CHLORDIMEFORM        JMPR 1978

    Explanation

         Chlordimeform was evaluated in 1971 and 1975 (FAO/WHO 1972b,
    1976b) and a temporary ADI was established. Temporary MRLs were
    recommended for a number of foods and five requirements for further
    information were listed.

         In 1976 the manufacturer temporarily suspended, the sale of the
    pesticide. This action was noted by the 1976 and 1977 Joint Meetings
    (FAO/WHO 1977a, 1978a).

         Having completed a number of long-term toxicological, metabolism
    and residue studies, in 1978 the manufacturers approached authorities
    in a number of countries to allow limited commercial use in cotton
    crops. The results of these studies were evaluated by the meeting.

    EVALUATION FOR ACCEPTABLE DAILY INTAKE

    BIOCHEMICAL ASPECTS

    Effects on Enzymes and other biochemical parameters

         Chlordimeform and its major metabolites were tested for their
    ability to affect DNA, RNA and protein synthesis. In vitro studies
    using HeLa cells exposed to a variety of concentrations of
    chlordimeform and its major metabolites showed that the synthesis of
    macromolecules were virtually unaffected except at extremely high
    concentrations at which RNA synthesis was reduced (Murakami and
    Fukami, 1974).

         Several studies have been reported to define the mode of action
    of chlordimeform. Chlordimeform has shown a complex mechanism of
    action eliciting a variety of unusual biochemical and pharmacological
    responses. Chlordimeform has shown little activity on cholinergic
    transmission although in frog muscle preparations a decrease in
    acetylcholine reception sensitivity has been observed. Chlordimeform
    has been shown to be an effective uncoupler of oxidative
    phosphorylation and inhibitor of electron transport. Physiologically,
    chlordimeform acting as a direct depressant on cardiac and vascular
    muscle, induced a hypotensive state in dogs. Chlordimeform did not
    interfere with the autonomic nervous system. The mechanism of
    cardiovascular depression may be related to that noted with frog nerve
    preparations treated with procaine, a local anesthetic.
    4-chloro-o-toluidine has been shown to interfere with rat cardiac
    receptors (Lund et., al., 1978; Chinn et al., 1976; Wang et al., 1975;
    Watanabe et al., 1975; Matsumura and Beeman, 1976; Knowles, 1976;
    Hollingworth, 1976).

         It has been suggested that the increase in biogenetic amines
    resulting from inhibition of monoamine oxidase (MAO) activity by
    chlordimeform and its metabolites may be a factor in expressing the
    complex mode of action. Matsumura and Beeman (1976) have shown that
    rats treated with chlordimeform accumulated seratonin and
    norepinephrin in the brain while Benezet and Knowles (1978) showed
    dopamine to also accumulate. While, as had been suggested in the past
    that MAO inhibition is not the primary mechanism in chlordimeform
    insecticidal activity, inhibition of this process in concert with
    several other biological events has provided some data that may
    resolve the toxic mode of action in mammals. In further studies
    towards this goal, Knowles and his coworkers have continued to examine
    the metabolic fate of chlordimeform. They have also investigated the
    physiological significance and toxic action of the metabolic product.
    These workers have proposed that sequential oxidative and
    N-demethylation of chlordimeform results in the formation of two
    formamidine derivatives, the N-mono-desmethyl and the N-didesmethyl
    formamidines. These two metabolites appear to be more acutely toxic
    and more biologically active than chlordimeform. The onset of acute
    signs of poisoning was shown to be faster with these metabolites than
    with chlordimeform itself. Chlordimeform and the two formamidine
    metabolites have been shown to increase sleeping time in mice.
    Chlordimeform may indirectly influence chemically induced sleep or it
    may be an active sleep inducer especially through the action of some
    of its metabolites. Based on preliminary acute toxicity data, the two
    formamidine chlordimeform metabolites appear to be rapidly formed and
    may serve to contribute directly to the overall toxic action in rats.
    In contrast, two additional metabolites, the N-formyl-4-
    chloro-o-toluidine and the 4-chloro-o-toluidine were not as
    acutely toxic as chlordimeform but still appear to have contributed to
    the complex mode of action. The latter toluidine metabolites have been
    shown in vivo to induce a sedative and an anesthetic effect in
    addition to their biochemical ability to inhibit MAO activity. To
    define the multiple stress on animals treated with chlordimeform,
    Knowles and coworkers reported the acute poisoning response in rats of
    all of these metabolites following a variety of administrative routes.
    The signs of poisoning following acute Intoxication with chlordimeform
    and its two major formamidine metabolites included a marked
    hyperexcitability which increased with time and was accompanied by a
    high degree of sensitivity to external stimuli. Stimulation caused
    rapid running and elicited an intense escape behavior in rats. At the
    latter stages of poisoning, prostration and hyperextension of the hind
    legs occurred and death followed shortly. Signs of poisoning were
    observed to be the most rapid with the didesmethyl (the NH2
    derivative) formamidine, Hyperexcitability was evident almost
    immediately after poisoning and within five to six minutes convulsions
    were observed. In sharp contrast, rats treated with N-formyl and the
    4-chloro-o-toluidine metabolises responded rapidly as though they
    had been sedated. This condition was soon followed by one which
    resembled an anesthetic effect (Benezet et al., 1978).

         As mentioned above, the increase in biogenic amines resulting
    from inhibition of MAO activity undoubtedly has some physiological
    significance in defining the overall toxic response. Previous studies
    have shown that inhibition of rat MAO activity was relatively rapid
    and reversible following chlordimeform intoxication. In contrast to
    classical MAO inhibitory chlordimeform exhibited a readily reversible
    Inhibition (Maitre et al., 1978). Recovery was complete within two
    days in liver and four days in brain following an acute
    intraperitoneal dose of 100 mg/kg. In evaluating the sensitivity of
    the two types of MAO activity to inhibition with chlordimeform, the
    type B seemed to be more sensitive. Subacute, intraperitoneal
    administration of 10 mg/chlordimeform/kg body weight for four days did
    not appear to result in brain M0 inhibition and only marginally
    reduced liver MAO activity. This suggested the low cumulative
    potential for MAO inhibition at low or threshold doses. In contrast,
    Kaloyanova et al., (1978) observed substantial inhibition when rats
    were administered 5 mg/kg daily for 40 days. This inhibition may have
    resulted from an observed cumulative effect of prolonged
    administration and delayed recovery of MAO activity over the 40 day
    treatment interval. Thus, evidently the complex mechanism of action
    has not been fully elucidated. Direct diverse actions of chlordimeform
    and its metabolites and the indirect action of many biologically
    active amines all interacting simultaneously appear to preclude a
    rapid resolution of this complicated event.

    TOXICOLOGICAL STUDIES

    Mutagenicity

         In a series of bioassays, chlordimeform and its principle
    metabolites were examined for mutagenic effects on
    histidine-auxotrophic mutants of Salmonella either alone or after
    biological activation. Chlordimeform tested in a series of five tester
    strains (TA-98, TA-100, TA-1535, TA-1537 and TA-1538), was found to be
    negative at concentrations up to and including 20 mg/ml. The desmethyl
    metabolite of chlordimeform was slightly mutagenic when assayed
    against TA-1535. The Formyl-metabolite was negative when tested
    against the five tester strains although in one instance a weak
    response was elicited with TA-100.  In contrast, the 4-chloro-o-
    toluidine metabolite was found to elicit a positive mutagenic response
    at high concentrations with the tester strains TA-100 and TA-1535
    (Arni and Muller, 1976a, b and c; Konopka and Haymann, 1977a and b;
    Rashid, 1974). Chlordimeform did not elicit a mutagenic response when
    examined utilizing a streptomycin dependent E. coli-SD 4 tester
    strain (Hurni and Ohder 1970), A potential metabolite of chlordimeform
    formed in soil and in the presence of sunlight from the
    4-chloro-o-toluidine (4,4-dichloro-2,2-dimethylazobenzene) was found
    to be inactive when examined against two tester strains of Salmonella,
    TA-1535 and TA-1538 (Rashid, 1974).

    Mouse - Dominant Lethal

         Chlordimeform was administered to groups of 20 male mice at
    dosage levels of 0, 22 and 66 mg/kg/body weight. The males were mated
    with two untreated females weekly for six consecutive weeks in a
    standard dominant lethal assay. No evidence of dominant lethal effects
    was observed in the progeny of the male mice treated with
    chlordimeform (Fritz, 1978).

         Chlordimeform was bioassayed as a potential carcinogen using the
    C3H7OT 1/2 CLB mouse embryo fibroblast oncogenic transformation assay
    Nesnow and Heidelberger, 1976) and found to be negative over a range
    of 1 to 100 µg/ml. This assay has been shown to have a low rate for
    false positive responses suggesting that a positive result is highly
    likely to be an animal carcinogen (Nesnow, 1978).

         Groups of Chinese hamsters (3 males and 3 females per group) were
    administered chlordimeform by gavage on two consecutive days at dose
    levels of 0, 60, 120 and 240 mg/kg. A positive control using 
    cyclophosphamide (128 mg/kg) and a negative control of
    carboxymethylcellulose was used in an in vivo, mutagenesis assay
    evaluating bone marrow cells. In all dosage groups (sacrificed 24
    hours after the second dose) the percentage of cells displaying
    changes in nuclei did not differ from the negative controls. The
    positive controls showed a marked increase in the percentage of cells
    with anomalies of the nuclei (Langeuer and Muller, 1977).

    Teratogenicity

         Groups of rabbits (group size ranged from 17 to 38 dams per
    group) were administered chlordimeform orally from day 6 to day 18 of
    pregnancy at dose levels of 0, 10, 30 and 100 mg/kg/body weight. The
    administration of chlordimeform at 100 mg/kg/body weight produced a
    distinct adverse effect on dams in the first four days of treatment
    within four hours of administration of chlorphenamide. Examination of
    dams and fetuses (fetuses were removed by caesarian section on day 28
    of pregnancy) suggested that the low dose had no teratogenic or embryo
    toxic effect. In the high dose group, the number of incompletely
    ossified sternebrae showed a slight increase over that observed in the
    controls and in the other groups. In addition, the number of live
    fetuses with malformations was slightly increased. These malformations
    included a median cleft palate and exencephaly and an omphalocele.
    Further examination of spontaneous malformations observed in a 
    cumulative control of 2495 rabbit fetuses suggested that these
    abnormalities may be spontaneous and not as a consequence of the
    administration of chlordimeform (Fritz, 1971).

         Groups of rats (25 females/group, 30 females were used as
    controls) were administered chlordimeform in carboxymethylcellulose at
    dose levels of up to 0, 10, 25 and 50 mg/kg/day from six to day
    fifteen of pregnancy. Examination of fetuses removed by caesarian
    section on day 21 showed that a slight delay in growth of the fetuses
    had occurred at the two upper treatment levels. This effect was

    probably a direct result of a slightly toxic response in the dams as
    evidenced by reduced body weight gain and reduced food intake and
    somnolence noted on days 6, 7 and 8 of pregnancy. No teratogenic
    events were observed in the offspring although an increased incidence
    of eternal ossification defects occurred at 25 mg/kg (Fritz, 1975).

    Short Term Studies

         Monoamineoxidase (MAO) activity of liver, brain and serum was
    examined from rats treated with chlordimeform at dosages of 0, 5, 10
    and 50 mg/kg/day for 40 consecutive days. The MAO activity was
    determined with a variety of substrates. A dose-dependent inhibitory
    promotion was observed using several of the substrates representing
    different forms (isoenzymes) of MAO activity. The liver, brain and
    serum MAO activity was significantly inhibited at 5 mg/kg/day, the
    lowest dose level tested. A distinct, dose-response relationship was
    observed in both liver, brain and serum with two of the substrates
    used in the assay (Kaloyanova et al., 1978).

    Mice - 4-chloro-o-toluidine

         Groups of mice (30 males and 30 females/group, TIF.: NMRI strain)
    were bred and maintained under SPF conditions and fed a diet
    containing 4-chloro-o-toluidine at concentrations of 0, 750, 1500,
    3000 and 6000 ppm for 60 days. Mortality 50% was observed in the 6000
    ppm group. Food intake and growth were retarded at the two highest
    dose levels. No abnormal clinical signs of poisoning were observed.
    Eye examinations did not indicate adverse occular changes. A toxic
    Hemolytic anemia in both sexes of all treated groups was characterized
    by reticulocytosis and heinz body formation. In the male mice of all
    treated groups, hemoglobin concentration, packed cell volume and
    erythrocyte count was slightly below that of controls. In addition,
    Leukocytosis was observed in all animals of all dosage groups with the
    exception of female at the 750 ppm level. In both sexes at 6000 ppm
    and in the females at 3000 ppm total protein concentration was
    reduced, blood glucose and urea nitrogen values were increased. Plasma
    GPT was increased in male mice at 3000 ppm and in female mice at 1500
    ppm. Urinalysis revealed a reduced specific gravity in both sexes at
    3000 ppm and above and in females at 1500 ppm. Microscopic examination
    of tissues and organs at the conclusion of the studies showed slight
    to moderate vacuolar changes of Hepatocytes, which was pronounced in
    animals at the 3000 ppm levels and above. There was also a marked
    congestion of the spleen at these high dose levels. In addition, the
    urinary bladder revealed hyperemia and dilation of the capillaries in
    the mucosal layer. These changes were accompanied by edema, multiple
    intraepithelial hemorrhage and focal proliferation of the transitional
    cell epithelium. On occasion, these changes in the urinary bladder
    were noted at the lowest concentration (Suter et al., 1976b).

    Rat - 4-chloro-o-toluidine

         Groups of rats (30 males and 30 females per group Tif/Rai,
    strain) were maintained under SPF-barrier conditions and fed
    4-chloro-o-toluidine in the diet at concentrations of 0, 750, 1500,
    3000 and 6000 ppm for 60 days. There was no mortality over the course
    of the study and clinical signs of poisoning were not observed.
    Ophthamological examinations did not suggest changes related to the
    presence of 4-chloro-o-toluidine in the diet. Growth was reduced in
    the upper three dosed groups at dietary levels of 1500 ppm and above.
    Toxic hemolytic anemia in both sexes of all treated groups was
    characterized by a variety of hematological parameters including:
    reduced hemoglobin content, reduced hematocrit content, reduced blood
    cell count, increased methemoglobin content, heinz body formation,
    reticulocytosis and polychromatophilia. In the highest dose group, an
    increased number of immature red blood cells (normoblasts) were
    observed. An increased leukocyte count and prothrombin time was
    recorded at the 3000 and 6000 ppm level. Total protein was slightly
    reduced at 3000 and 6000 ppm and there was a shift in the globulin
    content as observed by electrophoresis. Plasma lambda-glutamyl
    transpeptidase of males and alkaline phosphatase of females was
    increased at 6000 ppm. Urinalysis was not significantly affected. In
    all treated animals, the liver showed an increase in size accompanied
    by hypertrophy of the hepatocytes. In the two highest dose groups, the
    spleen was enlarged and microscopic examination showed pronounced
    congestion and hemorrhage. In the highest dose group, slight or
    moderate proliferation of the transitional cell epithelium was noted
    in the urinary bladder (Suter et al., 1976b).

    Long Term Studies

    Mice - Chlordimeform Hydrochloride

         Groups of mice (50 males and 50 females per group, Tif: MAG
    strain, SPF derived) were fed chlordimeform in the diet at
    concentrations of 0, 20, 100 and 500 ppm for 24 months. At the
    conclusion of the dietary feeding interval animals were maintained on
    control diet until 90% of a group had died, at which time the
    remaining animals of the group were sacrificed. There were no acute
    signs of toxicity related to chlordimeform in the diet over the course
    of the feeding trial. Growth and food consumption were similarly
    unaffected by the presence of chlordimeform in the diet. Mortality was
    significantly increased in females after 60, and 90 weeks at 500 and
    100 ppm respectively. In males, significantly increased mortality was
    observed after 70 and 110 weeks at 500 and 100 ppm. However, lifespan
    was not significantly affected in males at 100 ppm. The animals fed
    dietary levels of 100 ppm and above displayed an increased incidence
    of hemorrhagic tissue masses in subcutaneous tissues, retroperitoneum
    and some internal organs (kidney, liver and spleen) which upon
    examination were classified as malignant haemangioenditheliomas. These
    malignancies which were reported to occur rarely in control
    populations mere found predominantly in the 100 and 500 ppm dietary
    groups. In some animals the tumors were of multiple origin and

    metastases were observed in the lungs. There were no other types of
    neoplasm observed in the study which were attributable to
    chlordimeform in the diet. Under the conditions of this study, 20 ppm
    in the diet appears to be a no effect level (Suter et al., 1978).

    Mice - N-formyl-4-chloro-o-toluidine

         Groups of mice (50 male and 50 female/group, Tif: MAG strain)
    were bred and maintained under standard SPF conditions and fed
    N-formyl-chloro-o-toluidine in the diet at concentrations of 0, 20,
    100 and 500 ppm for 24 months. Alter 24 months all animals were fed a
    control diet until the study was concluded when 90% of a group of
    animals had died.

         There was no sign of adverse behavior and acute mortality was not
    noted. Growth and food consumption were unaffected. There were
    significant differences noted in survival after one year of age. Both
    males and females showed an increased mortality after at 100 and 500
    ppm approximately one year of feeding. The onset of increased
    mortality occurred earlier in females. The females at the 20 ppm
    dietary level showed a slightly higher, non-significant mortality
    during the same period. Detailed gross and microscopic examination of
    a variety of tissues and organs showed the presence of numerous gross
    anatomical lesions. There was an increased number of hemorrhagic
    masses in the subcutaneous tissues in the retroperitoneum and in some
    internal organs of mice at all treatment levels. Detailed microscopic
    examination confirmed that the increased incidence of hemorrhagic
    masses were malignant tumors of vascular origin. These tumors were
    histologically classified as malignant haemangioenditheliomas. In
    addition to the occurrence of tumors, the time to tumor relationship
    was decreased as the dietary concentration was increased. Apart from a
    significantly higher incidence and earlier appearance of malignant
    vascular tumors noted at all dose levels, other neoplasms occurring in
    the study were not influenced by the dietary concentration; a
    non-effect level was not demonstrated under the conditions of this
    experiment (Sachsse at al., 1978a).

    Mice - 4-chloro-o-toluidine

         Groups of mice (30 males and 30 females per group, ICR-strain)
    were fed dietary concentrations of 4-chloro-o-toluidine for 80 weeks
    at dietary levels of 0, 20, 100 and 500 ppm. At the conclusion of the
    study, gross and microscopic examination of tissues and organs was
    performed. Data were not presented with respect to general
    observation, behavior changes, food consumption and growth. There
    appeared to be no significant changes with respect to organ and tissue
    weight in animals sacrificed at 80 weeks. The results of the
    histological examination of tissues and organs suggest that there was
    a dose-related increase in the number of tumors observed when data
    from both sexes are combined. These have been characterized as
    reticulum cell sarcoma. In addition, there was a dose-related increase
    in the overall tumor incidence at all treatment levels. The study
    showed a high incidence of spleenic effects and lung problems,

    indicative of probable infection in the colony. A no-effect level has
    not been demonstrated in the study based upon the dose-related
    increase in tumor incidence at all dietary levels tested.
    Unfortunately, a complete analysis of the study was  precluded in part
    due to the overall tumor incidence in control animals. The control
    tumor incidence ranged from 56 to 73% of the animals at eighty weeks.
    The data upon which statistical analyses were based utilized a
    modified life table (acturial) method. On the basin of the actual
    incidence of tumors, the dose/effect relationship is still apparent
    when use is made of total tumor incidence data (Ezumi and Nakao,
    1974b).

    Mice - 4-chloro-o-toluidine

         Groups of mice (50 males and 50 females/group, Tif: MAG strain)
    were derived and maintained under standard SPF-barrier conditions and
    fed a diet containing 4-chloro-o-toluidine at concentrations of 0,
    2, 20, 100 and 500 ppm for 24 months. After 24 months all animals were
    fed control diets until the study was concluded when 90% of a group of
    animals had died.  There were no overt toxicological signs of
    poisoning. Growth and food consumption were unaffected. An adverse
    effect on longevity (lifespan) was noted in both males and females at
    the two highest dietary levels. At the conclusion of the study upon
    gross examination there was a marked increase number of hemorrhagic
    masses in subcutaneous tissue, in the retroperitoneum and in some
    internal organs. Microscopic examination revealed an increased
    incidence of hemorrhagic malignant tumors of vascular origin at dosage
    levels of 20 ppm and above. The incidence in control exceeded the
    incidence observed at 2 ppm. The tumors were histologically classified
    as malignant haemangioenditheliomas and on occasion metastases were
    observed. There was not only a significant dose-dependent increase in
    the total incidence of malignant tumors but, the tumors occurred in
    animals at the higher concentrations at a markedly earlier date than
    those at the lower concentrations. (The time-to-tumor calculations
    were dose related). A benign variant of the haemangioma was observed
    in all groups. The occurrence of other types of neoplasms in the study
    was apparently not influenced by the presence of
    4-chloro-o-toluidine in the diet. Under the conditions of this
    experiment, 2 ppm in the diet appears to be a non-effect level
    (Sachsse et al., 1978b).

    Rat - 4-chloro-o-toluidine

         Groups of rats (30 males and 30 females/group, Sprague-Dawley
    strain) were fed a diet containing 4-chloro-o-toluidine at
    concentrations of 0, 20, 100 and 500 ppm for periods of 94 weeks for
    the males and 104 weeks for the females. There was a slight increase
    in mortality over the course of the study that appeared to be dose
    related when calculations of combined mortality data with both sexes
    were made. There was no indication of a reduced lifespan as a
    consequence of 4-chloro-o-toluidine in the diet. At the conclusion
    of the study there seemed to be an increased liver weight observed at

    500 ppm. A dose-related decrease in ovarian weight was also observed
    at all dose levels. There was a dose-related increase in tumors in the
    study. In the animals sacrificed at the conclusion of the study, liver
    nodules, described as tumor-like, were observed at 100 and 500 ppm but
    not in the controls and in the 20 ppm group. Upon microscopic
    examination these nodules were found to be non-malignant. In those
    animals that died during the course of the study there were a
    significant number of tumor-like nodules and swellings in the liver
    which were classified as benign or malignant hepatomas upon
    histological examination. On an overall evaluation of the study, it
    was concluded that liver tumors were found in all dose groups but not
    in controls. The total incidence of liver tumor was related to dietary
    dosage and found to be higher in females than in males. However,
    incidence of malignant tumors while increasing with dose was not sex
    related. Additionally, adenoma of the adrenal gland was found in the
    treated groups. A "no-effect level" was not seen in this study (Ezumi
    and Nakao, 1974a). As in the previous mouse study, the data
    calculations are based on the modified life table (actuarial) method.

         Two studies, each using both rats and mice in the bioassay, have
    been undertaken under the U.S. National Cancer Institute
    carcinogenicity program to evaluate the carcinogenic potential of
    4-chloro-o-toluidine. Full reports of these studies are not available.

    Mice

         In an initial study reported only in summary, groups of 25 male
    mice were fed 4-chloro-o-toluidine in the diet for 2 years at levels
    of 0, 750 and 1500 ppm. Similar groups of female mice were fed dietary
    concentrations of 0, 2000 and 4000 ppm. Further details on the
    parameters used in the study are unavailable. There appear to be a
    significant number of vascular tumors observed in both male and female
    mice. There were no differences reported between the treated and
    control animals with regard to other tumors.

         In a more well-defined program, reported as a draft report,
    groups of mice (50 males and 50 females per group, B6C3F1 strain) were
    fed dietary concentrations of 4-chloro-o-toluidine in a lifetime (92
    to 99 weeks) carcinogenic bioassay. The females were fed dietary
    levels of 0, 3750 and 15,000 ppm while the males were fed dietary
    levels of 0, 1250 and 5000 ppm. Body weight of treated animals was
    reduced. Mortality was not affected. A significant number of tumors
    designated as hemangiosarcoma of the epididimus, testes, urinary
    bladder, seminal vesicle and prostate were recorded in the high dosed
    male mice receiving 5000 ppm in the diet. A similar neoplastic
    condition existed in the reproductive organs of both high and
    low-dosed groups of female mice.

    Rat

         Groups of 25 male rats/group were fed dietary levels of 0, 2000
    and 4000 per four months at which time the dietary level was decreased

    to 500 and 1000 ppm for the remainder of the study. The rat study
    showed no significant differences between the treated and control
    animals.

         Groups of rat (50 males and 50 females per group, 20 males and 20
    females were used in controls, F344 strain) were fed dietary
    concentration of 4-chloro-o-toluidine at levels of 0, 1250 and 5000
    ppm. A significant number of rats survived the two-year feeding trial
    although growth was retarded at the high dose. Gross and microscopic
    pathological examinations of the animals revealed no tumours
    associated with the presence of 4-chloro-o-toluidine in the diet. It
    was concluded that 4-chloro-o-toluidine hydrochloride was not
    carcinogenic for F344 rats (Anonymous, 1978b).

    Rat - 2 Year dietary-lifetime studies - Interim reports

         A series of three one-year interim reports entitled "Lifetime
    Feeding Study in Rats" indicate that studies are being performed with
    chlordimeform, N-formyl-4-chloro-o-toluidine and with
    4-chloro-o-toluidine. Details of the studies are not defined. The
    reports, designated "interim reports", contain some brief information
    on hematology, blood chemistry, urinalysis urine electrophoresis and
    growth and food consumption data for a one year period (Anonymous,
    1978a, b and c).

         For chlordimeform, the data show a slight growth retardation at
    the highest dose level accompanied by a toxic hemalytic anemia.
    Methaemoglobin was increased at 4 weeks in a dose related response
    from 20 ppm. However, no effects were noted at 13, or 26 weeks, and
    minor changes were observed only at 500 ppm after 52 weeks. Total
    serum protein was reduced at 500 ppm. Heinz bodies, generally
    associated with methemoglobinemia were not observed in male rats but
    were observed in females. The dose levels used in these studies are 0,
    2, 20, 100 and 500 ppm in the diet with groups of 108 rats/sex/dose
    being used. There may have been an interim sacrificed but data are not
    reported.

         The data for the N-Formyl-metabolite appear to be similar to that
    described for chlordimeform. At the highest dose level, hemaglobin
    concentration was significantly reduced in both males and females. In
    addition, packed cell volume and red blood cell count were decreased
    and there was a significant reticulocytosis. There was an increased
    methemoglobin formation but heinz bodies were not observed. Blood
    chemistry, urine analysis and urine elcotrophoresis were not unusual.
    As with the previous study, growth was slightly retarded at the 500
    ppm dose level although food consumption did not appear to be
    affected.

         Preliminary data presented for the dietary 4-chloro-o-toluidine
    study show a reduction in weight gain in both sexes at the highest
    dietary concentration. This was again associated with a toxic
    hemolytic anemia. A marginal decrease was observed in the hemoglobin

    concentration in males and females at the highest dose level.
    Methemoglobin concentration at the end of one year was increased in
    females at the top three dose levels while with the males it was
    increased only at the top dose level. Again, heinz bodies were not
    noted in male rats but were noted in females. Against blood chemistry,
    urine analysis and urine electrophoresis was not affected.

         In all studies, there was a suggestion that an interim sacrifice
    was made. A statement was made that neither toxic symptoms nor gross
    pathology changes were found at this interim sacrifice time. The
    reports of these three studies are incomplete and the data could not
    be evaluated at the present time. The studies should be considered
    when full reports are available (Anonymous, 1978a, b and c).

    COMMENTS

         Chlordimeform was reviewed and evaluated for acceptable daily
    intake in 1971 and 1975 and a temporary ADI was allocated. The
    metabolic pathway in animals has been well-defined but no information
    is available on either the metabolic fate in man or the primary mode
    of toxic action.

         In studies performed subsequent to the initial reviews, adverse
    effects of long-term exposure were observed. Results of a preliminary
    report suggested a carcinogenic response in mice with
    4-chloro-o-toluidine, a major metabolite of chlordimeform. At that
    time voluntary withdrawal of all agricultural uses of chlordimeform
    was instituted by the major manufacturers to allow time to complete
    ongoing studies which would permit a complete evaluation of the
    hazards associated with continued agricultural use.

         An extensive series of short-term, high-level and long-term,
    low-level studies in both rats and mice, were reviewed and considered
    by the Meeting. Results of long-term studies demonstrate that
    chlordimeform and its principal metabolites, the N-formyl
    chlordimeform and 4-chloro-o-toluidine, are carcinogenic in the
    mouse, producing a dose-related malignancy histologically
    characterized as a haemangioendothelioma. In carcinogenicity studies
    with rats, results have been conflicting. Chlorimeform was not
    mutagenic to bacteria but its main metabolite 4-chloro-o-toluidine was
    mutagenic in the same bioassay systems. Because of these concerns, the
    temporary ADI was reduced.

         The Meeting was informed that there were currently uses of
    chlordimeform on cotton from which food residues have not been
    detected at the limit of the existing method of analysis. In
    consideration of this and the new data expected within one year, the
    Meeting recommended chlordimeform be reviewed in 1979.

    TOXICOLOGICAL EVALUATION

    Level causing no toxicological effect

         Rat: 100 ppm in the diet equivalent to 5 mg/kg bw

         Dog: 250 ppm in the diet equivalent to 6.25 mg/kg bw

    ESTIMATE OF ACCEPTABLE DAILY INTAKE FOR HUMANS

         0-0.0001 mg/kg body weight

    RESIDUES IN FOOD AND THEIR EVALUATION

    USE PATTERN

         The manufacturers were currently limiting their proposals to
    aerial applications to cotton crops under supervised conditions  which
    limit the uptake by operators, bystanders and consumers in Guatemala,
    Colombia, Nicaragua, Salvador and USA. Further such applications to
    cotton in Australia, Egypt, Mexico and Sudan were under consideration.

    Use pattern on cotton

         In 1972, chlordimeform was recommended in cotton at rates of 
    0.5 - 1.0 kg/ha. Further research revealed that useful control could 
    be achieved at rates of 0.125 to 0.25 kg/ha by killing eggs and early
    instar larvae and in 1973 these rates were recommended. Emulsifiable
    solutions of chlordimeform base, or solutions of the hydrochloride in
    Australia, USA and several other countries, are need on a 3 - 5 day
    schedule when moth flights begin or when eggs appear. In some other
    countries including Colombia and Guatamala it has been found necessary
    to use 1 kg/ha when mites are to be controlled. The compound is active
    again a narrower spectrum of insects than the organo-chlorine
    compounds and other compounds used on cotton and is being used
    successfully in integrated pest management programmes in the USA and
    other countries. Observations have shown that it does little harm to
    many beneficial insects and its removal in 1976 created a serious
    situation because acceptable alternatives were not available.

    RESIDUES RESULTING FROM SUPERVISED TRIALS

    Cotton

         Reports on supervised trials carried out between 1969 and 1977
    (Ciba-Giegy/Schering, 1978) and showing residues in cottonseed and
    cotton products processed from seed cotton grown under a range of
    treatment regimes were considered.

         The analytical methods used were based on the determination of
    4-chloro-o-toluidine which is a moiety common to chlordimeform and
    its intermediate metabolites as-well as being the principal metabolite
    in plants and animals (Morton, 1968; Nor-Am, 1969; Ciba-Geigy, 1976;
    Schering, 1973). The limits of determination were in the range 0.01 -
    0.05 mg/kg of chlordimeform equivalents with recoveries usually above
    80% (69-106%). A gas-chromatographic, method which allows separate
    determination of chlordimeform and its metabolites and has a limit of
    determination of 0.01 mg/kg and recoveries in the range of 74 - 106%
    was also used (Schering, 1975).

         Most determinations were concerned with residues in intact
    cottonseeds. The application rate ranged from 125 g to 3.6 kg/ha. From
    1 to 20 applications were made and the interval between the last
    application and harvest ranged from 0 to 89 days. Table 1 summarizes
    the results from the many trials reported Ciba-Geigy/Schering, 1978).

         There was no direct relationship between the residue levels and
    the experimental conditions such as application rate, number of
    applications, length of waiting period etc. However, the results
    suggest that the application rate had the strongest influence on the
    residue level, followed by the interval between last application and
    harvest. Tables 2 and 3 summarize the results of typical trials
    carried out in the USA.

         As shown in Table 3, the decrease of residues with time is most
    pronounced during the first 10 days after the last treatment of the
    cotton plants. The number of applications in a season appears to have
    little influence on the residue present at harvest. When comparing the
    dates of application with the residue levels in seed, it was thought
    that the later the last application was made in the season, the higher
    the residues. To test this hypothesis a study was conducted in South
    Carolina in 1977 (Ciba-Geigy, 1978b) in which the interval between
    applications and the pre-harvest interval were varied. The date of the
    last application was compared with the proportion of open bolls. With
    only one exception, the 0.25 kg/ha rate resulted in residues
    proportionally higher than the 0.125 kg/ha rate. In addition, the data
    showed that the timing of the last application (number of bolls open)
    influences the level of residues more than the total number of
    applications or the interval between applications.

         Though most of the data are from trials using the emulsifiable
    concentrate base, in some studies the emulsifiable concentrate and the
    soluble powder (hydrochloride) formulations were compared. The results
    were essentially the same (see Table 3).  Trials in which the fate of
    residues were traced through all stages of cottonseed processing
    showed that this similarity remains at each stage (Table 4).

    TABLE 1. Effect of application rate on chlordimeform residues in
             cottonseed.

    Application rate,        No. of                  Residue, mg/kg
    kg/ha                    trials                                       

                                              Range                  Mean

    0.125                      32             <0.05 - 0.49           0.1

    0.25                        2             <0.05                  <0.05

    0.45                        2             2.0 - 2.8              2.4

    0.5                         9             <0.05 - 3.0            1.7

    0.6                         5             <0.05 - 3.0            1.5

    0.75                        1             1.3                    1.3

    1.0                        23             <0.05 - 5.7            2.3

    1.5                         2             0.25 - 3.7             2.0

    2.0                         6             0.05 - 5.6             1.9

    2.25                        1             12.0                   12.0

    3.6                         1             13.1                   13.1

    NOTE:  The high figures in the range represent residues in cottonseed
           harvested immediately after application.

    
    TABLE 2. Chlordimeform residues in cottonseed, USA.

                                        Application
                                                            
    State          Year      No.    Rate,        Formulation    20        30        40        50        60        70        > 70
                                    kg/ a.i./ha

    Ala.           1970       8     0.5          EC                                                                         0.46
    Miss.          1970       9     0.5          EC                       0.95
    Ark.           1969       9     0.5          EC                       < 0.05
    Tex.           1970       9     0.5          EC                                                     0.22
    Cal.           1969      10     0.5          SP                                                                         2.7
    Cal.           1969      10     0.5          EC                                                                         2.3
    Geo.           1970      14     0.5          EC             3.0
    Cal.           1970       7     0.75         SP             2.6
    S. Car.        1969       5     1.0          EC                                                                         <0.05
    Okla.          1970       6     1.0          EC                                                     < 0.05
    Tex.           1969             1.0          EC                                                               0.22
    Cal.           1968      10     1.0          SP                       3.3
    Miss.          1969      11     1.0          EC                                 0.81
    Tex.           1970       7     1.5          EC             0.25
    Tex.           1970       9     1.5          EC             3.7
    Tex.           1969       7     2.0          EC             0.05
    Miss.          1970       9     2.0          EC                       5.6
    Miss.          1969      11     2.0          EC                                 0.62
    Tex.           1970      11     2.0          EC                                           1.5
    Tex.           1970       9     2.25         EC             12.0
                                                                (3 days)

    EC = Emulsion concentrate
    SP = Soluble powder

    TABLE 3. Effect of pre-harvest interval on level of chlordimeform residues in cottonseeds

                                            Application
                                                                 
    State          Year       No.     Rate, kg        Formulation    3         7         10        14        21
                                      a.i./ha

    Ariz.          1970       6       1.0             EC             1.8       1.6       2.3       1.9

    Ariz.          1970       6       1.0             SP             2.0       1.8       2.4       1.6

    Florida        1970       14      1.0             EC             3.7       3.0       2.0       3.0

    Florida        1970       14      1.0             SP             5.7       5.1       2.3       2.9

    Georgia        1975       20      0.5             EC             2.0       1.0                 1.0       0.81

    Miss.          1975       20      0.5             EC             3.0       1.6                 1.3       0.92

    EC = Emulsion concentrate
    SP = Soluble powder

    TABLE 4. Effect of Processing on chlordimeform residues in cottonseed and cottonseed products.

                             333/     333/     333/     333/     333/     333/     333/      13/      13/     13/       13/      13/
    Trial no.                107      92       92       105      86       109      08        77       77      77        77       77

    Location                 Texas    Calif.   Calif.   Calif.   -        Miss.    Texas     Calif.   Calif.  Calif.    Calif.   Miss.

    Rate of application
      kg/ha                  0.5      0.5      0.5      1.0      -        2.0      2.0       1.0      1.0     2.0       2.0      2.0

    Formulation              EC       EC       EC       SP       -        EC       EC        EC       SP      EC        SP       EC

    No. of applications      9        10       10       10       -        9        11        15       15      15        15       15

    Pre-harvest interval
    (days after last appl.)  57       84       84       28       -        24       45        21       21      21        21       -

    Cottonseed               0.22     2.3      2.7      3.3      -        5.6      1.5       2.9      3.4     -         -

    Lint cotton              0.37     -        -        -        -                 5.2       -        -       -         -        

    Delinted cottonseed      0.19     -        -        -        -                 1.4       -        -       -         -

    Linters                  0.47     -        -        -        -                 2.3       -        -       -         -

    Cottonseed hulls         0.20     3.1      3.9      8.0      -        8.5      2.2       3.75     4.0     -         -

    Solvent-extracted
    meal                     0.15     1.6      2.5      2.0      -        1.4      0.32      0.15     0.2     -         -

    Screw press
    extracted meal           0.09     0.92     1.1      1.1      -        0.83     0.39      -        -       -         -

    Crude
    solvent-extracted meal   0.11     -        -        -        1.2      0.63     0.28      0.41     0.52    0.8       1.1      3.8

    TABLE 4. Cont'd.

                             333/     333/     333/     333/     333/     333/     333/      13/      13/     13/       13/      13/
    Trial no.                107      92       92       105      86       109      08        77       77      77        77       77

    Location                 Texas    Calif.   Calif.   Calif.   -        Miss.    Texas     Calif.   Calif.  Calif.    Calif.   Miss.

    Refined
    solvent-extracted meal   0.09     0.73     0.78     -        1.0      0.21     0.33      0.37     0.38    0.8       0.8      3.3

    Crude screw
    press-extracted oil      0.17     -        -        -        4.0      1.7      1.3       -        -       -         -        -

    Refined screw
    press-extracted oil      0.14     1.3      1.7      -        3.6      0.85     0.72      -        -       -         -        -

    Solvent-extracted
    soapstock                -        1.2      1.8      -        -        -        -         -        -       -         -        -

    Screw press-extracted
    soapstock                -        2.8      2.8      -        -        -        -         -        -       -         -        -

    Refined, bleached,
    deodorised oil           -        -        -        -        -        -        -         <0.05    <0.05   0.05      0.06     <0.05

    Refined, bleached,
    deodorised,
    hydrogenated oil         -        -        -        -        -        -        -         <0.05    <0.05   <0.05     <0.05    <0.05

    FATE OF RESIDUES

    General

         Since the 1971 Meeting FAO/WHO, 1972b there have been a number of
    detailed studies on metabolism in cotton plants and on the fate of
    residues in cottonseed and products subjected to commercial
    processing. There have also been studies of the nature and fate of
    residues in lactating cows and a balance and characterization study of
    radio-labelled metabolites in a lactating goat. Studies have also been
    made into the fate of residues fed to poultry.

    Fate in plants

         The metabolic behaviour was studies in greenhouse-grown cotton
    plants (Gross, 1977). Selected leaves of the plants were treated at a
    rate equivalent to 0.6 kg a.i./ha with 14C-phenyl-labelled
    chlordimeform at the "first flower" stage and the treatment was
    repeated twice at one-week intervals. The distribution of
    radioactivity and pattern of metabolites were determined 30 minutes
    and one, two, eight and eleven weeks after the first treatment. The
    last period corresponded to the normal pre-harvest interval.
    Metabolism was relatively slow: six weeks after the last treatment,
    18.2% of the radioactivity was still unchanged chlordimeform.

         The treated leaves were exhaustively extracted with
    acetonitrile/water (8:2). The extractable radioactivity was further
    fractionated into hexane, methylene chloride- and water-soluble
    fractions. The radioactivity of the organic fractions was found to
    consist of at least seven different substances. Four of them were
    characterized by TLC comparison with reference compounds (Figure 1) as
    chlordimeform, N-demethyl-chlordimeform, 4-chloro-o-toluidine and
    N-formyl-4-chloro-o-toluidine.  Their identIty was confirmed by gas
    co-chromatography. A fifth compound, also found by Bull (1973) was not
    identical to any of the available reference compounds.

         Fifty six per cent of the foliar-applied dose was found in the
    plant after one week, the balance being lost by volatilization. Most
    of the absorbed radioactivity remained in the treated plant parts
    during eight weeks accounted for only 1.8% of the dose. At harvest the
    radioactivity calculated as chlordimeform was 0.45 mg/kg in the open
    bolls, 0.1 mg/kg in the fibre and 0.3 mg/kg in the seeds. No parent
    insecticide was found in the seeds and only 4.5% of the radioactivity
    in the seeds still contained the intact aniline moiety. Hydrolysis of
    the parent compound to yield N-formyl-4-chloro-o-toluidine and
    4-chloro-o-toluidine was the main degradation pathway.
    N-demethylation of chlordimeform was of more importance only at later
    sampling times. Strong evidence for the formation of the 
    N-hydroxymethyl derivative of chlordimeform as an intermediate was
    found. Some water-soluble and unextractable compounds were produced.
    As previously demonstrated by Bull (1973), this metabolic behaviour of
    chlordimeform in cotton is similar to that reported in apple seedlings
    (Sen Gupta and Knowles 1969) and grapefruit seedlings (Ehrhardt and
    Knowles, 1970).

         After hydrolysis and analysis by TLC and GLC, 45% of the
    water-soluble radioactivity was found as 4-chloro-o-toluidine.
    Another 42% of the radioactivity remaining in the aqueous hydrolysate
    was extractable with n-butanol at pH 1 but was not further
    characterized.

         The unextractable radioactivity in the treated leaves was low at
    all time intervals. On the other hand, when the untreated leaves,
    stems and green bolls at the 8 week sampling interval were extracted
    exhaustively with acetonitrile/hexane (7:3), then 80% aqueous methanol
    and finally hot methanol only 15% to 23% of the radioactivity of each
    plant part could be extracted. No unchanged chlordimeform was present
    in the untreated plant parts. When the plant material was hydrolysed
    45%, 17% and 10.5% of the radioactivity of the untreated leaves, stems
    and bolls, respectively, was extractable by iso-octane.

         Extraction of cottonseeds, taken from mature bolls 11 weeks after
    the first application, showed 8% of the radioactivity in the
    acetonitrile/hexane phase and an additional 7% of the radioactivity in
    the aqueous methanol. Hydrolysis of the seeds showed that only 4.5% of
    the radioactivity could be converted to 4-chloro-o-toluidine. When
    the nonextractable radioactivity of the weeds was further fractionated
    by chemical means into pectin, lignin, hemicellulose and cellulose
    fractions, the main part of the radioactivity (78%) was found to be
    associated with the cellulose fraction and 14%, 2% and 6% in the
    hemicellulose, lignin and pectin fractional respectively.

         Gross (1977) concluded that when 14C-chlordimeform was topically
    applied to cotton leaves, there was little movement of the
    radioactivity and none of chlordimeform itself into the untreated
    plant parts. The translocated radioactivity consisted exclusively of
    polar, mainly non-extractable substances, suggesting a possible
    incorporation of radioactivity into natural products.

         Fischer and Cassidy (1976a) reported an experiment in which the
    soil of a cotton field plot was treated with 14C-chlordimeform when
    the cotton was ten weeks old. The rate of application to the soil
    surface was equivalent to 1.0 kg a.i./ha.

         Radioactivity in the cotton plants seven weeks after treatment
    was equivalent to 0.13 mg/kg chlordimeform. Biphasic extraction showed
    42.4% in the organic fraction, 34.9% in the polar fraction and 25.8%
    was non-extractable. Thirteen weeks after treatment the mature cotton
    contained 0.09 mg/kg in the leaves. The radioactive content of these
    mature samples was too low to fractionate for balance determinations.
    Treatment of soil surrounding a growing cotton plant showed that
    14C-chlordimeform and its soil metabolites are taken up, in small
    quantities by cotton, especially the seeds and fibres.

         In another experiment Fischer and Cassidy (1976b) treated a
    cotton field plot over-the-top with formulated 14C-chlordimeform at a
    rate of 1.0 kg/ha when the plants were 10, 12 and 14 weeks old.

    Radioactivity in the cotton plants immediately after treatment was
    equivalent to 2.44 mg/kg chlordimeform. At harvest the radioactivity
    calculated as 14C-chlordimeform was 12.91 mg/kg in the leaves, 0.99
    mg/kg in stalks, 0.03 mg/kg in the fibre and 0.26 mg/kg in the seed
    with 0.07 mg/kg in the oil and 0.19 mg/kg in the meal. The balance
    could be done only on the leaves and stalks at maturity because of the
    low radioactivity in the other plant parts.  From the leaves and
    stalks respectively, the organic fractions contained 57.6% and 72.9%,
    the polar fractions 18.9% and 11.4% and the non-extractable fractions
    18.8% and 17.8%. Parent 14C-chlordimeform accounted for 31.0% and
    45.2% in leaves and stalks respectively. The radioactivity
    co-chromatographing with other known standards was minor, accounting
    for no more than 8% of the total in either leaves or stalks. This
    indicates that there is a large percentage of polar conjugates. The
    data show that, although leaf radioactivity is high, there is little
    translocation of 14C-chlordimeform metabolites to the seed or fibre.

         Application of the method for determining chlordimeform and its
    metabolites to cotton leaves showed that 88.9% of the radioactivity in
    the leaves but only 29.9% of that in the seeds could be accounted for
    as 4-chloro-o-toluidine. This indicates extensive further metabolism
    of the translocated metabolites of 14C-chlordimeform.

         Fischer and Cassidy (1976c) identified the radioactive
    metabolites in leaves after phenyl-14C-chlordimeform was sprayed
    over-the-top according to standard agricultural practice. At mature
    harvest the total radioactivity in the leaves consisted of parent
    chlordimeform 60.3%, demethyl-chlordimeform 4.1%,
    4-chloro-o-toluidine 7.6% and its N-formyl derivative 7.0%.
    Identification was based on partitioning characteristics and
    cochromatography with standards. The metabolism of chlordimeform by
    cotton leaves involves, therefore, sequential hydrolysis of terminal
    groups with 4-chloro-o-toluidine as the final non-polar product.
    These data show that parent chlordimeform will account for the main
    part of the material containing the chlorotoluidine moiety in mature
    cotton foliage.

         Cotton grown in a greenhouse was injected with ring-labelled
    chlordimeform in order to characterize the radioactive metabolites,
    especially those not extractable with methanol and water, and to
    elucidate the metabolic pathway of chlordimeform in cottonseed
    (Honeycutt and Cassidy, 1977). The injections were made into the stem
    69, 73 and 81 days after planting, the plants were grown to maturity
    and the cotton harvested 157 days from planting. The cottonseed
    contained radioactivity equivalent to 0.65 mg/kg of chlordimeform at
    maturity. Polar, extractable material accounted for 58% of the total
    radioactivity in the seed and consisted of 5-chloro-2-formamidobenzoic
    acid (N-formyl-5-chloroanthranilic acid) (3%) and eight unknown polar
    metabolites, none of which made up more than 8% of the total.

         Forty per cent of the radioactivity in the cottonseed was not
    extractable and was further fractionated into base-soluble (30%) and
    base-insoluble (23%) radioactivity. The base-soluble radioactivity was

    characterized as low molecular weight degradation products "A" (1.2%)
    and "B" (5.7%), as well as high molecular weight (> 1,000) components
    (20%) physicochemically similar to lignin. The base-insoluble
    radioactivity was characterized mainly as radioactive conjugates
    (17.6%) covalently linked to natural products physicochemically
    similar to cellulose. Three percent of the total radioactivity in the
    cottonseed could not be solubilized either by strong acid or base
    hydrolysis. Application of the total hydrolysis method to both the
    polar extractable and the non-extractable radioactivity showed that a
    total of only 19.8% of the radioactivity in the cottonseed could be
    converted to 4-chloro-o-toluidine (5-CAT).

         The above data confirm that the metabolism of chlordimeform in
    cottonseed is extensive, processing through 5-CAT, its N-formyl
    derivative and N-formyl-5-chloroanthranilic acid to hydroxylated
    metabolites which become conjugated to natural products such as
    lignin, pectin, cellulose or hemicellulose. This explains why attempts
    to define a moiety common to chlordimeform and its metabolites in
    cottonseed have not been successful.

         The authors have deduced the scheme set out in Figure 2 to
    explain the metabolism of chlordimeform in cotton. They contend that
    4-chloro-o-toluidine is incorporated into lignin according to the
    schemes outlined in Figure 2.

    In animals

         Burkard (1971a) studied the fate following the application of a
    0.5% solution of chlordimeform to the hindquarters of cows. The
    treatment was applied three times at 7-day intervals and the results
    were determined in milk collected daily until 33 days after the last
    treatment. At the end of that period the cows were slaughtered and
    samples of meat, liver and fat were analysed.

         Generally the total residue in milk was below the limit of
    determination (0.03 mg/kg) except on the day following treatment when
    it rose to a maximum of 1.0 mg/kg. Thereafter the concentration
    declined rapidly reaching the limit of determination on the third day.
    The meat and fat contained no detectable residues but liver contained
    from 0.3 to 0.6 mg/kg 33 days after the last treatment.

         In a further study Burkard (1971b) fed Swiss cows daily with 1 kg
    of a concentrate containing from 40 to 240 ppm chlordimeform for
    periods up to 42 days. Residues were determined in samples of milk
    collected throughout the trial and in samples of meat, fat, liver and
    kidney obtained from cows slaughtered after 7, 14, 21 and 42 days
    continuous administration.

         Total residues of chlordimeform and its metabolites in all milk,
    meat and fat samples analysed were below the limit of determination
    (0.03 mg/kg). However, the residues in the kidney and liver samples
    (mean values of duplicate determinations) were as shown in Table 5.

    TABLE 5. Residues of chlordimeform and its metabolites in cows.

                        Chlordimeform and metabolites, mg/kg
    Duration of                                                        
    feeding (days)
                        In concentrate      In liver          In kidney

    21                        40              0.09             < 0.03

    21                       120              0.38               0.07

    7                        240              0.45               0.05

    14                       240              0.58               0.13

    21                       240              0.50               0.13

    42                       240              0.44               0.09

         Honeycutt and Cassidy (1976) fed a lactating goat with cottonseed
    containing biosynthesized metabolites of phenyl-14C-chlordimeform at
    a level equivalent to 0.032 ppm chlordimeform daily for 4 consecutive
    days. The total recovery of radioactivity was 97.1%. It was primarily
    distributed between the urine (21.3%), which reached a plateau by the
    2nd day, the faeces (35.9%), which reached a plateau by the third day,
    and the rumenintestinal contents (38.7%). There was no deposition of
    radioactivity in tissues (< 0.002 mg/kg), even though uptake of the
    biosynthesized metabolites occurred.

         The milk contained 0.9% of the total dose, its radioactivity
    plateaued by the second day (0.0004 ± 0.0001 mg/kg chlordimeform
    equivalents). The ratio of the concentration in milk to that in the
    feed was 0.013. Blood contained < 0.2% of the total dose.

         Comparison of the 14C-extractables of the original cottonseed
    with those in the faeces, urine and rumen-intestinal contents,
    suggests that the goat metabolized the biosynthesized metabolites in
    cottonseed. However, metabolism to natural products was not
    significant since expired 14CO2 accounted for only 0.3% of the total
    dose.

         Ciba-Geigy Corp. (1971) conducted experiments in which white
    Leghorn hens in active egg production were fed a commercial egg laying
    mash to which chlordimeform was added at rates equivalent to 0, 0.25,
    0.75 and 1.50 mg/kg of feed. Groups were sacrificed after 7, 14, 21
    and 28 days feeding and were subjected to post-mortem examination and
    assessment of egg production, body-weight change and food consumption.

    There were no significant differences between treated and control
    binds in any of the features examined.

         Residues in eggs and tissues (breast, fat and liver) were
    determined by Jenny (1971). Eggs collected at -3 (pre-treatment), -1
    (pre-treatment), 3, 7, 14, 21 and 28 days from each group contained no
    detectable amounts of chlordimeform and/or its metabolites containing
    the 4-chloro-o-toluidine moiety.

         The breasts of birds sacrificed at 7, 14, 21 and 28 days of
    feeding treated feed as well as 49 days after returning to untreated
    feed contained no significant residues. Average residues of 0.11, 0.09
    and 0.22 mg/kg were detected in fat at the feeding levels of 0.25,
    0.75 and 1.50 ppm respectively, at the 21-day interval only. No
    residues were detected in livers from chickens fed 0.25 ppm
    chlordimeform at any of the intervals tested and residue of 0.09 mg/kg
    was detected in liver from chickens fed 0.75 ppm only at the 21-day
    interval. Residues of 0.20, 0.17, and 0.10 mg/kg were detected in
    livers from the 1.5 ppm level at intervals of 7, 14, 21 and 28 days.
    Nine days after withdrawal from the feed containing 1.5 ppm, no
    residues could be detected in any of the tissues.

    In processing

         The Meeting reviewed many data on the level and fate of residues
    during processing. Cottonseed is subjected to well-defined sequential
    processes, details of which were available (Ciba-Geigy, 1978). The
    results of some typical trials are set out in Table 4 and 6. The
    distribution is independent of the pattern of application as is
    apparent from Table 6. Although the major fraction of the residue
    remains in the hulls and the meal from which the oil has been removed
    there is still a significant concentration in the crude oil. Treatment
    of crude cottonseed oil with caustic soda in the refining process does
    not reduce the residue concentration significantly but the additional
    refining procedures including bleaching, hydrogenating and deodorizing
    reduce the residue in the oil to a level below the detection limits of
    present analytical methods (< 0.05 mg/kg). Since all the oil used for
    human consumption is at least subjected to the bleaching and
    deodorizing process, virtually all the chlordimeform will be removed
    before the cottonseed oil is consumed.

    In soil

         In the course of studies on the uptake of chlordimeform and its
    soil metabolites from treated soil into cotton Fischer and Cassidy
    (1976a) applied radio-labelled chlordimeform to a cotton field plot
    thirteen weeks before harvesting the cotton. The rate of application
    was equivalent to 1 kg/ha.

    
    TABLE 6. Effect of use pattern on Chlordimeform residues in cottonseed products.

                            AGA       AGA        AGA        AGA       AGA       AGA       AGA       AGA       AGA       AGA
    Trial No.               4397      4408       4409       4449      4450      4502      4981      4981      4981      4981

    Location                Alab.     Ariz.      Calif.     Georg.    Georg.    Miss.     S.C.      S.C.      S.C.      S.C.

    Rate of application     0.125     0.125      0.25       0.125     0.25      0.125     0.125     0.25      0.125     0.25

    Formulation             EC        EC         EC         EC        EC        EC        EC        EC        EC        EC

    No. of applications     7         7          6          11        12        9         12        12        17        17

    Pre-harvest interval    28        39         31         22        16        31        21        21        21        21

    Cottonseed              0.2       0.82       0.35       0.35      0.35      0.30      0.29      0.20      0.22      0.40

    Hulls                   0.36      1.8        0.85       0.58      0.55      0.33      0.86      0.50      0.90      0.75

    Meal                    0.19      1.2        0.45       0.24      0.32      0.45      0.18      0.10      0.17      0.15

    Soap stock              0.10      0.84       0.34       0.17      0.23      0.22      0.09      0.07      0.10      0.10

    Crude oil               0.10      0.46       0.21       0.15      0.23      0.16      0.10      0.07      0.11      0.08

    Refined oil             0.12      0.66       0.32       0.17      0.19      0.19      0.07      <0.05      0.08      0.07

    Refined bleached
    deodorised oil          <0.05     <0.05      <0.05      <0.05     <0.05     <0.05     <0.05     <0.05     <0.05     <0.05

    Refined bleached
    deodorised
    hydrogenated oil        <0.05     <0.05      <0.05      <0.05     <0.05     <0.05     <0.05     <0.05     <0.05     <0.05

         Radioactivity in the top 75 mm layer of silt loam soil accounted
    for 1.23 mg/kg chlordimeform equivalents after treatment. Seven weeks
    later this level had decreased to 0.33 mg/kg and at 13 weeks to 0.20
    mg/kg. When the 0-75 mm layer was extracted after seven weeks, 32% of
    the radioactivity partitioned into the organic fraction, 20%
    partitioned into the polar fraction and 44% remained non-extractable,
    showing that the chlordimeform had degraded rapidly. The radioactivity
    in the samples taken 13 weeks after treatment was too low to
    fractionate. Except for one sample, the level of radioactivity as
    chlordimeform equivalents in the 75-150 mm and 150-200 mm layers was
    less than 0.01 mg/kg; thus leaching did not occur in this silt loam.

         In a later study when radio-labelled chlordimeform was applied at
    the rate of 1 kg/ha over cotton plants and soil in field plots,
    Fischer and Cassidy (1976b) found that the radioactivity in the 0-75
    mm layer of silt loam was 1.24 mg/kg immediately after the first
    treatment but only 0.21 mg/kg after the final treatment and 0.20 mg/kg
    at harvest of the mature cotton. This decrease is due to (1) less
    drainage from the leaves onto the soil at the later sprayings because
    of the increased cover with plant growth, (2) volatility of the
    chlordimeform and (3) soil metabolism. Radioactivity was less than
    0.01 mg/kg in the 75-150 and 150-225 mm layers at all-sampling dates.
    These data show that metabolism of 14C-chlordimeform in silt-loam is
    rapid and that the metabolites are not leached.

         Analysis of the 0-75 mm layer of soil taken at harvest showed
    that 91.0% of the radioactivity could be converted to 
    4-chloro-o-toluidine. Therefore, although 57% of the soil 
    metabolites were polar or non-extractable, they contained this moiety.

         Following this study Fischer and Cassidy (1976b) planted soybeans
    at a rotation crop 38 weeks after the original treatment with
    chlordimeform at 1 kg/ha. At maturity the soybeans were found to
    contain radioactivity equivalent to 0.01 mg/kg chlordimeform in the
    stalks and less than 0.01 mg/kg in the beans themselves.

         Wheat planted in the plots was examined for radioactivity at
    maturity. The radioactivity was equivalent to 0.16 mg/kg in the straw
    and 0.03 mg/kg in the grain.

    METHODS OF RESIDUE ANALYSIS

         Although methods have been developed for the determination of
    chlordimeform alone, most of the residue data have been based on the
    determination of chlordimeform and its metabolites as
    4-chloro-o-toluidine.

         Geissbuhler et al. (1971) published the method used by Ciba-Geigy
    Ltd. and Schering A.G. for the development of chlordimeform
    pesticides. This method has been utilized with and without
    modification for much of the work reported in this monograph. In this

    method chlordimeform and its potential metabolites and/or conjugates
    are hydrolyzed to 4-chloro-o-toluidine in the presence of plant and
    soil material by successive treatments with acetic acid and sodium
    hydroxide. The aromatic amine is steam distilled and extracted into
    iso-octane. Diazotization and coupling of 4-chloro-o-toluidine with
    N-ethyl-1-naphthylamine yields a stable purple dye, which, after
    column chromatography on cellulose, is measured colorimetrically. To
    verify the identity of chlordimeform, the azo-dye is separated and
    visualized on a cellulose thin-layer plate. Alternatively,
    4-chloro-o-toluidine is diazotized and iodinated. The iodine
    derivative is determined by electron capture gas chromatography. A
    further potential metabolite of chlordimeform
    2,2-dimethyl-4-4-dichloroazobenzene, is reduced to
    4-chloro-o-toluidine in the presence of zinc and acid 1,4-dioxane
    and likewise determined by gas chromatography. Recoveries in various
    plant and soil materials have been observed to vary between 75 and
    100% for all three quantitative methods described. The limit of
    determination is 0.05 mg/kg.

    APPRAISAL

         Chlordimeform products which had been widely used for many years
    as insecticides/acaricides on a large variety of crops and on
    livestock, were withdrawn by the manufacturers in 1976. Additional
    studies have since been carried out and new proposals have been made
    for commercial use on cotton crops under restricted conditions.
    Information on those proposed uses in various countries and data on
    the nature, level and fate of residues, and including the fate of
    residues in cottonseed products used in the rations of dairy cattle
    and poultry was also examined.

         The influence of rate of application pre-harvest interval and
    number of applications were investigated. At the maximum application
    rate of 1 kg/ha the residue rarely exceeded 2 mg/kg in cottonseed,
    seed meal or crude oil. Solvent-extracted oil contained significantly
    lower residues than screwpress-extracted oil. Treatment of crude oil
    with caustic sodal did not reduce residues markedly but refining
    procedures such as bleaching and deodorization as normally undertaken
    in commercial practice reduced the levels to below the analytical
    detection limits (< 0.05 mg/kg).

         Eggs from hens fed for 28 days on rations containing up to 1.5
    ppm contained no detectable residues. Meat, fat and liver from hens
    slaughtered at various intervals throughout the study was analysed.
    Although nothing was found in any of the meat, small residues were
    detected in some samples of fat and liver. Cows fed for up to 42 days
    on rations containing excessive levels of chlordimeform (up to 240
    ppm) showed no residues (< 0.3 mg/kg) in milk, fat or meat at any
    stage during the feeding; but residues in some animals fed at the
    highest rates, ranged up to 0.6 mg/kg in liver and 0.1 mg/kg in

    kidney. Since there appears to be no likelihood of livestock rations
    containing more than 0.5 mg/kg of residue however, detectable residues
    are unlikely to occur in foods of animal origin.

         The analytical methods used determined chlordimeform, the
    intermediate metabolite N-formyl-4-chloro-o-toluidine and the final
    metabolite 4-chloro-o-toluidine together as 4-chloro-o-toluidine. No
    attempt was made to quantify the amounts of each present and the limit
    of determination was 0.05 mg/kg.

         Studies of the metabolism of chlordimeform in cotton plants and
    goats showed that metabolism in plants and animals proceeds through
    sequential hydrolysis of terminal groups to 4-chloro-o-toluidine. In
    cotton metabolism proceeds to the formation of polar metabolites which
    apparently conjugate to natural products. Sprays applied after the
    bolls open would partly avoid translocation of residues as metabolites
    from the treated leaves to seed but would result in the retention of
    significant residues of the parent compound in fibre and seed. In a
    study involving the feeding of cottonseed containing biosynthesised
    radioactive metabolites to a goat, no residues were deposited in the
    animal tissue (limit of determination 0.002 mg/kg).

         In the light of the new data examined, the Meeting decided that
    there was no need to amend the temporary MRL formerly proposed for
    cottonseed and crude cottonseed oil; but unless direct application to
    cattle were to be re-installed, the MRL for foods of animal origin
    should be based on the data reflecting the feeding of cottonseed
    hulls, cottonseed meal and similar products. As chlordimeform is no
    longer used on crops other than cotton, the recommendations for
    temporary MRLs previously made for crops other than cotton should be
    held in abeyance until such time as the uses on those crops should be
    reinstated.

    RECOMMENDATIONS

         The following new or amended recommendations for temporary MRLs
    replace all previous recommendations.

              Commodity                     Temporary MRL mg/kg

              Cottonseed                    2
              Cottonseed (crude)            2
              Cottonseed oil (edible)  )    No residues are to
                                       )    occur at the current
              Meat of cattle           )    limit of detection
                                       )    (0.05 mg/kg)
              Meat of pigs             )
              Meat of poultry          )    "     "     "     "
              Meat of sheep            )
              Milk                     )
              Milk products            )

    FURTHER WORK OR INFORMATION

    Required (by 1979)

    1.   Full submission of all toxicological data.

    2.   Continued epidemiology data on people exposed to chlordimeform.

    3.   Reports of continued observation of the possible occurrence of
         haemorrhagic cystitis.

    REFERENCES

    Anonymous Chlordimeform HC1 Life Time Feeding Study in Rats - One
    (1978a)                  Year Interim Report. Unpublished report from
                             CIBA-GEIGTY, Ltd., Basal, Switzerland
                             submitted by CIBA-GEIGY, Ltd. to the WHO.

    Anonymous N-Formyl-4-chloro-o-toluidine Life Time Feeding Study in
    (1978b)                  Rats One Year Interim Report. Unpublished
                             report from CIBA-GEIGY, Ltd., Basel,
                             Switzerland submitted by CIBA-GEIGY, Ltd. to
                             the WHO.

    Anonymous 4-chloro-o-toluidine HC1 Life Time Feeding Study in Rats
    (1978c)                  - One Year Interim Report. Unpublished report
                             from CIBA-GEIGY, Ltd., Basel, Switzerland
                             submitted by CIBA-GEIGY, Ltd. to the WHO.

    Anonymous Bioassay of 4-chloro-o-toluidine Hydrochloride for
    (1978d)                  Possible Carcinogenicity.  NCI Carcinogenic
                             Programme Preliminary Draft Report [ NIH ]
                             79-1721).

    Arni, P. and D. Muller Salmonella/Mammalian-Microsome Mutagenicity
    (1976a)                  Test with C-8513 Chlordimeform HC1.
                             Unpublished report from CIBA-GEIGY, Ltd.,
                             Basel, Switzerland submitted by CIBA-GEIGY,
                             Ltd. to the WHO.

    Arni, P. and D. Muller Salmonella/Mammalian-Microsome Mutagenicity
    (1976b)                  Test with CGA 72651
                             (N-formyl-4-chloro-o-toluidine).
                             Unpublished report from CIBA-GEIGY, Ltd.,
                             Basel, Switzerland submitted by CIBA-GEIGY,
                             Ltd. to the WHO.

    Arni, P. and D. Muller Salmonella/Mammalian Microsome Mutagenicity
    (1976c)                  Test with CGA 72647 (4-chloro-o-toluidine
                             HC1). Unpublished report from CIBA-GEIGY,
                             Ltd., Basil, Switzerland submitted by
                             CIBA-GEIGY, Ltd. to the WHO.

    Benezet, H.J. and C.O. Knowles Inhibition of Rat Brain
    (1976)                   Monoamine Oxidase by Formamidine and Related
                             Compounds. Neuropharmacol. 15:369.

    Benezet, H.J., K.M. Chang and C.O. Knowles Formamidine Pesticides-
    (1978)                   Metabolic Aspects of Neurotoxicity in
                             Pesticide and Venom Neural Toxicity. Ed. D.L.
                             Shankland, R.M. Hollingworth and J. Smyth,
                             Jr. Plenum Press, N.Y. pg. 189-206.

    Bull, D.L., Environmental Entomology 2, 869.
    (1973)

    Burkhard, N. Residues in milk, meat, liver and fat of a Swiss cow
    (1971a)                  washed with a 0.5%  solution of
                             chlordimeform. Report RVA 14/71, Agrochemical
                             Division, CIBA-GEIGY, Ltd., Basel,
                             Switzerland. 1 February 1971.

    Chinn, C., W.R. Pfister and G.K.W. Yim Local Anesthetic-Like Actions
    (1976)                   of the Pesticide Chlordimeform. Fed. Proc.
                             35:729.

    Ciba-Geigy Chlorphenamidime residues in eggs and poultry tissues.
    (1971)                   Research Report C79910, CIBA-GEIGY, Corp.
                             Greensboro N.C.

    Ciba-Geigy Method for determination of chlordimeform residues -
    (1976)                   Method REM 4/76.

    Ciba-Geigy Submission of data for the reevaluation of chlordimeform by
    (1978)                   the 1978 FAO/WHO Joint Meeting on Pesticide
                             Residues, CIBA-GEIGY, Ltd., Basel and
                             Schering AG, Berlin. July 24, 1978.

    Ciba-Geigy Residues resulting from chlordimeform application to
    (1978a)                  cotton. Report by H.B. Camp and B.G. Tweedy
                             No ABR-78004 (4 January 1978).

    Ehrhardt, D.A. and Knowles, C.O. J. Econ. Entomol. 63, 1306.
    (1970)

    Ezumi, K. and H. Nakao Effects of 4-chloro-o-toluidine in Oral
    (1974a)                  Prolonged Administration to Rat for 94 (Male)
                             and 104 (Female) Weeks. Unpublished report
                             from Nihon Schering Inc., Osaka, Japan
                             submitted by CIBA-GEIGY, Ltd. to the WHO.

    Ezumi, K. and H. Nakao Effects of 4-chloro-o-toluidine in Oral
    (1974b)                  Prolonged Administration to Mice for 80 Weeks
                             Unpublished report from Nihon Schering, Inc.,
                             Osaka, Japan submitted by CIBA-GEIGY, Ltd. to
                             the WHO.

    FAO/WHO 1971 Evaluations of some pesticide residues in food.
    (1972b)                  AGP:1971/M/9/1; WHO Pesticide Residues Series
                             No 1.

    FAO/WHO 1975 Evaluations of some pesticide residues in food. AGP:
    (1976b)                  1975/M/13; WHO Pesticide Residue Series No 5.

    FAO/WHO Pesticide residues in food. Report of the 1976 Joint Meeting,
    (1977a)                  of the FAO Panel Experts on Pesticide.
                             Residues and the Environment and the WHO
                             Expert Group on Pesticide Residues. FAO Food
                             and Nutrition Series, No 9; FAO Plant
                             Production and Protection Series, No 8; WHO
                             Technical Report Series, No 612.

    FAO/WHO Pesticide residues in food - 1977. Report of the FAO Panel of
    (1978a)                  Experts on Pesticide Residues and Environment
                             and the WHO Expert Committee on Pesticide
                             Residues. FAO Plant Production and Protection
                             Paper 10 Rev. par 4.13.

    Fischer, W.C. and Cassidy, J.E. Uptake of radio-labelled (1976a)
                             chlordimeform and its soil metabolites in
                             cotton in a field prepared for rotation
                             crops. Report No GAAC-76014 CIBA-GEIGY
                             Corporation, Greensboro, N.C. March 15, 1976.

    Fischer W.C. and Cassidy, J.E. Uptake and characterization of
    (1976b)                  the metabolites of radio-labelled
                             chlordimeform in spray treated cotton and
                             field soil. Report of Biochemistry Department
                             CIBA-GEIGY Corporation Report ABR - 76056
                             July 27, 1976.

    Fischer W.C. and Cassidy, J.E. Identification of the metabolites
    (1976c)                  of chlordimeform in mature leaves of spray
                             treated field cotton. Report No ABR - 76073
                             from Biochemistry Department, CIBA-GEIGY,
                             Corporation, Greensboro, N.C.

    Fritz, H. Reproduction study on C8514 Technical Rabbit - Seg II. (Test
    (1971)                   for teratogenic and embryotoxic effects).
                             Unpublished report from Toxicology/ Pathology
                             CIBA-GEIGY, Ltd., Basely Switzerland,
                             submitted by CIBA-GEIGY Ltd to the WHO.

    Fritz, H. Reproduction Study on C8514 Technical Rat-Seg II (Test for
    (1975)                   Teratogenic or Embroyotic Effects).
                             Unpublished report from Toxicology/Pathology
                             CIBA-GEIGY, Ltd., Basely Switzerland
                             submitted by CIBA-GEIGY, Ltd. to the WHO.

    Geisebuhler, H., Kossmann, K., Baunck, I. and Boyd, V.F.
    (1971)                   Determination of total residues of
                             chlorphenamidine in plant and soil material
                             by colorimetry and thin layer and electron
                             capture gas-chromatography. Agric. Food Chem.
                             19 (2):365.

    Gross Metabolism of chlordimeform in cotton. Project Report 19/77 from
    (1977)                   Biochemistry Department, CIBA-GEIGY, Ltd.,
                             Basel, Switzerland (April 1977).

    Hollingworth R.M. Chemistry Biological Activity and Uses of
    (1976)                   Formamidine Pesticides. Environ. Health
                             Perspect. 14: 57-69.

    Honeycutt, R.C. and Cassidy, J.E. Extraction and characterization
    (1977)                   of cottonseed metabolite  from cotton
                             injected with o 14C-chlordimeform. Report No
                             ABR - 77087 from Biochemistry Department,
                             CIBA-GEIGY, Corporation, Greensboro, N.C.
                             Sept. 14, 1977.

    Hurni, H. and H. Ohder Report on the Mutagenic Effect of Technical
    (1970)                   C-8514-N. Unpublished report from Tierfarm AG
                             Biomedical Research, Sisseln, Switzerland
                             submitted by CIBA-GEIGY, Ltd., to the WHO.

    Jenny, N.A. Chlorphenamidine residues in eggs and poultry 
    (1971)                   tissues/Analysis of tissue and egg samples.

    Knowles, C.O. Chemistry and Toxicology of Quinoxaline Organotin
    (1976)                   Organofluorine and Formamidine Acaracides.
                             Env. Health Perspect. 14:93-102.

    Kalyonova, F.P., Z. Zapryanav and A.I. Baynova Influence of the
    (1978)                   Insecticide Chlordimeform of the Activity of
                             the Monoamine Oxidase (MAO) of Albino Rats.
                             Comptes Rendus de l'Academi Bulgare des
                             Sciences 31 (4): 491-493.

    Konopa, E.A. and H. Heymann Salmonella/Microsome Mutagenicity
    (1977)                   Test with Chlordimeform. Unpublished report
                             from Pharmaceuticals Division, CIBA-GEIGY,
                             Corporation, Surnmity N.J. submitted by
                             CIBA-GEIGY, Ltd., to the WHO.

    Konopka. E.A. and H. Heymann Salmonella/Microsome Mutagenicity
    (1977b)                  Test with Chlordimeform.  Unpublished report
                             from Pharmaceuticals Division, CIBA-GEIGY
                             Corporation, Summit, N.J. submitted by
                             CIBA-GEIGY, Ltd. to the WHO.

    Langauer, M. and D. Muller Nucleus Anomaly Test on Somatic
    (1977)                   Interphase Nuclei-C-8513- Chinese Hamster.
                             Unpublished report from Toxicology/Pathology,
                             CIBA-GEIGY, Ltd., Basel, Switzerland
                             submitted by CIBA-GEIGY, Ltd. to the WHO.

    Lund, A.E., G.K.W. Yim and D.L. Shankland The Cardiovascular
    (1978)                   Toxicity of Chlordimeform, a Local
                             Anesthetic-Like Action in Pesticide and Venom
                             Neurotoxicity. Ed. D.L. Shankland, R.M.
                             Hollingworth and T. Smyth, Jr. Plenum Press,
                             N.Y., pgs. 171-177.

    Maitre L., A. Felner, P. Waldmeier and W. Kehr Moccasins Oxidase
    (1978)                   Inhibition in Brain and Liver of Rats Treated
                             with Chlordimeform. J. Agr. Fd Chem. 26
                             (2): 442-446.

    Matsumura, F. and R.W. Beeman Biochemical and Physiological
    (1976)                   Effects of Chlordimeform.  Environ. Health
                             Perspect. 14: 71-82.

    Morton Method for the determination of chlordimeform. Method 333/7
    (1968)                   (7/12/68)

    Murakami, J. and J. Fukami Effects of Chlophenamidine and Its
    (1974)                   Metabolites on HeLa Cells.  Bull. Env.
                             Contam. and Tox. 11: 184-188.

    Nesnow, S. and C. Heidelberger The Effect of Modifiers of
                             Microsomal Enzymes on Chemical Oncogenesis in
                             Cultures of C3H Mouse Cell Lines. Cancer
                             Res. 36:1801-1808.

    Nesnow S. Unpublished bioassay submitted to the WHO.
    (1978)

    Nor-Am Method of analysis for chlordimeform residues - Research Method
    (1969)                   333/44. 5/6/69.

    Sachsse, K., P. Suter, F. Zak and R. Hess Lifespan Feeding Study in
    (1978b)                  Mice with 4-chloro-o-toludine. Unpublished
                             report from Toxicology, CIBA-GEIGY, Ltd.,
                             Basel, Switzerland submitted by CIBA-GEIGY,
                             Ltd. to the WHO.

    Schering Method of analysis of chlordimeform residues. Method 36 268/8
    (1973)                   (24/4/73).

    Schering Method of determination of chlordimeform and its degradation
    (1975)                   products. Method 36268/9 17/11/75.

    Sen Gupta, A.K. and Knowles, C.O. J. Agric. Food Chem. 17, 595.
    (1969)

    Suter, P., H. Luetkemeier, K. Sachsse and F. Zak 60-day Pending
    (1976a)                  Study in the Rat with chloro-o-toluidine
                             HC1. Unpublished report from
                             Toxicology/Pathology, CIBA-GEIGY, Ltd.,
                             Basel, Switzerland, submitted by CIBA-GEIGY,
                             Ltd. to the WHO.

    Suter, P., H. Luetkemeier, K. Sachsse and F. Zak 60-day Pending
    (1976b)                  Study in Mice with 4, chloro-o-toluidine
                             HC1. Unpublished report from
                             Toxicology/Pathology, CIBA-GEIGY, Ltd.,
                             Basel, Switzerland submitted by CIBA-GEIGY,
                             Ltd. to the WHO.

    Suter, P., F. Zak, K. Sachsse and R. Hess Lifespan Feeding Study
    (1979)                   in Mice with Chlordimeform HC1. Unpublished
                             report from Toxicology/Pathology, 
                             CIBA-GEIGY, Ltd., Basel, Switzerland, 
                             submitted by CIBA-GEIGY, Ltd. to the WHO.

    Wang, C.M., T. Narahashi and J. Fukami Mechanism of Neuromuscular
    (1975)                   Block by Chlordimeform.  Pest. Biochem.
                             Physiol. 5: 119-125.

    Watanabe, H., S. Tsuda and J. Fukami. Effects of Chlordimeform
    (1975)                   on Rectusabdominum Muscle in Frog. Pest.
                             Biochem. Physiol. 5: 150-154.

    See Also:
       Toxicological Abbreviations
       Chlordimeform (EHC 199, 1998)
       Chlordimeform (ICSC)
       Chlordimeform (WHO Pesticide Residues Series 1)
       Chlordimeform (WHO Pesticide Residues Series 5)
       Chlordimeform (Pesticide residues in food: 1979 evaluations)
       Chlordimeform (Pesticide residues in food: 1980 evaluations)
       Chlordimeform (Pesticide residues in food: 1985 evaluations Part II Toxicology)
       Chlordimeform (Pesticide residues in food: 1987 evaluations Part II Toxicology)
       Chlordimeform (IARC Summary & Evaluation, Volume 30, 1983)