First draft prepared by
    Dr K.N. Woodward
    Veterinary Medicine Directorate
    Ministry of Agriculture, Fisheries and Food
    Weybridge, Surrey, United Kingdom


         Nitrofural also known as nitrofurazone, is a broad-spectrum
    bactericidal drug. It also possesses some antiprotozoal activity. It
    is used both therapeutically and prophylactically in a number of
    food-producing species including pigs, sheep, goats, cattle,
    chickens and turkeys (Orphahell, 1991). The chemical structure is
    shown below in Figure 1.

    FIGURE 1

         Nitrofural had not been previously evaluated by the Joint
    FAO/WHO Expert Committee on Food Additives.


    2.1  Biochemical Aspects

    2.1.1  Absorption, distribution and excretion  Rats

         Nitrofural appeared to be well absorbed in the rat after oral
    administration of 100 mg/kg bw. At 4 h after administration plasma
    levels had reached 4.5 mg/l. No other time points were investigated.
    Around 5% and 0.5% of the dose were excreted in the 24 h urine and
    faeces, respectively. As acknowledged by the authors, the values are
    likely to be underestimates as nitrofurazone was shown in this study
    to be rapidly metabolized after incubation with rat liver slices.
    There was no evidence of deposition in any organ (Paul  et al.,
    1960).  Cattle

         A single oral dose of 14 mg/kg bw nitrofural administered to
    five pre-ruminant calves resulted in maximum plasma concentrations
    of 3.5 mg/litre at 3 h after administration. The final elimination
    half-life was 5 h and around 2% of the administered dose was
    recovered in the urine (Nouws  et al., 1987).

    2.1.2  Biotransformation

         As described in paragraph, nitrofural is rapidly
    metabolized by rat liver slices. Nitrofurans, including nitrofural,
    undergo metabolic reduction at the nitro group to generate reactive
    species which can covalently bind to cellular macromolecules
    (Polnaszek  et al., 1984; Kutcher & McCalla, 1984; McCalla 1979;
    McCalla  et al., 1975). Data obtained from studies with isolated
    perfused rat liver suggested that conjugation to glutathione
    followed by biliary excretion was important; only 0.27% of the
    administered perfusate was excreted unchanged (Sorrentino  et al.,
    1987). Conjugation with amino acids such as methionine may also
    occur (Hoener, 1988).

         Ring opening of nitrofurans is known to occur (Swaminathan &
    Lower, 1978) but there are insufficient data to demonstrate this
    with nitrofurazone.

    2.1.3  Effects on enzymes and other biochemical parameters

         In chickens nitrofural increased gamma-aminobutyric acid and
    glutamate concentrations in the brain at oral doses of 25 and 50
    mg/kg bw for 5 days (Ali, 1988). Nitrofural, given orally at doses
    of 10 and 20 mg/kg bw for 7 days, induced thiamine deficiency in
    chickens (Ali 1983).

         Nitrofural was found to inhibit the uptake of oxygen by various
    rat tissues (Paul  et al., 1952). Nitrofural inhibited respiration
    in isolated mouse liver mitochondria suggesting that these may be
    potential sites for toxicity (Lim  et al., 1986).

         Nitrofural and other nitrofurans are relatively potent
    inhibitors of MAO in mammalian and avian species (Hoogenboom, 1991).

         As with other nitrofurans, nitrofural can affect the endocrine
    system resulting in alterations of hormone levels. In laying
    turkeys, nitrofural decreased luteinizing hormone and prolactin in
    plasma (Ali  et al., 1987). Similar effects were noted in young
    male turkeys (Ali  et al., 1988).

         Nitrofural has been shown to inhibit utilization of
    progesterone as a steroid percursor by the adrenal in rats. In this
    study it was shown that nitrofurans, including nitrofural, block
    11-hydroxylation of deoxycorti-costerone to corticosterone.
    Sprague-Dawley rats (1 year old) given nitrofurans for 325 days
    underwent constant oestrous. The authors concluded that nitrofurans
    were antagonists of normal adrenal function and believed that this
    could lead to endocrine stress (King & Currie, 1972).

    2.2  Toxicological studies

    2.2.1  Acute toxicity studies

         The LD50 values for nitrofural are shown in Table 1. Signs of
    toxicity included roughened coats, hyperexcitability, tremors,
    reduced activity, weakness, convulsion, and respiratory collapse
    (Krantz & Evans 1945; Anderson, 1983).

        Table 1. Acute toxicity of nitrofural

    Species/strain          Sex     Route        LD50       Reference
                                              (mg/kg bw)

    Rat (unspecified)        M      oral          590       Krantz & Evans,

    Rat (Donryu)            M&F     oral          590       Miyaji, 1971

    Rat (Wistar)             M      oral          800       Anderson, 1983

    Mouse                    ?      oral          380       Krantz & Evans,
    (unspecified)                                           1945

    Mouse (ICR Jcl)         M&F     oral          590       Miyaji, 1971

    Mouse                    ?      i.p.          300       Smith et al.,
    (unspecified)                                           1963
    2.2.2  Short-term toxicity studies  Mice

         Groups of 5 male and 5 female B6C3F1 mice were given
    diets containing 0, 630, 1250, 2500, 5000 or 10 000 ppm nitrofural,
    equivalent to 0, 94.5, 188, 375, 750 or 1500 mg/kg bw/day for 14

         All mice given the three highest dietary levels died during the
    study, as did 3/5 given 188 mg/kg bw/day. Animals in all the groups
    except females given 94.5 mg/kg bw/day lost weight and all dosed
    animals displayed rough coats and convulsive seizures (National
    Toxicology Program (NTP), 1988).

         Groups of 10 male and 10 female B6C3F1 mice were given
    diets containing 0, 70, 150, 310, 620 or 1250 ppm nitrofural,
    equivalent to 0, 10, 23, 47, 93 or 188 mg/kg bw/day for 13 weeks.

         High mortality was noted at the highest dietary level (6/10
    males and 9/10 females). Mortality was relatively high at the
    penultimate dietary level (3/10 males and 5/10 females). Elevated
    relative liver weights were noted in mice given the two highest

    dietary levels. Animals given the two highest dietary levels also
    displayed hyperexcitability and convulsive seizures, and males in
    these groups had a very high incidence (80-90%) of testicular
    hypoplasia (NTP, 1988).  Rats

         Groups of 10 male and 10 female F344/N rats were given diets
    containing 0, 630, 1250, 2500, 5000 or 10 000 ppm nitrofural,
    equivalent to 0, 63, 125, 250, 500 or 1000 mg/kg bw/day, for 14

         All rats given 500 or 1000 mg/kg bw/day died. Feed consumption
    was decreased at all levels in excess of the lowest level. Signs of
    toxicity included lethargy and roughened coats. Seizures occurred in
    animals given 250 mg/kg bw/day and above. Histopathologic
    examination revealed aspermatogenesis in dosed males (NTP, 1988).

         Groups of 10 male and 10 female F344/N rats were given diets
    containing 0, 150, 310, 620, 1200 or 2500 ppm nitrofural, equivalent
    to 0, 15, 31, 62, 120 or 250 mg/kg bw/day, for 13 weeks.

         All rats survived until the end of the study but final body
    weights of animals given 62, 120 or 250 mg/kg bw/day were 11%, 32%
    and 55% lower, respectively, than control values. Relative liver
    weights for treated rats were significantly higher than control
    animals. Atrophy constricture of the hind-quarters occurred in males
    and females given the highest dietary level.

         Moderate to severe degeneration of the seminiferous epithelium
    of the testes was noted in males given the 4 highest doses. The
    no-effect-level was 15 mg/kg bw/day. Males and females given the two
    highest dietary levels had osteoporosis in the metaphyseal region.
    (NTP, 1988).

         In a study designed to investigate effects on the testes,
    45-day-old male Donryu rats were given a diet containing 0.2%
    nitrofural (equal to 300 mg/kg bw/day) for 9 days. Three to 6 rats
    given nitrofural were sacrificed every day after feeding began.

         A decrease in testis weight occurred by day 5 but
    histologically, a decrease in spermatozoa and spermatids occurred at
    day 1, with disappearance by days 3 and 4. Here, the stratified
    appearance of the normal testis had disappeared and only basal
    spermatogonia and Sertoli cells remained. Slight oedema and
    exudation occurred in interstitial tissue but Leydig cells appeared
    normal (Miyaji  et al., 1964).

    2.2.3  Long-term/carcinogenicity studies  Mice

         Groups of 50 male and 50 female B6C3F1 mice were given
    diets containing 0, 150 or 310 ppm nitrofural equal to 0, 14 or 29
    mg/kg bw/day for 2 years.

         There were no effects on body weights and survival was only
    slightly affected at the highest dose in males and females (27/50
    and 35/50 for males and females, respectively) compared with
    controls (39/50 for each sex).

         Females given nitrofural were found to have increased
    incidences of ovarian atrophy (44/50 and 38/50 in low- and high-dose
    groups respectively) compared with controls (7/47) and tubular cell
    hyperplasia (23/50 and 21/50 in low- and high-dose groups) when
    compared with controls (1/47). However, there were no effects on the
    testes in males.

         There were increased incidences of ovarian granulosa cell
    tumours in low- (4/50) and high-dose (9/50) females when compared
    with control values (1/47). Similarly, there were increased
    incidences of benign mixed tumours of the ovary in low- (17/50) and
    high-dose (20/50) females compared with controls (0/47). There were
    no elevated incidences of any tumour type in male mice (NTP, 1988;
    Kari  et al., 1989).

         In a study of transplacental carcinogenesis, a group of 20
    pregnant ICR/Jcl mice was given 3 subcutaneous injections at
    75 mg/kg bw nitrofural on days 13, 15 and 17 of gestation. The
    incidence of tumours in offspring was not increased when compared
    with untreated controls when they were examined 32 weeks after

         In the same study, newborn mice (61) were given a subcutaneous
    injection at 75 mg/kg bw 12 h after birth and on days 7, 14 and 21.
    The offspring were examined at 32 weeks after birth when an
    increased incidence of pulmonary papillary adenomas was noted
    (Nomura  et al., 1984).  Rats

         A group of 20 female Holtzman rats was fed a diet containing
    1000 ppm nitrofural (150 mg/kg bw/day) for 36 weeks. A similar group
    of 30 females was fed the same level for 44.5 weeks. Animals were
    killed 15-19 weeks after dosing was completed. At termination, there
    was an elevated incidence of mammary tumours in rats fed
    nitrofural-containing diets when compared with controls (Morris
     et al., 1969; Morris, 1965).

         Groups of 35 female Holtzman rats were fed diets containing
    0, 500 or 1000 ppm nitrofural (0, 75 or 150 mg/kg bw/day) for 45
    weeks, and then a basal diet for a further 8 weeks. Rats consuming
    nitrofural at 75 mg/kg bw/day nitrofural had no increased tumour
    incidence but in those given 150 mg/kg bw/day the numbers of
    tumour-bearing rats were increased. At week 53, the numbers with
    detectable mammary tumours were significantly increased in both the
    75 (10/35) and 150 (12/35) mg/kg bw/day groups, when compared with
    untreated controls (2/35). None of the reported tumours were
    malignant (Siedler & Searfoss, 1966).

         Groups of 20 male and 20 female CFE rats were fed diets
    containing nitrofural to give a daily intake of 50-55 mg/kg bw/day
    for 45 weeks. At the end of this period the animals were returned to
    the basal diets for a further 7 weeks. There was no increased
    incidence of any tumours in the male rats. A significant increase in
    the incidence of benign mammary tumours was noted at week 52 (12/20)
    compared with untreated controls (0/20) (Siedler & Searfoss, 1967).

         Groups of 50 male and 50 female F344/N rats were fed diets
    containing 0, 310 or 620 ppm nitrofural equal to 0, 11 or 24 mg/kg
    bw/day for 2 years. There were reductions in the rate of weight gain
    and final body weights were reduced in males and females given
    dietary nitrofural. Mortality was increased in males given the high
    dietary level (30/50; 60%) compared with controls (17/50; 34%).

         The major non-neoplastic finding was degeneration of joint
    articular cartilages in males and females in several areas of the
    body, notably, the vertebral and knee joints. A no-effect-level was
    not identifiable. Testicular degeneration occurred in 49/50 low
    dietary level and 47/50 high dietary level males compared with 12/50
    in controls.

         There was an increased incidence of mammary fibroadenomas in
    both low and high dietary level females (36/50; 72% in both groups)
    when compared with controls (8/49; 16%) but the incidence of
    adenocarcinomas was not elevated. In males, the incidence of
    mesotheliomas in the tunica vaginalis was increased but not in a
    dose-related manner (0/50, 7/50 and 2/50 in the control, low and
    high dietary level groups). There was also an increased incidence of
    sebaceous adenoma of the skin of male rats but only at the high
    dietary level (0/50, 0/50 or 5/50 for controls, low and high dietary
    levels, respectively). A slightly increased incidence of carcinoma
    of the preputial gland was seen in males (1/50, 8/50 and 5/50 in the
    control, low and high dietary groups, respectively). The incidence
    of testicular tumours was reduced in dosed males (NTP, 1988; Kari
     et al., 1989).

         It was concluded by the authors that the data from male rats
    provided only equivocal evidence of carcinogenicity. The increases
    in incidences of the various tumours were small and not

         The induction of ovarian tumours noted in mice treated with
    chemicals is relatively rare. Such an effect had been noted with
    only 8 chemicals from the NTP studies among which were nitrofural
    itself, 1,3-butadiene, 4-vinylcyclohexene, nitrofurantoin, benzene
    and delta-9-tetrahydrocannabinol. The tumours were mainly in mice
    but rats too were also affected (Maronpot, 1987).

         Nitrofural is unusual in that of 143 chemicals which had been
    tested by the NTP by 1989, only 5, including nitrofural, produced
    only benign neoplastic lesions (Huff  et al., 1989). Many other
    nitrofurans tested have been shown to be carcinogenic in rodents and
    many of these have produced malignant tumours (Cohen & Bryan, 1973;
    Cohen, 1978; Cohen  et al., 1973).

         Nitrofural and other nitrofurans inhibited the
    hepatocarcinogenesis by 4-(dimethylamino) azobenzene in rats (Akao
     et al., 1971). It also inhibited the growth of fibrosarcomas in
    mice and demonstrated antineoplastic properties against seminomas
    and carcinoma of the prostate in humans (Friedgood & Green, 1950;
    Friedgood & Ripstein, 1951).

         The International Agency for Research on Cancer (IARC) has
    concluded that there was limited evidence for the carcinogenicity of
    nitrofurazone in experimental animals and inadequate evidence for
    its carcinogenicity in humans. In its overall evaluation, IARC
    placed nitrofural in Group 3 (not classifiable as to its
    carcinogenicity) (IARC, 1990).

    2.2.4  Reproduction studies

         No information available.

    2.2.5  Special studies on teratogenicity  Mice

         A group of 16 pregnant ICR/Jcl mice was given subcutaneous
    injections at 100 mg/kg bw nitrofural on days 9, 10 and 11 of
    gestation. A group of 6 pregnant mice was given a single
    subcutaneous injection at 300 mg/kg bw nitrofural on day 10 of
    gestation. Controls were given water only on days 9, 10 and 11 of

         The 100 mg/kg bw regime did not produce an elevated incidence
    of malformations in 185 fetuses examined. However, the 300 mg/kg bw
    dose on day 10 of gestation led to an overall incidence of
    malformations of 21% compared with 0.3% in controls. These were
    mainly tail anomalies, oligodactyly, and leg defects (unspecified)
    (Nomura  et al., 1984).

         In a prescreening developmental test, female CD-1 mice were
    given nitrofural (100 mg/kg bw/day) by gavage on days 6-13 of
    gestation. Nitrofural caused a decrease in the numbers of viable
    litters (28/35 compared with 45/45 in controls) and slight
    reductions in birth weights. By the authors' criteria, these effects
    constituted positive results (Hardin  et al., 1987).

         In a preliminary teratogenicity study, groups of 6-7 CD-1 mice
    were given diets containing 0, 100, 500, 1000, 2000 or 3000 ppm
    nitrofural, equivalent to 0, 15, 75, 150, 300 or 450 mg/kg bw/day,
    on days 6-17 of gestation.

         Dams given the two highest dietary levels died during the
    study. There were no differences in food consumption in the other
    groups and no effects on weight gain. There were no differences in
    implantation site and resorption parameters except in those given 75
    mg/kg bw/day where an increased incidence of resorptions was noted
    (Price  et al., 1985).

         In a second preliminary teratology study, groups of 12 pregnant
    CD-1 mice were given diets containing 0, 75, 250, 500 or 750 ppm
    nitrofural equivalent to 0, 11, 38, 75 or 112 mg/kg bw/day, on days
    6-17 of gestation. Approximately 8% mortality occurred at the
    highest dietary level and hyperactivity, tremor, paralysis and
    convulsions were seen at the two highest levels. There were
    dose-related decreases in food consumption and body weights. Gravid
    uterine weights decreased with increasing nitrofural intake. There
    were no effects on implants/dam but the numbers of resorptions per
    litter increased with dose. There were no increases in the
    incidences of malformations (Price  et al., 1985).

         In the main teratogenicity study, groups of 26 pregnant CD-1
    mice were given diets containing 0, 38, 75, 250 or 500 ppm
    nitrofural, equivalent to 0, 6, 11, 38 or 75 mg/kg bw/day on days
    6-15 of gestation. There were no maternal deaths and no effects on
    food and water consumption. Gravid uterine weights were unaffected
    by compound intake and there were no effects on reproductive
    parameters except for an increased incidence of late fetal deaths at
    marginally maternally toxic doses. The NOEL was 11 mg/kg bw/day
    (Price  et al., 1985).  Rabbits

         Groups of 22-27 New Zeeland white rabbits were given gavage
    doses of 0, 5, 10, 15 or 20 mg/kg bw/day nitrofural in corn oil on
    days 6-19 of gestation.

         At sacrifice, the uterine contents of 121 pregnant animals were
    examined. There was no evidence of any teratogenic effect nor
    materno- or embryo/feto-toxicity at doses of 5-15 mg/kg bw/day. At
    20 mg/kg bw/day, two dams died, there was reduced weight gain,
    relative and absolute maternal liver weights were increased and an
    increased incidence of fetal malformations/litter was observed,
    which was suggestive of fetotoxicity. Hence, effects on the
    developing fetus occurred only at maternally toxic levels
    (Price  et al., 1987). The NOEL was 15 mg/kg bw/day.

    2.2.6  Special studies on genotoxicity

         Nitrofural has been tested in a range of studies (Table 2). In
    general,  in vitro studies of forward mutation in bacteria and
    mammalian cells, sister chromatid exchange, DNA damage and tests for
    clastogenicity have proved positive while  in vivo studies,
    including those for cytogenetic effects and the micronucleus test,
    have proved negative.

         In studies where the metabolic activation was incorporated into
    the agar layer, the mutagenic activities of a number of nitrofurans
    (and nitroimidazoles) were reduced, suggesting that the metabolites
    of reductive metabolism may be less mutagenic than those formed
    after oxidative metabolism (Skeggs  et al., 1984). Metabolic
    reduction is believed to be the major route of biotransformation of
    nitrofurans and nitroimidazoles and the metabolites so formed may
    account for the radiosensitizing properties of these agents (Olive,
    1979, 1980, 1978b).

         The results obtained with nitrofural are similar to those noted
    with a wide range of nitrofurans, and in particular, the propensity
    of these compounds for activity in  in vitro systems is evident
    (Klemencic & Wang, 1978). The mechanism of mutagenicity is unclear,
    as is the route of activation, although nitroreduction appears to be
    an early step (McCalla, 1981, 1983; Matsuoka  et al., 1979; McCalla
     et al., 1970, 1971). Nitrofural and other nitrofurans are
    cytotoxic to hepatocytes  in vitro (Hoogenboom  et al., 1991), but
    as the tumours caused by nitrofural are largely in endocrine organs,
    this is unlikely to be relevant to carcinogenesis by this substance.

    2.3  Observations in humans

         Nitrofural is a skin sensitizer in humans and there have been
    reports of contact dermatitis following exposure to the drug (Bajaj
    & Gupta, 1986; Laubstein and Niedergesass, 1970; Lo  et al., 1990).

         When used as an experimental antineoplastic agent in humans
    with prostatic or testicular tumours, testicular degeneration
    occurred similar to that noted in experimental animals (Friedgood &
    Ripstein, 1951). The doses used were unclear in the original text.

         In the Collaborative Perinatal Project conducted in the USA by
    the National Institute of Neurological and Communicative Diseases
    and Stroke in 1958-1965, only 0.5% (234/50282) of mother-child pairs
    had been exposed to nitrofural during months 1-4 of gestation. There
    was no increased risk of malformations in this group (Heinonem
     et al., 1977). 

        Table 2. Results of genotoxicity assays on nitrofurazone

    Test system          Test object           Concentration          Results     Reference

    Ames test1           S. typhimurium        0-75 g/ml             Positive    Baars et al.,
                         TA1538                                                   1980

    Ames test1           S. typhimurium        0-5 g/plate           Positive    Goodman et al.,
                         TA98, TA100                                              1977

    Ames test1           S. typhimurium        0-100 g/plate         Positive    NTP, 1988
                         TA100, TA1535,
                         TA1537, TA98

    Ames test1           S. typhimurium        1-20 g/plate          Positive    Ni et al., 1987
                         TA98, 98NR, 98/

    Ames test1           S. typhimurium        0.1-10 g/plate        Positive    Yahagi et al.,
                         TA1535, TA100                                            1976

    Ames test1           S. typhimurium        0-250 nmole/plate      Positive    Gajewska et al.,
                         TA97, TA102                                              1990

    Ames test1           S. typhimurium        0.1-100 g/plate       Positive    Chandler &
                         TA98                                                     Parenti, 1982

    Forward mutation     S. typhimurium        0-0.5 g/ml            Positive    Karube et al.,
                         TA100 electrode                                          1982

    Forward mutation1    E. coli               0-75 g/ml             Positive    Baars et al.,
                         343/113/(R-9                                             1980

    Table 2. cont'd

    Test system          Test object           Concentration          Results     Reference

    Forward mutation1    E. coli WP2           0-240 M               Weak        Bryant & McCalla,
                                                                      Positive    1980

    Forward mutation1    E. coli K12           0-10 mm                Positive    Chessin et al.,

    Forward mutation1    E. coli W2            10-300 M              Positive    Lu et al., 1979
                         WP2 uvrA              0-250 M               Positive

    Forward mutation2    E. coli K12           not specified          Positive    Maier et al.,
                         343/13                                                   1979

    Forward mutation1    E. coli WP2           30-120 M              Positive    McCalla &
                         uvrA-                                                    Voutsinos, 1974

    Forward mutation1    E. coli B and S       unspecified            Positive    Zampieri &
                         strains                                                  Greenberg, 1974

    Fluctuation test1    S. typhimurium        0-1 ng/ml              Positive    Green et al., 1977
                         TA1535, TA100
                         E. coli WP2 uvrA      0-100 ng/ml

    Forward mutation1    A. nidulans           0-1000 g/plate        Negative    Bignami et al.,

    Forward mutation1    N. crassa             0-200 g/ml            Positive    Ong, 1977

    SOS Chromotest       E. coli PQ37          0.01-01 ng/sample      Positive    Gajewska et al.,

    Table 2. cont'd

    Test system          Test object           Concentration          Results     Reference

    DNA damage           Mouse L cells         500 M                 Positive    Olive & McCalla,

    DNA damage           Mouse L929 cells      )0-500 M              Positive    Olive & McCalla,
                         Hamster BHK21         )                                  1975
                         cells                 )
                         Human KB cells        )

    DNA damage           Swiss mice            0.1% in diet           Positive    Olive, 1978a
    in vitro                                   (150 mg/kg bw/day)                 

    Unscheduled DNA      Rat thymocytes        unspecified            Positive    Tempel, 1980

    Unscheduled DNA      Rat and mouse         1.1 x 10-3-1.1 mg/ml   Negative    Mori et al., 1987
    synthesis            hepatocytes

    Sister chromatid     Chinese hamster       0-15 g/ml             Positive    NTP, 1988
    exchange             ovary cells

    Forward mutation1    Mouse lymphoma        0-300 g/ml            Positive    NTP, 1988
                         L5178Y cells          0-400 g/ml

    Forward mutation1    Chinese hamster       100 g/ml              Positive    Olive, 1981
                         V79 cells

    Forward mutation1    Chinese hamster       0-200 g/ml            Negative    Phillips, 1983
                         ovary - HGPRT

    Forward mutation1    Chinese hamster       0-200 g/ml            Negative    Anderson &
                         ovary - HGPRT                                            Phillips, 1985

    Table 2. cont'd

    Test system          Test object           Concentration          Results     Reference

    Sex-linked           D. melanogaster       5 mM                   Negative    Kramers, 1982
    recessive lethal

    In vitro             Chinese hamster       0-200 g/ml            Positive    Anderson &
    cytogenetics         bone marrow                                              Phillips, 1985

    In vitro             Chinese hamster       0-600 g/ml            Positive    NTP, 1988
    cytogenetics         ovary

    In vitro             Chinese hamster       0.1 mg/ml              Positive3   Matsuoaka et al.,
    cytogenetics         cells (male                                              1979

    In vitro             Human lymphocytes     not specified          Positive    Tonomura &
    cytogenetics                                                                  Sasaki, 1978

    In vivo              Chinese hamster       0-400 mg/kg bw         Negative    Anderson &
    cytogenetics         bone marrow                                              Phillips, 1985

    In vivo              Wistar rat bone       0-400 mg/kg bw or      Negative    Anderson, 1983
    cytogenetics         marrow                1-150 mg/kg
                                               bw/day for 5 days

    Micronucleus test    Sprague-Dawley rat    )0-60 mg/kg bw;        Negative    Goodman et al.,
                                               )half dose 30 h and                1977
                         Long Evans rat        )half dose 6 h,
                                               )prior to sacrifice

    1    With and without hepatic metabolic activation.
    2    With and without testicular metabolic activation.
    3    In presence but not absence of metabolic activation.


         The results of acute and short-term toxicity studies,
    carcinogenicity studies, and teratogenicity studies were considered
    by the Committee, in addition to several genotoxicity studies. Some
    limited pharmacokinetic data were also available which demonstrated
    good absorption of the drug in rats and cattle, but no data were
    available on the extent of biotransformation nor on the identities
    of any metabolites formed. A comparison with other nitrofurans
    suggests that nitrofural probably undergoes extensive

         In acute oral toxicity studies, nitrofural was slightly toxic
    to mice (LD50 = 300-600 mg/kg bw) and rats (LD50 = 600-800 mg/kg

         Testicular hypoplasia was noted in rats and mice given dietary
    nitrofural for 13 weeks. The NOEL for this effect was 15 mg/kg
    bw/day in a 13-week study in rats.

         Nitrofural was tested for its ability to induce forward
    mutations in bacteria, yeast, and mammalian cells  in vitro. In the
    majority of these studies it produced positive results. Positive
    results were also noted in tests for DNA damage in bacteria and in
    mammalian cells  in vitro, in  in vitro studies for clastogenic
    effects in Chinese hamster ovary or bone marrow cells, and in human
    lymphocytes. However,  in vivo studies of clastogenic potential in
    Chinese hamster or rat bone marrow were negative. The results
    suggest that nitrofural is genotoxic  in vitro but not  in vivo.

         Carcinogenicity studies with nitrofural were conducted in rats
    and mice. In a 2-year dietary study in B6C3F1 mice, the
    animals were given nitrofural equal to daily oral doses of 14 or
    29 mg/kg bw/day. In female mice, nitrofural induced a high incidence
    of ovarian atrophy and of ovarian tubular-cell hyperplasia in both
    dose groups when compared with controls. There were also increased
    incidences of ovarian granulosa-cell and benign mixed tumours in
    low-and high-dose groups. There were no elevated incidences of any
    tumour type in male mice.

         In a 2-year carcinogenicity study in rats, testicular
    degeneration was one of the major non-neoplastic findings. There
    were also elevated incidences of degeneration of the joint articular
    cartilages at several sites in both males and females. No-effect
    levels could not be identified for these findings. There was an
    increased incidence of mammary fibroadenomas in females given the
    low and high doses of 11 and 24 mg/kg bw/day.

         Teratogenicity studies were conducted in mice and rabbits. In
    mice, the rate of late fetal death was increased in treated animals
    at maternally toxic doses; the NOEL was 11 mg/kg bw/day. Pregnant

    rabbits were given gavage doses of up to 20 mg/kg bw/day. At the
    highest dose, there was an elevated incidence of minor fetal
    malformations per litter but this dose produced maternal toxicity.
    The NOEL was 15 mg/kg bw/day. The data suggest that nitrofural is
    not a direct teratogen but produces fetotoxic effects at maternally
    toxic doses in mice and rabbits.

         Although nitrofural was tumorigenic in rats and mice, the
    tumours it produced were benign and were restricted to endocrine
    organs and the mammary gland. Moreover, the mutagenicity studies
    suggest that nitrofural is genotoxic  in vitro but not  in vivo.
    Taken together, the data suggest that nitrofural is a secondary
    carcinogen producing its effects in endocrine-responsive organs by a
    mechanism that remains to be elucidated. In rats, nitrofural has
    been shown to disrupt the utilization of progesterone and blocks the
    synthesis of cortisone from deoxycortisone. Effects on
    steroidogenesis may therefore be involved in the process of tumour
    formation. The closely related compound nitrofurantoin also produced
    benign ovarian tumours in the same strain of mouse, which were
    thought to be secondary to drug-induced ovarian atrophy. This type
    of effect cannot be excluded in the biogenesis of the ovarian
    tumours produced by nitrofural. However, the Committee recognized
    that no-effect levels for these effects could not be identified. The
    Committee noted that nitrofural had been reviewed by the
    International Agency for Research on Cancer, which concluded that
    there was limited evidence for the carcinogenicity of nitrofural in
    animals but inadequate evidence for humans.


         The Committee concluded that it could not establish an ADI for
    nitrofural because no-effect levels had not been established for the
    tumorigenic effects. It noted that, although a no-effect level
    (15 mg/kg bw/day) had been established in rats for testicular
    hypoplasia in a 13-week study, a slightly lower dose (11 mg/kg
    bw/day) resulted in a high incidence of testicular degeneration in
    the 2-year carcinogenicity study. There was no NOEL for this effect
    in the 2-year study and, moreover, no study on reproductive
    performance was available. The degenerative changes in the joints of
    rats were seen in both males and females but a no-effect level could
    not be established.

         Before reviewing nitrofural again the Committee would wish to

         1.   Further data from long-term studies in rats which would
              allow the identification of no-effect levels for the
              effects on joint articular cartilages and for testicular

         2.   Data to support the view that tumour formation in rodents
              following nitrofural administration has an endocrine
              origin. If a mechanism were demonstrated, no-effect levels
              for suitable end-points would need to be identified.

         3.   Additional data on the identity, quantity and biological
              characteristics of nitrofural metabolites.


    AKAO, M., KURODA, K., KANISAWA, M. & MIYAKI, K, (1971). Influence of
    some nitrofurans on carcinogenesis in rats fed 4-(dimethylamino)
    azobenzene.  Gann, 62: 479-484.

    ALI, B.H. (1983). The effect of nitrofurazone on the thiamine status
    of chickens.  Comp. Biochem. Physiol., C.76: 131-134.

    ALI, B.H., SILSBY, J. & EL HALAWANI, M.E. (1987). Effect of
    furazolidone and nitrofurazone on egg production, on plasma
    luteinising hormone and on prolactin concentrations in turkeys.
     Brit. Poultry Sci., 28: 613-621.

    ALI, B.H. (1988). Effect of furazolidone and nitrofurazone on brain
    gamma-aminobutyric acid and glutamate concentrations in chickens.
     Clin. Exp. Pharmacol. Physiol., 15: 959-963.

    ALI, B.H., SILSBY, J.L., EDENS, F. & EL HALAWANI, M.E. (1988).
    Effects of furazolidone or nitrofurazone on the concentrations of
    hypothalamic amines and plasma luteinizing hormone (LH) and
    prolactin (PRL) levels in young turkeys.  Gen. Pharmacol., 19:

    ANDERSON, D. (1983) Cytogenetic analysis in rat bone marrow with
    nitrofurazone. Project No. 466.59.63. Unpublished report for Norwich
    Eaton Pharmaceuticals, Inc. Submitted to WHO by Orphahell B.V.,
    Mijdrecht, Netherlands.

    ANDERSON, D. & PHILLIPS, B.J. (1985). Nitrofurazone - genotoxicity
    studies in mammalian cells  in vitro and  in vivo. Food Chem.
     Toxicol., 23: 1091-1098.

    BREIMER, D.D. (1980). Preliminary studies on the ability of
     Drosophila microsomal preparations to activate mutagens and
    carcinogens.  Mutat. Res., 72: 257-264.

    BAJAJ A.K. & GUPTA, S.C. (1986). Contact hypersensitivity to topical
    antibacterial agents.  Int. J. Dermatol., 25: 103-105.

    FABRIZI, M. (1982). Evaluation of 2 different genetic markers for
    the detection of frameshift and missense mutagens in  A. nidulans.
     Mutat. Res., 97: 293-302.

    BRYANT, D.W. & McCALLA, D.R. (1980). Nitrofuran-induced mutagenesis
    and error prone repair in  Escherichia coli. Chem-Biol. Interact.,
    31: 151-166.

    CHANDLER, A.D. & PARENTI, R.R. (1982). Isolation and
    characterisation of a nitrofurazone resistant strain of  Salmonella
    TA98.  Environ. Mutagen., 4: 319-320.

    (1978). Radiosensitization, mutagenicity, and toxicity of
     Escherichia coli by several nitrofurans and nitroimidazoles.
     Radiat. Res., 75: 424-431.

    COHEN S.M. & BRYAN, G.T. (1973). Carcinogenesis caused by nitrofuran
    derivatives. Pharmacology and the Future of Man.  Proc. Vth Int.
     Congr. Pharmacol., San Francisco, Karger, Basel, 161-170.

    (1973). Carcinogenicity of 5-nitrofurans, 5-nitromidazoles,
    4-nitrobenzenes and related compounds.  J. Natl. Cancer Inst., 51:

    COHEN, S.M. (1978). Toxicity and carcinogenicity of nitrofurans.
     Carcinogenesis, 4: 171-231.

    FRIEDGOOD, C.E. & GREEN, M.N. (1950). The effect of nitrofurazone on
    the growth of fibrosarcoma in mice.  Cancer Res., 10: 613-615.

    FRIEDGOOD, C.E. & RIPSTEIN, C.B. (1951). The effect of nitrofurazone
    on tumours of the human testis and prostate gland.  Surg. Forum,

    GAJEWSKA, J., SZCZYPKA, M., TUDEK, B. & SZYMCZYK, T. (1990). Studies
    on the effect of ascorbic acid and selenium on the genotoxicity of
    nitrofurans : nitrofurazone and furazolidone.  Mutat. Res., 232:

    GOODMAN, D., AUFRERE, M., VORE, M. & MEYERS, F. (1977). Metabolism
    of nitrofurazone. 61st Annual Meeting of the Federation of American
    Societies for Experimental Biology, Chicago, Illinois.

    D.R. (1977). Use of a simplified fluctuation test to detect and
    characterize mutagenesis by nitrofurans.  Mutat. Res., 44: 139-143.

    K.P., MacKENZIE, K.M., PICCIRILLO, V.J., & SMITH, K.N. (1987).
    Evaluation of 60 chemicals in a preliminary developmental toxicity
    test.  Terat. Carcinogen. Mutagen., 7: 29-48.

    HEINONEN, O.P., SLONE, D. & SHAPIRO, S. (1977). (Eds). Antimicrobial
    and antiparasitic agents. In: Birth Defects and Drugs in Pregnancy,
    Publishing Sciences Group, Inc, Littleton, MA, 296-313.

    HOENER, B-A. (1988). Nitrofurazone : Kinetics and oxidative stress
    in the singlepass isolated perfused rat liver.  Biochem. Pharmacol.,
    37: 1629-1636.

    (1991). The use of pig hepatocytes for cytotoxicity studies of
    veterinary drugs: A comparative study with furazolidone and other
    nitrofurans.  Toxicol. in Vitro, 5: 31-38.

    HOOGENBOOM, L.A. (1991). The use of pig hepatocytes for
    biotransformation and toxicity studies. Thesis,
    Landbouwuniversiteit, Wageningen, Netherlands.

    HUFF, J.E., EUSTIS, S.L. & HASEMAN, J.K. (1989). Occurrence and
    relevance of chemically-induced benign neoplasms in long-term
    carcinogenicity studies.  Cancer Metastasis Rev., 8: 1-22.

    IARC (1990). Monographs on the evaluation of carcinogenic risks to
    humans.  Pharmaceutical Drugs, IARC, Lyon, 50: 195-209.

    (1989). Toxicity and carcinogenicity of nitrofurazone in F344/N rats
    and B6C3F1 mice.  Food Chem. Toxicol., 27: 129-137.

     Salmonella electrode for screening mutagens.  Anal. Chem., 54:

    KING, C.D. & CURRIE, G.N. (1972). Effects of nitrofurans on
    steroidogenesis and mammary hyperplasia. Unpublished report. Norwich
    Pharmacal Company, Project No. 596.09. Submitted to WHO by Orphahell
    B.V., Midrecht, Netherlands.

    KLEMENCIC, J.M. & WANG, C.Y. (1978). Mutagenicity of nitrofurans.
    In: Bryan, G.T. ed.  Carcinogenesis, 4: 99-130, New York, Raven

    KRAMERS, P.G.N. (1982). Studies on the induction of sex-linked
    recessive lethal mutations in  Drosophila melanogaster by
    nitroheterocyclic compounds.  Mutat. Res., 101: 209-236.

    KRANTZ, J.C. & EVANS, W.E. (1945). A contribution to the
    pharmacology of 5-nitro-2-furaldehyde semicarbazone.  J. Pharmacol.
     Exp. Therap., 85: 324-331.

    KUTCHER, W.W. & McCALLA, D.R. (1984). Aerobic reduction of
    5-nitro-2-furaldehyde semicarbazone by rat liver xanthine
    dehydrogenase.  Biochem. Pharmacol., 33: 799-805.

    LAUBSTEIN V.H. & NIEDEGESASS, G. (1970). Untersuchungen uber
    gruppensensibilisierungen bei nitrofuranderivaten.  Derm. Mschr.,
    156: 1-8.

    LIM, L.O., BORTELL, R. & NEIMS, A.H. (1986). Nitrofurantoin
    inhibition of mouse liver mitochondrial respiration involving
    NAD-linked substrates.  Toxicol. Appl. Pharmacol., 84: 493-499.

    LO, J.S., TAYLOR, J.S. & ORIBA, H. (1990). Occupational allergic
    contact dermatitis to airborne nitrofurazone.  Dermatologic Clinics,
    8: 165-168.

    LU, C., McCALLA, D.R. & BRYANT, D.W. (1979). Action of nitrofurans
    on  E. coli (mutation and induction and repair of daughter-strand
    gaps in DNA).  Mutat. Res., 67: 133-144.

    MAIER, P., PHILPOT, R.M., MOHN, G.R. & MALLING, H.V. (1979).
    Influence of subcellular fractions of mammalian testes on the
    mutagenic activity of nitrofurans toward  Escherichia coli. Mutat.
     Res., 63: 233-243.

    MARONPOT, R.R. (1987). Ovarian toxicity and carcinogenicity in eight
    recent National Toxicology Program studies.  Environ. Health
     Perspect., 73: 125-130.

    MATSUOKA, A., HAYASHI, M. & ISHIDATE, M.J, (1979). Chromosomal
    aberration tests on 29 chemicals combined with S9 mix  in vitro.
     Mutat. Res., 66: 277-290.

    McCALLA, D.R. (1979). Nitrofurans : antibiotics. In: Hahn F(Ed),
    Mechanism of Action of Antibacterial Agents (Vol IV). Berlin,
    Springer Verlag, 176-213.

    McCALLA, D.R., OLIVE P.L., TU, Y. & FAN, M. (1975). Nitrofurazone -
    reducing enzymes in  E. coli and their role in drug activation
     in vivo. Canad. J Microbiol., 21: 1484-1491.

    McCALLA D.R. & VOUTSINOS, D. (1974). On the mutagenicity of
    nitrofurans.  Mutat. Res., 26: 3-16.

    McCALLA, D.R. (1981). Metabolic activation of nitroheterocyclic
    compounds in bacteria and mammalian cells. In:  Short-Term Tests for
     Chemical Carcinogens, Springer, New York, 36-47.

    McCALLA, D.R. (1983). Mutagenicity of nitrofuran derivatives:
    Review.  Environ. Mutagen., 5: 745-765.

    McCALLA, D.R., REUVERS, A. & KAISER, C (1971). "Activation" of
    nitrofurans in animal tissues.  Biochem. Pharmacol., 20: 3532-3537.

    McCALLA, D.R., REUVERS, A. & KAISER, C. (1970). Mode of action of
    nitrofurazone.  J. Bacteriol., 104: 1126-1134.

    MIYAJI, T., MIYAMOTO, M, & UEDA, Y. (1964). Inhibition of
    spermatogenesis and atrophy of the testes caused by nitrofuran
    compounds.  Acta Path. Jap., 14: 261-273.

    MIYAJI, T. (1971). Acute and chronic toxicity of furylfuramide in
    rats and mice.  Tohoku J. Exp. Med., 103: 331-369.

    MORI, H., SUGIE, S., YOSHIMI, M., KINOUCHI, T. & OHNISHI, Y. (1987).
    Genotoxicity of a variety of nitroarenes and other nitro compounds
    in DNA-repair tests with rat and mouse hepatocytes.  Mutat. Res.,
    190: 159-167.

    MORRIS, J.E. (1965). The carcinogenic activity in the rat of some
    derivatives of 5-nitrofurn. PhD Thesis, University of Wisconsin,
    Madison, USA.

    MORRIS, J.E., PRICE, J.M., LALICH, J.J. & STEIN, R.J. (1969). The
    carcinogenic activity of some 5-nitrofuran derivatives in the rat.
     Cancer Res., 29: 2145-2156.

    NI, Y.C., HEFLICH R.H., KADLUBAR, F.F. & FU, P.P. (1987).
    Mutagenicity of nitrofurans in  Salmonella typhimurium TA98, TA98NR
    and TA98/1,8-DNP-6.  Mutat. Res., 192: 15-22.

    MASADA, K. & SAKAMOTO, Y. (1984). Induction of tumours and
    malformations in mice after prenatal treatment with some antibiotic
    drugs.  Med. J. Osaka Univ., 35: 13-17.

    (1987). Some pharmacokinetic data about furaltadone and
    nitrofurazone administered orally to preruminant calves.  Vet. Q.,
    9: 208-214.

    NTP (1988). Toxicology and carcinogenesis studies of nitrofurazone
    (CAS No. 59-87-0) in F344/N rats and B6C3F1 mice. Technical
    Report Series No. 337. US Department of Health and Human Services,
    Public Health Service/National Institutes of Health National
    Toxicology Program.

    OBASEIKI-EBOR, E.E. & AKERELE, J.O. (1986). Nitrofuran mutagenicity:
    Induction of frameshift mutations.  Mutat. Res., 175: 149-152.

    OLIVE, P.L. & McCALLA, D.R. (1975). Damage to mammalian cell DNA by
    nitrofurans.  Cancer Res., 35: 781-784.

    OLIVE, P.L. & McCALLA, D.R. (1977). Cytotoxicity and DNA damage to
    mammalian cells by nitrofurans.  Chem.-Biol. Interact., 16:

    OLIVE, P.L. (1978a) Nitrofurazone-induced DNA damage to tissues of
    mice.  Chem.-Biol. Interact., 20: 323-331.

    OLIVE, P.L. (1978b). Macromolecular, antineoplastic, and
    radiosensitization effects of nitrofurans.  Carcinogenesis, 4:

    OLIVE, P.L. (1979). Influence of cellular environment on toxicity of
    nitroheterocycles.  Chem.-Biol. Interact., 27: 281-290.

    OLIVE, P.L. (1980). Mechanisms of the  in vitro toxicity of
    nitroheterocycles, including flagyl and misonidazole. Section of
    Radiobiology, The Johns Hopkins Oncology Center, Baltimore, MD,
    Radiation Sensitizers: Their Use in the Clinical Management of
    Cancer, 5: 39-45.

    OLIVE, P.L. (1981). Correlation between the half-wave reduction
    potentials of nitroheterocycles and their mutagenicity in Chinese
    hamster V79 spheroids.  Mutat. Res., 82: 137-145.

    ONG T-M. (1977). Mutagenic activities of nitrofurans in  Neurospora
     crassa. Mutat. Res., 56: 13-20.

    ORPHAHELL, B.V. (1991). Furazolidone-nitrofurazone. Summary volume
    1. Unpublished report. Submitted to WHO by Orphahell B.V.,
    Mijdrecht, Netherlands.

    ELLS, V.R. & BUZARD, J.A. (1960). Studies on the distribution and
    excretion of certain nitrofurans.  Antibiotics and
     Chemotherap., 10: 287-302.

    PAUL, H.E., PAUL, M.F., KOPKO, F. & MAIN, R.J. (1952). Effect of
    furacin (5-nitro-2-furaldehyde semicarbazone) on the  in vitro
    metabolism of mammalian tissues.  Proc. Soc. Exp. Biol. Med., 79:

    PHILLIPS, B.J. (1983). Study of mutation at HGPRT locus in cultured
    Chinese hamster ovary cells with nitrofurans. Unpublished report
    (466.59.62), Norwich Eaton Pharmaceuticals, Inc. Submitted to WHO by
    Orphahell B.V., Mijdrecht, Netherlands.

    (1984). No detectable reaction of the anion radical metabolite of
    nitrofurans with reduced glutathione or macromolecules.  Chem.-Biol.
     Interact., 51: 263-271.

    PRICE, C.J., GEORGE, J.D. MARR, M.C. (1985). Teratologic evaluation
    of nitrofurazone (CAS No 59-87-0) administered to CD-1 mice on
    gestational days 6 though 15. National Toxicology Program,
    NTP-85-335, PB86-145844/GAR.

    PRICE, C.J., GEORGE, J.D. & MARR, M.C. (1987). Teratologic
    evaluation of nitrofurazone (CAS No 59-87-0) administered to New
    Zealand white rabbits on gestational days 6 through 19. National
    Toxicology Program, NTP-87-200, PB88-130984/GAR.

    SIEDLER, A.J. & SEARFOSS, W. (1966). Effects of long-term feeding of
    various nitrofurans to rats. Unpublished report (Problem No. 440.07)
    (Special Report), Norwich Pharmacal Company. Submitted to WHO by
    Orphahell B.V., Mijdrecht, Netherlands.

    SIEDLER, A.J. & SEARFOSS, W. (1967). Effects of long-term feeding of
    various nitrofurans to rats. Unpublished report (Problem No.440.07)
    (Addendum to Special Report), Norwich Pharmacal Company. Submitted
    to WHO by Orphahell B.V., Mijdrecht, Netherlands.

    WISLOCKI, P.G. & WOLF, F.J. (1984). Effect of liver enzymes on the
    mutagenicity of nitroheterocyclic compounds : activation of 3a, 4,
    5, 6, 7, 7a - hexahydro-3-(1-methyl-5-nitro-1H-imidazole-2-yl)-
    1,2-benzisoxazole and deactivation of nitrofurans and
    nitroimidazoles in the Ames test.  Mutat. Res., 136: 1-8.

    SMITH, C.G., GRADY, J.E. & NORTHAN, J.I. (1963). Relationship
    between cytotoxicity  in vitro and whole animal toxicity.  Cancer
     Chemotherapy Reports, 20: 9-12.

    SORRENTINO, D., BODE, W. & HOENER, B-A. (1987). Nitrofurazone
    disposition by perfused rat liver. Effect of dose size and
    glutathione depletion.  Biochem Pharmacol., 36: 915-918.

    STITZEL, K.A., McCONNELL, R.F. & DIERCKMANN, T.A. (1989). Effects of
    nirofurantoin on the primary and secondary reproductive organs of
    female 2B6C3F1 mice.  Toxicol. Pathol., 17: 774-781.

    SWAMINATHAN, S. & LOWER, G.M. (1978). Biotransformation and
    excretion of nitrofurans.  Carcinogenesis, 4: 59-97.

    TEMPEL, K. (1980). Unscheduled DNA synthesis of rat thymocytes under
    the influence of radioprotective and radiosensitizing agents.
    Naunyn-Schmiedebergs  Arch. Pharmacol., 311: R7.

    TONOMURA, A. & SASAKI, M.S. (1973). Chromosome aberrations and DNA
    repair synthesis in cultured human cells exposed to nitrofurans.
     Japan J. Genetics, 48: 292-294.

    BRYAN, G.T. (1976). Mutagenicities of nitrofuran derivatives on a
    bacterial tester strain with a R factor plasmid.  Mutat. Res., 40:

    ZAMPIERI, A. & GREENBERG, J. (1964). Nitrofurazone as a mutagen in
     Escherichia coli. Biochem. Biophys. Res Commun., 14: 172-176.

    See Also:
       Toxicological Abbreviations
       Nitrofural (Nitrofurazone) (IARC Summary & Evaluation, Volume 50, 1990)