613. Carbendazim (Pesticide residues in food: 1983 evaluations)

PESTICIDE RESIDUES IN FOOD - 1983

Sponsored jointly by FAO and WHO

EVALUATIONS 1983

Data and recommendations of the joint meeting
of the FAO Panel of Experts on Pesticide Residues
in Food and the Environment and the
WHO Expert Group on Pesticide Residues
Geneva, 5 - 14 December 1983

Food and Agriculture Organization of the United Nations
Rome 1985

CARBENDAZIM

Explanation

Carbendazim (methyl 2-benzimidazole carbamate) was evaluated by
the Joint Meetings of 1973, 1976, 1977 and 1978 (FAO/WHO 1974, 1977,
1978, 1979).¹ The data available were not considered adequate for
the estimation of an acceptable daily intake (ADI). Data necessary for
estimating an ADI were identified in 1976 and 1978 and listed under
"Further Work or Information". These data have been provided and
reviewed in this monograph addendum.

New, additional, or updated information on use patterns,
environmental chemistry and plant metabolism was also made available
to the Meeting, along with new or additional data on crop residues
from supervised trials.

TOXICOLOGY

EVALUATION FOR ACCEPTABLE DAILY INTAKE

BIOCHEMICAL ASPECTS

Absorption, Distribution and Excretion

NMRI mice and Wistar rats were given carbendazim, via
intragastric incubation, as a single dose of 3 mg/kg b.w. and one of
300 mg/kg b.w. Urine was collected during the first 6 h, after which
the animals were killed. Analyses revealed no sex differences. Almost
all metabolites in urine were conjugated as sulphate esters. Cleavage
of these conjugates by B-glucuronidase/arylsulphatase released 5-HBC
as the only extractable metabolite from water. Urine of mice contained
a higher portion of compounds that remained polar after enzyme
treatment than the corresponding urine of rats. Polarity was caused by
a functional group (e.g. phenolic hydroxyl group) that was introduced
into the MBC molecule by a conjugation reaction. These water-soluble
compounds were not identified. Essentially the same metabolites were
found in both mouse and rat urine samples with only quantitative
differences observed between species (Dorn et al. 1983).

Effects on Enzymes and Other Biochemical Parameters

Groups of Wistar-SPP male rats and Swiss-SPF male mice were
administered carbendazim in the diet at dosage levels of 0 to

¹ See Annex 2 for FAO and WHO documentation.

10 000 ppm for 60 days. The induction of liver enzyme activities by
carbendazim was examined and compared with a positive control, which
received phenobarbital sodium (administered via drinking water).

Growth and food consumption were decreased in rats at 10 000 ppm
but not in mice administered up to 5 000 ppm in the diet. Relative
liver weights were increased in rats fed 2 000 and 10 000 ppm and in
mice receiving 1 000 and 5 000 ppm of carbendazim. Phenobarbital
groups were similarly affected. Protein concentrations in the total
liver homogenates and in post-mitochondrial fractions of rate were not
affected by carbendazim, whereas in mice both fractions were increased
at 5 000 ppm.

The feeding of carbendazim to rats at dose levels of 2 000 ppm
and higher resulted in slight to moderate induction of several
drug-metabolizing enzymes of phase 1 (7-ethoxy-coumarin-O-deethylase,
biphenyl-4-hydroxylase, aniline hydroxylase,
4-methoxybiphenyl-N-demethylase and cytochrome-o-reductase). Similar
increased activities of phase 2 drug metabolizing enzymes (glucuronyl
transferase I and II) and glutathione content were moderately to
markedly increased at this dose.

The feeding of carbendazim to mice at dose levels of 1 000 ppm
and higher resulted in moderate to marked increases in drug
metabolizing enzymes of phase 1 (including cytochrome P-450 and
aminopyrine-N-demethylase). Cytochrome-c-reductase activity was
decreased. Glucuronyl transferase and glutathione-S-transferase
activities, along with glutathione content, were slightly increased.

There were no measurable differences noted between rats and mice
in regard to the metabolism of the test substance, although exhaustion
of the detoxification mechanism was more evident in the mouse at the
higher dose levels. The detoxification and elimination of carbendazim
and its metabolites proceed more rapidly in the rat than in the mouse.
This is reinforced by the increased glutathione content in rat liver
and increased activity of phase 2 enzymes (Falke et al. 1982a,b).

TOXICOLOGICAL STUDIES

Acute Toxicity

The acute toxicity of carbendazim in several animal species is
summarized in Table 1.

Gross and histopathological changes were observed in the testes
and epididymides of male rats orally dosed with carbendazim at doses
of 1 000 mg/kg b.w. and greater. Testes were small, soft and
discoloured with greater than 70 percent of the tubules showing
degenerative changes. Sperm was reduced or absent in the epididymides
examined.

Table 1 Acute Toxicity of Carbendazim in Animals

Chemical Species Sex (Number) Route Vehicle ADL/LD₅₀¹ Reference
(mg/kg b.w.)

Carbendazim Rat M/F (10/dose) Oral Corn oil LD₅₀ > 10 000 Goodman 1975
(MBC) Rat M (1/dose) Oral Peanut oil ALD > 11 000 Sherman 1965
Rat M/F (10/dose) Oral Sesame oil LD₅₀ > 15 000 Kramer &
Weigand 1971
Rat M (1/dose) Oral Peanut oil ALD > 17 000 Sherman &
Krause 1966
Mouse M (10/dose) Oral Propylene LD₅₀ > 15 000 Til & Beems
glycol 1981
Dog M/F (2/dose) Oral Sesame oil ALD₅₀ > 5 000 Scholz &
Weigand 1972
G. pig M (10/dose) Oral Corn oil LD₅₀ > 5 000 Dashiell 1975a
Mouse M/F (10/dose) I.P. Sesame oil LD₅₀ > 15 000 Scholz &
Weigand 1972
Rat M (10/dose) I.P. 0.9% saline LD₅₀ > 2 000 Scholz &
& Tween 80 Weigand 1972

Rat M (6/dose) Inhal. Dust ALC>5.9 mg/l Server 1975
(1 h. ) (time weighted
concentration)
Rabbit M (10/dose) Dermal 50/50 Aqueous LD₅₀>10 000 Edwards 1974a
paste
Rat F (5/dose) Dermal Sesame oil ALD > 2 000 Kramer &
Weigand 1971
75% wettable Rat M/F (5/dose) Oral Corn oil LD₅₀ > 5 000 Hinckle 1981
powder
75% Wettable Rat M/F (10/dose) Inhal. Dust LC₅₀ > mg/l Nash 1982
Powder (4 h.)
75% Wettable Rabbit M/F (5/dose) Dermal Physiologic LD₅₀ > 2 000 Ford 1982
Powder saline

¹ Based on mg/kg a.i.
Special Studies on Eye and Skin Irritation

The eye irritation evaluation of technical carbendazim was
negative in albino rabbits. The 75 percent wettable powder formulation
when tested in rabbits produced transient corneal opacity in 6/6
unwashed and 2/3 washed eyes. Biomicroscopic examination confirmed
this finding as mild to moderate. Conjunctival irritation (redness,
swelling, discharge) was also transient. All eyes were normal at day
4 of the observation period. Irritation response was probably related
to the inert ingredients in the wettable powder formulation (Edwards
1974b; Henry 1982).

A 75 percent wettable powder formulation produced transient
slight irritation when applied to the intact and abraded skin of
albino rabbits (Ford 1981a). A 55 percent suspension of the 75 percent
wettable powder formulation in dimethyl phthalate produced mild
irritation to the shaved intact skin of albino guinea pigs. A 5.5
percent concentration produced no irritation (Ford 1981b).

Special Study on Sensitization

Albino guinea pigs (10 males) exposed to carbendazim, either
technical material or a 75 percent wettable powder formulation,
presented no evidence of dermal sensitization following both
intradermal injections and repeat applications to shaved intact skin
(Ford 1981b).

Short-Term Studies

Rat

Groups of ChR-CD male rats (6/dose) were intubated with 200,
3 400 or 5 000 mg carbendazim/kg/day, five times/week for two weeks.
Mortality occurred in 2/6 rats at the 3 400 mg/kg dose only. Animals
at all levels demonstrated gross and microscopic evidence of adverse
effects on testes and reduction or absence of sperm in the
epididymides. Testes were small and discoloured, with tubular
degeneration and evidence of aspermatogenesis. Morphological changes
were also reported at 3 400 mg/kg for the duodenum, bone marrow and
liver (Sherman 1965; Sherman & Krauss 1966).

Groups of ChR-CD rats (16 males and 16 females per group) were
fed carbendazim (72 percent a.i.) in the diet for 90 days at dosage
levels of 0, 100, 500 and 2 500 ppm. Animals were observed daily for
behavioural changes and body weight and food consumption were recorded
at weekly intervals. Haematological examinations were conducted on 10
male and 10 female rats in each group at 30, 60 and 90 days. Routine
urinalyses were performed on the same animals, as well as plasma
alkaline phosphatase and glutamic pyruvic transaminase levels. After
90-98 days of continuous feeding, 10 male and 10 female rats in each

group were killed and selected organs weighed. Additional organs were
preserved for microscopic examination. The six male and six female
rats remaining in each group after terminal sacrifice were subjected
to a reproduction study.

There were no gross toxic signs of poisoning and no compound-
related effects on weight gain, food consumption, food efficiency or
haematology. There were no control data for biochemistry, urinalysis
determinations or differential white blood counts. The average daily
dose for the high dose group animals was 360 mg/kg b.w./day,
initially, and 123-152 mg/kg b.w./day at sacrifice. Liver to body
weight ratio in females at 2 500 ppm was slightly increased compared
with control rats. There were no effects on testicular weights in any
of the treatment groups. Microscopic examination of selected tissues
and organs in the control and high dose groups demonstrated no adverse
effects attributable to the presence of carbendazim in the diet
(Sherman 1968).

Rabbit

New Zealand Albino rabbits (6 males/group) were treated with
0 and 2 000 mg/kg of carbendazim, applied as a 50 percent aqueous
paste to the shaved intact dorsal skin. Material was applied
repeatedly, six hours/day for ten consecutive days. There were no
untoward effects on body weight, clinical symptoms, organ weights,
gross or histopathology of selected organs. There was focal necrosis
of the epidermis and polymorphonuclear cell infiltration of the dermis
in 5/6 rabbits exposed to carbendazim. No other effects were observed
(Dashiell 1975b).

Dog

Groups of one-year-old beagles (4 males and four females per
group) were administered carbendazim (53 percent a.i.) in the diet for
three months at dosage levels of 0, 100, 500 and 2 500 ppm. The
2 500 ppm level was reduced to 1 500 ppm because of loss of appetite
and decreased body weight. However, compound administration was
interrupted when animals were placed on a control diet for a few days
and then restarted at the 1 500 ppm dietary level. Therefore, the data
generated from the high dose group are not considered in the
evaluation of this study.

Food consumption and body weight data were recorded weekly.
Clinical laboratory examinations, including haematological,
biochemistry and urinalysis measurements were performed pre-test and
after 1, 2 and 3 months of feeding. At the conclusion of the study all
animals were killed, selected organs weighed and additional organs
subjected to gross and microscopic evaluations.

There was no mortality or adverse cageside observations over the
course of the study and growth and food consumption were normal,
except as noted at the high dose level (1 500-2 500 ppm). Urinalysis
measurements were unaffected by treatment. There were no dose-related
effects on the haematological measurements. Females at the mid-dose
level showed a trend toward increased cholesterol levels at 1, 2 and
3 months compared with pre-test and control. High-dose females had
similarly elevated cholesterol levels. Organ-to-body weight changes
were observed for the thymus of low- and mid-dose males and for the
prostate of mid-dose males. All weights for these organs were
increased compared with control values. However, only liver, kidney
and testes were examined histologically in low- and mid-dose group
dogs. Limited histopathology data did not indicate compound-related
effects (Sherman 1970).

Groups of beagles (four males and four females per group) were
administered carbendazim in the diet at dosage levels of 0, 100, 300
and 1 000 ppm for 13 weeks. The 1 000 ppm level was increased to
2 000 ppm after six weeks of treatment. Body weights, haematological
and blood chemistry measurements, urinalysis and liver/kidney function
tests were examined periodically during the test. Gross and
microscopic examinations of all animals were performed at the
conclusion of the study.

There were no reported compound-related effects on clinical
behaviour, body weight, food consumption, haematology, kidney function
(phenol red excretion) or liver function (BSP retention) examinations.
Blood chemistry measurements were normal, except for a slight decrease
in albumin in mid- and high-dose males at 12 weeks. These values
differed from week 0 measurements only in high-dose males. Urinalysis
was normal except for a high bacteria count in high-dose females at
week 13. Blood clotting time was slightly reduced in high-dose dogs at
week 12. There were slight increases in relative liver and thyroid
weights and a decrease in relative heart weights in the 2 000 ppm
group compared with control. There were no microscopic changes in
these organs or any other organs that could be associated with
treatment. There was an increase in submucosal lymphocytic infiltrates
in female dogs in all groups, which was significant at the high dose.
Carbendazim appeared to be without adverse effects on beagles when
incorporated in the diet for 13 weeks at dietary levels of 300 ppm or
less (Til et al. 1972). (NOTE: All measurements for males and
females were combined and averaged. This practice can complicate
interpretation of the results when there are slight or marginal
effects in one sex, such as in this study.)

Special Study on Reproduction

Rat

A one-generation reproduction study in rats was performed with 24
male and 24 female rats that had been removed from a 90-day dietary
feeding study. Dietary levels of carbendazim administered were 0, 100,
500 and 2 500 ppm. There were six male and six female rats per group.
After each female had been exposed to three males from the same dosage
group they were separated to produce the F_1A generation. Litters were
reduced to 10 pups/litter on the fourth day after birth. The F₀
animals were mated again after approximately one week in order to
produce the F_1B litter. Reproduction indices, litter data at birth
and on days 4, 12 and 21 were recorded, along with body weights at
weaning.

Data presented were extremely limited and submitted as group data
only. There were no pregnancies at 100 ppm for either F_1A or F_1B
matings. There were no apparent effects on the reproduction indices or
weanling weights. However, the fertility index for all groups, which
was 33-67 percent, prevents any meaningful interpretation of the data
(Sherman 1968).

Groups of ChR-CD rats (three male and 16 female rats/group; high-
dose group was 20 females) were fed carbendazim in the diet at dosage
levels of 0, 100, 500, 5 000 and 10 000 ppm and subjected to a
standard two-litter per generation, three-generation reproduction
study. Animals were fed from 21 days of age until 100 days of age,
when they were mated to initiate the study. Number of matings,
pregnancies and young in each litter at birth, and on days 4, 12 and
21 were recorded, along with body weights of pups at weaning. Litters
were culled to 10 pups/litter on day 4. After one week, F₀ parents
were mated again to produce the F_1B litters, after the F_1A had been
sacrificed. The F_1B litters were maintained on their respective diets
for 110 days and then mated to produce the F_2A and F_2B litters. The
F_3A and F_3B litters were similarly produced. Gross and
histopathological examination of selected tissues and organs were
performed on two males and two females in each of five litters from
the control, 5 000 and 10 000 ppm dose group pups from the F_3B
litter.

Reproduction indices, including mating, fecundity, fertility,
gestation, viability and lactation, were calculated and compared with
control values. There were no compound-related effects on any of the
reproduction indices other than reduced average litter weights at
5 000 and 10 000 ppm in all generations at weaning. Histopathological
examination of F_3B weanlings did not reveal any effects that were
considered compound related. MBC is considered to be without adverse
effects on reproduction in the rat when administered in the diet at
dose levels up to and including 500 ppm (Sherman 1972).

Carbendazim was administered to groups of Wistar Rats (10 males
and 20 females/group) at dietary levels of 0, 150, 300 and 2 000 ppm
for three generations. Two successive litters were reared from each
female. General condition and behaviour were routinely observed and
individual body weights were recorded throughout the study. The number
of pups in each litter were recorded and culled to eight on day 1.
Total weight of each litter was measured at days 1, 10 and 20. The
F_1A and F_2A litters were discarded at weaning and the F_1B and
F_2B litters were used to produce succeeding generations. The F_3A
offspring were selected for use in a teratology study, while F_3B
offspring were used in a 4-week short-term toxicity evaluation.

Health and body weight gain were not affected by carbendazim.
However, treatment groups weighed significantly more than controls in
all generations. There were no compound-related effects on fertility
or survival at birth, day 10 or day 20. Litter size was not affected
by treatment, except for a marginal decrease in F_2A litters in all
dose groups. There were no differences in the F_2B litters at 300 and
2 000 ppm; however, there was a decrease in litter size and increase
in mortality at birth at 150 ppm. Birth weight during the lactation
period was comparable among all groups. There were no gross
abnormalities related to treatment.

Autopsy of rats in the four week short-term feeding study
demonstrated increased relative liver weights and decreased relative
spleen weights in females fed 2 000 ppm. There were also significant
decreases in relative ovarian weights for females in all dose groups.
Histopathology of the livers did not indicate any compound-related
changes. There was no histopathology of the other organs presented.

There was no maternal or feto-toxicity evident in the teratology
portion of the study. There were similarly no differences in visceral
anomalies at 0 or 2 000 ppm (only groups examined). Thoracic vertebral
bodies were reduced at 2 000 ppm and a significant reduction of the
cervical vertebral bodies at 2 000 ppm. However, controls presented
more significant changes throughout with regard to absent or delayed
ossification of skeletal structures.

Although there were no apparent adverse effects on reproduction
and no teratogenic effects at dietary levels of carbendazim up to and
including 2 000 ppm, there were no individual animal data presented.
Histopathology of animals in the four-week study was incomplete and
did not include evaluations of spleen or ovaries. Such additional data
are needed to confirm the absence of adverse effects in this three-
generation reproduction study in rats (Koeter et al. 1976).

Special Studies on Teratogenicity

Rat

Groups of ChR-CD rats (27-28 pregnant rats/group) were
administered carbendazim (53 percent a.i.) in their diet at dosages of
0, 100, 500, 2 500, 5 000, 7 500 and 10 000 ppm, from day 6 through
day 15 of gestation. Average doses were equivalent to 0, 8.9, 45.9,
218.4, 431.6, 625.5 and 746.9 mg/kg day, respectively. On day 20 of
gestation all pregnant animals were sacrificed and foetuses delivered
by Caesarean section.

Determination of the number and location of live/dead foetuses
and resorption sites were performed, as well as body weights,
crown-rump length, sex and external examination for visible
abnormalities. Two thirds of the foetuses were prepared for
examination of skeletal abnormalities and the remaining ones were
examined for visceral and soft tissue anomalies.

There was no mortality, no adverse effect on body weight or
clinical signs of toxicity. Food intake was reduced in the 10 000 ppm
group during the period the test diet was administered, but returned
to comparable control levels from days 16 to 20. The data related to
reproduction (implantation sites, resorption sites and live/dead
foetuses) were not adversely affected by carbendazim. There were no
external or internal abnormalities reported that were considered
compound related. There was no individual litter data presented. It
was concluded that carbendazim was not teratogenic when administered
to ChR-CD rats at dietary levels up to and including 10 000 ppm during
the critical period of organogenesis (Sherman et al. 1970).

Groups of pregnant Wistar-SPF rats (18-22 females per group) were
administered carbendazim in the diet at dosage levels of 0, 600, 2 000
and 6 000 ppm from days 6 through 15 of gestation. On day 21 of
gestation all pregnant rats were sacrificed and pups delivered by
Caesarean section.

Dams were weighed periodically during the test and food
consumption measured for specific periods. The number of corpora lutes
were determined, ovaries weighed and foetuses weighed and examined.
The number of implantation and resorption sites were recorded and the
empty uterine horns weighed. One third of the foetuses were fixed and
stained for skeletal examination and the remaining two thirds were
examined for soft tissue anomalies.

Although 23 females per group were mated, the pregnancy rate was
variable, with 18, 22, 20 and 18 pregnant in the 0, 600, 2 000 and
6 000 ppm groups, respectively. The mean body weight gain and food
consumption for the high-dose females were significantly decreased in

comparison with controls. The number of live/dead foetuses,
implantation sites, embryonal resorptions, foetal resorptions and
corpora lutea/dam were comparable among all groups. Ovarian weights
and weight of the empty uterus were not affected by treatment. The
mean foetal weight/litter and sex ratio were comparable among all
groups. Pre- and post-implantation losses were not affected by
treatment.

No visceral anomalies were reported that were significantly
different from the control response. Misshapen and fused bones were
much more frequent occurrences in the high-dose groups then any of the
other treatment or control groups. Supernumery ribs were also
significantly increased in high-dose females. Ossification was
significantly delayed or absent in high-dose group pups, particularly
for forelimb, hindlimb, sternebrae and skull bones. Ossification was
significantly delayed or absent in cervical vertebral bodies in all
treatment groups when compared with control pups.

There were no individual animal or litter data and variations in
ossification and other skeletal abnormalities were presented as
percentages. The teratogenic or fetotoxic potential of carbendazim to
pregnant Wistar-SPF rats, therefore, cannot be determined from the
results and data presented (Koeter 1975a).

Rabbit

Groups of pregnant New Zealand albino rabbits (3-11 females/
group) were administered carbendazim in the diet at dosage levels of
0, 600, 2 000 and 6 000 ppm from day 6 through day 18 of gestation. On
day 29 of gestation, all pregnant animals were sacrificed and foetuses
delivered by Caesarean section.

Does were weighed periodically and food consumption was
determined for specific periods. The number of corpora lutea were
determined, ovaries weighed and foetuses weighed and examined. The
number of implantation sites and resorption sites were recorded and
the empty uterus weighed. One half of foetuses were stained and
sectioned for skeletal anomalies and the other half examined for soft
tissue abnormalities. Only the foetuses in the high-dose and control
groups were examined for visceral anomalies.

Although 18 females per group were artificially inseminated the
pregnancy rate was extremely variable among groups, with 9 in control,
8 in 600 ppm, 3 in 2 000 ppm, and 11 in the 6 000 ppm group. The mean
body weight gain was significantly decreased in the high-dose group,
although food consumption did not vary among groups. The number of
live/dead foetuses, implantation sites, embryonal resorptions, foetal
resorptions and corpora lutea/dam were comparable among all groups.
The ovarian and uterine weights in the high-dose group were depressed
in comparison with control females. Pre- and post-implantation losses
were not affected by treatment.

There were apparent differences between high-dose and control
groups for visceral anomalies. However, too few litters and foetuses
were examined to enable making any conclusions.

There was a significant increase in the number of supernumery
ribs (bilateral) and skull bones in the high-dose group. Ossification
was significantly delayed or absent in high-dose group foetuses, most
notably in the forelimb metacarpals and phalanges. There was also
incomplete ossification of the sternebrae and skull bones, which was
significant at 600 ppm and 6 000 ppm. Misshapen sternebrae were also
present in the 6 000 ppm group.

There were no individual animal or litter data, variations in
ossification were presented as percentages and visceral anomalies were
evaluated in only 2/4 of the groups. The teratogenic potential of
carbendazim to pregnant New Zealand albino rabbits, therefore, cannot
be ascertained from the results and data presented (Koeter 1975b).

Special Study on Neurotoxicity

A neurotoxicity study performed using chickens gave no indication
of neurotoxic potential at single oral doses up to and including
5 000 mg/kg (Goldenthal et al. 1978).

Long-Term Studies

Rat

Groups of weanling rats (36 male and 36 female ChR-CD albino
rats/group) were administered carbendazim (50-70 percent a.i.) in the
diet for 104 weeks at dosage levels of 0, 100, 500, 2 500-10 000, and
5 000 ppm. Growth was observed by body weight changes and food
consumption data, which were recorded weekly for the first year and
twice a month thereafter. Daily observations were made with respect to
behavioural changes and mortality. At periodic intervals throughout
the study, haematologic, urinalysis and selected clinical chemistry
examinations were performed. After one year each group was reduced to
30 male and 30 female rats by interim sacrifice for gross and
microscopic evaluations. At the conclusion of the study all surviving
animals were sacrificed and gross pathological examination of tissues
and organs was made. Microscopic examination of all tissues and organs
from the control and 2 500 ppm groups were conducted, along with liver
only from the 100 and 500 ppm groups, and liver, kidney testes and
bone marrow from the 5 000 ppm group animals.

The few mortalities observed in the first year were not
attributable to the presence of carbendazim in the diet. Survival
decreased during the second year to approximately 50 percent for males
and 39 percent for females, but was not related to treatment. Body
weight gain was depressed for males and females in the 2 500-

10 000 ppm group and for females in the 5 000 ppm group when compared
to control groups. Food consumption did not differ among groups. The
average daily dose for the 500 ppm group was 65 mg/kg b.w./day
(initially, M and F), 18 mg/kg (at one year) and 15 mg/kg (at two
years). Haematologic examinations demonstrated reduced erythrocyte
count, haemoglobin and haematocrit values for females at 9-24 months
in the 2 500 and 5 000 ppm groups; and for males at 24 months in the
2 500 ppm group. There were no compound-related clinical
manifestations of toxicity and no effects observed in urinalysis
examination. Alkaline phosphatase and glutamic pyruvic transaminase
activities varied throughout the test at 2 500 and 5 000 ppm but did
not demonstrate a consistent dose response. There were no apparent
differences in the organ weights or organ-to-body weight measurements,
except for female livers in the 2 500 and 5 000 ppm group. This
increase in the liver-to-body weight ratio was reflective of lower
body weights for both groups and therefore, not compound related.
Histopathologic evaluation of the livers did not demonstrate any
compound-related effects. Histopathologic examinations demonstrated an
increased incidence of pigment deposition in spleen and bone marrow
for both males and females at the 5 000 ppm level. This is consistent
with the haematology data for the same group. Males in the 2 500-
10 000 ppm group presented marginal increases for diffuse testicular
atrophy and prostatitis. Carbendazim is considered to be without
adverse effects on ChR-CD rats when incorporated in the diet at levels
up to and including 500 ppm (Sherman).

Groups of Wistar rats (60 males and 60 females/group) were
administered carbendazim (99 percent pure) in the diet at dosage
levels of 0, 150, 300 and 2 000 ppm for two years. The 2 000 ppm dose
was increased to 5 000 ppm after one week and then to 10 000 ppm after
two weeks for the remainder of the study. Clinical signs of toxicity
and general health were determined daily. Body weight and food
consumption were measured regularly throughout the study. Haematology
(peripheral blood), blood chemistry (orbital sinus) and urinalysis
evaluations were periodically conducted during the study. All animals
were subjected to complete gross necropsy and selected organs weighed.
A complete list of tissues and organs was prepared and examined
microscopically in 20 male and 20 female rats of the control and high-
dose groups. All tumours and gross abnormalities were also examined
histologically.

There were no differences between test groups and control animals
concerning clinical signs of toxicity or food consumption. Body
weights were significantly reduced in low-dose males at week 88 to
term and in high dose females at week 12 to term. Urinalyses and
kidney function (specific gravity) were comparable among all groups.
Of the haematological measurements examined, Hgb was depressed in
high-dose females at week 26, 52 and 103 and PCV was depressed in
high-dose females at week 103. There were no compound related effects
in males. SGOT activity was decreased in high-dose males at term, but

not in females. High-dose females had increased SGPT activity and
decreased total serum protein at term. There were no compound-related
effects on organ weights except for increased relative liver weights
in high-dose females. There were also no compound-related effects on
mortality, with 50 percent mortality in control males at week 76, and
at week 92 in treated group males. There was 50 percent mortality in
control and low-dose females at week 88 and at 92-96 weeks in mid- and
high-dose females. Survival at termination of the study was comparable
among all groups.

There were no measurable histological differences between control
and treated groups, except for an increased incidence of diffuse
proliferation of parafollicular cells of the thyroid in the high-dose
females. The number of tumour-bearing animals and total number of
primary tumours were comparable among all groups, and there were no
compound-related oncogenic effects reported. (NOTE: All data presented
were group mean values with SD. There were no data on individual
animals.)

The no observed effect level (NOEL) is 300 ppm, based on body
weight changes, decreased Hgb and PCV values and increased relative
liver weight in high-dose females. There was no tumourigenic effect in
this strain of rat at doses up to and including 10 000 ppm for 104
weeks (Til et al. 1976).

Dog

Groups of beagles (four males and four females/group) were
administered carbendazim (53 percent a.i.) in the diet at dosage
levels of 0, 100, 500 and 2 500 ppm for two years. Dogs were one to
two years of age at the start of the test. Some dogs in the high-dose
group received only 1500 ppm. Food consumption and body weight data
were obtained weekly and animals were examined daily for clinical
signs of toxicity. Haematological, biochemical and urinalysis
examinations were performed periodically throughout the study. Interim
sacrifice after one year was performed on one male and one female from
the control and 500 ppm groups, as well as one female from the high-
dose group. One male from the high-dose group was sacrificed in
extremis after 42 weeks on the test diet. Organ weights, gross
necropsy and histopathological evaluations were performed at the
conclusion of the study. Only the livers and testes were examined
histologically in the 100 and 500 ppm dose groups.

There was no mortality reported for the control or 100 and
500 ppm dose groups. However, three males in the high-dose group were
sacrificed after 22 and 42 weeks because of poor nutrition. No females
in the high-dose group died. Body weight and food consumption were all
adversely affected in the high-dose group animals, but not at lower
levels. The average daily intake for the 500 ppm dose group was

15.0-20 mg/kg (initially, M and F), 14-18 mg/kg (one year) and
10-16 mg/kg (two years). Dogs in the highest dose group developed
anorexia, distended abdomens and overall poor nutritional condition.
Haematological evaluations and urinalyses were not apparently affected
by treatment. The dogs in the 500 ppm and 1 500-2 500-dose groups had
increased cholesterol, BUN, total protein, GPT and APase levels and
presented evidence of a decreased A/G ratio throughout the study. This
biochemical evidence of liver effect was supported by liver pathology,
with incidences of hepatic cirrhosis, swollen vacuolated hepatic cells
and mild chronic hepatitis in dogs fed 500 ppm or more of carbendazim.
There were no noticeable effects on organ weights and organ-to-body
weight ratios. Diffuse testicular atrophy (which was marked) and
aspermatogenesis were observed in 2/4 males at 100 ppm but were not
present in the other dose group or in control males. Based on the lack
of supporting data in the other dose group males, these findings are
not considered as being compound-related.

The NOEL in this study appears to be 100 ppm. based on the liver
effects noted at 500 ppm and greater (Sherman 1972).

Groups of beagles (four males and four females/group) were fed
carbendazim in the diet at dosage levels of 0, 150, 300 and 2 000 ppm
for 104 weeks. After 33 weeks the 2 000 ppm dose was increased to
5 000 ppm. Dogs were 22-27 weeks old at the start of the study. Daily
examinations were made for clinical signs of poisoning and adverse
behaviour. Growth, as evidenced by body weight, was recorded regularly
throughout the study, as were food consumption data. At periodic
intervals (weeks 13, 26, 52, 78 and 104), haematology, blood chemistry
and urinalysis were performed. Liver function (BSP retention) and
kidney function (phenol red excretion) tests were evaluated at weeks
26, 52 and 104. At the conclusion of 104 weeks of dietary
administration, each dog was sacrificed and gross and microscopic
examination of tissues and organs were performed.

There was no mortality in any group except for one female in the
high-dose group which was killed in a moribund state after week 36.
Growth, as measured by body weight, was decreased in mid-dose males
and high-dose males and females. Food consumption was comparable among
all groups. Blood clotting times were significantly reduced in high-
dose males from week 13 to term, with slight decreases noticed in
high-dose females. Serum alkaline phosphatase activity was increased
in the high-dose group dogs throughout the study. There were no
compound-related effects on SGPT or SGOT levels. All other
haematological and blood chemistry measurements were comparable with
control groups. There were no differences among groups for BSP
retention, phenol red excretion or urine analyses.

Absolute liver and thyroid weights were significantly increased
in high-dose group dogs. Relative liver, thyroid and pituitary weights
were also significantly increased at the high dose. There were no

reported microscopic changes in these organs related to treatment.
There was an increased incidence of prostatitis (3/4 vs 1/4) in high-
dose males compared with controls. Also noted in 1/4 high dose males
was interstitial mononuclear inflammatory cell infiltrates and
atrophic tubules of the testes.

(Summary tables only were provided for the number of dogs with
the indicated pathological response. Severity of response, identity of
dog involved, gross and histopathology reports of individual animals
were not provided. Data provided were generally not separated
according to sex.)

The feeding of carbendazim in the diet to dogs for two years was
without apparent adverse effects at levels up to and including 300 ppm
(Reuzel et al. 1976).

Special Studies for Carcinogenicity

Groups of CD-1 mice (80 males and 80 females/group) were
administered carbendazim (99 percent a.i.) in the diet at dose levels
of 0, 500, 1 500 and 7 500 ppm for two years. The 7 500 ppm dose was
reduced to 3 750 ppm after 66 weeks for the males because of increased
mortality. Females received 7 500 ppm throughout the study period.
Animals were 6-7 weeks old at the start of the study. Mice were
examined daily for behaviour and clinical signs of toxicity, biweekly
for palpable masses and regularly weighed for body weight changes.
Food consumption was similarly determined on a routine basis.
Mortality was noted and recorded. Peripheral blood was collected
periodically throughout the study for haematological examinations.
Selected organs were weighed, including brain, heart, lungs, liver,
spleen, kidney, testes and thymus. Microscopic examination was
performed on a complete list of tissues and organs. Urine and faecal
samples were also analysed.

Mortality was compound related in male mice. The high-dose group
males terminated at week 73 because of significant increase in
mortality. Only nine males in the 1 500 ppm group survived to week
104, compared with 18 for control males. Females were unaffected by
treatment in this respect.

There were no dose related effects on body weight or food
consumption throughout the study, although terminal body weights for
low- and mid-dose group males were less than control and high-dose
group males. Clinical signs of toxicity were similar among all
treatment and control groups. Haematological determinations in males
were unaffected by treatment. Females in the 7 500 ppm group had
reduced erythrocyte counts and marginal decreases in haemoglobin
concentration.

Both absolute and relative thymus weights were significantly
decreased in females in the 500 and 1 500 ppm groups, but not in the
high-dose group. Absolute liver weight was increased in the 7 500 ppm
females, with relative liver weight increased in the 1 500 and
7 500 ppm groups. Organ weights for the males were variable with only
the kidney and thymus weights apparently decreased by treatment.
Absolute kidney and thymus weights were depressed in all male
treatment groups. However, relative kidney and thymus weights were
significantly decreased in the high-dose males only. The lower
absolute kidney and thymus weights in the low- and mid-dose group
males were probably a reflection of reduced terminal body weights.

Histological examination revealed dose-related changes in the
thymus (lymphoid depletion) and kidneys (bilateral/unilateral
accumulation of yellow-brown pigment in the tubules) for mid- and
high-dose group male mice. Examination of the testes demonstrated a
marginal increase in the finding of sperm stases (bilateral/unilateral
combined) in treated males, with a similar finding of increased
germinal cell atrophy (bilateral only). There was an opposite trend,
however, for unilateral germinal cell atrophy, where the incidence in
controls was greater or equal to treated males. These effects are,
therefore, not considered compound related.

Examination of livers of male mice revealed a significant
hepatotoxic effect at 1 500 and 7 500 (3 750) ppm, demonstrated by
centrilobular hypertrophy, necrosis and swelling. There was no
increase in the finding of hepatocellular adenoma, which occurred with
equal frequency in control and treatment groups. A significant
increase occurred for hepatocellular carcinomas at the 1 500 ppm dose
only. However, too few high-dose males survived to 17 months
(510 days) to support the conclusion of no oncogenic effect at that
dose level.

Histomorphic evaluation of the female mice revealed an increased
incidence of lymphoid depletion in the thymus in the mid- and high-
dose groups. There was a significant accumulation of yellow-brown
pigment in the macrophages and tubules in the kidneys, as well as an
increase in cystic tubules for high-dose group females. The occurrence
of hepatocellular carcinomas was significantly increased in the mid-
and high-dose females. However, there was no apparent compound-related
effect on the latency period for this finding. The finding of
hepatocellular adenomas was marginally increased in the low- and mid-
dose females, but not in the high-dose group, compared with control.
Other findings indicative of hepatotoxicity were more prominent in the
control females than in the treatment groups. Hepatocellular chromatin
aggregation and necrosis (focal, multifocal, single cell) were
increased in controls. There was also a significant increase for the
incidence of macrophages containing yellow-brown pigment. These
findings appear to indicate different metabolic or detoxification
mechanisms, which are sex dependent. The carcinogenic response in the

liver, although significant at 1 500 and 7 500 ppm for females and at
1 500 ppm for males, is considered a weak response in light of the
histomorphic changes in the livers in male and female control group
mice (Wood 1982).

Carbendazim was administered in the diet to groups of SPF Swiss
mice (100 males and 100 females/group) at dosage levels of 0, 150, 300
and 1 000 ppm for 80 weeks. The 1 000 ppm dose was increased to
2 000 ppm at week 4 and to 5 000 ppm at week 8 for the remainder of
the study. Animals were observed for behaviour and clinical signs of
toxicity. Body weight measurements were determined throughout the
study. Gross necropsies were performed on all animals, liver and
kidney weights recorded, and a complete list of organs and tissues was
examined microscopically.

There were no compound-related effects on general condition,
mortality or body weight. Survival at term was 70 percent for males
and 80 percent for females. Relative liver weights in high-dose males
and females were significantly different from controls. There were no
changes in kidney weights. Gross and histopathology examinations
demonstrated a compound-related effect on the livers of both male and
female mice in the high-dose group. There was a significant increase
in the number of mice with clear cell foci in high-dose males and
females and in mixed cell foci for high-dose males. Neoplastic nodules
were reportedly increased in high-dose females, while the incidence of
hepatoblastoma was increased in high-dose males. There were no
differences between control and treatment groups for the finding of
hepatocellular carcinoma. It was concluded that carbendazim is
oncogenic to this strain of mouse at dietary doses greater than or
equal to 5 000 ppm (Beems et al. 1976). (NOTE: All data presented
were group mean values. There were no data on individual animals.)

Carbendazim was administered in the diet to groups of HOE NMRKf
(SPF 71) mice (100-120 males and females/group) for 96 weeks at dosage
levels of 0, 50, 150, 300 and 1 000 ppm. The 1 000 ppm dose was
increased to 2 000 ppm at week 4 and to 5 000 ppm at week 8 for the
remainder of the study. Animals were observed for behaviour and
general condition, as well as body weight, food/water consumption and
mortality. Gross necropsies were performed on all animals, liver and
lung weights were recorded, and a complete list of organ and tissues
was examined microscopically. An interim sacrifice was made at 18
months on 20 males and 20 females from the control group and the
5 000 ppm group.

There were no compound-related effects on behaviour, body weight
gain, food/water consumption or mortality. At 22 months there was
24-31 percent mortality in male mice and 37-52 percent mortality in
females, for all groups. Mean daily consumption of carbendazim in
mg/kg was 5.8-7.1 at 50 ppm, 17.1-21.2 at 150 ppm, 34.4-41.9 at
300 ppm, and 548.4-682.3 at 5 000 ppm.

Examination of lung and liver weights at 18 and 22 months
demonstrated an increase in absolute and relative liver weights in
both male and female mice at 5 000 ppm.

Macroscopic and microscopic examination of animals at 18 months
revealed compound-related effects on the liver at 5 000 ppm. There
were reported increases in centrilobular hypertrophy of liver cells,
single cell necroses, liver cells in mitosis and pigment in Kupffer
cells. Controls presented evidence of fatty change of liver cells
only. Microscopic evaluation of tissues/organs at 22 months
demonstrated a definite compound-related effect on liver at 5 000 ppm
in both males and females. There was marked liver cell hypertrophy,
clear cell foci, liver cells in mitosis, abundant inclusion bodies in
enlarged cell nuclei, multiple cell necroses and greenish yellow
pigment in Kupffer cells. Neoplastic nodules (adenomas), carcinomas,
fibrosarcomas and other tumourigenic responses in the liver were
equally distributed among all groups. The occurrence of hemangiomas,
evident in treated groups with none in the control groups, was
randomly distributed (both by dose and sex), not dose related, not
significantly different from control and, therefore, not considered
compound related. The finding of lung neoplasias (such as
adenomatosis) were equally distributed among all groups. There was no
effect on incidence or time of onset of tumours by carbendazim and the
total number of benign and malignant tumours were comparable among
groups. There was a significant increase in liver toxicity in both
males and females at 5 000 ppm. However, there was no evidence of a
carcinogenic effect from carbendazim when administered in the diet to
mice at doses up to and including 5 000 ppm for 22 months (Kramer &
Weigand 1982).

Special Studies on Mutagenicity

Results of the various mutagenicity assays are summarized in
Table 2.

Bacteria

MBC was examined for mutagenic activity in Salmonella
typhimurium following the plate incorporation protocol of Ames
et al. (1975), at concentrations between 1 and 325 µ/g plate, with
and without activation. MBC increased the reversion frequency 3-5
times the control frequency, without activation. Doubtful activity was
found with activation. Source and purity of test substance were not
provided and there were no actual data presented (Rashid & Ercegovich
1976; Ercegovich & Rashid 1977).

MBC was tested for mutagenic activity on S. typhimurium,
strains TA1535, TA1537, TA98 and TA100, with and without activation.
MBC, dissolved in DMSO, was not mutagenic at concentrations between 4
and 2 500 µg/plate (Hoechst 1977).

Table 2 Mutagenicity Assays

Test Organism Test Result Reference
Substance

Gene Mutation Studies

Bacteria

Salmonella typhimurium MBC Bacterial assays with MBC. Ercegovich & Rashid
Strains TA98, TA100, TA- 1977
1535, TA1537, and TA1538.
Doubtful mutagenic activity Rashid & Ercegovich
was reported for MBC both 1976
with and without metabolic
activation.

Negative Shirasu et al. 1977

Negative. Results dependent Russell 1977a,b,
on sample source and purity. 1978

Negative. Hoechst 1977

Series of tests: spot and Ficsor et al. 1978
liquid culture assays using
strains his G46 and TA1530,
TA1535, TA1950. No mutagenic
activity except one weak
positive in his G46.

Table 2 (con't)

Test Organism Test Result Reference
Substance

Plates treated with 100 to Donovan 1981a
10 000 ug MBC, with activation.
The number of revertants/
plate increased from
4.2 to 8.95 times in the
trials with positive responses
in TA98 and from 3.7 to
6.4 times in TA1537.

MBC Negative. Same as previous Donovan 1981b
(Hoechst) citation, with and without
activation. Results dependent
on sample source and purity.

S. typhimurium
(host mediated assay) MBC Negative Shirasu et al. 1977

Yeast and Fungi

Aspergillus nidulans MBC Positive at pH 5.2 and 5.3 Speakman & Nirenberg
1981
Nirenberg & Speakman
1981

MBC Positive. Kappas et al. 1974
MBC Positive. Davidse 1973

Table 2 (con't)

Test Organism Test Result Reference
Substance

Cladosporium
cucumerinum MBC Positive at pH 6.8 Speakman & Nirenberg
1981
Nirenberg & Speakman
1981

Cultured Mammalian Cells

Chinese hamster ovary MBC Negative Waterer & Krahn 1980
cells in vitro

Insects

Drosophila melanogaster MBC Noted sterility in some Lamb & Lilly 1980
broods. This was considered
to be consistent with
spindle effects of MBC.

Mammals

Mouse, in utero MBC Positive - coat colour Fahrig & Seiler 1979
changes

Chromosomal effects

Cytogenetics-in vitro
Human lymphocytes MBC Grown in culture medium Lamb & Lilly 1980
containing 0.5 mg MBC.
No compound related chromosome
aberrations.

Table 2 (con't)

Test Organism Test Result Reference
Substance

Mouse lymphoma L5178Y cells MBC Dose-related increase in Jotz et al. 1980
mutation frequency with
metabolic activation at
TK+/- locus at 100 µM.

Mouse lymphoma L5178Y cells MBC MBC was not mutagenic at the Krahn et al. 1983
TK+/- locus with or without
activation at concentrations
up to and including 200 µM.

Cytogenetics - in vitro

Rat bone marrow MBC Negative. BASF 1975a

Chinese hamster bone marrow MBC Negative. Seiler 1976

Mouse bone marrow MBC Negative. Seiler 1976

Dominant lethal-rodents

Rat MBC Negative. Benes & Sram 1976
Mice MBC Negative. BASF 1975b
Negative. Hoechst 1974

Micronucleus Test

Mouse bone marrow MBC Positive. Seiler 1976

Table 2 (con't)

Test Organism Test Result Reference
Substance

DNA Damage and Repair
Mice B6C3F1 and F344 MBC MBC was tested for DNA Tong 1981a,b
repair using primary hepatocyte
cultures. MBC did not
induce DNA repair in either
rat or mouse.
Differential Toxicity-
Bacteria

Bacillus subtillis MBC Negative. Shirasu et al. 1977

Gene Mutation-Bacteria

S. typhimurium 5-hydroxy-MBC Negative. Cannon Laboratories
1978

Negative Russell 1977b
Plant Studies

Allium cepa MBC Positive. Richmond & Phillips
1975

MBC and some of its commercial preparations were examined for
mutagenic potential in S. typhimurium following different
treatment protocols. An overlay spot test was used to test MBC at
concentrations of 50 and 100 µg/spot in strains his G46, TA1530 and
TA1950. Only one sample of MBC (Seiler) exhibited weak mutagenic
activity at 100 µg/spot. In a plate incorporation assay using strain
TA100, MBC was not mutagenic at concentrations between 50 and
200 µg/plate. Liquid culture assays with 1 000 µg/ml of MBC showed no
evidence of mutagenicity in his G46 and TA1950 (Ficsor et al.
1978).

MBC was non-mutagenic in S. typhimurium strains TA1535,
TA1537, TA1538, TA98 and TA100, and in Escherichia coli strain WP2
hcr. Concentrations between 5 and 1 000 µg/plate were tested in a
plate incorporation assay with DMSO as the solvent both in the
presence and absence of an activation system, which included a
9 000 x g supernatant fraction of homogenized livers from Aroclor
1254-treated Sprague-Dawley rats (Shirasu et al. 1977).

MBC and 5-hydroxy-MBC were tested for mutagenic activity in
S. typhimurium strains TA1535, TA1537, TA1538, TA98 and TA100
according to the plate incorporation procedure of Ames et al.
(1975). DMSO was the solvent. Each sample was evaluated in the
presence and absence of an activation system. Five different samples
of MBC were tested at concentrations up to 10 mg/plate. A sample of
technical grade MBC was mutagenic in the presence of rat liver
homogenate in strains TA1537, TA1538 and TA98. A 99.6 percent pure MBC
sample, in the presence of rat and mouse activation systems, was
mutagenic in TA1537, TA98, TA1535 and TA100. Rat and mouse liver
activation systems gave essentially identical results in TA1537 and
TA98. A third sample (97.6 percent, Czech-Polish) was mutagenic with
rat liver activation in strains TA1537, TA98 and TA100. An MBC sample
from Hoechst (95-100 percent pure) was not mutagenic in any of the
Salmonella strains. Analytical grade MBC (99.5 percent pure) was
weakly mutagenic in TA1537 at concentrations up to 15 mg/plate. A
sample of 5-hydroxy MBC was not mutagenic at concentrations up to
20 mg/plate (Russell 1977 a, b, 1978; Donovan 1981 a, b).

In a host mediated assay in male ICR mice given total doses of
either 1 000 or 4 000 mg MBC/kg the mutation frequencies observed in
the S. typhimurium strain his G46 from treated animals were
identical to mutation frequencies in bacteria from control animals
(Shirasu et al. 1977).

Yeast and fungi

MBC was evaluated for mutagenic activity in Aspergillus
nidulans and Cladosporium cucumerinum. In A. nidulans, MBC
at a concentration of 2.77 µM (0.53 µg/ml), caused an increase in the

frequency of colonies resistant to MBC (2 µg/ml) but did not increase
the frequency of colonies resistant to carboxin (20 µg/ml). In C.
cucumerinum, MBC at 0.58 µM (0.11 µg/ml) caused an increase in
carboxin (20 µg/ml) resistant colonies, but had no effect on the
frequency of colonies resistant to MBC (0.8 µg/ml). An activation
system was not used for any of these studies (Speakman & Nirenberg
1981).

Experiments were conducted in A. nidulans and C.
cucumerinum to evaluate the effect of altering the pH of the agar
medium, using pHs of 5, 5.2-5.3 and 6.8. The pH of the treatment
medium had a significant effect on the activity of MBC, which was
mutagenic in A. nidulans only at a pH of 5.2-5.3 (MBC resistance)
and in C. cucumerinum only at pH 6.8 (carboxin resistance). The
concentrations of MBC exhibiting mutagenic activities were 0.53 µg/ml
and 0.11 µg/ml for A. nidulans and C. cucumerinum,
respectively. The effect of an activation system was not studied. The
authors concluded that MBC had weak mutagenic activity (Nirenberg &
Speakman 1981).

Cultured mammalian cells

Chinese hamster ovary cells in culture were exposed to varying
concentrations of MBC without activation (3 to 654 µM) and with
activation (3 to 628 µM), to detect mutations at the gene locus coding
for hypoxanthine guanine phosphoribosyl transferase (HGPRT). A dose-
related cytotoxic response was evident in cultures exposed to MBC
without activation, with a decreased survival at 16 µM. No
statistically significant differences in mutation frequency were noted
and MBC was not mutagenic under the test conditions used (Waterer &
Krahn, 1980).

Insects

MBC dissolved in DMSO at 0.5 mg/ml did not cause a significant
increase in the frequency of sex-linked recessive lethals when given
to Drosophila melanogaster. The only indication that the substance
may have some potential for damaging germ cells comes from the
observation that it caused an increased incidence of sterility in the
later broods from treated Oregon-R males. However, this effect was
not observed in treated yw^{+B^{sZy⁺ males. No compound-related}}
effects were noted when chromosomes were examined for breakage in a
second set of experiments and the overall incidence of recessive
lethal mutations reported was 5/4807 (0.1 percent). The sterility
observed in broods from matings involving mitotic spermatogonial cells
is consistent with the suspected spindle effects of the chemical (Lamb
& Lilly 1980).

Mouse embryo

Mouse embryos, heterozygous for four different recessive coat
colour genes, were treated in utero by dosing the mother orally
with MBC at dose levels of 100 to 300 mg/kg b.w. If mutations are
induced in pigment precursor cells in a wild type allele of one of the
genes under study, a spot of an altered colour may appear on the coats
of the offspring. At 200 mg/kg the number of spots was significantly
different from controls (Fahrig & Seiler 1979).

Cytogenetics

The ability of MBC to cause chromosomal aberrations was evaluated
in human lymphocytes in culture. MBC, at a concentration of 0.5 mg/ml,
did not increase the frequency of chromosome aberrations over the DMSO
control in a system without activation. Cells treated with MBC did
exhibit grossly contracted chromosomes, an effect induced by spindle
poisons such as colchicine (Lamb & Lilly, 1980).

MBC, with and without metabolic activation, was evaluated for its
ability to induce forward mutations at the thymidine kinase (TK) locus
in mouse L5178Y lymphoma cells. Metabolic activation was accomplished
by microsomal enzymes obtained from induced rat liver preparations
(S-9 mix). Ethylmethane sulphonate (EMS) and 3-methylcholanthrene were
used as positive controls.

MBC was mutagenic in this test system with activation, the
mutation frequency being increased in a dose-related manner. There was
no mutagenic response without metabolic activation at doses of 50 to
250 µM. The results indicated that metabolic activation enhanced MBC's
mutagenic activity at 100 µM (Jotz et al. 1980).

The effects of MBC on the mouse lymphoma L5178Y cell line at the
thymidine kinase (TK^+/-) locus were examined both with and without
metabolic activation. Concentrations tested included 12.5 to 200 µM
of MBC, DMSO as negative control, 2 000 µM of ethylmethane
sulphonate (positive control without activation) and 15 µM of
3-methylcholanthrene (positive control with activation). The positive
controls gave the expected response. However, MBC, both with and
without activation tested in replicate trials, was not mutagenic at
levels up to and including 200 µM (Krahn et al. 1983).

Technical grade MBC was tested for its ability to cause
chromosome aberrations in rat bone marrow cells in vivo. Male and
female Sprague-Dawley rats were given a single oral dose of 300 mg/kg.
Metaphase cells from treated animals, sacrificed at 6, 24, and
48 hours after treatment, did not exhibit an increased frequency of
chromosome aberrations (BASF 1975a).

MBC did not produce chromosome breakage in bone marrow cells from
Chinese hamsters given oral doses of 1 000 mg/kg. Only one chromatid
break was observed in a total of 500 metaphases from four animals. The
mitotic figures were examined in bone marrow cells from ICR mice given
two oral doses of MBC at 1 000 mg/kg each. Twelve of the 1 000
nucleated anaphase cells exhibited lagging chromosomes, bridge
formation, tripolar spindle formation or unequal chromatin
distribution. MBC does not break chromosomes but probably exerts its
effect by interfering with spindle function (Seiler 1976).

Rodent dominant lethal test

Twenty NMRI mice were given intraperitoneal injections of MBC
(500 mg/kg) on five successive days. A 0.5 percent solution of the
vehicle CMC was given to 20 mice and served as controls. Animals
receiving MBC did not exhibit any clinical signs of toxicity. The body
weights of control and MBC-treated animal groups were identical after
the first week of mating. No macroscopically observable pathological
changes were seen in dissected mice from the MBC-treated group. MBC
did not exhibit a dominant lethal effect (Hoechst 1974).

Twenty-two male NMRI mice were given MBC by stomach tube
(300 mg/kg) on five successive days. The vehicle for delivering MBC
was not given. The same number of untreated mice served as controls.
MBC-treated mice did not exhibit clinical symptoms of toxicity, body
weight changes or macroscopically recognizable pathological changes in
the internal organs. MBC did not cause a change in the mutagenicity
index over that of the control (BASF 1975b).

DNA damage and repair

DNA repair assays in rat (F344) or mouse (B6C3F1) hepatocyte
primary cultures (HPC) were evaluated for MBC along with
dimethylnitrosamine and 2-amino-fluorene, which were used as positive
controls. MBC and tritiated thymidine (10 µCi) were added to the
culture medium. After 18 to 20 hours of incubation they were fixed and
examined microscopically for morphological changes and absence of
S-phase nuclei indicative of cytotoxicity. Autoradiographic techniques
were used to determine the number of nuclei grains induced. MBC did
not induce DNA repair in rat or mouse hepatocytes. The positive
controls gave the expected response (Tong 1981a,b).

Differential toxicity to bacterial strains with different repair
capacities

MBC (typically 99 percent pure, sources unspecified) was tested
for toxicity to recombination repair-proficient and repair-deficient
strains of Bacillus subtilis. MBC was tested at concentrations
between 20 and 1 000 µg/disk. MBC did not cause a zone of killing in
either strain and, thus, was negative in the assay. The absence of

toxicity to either strain indicated that MBC was either non-toxic
under the test conditions or the limited solubility prevented
diffusion from the disk. An activation system was not used (Shirasu
et al. 1977 ).

COMMENTS

Carbendazim follows a similar metabolic pathway to benomyl in
rate and mice, being excreted in urine as 5-hydroxy carbendazim
(5-HBC). Enzyme induction studies demonstrate that the rat is more
efficient than the mouse in metabolizing and eliminating carbendazim
and its metabolites.

Carbendazim is not acutely toxic to mammals as demonstrated by
acute oral and dermal LD₅₀s of >10 000 mg/kg in rat and rabbit,
respectively. Gross and histopathological examinations performed in
many of these acute studies indicated that doses >1 000 mg/kg
produced adverse effects on the testes (small, soft, discoloured,
degenerative changes of the tubules) and epididymides (reduced or
absent sperm).

A three-generation reproduction study in rats demonstrated a NOEL
of 500 ppm, with higher doses resulting in reduced average litter
weights.

Teratology studies in which the test material was administered in
the diet of rats indicated in one study the absence of induction of
terata at 10 000 ppm and in the second study a low incidence of
misshapen, fused or incompletely ossified bones at 6 000 ppm. A
limited study in rabbits did not indicate the induction of terata
following dietary administration at 6 000 ppm (see also benomyl).

A short-term dietary study in rats indicated increased liver to
body weight ratios in females at 2 500 ppm, although no compound-
related histomorphic changes were evident. There were no effects on
testicular weight and the NOEL was 500 ppm. Two short-term dietary
studies in dogs demonstrated that 300 and 500 ppm, respectively,
caused no adverse effects. However, at doses of 1 000 and 2 500 ppm
animals lost their appetite, lost weight and had increased cholesterol
levels and relative liver weight increases.

In two separate long-term feeding studies in rats, carbendazim
produced relative liver weight increases, deposition of pigment in the
spleen and bone marrow, and decreased haemoglobin, haematocrit and red
blood cell counts at the higher doses. It was without adverse effects
at 300 and 500 ppm, respectively, and there was no oncogenic response
at doses up to 10 000 ppm.

Beagles appeared to be more sensitive than rats to dietary
exposure to carbendazim. Hepatic cirrhosis, vacuolated hepatic cells
and increased levels of cholesterol, BUN, total protein, GPT and
alkaline phosphatase, with decreased A/G ratio, were evidence of liver
toxicity at levels greater than 100 ppm for two years.

Oncogenicity studies were performed using three strains of mice
(CD-1, Swiss SPF and HOE-NMR). In CD-1 mice there was a significant
increase in hepatocellular carcinomas at 1 500 and 7 500 ppm in
females and 1 500 ppm in males. However, there were also substantial
histomorphic changes in the livers of male and female control animals.
There was no oncogenic response at 500 ppm. Swiss mice, exposed for 80
weeks to 150, 300 and 1 000-5 000 ppm carbendazim, showed an oncogenic
response at 5 000 ppm, which was evidenced by significant increases in
the incidence of neoplastic nodules and hepatoblastomas. There were no
compound-related effects in this study at 300 ppm. HOE-NMR mice
exposed to 50-5 000 ppm carbendazim for 96 weeks presented no evidence
of an oncogenic response. It was concluded that carbendazim was
hepatocarcinogenic to mice at high dose levels.

Mutagenicity studies with carbendazim gave both positive and
negative results. Carbendazim was positive in the micronucleus, yeast,
fungi and Drosophila tests. Conflicting negative and positive results
in other tests prevented evaluation of the mutagenic potential. The
potential impact of these results on human health cannot be adequately
assessed at this time.

The data for benomyl and carbendazim have indicated that the
metabolism of the two compounds is essentially the same, with benomyl
converted rapidly to carbendazim in mammals. Accordingly, the
available data for benomyl and carbendazim should be considered
collectively for the evaluation of specific studies such as
teratology, reproduction, chronic toxicity and oncogenicity, taking
into account the different molecular weights of the two compounds.

Previous Meetings have considered the aetiology and pathogenesis
of liver tumours in certain strains of mice, with particular emphasis
on organochlorine pesticides (FAO/WHO 1970, 1973, 1976). It was
recognized that liver tumours are known to develop spontaneously in
many strains of mice, at relatively high incidence and without
intentional exposure to chemicals. Evidence of such tumours in several
strains of mice has been found in many of the oncogenicity studies
performed with benomyl and carbendazim. Furthermore, one strain of
mouse used (HOE-NMR) is known to have a low background incidence of
liver tumours (1-2 percent) and did not provide evidence for
oncogenicity when exposed to carbendazim at doses up to and including
5 000 ppm. Two additional studies have been carried out in rats using
both benomyl and carbendazim. Both studies were negative for
oncogenicity at doses up to and including 2 500 and 10 000 ppm,
respectively. The hepatic tumours produced in mice, therefore, appear
to be a species-related phenomenon.

The Meeting expressed concern at the equivocal nature of the
results of a wide range of mutagenicity studies. The possibility that
conflicting results were due to variations in the type and amount of
impurities was considered, but the Meeting was informed that current
levels of the impurities in question are very low in technical
material.

In view of established NOEL determined in several studies,
including teratology, reproduction and chronic feeding, an ADI for
both benomyl and carbendazim could be estimated. However, a safety
factor of 200 was used to reflect the concern of the Meeting for the
paucity of individual animal data for many studies on carbendazim.

TOXICOLOGICAL EVALUATION

Level Causing no Toxicological Effect

Rat: 500 ppm in the diet, equivalent to 25 mg/kg b.w.

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

Rat: Teratology (see benomyl )

Estimate of Acceptable Daily Intake for Man

0-0.01 mg/kg b.w.

FURTHER WORK OR INFORMATION

Desirable

1. Data on individual animals used in studies on carbendazim
that have been identified in this evaluation addendum.

2. Additional data to elucidate the mechanism of degenerative
testicular effects on mammalsœ

3. Elucidation of the variability of the mutagenicity data.

REFERENCES- TOXICOLOGY

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1975a (>95% MBC). Report HLR No. 847-74 submitted to WHO by
DuPont. (Unpublished)

Dashiell, O.L. Ten-day subacute exposure of rabbit skin to technical
1975b MBC (>98% MBC). Report submitted to WHO by DuPont.
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Davidse, L.C. Antimitotic activity of methyl benzimidazol-2-yl
1973 carbamate (MBC) in Aspergillus nidulans. Pestic.
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Donovan, S.M. Mutagenicity evaluation in Salmonella typhimurium using
1981b MBC. Report HLR 507-81 submitted to WHO by DuPont.
(Unpublished)

Dorn, E. et al. HOE 017411-14-C (carbendazim-14-C) metabolic fate in
1983 rats and mice, a comparison. Report submitted by BASF to
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Edwards, D.F. Acute dermal LD₅₀ test on rabbits using technical MBC
1974a (>98% MBC). Report HLR No. 799-74 submitted to WHO by
DuPont. (Unpublished)

Edwards, D.F. Eye Irritation test in rabbits using technical MBC (>98%
1974b MBC). Report HLR No. 799-74 submitted to WHO by DuPont.
(Unpublished)

Ercegovich, C.D. & Rashid, K.A. Mutagenesis induced in mutant strains
1977 of Salmonella typhimurium by pesticides. 174th
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Fahrig, R. & Seiler, J.P. Dose and effect of methyl
1979 2-benzimidazolylcarbamate in the "Mammalian Spot Test", an
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Falke, H.E., Beems, R.B. & Spit, I.B. Carbendazim - technical grade
1982a 59-day enzyme induction study in rats. Report from Central
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Falke, H.E., Beems, R.B. & Spit, I.B. Carbendazim - technical grade
1982b 60-day enzyme induction study in mice. Report from Central
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Ficsor, G., Bordas, S. & Stewart, S.J. Mutagenicity testing of
1978 benomyl, methyl 2-benzimidazolecarbamate, streptozotocin and
N-methyl-N'-nitro-N-nitro-soguanidine in Salmonella -
typhimurium in vitro and in rodent host-mediated
assays. Mutat. Res. 51: 151-164.

Ford, L.S. Skin irritation test on rabbits using a wettable powder
1981a formulation (75% MBC). Report HLR No. 728-81 submitted to
WHO by DuPont.(Unpublished)

Ford, L.S. Primary skin irritation and sensitization of guinea pigs
1981b using a wettable powder formulation (75% MBC). Report HLR
No. 729-81 submitted to WHO by DuPont. (Unpublished)

Ford, L.S. Acute dermal LD₅₀ test on rabbits using a wettable powder
1982 formulation (75% MBC). Report HLR No. 822-81 submitted to
WHO by DuPont. (Unpublished)

Goldenthal, E.I. et al. Neurotoxicity study in hens using technical
1978 MBC (>98% MBC). Report HLO-0027-79 by International
Research & Development Corp. submitted to WHO by DuPont.
(Unpublished)

Goodman, N.C. Intraperitoneal LD₅₀ test in rats using technical MBC
1975 (>98% MBC). Report HLR No. 845-74 submitted to WHO by
DuPont. (Unpublished)

Henry, J.E. Eye irritation test in rabbits using a wettable powder
1982 formulation (>75% MBC). Report HLR No. 66-82 submitted to
WHO by DuPont. (Unpublished)

Hinckle, L. Acute oral LD₅₀ test in rats using a wettable powder
1981 formulation (>75% MBC). Report HLR No. 769-81 submitted to
WHO by DuPont. (Unpublished)

Hoechst. Mouse dominant lethal study on MBC. Report submitted to WHO
1974 by BASF. (Unpublished)

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1977 submitted to WHO by BASF. (Unpublished)

Jotz, M.M. et al. An evaluation of mutagenic potential of MBC (>98%
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(Unpublished)

Kappas, A., Georgopoulos, S.G. & Hastie, A.C. On the genetic activity
1974 of benzimidazole and thiophanate fungicides on diploid
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Koeter, H.B.W.M. Effect of HOE 17411F on pregnancy of the rat. Report
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1975b from Central Institute of Nutrition and Food Research
submitted by BASF to WHO. (Unpublished)

Koeter, H.B.W.M., Til, H.P. & vander Heijden, C.A. Multigeneration
1976 study with carbendazim in rats. Report from Central
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Krahn, D.F. et al. L5178Y mouse lymphoma cell assay for mutagenicity.
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1982 determination of the cancerogenic effect of HOE 17411
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1980 toxicological effects of the fungicide benomyl. Toxicology,
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1981 of methyl benzimidazol-2-yl carbamate (MBC) towards
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Russell, J.F. Mutagenic activity of analytical grade MBC (>99% MBC) in
1978 the Salmonella/microsome assay. Report HLR No. 55-78
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1976 derivatives. Mutat. Res., 40: 339-348.

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(Unpublished)

Sherman, H. Ninety-day feeding study in rats using a wettable powder
1968 formulation (70% MBC). Report HLR No. 95-68 submitted to WHO
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Sherman, H. Three-month feeding study in dogs using a wettable powder
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1972 benzimidazolecarbamic acid, methyl ester (INE-965) (50% and
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1966 subacute oral test in rats using technical MBC (>98% MBC).
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(Unpublished)

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Environmental Toxicology submitted to WHO by DuPont.
(Unpublished)

Speakman, J.B. & Nirenberg, H.I. Mutagenicity of methyl benzimidazol-
1981 2-yl carbamate (MBC) towards Aspergillus nidulans
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1976 with carbendazim in rats. Report from Central Institute for
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(Unpublished)

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1981 carbendazim in mice. Report from Central Institute for
Nutrition and Food Research submitted by BASF to WHO.
(Unpublished)

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1981a 11,201-01 (>98% MBC) using mouse hepatocytes in culture.
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Tong, C. Hepatocyte primary culture/DNA repair assay on compound
1981b 11,201-01 (>98% MBC) using rat hepatocytes in culture.
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Waterer, J.C. & Krahn, D.F. Chinese hamster ovary cell assay for
1980 mutagenicity using MBC (100% MBC). Report HLR No. 660-80
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Wood, C.K. Long-term feeding study with 2-benzimidazole-carbamic
1982 acid, methyl ester (MBC, INE-965) in mice Parts I and II
(>99% MBC). Report HLR No. 70-82 submitted to WHO by
DuPont. (Unpublished)

RESIDUES

RESIDUES IN FOOD AND THEIR EVALUATION

USE PATTERN

Information on the registered uses of carbendazim in the United
Kingdom, France, Australia, China (Taiwan Province), New Zealand,
Colombia and Argentina (DuPont 1983a) and from 22 other countries
(Hoechst 1983) was made available to the Meeting. This is summarized
in Table 1.

Table 1 Use Pattern of Carbendazim in Certain Countries

Application Preharvest
Country interval
and Rate Number (days)
Crop (g a.i./ha or as
specified)

Argentina
Apple 15-30 g a.i./100 l 2 7
Pear 15-30 g a.i./100 l immerse or spray postharvest
Citrus 15-30 g a.i./100 l immerse or spray postharvest
Apple 9-15 g a.i./100 l 4 or more as needed 7
Pear 9-21 g a.i./100 l 4 or more as needed 7
Peach 15-17 g a.i./100 l at 10 to 20-day intervals 7
Peanut 73 g a.i./100 l at 15 to 20-day intervals 7
Beans 21 g a.i./100 l 2-3 7
Grape 15-30 g a.i./100 l 3-5 14
Tomato 15 g a.i./100 l at 15-day intervals 7
Celery 15-22 g a.i./100 l at 10 to 15-day intervals 7
Lettuce same as above same as above 7
Strawberry same as above 3 7

Australia
Peanuts¹ 100 14-day intervals 28
Pome fruit² 15-30 g/100 l 10 to 14-day intervals 7
Stone fruit³ 15-60 g/100 l up to 4 1
Roses

Table 1 (con't)

Application Preharvest
Country interval
and Rate Number (days)
Crop (g a.i./ha or as
specified)

China (Taiwan Prov.)
Tobacco 500 g/kg
Chrysanthemums

Colombia
Rice 200-300 not specified 10
Sorghum 200-300 not specified 10

France
Cereals 270 1 or more apply between stem
erection and first
node stage

New Zealand
Stone fruit 20-30 g/100 l 7 to 14-day intervals 1
Beans 1 125 2 at specified times 14
Field tomatoes 150 g/100 l 14-day intervals 3
Wheat 150-200 g a.i. 1 at specified times

United Kingdom
Winter wheat 333 1 apply between
leaf
Winter barley 333 1 sheaths erect
and first node
detectable
Barley 333 1 to 2 same as above
Winter rye 333 1 same as above

¹ South QLD only.
² All states.
³ QLD, NSW, VIC, SA and TAS only.

RESIDUES RESULTING FROM SUPERVISED TRIALS

Additional information and data were received from the
manufacturers, which are summarized in Table 2 (DuPont 1983b; BASF
1983).

Table 2 Carbendazim Residues in Crops

Application Interval Carbendazim residues
Country after last
and Rate (kg/ha) Frequency application Range No. of
crop (days) (mg/kg) samples

Belgium
Wheat
grain 0.28 - 0.48 3X 0 - 28 <0.05 30

Brazil
Beans
(carioca) 340 - 680 3X 31 <0.05 2
beans 340 - 680 1X 17 <0.05 2

Onions 0.16 - 0.32 2X 1 0.39 - 1.10 2
0.16 - 0.32 3X I 0.24 - 1.30 2
0.16 - 0.32 4X 1 0.13 - 0.71 2

Rice

grain 0.35 2X 38 0.12 1
grain 1.2 kg/ha 4X 36 <0.05 1
grain 1.7 4X 36 0.05 1
grain 0.8 2X 41 <0.05 1
grain 1.7 3X 36 0.07 1
grain 0.8 2X 41 <0.05 1
grain 1.7 2X 41 <0.05 1

Soybeans

beans 0.25 - 0.5 2X 19 - 35 <0.05 3

Table 2 (con't)

Application Interval Carbendazim residues
Country after last
and Rate (kg/ha) Frequency application Range No. of
crop (days) (mg/kg) samples

Wheat
grain 0.34 - 0.68 2 - 3X 14 - 38 <0.05 - 0.22 1

Costa Rica
Bananas

whole 0.25 - 0.5 6X 1 <0.05 - 0.08 8
pulp 0.25 - 0.5 6x 1 <0.05 8
whole 0.25 8X 1 <0.05 4
pulp 0.25 8X 1 <0.05 4
whole 0.25 - 0.5 10X 1 <0.05 2
pulp 0.25 - 0.5 10X 1 <0.05 2
whole 325 - 650 mg/ha 1X postharvest 1 0.15 - 2.4 18
pulp 325 - 650 mg/ha 1X 1 <0.05 - 0.11 18

Pineapple
flesh 0.015- 0.025 g/dip 1 dip 15 - 16 0.25 - 1.10 9
skin same as above 1 dip 15 - 16 8.3 - 60 9

Fed. Rep. Germany
Hops
dry 225 - 375 g/ha 2X 17 - 26 16 - 49 4
Potatoes
unwashed 30 mg/1 000 kg 1X 0,28,56,135 0.3 - 1.2 4
washed 30 mg/1 000 kg 1X 0,28,56,135 0.3- 0.8 4
peel 30 mg/1 000 kg 1X 0,28,56,135 0.3 - 1.6 4
peeled 30 mg/1 000 kg 1X 0,28,56,135 <0.2 - 0.3 4

Table 2 (con't)

Application Interval Carbendazim residues
Country after last
and Rate (kg/ha) Frequency application Range No. of
crop (days) (mg/kg) samples

South Africa
Pineapple
fruit 0.05 - 0.125% 1 dip 26 4.6 - 6.5 6
flesh 0.05 - 0.125% 1 dip 26 0.2- 0.5 6

United Kingdom

Swedes 125 g/ha 1X 52 <0.06 1

United States

Apples 0.1 - 0.25 11X 41 0.36 - 0.55 2
0.1 - 0.25 3x 81 <0.05 2
0.1- 0.2 0 - 17 0.35 - 1.4 6

Apricots 1 - 1.5 3 - 4X 0 - 10 <0.05 - 7.1 3

Beans 0.75 - 1.5 2x 8 - 25 1.4 - 42 12

beans 1.3 - 2.6 3X 10 - 21 0.11 - 0.79 6
foliage 1.3 - 2.6 3X 10 - 21 6.7 - 28 6
beams 0.75 - 2.0 1X 29 - 41 <0.05 6

Blueberries 1 - 2 6X 14 - 28 0.96 - 5.6 6

Celery 0.5 14X 7 - 14 0.76 - 1.6 3
2.0 10X 7 - 10 1.3 - 2.3 2

Table 2 (con't)

Application Interval Carbendazim residues
Country after last
and Rate (kg/ha) Frequency application Range No. of
crop (days) (mg/kg) samples

Cherries 0.1 8X 1 - 7 1.3 - 7.0 3
0.2 - 0.4 5X 1 - 7 0.32 - 0.66 6
1.5 3X 7 <0.05 1
1.5 3X 14 - 21 <0.05 - 0.46 2
0.4 - 0.5 5x 0 - 7 0.3 - 11.0 5

Cucumbers 0.5 10X 5 0.33 1
0.5 6X 0 - 2 0.08 - 0.26 3

Eggplant 0.5 - 1.0 4X 1 - 14 <0.05 - 0.44 6

Grapes 0.5 9X 8 - 20 5.0 - 6.5 6
1.33 5X 1 - 7 2.7 - 4.1 4
0.75- 1.0 4x 28 - 138 0.48 - 4.4 2
1 3X 5 - 7 1.7 - 2.3 2

Grapefruit 1 kg/ha + dip 4X 1 0.88 1
1.5 4X 1 0.80 1
3.0 4X 1 2.1 1
4.0 kg/ha + dip 4X I 3.5 1

Oats 0.5 - 1 4X 3 - 21 0.18 - 7.6 8

Oranges 1 - 2 1X foliar 0 - 7 0.06 - 7.3 4
1 lb/100 g 1X dip 0 1.8 1

Table 2 (con't)

Application Interval Carbendazim residues
Country after last
and Rate (kg/ha) Frequency application Range No. of
crop (days) (mg/kg) samples

Peaches 0.5 2X 0 - 7 1.5 - 2.6 3
1.5 4X 3 1.9 1
1.5 4X 0 1.7 1
0.2 - 0.4 10X 0 - 14 1.4 - 5.0 6
0.75 4X 1 - 14 0.36 - 3.1 6

Pears 1 - 1.5 6X 125 <0.05 - 0.08 3

Peanuts

nut 0.5 7X 7 <0.05 1
bulb 0.5 7X 7 0.44 1
hay 0.5 7x 7 3.3 1
whole nut 0.25 1X 30 <0.05 1
meat 0.25 1X <0.05 1

Peppers, bell 0.5 - 1.0 4X 1 - 14 0.18 - 2.2 6

Rice
grain 1.0 1X 49 <0.05 1
straw 1.0 1X 49 3.0 1
head 1.0 2X 26 1.4 1
head 1.0 1X 14 0.18 - 0.57 2
forage 1.0 1X 14 0.08 - 6.6 2
head 1.0 2X 18 0.24 1
forage 1.0 2X 18 11 1
head 2 kg/ha 2X 18 0.37 1
forage 2 kg/ha 2X 18 25 1

Table 2 (con't)

Application Interval Carbendazim residues
Country after last
and Rate (kg/ha) Frequency application Range No. of
crop (days) (mg/kg) samples

Soybeans
beans 0.25 - 1.0 2X 48 - 51 <0.05 - 0.11 3
hay 0.25 - 1.0 2X 48 - 51 <0.05 2

Strawberries 0.5 4X 1 0.54 - 0.75 2
0.5 - 1 5X 0 - 7 1.0 - 3.0 8

Sugarbeets
tops 0.2 - 0.4 2X 7 - 21 <0.05 - 8.3 15
root 0.2 - 0.4 2X 7 - 21 <0.05 - 8.3 15

Wheat

straw 0.5 1X 129 - 134 <0.05 4
grain 0.5 4X 129 - 134 <0.05 4
straw 0.1 - 1 2X 60 0.42 - 4.4 4
grain 0.1 - 1 2X 60 <0.05 4
straw 0.1 - 0.25 3X 45 <0.05 - 0.07 2
grain 0.1 - 0.25 3X 45 <0.05 2
straw (green) 0.25 - 1 3X 22 1.8 - 16 3
grain (immature) 0.25 - 1 3X 0.71 - 6.8 3

West Indies
Bananas
whole 0.15 4X 0 - 3 <0.05 - 0.07 6
whole 0.15 1X 0 - 5 <0.05 6

FATE OF RESIDUES

In Plants

Alfalfa, soybean, and ryegrass, which were grown in 1 cu ft
(1 cu ft = 0.0283 cu m) containers in a greenhouse in soil treated
with 80:20 mixtures of carbendazim (MBC) and 2-aminobenzimidazole
(2-AB), contained small but detectable residues of both compounds.
Both ¹⁴C-labelled and non-labelled mixtures were applied at the rate
of 2 kg/ha uniformly incorporated in the 0-10 cm layer of soil. In the
¹⁴C studies, alfalfa contained total ¹⁴C residues equivalent to
0.13 - 0.30 mg/kg of MBC/2-AB. Soybean plants contained 0.32 -
0.53 mg/kg and ryegrass (20-183 days) had 0.09 - 0.19 mg/kg. Each
plant contained approximately equal amounts of MBC, 2-AB and a polar
unknown fraction. Alfalfa from the non-labelled series contained 0.05
and 0.08 mg/kg, respectively, of MBC and 2-AB at the first cutting and
<0.05 mg/kg of either compound at the second and third cuttings.
Soybean plants contained <0.1 mg/kg of 2-AB and 0.59 mg/kg of MBC.
Ryegrass from six cuttings (20-149 days) contained 0.08 - 0.48 mg/kg
of MBC and <0.05 mg/kg of 2-AB. All data were on a fresh weight basis
(Rhodes et al. 1983).

Bean plants grown to maturity in Delaware (United States)
contained less than 0.1 mg/kg total ¹⁴C-residue in the edible beans
following two foliar applications of 1 kg a.i./ha of 2-¹⁴C-MBC at 25
percent and 50 percent bloom. Total ¹⁴C-residues in the bean foliage
decreased from about 5 mg/kg one week after the second spray to
0.2 mg/kg three weeks later. Of the total ¹⁴C in the edible beans and
foliage, 89-95 percent was intact free MBC and 2-8 percent was free
2-AB. An additional 1-3 percent of the ¹⁴C was found as ß-glycosidic
conjugates of MBC and 2-AB (Han 1983a).

Benomyl was sprayed on apple foliage at 1.68 kg/ha leaving
deposits ranging from 95-120 µg/g of leaf. Twelve days after
application at least 15 percent of the original deposit existed as
intact benomyl. The MBC concentration in leaves gradually increased,
remaining above 17 µg/g for approx. 80 days as a result of three
applications (of benomyl). The maximum concentration of 55.8 µg/g,
occurred eight days after the third (and last) application (Chiba &
Veres 1981).

In Soil

In greenhouse studies to determine run-off and leaching of
2-¹⁴C-MBC on soil, a container of Keyport silt loam was treated
(spray) with labelled MBC, at a rate of 10 kg/ha, by spraying the
upper one-third (0.093 sq m) of the plot and allowed to stand 24h.
Artificial rain was then applied at 3.75 cm the first day after
treatment and 2.5 cm on the third and seventh days. All water that ran
off or leached through the soil was collected and analysed for total

¹⁴C by counting 2 ml aliquots in a liquid scintillation counter. Soil
in the plot was divided into increments for analysis, air dried and
1 g aliquots analysed for total ¹⁴C. After each of the three rain
applications, 0.05 - 0.39 percent of the applied ¹⁴C was found in
run-off water; <0.01 percent was found in the leach water after two
rains and 0.19 percent after the third one. Soil analyses showed that
90.6 percent of the applied activity remained in the treated area and
93.1 percent in the first 10 cm of soil (Rhodes & Long 1983).

Degradation studies of labelled (2-¹⁴C) 2-amino benzimidazole,
the primary degradation product of carbendazim, in soil showed that
¹⁴C evolution increased exponentially from 1 to 22°C, reached a
maximum at 22°, remained almost constant up to 35°, then became almost
nil at 40°, when the soil water content was 100 percent of field
capacity. At 25°C evolution increased exponentially from 28 to 94
percent of field capacity of water; evolution decreased slightly at
about this temperature. These and other results indicate the presence
of organisms that are able to decompose 2-AB (Helweg 1979).

Laboratory studies in two soil types under anaerobic conditions
using 2-¹⁴C-labelled carbendazim showed only a small amount of 2-AB
(<2 percent) and no other postulated degradation products (<0.05
percent). Reincorporation of ¹⁴C into soil humus was indicated by
fractionation studies, which showed that the unextracted ¹⁴C-residue
was widely distributed in various organic soil components (Han 1983b).

Laboratory experiments with flooded soils treated at various
levels (0-1 000 ppm) with benomyl, MBC and 2-AB showed that benomyl
exerts a strongly inhibitory effect on nitrofication at 1 000 ppm, as
measured by nitrate production. No inhibition of nitrification by MBC
occurred at 10 or 100 ppm but slight retardation was noted at
1 000 ppm. Substantial inhibition occurred for 2-AB at the 100 and
1 000 ppm levels. Similar results were obtained in studies involving
pure cultures of nitrifying bacteria (Ramakrishna et al. 1979).

METHODS OF RESIDUE ANALYSIS

The majority of the data presented in Table 2 were obtained using
the method of Pease and Holt (1971), which is still suitable for
regulatory purposes.

Since the last evaluation of carbendazim in 1978, a simple high
performance liquid chromatographic (HPLC) method has been developed
to determine individually residues of benomyl and MBC on apple
leaves without clean-up. In this procedure, sample leaves are
freeze-dried and tumble-extracted with CHCl₃ containing 5 mg/ml
of n-propyl isocyanate at 1°C. The latter converts the MBC into
1-(n-propylcarbamoyl)-2-benzimidozole carbamate, which is more easily
extracted and allows its distinction from intact benomyl residues

during HPLC using a UV detector set at 280 nm. Recoveries of both
compounds ranged from 78 to 86 percent and a detection level of
0.2 mg/kg was achieved (Chiba & Veres 1980).

APPRAISAL

New information was received on use patterns and registered uses
for carbendazim in the United Kingdom, France, Australia, China
(Taiwan province), New Zealand, Colombia, Argentina and 22 other
countries. Some additional information was available on the fate of
residues in plants and soil.

Systemic uptake of labelled carbendazim from treated soil
(2 kg/ha) was limited, reaching maximum levels of total ¹⁴C of
0.3 mg/kg in alfalfa, 0.53 mg/kg in soybean plants and 0.19 mg/kg in
ryegrass. Each plant contained approximately equal amounts of
carbendazim, 2-aminobenzimidazole (2-AB) and a polar unknown fraction.

Soil mobility of carbendazim is limited, with a maximum of 0.4
percent of applied ¹⁴C being found in run-off water from artificial
rain and 0.2 percent in leach water. The soil contained 91 percent of
applied activity in the treated area and 93 percent in the first
10 cm.

Degradation of carbendazim in soil under anaerobic conditions
resulted in only a small amount (<2 percent of the applied dose) of
2-AB and no other products. The carbendazim was apparently mostly
incorporated into soil humus. In contrast to benomyl, which is
strongly inhibitory to soil nitrification at high doses (1 000 ppm),
carbendazim showed no effects at low levels and only slight inhibition
at 1 000 ppm. Substantial inhibition was shown by 2-AB at medium
(100 ppm) to high (1 000 ppm) levels.

New or additional data on residue levels in various crops
resulting from supervised trials with carbendazim were received from
the Federal Republic of Germany and the United States and from trials
with benomyl from those countries as well as Australia, Japan, Kenya
and the United Kingdom. The data supported previously recommended
levels except for bean fodder (formerly bean vines), which should be
increased from 30 to 50 mg/kg; apricots, grapes and sugarbeet tops
from 5 to 10 mg/kg; rice straw from 2 to 15 mg/kg; peanut hay from 2
to 5 mg/kg; and peanut hulls from 0.2 to 1 mg/kg. Additional
commodities for which recommendations can be made include dry hops
(50 mg/kg), peppers and blueberries (5 mg/kg), onions (2 mg/kg),
eggplant and oats (0.5 mg/kg), chestnuts and soybeans (0.2 mg/kg) and
asparagus, rutabagas and soybean hay (0.1 mg/kg).

The improved residue analytical method of Pease and Holt is still
recommended for regulatory use. A simple HPLC method, which
individually determines residues of benomyl and carbendazim on apple
leaves without clean-up, has been published that appears to be useful
for research purposes.

RECOMMENDATIONS

The guideline levels listed in 1978 are now converted to maximum
residue limits on the basis of the establishment of an ADI for
carbendazim.

The following maximum residue levels, amending or additional to
those of 1978, are proposed. Those levels which are based on benomyl
trials (see benomyl) are indicated with a ¹ beside the commodity and
were derived by dividing benomyl tabular data by the conversion factor
of 1.52.

Commodity Maximum residue Preharvest intervals
limits on which levels are
(mg/kg) based (days)

Bean fodder 50 (increased from 30) 8
*Hops, dry 50 17
Rice straw 15 (increased from 2) 18
Apricots 10 (increased from 5) 0
Grapes 10 (increased from 5) 8
Sugarbeet tops 10 (increased from 5) 7
*Pineapple¹ 20 Postharvest dip
Peanut hay 5 (increased from 2) 7
Wheat straw¹ 5 (increased from 2) -
*Blueberries 5 14
*Peppers 5 7
Melons1 2 (increased from 0.5) Postharvest dip
*Onions 2 1
*Sweet potatoes¹ 1 90
Peanut hulls¹ 1 (increased from 0.2)
*Eggplant 0.5 1
*Oats 0.5 21
*Soybeans 0.2 48
*Chestnuts¹ 0.2 1
*Soybean hay 0.1** 48
*Asparagus¹ 0.1** 260
*Taro¹ 0.1** 162
*Rutabagas 0.1** 52

* New NRLs.
** At or about the limit of determination.
¹ Based on benomyl data.

REFERENCES- RESIDUES

BASF. Residue data on pineapples, hops and swedes. (Unpublished)
1983

Chiba, M. & Veres, D.F. High performance liquid chromatographic method
1980 for simultaneous determination of residual benomyl and
methyl 2-benzimidazole carbamate on apple foliage without
cleanup. J. Assoc. Off. Anal. Chem. 63(6): 1291-1295.

Chiba, M. & Veres, D.F. Fate of benomyl and its degradation compound
1981 methyl 2-benzimidazolecarbamate on apple foliage. J. Agric.
Food Chem., 29: 588-590.

DuPont. Samples of registered labels for carbendazim in the United
1983a Kingdom, France, Australia, Taiwan, New Zealand, Colombia
and Argentina.

DuPont. Analytical reports on residue data in various crops.
1983b (Unpublished)

Han, J. C-Y. Characterization of residues in bean plants following
1983a foliar spray application with MBC. DuPont Report.
(Unpublished)

Han, J. C-Y. Anaerobic soil metabolism of 2-¹⁴C-benomyl and methyl
1983 2-¹⁴C-benzimidazolecarbamate. Du Pont report. (Unpublished)

Helweg, A. Influence of temperature, humidity, and inoculation on the
1979 degradation of ¹⁴C-labeled 2-aminobenzimidazole in soil.
Water, Air, Soil Pollut., 12: 275-281.

Hoechst. Computer printout of registered or permitted uses of
1983 carbendazim in 29 countries submitted to the Meeting.

Pease, H.L. & Holt, R.F. Improved method for determining benomyl
1971 residues. J. Assoc. Off. Anal. Chem., 54(6): 1399-1402.

Ramakrishna, C., Gowda, T.K.S. & Sethunathan, N. Effect of benomyl and
1979 its hydrolysis products, MBC and AB, on nitrofication in a
flooded soil. Bull. Environ. Contam. Toxicol., 21: 328-333.

Rhodes, R.C. & Long, J.D. Run-off and leaching studies with methyl-2
1983 ¹⁴C-benzimidazole-carbamate on soil. DuPont report.
(Unpublished)

Rhodes, R.C., Pease, A.L. & Holt, R.F. Greenhouse studies on crop
1983 uptake of MBC and 2-AB from soil. DuPont report.
(Unpublished)

    See Also:
       Toxicological Abbreviations
       Carbendazim (EHC 149, 1993)
       Carbendazim (HSG 82, 1993)
       Carbendazim (ICSC)
       Carbendazim (WHO Pesticide Residues Series 3)
       Carbendazim (Pesticide residues in food: 1976 evaluations)
       Carbendazim (Pesticide residues in food: 1977 evaluations)
       Carbendazim (Pesticide residues in food: 1978 evaluations)
       Carbendazim (Pesticide residues in food: 1985 evaluations Part II Toxicology)
       Carbendazim (Pesticide residues in food: 1995 evaluations Part II Toxicological & Environmental)
       Carbendazim (Pesticide residues in food: 1995 evaluations Part II Toxicological & Environmental)
       Carbendazim (JMPR Evaluations 2005 Part II Toxicological)