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    DICLORAN            JMPR 1974

    IDENTITY

    Chemical name

         2,6-dichloro-4-nitroaniline

    Synonyms

         Dicloran (B.S.I.), DCNA, Botran(R), Allisan(R), Ditranil(R)

    Structural formula


        CHEMICAL STRUCTURE 8


    Other information on identity and properties

         Molecular weight:     207

         State:                Yellow, crystalline solid with practically
                               no odour

         Melting point:        192-194C

         Vapour pressure:      1.2 x 10-6 mm Hg (20C)

             Solubility:               Water7 mg/l
             Cyclohexane               0.006 g/100 ml
             Petroleum ether           0.02 g/100 ml
             Carbon tetrachloride      0.06g/100 ml
             Ethanol                   0.29 g/100 ml
             Benzene                   0.46 g/100 ml
             Glacial acetic acid       8.80 g/100 ml
             Chloroform                1.2 g/100 ml
             Acetone                   3.4 g/100 ml

         Stability:            Stable to hydrolysis and relatively stable
                               to oxidation. Readily reduced to
                               phenylenediamine by zinc and acid. Stable
                               to light and heat.

         Composition and
         purity:               The technical product contains
                               2,6-dichloro-4-nitroaniline not less than
                               96% (on dry weight basis);
                               2,4-dichloro-6-nitroaniline not more than
                               2%; 2-chloro-4-nitroaniline not more than
                               l%; Chloranil not more than 1%;
                               P-nitroaniline not more than 0.1%; Loss on
                               drying not more than 1% (at 60C, <5 mm
                               Hg); Sulphated ash not more than 0.25%;
                               Sodium chlorate not more than 100 ppm.

    EVALUATION FOR ACCEPTABLE DAILY INTAKE

    BIOCHEMICAL ASPECTS

    Biotransformation

         2,6-Dichloro-4-nitroaniline-14C administered to three male human
    subjects at a level of 50 mg was found to be rapidly absorbed and
    excreted. Excretion was somewhat slower than measured in the rat with
    the major quantity of material being excreted within 1.5 days.
    Preliminary studies suggest that 2,6-dichloro-4-nitroaniline
    metabolites are similar to those obtained from the rat. Approximately
    85% of the total urinary excretion from the rat was found to be of
    2,6-dichloro-4 hydroxy aniline sulfate.
    2,6-Dichloro-4-nitroaniline-14C administered to male rats at a
    dosage of either 1.7 mg/kg or 8 mg/kg orally was rapidly excreted
    from the body. Urinary excretion accounted for approximately 90% of
    the recovered material with the remainder being located in the
    faeces. The majority of material (about 90%) was recovered within 48
    hours and over half was recovered within 8 hours after treatment.
    2,6-Dichloro-4-nitroaniline was not observed in any body tissues with
    the exception of small quantities detected in the G.I. tract, urinary
    tract, and liver (Eberts, 1965).

         2,6-Dichloro-4-nitroaniline administered to rats (ip or orally,
    20 mg/kg) was metabolized to the dichloroamimophenol and the
    dichlorophenylenediamine derivatives. Following both routes of
    administration the majority of material was recovered from the urine
    within 24 hours and total recovery was noted within 72 hours. Very
    small quantities were obtained in the faeces. In vitro studies using
    mouse liver microsomes showed limited conversion of
    2,6-dichloro-4-nitroaniline to the same two metabolites (Mt et al.,
    1967).

         Biotransformations in plants and micro-organisms are discussed
    under "Fate of residues".

    Effects on enzymes and other biochemical parameters

         Following high level subacute oral administration of
    2,6-dichloro-4-nitroaniline to rats, an increase of hepatic
    demethylase and desulfurase activity was noted. Liver mitochondrial
    O2 consumption was also increased in rats. The liver enzymes were
    not stimulated in monkey following similar treatment (Serrone, 1967).

    TOXICOLOGICAL STUDIES

    Special studies on carcinogenicity

    Mouse

         A carcinogenic screening using high levels of
    2,6-dichloro-4-nitroaniline administered to susceptible mice was
    negative. Groups of mice (18 males and 18 females of each of two
    hybrid strains) were administered 2,6-dichloro-4-nitroaniline at 215
    mg/kg/day for three weeks from day seven after birth. Thereafter for
    18 months the mice were fed 603 ppm in the diet, sacrificed and
    examined for tumours. 2,6-Dichloro-4-nitroaniline did not cause a
    significant increase in tumours (Innes et al., 1969).

    Effects on blood and blood forming tissues

    Cat

         In studies to substantiate the difference between effects of
    4-nitroanaline and 2,6-dichloro-4-nitroaniline, a study on the effect
    of these materials on methaemoglobinemia in the cat was performed
    (Gurd, 1974). After a single oral dose of 2,6-dichloro-4-nitroaniline
    (500 mg/kg), no methaemoglobin was noted at any time between 1 and 48
    hours after dosing. Administration of 4-nitroaniline at a single dose
    of 100 mg/kg resulted in methaemoglobin, observed over the same time
    course. In addition, the cats subjected to this experiment were noted
    to be cyanotic and have extensive muscle weakness following
    4-nitroaniline.

    Ocular toxicity

    Dog and miniature swine

         Dogs have been shown to develop lesions in the cornea and lens
    following prolonged oral administration of
    2,6-dichloro-4-nitroaniline. It has been suggested that a
    photochemical product reaction may be responsible for the lesion as
    it occurred only when dogs were exposed to sun light.

         Dogs and miniature swine were fed 2,6-dichloro-4-nitroaniline in
    the diet at levels of 0, 0.75, 6, 24, 48 and 192 mg/kg/day for periods
    of time varying from 50 to 306 days. Corneal opacity appeared in dogs
    within 53 to 104 days after administration of 24 or 48 mg/kg, and when
    exposed to sunlight. Dogs unexposed to sunlight and those with one eye

    sutured closed failed to develop lesions in the unexposed eyes. Dogs
    administered 192 mg/kg refused to eat after 38 days and were
    administered 2,6-dichloro-4-nitroaniline by capsule. All of these
    animals died 49 - 53 days after the study began. Eye lesions were not
    detected in this high level group. Several dogs showing eye damage
    were maintained for four months after 2,6-dichloro-4-nitroaniline
    administration had ceased. Pathological changes seen in the cornea and
    lens were not reversible. Administration of
    2,6-dichloro-4-nitroaniline at all levels did not appear to affect
    miniature swine. Sporadic instances of the presence of Heinz bodies
    in blood were observed in both swine and dogs. Administration of
    2,6-dichloro-4-nitroaniline as dust or 5% solution directly to the
    eyes for three months had no effect on corneal opacity or irritation
    of the conjunctiva (Earl et al., 1971; Bernstein et al. 1970; Curtis
    et al. 1968).

    Special studies on reproduction

    Rat

         In a standard three generation, two litters per generation,
    reproduction study, 2,6-dichloro-4-nitroaniline was administered to
    rats (20 males and 20 females per group) at levels of 0 and 100 ppm in
    the diet. On the basis of the reproduction parameters examined,
    including number of litters, stillbirths, live births, birthweight,
    lactation indices, etc. no evidence of an effect of dicloran on
    reproduction was indicated (Lobdell and Johnston, 1965).

         Male rats were fed 2,6-dichloro-4-nitroaniline in the diet at
    levels of 0, 1000, and 2000 ppm for 90 days. The males were mated with
    untreated females. There were no differences observed in the number of
    litters or in the number of animals born or weaned. Feeding
    2,6-dichloro-4-nitroaniline in the diet to male rats resulted in an
    increased liver weight at 1000 ppm. An increased kidney weight was
    seen at 1000 ppm (EPA, 1974).

         Female rats were fed 2,6-dichloro-4-nitroaniline in the diet at
    levels of 0, 500, and 1000 ppm for 188 days prior to mating. The rats
    were continued on the diet through gestation and lactation. From the
    small number of animals in this experiment (10 females/group), it is
    difficult to make definitive conclusions concerning the effect on
    reproduction of 2,6-dichloro-4-nitroaniline administered to females.
    The data suggested a reduced number of pups when the females were fed
    a level of 1000 ppm in the diet. There was no apparent effect on
    survival of pups, although the mean body weight of pups at 1000 ppm
    was slightly reduced. It might be considered that 1000 ppm in the diet
    of females for six months might have a slight effect on reproduction.
    No effects were seen at 500 ppm (EPA, 1974).

    Rabbit

         Groups of pregnant New Zealand white rabbits (10, 12, and 14 does
    respectively) were fed 2,6-dichloro-4-nitroaniline in the diet at 0,

    100, and 1000 ppm from day 8 until day 16 of gestation. In no case was
    there evidence of an adverse effect of 2,6-dichloro-4-nitroaniline on
    reproduction, affecting either the parents or the offspring (Anonymous
    1974).

    Sensitization

    Rat, guinea pig, rabbit

         An examination was made of the potential skin sensitization
    properties of 2,6-dichloro-4-nitroaniline in guinea pigs. Ten
    subcutaneous injections were administered to male guinea pigs (total
    dose 0.95 mg). Two weeks after the last injection a re-injection of
    0.05 mg was made. Twenty-four hour readings showed no apparent
    sensitization (Johnston and Sweickert, 1963)

         Rats, guinea pigs and rabbits were administered
    2,6-dichloro-4-nitroaniline via inhalation exposure to an 8% dust
    for seven hours. It was estimated that the exposure averaged 0.4
    mg/l. No deaths were observed although reddening of the lungs and
    pale kidneys were seen in rats and guinea pigs (Horn, 1961).

    TABLE 1  Acute toxicity of 2,6-dichloro-4-nitroaniline
                                                                           
                                  LD50
    Species          Route        (mg/kg)                  References
                                                                           
    Rat              oral         4000-10 000              Serrone, 1967
                                                           Lessel, 1974a
                                                           Feenstra, 1961

                     Ip           1460-5471                Serrone, 1967
                                                           Lessel, 1974a
                                                           Feenstra, 1961

                     SC           >5000                    Lessel, 1974a

    Guinea Pig       oral         1450                     Lessel, 1974a

    Mouse            oral         1500-2500                Lessel, 1974a
                                                           Feenstra, 1961

                     Ip           2500-8000                Lessel, 1974a
                                                           Feenstra, 1961

                     SC           >6000                    Lessel, 1974a

                     dermal       >5000                    Lessel, 1974a

    Cat              oral         >500                     Lessel, 1974a

                     oral         200 x 7 daily doses      Lessel, 1974a
                                                                           

         Signs of poisoning in the mouse included defaecation and
    urination, depression and lethargy leading to sleep. In rats the same
    signs were noted including nasal haemorrhage and paralysis. Death
    occurred up to four days after administration of 2,6-dichloro-4-
    nitroaniline (Feenstra, 1961). Cyanosis, muscle weakness, and other
    typical signs of aniline poisoning were not observed.

         Two formulations (8% dust and 50% W.P.) were applied to intact
    and abraded skin of rabbits daily for five days. No irritation of skin
    was observed (Johnston and Schwikert 1961a). When these materials were
    instilled into the conjunctival sac of rabbits, no ocular irritation
    was noted (Johnston and Schwikert, 1961b).

    Short Term Studies

    Rat

         Groups of rats (either 10 males and females or 15 males and
    females per group) were treated with 2,6-dichloro-4-nitroaniline to
    examine potential haematopoietic effects. One series of animals was
    administered 2,6-dichloro-4-nitroaniline at levels of 0, 5, 20 and 100
    mg/kg/day by gavage for four months. Another group was administered
    2,6-dichloro-4-nitroaniline in the diet at levels of 0 and 20 ppm for
    four months. Haematological examinations (RBC, total and differential
    leucocyte, platelet, haematocrit and haemoglobin concentration), blood
    sugar, as well as growth and food consumption data indicated no
    significant effects attributable to 2,6-dichloro-4-nitroaniline at any
    of the dose levels or treatments (Evans et al., 1963).

         Groups of rats were fed 2,6-dichloro-4-nitroaniline (either
    technical or recrystallized material) in the diet for six months.
    Groups of 15 males and 15 females were fed 30 and 300 ppm, groups of
    10 males and 10 females were fed 3000 ppm of either the technical or
    pure material; and groups of 25 males and 25 females were fed a
    control diet. At 3000 ppm in the diet, growth of both males and
    females fed technical 2,6-dichloro-4-nitroaniline was impaired with
    only the males fed the purified material showing a slight reduction in
    growth. In both high level groups livers were enlarged at 3000 ppm.
    There was no effect on haematology or on tissues and organs. There was
    no sign of damage when the tissues were examined microscopically. No
    effects were noted at 300 ppm over the six month period (Lessel,
    1974d).

         Groups of rats (5 males and 5 females per group) were
    administered 2,6-dichloro-4-nitroaniline orally at doses of 0, 140 and
    350 mg/kg/day, 6 days/week, for four weeks. Growth of males was
    reduced at 350 mg/kg. At both doses, liver enlargement and
    histological changes were noted. There was no apparent effect on blood
    parameters or on the kidneys when examined at the termination of the
    experiment (Lessel, 1974c).

         Groups of rats (10 males and 10 females per group, 20 of each sex
    were used in the controls) were administered
    2,6-dichloro-4-nitroaniline orally at dose levels of 0, 35, 140 and
    350 mg/kg/day, 6 days/week, for four weeks. Growth depression was
    observed in both males and females at the highest dose level. Growth
    depression was also noted at the intermediate level in males only.
    Liver enlargement was again observed at 140 mg/kg. No effects were
    noted at 35 mg/kg. Microscopic examination revealed the presence of
    enlarged liver cells with increased vacuolization especially at the
    outer lobes. Some animals were maintained for two weeks after the
    conclusion of the treatment. After this two week period, liver size
    was normal in all but the highest male dosage level. Liver hypertrophy
    caused by repeated short term dosing is apparently reversible within a
    two week period on cessation of treatment. In this study daily acute
    administration 35 mg/kg was observed to have no effect on the rat
    (Lessel, 1974c).

         Because of the known effect of 4-nitroaniline in inducing
    specific blood dyscrasias, subacute feeding experiments were performed
    to compare 2,6-dichloro-4-nitroaniline with this material. Groups of
    weanling rats (5 males and 5 females/group) were administered
    2,6-dichloro-4-nitroaniline by gavage at 0 and 400 mg/kg, 5 days/week
    for four weeks. 4-Nitro-aniline was administered at 200 and 400 mg/kg
    to two other comparable groups over the same interval. In a second
    experiment, groups of male weanling rats (6 rats per group) were
    administered 2,6-dichloro-4-nitroaniline by oral gavage at 0 and 400
    mg/kg and 4-nitroaniline at 200 and 400 mg/kg, twice daily, 5
    days/week, for two weeks. With 2,6-dichloro-4-nitroaniline at 400
    mg/kg haematology was normal - no Heinz bodies were detected and the
    reticulocyte count was normal. At the 800 mg/kg dose there was a
    slight weight loss. The red blood cell and haemoglobin counts were
    normal while lymphocyte counts were slightly reduced. It was noted at
    the conclusion of the study that there was no effect of this compound
    on the spleen. In contrast, 4-nitroaniline had definitive effects on
    growth at 200 mg/kg per day. Heinz bodies were identified in the blood
    and the reticulocyte count was greatly elevated (marked
    reticulocytosis). Bone marrow was not affected. At 200 mg/kg (2x/day)
    there was a reduction of growth, reduced RBC count (with polychromasia
    and nucleation) accompanied by an enlarged spleen. These effects were
    more pronounced at the higher dose where, in addition, a haemoglobin
    was reduced and the lymphocyte count greatly increased. At high
    levels, although 2,6-dichloro-4-nitroaniline caused lymphopenia, the
    hemotoxic effects normally associated with 4-nitroaniline were not
    observed (Lessel, 1974b).

         Mortality was observed when rats were administered
    2,6-dichloro-4-nitroaniline at 1000 mg/kg. No mortality was noted when
    400 mg/kg was administered for three months. Liver and kidney changes
    were observed with light and electron microscopic examinations
    (Serrone, 1967).

    Dog

         Groups of dogs (8 males and 8 females per group) were fed
    2,6-dichloro-4-nitroaniline in the dry diet at levels of 0, 20, 100
    and 3000 ppm for two years. One female dog at 3000 ppm died at 74
    weeks. This death war attributable to the presence of
    2,6-dichloro-4-nitroaniline in the diet. One male control dog lost
    considerable weight but survived to the end of the experiment. No
    compound-related changes in behaviour, food consumption, or growth
    were observed. A watery lacrimation was noted for all dogs at 3000 ppm
    which persisted during the entire testing interval. A yellowing of the
    sclera, mucous membranes, and abdominal skin was noted at the high
    level of feeding. The dog that died showed a picture of haemolytic
    anemia prior to death (reduced haemoglobin, immature erythrocytes,
    polychromophylic macrocytes, increased leucocyte count and increased
    M:E ratio in the bone marrow). At 3000 ppm, clinical chemistry was
    altered in both males and females with an elevation observed in the
    activities of the SGOT and SGPT enzymes, a reduced blood protein,
    increased prothrombin time, BUN, BSP and urinary albumin content. At
    the conclusion of the study, gross and microscopic examination
    revealed an increase in liver weight accompanied by histological
    changes at 3000 ppm in the diet. Histological changes in the animals
    fed 3000 ppm in the diet included irregular hepatic cell size, hepatic
    cell hypertrophy and increased pigmentation of hepatic cells and of
    liver macrophages. Slight changes at 100 ppm were noted in two dogs. A
    no-effect level in this study is estimated to be between 100 and 3000
    ppm in the dry diet (Woodard et al., 1964).

    Monkey

         Daily oral administration to Rhesus monkey at 160 mg/kg was
    lethal within three months with a greater effect noted on females than
    males. Coloration of monkey urine differed from rat urine suggesting a
    difference in metabolism in the two species. Liver and kidney changes
    were observed after light and electron microscopic examination.
    Centrolobular fatty degeneration was observed. Swelling of
    mitochondria with distortion of the cristal was also observed. There
    were differences in the comparative effects of
    2,6-dichloro-4-nitroaniline on liver metabolizing enzymes of monkey
    and rat (Serrone, 1967).

    Long-Term Studies

    Rat

         Groups of rats (35 males and 35 females/group) were fed
    2,6-dichloro-4-nitroaniline in the diet at levels of 0, 20, 100 and
    3000 ppm for two years. At 100 ppm there was no effect on behaviour,
    mortality or growth. At this level all values from treated animals
    were comparable to control values. Growth and food consumption of both
    males and females was depressed at 3000 ppm. Haematological parameters
    (haemoglobin and packed cell volume) were reduced at 3000 ppm. These
    haematological changes were noted only after the first year of

    treatment. Gross and microscopic examination performed at 13 weeks and
    at the conclusion of study showed slightly higher liver weights,
    kidney weights, testicular weights and (possibly) thyroid weights at
    3000 ppm. The incidence and location of neoplasms in all treatments
    did not differ from those in the controls. Histological examination
    revealed liver changes at 3000 ppm, characterized by hepatic cell
    enlargement, glycogen depletion, increased basophilia of the
    cytoplasm, and the presence of necrobiotic hepatic cells. Histological
    examination performed at 13 weeks also indicated hepatic cell changes
    and slight adrenal cortical atrophy in several animals at 3000 ppm.
    The adrenal changes were not noted at 104 weeks. An estimated
    no-effect level in this study is between 100 and 3000 ppm in the diet
    (Woodard et al., 1964).

         Groups of rats (25 males and 25 females/group, Boots-Wistar
    strain) were fed 2,6-dichloro-4-nitro-aniline in the diet at
    concentrations of 0 and 1000 ppm for two years. There was no effect on
    survival, food consumption, growth, haematology, or upon gross and
    histological appearance of tissues and organs at the conclusion of the
    study. There were no differences in the size or cellular makeup of
    liver, kidney, or spleen. The incidence of tumors in the control and
    treatment group was similar. From the results of this experiment a
    suggested no-effect level is greater than 1000 ppm (Lessel, 1974e).

    OBSERVATIONS IN MAN

         In a clinical double blind study, two groups of adult males were
    administered 2,6-dichloro-4-nitroaniline (20 individuals) or a placebo
    (10 individuals) once a day for ninety days.
    2,6-Dichloro-4-nitroaniline was administered at a level of 10 mg per
    day. Hematological, liver function, and kidney function tests were
    performed at various intervals over the course of the study and were
    found to be normal. There were no indications that administration of
    2,6-dichloro-4-nitroaniline at 10 mg per day to adult males had any
    adverse effect (Stough, 1962).

         Extensive examinations were made on one industrial worker
    occupationally exposed to 2,6-dichloro-4-nitroaniline over a period of
    three years. It was reported that for about 60 days per year
    considerable inhalation and dermal exposure had occurred. No adverse
    effects were observed with the individual (Brooks and Boyack, 1963).

         Another investigation in man on the potential ocular problem
    associated with 2,6-dichloro-4-nitroaniline was again negative
    (Manger, 1972).

    COMMENTS

         2,6-Dichloro-4-nitroaniline has a low order of toxicity to
    mammals, including man. 2,6-Dichloro-4-nitroaniline is rapidly
    metabolized in plants, and fragments of the molecule are
    reincorporated as natural plant constituents. In mammals,
    2,6-dichloro-4-nitroaniline is rapidly absorbed, metabolized and
    excreted. Metabolism in mammals results in formation of the
    chlorinated phenylenediamine and aminophenol which are conjugated and
    excreted. 2,6-Dichloro-4-nitroaniline does not induce
    methemoglobinemia as evidenced with 4-nitroaniline.
    2,6-Dichloro-4-nitroaniline does not affect reproduction in rodents
    and has shown no evidence of being a teratogen under the experimental
    protocol used. Following subacute feeding, dogs exposed to sunlight
    developed cataracts. Specific experiments with rabbits, rats and
    swine did not duplicate these results. Long term feeding studies in
    rat and two year feeding studies in dog resulted in growth
    retardation accompanied by an increased liver and kidney size at
    high levels. No-effect levels based on a two year dog study and short
    and long term rat studies formed the basis for allocating a temporary
    ADI for man. A short term study in man is reassuring in estimation of
    the temporary ADI although no conclusions could be drawn on the
    possibility of ocular damage to man.

    TOXICOLOGICAL EVALUATION

    Level causing no toxicological effects

         Rat: 1000 ppm in the diet, equivalent to 50 mg/kg bw.

         Dog: 100 ppm in the diet, equivalent to 2.5 mg/kg bw.

    ESTIMATE OF TEMPORARY ACCEPTABLE DAILY INTAKE FOR
    MAN

         0 - 0.03 mg/kg bw.

    RESIDUES IN FOOD AND THEIR EVALUATION

    USE PATTERN

         Registration of 2,6-dichloro-4-nitroaniline as a commercial
    fungicide is recorded in Canada, France, Holland, Italy, Japan, New
    Zealand, South Africa and USA.

         It is claimed to be effective against several Basidiomycetes
    and Deuteromycetes species of fungi, being fungistatic to the
    mycelium and spores of Botrytis cenera (Clark et al., 1960; Clark
    and Hams, 1961; Sharples, 1962) and to several Rhizopus fruit rot
    fungi (Ogawa et al., 1961, 1962; Cappelini and Stretch, 1962).

         It is mostly marketed as 4% or 8% (w/w) dust formulations or as
    50% (w/w) wettable powders, alone or mixed with thiram (TMTD) or
    pentachloronitrobenzene. Smoke formulations containing 40% dicloran
    are also available.

         Pre-harvest treatments recommended by the manufacturers include
    soil treatments for lettuce under glass, dusting or spraying of soft
    fruits, cotton, leafy vegetables, strawberries, onions, garlic,
    tomatoes and ornamentals. Post-harvest dips of peaches, nectarines and
    carrots are practised at rates of 750-1000 mg/kg a.i.

    Phytotoxicity

         Under extreme experimental conditions a "bronzing" discolouration
    of lettuce leaves and marginal "off-taste" taints in fruits have
    occasionally been found, but none of these effects have ever been
    reported in practical use.

    RESIDUES RESULTING FROM SUPERVISED TRIALS

         Residue data on 2,6-dichloro-4-nitroaniline in several fruit and
    vegetable crops have been presented by the originating company (The
    Boots Company Ltd., 1972). The data derive from field trials and
    post-harvest experiments carried out mainly in the USA and UK.
    Summaries of residue ranges and experimental conditions extracted from
    these data are presented in Tables 2, 3 and 4.

    Peaches and apricots

         Extensive trials with single or repeated spraying of peaches and
    apricots at recommended dosage rates (usually 0.12-0.18% a.i.) and
    higher rates have been carried out during the period from 1960 to 1965
    (Tables 2 and 4). The initial deposits of 2,6-dichloro-4-nitroaniline
    on peaches are generally between 5 and 15 mg/kg with occasional values
    up to 25-30 mg/kg. A typical residue was 5 mg/kg after 6 days, 3 mg/kg
    after 9 days and 1.5 mg/kg after 14 days. The highest recorded residue
    14 days after treatment at a dosage rate of 0.18% was 4.9 mg/kg.

         The average half-life of 2,6-dichloro-4-nitroaniline on peaches
    has been calculated to be 5 days.

         Spraying apricots gave residues very similar to those found in
    sprayed peaches (Table 4).

         Dusting peaches using 8% a.i. dicloran powders at rates of 3-5 kg
    per ha gave maximum initial deposits of 3.7 mg/kg.

         A suggested practice of wrapping apricots and peaches in tissues
    impregnated with 2,6-dichloro-4-nitroaniline (from 500-3000 mg/kg, in
    the wrap) showed a definite transfer of the chemical to the fruits.
    Residues from such post harvest application were from 1.6 - 3.5 mg/kg
    in peaches and 1.9 - 3.3 mg/kg in apricots (Table 3).

    TABLE 2  Residues of dicloran in peaches
                                                                             

    Application                 Number         Number    Number     Range of
    (% a.i. w/v or    PHI       of             of        of         results
    kg a.i./ha)       (days)    applications   trials    results     mg/kg
                                                                             

    0.06%             3         3              1         4          0.1-0.3

    0.09%             0         1-3            3         9          6.1-16.5

                      1         1-3            5         9          1.5-11.6

                      2-3       1-3            2         7          0.2-4.7

                      7         1              1         1          2.3

    0.12%             0         1-3            1         6          10.8-14.0

                      1         1-2            1         2          3.4-5.5

                      3         1-3            2         10         0.2-6.3

                      7         1              1         1          3.2

    0.18%             0         2              1         6          6.0-11.4

                      1         2              1         6          9.2-13.7

                      2         2              1         6          5.7-13.8

                      4         2              1         6          4.2-9.4

                      6         2              1         6          4.5-7.8

                      9         2              1         6          1.9-6.0

                      14        2              1         6          0.6-4.9

    0.18%             0         3              1         4          18.9-29.5

                      3         3              1         4          22.4-29.6

    0.24%             1         3              1         4          3.0-8.3

                      3         3              1         4          2.7-11.1

    3.1 kg/ha
    (8% dust)         0         1              1         2          0.14-0.35

    4.5-4.9 kg/ha
    (8% dust)         0         1              3         21         0.22-3.7
                                                                             

    TABLE 3  Residues in fruits wrapped in tissues impregnated with
             dicloran

                                                                            

    Fruit               Dicloran in wrap mg/kg         Dicloran in fruits
                                                                            

    Apricot             500                            1.9

                        1000                           2.6

                        2000                           3.3

    Peach               500                            1.6

                        1000                           2.3

                        2000                           3.8

                        3000                           3.5
                                                                            

    
    TABLE 4  Residues of 2.6-dichloro-4-nitroaniline in fruit and vegetables

                                                                                                 

                                               Number          Number    Number      Range of
                                   PHI         of              of        of           results
    Crop            Application    (days)      application     trials    results       (ppm)
                                                                                                 

    Apricots        0.09%          0           1               1         12          1.9-7.9

                                   2           3               1         2           0.9-1.4

                                   4           1-2             1         4           1.4-3.7

                                   11          1               1         2           0.1-0.7

    Apricots        0.12%          0           1               1         1           10.1

                                   1           1               1         1           8.0

                                   7           1               1         1           3.8

    Cherry          0.12%          0           1               1         1           10.3

                                   1           1               2         5           6.4-14.8

                                   7           3               1         1           4.9

    TABLE 4  (Cont'd.)

                                                                                                 

                                               Number          Number    Number      Range of
                                   PHI         of              of        of           results
    Crop            Application    (days)      application     trials    results       (ppm)
                                                                                                 

                    0.24%          1           1               1         2           0.7-11.2

                                   7           1               1         1           2.9

                    0.36%          1           5               1         2           2.7-3.1

                    1200 ppm       post-harvest                6         21          1.5-6.4

                    1200 ppm       post-harvest
                                   and washing                 1         2           0.3-0.3

                    1350 ppm       post-harvest                1         2           10.2-13.7

                    1800 ppm       post-harvest                1         2           7.7-8.4

    Grapes          1.7 kg/ha
                    (dust)         0           2-4             2         6           0.2-3.9

                                   7           2               1         2           0.3-1.1

                    2.3-3.3
                    kg/ha (dust)   1           4               1         4           1.1-1.3

                                   7           1-2             2         4           0.2-1.1

                    1000 ppm       post-harvest dip            1         4           0.6-5.1
    Plums
    (dried)         1000 ppm       post-harvest dip            1         3           1.0-4.7*

                                   post-harvest dust           1         3           2.0-8.1

                    0.12%          1           1               1         1           2.5

    Strawberry      0.16-0.18%     1           4               1         1           0.32

                                   3           1               1         2           4.0-5.7

                                   9-11        3-4             2         4           0.2-1.6

                    0.24%          1           3               1         2           0.9-1.7

    Blackberry      1.1 kg/ha
                    (spray)        4-11        1               1         2           1.2-2.3

    TABLE 4  (Cont'd.)

                                                                                                 

                                               Number          Number    Number      Range of
                                   PHI         of              of        of           results
    Crop            Application    (days)      application     trials    results       (ppm)
                                                                                                 

                    1.8 kg/ha
                    (dust)         4-11        1               1         2           <0.05-0.3

    Currants        0.11-0.15%     33          4               1         3           1.7-3.3

    Raspberry       0.15-0.18%     5           4               2         2           17.0-20.2

                                   9-10        4-5             2         2           2.2-11.8

                                   13-14       3-4             2         2           2.6-7.5

                    0.36%          5           4               1         1           39.0

                                   10          4               1         1           28.4

                                   14          4               1         1           16.3

    Carrots         900-           post-harvest dip
                    1000 ppm       (stored 0-3 days)           2         12          3.6-9.4

                                   peeled                      1         1           2.1

                                   canned                      1         7           <0.05-0.2

    Lettuce         2.2 kg/ha      0           2               1         4           50.1 (average)

                                   7           2               1         9           18.0     "

                    RL50 approx    14          2               1         9           3.0      "
                    3-4 days

                                   21          2               1         9           2.4      "

                                   28          2               1         8           0.8      "

                                   35          2               1         8           0.04     "

                    1.3 g/m2
                    (dust)         120         1               1         1           0.1

                    1.3 g/m2 (soil
                    treatment;
                    pre-planting)  114-140     1               3         5           0.2-2.1

    TABLE 4  (Cont'd.)

                                                                                                 

                                               Number          Number    Number      Range of
                                   PHI         of              of        of           results
    Crop            Application    (days)      application     trials    results       (ppm)
                                                                                                 

    Gherkins
    (indoor)        0.05%          0           1               1         1           5.5

                                   2           1               1         1           1.1

                                   4           1               1         1           0.6

                                   5           1               1         1           0.35

    Tomato
    (indoor)        0.1%           2           1               1         1           0.25

                                   4           1               1         1           0.2

                                   6           1               1         1           0.04

    Beans,
    French          3 kg/ha        12          4               1         2           1.3-1.9

                                   20          2               1         2           0.8-1.5
                                                                                                 

    *  Loss by drying.
    

    Cherries, grapes and plums

         The residue levels of 2,6-dichloro-4-nitroaniline from
    recommended uses on these fruits are similar to those in peaches and
    apricots (Table 4).

         After dipping procedures or post-harvest dusting of plums,
    dicloran was retained on the fruits at levels of 4.7 and 8.1 mg/kg
    respectively. These residues were reduced to 1.0 and 2.0 mg/kg
    respectively, when the fruits were subjected to an 8 day air-drying
    period at ambient temperature.

         A post-harvest treatment of cherries by spraying on the packing
    line with 1200-1800 ppm suspensions gave residues of
    2,6-dichloro-4-nitroaniline from 1.5 - 13.7 mg/kg. In storage
    experiments these residues were reduced at a rate corresponding to a
    half-life of 11 days at 20C.

    Small fruits and berries

         Repeated spraying of strawberries at different locations (0.06
    - 0.24% a.i.) have been reported to give maximum initial deposits of
    dicloran of 9 mg/kg. However, residues at harvest after a 9 - 11 day
    interval from the last application were generally well below 1 mg/kg,
    with an occasional individual sample at 1.6 mg/kg. Among the berries,
    raspberries in some cases showed higher residues. After application of
    0.36% a.i. the initial residue was 39 mg/kg which was reduced to 16.3
    mg/kg at 14 days. At lower application rates, residues were
    proportionately lower.

         Experiments with other berries (blackberries, currants and
    boysenberries) generally gave residues below 5 mg/kg.

    Lettuce

         Roburn (1960) followed the disappearance of
    2,6-dichloro-4-nitroaniline after post-planting treatments of lettuce
    plants with 1/4 oz of 4% dust per sq. yd. and found a reduction in
    deposits greater than that due to growth dilution. Volatilization and
    chemical degradation were suggested as contributing factors.

         The half-life values decreased progressively in these trials,
    being about 3-5 days during the last weeks of the growing season. A
    three weeks pre-harvest interval for dicloran treatments of lettuce is
    recommended on this basis in the United Kingdom.

         Later experiments by Boyack and Boot (1962 a, b, c) confirmed
    Roburn's findings, indicating half-life values of 3-4 days on
    greenhouse lettuce with average residues of 2.4 mg/kg after an
    interval of 21 days (Table 4).

         Uptake of 2,6-dichloro-4-nitroaniline through the roots of
    lettuce plants after pre-planting soil treatments has been
    demonstrated by Boyack and Boot (1962a), Lemin and co-workers (Lemin,
    1963, 1965; Moe and Lemin, 1963a, 1964) and Groves and Chough (1970).
    Residues from such practices at the time of harvest will be near or
    below the detection limit, i.e. less than 0.05-0.1 mg/kg.

         In recent experiments in the Netherlands lettuces have been
    treated both by a pre-planting soil application and by dusting the
    young plants immediately after planting. (Pieters, 1974). Residues in
    these experiments ranged from 0.1 - 2.1 mg/kg (Table 4).

    Carrots

         Post-harvest dipping in 900-1000 ppm suspensions caused residues
    of 3.6-9.4 mg/kg at 0-3 days. After storage for 5 months the remaining
    residues were 2.1-3.7 mg/kg.

    Other vegetables

         Residue data on a few other vegetable crops, e.g. tomatoes,
    gherkins and French beans, mostly grown under glasshouse conditions,
    are available (Pieters, 1974). The data are insufficient to evaluate
    quantitatively the rates of dissipation, but residues are generally
    low at the time of harvest, i.e. from a few days to 1-2 weeks after
    treatment (Table 4).

    FATE OF RESIDUES

    In animals

         The metabolism of 2,6-dichloro-4-nitroaniline by the rat, mouse
    and to a limited extent man, has been described above under
    "Biotransformation". No such experimental evidence is available on
    livestock animals, nor on the excretion of dicloran or its metabolites
    in milk.

    In plants

         Studies by Lemin (1965), Lemin et al. (1963) and Moe and Lemin
    (1963a, 1964) have shown that 2,6-dichloro-4-nitroaniline is absorbed
    by plant roots and translocated to the plant material in tomato and
    lettuce. These experiments include uptake from both nutrient solutions
    and treated soils.

         Application of 14C-labelled 2,6-dichloro-4-nitroaniline gave
    evidence of rapid degradation into polar metabolites and 14C was
    found in carbohydrate constituents of the plant tissue, presumably
    owing to the incorporation of degradation fragments. Transitional
    metabolites such as dechlorinated components, reduction products or
    2,6-dichloro-p-phenylene-diamine were not detected.

         Amino acids, chlorophyll or uronic acid did not contain
    radioactivity. Neither could 2,6-dichloro-4-aminophenol, known to be
    formed by animal metabolism (see previous section), be found in plant
    tissues.

         Exhaustive extractions of peaches 13 days after treatment with
    14C-labelled 2,6-dichloro-4-nitroaniline revealed (in addition to
    unchanged parent compound) 14C-labelled phenylalanine and labelled
    flavonoid glycosides (Moe and Lemin, 1963b). As in the above
    experiments, no transitional metabolites could be traced.

         Groves and Chough (1970) report rapid absorption of dicloran and
    incorporation of fragments into tissue constituents by a number of
    plants.

    In processing and storage

         Washing or canning processes have been shown to reduce residues
    of 2,6-dichloro-4-nitroaniline considerably (Boots Company Ltd.,
    1972). On lettuce, for example, residues of 18-25 mg/kg still
    remaining two weeks after the last application were reduced to 5.6-6.0
    by washing with water (Roburn, 1958). In peaches, freshly treated with
    2,6-dichloro-4-nitroaniline, the residues which still remained after
    an industrial canning procedure were below 0.1 mg/kg.

         Carrots treated by post-harvest dipping showed only moderate
    reduction of the residues by peeling, from 3.7 to 2.1 mg/kg,
    indicating a significant penetration into the inner parts. A
    subsequent canning procedure, however, reduced the levels to 0.1-0.2
    mg/kg. Storage of dipped carrots for 5 months at 40F gave a decrease
    in the residue of about 25%.

    In soils

         2,6-Dichloro-4-nitroaniline is generally regarded as being
    relatively stable in soils at field moisture capacity. Its
    persistence, however, is greatly affected by conditioning factors such
    as soil composition, water content, microculture etc. (Wang and
    Broadbent, 1973). The absorption of the chemical from soils by oats
    was found by Groves and Chough (1970) and Wang (1972) to be inversely
    related to the clay and organic matter content of the soils,
    suggesting a binding of the chemical to these soil constituents. Later
    exhaustive extraction experiments showed that a reversible binding
    probably occurs (Groves and Chough, 1971).

         The same authors (Groves and Chough, 1970, 1971) showed that
    14C-labelled dicloran can be metabolized by soil organisms which
    degrade the compound to 14C-carbon dioxide and other volatile
    compounds. The rate of microbial breakdown could be increased by
    repeated applications of dicloran to the same soil or by incubation of
    the soil with dicloran. A culture of rod-shaped bacteria was isolated
    and found active in the decomposition of the fungicide. Microbial
    breakdown is also held responsible for the rapid degradation of DCNA
    which is seen in soil under flooded conditions (Wang and Broadbent,
    1973).

         Van Alfen and Kosuge (1974) have identified both
    2,6-dichloro-p-phenylenediamine and its acetylated derivative,
    4-amino-3,5-dichloroacetanilide, in cultures of Pseudomonas cepacia
    and Escherichia coli B. In soil and through the action of
    horseradish peroxidase, the phenylenediamine was converted to one of
    three isomers of the azine resulting from oxidative dimerization (Van
    Alfen, 1973).

    EVIDENCE OF RESIDUES IN COMMERCE OR AT CONSUMPTION

         Market sample surveys carried out in the Netherlands in 1973 on
    domestically grown fruit and vegetables showed that residues of
    2,6-dichloro-4-nitroaniline were frequently present in some crops
    (Table 5). In vegetables such as endive, lettuce and chicory sprouts
    about 50-80% of samples were positive with individual residues up to 8
    mg/kg. Crops such as cucumbers, tomatoes and paprika were less
    frequently positive (3-25%). At the time of this survey, the tolerance
    for vegetables in the Netherlands was 10 mg/kg.

    TABLE 5  Dicloran residues in marketed vegetables (Netherlands, 1973)

                                                                             

    Dicloran
    residues,
    mg/kg                                number of samples
    (range)                                                                  
                      Cucumber  Endive   Lettuce   Paprika   Tomato   Chicory

                                                                             

    not detectable    31        34       136       12        30       68

    0.01 - 0.1        1         16       139       2         4        50

    0.1 - 0.3         -         7        126       2         -        21

    0.3 - 1.0         -         8        180       -         -        3

    1.0 - 3.0         -         4        123       -         -        1

    3.0 - 10.0        -         1        35        -         -        -
                                                                             

    Total number      32        70       739       16        34       143

                                                                             

    [text missing]

    pre-harvest treatments on fruits, vegetables and ornamentals and
    post-harvest treatments on peaches, nectarines, cherries and carrots.
    It is registered in several countries, usually as 4 or 8% (w/w) dusts
    or as 50% (w/w) wettable powders.

         The technical grade typically contains about 96%
    2,6-dichloro-4-nitroaniline. The remaining parts consist of chemically
    related compounds and impurities which have been identified and
    quantified.

         Concentrations and rates of application vary, depending on the
    crop and method of application. Normal spraying concentrations are
    0.05% to 0.18% applied at rates of 2-5 kg a.i. per ha. Post-harvest
    dips are practised at rates of 1000 - 2000 mg per litre.

         The residue data available are mainly obtained from the USA and
    UK, in a few cases supplemented from other countries. Most of the data
    derive from field trials or experiments under practical conditions
    likely to represent the results of good agricultural practice.

         2,6-dichloro-4-nitroaniline is a non-systemic compound of
    relatively low persistence. The dissipation of the compound on fruits
    and vegetables is often faster than can be explained by normal
    weathering or growth dilution. Half-life periods of 11 days have been
    found for residues during storage of post-harvest treated cherries.

         2,6-dichloro-4-nitroaniline is absorbed by plant roots from the
    soil. The fate of residues in plant material has been followed with
    the radio-labelled compound, which indicated a rapid degradation into
    polar metabolites and 14CO2 followed by incorporation of 14C into
    normal metabolic plant constituents. No indications of transitional
    metabolites from such degradation have been found.

         Similar degradation of 2,6-dichloro-4-nitroaniline occurs in
    soils, although in this case microbial breakdown may also produce
    smaller amounts of reduction products and acetylated derivatives of
    the parent compound.

         Studies were available on the metabolic fate of residues in rats
    indicating a fairly rapid process of absorption, metabolism and
    excretion, especially as aminophenol and phenylenediamine derivatives,
    in the urine. There was no evidence of tissue storage in these
    experiments. Preliminary studies in man suggest a metabolic pattern
    similar to that of rats. No information, however, has been recorded on
    metabolism in livestock animals or on transfer of residues into meat
    or milk.

    METHODS OF RESIDUE ANALYSIS

         A colorimetric method of residue analysis, based on the
    development of a yellow colour characteristic of some mononitro
    aromatic compounds in the presence of strong alkali and acetone, is
    described by Kilgore et al. (1962). Extraction is with benzene,
    followed by clean-up on Florisil alone or in combination with an
    acetonitrile-petroleum ether partitioning procedure. Blank values for
    fruits were about 0.01-0.02 mg/kg and recoveries in the range 75 -
    100% (average 86%). The colorimetric method described by Groves and
    Chough (1966) and Heagy (1969) has a similar sensitivity, based on
    diazotization and coupling to N-(1-naphthyl)ethylenediamine has a
    similar sensitivity.

         An alternative method developed by Roburn (1961) utilizes the
    reduction of 2,6-dichloro-4-nitroaniline by zinc and hydrochloric acid
    to the corresponding 2,6-dichloro-p-phenylenediamine which, in the
    presence of aniline, can be oxidized to produce an intense blue
    colour. Less than 2 g can be detected by this method, corresponding
    to 0.05 mg/kg in fruit and 0.2 mg/kg in lettuce. Recovery of surface
    deposits was approximately 100%, but for macerated samples it was only
    71%. Chloronitroanilines in general give similar reactions but a
    series of other pesticide chemicals did not interfere, indicating a
    fair degree of specificity of the method.

         A thin-layer chromatographic spot test sensitive to 0.5 g of
    2,6-dichloro-4-nitroaniline is given by Groves et al. (1966) based on
    their colour reaction mentioned above.

         Gas-chromatographic methods for the determination of
    2,6-dichloro-4-nitroaniline have been developed by Beckmann and
    Bevenue (1962) and Kilgore (1964) using electron capture and
    microcoulometric detectors, in combination with the Kilgore extraction
    procedure already mentioned. Recoveries of 85-105% were established by
    Brewerton et al. (1967) for the analysis of dicloran in soils, fruit
    and vegetables down to the 0.1 mg/kg level by the GLC/EC technique.
    These methods could be adapted for regulatory purposes as part of a
    multi-residue procedure.

    NATIONAL TOLERANCES REPORTED TO THE MEETING

                                                                             

                                                                Tolerance
    Country       Crop                                          mg/kg
                                                                             

    Australia     Beans, lettuce, stone fruits, tomatoes        20

    Belgium       Fruit and vegetables (exc. potatoes)          10

    Canada        Apricots, nectarines, peaches
                  (pre-harvest and post-harvest),
                  sweet cherries pre-harvest and
                  post-harvest), snap beans                     20

                  Strawberries, raspberries                     15

                  Celery, grapes, lettuce, rhubarb,
                  sweet potatoes                                10

                  Carrots, cucumbers, garlic, onions,
                  plums, tomatoes                               5

    Germany       All fruits and vegetables                     0.1

                                                                             

                                                                Tolerance
    Country       Crop                                          mg/kg
                                                                             

    Italy         General                                       10

    Switzerland   Lettuce (tentatively)                         0.5

    Netherlands   Lettuce, endive                               3

                  Chicory (sprouts)                             1

                  Cucumbers, gherkins, melons, bell
                  peppers, tomatoes                             0.3

                  Other vegetables                              0.1

    USA           Apricots, nectarines (pre-harvest and
                  post-harvest), peaches (pre-harvest
                  and post-harvest), sweet cherries
                  (pre-harvest and post-harvest), snap
                  beans                                         20

                  Blackberries, boysenberries, celery,
                  plums (fresh prunes) (pre-harvest and
                  post-harvest), raspberries                    15

                  Carrots (pre-harvest and post-harvest),
                  grapes lettuce, rhubarb, sweet
                  potatoes (pre-harvest and post-harvest)       10

                  Cucumbers, garlic, onions, tomatoes           5

                  Potatoes                                      0.25

                  Cotton seed                                   0.1

                                                                             

    APPRAISAL

         2,6-dichloro-4-nitroaniline is a fungicide active against the
    mycelium and spores of Botrytis cinerea, B. sclerotinia and other
    fungi, including several Rhizopus fruit rots.

         Practical use patterns comprise foliar applications and soil
    treatments under indoor and outdoor growing conditions.

         The residue data from supervised trials on several crops were
    considered sufficiently extensive to enable recommendations to be
    made, including some for post-harvest treatments.

         Gas-chromatographic methods combined with appropriate extraction
    and clean-up procedures are available for specific determination.
    These methods are suitable for regulatory purposes.

    RECOMMENDATIONS

         Temporary tolerances are recommended for
    2,6-dichloro-4-nitroaniline in the following commodities at the
    levels indicated. The levels are not likely to be exceeded when
    2,6-dichloro-4-nitroaniline is applied in accordance with good
    agricultural practice including both pre-harvest and post-harvest
    applications and taking into account that preharvest intervals may
    apply for certain treatments.

    TEMPORARY TOLERANCES

                                                                             

                                Limit       Basis on which recommendations
    Crop                        (mg/kg)     are made
                                                                             

    Cherries                    15          pre and post-harvest

    Peaches                     15          pre and post-harvest

    Apricots                    10          pre and post-harvest

    Carrot                      10          post-harvest only

    Grapes                      10          pre and post-harvest

    Lettuce                     10          pre-harvest only

    Plums                       10          pre and post-harvest

    Raspberry                   10          pre-harvest only

    Strawberry                  10          pre-harvest only

    Blackberry                  5           pre-harvest only

    Currant (red, black
    and white)                  5           pre-harvest only

    Beans (French)              2           pre-harvest only

    Gherkins                    0.5         pre-harvest (indoor) only

    Tomato                      0.5         pre-harvest only
                                                                            

    FURTHER WORK OR INFORMATION

    REQUIRED (by 1977)

    1.   Further studies on the ocular disturbance observed in dogs to
    confirm and clarify this effect.

    2.   Information on fate in livestock animals in so far as plant
    material containing residues may be fed to animals.

    DESIRABLE

    1.   Effect on hepatic microsome systems in several species.

    2.   Further observations in man.

    3.   Further information on fate of residues during storage, transport
         and processing of fruit and vegetables.

    4.   Information on transfer of residues from grapes to wine and on
         possible influence on wine processing.

    5.   Further information on soil residues and their possible uptake
         into subsequent crops.

    6.   Further data to clarify inconsistencies in the residue levels
         found in different berries.

    REFERENCES

    Anonymous. (1974) "Somers test" in rabbits. Technical summary reports
    submitted to WHO by UpJohn Co. (Unpublished)

    Beckmann, H. and Bevenue, A. (1962) Pesticide residue analysis Gas
    chromatographic analysis of 2,6-dichloro-4-nitroaniline. J. Food Sci.,
    27:602-604.

    Bernstein, H.N., Curtis, J., Earl, F.L. and Kuwabara, T. (1970)
    Phototoxic corneal and lens opacities in dogs receiving a fungicide,
    2,6-dichloro-4-nitroaniline. Archs Ophthal. (Chicago), 83:336-348.

    Boots Co. Ltd. (1972) Dicloran. Registration Document Vol. I, II, III
    and VI. Reports submitted by The Boots Company Ltd. Agricultural
    Research, Nottingham. (Unpublished)

    Boyack, G. and Boot, D. (1962a) Assay for 2,6-dichloro-4-nitroaniline
    in lettuce grown in treated soil under glass. Dicloran, Registration
    document from The Boots Co. Ltd. Vol. II, Appendix 26.

    Boyack, G, and Boot, D. (1962b) Assay for 2,6-dichloro-4-nitroaniline
    on sprayed greenhouse lettuce. Dicloran, Registration document from
    The Boots Co. Ltd. Vol. II, Appendix 26.

    Boyack, G. and Boot, D. (1962c) Residue of 2,6-dichloro-4-nitroaniline
    (DCNA) on lettuce dusted with Botran. Dicloran, Registration document
    from The Boots Co. Ltd. Vol. II, Appendix 26.

    Brewerton, H.V., Clark, P.J. and McGrath, H.J.W. (1967) Dicloran
    analysis by gas chromatography. New Zealand J. Sci., 10:124-127.

    Brooks, B. and Boyack, G. (1963) Medical examination of a man after
    prolonged exposure to 2,6-dichloro-4-nitroaniline. Report submitted by
    The Upjohn Co. (Unpublished)

    Cappellini, R.A. and Stretch, A.W. (1962) Control of post-harvest
    decays of peaches. Pl. Dis. Reptr., 46:31-33.

    Clark, N.G., Hams, A.F., Higgons, D.J. and Stevenson, H.A. (1960) A
    new fungicide active against Botrytis spp. Chemy Ind., 1960:572-573.

    Clark, N.G. and Hams, A.F. (1961) Antifungal activity of substituted
    nitroanilines and related compounds. J. Sci. Food Agric., 12:751-757.

    Curtis, J., Bernstein, H.N., Earl, F.L. and Smalley, H., Jr. (1968)
    Corneal and lens opacities in dogs treated with
    2,6-dichloro-4-nitroaniline. Toxic. appl. Pharmac., 12:305.

    Earl, F.L., Curtis, J., Bernstein, H.N. and Smalley, H., Jr. (1971)
    Ocular effects in dogs and pigs treated with dichloran. Food Cosmet.
    Toxic., 9:819-828.

    Eberts, F.S. (1965) Fate of Botran (2,6-dichloro-4-nitroaniline) in
    rat and man. Dicloran, Registration Document, Vol. IV, Appendix 54.
    Boots Co. Ltd. (Unpublished). Botran Symposium, Augusta, Mich. 1965.
    Cited in Chemical Abstracts 1968, 68:abstract 11908g

    EPA. (1974) Data from EPA Laboratories. (Unpublished)

    Evans, J., Mengel, G. and Bostwick, L. (1963) Botran(R) (U-2069)
    Effect of oral administration. Final report, four months study. Report
    submitted by the Upjohn Co. (Unpublished)

    Feenstra, E. (1961) Report submitted by The Upjohn Co. (Unpublished)

    Groves, K. and Chough, K.S. (1966) Determination of small quantities
    of 2,6-dichloro-4-nitroaniline (dicloran). J. agr. Food Chem.,
    14:668-669.

    Groves, K. and Chough, K.S. (1970) Fate of the fungicide
    2,6-dichloro-4-nitroaniline (DCNA) in plants and soils. J. agr. Food
    Chem., 18:1127-1128.

    Groves, K. and Chough, K.S. (1971) Extraction of
    3-amino-1,2,4-triazole (Amitrole) and 2,6-dichloro-4-nitroaniline
    (DCNA) from soils. J. agr. Food Chem., 19:840-841.

    Gurd, P.R. (1974) Methaemoglobin formation in the cat. Report from
    Boots Pure Drug Company Limited. Submitted to WHO by the Boots Co.
    Ltd. (Unpublished)

    Heagy, J.A. (1969) Colorimetric determination of
    2,6-dichloro-4-nitroaniline (dicloran) residues in foods. J. Ass. off.
    analyt. Chem., 52:797-799.

    Horn, J. (1961) U-2069 - Acute inhalation toxicity. Study from Woodard
    Research Corporation submitted by The Boots Co., Ltd. (Unpublished)

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

    Johnston, R. and Schwikert, R. (1961a) Skin irritation in rabbits.
    Report from The Upjohn Co. submitted by The Boots Co., Ltd.
    (Unpublished)

    Johnston, R. and Schwikert, R. (1961b) Dichloran - eye irritation in
    rabbits. Report from The Upjohn Co. submitted by The Boots Co., Ltd.
    (Unpublished)

    Johnston, R. and Schwikert, R. (1963) Skin sensitization in guinea
    pigs. Report from The Upjohn Co. (Unpublished)

    Kilgore, W.W. (1964) 2,6-dichloro-4-nitroaniline. Analytical methods
    for pesticides, plant growth regulators and food additives. Gunter
    Zweig (Editor). Vol. III. p. 61-68.

    Kilgore, W.W., Cheng, K.W. and Ogawa, J.M. (1962) Extraction and
    determination of 2,6-dichloro-4-nitroaniline in processed fruits. J.
    agr. Food Chem., 10:399-401.

    Lemin, A.J. (1965) Translocation and metabolism of
    2,6-dichloro-4-nitroaniline by lettuce and tomato. J. agr. Food Chem.,
    13:557-560.

    Lemin, A.J. Moe, L.D. and Smith, A.H. (1963) The metabolism of
    2,6-dichloro-4-nitroaniline by bibb lettuce. Dicloran, Registration
    document from the Boots Co. Ltd. Vol. II, Appendix 34 A.

    Lessel, B. (1974a) Experimental data on toxicity. Report from The
    Boots Co. Ltd. (Unpublished)

    Lessel, B. (1974b) Effect on blood and blood forming tissues. Report
    from The Boots Co. Ltd. (Unpublished)

    Lessel, B. (1974c) Short term (4-week) oral toxicity. Report from The
    Boots Co. Ltd. (Unpublished)

    Lessel, B. (1974d) Chronic (6 month) oral toxicity. Report from The
    Boots Co. Ltd. (Unpublished)

    Lessel, B. (1974e) Two year feeding trial in rats on dicloran
    (2,6-dichloro-4-nitroaniline). Report submitted by the Boots Co. Ltd.
    (Unpublished)

    Lobdell, B. and Johnson, C. (1965) U-2069 - Effect on reproduction
    capacity through three generations in the rat. Report from Woodard
    Research Corporation submitted by The Boots Co. Ltd. (Unpublished)

    Manger, W.H. (1972) Use and handling of dicloran formulations and
    influence on human eyesight. Report from Biological Laboratory,
    Aseptafabrick B.V. Delft, Netherlands. (Unpublished)

    Mat, C., Ryan, A.J. and Wright, S.E. (1967) Metabolism of some
    4-nitroaniline derivatives in the rat. Food Cosmet. Toxic., 5:657-663.

    Moe, L.D. and Lemin, A.J. (1963a) The metabolism of
    2,6-dichloro-4-nitroaniline by bibb lettuce, Part II, dicloran,
    Registration document from The Boots Co., Ltd. Vol. II, Appendix 34 B.

    Moe, L.D. and Lemin, A.J. (1963b) The metabolism of
    2,6-dichloro-4-nitroaniline-C14 in peaches. Dicloran, Registration
    document from the Boots Co., Ltd. Vol. II, Appendix 35.

    Moe, L.D. and Lemin, A.J. (1964) The metabolism of
    2,6-dichloro-4-nitroaniline by bibb lettuce, Part III. Dicloran,
    Registration document from The Boots Co., Ltd. Vol. II, Appendix 34 C.

    Ogawa, J.M., Lyda, S.D. and Weber, D.J. (1961)
    2,6-dichloro-4-nitroaniline effective against Rhizopus fruit rot
    of sweet cherries. Pl. Dis. Reptr., 45:636-638.

    Ogawa, J.M., Mathre, J.M., Weber, D.J. and Lyda, S.D. (1963) Effects
    of 2,6-dichloro-4-nitroaniline on Rhizopus species and its
    comparison with other fungicides on control of Rhizopus rot of
    peaches. Phytopathology, 53:950-955.

    Pieters, A.J. (1974) Information on dicloran residues in The
    Netherlands submitted to the Joint Meeting 8 November 1974.
    (Unpublished)

    Roburn, J. (1958) Experiments to demonstrate the effect of washing
    lettuce treated with dicloran ("Alisan"). Dicloran, Registration
    document from the Boots Co., Ltd. Vol. II, Appendix 33.

    Roburn, J. (1960) Allisan residue trials on lettuce, February to April
    1960. Lenton memo nr. 5. Lenton Experimental Station, Nottingham.
    (Unpublished)

    Roburn, J. (1961) Colorimetric determination of
    2,6-dichloro-4-nitroaniline in plants and soil. J. Sci. Fd. Agric.,
    12:766-772.

    Serrone, D., Pakdaman, P., Stern, A. and Coulston F. (1967)
    Comparative toxicology of 2,6-dichloro-4-nitroaniline in rats and
    Rhesus monkey. Tox. appl. Pharmacol., 10:404.

    Sharples, R.O. (1962) The fungitoxic effects of dicloran on Botrytis
    cinerea. Lenton Experimental Station, Nottingham. (Unpublished)

    Stough, A.R. (1962) Results of Botran(R) clinical study. Results from
    The Upjohn Co. submitted by The Boots Co. Ltd. (Unpublished)

    Van Alfen, N.K. (1972) Microbial metabolism of the fungicide
    2,6-dichloro-4-nitroaniline in soil by micro-organisms. Dissertation,
    Dept. of Plant Pathology, Univ. of California, Davis.

    Van Alfen, N.K. and Kosuge, T. (1974) Microbial metabolism of the
    fungicide 2,6-dichloro-4-nitroaniline. J. agr. Food Chem., 22:221-224.

    Wang, C.H. (1972) The effect of soil properties on losses of two
    chloronitrobenzene fungicides from soils. Dissertation, Dept. of Soil
    Sciences, Univ. of California, Davis.

    Wang, C.H. and Broadbent, F.E. (1973) Effect of soil treatments on
    losses of two chloronitrobenzene fungicides. J. environ. Quality, 2
    (4):511-515.

    Woodard, M., Cockrell, K. and Woodard, G. (1964) Safety evaluation by
    oral administration to rats and dogs for 104 weeks. Report from
    Woodard Research Corporation submitted by The Boots Co. Ltd.
    (Unpublished)
    


    See Also:
       Toxicological Abbreviations
       Dicloran (ICSC)
       Dicloran (Pesticide residues in food: 1977 evaluations)
       Dicloran (JMPR Evaluations 1998 Part II Toxicological)