particular bushes but the brewed product
    of these particular leaves shows residues of 0.25 ppm or less. Since
    tea is blended prior to sale to the ultimate consumer, a tolerance on
    blended tea of not more than 1 ppm and a tolerance of 7 ppm on tea
    from a particular estate appear justified (Niagara, 1966a).

    In animals. Lactating cows fed up to 20 ppm of radioactive ethion in
    their diet showed no ethion residues in their milk, the bulk of the
    radioactivity being associated with the protein fraction. In meat, the
    highest radioactivity occurred in the liver which contained an average
    of 3.15 ppm ethion-P32 equivalents at the end of the 28-day feeding
    at 20 ppm. This dropped by 60 per cent within 12 days after
    termination of the feeding period. However, chemical analysis showed
    that the radioactive material was not ethion but products from its
    metabolism (Hazelton Laboratories Inc., 1960; Niagara, 1960). Thus
    ethion residues in citrus pulp fed to cattle would not contribute to
    residues in milk.

    When animals were dipped, residues arising from absorption through the
    skin consisted largely of ethion and were mainly in the fat. The
    active oxidation products, the monothiol and bisthiol derivatives,
    were found at less than 0.01 ppm and 0.02 ppm respectively.

    In storage and processing. No data have been made available.

    Evidence of residues in food, in commerce or at consumption

    Duggan and Weatherwax (1967), in their survey of dietary intake of
    pesticide chemicals, found that the incidence and intake of ethion was
    too low to be detected. Trace amounts contributing to less than 0.001
    mg per day were found in fruit, samples being taken from June 1964 to
    April 1966.

    Martin and Duggan (1968) found residues of ethion at 0.024 to 0.054
    ppm in three out of 30 composite samples of fruit collected from 30
    markets in 29 different cities in the United States of America from
    June 1966 to April 1967. In the same studies ethion was found at 0.025
    ppm in one out of 30 composite samples of oils, fats and shortening.

    Methods of residue analysis

    A colorimetric procedure is based on the extraction of ethion, and
    necessary clean-up, followed by hydrolysis to diethylphosphorodithioic
    acid. The colour complex then formed with copper sulfate is measured
    at 418 m. From the optical density the amount of ethion is determined
    from a standard curve (Graham, 1964).

    An enzymatic method is based on the oxidation of ethion with bromine
    water to the active phosphorothiolate and subsequent measurement of
    the extent of inhibition of the cholinesterase from human blood plasma
    (Cook, 1954; Fallscheer and Cook, 1956).

    The clean-up procedure associated with the colorimetric method is so
    designed that determination of ethion in the presence of other organo-
    phosphorus insecticides can be made. However, the method does not
    measure oxidized or metabolized derivatives that contain phosphoryl
    rather than phosphorothiono groups. The enzyme method on the other
    hand measures the oxidized and unchanged ethion but does not
    differentiate from other cholinesterase inhibitors. The two methods
    are in substantial agreement however in the determination of ethion
    residues, which implies that there is little of the residue in the
    oxidized state (Niagara, 1958).

    A microcoulometric gas chromatographic method (Cassil, 1962) has been
    used to determine both ethion and its oxygen analogues independently
    (Niagara, 1967). The methods are sensitive to 0.05 ppm in most foods.

    For meat samples, following suitable extraction of macerated samples,
    partitioning between solvents and further clean-up on Florisil
    columns, the residue is determined by a gas chromatograph using a
    sodium thermionic detector. Recoveries of 92 per cent are normal with
    a sensitivity to 0.001 ppm. This precise method does not measure the
    oxidation products which contribute an insignificant amount to the
    residues (see above). With a different column the oxidation products
    can be measured but at a lower recovery rate (Niagara, 1966b).

    National tolerances

                                                                    
                                                         Tolerance
    Country                   Crop                         (ppm)
                                                                    

    Canada           Raisins                                 4

                     Citrus, grapes, peaches, plums,
                     strawberries, nectarines, beans,
                     melons, tomatoes                       2

                     Apples, pears, eggplant, onions,
                     peppers                                1

                     Summer squash                          0.5

                     Almonds                                0.1

    United States    Citrus pulps, dehydrated for
    of America       cattle feed                           10

                     Dried tea                              7

                     Almond hulls                           5

                     Raisins                                4

    (continued)
                                                                    
                                                         Tolerance
    Country                   Crop                         (ppm)
                                                                    

                     Beef (fat basis)                       2.5

                     Apples, citrus, grapes, plums,
                     strawberries, beans, melons,
                     tomatoes, sorghum grain                2

                     Pears, peaches, nectarines,
                     eggplant, onions, peppers              1

                     Beef or beef products                  0.75

                     Cucumbers, summer squash               0.5

                     Almonds                                0.1

                     Milk                                  "0"
                                                                    

    RECOMMENDATIONS FOR TOLERANCES AND PRACTICAL RESIDUE LIMITS

    Appraisal

    Ethion is an insecticide and acaricide for use on both plants and
    animals. The technical product contains a trace of an active oxidized
    metabolite but it is not of significance in the final residue. Ethion
    is used as a pre-harvest application to a variety of food crops
    especially citrus and deciduous fruit for the control of aphids, scale
    insects, mites, leaf miners and leaf hoppers. It shows minor systemic
    activity and residues are reduced also by weathering. No residues of
    ethion were found in milk or meat from lactating cows fed citrus pulp
    containing ethion residues. However, where ethion was applied as a dip
    to animals for the control of ticks, after a three-day interval
    following treatment, the highest level was in the fat at 1.25 ppm with
    less than 0.01 ppm in internal organs and muscle. The two active
    oxidation metabolites were present in insignificant quantity.
    Accordingly, animals dipped in ethion emulsion should be held three
    days prior to slaughter to ensure that the recommended tolerances,
    applied at slaughter, are not exceeded.

    In market sample studies in 1966-67, residues were found at 0.024 to
    0.054 ppm in three out of 30 composite samples of fruit in the United
    States of America. The same survey on oils, fats and shortening showed
    residues of 0.025 ppm in one out of 30 composite samples. Methods are
    available for measuring residues for use in general regulatory
    laboratories but further work is necessary to specify a method for
    referee purposes.

    Recommendations

    The following temporary tolerances (to be in effect until 1972) are to
    apply to raw agricultural products moving in commerce unless otherwise
    indicated. In the case of fruit-and vegetables the tolerances should
    be applied as soon as practicable after harvest and in any event prior
    to actual retail to the public. In the case of commodities entering
    international trade, the tolerances should be applied by the importing
    country at the point of entry or as soon as practicable thereafter.

    Temporary tolerances

         Grapes                                       2 ppm
         Other fruit                                  1 ppm
         Vegetables                                   0.5 ppm
         Tea (from a particular estate)
              for blending only                       7 ppm
         Tea, blended                                 1 ppm
         Beef (fat basis) (at slaughter)              1.5 ppm

    Further work or information

    ETHION

    Required before 30 June 1972:

    1. Data from countries other than the United States of America on the
    required rates and frequencies of application, pre harvest intervals,
    and the resultant residues.

    2. Data on residue levels in raw agricultural commodities moving in
    commerce.

    3. Residue data in processed food, including meat, meat products and
    wine.

    Desirable:

    1. Collaborative studies to establish a referee method.

    2. Adequate observations of effects in man, including studies of the
       metabolic fate.

    3. Determination of the metabolic fate in animals.

    4. Long-term studies in at least two species.

    5. Cholinesterase depression studies at more frequent intervals in
       animals.

    REFERENCES

    Cook, J. W. (1954) Report on determination of insecticides by
    enzymatic methods. J. Assoc. Off. Agr. Chem., 37: 561-564

    Cassil, C. C. (1962) Pesticide residue analysis by microcoulometric
    gas chromatography. Residue Rev., 1: 37-65

    Duggan, R. E. and Weatherwax, J. R. (1967) Dietary intake of pesticide 
    chemicals. Science, 157: 1006-1010

    Fahey, J. E., Rodriquez, J. G., Rusk, H. W. and Chaplain, C. E. (1962) 
    Chemical evaluation of pesticide residues on strawberries. J. Econ.
    Entomol., 55: 179-184

    Fallscheer, N. O. and Cook, J. W. (1956) Report on enzymatic methods 
    for insecticides. Studies on the conversion of some thionophosphates
    and a dithiophosphate to in vitro cholinesterase inhibitors. J.
    Assoc. Off. Agr. Chem., 39: 691-697

    Graham, J. R. (1964) Vol. II, Anal. Methods for Pesticides, ed. G.
    Zweig,  Academic Press

    Hazleton Laboratories, Inc. (1958a) Nialate Tech. (1240) Final report.
    In vitro cholinesterase studies. Unpublished report

    Hazleton Laboratories, Inc. (1958b) Nialate Tech. (1240) Final report.
    Subacute feeding studies - rats. Unpublished report

    Hazleton Laboratories, Inc. (1958c) Nialate Tech. (1240) Final report. 
    Subacute administration - dogs. Unpublished report

    Hazleton Laboratories, Inc. (1958d) Nialate Techn. Potentiation study. 
    Acute oral administration - rats. Unpublished report

    Hazleton Laboratories, Inc. (1959) Nialate Technical (ethion) Final
    report. Ninety-day feeding study - rats. Unpublished report

    Hazleton Laboratories, Inc., (1960) Palo Alto, Calif. Unpublished
    report

    Hazleton Laboratories, Inc. (1961) Ethion. Acute oral administration
    - rats. Unpublished report

    Industrial Bio-Test Laboratories, Inc. (1961a) Effects of ethion on
    cholinesterase activity in the dog. Unpublished report

    Industrial Bio-Test Laboratories, Inc. (1961b) Addendum report.
    Effects of ethion on cholinesterase activity in the dog. Unpublished
    report

    Industrial Bio-Test Laboratories, Inc. (1965a) Acute oral toxicity of 
    ethion MR E423. Unpublished report

    Industrial Bio-Test Laboratories, Inc. (1965b) Three-generation 
    reproduction study in albino rats on ethion. First generation, second
    generation and final report. Unpublished reports

    Martin, R. J. and Duggan, R. E. (1968) Pesticide residues in total 
    diet samples (III). Pesticides Monitoring Journal, 1: 11-20

    May and Baker, Ltd. (1960) Ethion. Unpublished report submitted to 
    the Ministry of Health, United Kingdom

    Niagara. (1958) Unpublished report M-602. Niagara Chemicals Division,
    F.M.C. Corpn., Middleport, N.Y.

    Niagara. (1960) Unpublished report M-796. Niagara Chemicals Division,  
    F.M.C. Corpn., Middleport, N.Y.

    Niagara. (1966a) Petition No. 351, Niagara Chemicals Division, F.M.C.
    to U.S. Food and Drug Administration

    Niagara Chemicals Division, F.M.C. Corpn., Middleport, N.Y. (1966b)  
    Ethion food additive petition. Report No. C.T.B. 23/2/20

    Niagara. (1967) Unpublished report M-2131. Niagara Chemicals Division, 
    F.M.C. Corpn., Middleport, N.Y.

    Taschenberg, E. F., Avens, A. W., Parsons, G. M. and Gibbs. S. D.
    (1963)  Disappearance of spray deposits of DDT, methoxychlor,
    perthane, ethion and diazinon from Concord grapes. J. Econ. Entomol.,
    56: 431-438
    


    See Also:
       Toxicological Abbreviations
       Ethion (ICSC)
       Ethion (FAO/PL:1969/M/17/1)
       Ethion (AGP:1970/M/12/1)
       Ethion (WHO Pesticide Residues Series 2)
       Ethion (WHO Pesticide Residues Series 5)
       Ethion (Pesticide residues in food: 1982 evaluations)
       Ethion (Pesticide residues in food: 1983 evaluations)
       Ethion (Pesticide residues in food: 1986 evaluations Part II Toxicology)
       Ethion (Pesticide residues in food: 1990 evaluations Toxicology)