INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY
ENVIRONMENTAL HEALTH CRITERIA 148
BENOMYL
This report contains the collective views of an international group of
experts and does not necessarily represent the decisions or the stated
policy of the United Nations Environment Programme, the International
Labour Organisation, or the World Health Organization.
Published under the joint sponsorship of
the United Nations Environment Programme,
the International Labour Organisation,
and the World Health Organization
First draft prepared by Dr L.W. Hershberger and
Dr G.T. Arce, E.I. Du Pont de Nemours and
Company, Wilmington, Delaware, USA
World Health Orgnization
Geneva, 1993
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WHO Library Cataloguing in Publication Data
Benomyl.
(Environmental health criteria ; 148)
1.Benomyl - adverse effects 2.Benomyl - toxicity
3.Environmental exposure I.Series
ISBN 92 4 157148 9 (LC Classification: SB 951.3)
ISSN 0250-863X
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CONTENTS
ENVIRONMENTAL HEALTH CRITERIA FOR BENOMYL
1. SUMMARY AND CONCLUSIONS
1.1. Summary
1.1.1. Identity, physical and chemical properties, and
analytical methods
1.1.2. Sources of human and environmental exposure
1.1.3. Environmental transport, distribution and
transformation
1.1.4. Environmental levels and human exposure
1.1.5. Kinetics and metabolism
1.1.6. Effects on laboratory mammals; in vitro test
systems
1.1.7. Effects on humans
1.1.8. Effects on other organisms in the laboratory and
field
1.2. Conclusions
2. IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, AND ANALYTICAL
METHODS
2.1. Chemical identity
2.1.1. Primary constituent
2.1.2. Technical product
2.2. Physical and chemical properties
2.3. Analytical methods
3. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE
3.1. Natural occurrence
3.2. Anthropogenic sources
3.2.1. Uses
3.2.2. Worldwide sales
4. ENVIRONMENTAL TRANSPORT, DISTRIBUTION AND TRANSFORMATION
4.1. Transport and distribution between media
4.1.1. Air
4.1.2. Water
4.1.3. Soil
4.1.4. Leaching
4.1.5. Crop uptake
4.2. Transformation
4.2.1. Biodegradation
4.2.1.1 Water
4.2.1.2 Soil
4.2.1.3 Crops
4.2.2. Abiotic degradation
4.2.3. Bioaccumulation
5. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE
5.1. Environmental levels
5.1.1. Air, water and soil
5.1.2. Food and feed
5.1.3. Terrestrial and aquatic organisms
5.2. General population exposure
5.2.1. USA
5.2.2. Sweden
5.2.3. Maximum residue limits
5.3. Occupational exposure during manufacture, formulation or
use
5.3.1. Use
6. KINETICS AND METABOLISM
6.1. Absorption
6.2. Distribution and accumulation
6.3. Metabolic transformation
6.4. Elimination and excretion
6.5. Reaction with body components
7. EFFECTS ON LABORATORY MAMMALS; IN VITRO TEST SYSTEMS
7.1. Single exposure
7.2. Short-term exposure
7.2.1. Gavage
7.2.2. Feeding
7.2.2.1 Rat
7.2.2.2 Dog
7.2.3. Dermal
7.2.4. Inhalation
7.3. Skin and eye irritation; sensitization
7.3.1. Dermal
7.3.2. Eye
7.3.3. Sensitization
7.4. Long-term exposure
7.4.1. Rat
7.4.2. Mouse
7.5. Reproduction, embryotoxicity and teratogenicity
7.5.1. Reproduction
7.5.1.1 Rat feeding studies
7.5.1.2 Rat gavage studies
7.5.1.3 Dog inhalation studies
7.5.2. Teratogenicity and embryotoxicity
7.5.2.1 Mouse gavage studies
7.5.2.2 Rat gavage studies
7.5.2.3 Rat feeding studies
7.5.2.4 Rabbit feeding studies
7.6. Mutagenicity and related end-points
7.7. Carcinogenicity
7.7.1. Rat
7.7.2. Mouse
7.8. Special studies
7.8.1. Neurotoxicity
7.8.2. Effects in tissue culture
7.9. Factors modifying toxicity; toxicity of metabolites
7.10. Mechanisms of toxicity - mode of action
8. EFFECTS ON HUMANS
8.1. General population exposure
8.2. Occupational exposure
8.2.1. Acute toxicity
8.2.2. Effects of short- and long-term exposure
9. EFFECTS ON ORGANISMS IN THE LABORATORY AND FIELD
9.1. Microorganisms
9.2. Aquatic organisms
9.3. Terrestrial organisms
9.4. Population and ecosystem effects
10. EVALUATION OF HUMAN HEALTH RISKS AND EFFECTS ON THE ENVIRONMENT
10.1. Evaluation of human health risks
10.2. Evaluation of effects on the environment
10.3. Conclusions
11. FURTHER RESEARCH
12. PREVIOUS EVALUATIONS BY INTERNATIONAL BODIES
REFERENCES
RESUME ET CONCLUSIONS
RESUMEN Y CONCLUSIONES
WHO TASK GROUP ON ENVIRONMENTAL HEALTH CRITERIA FOR BENOMYL AND
CARBENDAZIM
Members
Dr G. Burin, Office of Pesticide Programmes, US Environmental
Protection Agency, Washington, D.C., USA
Dr R. Cooper, Reproductive Toxicology Branch, US Environmental
Protection Agency, Research Triangle Park, North Carolina, USA
Dr I. Desi, Department of Public Health, Albert Szent-Györgyi
University Medical School, Szeged, Hungary
Dr S. Dobson, Institute of Terrestrial Ecology, Monks Wood, Abbots
Ripton, Huntingdon, United Kingdom
Dr A. Helweg, Department for Pesticide Analysis and Ecotoxicology,
Danish Research Service for Plant and Soil Science, Flakkebjerg,
Slagelse, Denmark
Dr M. Lotti, Institute of Occupational Medicine, University of
Padua, Padua, Italy ( Chairman)
Dr K. Maita, Toxicology Division, Institute of Environmental
Toxicology, Kodaira-Shi, Tokyo, Japan
Dr F. Matsumura, Department of Environmental Toxicology, Institute
of Toxicology and Environmental Health, University of California,
Davis, California, USA
Dr T.K. Pandita, Microbiology and Cell Biology Laboratory, Indian
Institute of Science, Bangalore, Indiaa
Dr C. Sonich-Mullin, Environmental Criteria and Assessment Office,
US Environmental Protection Agency, Cincinnati, Ohio, USA
Dr P.P. Yao, Institute of Occupational Medicine, Chinese Academy of
Preventive Medicine, Beijing, China
a Invited but unable to attend the meeting
Secretariat
Dr B.H. Chen, International Programme on Chemical Safety, World
Health Organization, Geneva, Switzerland ( Secretary)
Dr L.W. Hershberger, Dupont Agricultural Products, Walker's Mill,
Barley Mill Plaza, Wilmington, Delaware, USA ( Rapporteur)
Mr P. Howe, Institute of Terrestrial Ecology, Monks Wood, Abbots
Ripton, Huntington, United Kingdom
NOTE TO READERS OF THE CRITERIA MONOGRAPHS
Every effort has been made to present information in the
criteria monographs as accurately as possible without unduly
delaying their publication. In the interest of all users of the
Environmental Health Criteria monographs, readers are kindly
requested to communicate any errors that may have occurred to the
Director of the International Programme on Chemical Safety, World
Health Organization, Geneva, Switzerland, in order that they may be
included in corrigenda.
* * *
A detailed data profile and a legal file can be obtained from
the International Register of Potentially Toxic Chemicals, Palais
des Nations, 1211 Geneva 10, Switzerland (Telephone No. 7988400 or
7985850).
ENVIRONMENTAL HEALTH CRITERIA FOR BENOMYL
A WHO Task Group on Environmental Health Criteria for Benomyl
and Carbendazim, sponsored by the US Environmental Protection
Agency, met in Cincinnati, USA, from 14 to 19 September 1992. On
behalf of the host agency, Dr T. Harvey opened the meeting and
welcomed the participants. Dr B.H. Chen of the International
Programme on Chemical Safety (IPCS) welcomed the participants on
behalf of the Director, IPCS, and the three IPCS cooperating
organizations (UNEP/ILO/WHO). The Task Group reviewed and revised
the draft criteria monograph and made an evaluation of the risks for
human health and the environment from exposure to benomyl.
The first draft of this monograph was prepared by Dr L.W.
Hershberger and Dr G.T. Arce of E.I. Du Pont de Nemours and Company,
Wilmington, Delaware, USA. The second draft was prepared by Dr L.W.
Hershberger incorporating comments received following the
circulation of the first draft to the IPCS Contact Points for
Environmental Health Criteria monographs. Dr M. Lotti (Institute of
Occupational Medicine, University of Padua, Italy) made a
considerable contribution to the preparation of the final text. Dr
B.H. Chen and Dr P.G. Jenkins, both members of the IPCS Central
Unit, were responsible for the overall scientific content and
technical editing, respectively.
The efforts of all who helped in the preparation and
finalization of the monograph are gratefully acknowledged.
Financial support for the meeting was provided by the US
Environmental Protection Agency, Cincinnati, USA.
ABBREVIATIONS
ADI acceptable daily intake
a.i. active ingredient
BIC butyl isocyanate
BUB 2-(3-butylureido)benzimidazole
EEC European Economic Community
HPLC high performance liquid chromatography
Koc Distribution coefficient between pesticide adsorbed to soil
organic carbon and pesticide in solution
Kom Distribution coefficient between pesticide adsorbed to soil
organic matter and pesticide in solution
MRL maximum residue limits
NOEL no-observed-effect level
OECD Organisation for Economic Co-operation and Development
STB 3-butyl-1,3,5-triazino[1,2a]-benzimidazol-2,4(1H,3H)dione
2-AB 2-aminobenzimidazole
5-HBC methyl (5-hydroxy-1H-benzimidazol-2-yl)-carbamate
1. SUMMARY AND CONCLUSIONS
1.1 Summary
1.1.1 Identity, physical and chemical properties, and analytical
methods
Benomyl, a tan crystalline solid, is a systemic fungicide
belonging to the benzimidazole family. It decomposes just above its
melting point of 140 °C and has a vapour pressure of < 5 x 10-6
Pa (< 3.7 x 10-8 mmHg) at 25 °C. Benomyl is virtually insoluble
in water at pH 5 and 25 °C, the solubility being 3.6 mg/litre. It is
stable under normal storage conditions but decomposes to carbendazim
in water.
Residual and environmental analyses are performed by extraction
with an organic solvent, purification of the extract by a
liquid-liquid partitioning procedure, and conversion of the residue
to carbendazim. Measurement of residues may be determined by high
performance liquid chromatography or immunoassay.
1.1.2 Sources of human and environmental exposure
In 1988, the estimated worldwide use of benomyl was
approximately 1700 tonnes. It is a widely used fungicide registered
for use on over 70 crops in 50 countries. Benomyl is formulated as a
wettable powder.
1.1.3 Environmental transport, distribution and transformation
Benomyl is rapidly converted to carbendazim in the environment
with half-lives of 2 and 19 h in water and in soil, respectively.
Data from studies on both benomyl and carbendazim are therefore
relevant for the evaluation of environmental effects.
Carbendazim decomposes in the environment with half-lives of 6
to 12 months on bare soil, 3 to 6 months on turf, and 2 and 25
months in water under aerobic and anaerobic conditions,
respectively.
Carbendazim is mainly decomposed by microorganisms.
2-Aminobenzimidazole (2-AB) is a major degradation product and is
further decomposed by microbial activity.
When phenyl-14C-labelled benomyl was decomposed, only 9% of
the 14C was evolved as CO2 during 1 year of incubation. The
remaining 14C was recovered mainly as carbendazim and bound
residues. The fate of a possible degradation product
(1,2-diaminobenzene) may further clarify the degradation pathway of
benzimidazole fungicides in the environment.
Field and column studies have shown that carbendazim remains in
the soil surface layer. There is no available determination of
carbendazim adsorption in soil, but it is expected to be as strongly
adsorbed to soil as benomyl, with Koc values ranging from 1000 to
3600. The log Kow values for benomyl and carbendazim are 1.36 and
1.49, respectively.
No risk of leaching was apparent when this was evaluated in a
screening model based on adsorption and persistence data. This
statement is supported by analyses of well-water in the USA where
benomyl was not found in any of 495 wells and carbendazim not in any
of 212 wells (limit of detection not available). Surface run-off of
benomyl and carbendazim is expected to consist only of fungicide
adsorbed to soil particles, and these compounds are expected to be
strongly adsorbed to sediments in the aqueous environment.
Benomyl in solutions, plants and soil degrades to carbendazim
(methyl-1H-benzimidazol-2-carbamate) and to 2-AB, STB
(3-butyl-1,3,5-triazino[1,2a]-benzimidazol-2,4(1H,3H)dione) and BBU
(1-(2-benzimidazolyl)-3- n-butylurea). There is little or no
photolysis of benomyl.
In animal systems, benomyl is metabolized to carbendazim and
other polar metabolites, which are rapidly excreted. Neither benomyl
nor carbendazim has been observed to accumulate in any biological
system.
1.1.4 Environmental levels and human exposure
No environmental monitoring data for benomyl appear to be
available. However, the following can be summarized from
environmental fate studies.
Since benomyl and carbendazim remain stable for several weeks
on plant material, they may become accessible to organisms feeding
on leaf litter. Soil and sediments may contain residues of
carbendazim for up to 3 years. However, the strong adsorption of
carbendazim to soil and sediment particles reduces the exposure of
terrestrial and aquatic organisms.
The main source of exposure for the general human population is
residues of benomyl and carbendazim in food crops. Dietary exposure
analysis in the USA (combined benomyl and carbendazim) and the
Netherlands (carbendazim) yielded an expected mean intake of about
one-tenth of the recommended Acceptable Daily Intake (ADI) for
benomyl of 0.02 mg/kg body weight and for carbendazim of 0.01 mg/kg
body weight.
Occupational exposures during the manufacturing process are
below Threshold Limit Values. Agricultural workers engaged in
pesticide mixing and loading or re-entering benomyl-treated fields
are expected to be exposed dermally to a few mg of benomyl per hour.
This type of exposure could be reduced by the use of protective
devices. Furthermore, since dermal absorption is expected to be low,
the probability of benomyl having systemic toxic effects on human
populations through this route is very low.
1.1.5 Kinetics and metabolism
Benomyl is readily absorbed in animal experiments after oral
and inhalation exposure, but much less so following dermal exposure.
Absorbed benomyl is rapidly metabolized and excreted in the urine
and faeces. In rats fed 14C-labelled benomyl, its metabolites
carbendazim and methyl(5-hydroxy-1H-benzimidazol-2-yl)-carbamate
(5-HBC) were found in the blood and in small amounts in the testes,
kidneys and livers. The tissue distribution showed no
bioconcentration. In urine the primary metabolite was 5-HBC, some
carbendazim also being present. By 72 h after administration, 98% of
the given amount had been excreted. In cows dosed by capsule for 5
days with radiolabelled benomyl at a dose equivalent to 50 mg/kg
diet, there was a benomyl equivalent level of 4 mg/kg in the liver,
0.25 mg/kg in the kidney and no significant levels in other tissues
or fat. During feeding, 65% of the radiolabel was excreted in the
urine, 21% in the faeces and 0.4% in the milk. The major metabolite
in the milk was 5-HBC. Similar metabolism and elimination patterns
were found in other animals.
Benomyl does not inhibit acetyl cholinesterase in vitro. It
has been shown to induce liver epoxyhydrolase, gamma-glutamyl
transpeptidase and glutathione- S-transferase in in vivo studies
on mice and rats.
1.1.6 Effects on laboratory mammals; in vitro test systems
1.1.6.1 Single exposure
Benomyl has low acute toxicity with an oral LD50 in the rat
of > 10 000 mg/kg and an inhalation 4-h LC50 of > 4 mg/litre.
Carbendazim, like its parent compound benomyl, has an LD50 in rats
of > 10 000 mg/kg. Dogs, exposed via inhalation for 4 h at 1.65
mg/litre and examined 28 days after exposure, showed decreased liver
weight. A single dose of benomyl to rats by gavage showed
reproductive effects at 70 days after exposure (see section
1.1.6.5).
1.1.6.2 Short-term exposure
Short-term gavage, dietary or dermal administration of benomyl
for up to 90 days slightly increased liver weights in the rat (125
mg/kg per day, dietary) and produced effects on male reproductive
organs (decreased testis and epididymal weights, decreased sperm
production) in the rat (45 mg/kg per day, gavage; no-observed-effect
level (NOEL) = 15 mg/kg), rabbit (1000 mg/kg per day, oral; 500
mg/kg body weight per day, dermal) and beagle dog (62.5 mg/kg; NOEL
= 18.4 mg/kg per day, dietary). Liver and testicular effects were
not observed in rats exposed via inhalation to benomyl
concentrations of up to 200 mg/m3 for 90 days.
1.1.6.3 Skin and eye irritation and sensitization
Application to the skin of the rabbit and guinea-pig produced
either mild or no irritation and moderate skin sensitization.
Application to the eyes of rats produced temporary mild conjunctival
irritation.
1.1.6.4 Long-term exposure
A long-term feeding study in rats did not demonstrate any
compound-related effects at dose levels up to and including 2500
mg/kg diet (125 mg/kg body weight per day). This study was not
considered adequate to evaluate reproductive effects. In the CD-1
mouse, liver weights were increased at dose levels of 1500 mg/kg
diet or more. Male mice had decreased absolute testes weights and
thymic atrophy at a level of 5000 mg/kg diet.
1.1.6.5 Reproduction, embryotoxicity, and teratogenicity
Benomyl causes a decrease in testis and epididymis weight, a
reduction in caudal sperm reserves, a decrease in sperm production,
and a lowering of male fertility rates. At higher doses, there is
hypospermatogenesis with generalized disruption of all stages of
spermatogenesis. Benomyl does not effect copulatory behaviour,
seminal vesicles, sperm mobility or related reproductive hormones.
The lowest benomyl concentration shown to induce a statistically
significant spermatogenic effect in male rats was 45 mg/kg per day.
The NOEL for these effects was 15 mg/kg per day.
A single dose of benomyl (100 mg/kg or more) administered to
rats by gavage showed effects, at 70 days aftr exposure, which
included decreased testis weight and seminiferous tubular atrophy.
When administered via gavage from days 7 to 16 of gestation to
ChD-CD rats and Wistar rats, benomyl was found to be teratogenic at
62.5 mg/kg for both strains, but not at 30 mg/kg for ChR-CD rats and
not at 31.2 mg/kg for Wistar rats. When Sprague-Dawley rats were
administered by gavage on days 7 to 21 of gestation, benomyl was
found to be teratogenic at 31.2 mg/kg. The effects were
microphthalmia, hydrocephaly, and encephaloceles. Postnatal
development of rats was adversely affected at dose levels greater
than 15.6 mg/kg.
In mice, gavage dosing at a concentration of 50 mg/kg or more
induced supernumery ribs and other skeletal and visceral anomalies.
A NOEL was not established in the mouse because no doses lower than
50 mg/kg were tested. Except for a marginal increase in supernumery
ribs in rabbits, no teratogenic effects were observed at dose levels
as high as 500 mg/kg diet.
1.1.6.6 Mutagenicity and related end-points
Studies in somatic and germ cells show that benomyl does not
cause gene mutations or structural chromosomal damage (aberrations)
and it does not interact directly with DNA (causing DNA damage and
repair). This has been demonstrated in both mammalian and
non-mammalian systems.
Benomyl does, however, cause numerical chromosome aberrations
(aneuploidy and/or polyploidy) in experimental systems in vitro
and in vivo.
1.1.6.7 Carcinogenicity
Benomyl or carbendazim caused liver tumours in two strains of
mice (CD-1 and Swiss (SPF)) that have a high spontaneous rate of
liver tumours. In contrast, carbendazim was not carcinogenic in
NMRKf mice, which have a low spontaneous rate of such tumours.
The first carcinogenicity study using CD-1 mice showed a
statistically significant dose-related increase of hepatocellular
neoplasia in females, and a statistically significant response was
also observed in the mid-dose (1500 mg/kg) males but not in the
high-dose males because of the high mortality rate. A second
carcinogenicity study of carbendazim in a genetically related mouse
strain, SPF mice (Swiss random strain), at doses of 0, 150, 300 and
1000 mg/kg (increased to 5000 mg/kg during the study) showed an
increase in the incidence of combined hepatocellular adenomas and
carcinomas. A third study carried out in NMRKf mice at doses of 0,
50, 150, 300 and 1000 mg/kg (increased to 5000 mg/kg during the
study) showed no carcinogenic effects.
Carcinogenicity studies with both benomyl and carbendazim were
negative in rats.
1.1.6.8 Mechanism of toxicity - mode of action
The biological effects of benomyl and carbendazim are thought
to be the result of their interaction with cell microtubules. These
structures are involved in vital functions such as cell division,
which is inhibited by benomyl and carbendazim. Benomyl and
carbendazim toxicity in mammals is linked with microtubular
dysfunction.
Benomyl and carbendazim, like other benzimidazole compounds,
display selective toxicity for species. This selectivity is, at
least in part, explained by the different binding of benomyl and
carbendazim to tubulins of target and non-target species.
1.1.7 Effects on humans
Benomyl causes contact dermatitis and dermal sensitization. No
other effects have been reported.
1.1.8 Effects on other organisms in the laboratory and field
Benomyl has little effect on soil microbial activity at
recommended application rates. Some adverse effects have been
reported for groups of fungi.
The 72-h EC50, based on total growth, for the green alga
Selenastrum capricornutum was calculated to be 2.0 mg/litre; the
no-observed-effect concentration (NOEC) was 0.5 mg/litre. The
toxicity of benomyl to aquatic invertebrates and fish varies widely,
96-h LC50 values ranging from 0.006 mg/litre for the channel
catfish (yolk-sac fry) to > 100 mg/litre for the crayfish.
Benomyl is toxic to earthworms in laboratory experiments at
realistic exposure concentrations and as a result of recommended
usage in the field. It is of low toxicity to birds and its
degradation product carbendazim is "relatively non-toxic" to
honey-bees.
1.2 Conclusions
Benomyl causes dermal sensitization in humans. Both benomyl and
carbendazim represent a very low risk for acute poisoning in humans.
Given the current exposures and the low rate of dermal absorption of
these two compounds, it is unlikely that they would cause systemic
toxicity effects either in the general population or in
occupationally exposed subjects. These conclusions are drawn from
animal data and the limited human data available, and are supported
by the understanding of the mode of action of carbendazim and
benomyl in both target and non-target species.
Further elucidation of the mechanism of toxicity of benomyl and
carbendazim in mammals will perhaps permit a better definition of
no-observed-effect levels. Binding studies on tubulins of target
cells (testis and embryonic tissues) will facilitate inter-species
comparisons.
Carbendazim is strongly adsorbed to soil organic matter and
remains in the soil for up to 3 years. Carbendazim persists on leaf
surfaces and, therefore, in leaf litter. Earthworms have been shown
to be adversely affected (population and reproductive effects) at
recommended application rates. There is no information on other soil
or litter arthropods that would be similarly exposed.
The high toxicity to aquatic organisms in laboratory tests is
unlikely to be seen in the field because of the low bioavailability
of sediment-bound residues of carbendazim. However, no information
is available on sediment-living species, which would receive the
highest exposure.
2. IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, AND ANALYTICAL
METHODS
2.1 Chemical identity
2.1.1 Primary constituent
Chemical structure:
Molecular formula: C14H18N4O3
Common name: Benomyl
CAS chemical name: Carbamic acid, [1-(butylamino)carbonyl]-1H-
benzimidazol-2-yl]-, methyl ester
IUPAC chemical name: Methyl 1-[(butylamino)carbonyl]-1H-
benzimidazol-2-ylcarbamate
CAS registry number: 17804-35-2
Relative molecular mass: 290.3
Synonym: Methyl 1-(butylcarbamoyl)-2-benzimida-
zolecarbamate
2.1.2 Technical product
Major trade names: Benlate, Tersan, Fungicide 1991, Fundazol
Purity: > 95% (FAO specifications)
2.2 Physical and chemical properties
Table 1. Some physical and chemical properties of Benomyl
Physical state Crystalline solid
Colour Tan
Odour Negligible
Melting point/boiling point/ Decomposes just after
flash point melting at 140 °C
Explosion limits LEL = 0.05 g/litre in air
Vapour pressure < 5.0 x 10-6 Pa (< 3.7 x
10-8 mmHg) at 25 °Ca
Density 0.38 g/cm3
Log n-octanol/water partition
coefficient 1.36
Solubility in water 3.6 mg/litre (at pH 5 and 25
°C)
Solubility in organic solvents Chloroform 9.4
(g/100 g solvent at 25 °C) Dimethylformamide 5.3
Acetone 1.8
Xylene 1.0
Ethanol 0.4
Heptane 40
Henry's constant < 4.2 x 10-9 atm-m3/mol
at pH 5 and 25 °C
Soil/water partition coefficient 1090 mg/g (Kom); 1860 mg/g
(Koc)b
a Barefoot (1988)
b Koc = Distribution coefficient between pesticide adsorbed
to soil organic carbon and pesticide in solution.
Kom = Distribution coefficient between pesticide adsorbed to
soil organic matter and pesticide in solution.
2.3 Analytical methods
Most methods for determining benomyl and its by-product
residues in plant and animal tissue and in soil involve isolation of
the residue by extraction with an organic solvent, purification of
the extract by a liquid-liquid partitioning procedure, and
conversion of the residue to carbendazim. Residues may be measured
by procedures using high-speed cation-exchange liquid
chromatography, reversed phase HPLC, and immunoassay. One method for
analysis of water samples can distinguish between benomyl and
carbendazim. Recoveries of benomyl, carbendazim and 2-AB
(2-aminobenzimidazole) from various types of soils average 92, 88
and 71%, respectively. The lower limit of sensitivity of the method
is 0.05 ppm for each of these components. The recoveries and
sensitivities for plant tissues are similar. Table 2 outlines
various analytical methods for soil, water, plant and animal tissue.
Table 2. Analytical Methods for Benomyl
Analytical method Medium Detection limit Comments Reference
Strong cation exchange/HPLC soil 0.05 mg/kg acidic methanol extraction converts Kirkland et al. (1973)
residual benomyl to carbendazim
Strong cation exchange/HPLC plant 0.05 mg/kg acidic methanol extraction converts Kirkland et al. (1973)
residual benomyl to carbendazim
Strong cation exchange/HPLC animal 0.01 mg/kg (milk) acidic aqueous hydrolysis followed Kirkland (1973)
0.05 mg/kg by organic extraction converts
(tissue) benomyl to carbendazim and frees
metabolites from conjugates
Reversed phase HPLC water 9.0 x 10-6 g/litre on-line HPLC with preconcentration; Marvin et al. (1991)
benomyl and carbendazim
determined separately
Reversed phase blueberries 0.03 mg/kg acidic methanol extraction converts Bushway et al. (1991)
HPLC/fluorescence detection residual benomyl to carbendazim
Radioimmunoassay plant 0.05-1.0 mg/kg ethyl acetate extraction converts Newsome & Shields
(dependent on crop) residual benomyl to carbendazim (1981)
Enzyme-linked immunosorbent plant 0.50 mg/kg ethyl acetate extraction converts Newsome & Collins
assay (ELISA) residual benomyl to carbendazim (1987)
3. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE
3.1 Natural occurrence
Benomyl does not occur naturally.
3.2 Anthropogenic sources
3.2.1 Uses
Benomyl is one of the most widely used members of a family of
fungicides known as benzimidazoles. It is registered in more than 50
countries for use on more than 70 crops, including cereals, cotton,
grapes, bananas and other fruits, ornamentals, plantation crops,
sugar beet, soybeans, tobacco, turf, vegetables, mushrooms and many
other crops, and is used under most climatic conditions. Registered
benomyl usage specifies rates from 0.1 to 2.0 kg a.i./ha and
applications from once per year to spray intervals ranging from 7 to
14 days (FAO/WHO, 1985a; 1988a). Benomyl is effective at low usage
rates against more than 190 different fungal diseases such as leaf
spots, blotches and blights; fruit spots and rots; sooty moulds;
scabs; bulb, corn and tuber decays; blossom blights; powdery
mildews; certain rusts; and common soilborne crown and root rots.
A key limitation to the use of benomyl and other benzimidazoles
is the development of fungal resistance. Resistance management can
be achieved by using benomyl in combination with a non-benzimidazole
companion fungicide as a tank mix or it may be used alternately with
a non-benzimidazole fungicide (Delp, 1980; Staub & Sozzi, 1984).
Benomyl is formulated as a wettable powder and dry flowable or
dispersible granules. In some countries the latter formulation is no
longer available.
3.2.2 Worldwide sales
In 1991, the estimated worldwide sales of benomyl was US$ 290
million. This was about 50% of the worldwide market for
benzimidazole products. Carbendazim (20%) and thiophanatemethyl
(20%) account for most of the rest of the benzimidazole market
(County NatWest WoodMac).
4. ENVIRONMENTAL TRANSPORT, DISTRIBUTION AND TRANSFORMATION
4.1 Transport and distribution between media
4.1.1 Air
Benomyl has a vapour pressure of < 5.0 x 10-6 Pa (< 3.7 x
10-8 mmHg) and a solubility in water of 3.6 mg/litre at pH 5 and
25 °C. As a result, it has a Henry's constant of < 4.2 x 10-9
atm-m3/mol. Benomyl is essentially non-volatile from water
surfaces.
4.1.2 Water
The half-life of benomyl in surface water and sediment under
aerobic conditions has been shown to be approximately 2 h. Its
metabolite carbendazim had a half-life of 61 days under non-sterile
conditions. After 30 days, 22% of the applied radioactivity was
bound to sediments and < 1% of the applied radioactivity was
evolved as carbon dioxide (Arthur et al., 1989a).
4.1.3 Soil
Radiolabelled benomyl was found to be strongly adsorbed (Ka =
6.1 and 13 µg/g) to two different sandy loam soils and very strongly
adsorbed (Ka = 50 and 90 µg/g) to two different silt loam soils.
Adsorption was not significantly affected by the benomyl
concentration over the range 0.2-2.3 ppm. Adsorbed radioactivity was
not readily desorbed from any of the test soils. The Ka, corrected
for the organic matter content of the soils, was 2-4 times higher on
the silt loam than on the sandy loam soil. This difference suggests
that variables other than percentage organic matter (i.e. cation
exchange capacity, particle size or compound degradation) influence
adsorption. The ease of desorption appears to be inversely related
to the organic content of the soils (Priester, 1985). The structure
of benomyl and its soil degradation products, i.e. carbendazim
(methyl 1H-benzimidazol-2-ylcarbamate), 2-AB (2-aminobenzimidazole),
STB (3-butyl-1,3,5-triazino[1,2a]-benzimidazol-2,4(1H,3H)dione), and
BBU (1-(2-benzimidazolyl)-3-n-butylurea), which is also known as
2-(3-butylureido)-benzimidazole (BUB), are given in Fig. 1. The
major proportion of each of the metabolites was found in the
uppermost (0-12.7 cm) soil layer. The extent of mobility correlated
with the type and characteristics of the soil to which benomyl was
applied. The 14C label was less mobile in soils of lower sand
content and higher silt or clay content. It was also found to be
less mobile on soils of higher organic content and lower pH (Chang,
1985). In a soil column leaching experiment in rice paddy soil,
benomyl did not leach significantly. Approximately 94% was found in
the top 5 cm, 9% in the next 5-10 cm, and less than 1% was detected
in any lower segments (Ryan, 1989). These data indicate that
benomyl, carbendazim, BUB and STB are highly immobile.
Similar mobility results have been observed in the field.
Benomyl and its degradates were studied on bare soil and turf in
four areas of the USA. Carbendazim and 2-AB were the major and minor
degradates, respectively. After 1 and 2 years of outdoor exposure,
the half-life of total benzimidazole-containing residues was about 3
to 6 months on turf and about 6 to 12 months on bare soil (Baude et
al., 1974). Under these conditions, benomyl, carbendazim and 2-AB
showed little or no downward movement.
4.1.4 Leaching
To evaluate the risk of pollution of ground and drainage water,
screening models based on adsorption and persistence can be used,
together with existing analyses of groundwater samples. Gustafson
(1989) proposed the use of the equation GUS = log T´ (4 - log
Koc); GUS values < 1.8 = "improbable leachers", GUS values of
1.8-2.8 = "transition" and GUS values > 2.8 = "probable leachers".
For benomyl, Kom values of 550, 620, 2100 and 1100 (mean 1093)
were found in four different soils (Priester, 1985). A Kom of 1093
is equal to a Koc of 1857 since Koc = Kom x 1.7. The half-life
of 320 days given by Marsh & Arthur (1989) seems in good agreement
with field half-lives of 6 to 12 months (Baude et al., 1974).
When the calculation of the GUS value is based on a Koc of
1857 and a T0.5 of 320 days, a value of 1.83 is obtained.
According to this value, benomyl/carbendazim lies between the
"improbable leachers" and "transition", and, therefore, would not be
expected to occur in ground water. The adsorption of benomyl and of
carbendazim is expected to be of the same order of magnitude since
the Kow values are almost identical (log Kow = 1.49 and 1.36 for
carbendazim and benomyl, respectively). In ground water studies in
the USA (Parsons & Witt, 1988), benomyl was not found in any of 495
wells tested and carbendazim not in any of 212 wells (detection
limit not reported).
In an EEC survey (Fielding, 1992), the presence of carbendazim
in groundwater in the Netherlands and in Italy was investigated.
Carbendazim was found in one of two samples from the Netherlands
(0.1 µg/litre), and the level was above 0.1 µg/litre in 23 of 70
samples in Italy. Detection of the non-polar DDT and lindane in many
wells in the Italian study may indicate macropore transport or
artifacts such as direct pollution of wells.
4.1.5 Crop uptake
Various greenhouse and outdoor tests, in which benomyl was
applied to several crops (apples, bananas, cucumbers, grapes and
oranges), indicate that benomyl and carbendazim remain on plant
surfaces as major components of the total residue (Baude et al.,
1973). Benomyl is primarily converted to carbendazim once inside
plant tissues.
Although benomyl is systemic when applied directly to plant
foliage, crop uptake of soil residues is extremely low, even when
the crop is planted in the same growing season as the benomyl
treatment. In a greenhouse crop-rotation study, [2-14C]-
carbendazim, the more persistent benomyl metabolite, was applied to
a loamy sand soil. Aging periods of 30, 120 or 145 days were used
and the crops studied were beets, barley and cabbage. Radioactivity
did not accumulate in these crops grown to maturity in a loamy sand
soil treated 30 days earlier with 1 kg a.i./ha or 120 to 145 days
earlier with 3 kg a.i./ha. Accumulation factors, calculated as the
ratio of radioactivity in the crop to that in the corresponding
soil, were very low in beet foliage (0.04) and beet roots (0.03),
low in cabbage and barley grain (0.2), and ranged from 0.9 to 1.2 in
barley straw (Rhodes, 1987).
4.2 Transformation
Numerous field studies to determine the fate and behaviour of
benomyl in soil have shown the instability of benomyl under various
conditions. In solutions, plants, and soil, it degrades to
carbendazim. The conversion of benomyl under alkaline conditions to
STB and BBU has also been reported (section 4.1). The environmental
fate of benomyl has been thoroughly reviewed by Zbozinek (1984).
4.2.1 Biodegradation
4.2.1.1 Water
Anaerobic aquatic degradation studies in pond water and
sediment showed a half-life of 2 h for benomyl and 743 days for its
degradation product carbendazim. Some (1-8%) transformation to STB
occurred. After one year 36% of the applied radioactivity was bound
to the sediment (Arthur et al., 1989b).
4.2.1.2 Soil
In a study by Marsh & Arthur (1989), non-sterile and sterile
samples of Keyport silt loam soil were treated with [phenyl(U)-
14C]benomyl at a concentration of approximately 7.0 mg/kg. This is
equivalent to the expected soil residues in the surface 10 cm of
topsoil when benomyl is applied at 8 kg a.i./ha. Distilled water was
added to each sample until it reached 75% of its moisture-holding
capacity at 0.33 bar. The soils were incubated in the dark at
approximately 25 °C. The non-sterile soil flasks were sampled after
0.1, 0.2, 1, 3, 7, 14, 30, 60, 120, 270 and 365 days. Samples of
sterilized soil were taken after 14, 30, 120, 270 and 365 days.
The half-life of benomyl in non-sterile silt loam was 19 h, but
this was not determined in the sterilized soil. Benomyl was rapidly
converted to carbendazim. The carbendazim had a half-life of 320
days under non-sterile aerobic conditions (Marsh & Arthur, 1989).
This is in close agreement with reported half-lives of 6-12 months
for benzimidazoles applied to bare soil (Baude et al., 1974).
After 365 days of incubation, 9% of the 14C was evolved as
14CO2, 34% could still be recovered as carbendazim, and 36% was
not extractable. The total recovery of 14C was 88%.
In the sterilized soil, the half-life of carbendazim was
approximately 1000 days (Marsh & Arthur, 1989).
When the degradation of 2-14C-carbendazim (20 mg/kg) was
determined, 33% of the 14C label added was evolved as 14CO2
during 270 days. Identical or even faster 14C evolution was
observed from 2-14C-labelled 2-AB (Helweg, 1977). The relatively
low 14C evolution from phenyl-14C-labelled benomyl/carbendazim
may be caused by the formation of strongly adsorbed degradation
products or compounds that are readily incorporated into soil
organic matter. Thus, most of the remaining radioactivity was
accounted for in the organic fraction of the soil.
To elucidate the reason for the low 14C evolution from
phenyl-14C-labelled fungicide, the fate of a possible degradation
product, 1,2-diaminobenzene, needs to be determined.
4.2.1.3 Crops
Metabolism studies in various crops (soybeans, rice, sugar beet
and peaches) using [phenyl(U)-14C]benomyl have shown that the only
species of significance in plant tissues are benomyl, carbendazim
and 2-AB. Soybeans were treated twice with 1 kg a.i./ha and
harvested 35 days later. Rice was treated twice with 2 kg a.i./ha
and harvested at 21 days, sugar beet was treated with 0.5 kg a.i./ha
five times and harvested at 21 days, and peaches were treated twice
at 1 kg a.i./ha and harvested 20 min after spraying. Soybeans, rice
and sugar beet were treated at twice the recommended application
rate. The concentration of radiolabelled compounds in mature
soybeans was 0.7 mg/kg and consisted of 0.42 mg 2-AB/kg, 0.05 mg
benomyl/kg and 0.14 mg carbendazim per kg (Bolton et al., 1986a).
Levels in the rice grain were 2.7 and 7.3 mg/kg for benomyl and
carbendazim, respectively (Bolton et al., 1986b). Sugar beet tops
retained 99% of the total recovered radioactivity, 6.8 mg/kg being
present as carbendazim and 0.4 mg/kg as benomyl. The roots retained
only 0.01 mg carbendazim per kg (Tolle, 1988). After the first
application to peaches, benomyl was present at 0.65 mg/kg and
carbendazim at 0.72 mg/kg. The second application resulted in 0.33
mg benomyl/kg and 0.92 mg carbendazim/kg. No other radioactive
metabolites were found in peaches (Stevenson, 1985).
Chiba & Veres (1981) applied benomyl to apple trees as Benlate
50% WP at a rate of 1.7 kg/ha. Three successive applications were
made in 1977 and a single spray was applied in 1979. Between 3 and 7
days after application there was a marked reduction of about 50% in
benomyl residues from an initial level of about 110 mg/kg. This fall
in benomyl was accompanied by a doubling in the level of carbendazim
residues over the same period due to benomyl degradation to
carbendazim. Within 46 days of the single application in 1979,
benomyl residues fell to 0.63 mg/kg foliage and carbendazim was
present at 1.2 mg/kg. Following the three sprayings in 1977 (at 0,
13 and 27 days after the initial application), residue levels were
2.6 and 17.1 mg/kg foliage for benomyl and carbendazim 83 days after
the first spraying. Both experiments showed an exponential fall in
benomyl residues but the rate of decline was much slower in the case
of the more persistent metabolite.
4.2.2 Abiotic degradation
In a study by Wheeler (1985), the hydrolysis of benomyl was
studied in sterilized aqueous solutions maintained at 25 °C in the
dark for 30 days at pH 5, 7 and 9. In pH 5 buffer, the major product
was carbendazim, whereas at pH 7 and 9 carbendazim and STB were the
major products. STB represented approximately 25% of the total
radioactivity at pH 7 and approximately 80% at pH 9. The half-lives
of benomyl in the pH 5, 7 and 9 solutions were approximately 3.5,
1.5 and less than 1 h, respectively. There was no further
degradation of carbendazim at pH 5 and 7 over 30 days. At pH 9,
however, carbendazim was slowly hydrolysed to 2-AB with a half-life
of 54 days (Priester, 1984).
Aqueous photolysis studies conducted in natural sunlight have
shown that benomyl is mainly degraded by hydrolysis rather than
photolysis (Powley, 1985).
4.2.3 Bioaccumulation
Although only low concentrations of benomyl or its metabolites
would be expected in natural waters, studies have evaluated the
metabolism and bioaccumulation in fish. Bluegill sunfish ( Lepomis
macrochirus) were exposed to radiolabelled carbendazim
concentrations of 0.018 or 0.17 mg/litre for 4 weeks in a dynamic
study designed to measure the bioaccumulation of 14C residues in
edible tissue, viscera, remaining carcass and whole fish. A two-week
depuration phase followed the exposure phase. Results were similar
at the two exposure concentrations, the peak whole fish
bioconcentration factors (BCFs) being 27 and 23 at the low and high
exposure levels, respectively. The radioactivity was concentrated
more in the viscera than in other tissues, the peak viscera BCFs
being 460 and 380 for the low and high exposure levels,
respectively. Very little bioconcentration occurred in the muscle
tissue (BCF = < 4) or the remaining carcass (BCF = < 7). During
the 14-day depuration phase, > 94% of the peak level of
radioactivity was lost from the whole fish, viscera and muscle. The
rate of loss from the carcass tissue was lower (77% and 82% loss for
the low and high exposure levels, respectively) (Hutton et al.,
1984).
When rainbow trout ( Oncorhynchus mykiss), channel catfish
( Ictalurus punctatus) and bluegill sunfish ( Lepomis macrochirus)
were injected intraperitoneally with carbendazim, branchial and
biliary excretion were the major pathways for the elimination
(Palawski & Knowles, 1986). In a separate experiment, the three fish
species were exposed to 45 µg carbendazim/litre for 96 h, except in
the case of catfish, which were exposed for 48 h. This was followed
by a 96-h depuration phase. Rainbow trout had the highest uptake
rate constant (1.78 per h) and bioconcentration factor (159) of the
three species. Much less carbendazim was accumulated by channel
catfish than by the other two species, but this residue level (0.44
µg/g) appeared to be lethal after 48 h of exposure. The elimination
rate constant and the biological half-life of carbendazim were
similar for rainbow trout and bluegill sunfish. However, the
elimination rate constant was greater and the biological half-life
shorter in channel catfish (13 h) than in the other two species.
5. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE
5.1 Environmental levels
5.1.1 Air, water and soil
The environmental levels in air, water and soil are discussed
in detail in section 4.
5.1.2 Food and feed
Levels of benomyl in food and feed are indicated in section
5.2.
5.1.3 Terrestrial and aquatic organisms
Benomyl levels in terrestrial and aquatic organisms are
discussed in detail in sections 4 and 6.
5.2 General population exposure
The principal exposure of the general population to benomyl is
through dietary exposure. It was recommended by the Joint FAO/WHO
Meeting on Pesticide Residues (JMPR) (FAO/WHO, 1988b) that all
maximum residue limits (MRLs) for benomyl, thiophanate-methyl and
carbendazim be listed as carbendazim (see Table 5).
5.2.1 USA
A system called the Dietary Risk Evaluation System (DRES),
which was developed by the US Environmental Protection Agency, was
used to quantify the intake of residues occurring in various
commodities. The system assumes a diet consistent with the 1977-1978
USDA Nationwide Food Consumption Survey. This survey was a
stratified probability survey in which 3-day dietary records of
approximately 30 000 individuals were collected. The dietary intake
of residues resulting from registered food crop uses of benomyl was
then estimated using mean residue levels found in controlled field
trials and adjusting for the effects of food processing, e.g.,
washing and cooking, on residues of benomyl and its metabolites.
Based on this analysis, the total dietary exposure was
determined for the general population and for a number of population
subgroups. The exposure of the average person to residues resulting
from benomyl use was estimated to be 0.218 µg/kg body weight per
day. The highest exposure was found in the population subgroup
entitled "non-hispanic other than black or white", the estimated
exposure being 1.479 µg/kg body weight per day. The lowest exposure
was found in the > 20-year-old males where the estimated exposure
was 0.144 µg/kg body weight per day (Eickhoff et al., 1989). These
estimates are below the benomyl ADI allocated by JMPR (0-0.02 mg/kg
body weight per day) (FAO/WHO, 1985a,b).
5.2.2 Sweden
Residue monitoring data for benzimidazole fungicides, i.e.
benomyl, carbendazim and thiophanate-methyl, on food crops from
Sweden is shown in Table 3 (FAO/WHO, 1988b). No further analysis to
determine dietary intake was performed.
5.2.3 Maximum residue limits
National MRLs for certain commodities are listed in Table 4
(FAO/WHO, 1988a).
A complete list of MRLs for carbendazim, including new
proposals and an indication of the source of the data (application
of benomyl, carbendazim, or thiophanate-methyl) on which the MRL is
based, is given in Table 5 (FAO/WHO, 1988b).
5.3 Occupational exposure during manufacture, formulation or use
The levels of inhalation exposure to benomyl and carbendazim
experienced by workers in a major manufacturing facility (DuPont)
were reviewed from 1986 to 1989. The average levels of benomyl and
carbendazim were less than 0.2 mg/m3 and 0.3 mg/m3,
respectively. Table 6 lists established inhalation exposure limits
for benomyl and carbendazim.
5.3.1 Use
Potential dermal and respiratory exposure to benomyl wettable
powder formulation under actual use situations has been determined
for: a) tank loading and mixing for aerial application; b) re-entry
into treated fields; and c) home use (garden, ornamental and
greenhouse). For crop treatments, approximately 17 kg benomyl
(formulation) was handled per cycle. Maximum exposure occurred in
the loading and mixing operation for aerial application, where
dermal exposure was 26 mg benomyl per mixing cycle, primarily to
hands and forearms (90%) and respiratory exposure averaged 0.08 mg
benomyl. Re-entry data revealed dermal and respiratory exposures of
5.9 mg/h and < 0.002 mg/h, respectively. Home-use situations
(application of 7 to 8 litres of benomyl in hand-held compressed air
sprayers) produced exposures of 1 mg and 0.003 mg per application
cycle for dermal and respiratory routes, respectively (Everhart &
Holt, 1982). Similar average dermal exposure levels (5.39 mg/h) for
strawberry harvesters were reported by Zweig et al (1983).
Table 3. Benomyl/carbendazim/thiophanate-methyl residues in food in Swedena
Samples Swedish/imported No. of samples Samples with residues Residue level Median value
>0.20 mg/kg (mg/kg) (mg/kg)
1986
Pineapples imported 3 1 0.69
Grapes imported 20 3 0.17-0.35 0.26
Strawberries imported 7 1 0.29
Mangoes imported 17 4 0.20-1.82 0.70
Papayas imported 5 2 0.25-0.45
Pears Swedish 17 3 0.32-0.62 0.43
imported 45 7 0.20-0.45 0.34
Apples Swedish 78 17 0.20-0.72 0.40
imported 91 30 0.21-0.74 0.39
1987
Grapes imported 28 3 0.52-0.87 0.60
Strawberries imported 7 0.23
Mangoes imported 14 5 0.29-1.30 0.66
Papayas imported 4 2 0.86-1.14
Pears Swedish 14 1 0.52
imported 62 13 0.21-0.45 0.29
Apples Swedish 61 25 0.20-1.17 0.45
imported 94 12 0.21-0.82 0.36
a From: FAO/WHO (1988b)
Table 4. National Maximum Residue Limits (mg/kg) for certain commoditiesa
banana cereal cherries citrus bean cucumber peach pome fruit strawberries grapes
Australia 1 0.05 5 10 3 3 5 5 6 2
Austria 0.2 0.5 7 1 0.5 2 1.5 3
Belgium 2 0.5 2 2 0.5 2 5 5 2
Brazil 1 0.5 10 10 2 0.5 10 5 5 10
Bulgaria 0.5 10 5 5 10
Canada 5 10 1 0.5 10 5 5 5
Denmark 2 0.1 2 5 2 2 2 2 5 5
France 1 1.5 6
Finland 0.2 1 2 0.5 0.5 1 1 1
Germany 0.2 0.5 2 7 1 0.5 2 2 3
Hungary 2 1
Israel 10 10 10 5 10
Italy 0.5 0.5 1 1
Mexico 10 2 1 15 7 5 10
Netherlands 3 0.1 3 4 3 3 3 3 3 3
New Zealand 5 1 5 5 2 2 5 5 5 5
Spain (guidelines) 1 0.5 5 7 2 2 5 5 1 5
Switzerland 1 0.2 3 7 0.2 0.1 3 3 3 3
United Kingdom 1 0.5 10 0.5 10 5 5 10
(proposed)
USA 1 0.2 15 10 2 1 15 7 5 10
USSR 1 0.5 10 10 2 0.5 10 5 5 5
Yugoslavia 0.1 7 0.5 0.1 2 0.5 2
a From: FAO/WHO (1988a)
Table 5. Proposed Maximum Residue Limits for carbendazim from any
sourcea
Commodity MRL (mg/kg) Applicationb
Apricot 10c B,C
Asparagus 0.1d B,T
Avocado 0.5 B
Banana 1c B,C,T
Barley straw and fodder, dry 2 B
Bean fodder 50 C
Beans, dry 2 B
Berries and other small fruit 5 B,C,T
Brussel sprouts 0.5 B
Broad bean 2 T
Carrot 5c C,T
Cattle meat 0.1d B
Celery 2 B,C
Cereal grains 0.5 B,C,T
Cherries 10c B,C,T
Citrus fruits 10c B,C,T
Coffee beans 0.1d C
Common beane 2 C
Cucumber 0.5 B,C,T
Eggs (poultry) 0.1d B,T
Egg plant 0.5 C
Gherkin 2 C,T
Hops, dry 50 C
Lettuce, head 5 B,C,T
Mango 2 B
Melons, except watermelons 2c B,C
Milk 0.1d B
Mushrooms 1 B,C,T
Nectarine 2 B
Onion, bulb 2 C,T
Peach 10c B,C,T
Peanut 0.1d B,C
Peanut fodder 5 B,C
Peppers 5 C
Pineapple 20c B
Plums (including prunes) 2c B,C,T
Pome fruit 5c B,C,T
Potato 3c,f B,C
Poultry meat 0.1d B,T
Rape seed 0.05d C
Rice straw and fodder, dry 15 B,C,T
Sheep meat 0.1d B
Soya bean, dry 0.2 C
Soya bean fodder 0.1d C
Squash, summer 0.5 B
Table 5 (contd).
Commodity MRL (mg/kg) Applicationb
Sugar beat 0.1d B,C,T
Sugar beat leaves on tops 10 B,C,T
Swedeg 0.1d C
Sweet potato 1 B
Taro 0.1d B
Tomato 5 B,C,T
Tree nuts 0.1d B
Wheat straw and fodder, dry 5 B
Winter squash 0.5 B
a From: FAO/WHO (1988b)
b B = benomyl; C = carbendazim; T = thiophanate-methyl
c MRL based on post-harvest use
d At or about the limit of detection
e JMPR recommended 2 mg/kg for dry, dwarf, lima and snap beans. These
are all covered by "VP 0526, Common bean" and "VP 0071, Beans,
dry" in the new classification
f washed before analysis
g Described as rutabagas in 1983 recommendation
Table 6. Established inhalation exposure limitsa
Country and agency Compound TWAb STELc
(mg/m3) (mg/m3)
Australia benomyl 10 -
Belgium benomyl 10 -
Denmark benomyl 5 -
Finland benomyl 10 30
France benomyl 10 -
Switzerland benomyl 10 -
United Kingdom benomyl 10 15
USA: ACGIHd benomyl 10 -
USA: NIOSHe/OSHAf benomyl 10 -
(inhalable dust)
USA: NIOSH/OSHA benomyl 5 -
(respirable dust)
USSR carbendazim - 0.1
a From: ILO (1991)
b Time-weighted average
c Short-term exposure limit
d American Conference of Governmental Industrial Hygienists
e National Institute of Occupational Safety and Health
f Occupational Safety and Health Administration
Air concentrations of benomyl ranged from 0.0074 to 0.053
mg/m3 (average 0.027 mg/m3) during its application in
greenhouses. Spraying tall plants (over 1.5 m) caused three times
higher concentrations in air than spraying low plants. No detectable
amounts of benomyl or its metabolites (carbendazim, 4-HBC and 5-HBC)
were found in the urine of applicators during the 48 h following the
application. However, information describing protective clothing,
ventilation, and other hygienic factors was not reported (Liesivuori
& Jääskeläinen, 1984).
6. KINETICS AND METABOLISM
Benomyl is extensively metabolized by animals, as described in
detail in section 6.3. Metabolite names and structures are given in
Table 6 and Figures 2 and 3.
6.1 Absorption
Absorption in ChR-CD male rats was monitored after dermal
application of 0.1, 1, 10, and 100 mg benomyl (as 2-14C-Benlate 50
WP) at 0.5, 1, 2, 4 and 10 h intervals. Four rats were used for each
treatment and time interval. Benomyl was slowly absorbed across an
area of skin (16% of the animal), appearing in the blood and urine
within 30 min after treatment and reaching a maximum between 2 and 4
h after dosing (Belasco, 1979b). The concentration of benomyl and
its metabolites in the blood peaked at 0.05 mg/litre (2 h sample) in
the low-dose group (0.1 mg) and at 0.10 mg/litre (4 h sample) in the
high-dose group (100 mg). This represented a 20-fold increase in
blood concentration after a 1000-fold dose increase. Thus,
absorption into the bloodstream was non-linear with respect to dose.
An in vitro study on the penetration of formulated benomyl
(Benlate 50 WP) through human skin showed that benomyl penetrates
human skin poorly when it is applied as a recommended spray strength
solution. Much less penetration was detected when dry concentrated
benomyl was applied (Ward & Scott, 1992).
In a rat gavage study, the absorption of carbendazim given in
the form of a corn oil suspension was estimated to be approximately
80% (Monson, 1990).
6.2 Distribution and accumulation
Blood levels of benomyl and its metabolites in male rats were
measured 6 and 18 h after exposure in male rats. The rats were
exposed to time-weighted averages of 0.32 and 3.3 mg/litre of air
for 0.5, 1, 2 and 6 h. The methodology did not distinguish between
benomyl and carbendazim. At both exposure levels, the blood
concentrations of benomyl/carbendazim were greater than that of
5-HBC 6 h after exposure; the levels were 0.39-2.3 mg/litre and
0.25-1.2 mg/litre, respectively. At 18 h after exposure, only 5-HBC
was detected in the blood (1.1 mg/litre) and this only at the
highest dose. Urinary metabolites consisted primarily of 5-HBC, and
limited amounts of benomyl/carbendazim were also detected (Turney,
1979).
Table 7. Chemical names of benomyl and its metabolites in animalsa
Common or abbreviated Chemical name
name
Benomyl Carbamic acid, [1-(butylamino)carbonyl]-
1H-benzimidazol-2-yl]-, methyl ester
Carbendazim (MBC) methyl (1-H-benzimidazol-2-yl)carbamate
5-HBC methyl (5-hydroxy-1H-benzimidazol-2-yl)-
carbamate
4-HBC methyl (4-hydroxy-1H-benzimidazol-2-yl)-
carbamate
5-HBC-Sb 2-[(methoxycarbonyl)amino]-1H-benzimidazol-5-
yl hydrogen sulfate
5-HBC-Gc [2-[(methoxycarbonyl)amino]-1H-benzimidazol-
5-yl] ß-D-glucopyranosiduronic acid
MBC-4,5-epoxide
MBC-5,6-epoxide
MBC-4,5-dihydrodiol (4,5-dihydro-4,5-dihydroxy-1H-benzimidazol-
2-yl) carbamate
MBC-5,6-dihydrodiol (5,6-dihydro-5,6-dihydroxy-1H-benzimidazol-
2-yl) carbamate
MBC-4,5-diol
MBC-5,6-diol
5-OH-6-GS-MBCd S-[5,6-dihydro-5-hydroxy-2-(methoxycarbonyl
amino)-1H-benzimidazol-4-yl]glutathione
5-OH-4-GS-MBC S-[4,5-dihydro-5-hydroxy-2-(methoxycarbonyl
amino)-1H-benzimidazol-4-yl]glutathione
5,6-HOBC-N-oxide methyl (6-hydroxy-5-oxo-5H-benzimidazol-2-
yl)-carbamate-N-oxide
Table 7 (contd).
Common or abbreviated Chemical name
name
5,6-HOBC-N-oxide-G [2-[(methoxycarbonyl)amino]-6-oxo-6H-
benzimidazol-5-yl] ß-D-glucopyranosiduronic
acid-N-oxide
5,6-DHBC methyl (5,6-dihydroxy-1H-benzimidazol-2-yl)
carbamate
5,6-DHBC-G [6-hydroxy-2-[(methoxycarbonyl)amino]-1H-
benzimidazol-5-yl] ß-D-glucopyranosiduronic
acid
5,6-DHBC-S 6-hydroxy-2-[(methoxycarbonyl)amino]-1H-
benzimidazol-5-yl 5-(hydrogen sulfate)
2-AB 2-aminobenzimidazole
2-AB dihydrodiol 2-amino-4,5-dihydro-4,5-dihydroxy-1H-
benzimidazol
5-HAB 5-hydroxy-2-aminobenzimidazole
a From: Krechniak & Klosowska (1986); Monson (1986a,b); Monson (1990)
b S = conjugate with sulfuric acid
c G = conjugate with glucuronic acid
d GS = conjugate with glutathione
In a study by Han (1979), ten male ChR-CD rats were given 1 and
10 µg benomyl intravenously (as 14C-Benlate 50% WP). Radioactivity
was found in the urine as 5-HBC at 6, 12 and 24 h after dosing, and
there was little radioactivity in the blood or faeces at these
sampling times. No radioactivity (< 0.1%) was found in any tissue
after 24 h except in blood, which contained trace quantities of
14C residues.
In a further study, three groups of five rats of each sex were
gavaged with [phenyl(U)-14C] carbendazim. One group received a
single dose of 14C-carbendazim (50 mg/kg). The second group
received a single dose of 14C-carbendazim (50 mg/kg) following 14
days of pre-conditioning with non-radiolabelled carbendazim (50
mg/kg per day). The third group received a single dose of
14C-carbendazim (1000 mg/kg). For all groups, > 98% of the
recovered radioactivity was excreted in the urine or faeces by the
time of sacrifice (72 h after 14C dosing). The 14C remaining in
tissues was < 1% of the applied dose (Monson, 1990).
In a study by Belasco et al. (1969), 14C-benomyl was
administered to male ChR-CD rats and the blood and testes were
analysed. The fungicide was given by gavage as: (a) a single dose of
1000 mg/kg to five rats, which were sacrificed either 1, 2, 4, 7 or
24 h later; (b) 10 repeated doses of 200 mg/kg per day to two rats,
which were sacrificed either 1 or 24 h after the last dose. In
addition, blood and testes from rats fed 2500 mg/kg diet for one
year were analysed. In rats given 1000 mg/kg, results show that: (a)
the total 14C radioactivity (calculated as benomyl) ranged from 3
to 13 ppm in the blood and from 2 to 4 ppm in the testes; (b) 5-HBC
appeared in the blood and testes as early as 1 h after dosing; and
(c) the concentration of benomyl and/or carbendazim decreased with
time and there was a corresponding increase in the concentration of
5-HBC in the blood and testes. Analyses of blood and testes from
rats given 10 repeated oral doses of 200 mg/kg per day showed that
one hour after the last dose no benomyl/carbendazim (< 0.1 ppm) was
detected and only low levels of 5-HBC were found (1.5 ppm blood and
0.3 ppm in testes). No benomyl/carbendazim or 5-HBC (< 0.1 ppm) was
found 24 h after the last dose. With rats fed 2500 mg benomyl/kg
diet for one year, no benomyl/carbendazim (< 0.1 ppm) was detected
in blood or testes. Only a minimal amount of 5-HBC was found in
blood (0.2 ppm) and none was found in the testes (< 0.1 ppm)
(Belasco et al., 1969).
In a series of metabolic studies, benomyl and/or Benlate (50%
benomyl formulation) were administered either by gavage or in the
diet to pregnant ChR-CD rats to determine the concentrations of
benomyl, carbendazim and two carbendazim metabolites (4-and 5-HBC)
in maternal blood and embryonic tissue (Culik, 1981a,b). Dosing took
place on days 7 to 16 of gestation at levels of 125 mg/kg body
weight per day via gavage or 5000-10 000 mg/kg diet (approximately
400-800 mg/kg body weight). Blood samples from the dams and tissue
samples from their embryos were examined on the first, sixth and
tenth days of dietary administration and on days 12 and 16 of gavage
administration. Embryos and maternal blood were analysed 1, 2, 4, 8
and 24 h after gavage.
The levels of benomyl/carbendazim in maternal blood and
embryonic tissues, 24 h following each dose, markedly decreased with
the number of treatments. The level of benomyl (one hour after
treatment) ranged from 0.98 to 8.4 mg/kg with a mean value of 5.0
mg/kg on the first day of treatment. After 10 treatments, the levels
of benomyl/carbendazim ranged from < 0.12 to 0.39 mg/kg (one hour
after last treatment). In the embryo there was 0.13 mg/kg
benomyl/carbendazim after the tenth treatment compared with a mean
of 1.9 mg/kg after the first treatment. The half-life of benomyl in
maternal blood was approximately 45 min and was less in the embryos.
The level of 5-HBC (0.84-2.9 mg/kg) 2 h following the last gavage
increased with the number of exposures, the half-life in the blood
being 2-3 h in the dam and 4-8 h in the embryo. 4-HBC was not
detected.
In the dietary studies, the levels of benomyl, carbendazim and
4-HBC were too low to be measured in the embryonic tissue. 4-HBC
could not be detected in the dams. Irrespective of the dose level
(5000 and 10 000 mg/kg diet active ingredient) of benomyl or
Benlate, the level of benomyl/carbendazim in maternal blood was very
low. In three separate groups of animals, the mean highest blood
concentrations of benomyl/carbendazim were 0.35, 0.61 and 0.23 mg/kg
in each group of dams. The mean highest value of 5-HBC (5000 mg
benomyl/kg diet) was 0.44 mg/kg in the blood and 0.33 mg/kg in the
embryos. Animals fed benomyl or Benlate at a level of 10 000 mg/kg
diet had 5-HBC levels an order of magnitude higher (Culik, 1981a,b).
A lactating Holstein cow was dosed by capsule twice daily (515
mg [2-14C]-benomyl each dose), equivalent to 50 mg/kg in the
average total daily feed, for 5 consecutive days, and samples of
urine, faeces and milk were collected at each dosing. Approxi mately
17 h after the tenth dose, the cow was sacrificed and organ, tissue
and blood samples were subsequently collected. 14C residue levels
in the milk averaged 0.2 mg/kg (calculated as benomyl), 49% of the
radioactive metabolites being extractable in ethyl acetate, 36%
soluble in water, and 8% isolated as solids. Small amounts of
radioactivity were detected in the liver (4.12 mg/kg) and kidney
(0.25 mg/kg), most of which was bound. No significant levels of
radioactivity (0.06 mg/kg) were detected in other tissues or fat
(Monson, 1985).
Lactating and non-lactating goats were given daily capsule
doses of [2-14C]-benomyl, equivalent to 36 and 88 mg/kg,
respectively, in the total daily diet, for five days. Milk residues
accounted for approximately 2% of the total dose. Approximately 25%
of the milk radioactivity was incorporated into the natural milk
components casein and whey protein. There were no detectable
residues in muscle tissue and fat (< 0.01 mg/kg). However,
radioactivity detected in liver and kidney amounted to 3.8 and 0.09
mg/kg (calculated as benomyl equivalents), respectively (Han, 1980).
In a study by Johnson (1988), the total 14C residue and
metabolic fate of carbendazim in the liver was examined in
non-lactating female goats. Twelve goats were administered a
feed-rate-equivalent dose of [phenyl(U)-14C]-carbendazim (> 50
mg/kg), once a day, for up to 30 days. Within 2 weeks of dose
initiation, a plateau of 14C residues in the liver was achieved at
a level of 9.48 mg/kg (group mean of the total radiolabelled liver
residues for goats sacrificed 2, 3, and 4 weeks after initiation of
dosing). The total 14C residue levels in the liver decreased to
5.17, 3.55 and 1.67 mg/kg (calculated as carbendazim equivalents) 1,
2 and 3 weeks, respectively, after dosing ceased. The elimination
half-life for total 14C residues from the liver, based on this
depuration data, was calculated to be approximately 9 days. The
half-life for removal of carbendazim from the general circulation,
based on 14C-carbendazim equivalent whole blood levels, was
approximately 10 h. The level of bound, non-extractable 14C
residues in the liver of goats sacrificed after 28 days was 1.0
mg/kg.
The results of this study suggest that levels of carbendazim-
derived residues do not accumulate beyond 2 weeks when goats are
exposed to a constant feed level of 50 mg carbendazim/kg.
Furthermore, discontinuation of exposure results in a clearing of
residues from the liver (Johnson, 1988).
The metabolism of benomyl was studied in laying hens by Monson
(1986a). Two hens were individually dosed daily for three
consecutive days with 3.5 mg [2-14C]-benomyl at a rate equivalent
to 29 mg/kg in the daily feed, and two hens were individually dosed
with 3.29 mg [phenyl(U)-14C]-benomyl at rates equivalent to 27
mg/kg in the daily feed. Faeces and eggs from the previous 24 h were
collected just before each dosing. Twenty-two hours after the third
dose, the hens were killed and samples of muscle (breast and thigh),
liver, kidney and fat were analysed. The concentration of
radioactivity (calculated as benomyl equivalents) in the tissues and
day-3 eggs of the [2-14C]-benomyl- and [phenyl(U)-14C]-benomyl-
dosed hens, respectively, was as follows: liver (0.54 and 0.41
mg/kg), kidney (0.28 and 0.16 mg/kg), thigh and breast muscle (both
0.01 mg/kg), fat (0.05 and 0.02 mg/kg) and eggs (0.08 and 0.05
mg/kg).
The distribution of benomyl in this study was comparable to
that in a 20-hen [2-14C]-carbendazim metabolism study. The
concentrations of radioactivity, calculated as mg carbendazim/kg, in
the high-dose laying hens (dose equivalent 120 mg/kg carbendazim in
the diet) were liver (2.63), kidney (1.74), thigh muscle (0.06),
breast muscle (0.05), fat (0.03), day-6 eggs (0.63) (Monson, 1986b).
This study is discussed in detail in the Environmental Health
Criteria monograph on Carbendazim (WHO, 1993).
When bluegill sunfish were exposed to benomyl, carbendazim and
2-AB at nominal concentrations of 0.05 mg/litre (measured
concentrations of 0.01 to 0.04 mg/litre) and 5.0 mg/litre (measured
concentrations of 2 to 5 mg/litre), no residues were found in the
tissues of fish exposed to low levels of these three compounds.
Detectable residues were found in the tissues of fish exposed to the
high levels, but there was no build-up or bioconcentration with time
(DuPont, 1972).
6.3 Metabolic transformation
Benomyl is extensively metabolized by rats to carbendazim,
which is then further metabolized. Studies with rats administered
benomyl intravenously (Han, 1979), dermally (Belasco, 1979b) or by
inhalation (FAO/WHO, 1985a) showed that 5-HBC is the main urinary
metabolite, some carbendazim also being present.
In a rat gavage study (Monson, 1990; see section 6.2),
carbendazim was extensively metabolized. Three dosing regimens (five
rats of each sex per group) were used: a single oral dose of 50
mg/kg (low dose); a single oral dose of 50 mg/kg following
pre-conditioning gavage with non-radiolabelled carbendazim at 50
mg/kg for 14 days (pre-conditioned low dose); and a single oral dose
of 1000 mg/kg (high dose). The 48-h urine from the low-dose and the
high-dose rats and the 14-day urine from the pre-conditioned
low-dose group were collected. The total recovery from urine was
61.5 and 61.7% of given doses for the low-dose and pre-conditioned
low-dose male groups, 53.2 and 59.3% for the low-dose and
pre-conditioned low-dose female groups, and 39 and 41% for both male
and female high-dose groups, respectively. 5-HBC-S (21-43% of given
dose) was identified as the main metabolite, except in the case of
the pre-conditioned low-dose and high-dose female rat groups
(5.5-10%), while in all female rat groups 5,6-HOBC-N-oxide-G
(10-19%) was predominant. Both 5,6-DHBC-S and 5,6-DHBC-G were
identified as minor metabolites.
In the same study, the faeces were collected at the same
periods as the urine. The total recovery from faeces was about 24%
for the low-dose and pre-conditioned low-dose male groups, 33-38%
for the low-dose and pre-conditioned low-dose female groups, and
higher (> 60%) for both male and female high-dose groups. Unchanged
carbendazim was about 10-15% of the given dose in the faeces of
high-dose rats (Monson, 1990). The proposed metabolic pathway for
benomyl in rats is given in Fig. 2.
When a lactating Holstein cow was dosed by capsule twice daily
(515 mg per dose), equivalent to 50 mg/kg diet, for 5 consecutive
days with [2-14C]-benomyl, the major metabolites of whole milk
were 5-HBC (0.06 mg/litre), 4-HBC (0.03 mg/litre) and MBC-4,5-
dihydrodiol (< 0.07 mg/litre). The proportions of radioactive
residues in the urine were 46% 5-HBC, 3% 4-HBC, and 50% polar
aqueous-soluble metabolites, which included MBC-4,5-dihydrodiol,
2-AB-dihydrodiol and 5-OH-4-GS-MBC (Monson, 1985).
Lactating and non-lactating goats were given five consecutive
daily doses of 2-14C-benomyl by capsule at rates equivalent to 36
and 88 mg/kg, respectively, in the total daily diet. The main
metabolite in milk was 5-HBC, and there were minor amounts of 4-HBC
and 5-HAB. The principal metabolites in urine and faeces were 5-HBC
and 4-HBC. The main identified metabolite in the kidney and liver
was 5-HBC (about 6% of the residue). Much of the liver residue was
incorporated into glycogen, protein, fatty acids and cholesterol,
and accounted for approximately 35% of the liver residues. Further
characterization of the bound liver tissue residues following
enzymatic and trifluoroacetic anhydride hydrolysis identified
5-hydroxy-benzimidazole moieties as the principal (at least 77%)
14C residue in goat liver. No free benomyl, carbendazim or 5-HBC
was detected in the liver (Han, 1980; Hardesty, 1982).
In a further study, the total 14C residue and metabolic fate
of carbendazim in the liver were examined in non-lactating female
goats. Twelve goats were administered a dose equivalent to 50 mg/kg
feed once a day for up to 30 days. Extraction of composite liver
homogenate from goats sacrificed 4 weeks after initiation of dosing
("plateau level") indicated that the major ethyl acetate extractable
and identifiable radiolabelled residues in the liver were 5-HBC (2
to 3 mg/kg) and carbendazim (approximately 0.2 mg/kg) (Johnson,
1988).
The metabolism of [2-14C]-benomyl and [phenyl(U)-14C]-
benomyl has been studied in laying hens (see section 6.2 for a
detailed description of the study). Benomyl was extensively
metabolized to carbendazim, 5-HBC, MBC-4,5-dihydrodiol and a
metabolite tentatively identified as 5-OH-4-GS-MBC. The metabolic
profile observed in hens indicates that the benzimidazole ring is
not broken during metabolism (Monson, 1986a). The proposed metabolic
pathway for benomyl in the laying hen is given in Fig. 3.
Monson (1991) analysed the release and characterization of
bound benomyl and carbendazim metabolites in diary cow, goat, hen
and rat liver after treatment with 14C-benomyl or
14C-carbendazim via Raney nickel desulfurization and acid
dehydration. Using this technique, he was able to show that bound
14C residue was released from the liver of cows (76% bound before
desulfurization and 36% bound after desulfurization) and hens (58%
bound before desulfurization and 19% bound after desulfurization).
The major part of the reduced residue was identified as 5-HBC,
5,6-HOBC or carbendazim, suggesting that the bound liver residue
consisted of conjugates of benzimidazole-related products and not
natural products resulting from breakdown and incorporation.
In fish, benomyl and carbendazim are metabolized to 5-HBC
(Dupont, 1972).
6.4 Elimination and excretion
Absorbed benomyl and carbendazim are rapidly excreted in the
urine and faeces.
In a study where rats were administered 1 or 10 µg formulated
14C-benomyl (50% wettable powder) in a single intravenous dose by
tail injection, more than 80% of the dose was excreted in the urine
and faeces within 6 h after injection and the total urine and faeces
recovery was > 95% in 24 h (Han, 1979).
[Phenyl(U)-14C]-carbendazim was administered by gavage to
Sprague-Dawley rats using three dosing regimens: a single oral dose
of 50 mg/kg (low dose); a single oral dose of 50 mg/kg following
pre-conditioning gavage with unlabelled carbendazim of 50 mg/kg for
14 days (pre-conditioned low dose); and a single oral dose of 1000
mg/kg (high dose). Each dosing group consisted of five animals of
each sex. A preliminary study conducted with two rats of each sex,
each rat having received a single oral dose of 50 mg/kg,
demonstrated that 95% of the radioactivity excreted in the urine and
faeces was recovered within 72 h after dosing and that < 1% of the
dose was expired as volatile metabolites. In the full study, > 98%
of the recovered radioactivity was excreted by the time of sacrifice
(i.e. 72 h after dosing) for each dosing group. Urinary excretion
accounted for 62% to 66% of the dose in males and 54% to 62% of the
dose in low-dose and pre-conditioned low-dose female groups. In the
high-dose group, this pathway accounted for 41% of the dose in all
animals. Elimination of radiolabel in faeces accounted for virtually
all of the remaining radiolabel. There were no apparent differences
between male and female rats with respect to the extent of
absorption and extent and rate of elimination of 14C-carbendazim
equivalents within a given treatment group (Monson, 1990).
In a study by Han (1978), two male ChR-CD-1 mice were fed a
diet of non-radiolabelled benomyl (2500 mg/kg) for 21 days and were
then gavaged with 2.5 mg [2-14C]-benomyl in corn oil. An identical
experiment was performed with one male ChR-CD hamster. More than 90%
of the radioactivity was eliminated in the urine and faeces within
72 h (Han, 1978).
A lactating Holstein cow was dosed by capsule twice daily (515
mg [2-14C]-benomyl each dose), equivalent to 50 mg/kg in the
average total daily feed, for 5 consecutive days, and samples of
urine, faeces and milk were collected at each dosing. Approximately
17 h after the tenth dose, the cow was sacrificed and organs,
tissues and blood were collected for analysis. At sacrifice, 65% of
the radiolabel had been excreted in the urine, 21% in the faeces and
0.4% in the milk. Carbon-14 residue levels in the milk averaged 0.2
mg/litre (calculated as benomyl equivalents) with 49% of the
radioactive metabolites being extractable in ethyl acetate, 36%
soluble in water, and 8% isolated as solids (Monson, 1985).
Lactating and non-lactating goats were given 5 consecutive
daily doses of [2-14C]-benomyl by capsule at rates equivalent to
36 and 88 mg/kg, respectively, in the total daily diet. Most of the
radioactivity (96%) had been eliminated in the urine and faeces by
the time of sacrifice (Han, 1980).
The excretion of benomyl was studied in laying hens dosed daily
for three consecutive days with 3.5 mg [2-14C]-benomyl or 3.29 mg
[phenyl(U)-14C]-benomyl. At sacrifice (22 h after the last dose),
an average of 107% and 95% of the dose had been excreted for the
[2-14C]-benomyl- and [phenyl(U)-14C]-benomyl-dosed birds,
respectively (Monson, 1986a).
In a similar study on 14C-carbendazim, groups of laying hens
were fed at a rate equivalent to 5 and 120 mg/kg diet. At sacrifice,
24 h after the sixth daily dose, an average of 95% of the dose had
been excreted by the low-dose birds and 92% by the high-dose birds
(Monson, 1986b).
6.5 Reaction with body components
An in vitro study using acetyl cholinesterase from bovine
erythrocytes showed that benomyl did not inhibit this enzyme. The
acetyl cholinesterase inhibition constant (KI) for benomyl was
greater than 1 x 10-3 mol/litre (Belasco, 1970). Another in vitro
study by Krupka (1974) verified that benomyl did not inhibit either
acetyl cholinesterase or butyryl cholinesterase.
In a study by Guengerich (1981), the effects of benomyl and
carbendazim on hepatic enzymes were studied in male and female
Crl-CD rats and CD-1 mice. The treatment groups included animals fed
for 28 days with diets that contained benomyl or carbendazim at
concentrations of 0, 10, 30, 100, 300, 1000 or 3000 mg/kg. In these
studies, microsomal epoxide hydrolase and cytosolic glutathione- S-
transferase were monitored in subcellular fractions isolated from
the livers of animals in each treatment group. Liver weights were
also recorded. Elevated mean absolute liver weights were observed at
1000 and 3000 mg carbendazim/kg in both male and female rats and at
300 mg carbendazim/kg in female rats. However, the only
significantly elevated liver weight was found in females after a
dose of 3000 mg benomyl/kg. No apparent liver toxicity or effect on
body weight was observed. Both benomyl and carbendazim induced
epoxide hydrolase in both sexes of rats and mice at 1000 and 3000
mg/kg. Induction of glutathione- S-transferase was observed at 3000
mg/kg in the case of both benomyl and carbendazim. In general, the
level of induction seemed to be slightly greater in females than
males. There did not appear to be any substantial difference in
enzyme induction between rats and mice.
In a study by Shukla et al. (1989), levels of gamma-glutamyl
transpeptidase (GGT) were evaluated after benomyl exposure. Female
albino rats (eight per group) and female Swiss albino mice (eight
per group) were given 1000 and 4000 mg benomyl/kg feed for 15 days,
and blood and liver GGT levels were analysed. Benomyl exposure
increased the activity of both blood and liver GGT in both rats and
mice, and the degree of induction was dose related (Shukla et al.,
1989).
7. EFFECTS ON LABORATORY MAMMALS; IN VITRO TEST SYSTEMS
7.1 Single exposure
The acute toxicity of benomyl in several animal species is
summarized in Table 8. Benomyl has an oral LD50 in the rat of >
10 000 mg/kg and an inhalation 4-h LC50 > 4 mg/litre. Several
other minor metabolites were evaluated and the approximate lethal
doses were 3400 mg/kg for 2-AB, 7500 mg/kg for 5-HBC, 17 000 mg/kg
for BUB and 17 000 mg/kg for STB.
In a study by Littlefield & Busey (1969), three groups of male
dogs (around 10 dogs/group) were exposed to benomyl at air
concentrations of 0, 0.65 and 1.65 mg/litre. One half of the dogs in
each group were killed on day 14 and the remainder on day 28. The
liver weight of the high-dose dogs was significantly decreased on
day 28. For further discussion of single dose toxic effects, see
section 7.5.1.
7.2 Short-term exposure
7.2.1 Gavage
In a 14-day rat study, benomyl (200 and 3400 mg/kg in peanut
oil) was given by gavage five times a week for two weeks to six male
ChR-CD rats per group. Four out of six rats died after 5, 7, 8 and 9
doses, respectively, of 3400 mg/kg. No clinical signs of toxicity
were observed in the group treated with 200 mg/kg per day.
Degeneration of germinal epithelium, multinucleated giant cells and
reduction or absence of sperm were observed in the testes after
multiple doses of 3400 mg/kg per day. Less than 10% of the
testicular tubules were affected in only two out of six animals
dosed with 200 mg/kg. At the high dose level, there was erosion and
thickening of the squamous mucosa of the stomach with submucosal
inflammation and a decrease in the large globular-shaped vacuoles
located centrolobularly in the liver (Sherman & Krauss, 1966).
7.2.2 Feeding
7.2.2.1 Rat
In a 90-day study by Sherman et al. (1967), groups of rats
(4-week-old ChR-CD rats, 16 rats of each sex per group) were fed
Benlate 70 WP (72% benomyl) in the diet at levels of 0, 100, 500 and
2500 mg benomyl/kg. The animals were observed daily for behavioural
changes and body weights, and food consumption was recorded at
weekly intervals. Haematological examinations were conducted on six
male and six female rats in each group at 30, 60 and 90 days.
Routine urine and plasma alkaline phosphatase and glutamic pyruvic
transaminase activity analyses were performed on the same animals.
After 96-103 days of continuous feeding, 10 male and 10 female rats
in each group were killed, and selected organs were weighed and
examined microscopically. The remaining six male and female animals
in each group after the terminal sacrifice were used in a
one-generation reproductive study. No effect was observed with
respect to reproduction or lactation in the delivery or rearing of
the F1A litters. There were no compound-related effects on weight
gain, food consumption, food efficiency, clinical signs, or on
haematology, biochemistry or urinalysis determinations. The
liver-to-body weight ratio in females was slightly increased at 2500
mg/kg, compared with control rats. Gross and microscopic
examinations of tissues and organs showed no significant effects
attributable to the presence of benomyl in the diet at levels up to
and including 2500 mg/kg.
7.2.2.2 Dog
Groups of beagle dogs (four males and four females per group;
7-9 months old) were administered benomyl 50% wettable powder in the
diet at dosage levels of 0, 100, 500 and 2500 mg/kg diet (based on
active ingredient) for three months (this corresponded to treatment
levels of 0, 3.8, 18.4 and 84 mg/kg body weight). Food consumption
and body weight data were recorded weekly, and clinical laboratory
examinations (including haematology, biochemistry and urinalysis)
were performed pre-test and after 1, 2 and 3 months of feeding. At
the conclusion of the study, selected organs were weighed and
subjected to gross and microscopic examinations. No mortality or
adverse clinical effects were observed over the course of the study,
and growth and food consumption were not effected by the treatment.
Urine parameters showed no differences from the control, and there
were no dose-related effects on the haematological values. Alkaline
phosphatase and glutamic pyruvic transaminase activities were
increased in high-dose males and females. There were statistically
significant decreases in the albumin/globulin ratio in either males
or females fed 2500 mg/kg diet. Organ-to-body weight ratio changes
were observed in the high-dose males and females for the thymus
(decreased) and thyroid (increased). One of the four females fed
2500 mg/kg diet had an enlarged spleen at the end of the exposure
period, as well as a decreased erythrocyte count, haemoglobin
concentration and haematocrit value. Histopathological examination
revealed myeloid hyperplasia of the spleen and bone marrow and
erythroid hyperplasia of bone marrow. This did not appear to be
compound related since group mean values were not significantly
different. Three out of four males fed 2500 mg/kg diet had reduced
relative prostate weights when compared with controls. Microscopic
examinations of tissues and organs did not indicate changes in dogs
fed benomyl for 90 days. The no-observed-effect level (NOEL) was 500
mg/kg diet (Sherman, 1968).
Table 8. Acute toxicity of benomyl and its metabolites for laboratory mammals
Chemical Species Sex Number of Route Vehicle Concentrationa Reference
animals (mg/kg body weight)
Benomyl ratb M/F 10/dose oral peanut oil LD50 > 10 000 Sherman (1969a)
rabbitc M 1/dose oral 50% wettable powder ALD > 3400 Fritz (1969)
dogd M 1 oral evaporated milk and ALD > 1000 Sherman (1969b)
water (1:1)
Benlate OD (50% rat M 10/dose oral corn oil LD50 > 12 000 Hostetler (1977)
benomyl)
Fungicide 1991 rabbit M/F 4/dose dermal (4 h) 50% wettable powder LD50 > 10 000 Busey (1968a)
(50% benomyl)
rat M 6/dose inhalation (4 h) 50% wettable powder LC50 > 4.01 mg/litre Busey (1968b)
(analytical)
dog M 10/dose inhalation 50% wettable powder LC50 > 1.65 mg/litre Littlefield & Busey
(analytical) (1969)
Benlate fungicide rat M/F 10/dose oral aqueous suspension LD50 > 10 000 Sherman (1969a)
(52-53% benomyl)
rat M 5/dose inhalation 50% wettable powder LC50 > 0.82 mg/litre Hornberger (1969)
Benlate PNW (50% rabbit M/F 10/dose dermal 50% wettable powder LD50 > 2000 Gargus & Zoetis
benomyl) (1983c)
Benlate 50 DF (50% rat M/F 5/dose oral aqueous suspension LD50 > 5000 Sarver (1987)
benomyl)
rabbit M/F 5/dose dermal 50% dry flowable LD50 > 2000 Brock (1987)
Table 8 (contd).
Chemical Species Sex Number of Route Vehicle Concentrationa Reference
animals (mg/kg body weight)
Benomyl metabolites
2-Benzimidazole carbamic rat M/F 10/dose oral corn oil LD50 > 10 000 Goodman (1975)
acid, methyl ester
5-Hydroxy-2-benzimidazole- rat M 1/dose oral corn oil ALD > 7500 Snee (1969)
carbamic acid, methyl ester
2-Aminobenzimidazole rat M 1/dose oral peanut oil ALD > 3400 Fritz & Sherman
(1969)
Benzimidazole 2- rat M 1/dose oral corn oil ALD > 17 000 Dashiell (1972)
(3-butylureido)
S-Triazine, 3-butyl- rat M 1/dose oral corn oil ALD > 17 000 Barbo & Carroll
benzimidazole (1,2a), (1972)
-2,4(1H,3H)-dione
a Based on active ingredient; ALD = approximate lethal dose
b ChR-CD or Crl:CD rats
c New Zealand white rabbits
d Beagle dogs
7.2.3 Dermal
In a study on groups of five male and five female New Zealand
albino rabbits, weighing 2 to 2.4 kg, 15 dermal applications of a
50% benomyl formulation (equivalent to 1000 mg/kg) were made on both
abraded and intact abdominal skin sites. The animals were exposed
for 6 h/day, 5 days/week for 3 weeks. After each daily application,
the abdomen was washed with tap water. Observations were made daily
for mortality and toxic effects and weekly for body weight changes.
Gross necropsy and microscopic examinations were performed. Slight
erythema, oedema and atonia were observed at both abraded and intact
skin sites. Slight to moderate desquamation occurred throughout the
exposure period. No apparent compound-related body weight or organ
weight changes were reported. Microscopic examination of the males
demonstrated that benomyl produced degeneration of the spermatogenic
elements of the seminiferous tubules of the testes, the changes
including vacuolated and multi-nucleated spermatocytes (Busey,
1968d).
In a separate repeated-dose dermal study, groups of five male
and five female New Zealand albino rabbits, weighing 3 kg, were
exposed to doses of benomyl equivalent to 0, 50, 250, 500, 1000 and
5000 mg/kg applied to non-occluded abraded dorsal skin sites 6 h a
day, five days a week, for three weeks. Test material was removed by
washing the skin site and drying with a towel. There were decreased
body weight gains for both males and females at the two highest dose
levels. Mild to moderate skin irritation was reported for all groups
but was most notable at the highest dose level. Diarrhoea, oliguria
and haematuria were observed in males and females at 1000 and 5000
mg/kg. Decreased average testicular weights and testes-to-body
weight ratios were observed at 1000 mg/kg only. There were no
histopathological changes reported (Hood, 1969).
7.2.4 Inhalation
In an inhalation study, groups of 20 male and 20 female CD rats
were exposed, nose-only, 6 h a day for 90 days, to 0, 10, 50 and 200
mg benomyl/m3. At 45 and 90 days, blood and urine samples were
collected from 10 rats of each sex per group for clinical analysis
and then killed for pathological examination. After approximately 45
days of exposure, test-compound-related degeneration of the
olfactory epithelium was observed in all males and in eight of the
ten females exposed to 200 mg/m3. Two male rats exposed to 50
mg/m3 had similar but less severe olfactory degeneration. After
approximately 90 days of exposure, all of the animals showed
olfactory degeneration at 200 mg/m3, along with three males
exposed to 50 mg/m3. No other compound-related pathological
effects were observed. Male rats exposed to 200 mg/m3 had
depressed mean body weights compared to controls and this correlated
with a reduction in food consumption (Warheit et al., 1989).
7.3 Skin and eye irritation; sensitization
7.3.1 Dermal
A 50% wettable powder applied to the clipped intact and abraded
abdomen of albino rabbits produced moderate to marked erythema,
slight oedema and slight desquamation. Exposure was for 24 h to
occluded skin sites at doses > 0.5 g/animal. Albino guinea-pigs
similarly exposed to 10, 25 and 40% dilutions of technical grade
benomyl in dimethyl phthalate presented only mild irritation of both
intact and abraded skin sites (Majut, 1966; Busey, 1968a; Colburn,
1969; Frank, 1969).
When "Benlate" 50 DF (50% benomyl, 0.5 g Benlate 50 DF) was
evaluated for primary dermal irritation potential in six male New
Zealand white rabbits, no dermal irritation was observed at 4 or 24
h after application. By 48 h, slight to mild erythema was observed
in two rabbits and was still evident at 72 h. The primary irritation
scores ranged from 0-1 (not an irritant) (Vick & Brock, 1987).
In a study by Desi (1979), benomyl (98% purity) was applied to
a shaved area of the back of four albino rabbits at 5 mg/cm2.
Draize scores (Draize et al., 1944) were assessed 4 and 72 h after
application. Lesions produced by this method were classified as
"mild irritation".
7.3.2 Eye
The eye irritation properties of benomyl were examined in
albino rabbits in several tests using technical grade benomyl, 50%
wettable powder and a suspension in mineral oil. Mild conjunctival
irritation and minor transitory corneal opacity were reported after
48 to 96 h in all tests (Reinke, 1966; Frank, 1972). Similar results
were obtained with Benlate PNW (a 50% wettable powder) (Gargus &
Zoetis, 1983a,b).
Another eye irritation experiment was performed with 5 mg pure
benomyl using four albino rabbits (Desi, 1979). Results assessed
according to Draize (Draize et al., 1944) indicated that benomyl is
a mild eye irritant.
7.3.3 Sensitization
Albino guinea-pigs exposed to benomyl, either technical
material or a 50% sucrose formulation, produced mild to moderate
skin erythema during the challenge phase following both intradermal
injections or repeat applications to abraded skin (Majut, 1966;
Colburn, 1969; Frank, 1969).
In another sensitization study, Benlate PNW (50% benomyl
prepared as a 0.1% solution in saline) was injected weekly (four
injections) into ten albino guinea-pigs (Hartley strain). Ten
control animals were injected with saline. Fourteen days after the
final injection, 8% or 80% Benlate PNW saline solutions were applied
to the backs of the induced animals and saline was applied to the
backs of the control animals. No significant increase in score
occurred in any of the control animals at either challenge
concentration. Benlate PNW produced an unequivocal and significant
(two-step) increase at two of ten sites challenged with an 8%
suspension, and at seven of ten sites challenged with an 80%
suspension (Gargus & Zoetis, 1984).
Technical benomyl produced sensitization in all ten animals
tested in a guinea-pig maximization test (Matsushita et al., 1977).
7.4 Long-term exposure
7.4.1 Rat
Groups of weanling rats (36 male and 36 female Charles River
albino rats/group) were fed benomyl (50-70% a.i.) in the diet for
104 weeks at levels of 0, 100, 500 and 2500 mg/kg. Growth, as
observed by body weight changes and food consumption data, was
recorded weekly for the first year and twice a month thereafter.
Daily observations were made of clinical effects and mortality. At
periodic intervals during the study, haematological, urinalysis and
selected clinical chemistry examinations were performed. After one
year each group was reduced to 30 males and 30 females by interim
sacrifice for gross and microscopic evaluations. At the conclusion
of the study, all surviving animals were sacrificed and gross
examinations of tissues and organs were made. Initially, microscopic
examinations of tissues and organs from the control and 2500 mg/kg
groups were conducted, as were liver, kidney and testes examinations
of animals in the 100 and 500 mg/kg dose groups. In follow-up
pathological evaluations, all of the tissues and organs of the
control and low-, intermediate- and high-dose groups were examined
microscopically. There was no mortality attributable to benomyl in
the diet. Survival decreased to approximately 50% during the second
year, but was comparable among all groups. Body weight, food
consumption and food efficiency were unaffected by treatment. The
average daily dose for the 2500 mg/kg group was 330 mg/kg body
weight per day initially, 91-106 mg/kg body weight per day at one
year and 70-85 mg/kg body weight per day at two years. There were no
compound-related clinical manifestations of toxicity.
Haematological, urine and liver function tests were unaffected by
treatment. There were no differences in organ weight or organ-to-
body-weight ratios between control and treated groups (Sherman,
1969c; Lee, 1977).
7.4.2 Mouse
In a study by Weichman et al. (1982), male and female CD-1 mice
(80 males and 80 females per group) were administered benomyl (99%
a.i.) in the diet at levels of 0, 500, 1500 and 5000 mg/kg (the
highest levels was reduced from 7500 mg/kg after 37 weeks) for two
years. The mice were 6-7 weeks old at the start of the study. Median
survival time was unaffected by treatment. Male and female mice fed
1500 or 5000 mg/kg exhibited dose-related body weight decreases.
Food consumption was variable throughout the study, although
high-dose females appeared to consume less food. The average daily
intake of benomyl for males was 1079 mg/kg body weight per day
initially, 878 mg/kg body weight per day for 1 year and 679 mg/kg
body weight per day for 2 years; for females it was 1442 mg/kg body
weight per day initially, 1192 mg/kg body weight per day for 1 year,
and 959 mg/kg body weight per day for 2 years