WHO/FOOD ADD/71.42



    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
    Group on Pesticide Residues, which met in Rome, 9-16 November, 1970.



    Rome, 1971

    (other than mancozeb)


    The dithiocarbamate fungicides: ferbam, mancozeb, maneb, nabam,
    thiram, zineb and ziram were evaluated at the Joint FAO/WHO Meeting in
    1967. Although the biochemical data were limited, temporary acceptable
    daily intakes (ADI's) were established for all of these compounds, but
    it was pointed out that these ADI's are to be applicable to the parent
    compounds only (FAO/WHO, 1968). The following monograph addendum
    collectively summarizes data that have become available since that
    time. It is recognized that, in spite of their chemical similarity,
    the biochemical and toxicological properties of these fungicides vary
    considerably. However, the availability of new data did not justify
    producing a separate monograph for each compound. In the cast of
    mancozeb, however, important new information comparing the metabolism
    in animals with that in plants had become available, and this compound
    is the subject of a separate monograph.




    The dithiocarbamate compounds are thought to breakdown to
    alkylthioureas, carbon disulfide and hydrogen sulfide. The scheme
    shown in Figure 1 has been suggested for the breakdown of nabam (Falk
    et al., 1965).

    Breakdown of nabam, as well an zineb and maneb, in aqueous systems has
    been demonstrated to product ethylene thiourea and ethylene thiuram
    monosulfide (Vonk and Kaars Sijpesteijn, 1970).

    With maneb, zineb and nabam, ethylene thiourea and ethylenediamine
    were found in all cases, often as the end products of breakdown.
    Levels of the intermediates, such as ethylene-bis thiuram
    monosulfide and ethylene di-isothiocyanate, varied according to which
    compound was studied (Engst and Schnaak, 1970).

    Further work on the metabolism of various dithiocarbamate fungicides
    is reported to be in progress in The Netherlands (Verschuuren, 1970).

    Effect on enzymes and other biochemical parameters

    The effect on inhalation of maneb and zineb on five isoenzymes of
    lactic dehydrogenase in rat testes was studied. With maneb-poisoned
    animals, an increase in the isoenzyme related to aerobic metabolism
    and a decrease in that related to anaerobic metabolism was observed.
    There was no effect on those isoenzymes in the case of animals exposed
    to zineb (Izmirova et al., 1969).


    Information on the effect of zineb or maneb on various enzyme systems
    in the rat is reported under "Short-term studies" (Bankowska et al.,


    Special studies on carcinogenicity


    Groups of 18 mice of each sex from two hybrid strains were given
    various dithiocarbamate fungicides from seven days of age for 18
    months. The compounds were given daily by gavage from day seven of age
    until weaning; thereafter, the compounds were added to the diet in
    such a concentration that the animals received the corresponding
    amount. The compounds and the respective amounts administered were:
    ferbam, 10 mg/kg body-weight then 32 ppm in the diet; maneb, 46.4
    mg/kg, then 158 ppm; nabam 21.5 mg/kg then 73 ppm; thiram, 10 mg/kg
    then 26 ppm and zineb, 464 mg/kg then 1298 ppm. In all cases there was
    no significant increase in tumours compared with a control group of
    mice. However, when ethylene thiourea (a metabolite of some
    dithiocarbamates) was administered at 215 mg/kg body-weight and then
    after weaning incorporated into the diet at 646 ppm, the total
    incidence of tumours was 14 out of 16 for the males and 18 out of 18
    for the females in one strain; and 18 out of 18 and 12 out of 16,
    respectively, for the males and females in the other strain (Innes et
    al., 1969),

    Technical ziram (97.6 percent) was administered orally to mice
    (strains C57B1 and A) in doses 0 or 75 mg/kg body-weight twice a week
    over a period of 2.5 months. This regimen comprises a total of 20
    doses equivalent to 1500 mg/kg body-weight. A positive control group
    was exposed in the same way to urethane to enable comparison of
    results. The occurrence of adenomas in lungs and livers was studied
    after the end of the exposure at intervals of 1.5 months during six
    months. Adenomas were ascertained in the lungs only. The occurrence
    was: in strain A in the ziram group 51.2 percent, in the urethane
    group 100 percent and in the negative control group 42.6 percent. The
    differences are not statistically significant. The first adenomas were
    apparent after three months. In strain C57 in the ziram group, 7.4
    percent adenomas were found, in the urethane group 25.6 percent and in
    the negative control group 0 percent. The results are statistically
    significant according to the T test (T=2.08; P=0.05) but not according
    to the chi squared test (chi2= 2.18; P=0.15). The occurrence of
    adenomas was ascertained after six months. On the basis of the results
    of the experiments, ziram is considered a weak blastogen (Khicenko and
    Chernov, 1968).

    Mouse and rat

    Zineb, ziram, maneb and urethane have been administered orally once a
    week during six weeks to mice of strains A and C57 and to rats (SPF).
    The total dose of single substances in the rats (200-250 g in weight)

    was 120 mg (ca. 600 mg/kg body-weight/6 weeks), in mice (19-22 g) 60
    mg (ca. 3000 mg/kg/6/weeks), with the exception of ziram, where the
    total dose represented only half this amount i.e. 30 mg (ca. 1500
    mg/kg-6 weeks). Comparison was made between the activity of
    glucoso-6-phosphate dehydrogenase in liver and deoxyribonuclease in
    serum under development of proliferative changes in the lungs. With
    regard to zineb, ziram and urethane, the correlation was ascertained
    in rate only between the early blastogenic changes in lungs and the
    deoxyribonuclease activity in the serum. In case of the weak
    blastogens, zineb and ziram, maximum increase of ferment activity was
    apparent in the ninth month and lung changes in the 12th month after
    the exposure. In the urethane group, these reactions in rats were
    observed earlier (4,5 or 6 months and 9 months, respectively). Neither
    the activity of glucoso-6-phosphate dehydrogenase in the liver of mice
    and rats nor that of deoxyribonuclease in serum changed, and there was
    no correlation with proliferative changes in the lungs (Chepinoga et
    al., 1969).

    Special studies on mutagenicity

    The mutagenic activity of zineb and ziram is considered to be not
    significant using the Muller-5 test in Drosophila melanogaster, when
    compared with the spontaneous mutation rate of 0.14 percent (Benes and
    Sram, 1969). See also "Observations in Man".

    Special studies on reproduction

    Chick embryo

    The toxicity of the dithiocarbamate fungicides, ferbam, nabam, thiram
    and ziram, was determined for the chick embryo. It was determined that
    in order to avoid mortality from the solvent, propylene glycol, the
    injections must be made at a very early stage in the development of
    the embryo. The compounds were injected into the air chamber of
    fertile eggs prior to incubation. On days four and six, the eggs were
    candled to determine in which the embryos had died. The LD50 values
    in mg/egg were for: ferbam, 0.0022; nabam, 0.14; thiram, 0.0019 and
    ziram, 0.0021. The LD50 of thiram when injected into the yolk sac was
    much higher than when injected into the air chamber, namely 0.018
    mg/egg. The toxicity of thiram and ziram was reduced by simultaneous
    injection of cysteine. It was suggested that there was a reduction of
    these dithiocarbamates to less toxic materials (Gebhart and van
    Logten, 1968).


    Groups, each comprising 10 male and 10 female rats were given, twice
    weekly, oral doses of 0, 700 or 1400 mg/kg body-weight of maneb for
    four and a half months. (This dose regimen is equivalent to a daily
    intake of 0, 200 or 400 mg/kg body-weight.) No clinical abnormalities
    in the parent generation were observed. There were six births in the
    group given the lower dose. There was a moderately high incidence of

    stillbirths and imperfect skull development, but the actual numbers
    compared with the controls is not stated (Kaloyanova et al., 1967).

    The effect of the fungicide maneb on the embryonal development and the
    generative function of rate was studied. For studying the potential
    embryotoxic effect of maneb, mature white rat females with regular
    ovulation were used. During the whole pregnancy, these female rats
    were administered maneb by gavage every second day (as a suspension in
    milk) at a dose level of 50 mg/kg body-weight (0.001 LD50). An
    embryotoxic effect of the preparation was ascertained, i.e. abnormal
    development of the embryos (dead embryos and resorption), stillborn
    young and those incapable of living. In 21 percent of the pregnancies,
    the unfavourable effect of maneb on the course of the pregnancy was
    apparent when compared with 12 percent in the controls (Marcon, 1969).

    Immature female and male rats, of body-weight 80-100 g, were exposed
    to maneb for a month, the dose level being 50 mg/kg body-weight. When
    mature (after two and one half months from the beginning of the
    exposure), the control males were mated with the exposed females and
    vice versa. A decline of fertility in both sexes was observed. This
    effect, however, was not permanent, the function of the gonads being
    restored in three and one half months (Marcon, 1969).


    Groups of from nine to 14 pregnant female hamsters received an oral
    dose of 0, 31, 63, 125 or 500 mg/kg bodyweight of thiram in
    dimethylsulfoxide solution on day seven or eight of gestation. No
    litters were produced from the group given 500 mg/kg, and the
    mortality of the parents was 60 percent. Incidence of deformed young
    was significantly greater for the animals given 125 or 250 mg/kg of
    thiram compared to the controls given dimethylsulfoxide alone.
    Abnormalities included fused ribs, deformed tails and head defects
    including all degrees of exencephaly. A similar study using
    carboxymethyl cellulose as a solvent at 0, 125, 250, 300 or 500 mg/kg
    body-weight of thiram produced a lower incidence of malformation both
    in the treated and control animals, although the percentage incidence
    of terata was increased in the 250 mg/kg groups and higher compared to
    the controls (Robens, 1969).

    Special studies on the effects on thyroid function


    Groups, each comprising 10 rats of unspecified sex, received orally 0
    or 3 500 mg/kg body-weight of maneb or 2 400 mg/kg of zineb presumably
    as a single dose. After a period of 24 hours had elapsed following the
    administration of the compounds, the animals were injected
    intraperitoneally with 1 mCi of carrier free 131I. The animals that
    received zineb accumulated nine times less, and those given maneb 4.5
    times less 131I the control group. This goitrogenic effect of these

    dithiocarbamates was considered to be related to their metabolites
    which are derivatives of thiourea (Ivanova et al., 1967). See also
    "Observations in man".

    Acute toxicity

                                             LD50            Reference
    Compound     Animal            Route     mg/kg

    Zineb        mouse (M)         i.p.      2 400           Lessel and
                                             (approx.)       Cliffe, 1961

    Zineb        rat (M)           s.c.      >5 600          Lessel and
                                                             Cliffe, 1961

    Zineb        guinea-pig (F)    oral      >4 800          Lessel and
                                                             Cliffe, 1961
    Short-term studies


    Groups of 10 male and 10 female newly-weaned rats were given 0, 15,
    60, 250 or 1000 mg/kg body-weight of zineb by gavage for five days a
    week over a four week period. Blood samples were taken during the
    final week of feeding. Some of the animals were sacrificed for autopsy
    immediately upon termination of the period of administration of zineb;
    others were kept for two weeks without receiving zineb prior to
    sacrifice. No effects attributable to zineb were detected at levels of
    250 mg/kg or lower. At 1000 mg/kg the kidneys were enlarged in the
    animals of both sexes, but no histological changes were apparent in
    that organ. The thyroids were not enlarged, although histological
    examination indicated a slight hyperplasia in the females given 1000
    mg/kg. Weight gain in the animals was normal, as was the blood
    picture. There were no histological changes in any tissue other than
    in the thyroid. One female animal given 1000 mg/kg died of an unknown
    cause. Withdrawal of zineb appeared to result in a reversal of the
    effect on the kidney and thyroid, as evidenced by examination of the
    animals sacrificed two weeks after discontinuing treatment (Lessel and
    Cliffe, 1961).

    Male rats were maintained for six weeks on diets containing 0, 500 or
    5000 ppm of either maneb or zineb. Gross and histopathological changes
    in the thyroid gland, reduced assimilation of 124I and slightly
    reduced respiratory activity of the liver mitochondria at 5000 ppm
    were observed in the test groups. No significant alterations were
    observed with respect to the following parameters: mitochondrial

    cytochrome oxidase; glucose-6-phosphate dehydrogenase in the
    erythrocyte and liver homogenates; contents of cytochromes a3, b and
    c; content of flavoproteides in the mitochondria and content of
    oxidized and reduced nicotineamidoadenine in liver and kidney
    homogenates (Bankowska et al., 1970).

    Long-term studies


    Studies are reported to be underway in the Netherlands with rats.
    Preliminary results indicated that certain dithiocarbamate fungicides
    may affect the reticuloendothelial and haematopoietic systems as
    evidenced by the occurrence of slight anaemia, as well as effects on
    the thymus, mesenteric lymph nodes and thyroid function. Details are
    not yet available (Verschuuren, 1970).


    Mutagenic activity of ziram was studied in nine workers (four men and
    five women) exposed for three to five years to a concentration of 1.95
    - 3.7 mg/m3 air using the test of chromosome aberrations in
    peripheral leucocytes. The control group consisted of four humans
    (three women and one man). The number of aberrated cells in the group
    exposed to ziram was 5.9 percent and in the control group 0.75 percent
    (Pilinskaya, 1970).

    The damage of thyroid gland function has been observed not only in
    animals but in humans, too. Having examined 25 workers coming into
    contact with thiram, Ryznkova and Smirnova found a symptomology of
    hyperthroidosis in nine workers from this group; surgical treatment
    was necessary in three cases (Korablev, 1969).


    Since the last evaluation of the dithiocarbamate fungicides considered
    in this monograph, adequate information on the biotransformation in
    plants has not been forthcoming. Such information is needed to
    determine if the main metabolites are the same in animals as in
    plants. The Meeting was informed that a considerable amount of work on
    a number of dithiocarbamates is now in progress in The Netherlands.
    This work involves short and long term studies and studies on
    mechanisms of action and metabolism. Preliminary information from this
    work indicates the existence of some effects on the
    reticuloendothelial and haematopoietic system, as evidenced by a
    slight anaemia as well as changes in the thymus, mesenteric lymph
    nodes and thyroid function. Work reported from eastern Europe
    indicates that certain dithiocarbamate fungicides affect reproductive
    physiology, may have a carcinogenic potential and disturb the thyroid

    For these reasons it was considered that the temporary acceptable
    daily intakes should remain at the same figures as previously
    established (and be applicable to the parent compounds only), pending
    consideration of the results of the studies in progress.


    Levels causing no toxicological effects


          Rat:  250 ppm in the diet, equivalent to 12.5 mg/kg

          Dog:  5 mg/kg body-weight


          Rat:  250 ppm in the diet, equivalent to 12.5 mg/kg


          No long-term studies reported. See FAO/WHO, 1968


          Rat:  48 ppm in the diet, approximately equivalent to 2.5 mg/kg

          Dog: 5 mg/kg body-weight/day


          Not found in the long term study previously reported. See
          FAO/WHO, 1968


          Rat:  250 ppm in the diet, equivalent to 12.5 mg/kg

          Dog: 5 mg/kg body-weight/day


    0 - 0.025 mg/kg body-weight, applicable to the parent compounds only,
    and to the sum of all the dithiocarbamate fungicides if more than one
    is present.



    In the study of some ethylene-bis-dithiocarbamates, rapid
    decomposition of nabam, maneb and zineb in an aqueous environment,
    mainly to ethylene thiourea and ethylene thiram monosulfide, was
    confirmed (Vonk and Kaars Sijpesteijn, 1970). But only ethylene
    thiourea was taken up readily in the plant, being converted to an
    unidentified product to some extent: no ethylene thiram monosulfide
    was found in the plant.


    Currently, the most commonly used method is still the nonspecific one
    involving the determination of carbon disulphide evolved on treatment
    with acid. Modifications to improve recoveries have been made by
    various workers (McLeod and McCully, 1969; Rangaswamy, Poornima and
    Majumder, 1970).


    REQUIRED (by June 1973)

          1. Elucidation of the effect on the reticuloendothelial and
             haematopoietic systems.

          2. Further clarification of the possible carcinogenic effect of
             these compounds

          3. Elucidation of the effect on reproductive physiology

          4. Elucidation of the effect on thyroid function

    REQUIRED (before tolerances can be recommended)

    Studies on the biotransformation of the compound in plants to
    determine the chemical nature of residues, and appropriate
    toxicological studies on these residues.


    Bankowska, J., Bojanowska, A., Komorowska-Malewska, W., Krawczynski,
    K., Majle, T., Syrowatka, T. and Wiadrowska, B. (1970). The studies on
    the influence of zineb and maneb on the functional state of the
    thyroid gland and some related enzymatic systems (in Polish).
    Roczn.Zak.Hig. (Warsz.), 21 (2): 117-127

    Benes., V. and Sram, R. (1969) Mutagenic activity of some pesticides
    in Drosophila melanogaster. Industr.Med., 38 (12): 50-52

    Chepinoga, O.P., Zastavnyuk, N.P. and Zadorozhnaya, N.A. (1969).
    Blastogenic hazard of some derivatives of the dithiocarbamic acid and
    approaches to the detection (in Russian). Gigiyena primeneniya,
    toksikologiya pesticidov i klinika otravleniy (Kiev), 7: 153-166

    Engst, R. and Schnaak, W. (1970) Investigations of the metabolism of
    the fungicidal ethylene-bis-dithiocarbamates maneb and zineb. III
    The cause of the reaction of degradation. Z. Lebensm.
    Untersuch.-Forsch., 143: 99-103

    Falk, H.L., Thompson, S.J. and Kotin, P. (1965) Carcinogenic potential
    of pesticides. Arch. environm. Hlth., 10: 847-858

    FAO/WHO (1968) 1967 Evaluations of some pesticide residues in food
    FAO/PL: 1967/M/11/1; WHO/Food Add./68.30

    Gebhardt, D.O.E. and van Logten, M.J. (1968). The chick embryo test as
    used in the study of the toxicity of certain dithiocarbamates.
    Toxicol. appl. pharmacol., 13: 316-324

    Innes, J.R.M., Ulland, B.B., Valerio, M.G., Petrucelli, L., Fishbein,
    L., Hart, E.R., Pallotta, A.J., Bates, R.R., Falk, H.C., Gart, J.J.,
    Klein, M., Mitchell, I. and Peter, J. (1969) Bioassay of pesticides
    and industrial chemicals for tumorigenicity in mice: a preliminary
    note. J.nat.Cancer Inst., 42: 1101-1114

    Ivanova, L., Sheytanov, M. and Mosheva-Ismirova, N. (1967) Changes in
    the functional state of the thyroid gland upon acute intoxication with
    certain dithiocarbamates - zineb and maneb. C.R.Aced.bulg.Sci., 20
    (9): 1011-1013

    Izmirova, N., Izmirov, I. and Ivanova, L. (1969) The effect of zineb
    and maneb on the isoenzymes of lactic dehydrogenase in the testes of
    rats. C.R.Acad. bulg.Sci., 22 (2): 225-227

    Kaloyanava, F., Ivanova, L. and Alexiev, B. (1967) The influence of
    large doses of maneb on the progeny of albino rats. C.R.Acad.bulg.
    Sci., 20 (10): 1109-1112

    Khicenko, I.I. and Chernov, O.V. (1968) Experimental study of
    potential blastogenic effect of ziram (in Russian). Gigiyena
    primeneniya, toksikologya pesticidov i klinika otravleniy (Kiev),6:

    Korablev, M.V. (1969) Toxicological characteristics of dithiocarbamate
    acid derivative employed in the national economy and medicine
    (Literature survey) (in Russian). Farmakol. i Toksicol., 32 (3):

    Lessel, B. and Cliffe, E.E. (1961) The mammalian toxicity of metiram.
    Short term (4-week) oral toxicity. Unpublished report, submitted by
    Boots Pure Drug Co. Ltd.

    McLeod, H.A. and McCully, K.A. (1969) Head space procedure for
    screening food samples for dithiocarbamate residues. J.Assn.Off.
    Anal.Chem.,52: 1226

    Marcon, L.V. (1969) The effect of maneb on the embryonal development
    of generative function of rats (in Russian). Farmakol. i Toksikol.,
    32 (6): 731-732

    Pilinskaya, M.A. (1970) Chromosome aberrations in the persons
    contacted with ziram (in Russian). Genetika,6 (7): 157-163

    Rangaswamy, J.R., Poornima, P. and Majumder, S.K. (1970) Rapid
    colorimetric method for estimation of thiram residues in grain; also
    ferbam and ziram. J.Assn.Off.Anal.Chem., 53: 519-522, 1043-1044

    Robens, J.F. (1969) Teratologic studies of carbaryl, diazinon, norea,
    disulfiram and thiram in small laboratory animals. Toxicol.appl.
    Pharmacol., 15: 152-163

    Verschuuren, H.G. (1970) Information provided verbally to the 1970
    Joint FAO/WHO Meeting on Pesticide Residues on work being conducted on
    dithiocarbamate fungicides in The Netherlands

    Vonk, J.W. and Kaars Sijpesteijn, A., (1970) Fate in plants of
    ethylene-bis-dithiocarbamate fungicides and their decomposition
    products. Ann.appl.Biol., 65: 489-496

    See Also:
       Toxicological Abbreviations
       Dithiocarbamate fungicides (Pesticide residues in food: 1977 evaluations)