634. Mecarbam (Pesticide residues in food: 1983 evaluations)

PESTICIDE RESIDUES IN FOOD - 1983

Sponsored jointly by FAO and WHO

EVALUATIONS 1983

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

Food and Agriculture Organization of the United Nations
Rome 1985

MECARBAM

TOXICOLOGY

Explanation

Mecarbam was evaluated by the Meeting in 1980 (FAO/WHO 1981)^1/ and a
temporary acceptable daily intake (ADI) was allocated. Further studies
were required by the previous Meeting which included complete
metabolic studies in laboratory animals; a complete teratogenicity
assay; studies to define the mutagenic potential, and studies in hens
or other appropriate species to define the potential for delayed
neurotoxicity. Some of the required studies have become available and
are reviewed in this monograph addendum.

A temporary maximum residue level for mecarbam in oranges was
estimated by the 1980 Meeting. Several items of information on
residues were required for 1983, including additional data for citrus
and other fruit. Information on registered uses for citrus and residue
data for citrus and grapes were provided and are reviewed herein. Also
provided and evaluated are residue data from national monitoring for
mecarbam in citrus and information on national tolerances.

EVALUATION FOR ACCEPTABLE DAILY INTAKE

BIOCHEMICAL ASPECTS

Absorption, Distribution, Elimination and Biotransformation

Rat

Based on an interim report, male and female Sprague Dawley rats
(fasted 16 hours) given a single oral dose of 10 mg ¹⁴C-mecarbam/kg
b.w. excreted about 80, 2.8 and 0.2 percent of the administered dose
in urine, faeces and expired air, respectively, within 48 hours of
dosing. The radioactivity recovered in the urine was predominantly
associated with seven major polar metabolites. There was little or no
unchanged ¹⁴C-mecarbam or its potential metabolites, viz.
mercarboxon, diethoate and diethoxon, in the non-hydrolysed urine. A
significant sex difference in the metabolite profiles of ¹⁴C-mecarbam
in urine was not observed (Ward 1983).

^1/ See Annex 2 for FAO and WHO documentation.

TOXICOLOGICAL STUDIES

Special Study on Teratogenicity

Rat

Groups of 24 sexually mature (8-9 weeks old), time-mated rats (SPF,
CrL:COBS CD (SD) BR strain) were intubated with mecarbam (97.5 percent
pure) as a suspension in 0.2 percent Tween 80 and distilled water at
0, 1 or 3 mg/kg b.w./day from days 6 through 19 inclusive (day 0 = day
of positive vaginal smear) of pregnancy. The dosages selected were
based on the results of a preliminary study in pregnant rats.
The dams were sacrificed on day 20 of pregnancy and foetuses were
removed by Caesarean section for examination of external, skeletal and
visceral abnormalities. No mortality occurred. Dams at 3 mg/kg b.w.
exhibited cholinergic signs (salivation and intermittent body tremors)
and growth depression. Erythrocyte cholinesterase activity on day 20,
prior to sacrifice, was inhibited in a dose-dependent pattern at both
1 and 3 mg/kg b.w.

Terminal necropsy of dams revealed no compound-related changes. There
were no significant differences between control and treated groups in
pregnancy rate, mean number of live births, corpora lutea,
implantations, or embryonic deaths (early and late), post-implantation
loss, sex ratio and mean foetal weight. Multiple defects characterized
by for example facio-cranial schisis, ablepharia and domed palate were
similarly observed in 4/17 foetuses from 1/24 litters at 3 mg/kg b.w.
The fact that all four of the malformed foetuses were from a single
litter tends to indicate that these abnormal findings are unlikely to
be compound-related. Incidence of visceral and skeletal abnormalities
were not significantly affected by treatment. Under the conditions of
the experiment there were no indications of teratogenic activity of
mecarbam (Bottomley et al 1983).

Special Studies on Mutagenicity

Mecarbam (purity not specified) was tested for its genetic activity in
in vitro microbial systems (plate assay) in the presence or absence
of a mammalian metabolic activation preparation (S-9 mix from the
liver of rats induced with Aroclor 1254). Indicator organisms used
were Salmonella typhimurium strains TA 1535, TA1537, TA 1538, TA 98
and TA 100. Results indicated no evidence of mutagenicity of the
compound to any of the tester strains at concentrations ranging from
50 to 5 000 µg/plate, with or without the addition of the S-9 mix
(Richold et al 1982a).

The ability of technical mecarbam to induce DNA repair was tested in
cultured human epithelioid cells (HeLa). There was no indication of
unscheduled DNA synthesis in this in vitro test system at
concentrations up to 2 000 ug/ml, a level capable of causing some cell
death, with or without metabolic activation employing S-9 mix from
liver of rats induced with Aroclor 1254 (Richold et al. 1982c).

In a mouse lymphoma L5178Y cell TK locus mutation assay, technical
mecarbam, at concentrations (100 - 160 ug/ml) resulting in cell
survival of approximately 30 percent and above, induced no significant
increase in mutation frequency, with or without the presence of a
metabolic activation system (S-9 mix from liver of rats induced with
Aroclor 1254). At the most toxic concentrations (180 ug/ml and above)
when cell survival was only about 20 percent, a statistically
significant increase in mutation rate was observed in duplicate
experiments with the presence of S-9 mix and in one of the two
experiments without metabolic activation. Since mecarbam only appeared
to induce an increase in mutation frequency when present at highly
toxic dose levels, the compound possessed no significantly relevant
mutagenic potential (Richold et al. 1983a).

In a micronucleus test, groups of five male and five female mice (SPF,
CD-Swiss-derived strain) were intubated with technical mecarbam at a
total dosage of 0, 45, 90 or 180 mg/kg b.w. given as two equal doses
separated by an interval of 24 hours. The animals were sacrificed six
hours after the second dose and the femurs were removed for the
preparation of bone marrow smears. Four males and one female at 180
mg/kg b.w. and one female at 90 mg/kg b.w. died. There was no
significant treatment-related increase in the incidence of
polychromatic erythrocytes containing micronuclei. The ratio of
normochromatic to polychromatic erythrocytes was also unaffected
(Richold et al. 1982b).

Groups of five male and five female rats (SPF Sprague-Dawley CD
strain) were intubated with mecarbam at a total dosage of 0, 12.5, 25
or 50 mg/kg b.w. given as two equal doses separated by an interval of
24 hours. In a preliminary toxicity study, 50 mg/kg b.w. was found to
be a maximum tolerated dose in rats. Four hours after the second dose,
the animals were given i.p. 4 mg/kg b.w. of colchicine and then
sacrificed two hours later. Bone marrow cell preparations from the
animals were evaluated for the presence of chromosome aberrations in
50 metaphase figures per animal. Metaphase analysis showed no
significant difference between treated and control groups in the
incidence of aberrant cells, when gap damage was excluded, although a
non-dose-related increase in the number of chromosomal aberrations was
noted when gap damage was included. Examination of the data revealed a
number of deficiencies in the study. For instance, considerably less
than 50 metaphase figures per animal in the two high-dosage groups,
particularly at the top-dosage level, were analysed, thus leaving a
weak base of data to support any conclusion. In the top-dosage group,
two animals died and a total of only 62 metaphases were found in four
other animals. Notable also was the use of a single sacrifice time
after dosing. It is known that chemicals have different times for the
expression of a maximum effect in in vivo cytogenetic studies. The
seemingly negative response obtained in the study could be due to an
incorrect choice of time to terminate the experiment. Overall, the
study failed to demonstrate conclusively that the compound was not
mutagenic under the conditions of the experiment (Richold et al.
1983b).

Special Study on Neurotoxicity

Hen

Six "young" hens of the Sterling Ranger hybrid (1.48 - 2.3 kg), after
an acclimatization period of at least three weeks, were intubated with
a single dose of technical mecarbam (92 percent w/w active) in maize
oil at 200 mg/kg b.w. on day 1 and again on day 22. The birds were
observed for a total period of up to 44 days. The acute oral LD₅₀ of
mecarbam in this strain of hens was determined to be 200 mg/kg b.w.
prior to initiation of the neurotoxicity study. Hens showing overt
cholinergic signs after each dose on the day of treatment were given
intramuscularly 0.1 ml of atropine sulphate (8.909 percent w/v) and/or
up to 0.98 ml of PAM (5 percent w/v). In spite of the administration
of antidotes, one hen died during the first overnight period and
another was sacrificed on day 7 due to "progressive loss of locomotor
function such that on day 7 the legs were unable to support the body".
A third hen died three hours after the second dose. Two replacement
hens from the original acclimatized stock were similarly treated with
mecarbam six days after the beginning of the study. With the exception
of the bird sacrificed on day 7, all treated hens recovered from the
toxic symptoms, including muscle tremors, salivation, lethargic
movements, hyper noea, ataxia or an inability to stand within 24 hours
after each dose. Histopathological examination of brain (medulla,
pons, cerebellar cortex, optic chiasma, basal ganglia,cerebral cortex
and hippocampus), spinal cord and sciatic nerves from three surviving
hens and three vehicle controls at the end of the observation period
revealed no morphological changes characteristic of delayed neurotoxic
effects.Concurrent positive control hens, treated with TOCP, displayed
clinical signs and histopathological changes of the nervous system
typical of delayed neurotoxicity (Buch et al.1983)

Short-Term Studies

Rat

Groups of five male and five female rats (COBS CD strain) were exposed
dermally to technical mecarbam (undiluted) at 0, 250, 500 or 1000
mg/kg b.w. under an occlusive patch for a period of 6 hours/day 7
days/week for 21 days. There were no mortality and compound-related
toxic signs. Growth depression and reduced food consumption were noted
in males at 1000 mg/kg b.w. Terminal haematological and blood
chemistry studies showed decreased haemoglobin concentration,
erythrocyte counts, haematocrit values and erythrocyte cholinesterase
activity in females at both 500 and 1000 mg/kg b.w. Females of the
top-dosage group also had elevated blood urea nitrogen values.
Absolute weight and organ/body weight ratio of the liver were
increased in females at 500 mg/kg b.w. and above. Terminal necropsy of
all animals in the study and histopathological examination of a number
of selected tissues, including the skin at the application site, from
animals of control and top-dosage groups revealed no significant
findings that might be related to treatment (Woolley et al. 1983).

Comments

The 1980 Meeting estimated a temporary ADI with a requirement for
metabolic studies, a teratogenicity study, mutagenic studies and a
delayed neurotoxicity study.

An interim report of a study with ¹⁴C-labelled mecarbam indicated
rapid absorption, complete metabolism and excretion mainly via urine.
Over 80% excretion of the administered dose occurred within 48 hours,
the major excretory products comprising seven polar metabolites.

There was no evidence of teratogenic activity in rats dosed at 3
mg/kg. Mutagenicity studies, including in vitro microbial assays,
unscheduled DNA synthesis in cultured human epithelial cells and a
micronucleus test in mice, were negative. However, a mouse lymphoma
test, at highly toxic dose levels resulting in about 20 percent cell
survival, seemed to elicit a positive mutagenic response. An in vivo
cytogenic test in rats was inconclusive under the conditions of the
experiment. A delayed neurotoxicity study in young adult hens was
seemingly negative. However, histopathological examination of nerve
tissue was confined to only three of the six surviving hens. Moreover,
the dosage level used (equivalent to an oral LD₅₀) is considered
insufficient for a delayed neurotoxicity study.

Since the only metabolic studies available (presented as an interim
report) were carried out on the rat and the delayed neurotoxicity
study was not acceptable, the Meeting could only extend the temporary
ADI estimated in 1980.

TOXICOLOGICAL EVALUATION

Level Causing no Toxicological Effect

Rat: 5 ppm in the diet, equivalent to 0.21 mg/kg b.w.

Dog: 5 ppm in the diet, equivalent to 0.15 mg/kg b.w.

Estimate of Temporary Acceptable Daily Intake for Man

0-0.001 mg/kg b.w.

FURTHER WORK OR INFORMATION

Required (by 1985)

1. An adequate delayed neurotoxicity study in hens.

2. A full report of the metabolic studies in rats.

3. Complete metabolic studies in laboratory animals other than the
rat.

Desirable

Observations in humans.

REFERENCES - TOXICOLOGY

Bottomley, A.M., Mayfield, R. Effect of technical mecarbam &
Clark, R. on pregnancy of the rat. Report
1983 from Huntingdon Research Centre
submitted to WHO by Dow Chemical
(Netherlands) B.V. (Unpublished)

Buch, S.A., Gardner, J.R., Mecarbam: delayed neurotoxicity
Whitney, J.C. & study in the hen. Report from Life
Cavanagh, J.B. Science Research submitted to WHO
1983 by Dow Chemical (Netherlands) B.V.
(Unpublished)

Richold, M., Jones, E. & Ames metabolic activation test to
Hales, J.F. assess the potential mutagenic
1982a effect of mecarbam. Report from
Huntingdon Research Centre
submitted to WHO by Dow Chemical
(Netherlands) B.V. (Unpublished)

Richold, M., Richardson, J. C, & Micronucleus test on mecarbam.
Howell, A. Report from Huntingdon Research
1982b Centre submitted to WHO by Dow
Chemical (Netherlands) B.V.
(Unpublished)

Richold, M., Allen, J.A. & Autoradiographic assessment of DNA
Proudlock, R.J. repair in mammalian cells after
exposure to mecarbam. Report from
Huntingdon Research Centre
submitted to WHO by Dow Chemical
(Netherlands) B.V. (Unpublished)

Richold, M., Edgar, H.D., An assessment of the mutagenic
Ramsome. S.J. & Banks, S.J. potential of mecarbam using an
1983a in vitro mammalian cell test
system. Report from Huntingdon
Research Centre submitted to WHO
by Dow Chemical (Netherlands) B.V.
(Unpublished)

Richold, M., Allen, J.A., Metaphase analysis on mecarbam.
Richardson, J.C., Report from Huntingdon Research
Proudlock, R.J. & Morgan, N. Centre submitted to WHO by Dow
Chemical (Netherlands) B.V.
(Unpublished)

Ward, C. (¹⁴C)-Mecarbam: metabolic fate in
1983 the rat. Interim report from
Hazleton Laboratories Europe Ltd.
submitted to WHO by Dow Chemical
(Netherlands) B.V. (Unpublished)

Woolley, A.P.A.H., Hill, R.E. & Mecarbam. 21-day repeat dose
Wood, C.M. dermal toxicity study in the rat.
1983 Report from Huntingdon Research
Centre submitted to WHO by Dow
Chemical (Netherlands) B.V.
(Unpublished)

RESIDUES

RESIDUES IN FOOD AND THEIR EVALUATION

USE PATTERN

Information was provided (Ponena Chemicals 1983) on the registered use
of mecarbam on citrus to control red scale in the Republic of South
Africa. Even though mecarbam has been used for this purpose in South
Africa for 10 years, it is said not to be a "popular" control measure.
It is used as a 900 g/litre EC formulation at a rate of 55 ml/100 1.
water (0.05 percent a.i.) during the period of 80-100 percent petal
drop to eight weeks thereafter. A 100-litre spray mixture/tree and 200
trees/ha equals approximately 10 kg a.i./ha. A 200 day
last-treatment-to-harvest interval is imposed. This use is comparable
to high volume uses reviewed by the 1980 Meeting, although the
estimate for maximum residue limit (MRL) is based on a 14-day
pre-harvest interval, which appears to be the most common one.

RESIDUES RESULTING FROM SUPERVISED TRIALS

Residue data were available from supervised trials in the Republic of
South Africa for single mecarbam applications to citrus (Ponena
Chemicals 1983) according to approved usage in that country and for
three applications to grapes, for which it is not registered in that
country (Murphy Chemicals 1983). Also available were residue trials
data from Spain, but approved uses were not available (Dow Chemical
1983a, Dow Chemical 1983b).

Citrus

From a total of 32 samples, residues in South Africa on two varieties
of oranges ranged from 2.9-4.8 mg/kg in peel, <0.01-0.09 mg/kg in
pulp and 0.9-1.4 mg/kg on the whole fruit at intervals ranging from
7-67 days after treatment to <0.01 mg/kg in peel, pulp or whole fruit
220 days after treatment. Maximum residues in 16 grapefruit samples
were 2.4, 0.15 and 0.8 mg/kg in peel, pulp and whole fruit,
respectively, six days after the same 0.05 percent a.i. treatment and
0.5,<0.01 and 0.16 mg/kg, respectively, after 34 days. Residues were

less thereafter. Whole fruit residues in both cases were estimated by
the Meeting from peel and pulp residue, assuming a 70:30 pulp:peel
ratio. No information was provided on the analytical method used
except that residue determination was by gas chromatography utilizing
a flame photometric detector with a phosphorous filter. Residues are
presumed to be for mecarbam alone.

In residue field trials in Spain, conducted in 1981, both 50 percent
EC and 25 percent WP formulations were applied at a concentration of
0.1 percent a.i. to two varieties of lemons (Dow Chemical 1983a).
Although no good agricultural practice information is available for
Spain, the application rates are similar to those of North African and
Mediterranean countries, where pre-harvest intervals for citrus are
either unknown or typically 14 days. Whole fruit residues range from
0.05-0.67 mg/kg (0.32 ± 0.25 mg/kg mean) for the six samples 29-35
days after treatment and 0.06-0.69 mg/kg (0.32 ± 0.26 mg/kg mean) for
another six samples after 45-51 days. The relatively large variability
is due to 0.05 and 0.06 mg/kg residue levels from one of the four
sites. The analytical method is said to be similar to that previously
developed for oranges. Apparent residues in untreated controls are
0.31 mg/kg in two of the 12 samples analysed and 0.01 mg/kg in the
remainder. Both formulations resulted in similar residues at similar
intervals and the data confirm the relative persistence of this
compound.

When flesh and peel from the four trials in Spain were analysed for
mecarbam and and its metabolites diethoxon, diethoate and mecarboxon
29-30 days after treatment, no residues (<0.01 mg/kg were found in
the flesh (Dow Chemical 1983b). Peel residues were 0.5-1.7 mg/kg
mecarbam, <0.01 mg/kg diethoate and 0.1-0.22 mg/kg mecarboxon.
Diethoxon could not be determined, owing to interference by co-eluting
natural compounds. Calculations of whole fruit residues, based on
those in peel and flesh, gave results comparable to whole fruit
analyses.

Grapes

A 900 a.i./1 E.C. mecarbam formulation was applied in South Africa
three times to grapes at a spray concentration of either 0.045 or
0.068 percent. Residues (mg/kg) at the two application rates are shown
in Table 1.

No information was provided on the analytical procedure other than the
fact that a gas chromatographic detector was utilised. Residues are
presumed to be mecarbam alone and are obviously directly related to
dosage.

EVIDENCE OF RESIDUES IN FOOD IN COMMERCE OR AT CONSUMPTION

Information was available on mecarbam residues on citrus imported into
Sweden for the period 1/1/81 to 4/30/83 (Sweden 1983) (Table 2)

Table 1. Mecarbam residues in Grapes - South Africa

Interval after
Application rate last application Residue
(%) (days) (mg/kg)

0.045 19 0.04
0.05
0.068 0.3
0.3
0.045 26 0.04
0.05
0.068 0.14
0.16
0.045 33 0.02
0.03
0.068 0.04
0.06
0.045 40 0.02
0.01
0.068 0.04
0.03

Table 2. Mecarbam Residues in Imported Citrus - Sweden

Food Origin Number of Number of samples with residues within Maximum
samples given ranges (mg/kg) residue
analysed <0.41 0.41-1.03 1.03-2.05 (mg/kg)

Grapefruit Import 148 146 2 0.42

Lemon Import 164 163 1 0.55

Mandarin Import 292 290 1 1.8

NATIONAL MAXIMUM RESIDUE LIMITS

The following maximum residue limits (MRLs) were reported from Sweden
and South Africa.

MRL (mg/kg)

Commodity Sweden Republic of South Africa

Citrus 2 0.5

APPRAISAL

The Meeting reviewed additional good agricultural practice information
for citrus from one country, additional residue data for citrus
reflecting those practices, residue trials data for lemons from
another country, residue data for grapes and citrus import monitoring
data from another country. The good agricultural practice information
for citrus is comparable to that previously reviewed by the Meeting,
except that the pre-harvest interval is much longer.

These citrus data generally support the 2 mg/kg limit previously
estimated for oranges and, along with data previously reviewed,
indicate that the limit should also be suitable for the food group
"citrus". Swedish import monitoring data also give added assurance
that the limit is adequate for oranges, grapefruit and lemons and
demonstrates the need for a limit at that level for this relatively
persistent pesticide in citrus. The data also confirm that citrus
residues are mostly in the peel and demonstrate that up to 30 percent
of the terminal residue in peel is mercarboxon.

In the absence of information on approved or registered mecarbam uses
on grapes, as well as the relatively limited amount of residue data or
information on the analytical method used, the available data do not
support a limit for grapes.

The Meeting concluded that additional information is needed on the
fate of residues in ruminants and that a ruminant metabolism study
should be conducted. Depending on the results of that study, an
additional ruminant feeding study may be required.

RECOMMENDATIONS

The Meeting examined additional mecarbam residue data from supervised
trials on citrus which reflect established good agricultural practice.
From these data, the Meeting concluded that the previously estimated
temporary limit for oranges is confirmed and can be extended to other
citrus fruits when good agricultural practices and the reported
intervals between last application and harvest are observed. The limit
refers to mecarbam alone.

Interval between
Estimated last application
Commodity MRL (mg/kg) and harvest (days)

Citrus 2 14

FURTHER WORK OR INFORMATION

Required (by 1986)

1. A ruminant metabolism study.

2. If the metabolism study indicates any possibility of the presence
of significant residues in animal tissues or milk, other than those
identified in plants, data should be provided on such residues
occurring in meat and milk from feeding ruminants with a diet
containing residues found in treated citrus.

Desirable

Additional information on nationally registered or approved uses on
other commodities (especially olives, olive oil, fruits and
vegetables) and residue data from field trials that reflect those
uses.

REFERENCES - RESIDUES

Dow Chemical. Determination of mecarbam residues in lemon samples
1983a 1981 from trials in Spain, Data submitted to FAO by Dow
Chemical Co. Ltd. (Unpublished)

Dow Chemical. Determination of mecarbam and related metabolite
1983b residues in lemons. Information provided to FAO by
Dow Chemical Co. Ltd.

Murphy Chemicals. Data on mecarbam trials in grapes, Republic of
1983 South Africa. Submitted to FAO by Murphy
Chemicals. (Unpublished)

Ponena Chemicals. Data on mecarbam trials in citrus, Republic of
1983 South Africa. Submitted to FAO by Ponena
Chemicals. (Unpublished)

Sweden. Mecarbam residues in imported citrus. Information
1983 submitted to FAO by the Government of Sweden.

    See Also:
       Toxicological Abbreviations
       Mecarbam (ICSC)
       Mecarbam (Pesticide residues in food: 1980 evaluations)
       Mecarbam (Pesticide residues in food: 1986 evaluations Part II Toxicology)