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Description of key information

Supporting subacute oral toxicity studies were available in rats either by oral gavage (at 450, 500 and 1500 mg/kg bw) or in the diet (at 0.005, 0.05, and 0.5%, corresponding with 5, 50 and 500 mg/kg bw). Mortality was observed at 500 and 1500 mg/kg bw, whereas no mortality was seen at 450 mg/kg by oral gavage and in the diet study up to 0.5% (500 mg/kg bw). General signs of toxicity to rats are those of methemoglobinemia, such as cyanosis, enlarged dark spleen, and compensatory red blood cell (RBC) formation in the spleen and bone marrow. Dead animals generally show pulmonary edema, congestion, and hemorrhagy accompanied by anemia of liver, kidneys, and spleen. A NOAEL was not defined in the subacute toxicity studies; the dose of 5 mg/kg bw in the rat subacute toxicity study can be considered as Lowest Adverse Effect Level (LOAEL).

A key oral subchronic 13-week toxicity study in rats and supporting oral 13-week toxicity study in mice were available by dietary administration in both species. Effective concentrations in the rat were 0, 900, 1800, 3200, 7200 and 12400 ppm, corresponding with 72, 144, 256, 576 and 992 mg/kg bw. Effective concentrations were 3200, 7000, 12400, 28100 and 49800 ppm in the mouse, corresponding with 640, 1400, 2480, 5620 and 9960 mg/kg bw. Mortality and lymphoid depletion in the spleen, thymus and bone marrow were observed in both species at higher dose levels, whereas body weight changes (rats & mice) or observation in the lungs, stomach and adrenal glands (rats) were noted at lower dosages. As the studies were intended for dose selection of the carcinogenicity studies, A NOAEL was not defined in the subchronic toxicity studies. The dietary concentration of 900 ppm (72 mg/kg bw) in the key rat subchronic toxicity study can be considered as Lowest Adverse Effect Level (LOAEL).

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
1978
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Lot no. D-A330 obtained from the Hopkins Chemical Co. (Madison, WI) in one lot
- Expiration date of the lot/batch: The purity and identity determinations were made three times per year at the Study Laboratory and showed that the chemical maintained its identity and purity throughout the studies.
- Purity test date: Not provided.

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: A heat stability study performed by the analytical chemistry laboratory indicated that the compound was stable in storage. After receiving the study chemical from the analytical chemistry laboratory, the study laboratory stored it at 0° ± 5° C. Periodic characterization of the chemical at EG&G Mason Research Institute by infrared spectroscopy and thin-layer chromatography indicated that no detectable decomposition occurred during the studies.
- Stability under test conditions: Monuron was stable in feed when stored for 2 weeks at 25°C or below.
- Solubility and stability of the test substance in the solvent/vehicle: Monuron was stable in feed when stored for 2 weeks at 25°C or below.
The mixture of monuron in stock rodent feed at the 0.5% (5000 ppm) concentration was more homogenous after 15 minutes than after 10 minutes mixing in a 4-qt Patterson-Kelly Twin-Shell® blender with intensifier bar. The variations in the samples of the 15-minute mixture are within 10% of the target concentration of chemical in the feed.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing:After receiving the study chemical from the analytical chemistry laboratory, the study laboratory stored it at 0° ± 5° C.
- Final preparation of a solid: The study laboratory prepared formulated diets by layering a dry premix between portions of feed and blending the mixture for 15 minutes.
1.Premix: Monuron (7.54 g ± 0.01 g) was added directly to 100 g of Wayne Lab Blox® rodent feed.
This premixture was homogenized in a 1-qt large-mouth glass jar rotated for 15 minutes on a ball-mill type tumbler apparatus, with manual end-over-end tumbling every 5 minutes.
2.Bulk mixing: The above premix and 1400 g more feed were mixed in a Patterson-Kelly Twin-Shell® blender for 15 minutes. The blender was loaded from the top of the shells as follows: 700 g of feed
was poured in and allowed to settle and level at the bottom (vertex of the "V"); then the dried premix was poured in on top of the feed from each side; this layer was covered with the remaining 700 g
of feed poured in from each side. After 10-and 15minute mixing times, duplicate 5-g samples were removed from the top of each shell and the bottom trap of blender for subsequent analysis.
Species:
rat
Strain:
Fischer 344
Remarks:
F344/N
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Industries, Inc. (Indianapolis, IN)
- Age at study initiation (when placed on study): 7-8 wk
- Housing: Polycarbonate cages (Lab Products, Inc., Rochelle Park, NJ) with Nonwoven fiber cage filters (Lab Products, Inc., Rochelle Park, NJ) and Aspen Bed® hardwood chips (American Excelsior
Co., Baltimore, MD) as bedding material; 5 animals per cage
- Diet (e.g. ad libitum): Wayne Lab Blox® (Allied Mills, Inc., Chicago, IL); ad libitum
- Water (e.g. ad libitum): Tap, available ad libitum via automatic watering system (Edstrom Industries, Waterford, WI)
- Acclimation period: 18 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22° C average, range 19°-26° C
- Humidity (%): 23.9% average relative humidty; range 7%-55%
- Air changes (per hr): 10 changes room air/h
- Photoperiod (hrs dark / hrs light): fluorescent light 12 h/d

IN-LIFE DATES:
From: First dosing: 25 September 1978
To: 26 December 1978 – 29 December 1978 (killed at 20-21 wks old)
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): Maximum Storage Time: 14 days.
- Mixing appropriate amounts with (Type of food): Wayne Lab Blox® (Allied Mills, Inc., Chicago, IL).
The study laboratory prepared formulated diets by layering a dry premix between portions of feed and blending the mixture for 15 minutes.
1.Premix: Monuron (7.54 g ± 0.01 g) was added directly to 100 g of Wayne Lab Blox® rodent feed.
This premixture was homogenized in a 1-qt large-mouth glass jar rotated for 15 minutes on a ball-mill type tumbler apparatus, with manual end-over-end tumbling every 5 minutes.
2.Bulk mixing: The above premix and 1400 g more feed were mixed in a Patterson-Kelly Twin-Shell® blender for 15 minutes. The blender was loaded from the top of the shells as follows: 700 g of feed
was poured in and allowed to settle and level at the bottom (vertex of the "V"); then the dried premix was poured in on top of the feed from each side; this layer was covered with the remaining 700 g
of feed poured in from each side. After 10-and 15minute mixing times, duplicate 5-g samples were removed from the top of each shell and the bottom trap of blender for subsequent analysis.
- Storage temperature of food: The mixture was held at 5°C until use and was used within 13 days after being mixed.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical chemistry laboratory demonstrated the homogeneity of a formulated diet mixture. Further studies showed that monuron was stable in feed when stored for 2 weeks at 25°C or below.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
continuous exposure in the food
Dose / conc.:
0 ppm
Remarks:
target concentration
Dose / conc.:
750 ppm
Remarks:
target concentration
Dose / conc.:
1 500 ppm
Remarks:
target concentration
Dose / conc.:
3 000 ppm
Remarks:
target concentration
Dose / conc.:
6 000 ppm
Remarks:
target concentration
Dose / conc.:
12 000 ppm
Remarks:
target concentration
Dose / conc.:
0 ppm
Remarks:
experimental mean concentration
Dose / conc.:
900 ppm
Remarks:
experimental mean concentration
Dose / conc.:
1 800 ppm
Remarks:
experimental mean concentration
Dose / conc.:
3 200 ppm
Remarks:
experimental mean concentration
Dose / conc.:
7 200 ppm
Remarks:
experimental mean concentration
Dose / conc.:
12 400 ppm
Remarks:
experimental mean concentration
No. of animals per sex per dose:
10
Control animals:
yes, concurrent no treatment
Details on study design:
- Rationale for animal assignment: Distributed to groups so that average group weights approximately equal
Observations and examinations performed and frequency:
CLINICAL SIGNS AND MORTALITY
Animals were checked two times per day; moribund animals were killed, and necropsies were performed.

BODY WEIGHT AND WEIGHT GAIN
Individual animal weights were recorded once per week.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Feed consumption was measured once per week. One day before the animals were killed, formulated diets were replaced with control feed.

GROSS PATHOLOGY
At the end of the 13-week studies, survivors were killed. A necropsy was performed on all animals, except those excessively autolyzed or cannibalized.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes. At the end of the 13-week studies, survivors were killed. A necropsy was performed on all animals, except those excessively autolyzed or cannibalized.

HISTOPATHOLOGY: Yes. Histologic exam performed on control, 6000-, and 12000-ppm groups and early death rats: gross lesions and tissue masses, mandibular lymph nodes, mammary gland,
salivary glands, thyroid gland, parathyroids, sternebrae, liver, colon, small intestine, prostate/testes or ovaries/uterus, urinary bladder, lungs and bronchi, heart, esophagus, stomach, brain, thymus, trachea, pancreas, spleen, kidneys, adrenal glands, pituitary gland, spinal cord and eyes.
Partial histologic exam on 3000-ppm group: thymus, bone marrow, spleen, mesenteric and mandibular lymph nodes.
Mortality:
mortality observed, treatment-related
Description (incidence):
8/10 male rats and 9/10 female rats fed diets containing 12000 ppm monuron died during the first 2 weeks of the studies.
5/10 male rats and 6/10 female rats fed diets containing 6000 ppm monuron died during the first 4 weeks of the studies.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The final weights of the rats were inversely related to the concentration of monuron in the feed.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Female rats fed 750-12000 ppm ate less than did the controls.
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Discoloration and mottling of the lungs, smoothing and thinning of the stomach, and enlargement and discoloration of the adrenal glands were noted at necropsy and appeared to be compound related.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
A generalized atrophy of lymphocytic and hematopoietic tissues was observed in male and female rats fed diets containing 12000 ppm monuron. These changes included lymphoid depletion and
congestion of the splenic white pulp, lymphoid depletion with overall reduction in size of the thymus, myeloid depletion of bone marrow, and lymphoid depletion of B-and T-cell areas of lymph nodes
causing a marked reduction in the size of all examined lymph nodes. Similar but less severe changes were seen in male and female rats fed diets containing 6000 ppm monuron; changes seen in the l
ymphocytic and hematopoietic tissues of rats fed monuron at lower concentrations were considered to be slight or equivocal.
Key result
Dose descriptor:
LOAEL
Effect level:
5 mg/kg diet
Critical effects observed:
yes
System:
haematopoietic
Treatment related:
yes
Critical effects observed:
yes
Organ:
kidney
Treatment related:
yes
Critical effects observed:
yes
Organ:
liver
Treatment related:
yes
Conclusions:
In a 13 weeks feeding study, dietary uptake of monuron (>99% pure) of up to 12000 ppm in Fischer 344 rats, the kidney, liver and lympho/haematopoietic systems were targets for toxicity. The lympho/hematopoietic system of rats was the primary site affected. The Iymphocytic and haematopoietic tissue atrophy seen in rats at high doses in the 13-week studies was not seen in the two-year feeding studies.
Because of weight gain depression and histopathologic changes observed at higher dose levels, doses selected for the 2-year studies in rats were 750 and 1500 ppm monuron in feed.
Executive summary:

Thirteen-week studies were conducted to evaluate the toxicity associated with repeated ingestion of monuron and to determine the concentrations to be used in the 2-year studies.

Groups of 10 F344/N rats of each sex were fed diets containing 0, 750, 1500, 3000, 6000, or 12000 ppm monuron. All diets were available ad libitum. Formulated diets were first offered to each group on September 25, 1978. Necropsies were performed between December 26 and December 29, 1978.

Animals were checked two times per day, moribund animals were killed and necropsies were performed. Feed consumption was measured once per week. One day before the animals were killed, formulated diets were replaced with control feed. Individual animal weights were recorded once per week. At the end of the 13 week studies, survivors were killed. A necropsy was performed on all animals, except those excessively autolyzed or cannibalized.

Eight of 10 male rats and 9/10 female rats fed diets containing 12000 ppm monuron died during the first 2 weeks of the studies. Five of 10 male rats and 6/10 female rats fed diets containing 6000 ppm monuron died during the first 4 weeks of the studies. The final weights of the rats were inversely related to the concentration of monuron in the feed. Female rats fed 750-12000 ppm ate less than did the controls.

Discoloration and mottling of the lungs, smoothing and thinning of the stomach, and enlargement and discoloration of the adrenal glands were noted at necropsy and appeared to be compound related.

A generalized atrophy of lymphocytic and hematopoietic tissues was observed in male and female rats fed diets containing 12000 ppm monuron. These changes included lymphoid depletion and congestion of the splenic white pulp, lymphoid depletion with overall reduction in size of the thymus, myeloid depletion of bone marrow, and lymphoid depletion of B-and T-cell areas of lymph nodes causing a marked reduction in the size of all examined lymph nodes. Similar but less severe changes were seen in male and female rats fed diets containing 6000 ppm monuron; changes seen in the lymphocytic and hematopoietic tissues of rats fed monuron at lower concentrations were considered to be slight or equivocal.

Because of weight gain depression and histopathologic changes observed at higher dose levels, doses selected for the 2-year studies in rats were 750 and 1500 ppm monuron in feed.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LOAEL
5 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Reliable toxicity studies published by EPA, NPT, IARC.
System:
haematopoietic
Organ:
bone
bone marrow
kidney
liver
spleen
thymus

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Subacute oral toxicity

In a supporting study, doses of 1500 mg/kg (1/5 Approximate Lethal Dose) were administered by stomach tube to male albino rats, 5 times a week (U.S.EPA, 1975). By the eighth treatment, all 6 animals had died. During treatment the animals showed discomfort, weakness, and continuous weight loss. Autopsy revealed pulmonary edema and congestion as well as damage to liver, kidneys, and spleen. Because of the severe cumulative toxicity at 1500 mg/kg, the subacute dose was reduced to 500 mg/kg, 5 times per week for 2 weeks. Animals showed discomfort after treatment, lost weight continuously, and became cyanotic. During the post-treatment observation period, weight was regained to almost the original level. At 10 days post-treatment, all animals were sacrificed. Spleens were large, dark, and congested with small foci of blood formation in all cases. Deposition of brown granules of blood pigment (hemosiderin) in spleens was indicative of blood destruction. (US EPA, 1975).

In another subacute oral study, male and female rats were exposed to Monuron via the diet at 0.005, 0.05, and 0.5% for 6 weeks (U.S.EPA, 1975). Taking into account a mean body weight of 250 g and daily food consumption of 25 g in growing rats, these dose levels approximated 5, 50 and 500 mg/kg bw. At 0.005%, food consumption and weight gain were comparable to the control group; blood counts were normal, and no clinical signs were observed. After sacrifice, 2 of 5 males showed evidence of methemoglobinemia and/or compensatory RBC formation. No females showed significant pathology. At 0.05%, feed consumption and weight gain for the test group comparable to the controls. No clinical signs were observed and blood counts were within normal ranges . At sacrifice, signs of methemoglobinemia were apparent, and compensatory RBC formation was found in the spleen. The 0.5% group consumed much less food, and consequently gained much less weight than the other test groups or controls. The animals were pale and cyanotic with lowered blood counts . The blood was dark brown and contained many degenerate RBC's. At necropsy methemoglobinemia was apparent as well as possible blood destruction. Compensatory RBC formation was found in the spleen and bone marrow of all animals. No mortality occurred, even at the highest level fed (0.5%).

Finally, a short-term supporting (2-18 weeks) feeding of monuron at 450 mg/kg to rats caused hepatocyte mitochondrial changes associated with altered activity of glycolytic enzymes (IARC, 1991).

A NOAEL was not defined in the subacute toxicity studies; the dose of 5 mg/kg bw in the rat subacute toxicity study can be considered as Lowest Adverse Effect Level (LOAEL), however the study was done in a limited No. of rats.

 

Subchronic oral toxicity

A 13-week key toxicity study was conducted in rats via the diet containing target concentrations of 0, 750, 1500, 3000, 6000, or 12000 ppm and effective concentrations of 0, 900, 1800, 3200, 7200 and 12400 ppm Monuron (NTP, 1988; IARC, 1991). Taking into account a mean body weight of 250 g and daily food consumption of 20 g in adult rats, these dose levels approximated 72, 144, 256, 576 and 992 mg/kg bw.

8/10 male rats and 9/10 female rats fed diets containing 12000 ppm monuron died during the first 2 weeks of the studies. 5/10 male rats and 6/10 female rats fed diets containing 6000 ppm monuron died during the first 4 weeks of the studies. The final weights of the rats were inversely related to the concentration of Monuron in the feed. Female rats fed 750-12000 ppm ate less than did the controls. Discoloration and mottling of the lungs, smoothing and thinning of the stomach, and enlargement and discoloration of the adrenal glands were noted at necropsy and appeared to be compound related. A generalized atrophy of lymphocytic and hematopoietic tissues was observed in male and female rats fed diets containing 12000 ppm. These changes included lymphoid depletion and congestion of the splenic white pulp, lymphoid depletion with overall reduction in size of the thymus, myeloid depletion of bone marrow, and lymphoid depletion of B-and T-cell areas of lymph nodes causing a marked reduction in the size of all examined lymph nodes. Similar but less severe changes were seen in male and female rats fed diets containing 6000 ppm; changes seen in the lymphocytic and hematopoietic tissues of rats fed monuron at lower concentrations were considered to be slight or equivocal. Doses selected for the 2-year studies in rats were 750 and 1500 ppm in feed.

A 13-week supporting toxicity study was conducted in mice via the diet containing target concentrations of 3000, 6000, 12000, 25000, or 50000 ppm and effective concentrations of 3200, 7000, 12400, 28100 and 49800 ppm (NTP, 1988; IARC, 1991). Taking into account a mean body weight of 25 g and daily food consumption of 5 g in adult mice, these dose levels approximated 640, 1400, 2480, 5620 and 9960 mg/kg bw. 5/10 male and 8/10 female mice that received diets containing 49800 ppm monuron and 5/10 females that received 25000 ppm died. Final weights of dosed mice were lower than those of the controls. Female mice at the higher dose levels had a moderate initial loss of body weight. Apparent increases in feed consumption by mice in the higher dosed group may have been due to excessive scattering. Lymphoid depletion of the splenic white pulp, myeloid atrophy of bone marrow with marked reduction in stem cells and myeloblasts (approximately half the marrow was made up of band cells), lymphoid depletion of mandibular and mesenteric lymph nodes, and lymphoid depletion of the thymus were observed in males and females fed diets containing 50000 ppm monuron. The incidence and severity of the lesions in the lymphoid tissues and bone marrow were lower in mice fed 25000 ppm. Atrophy of hematopoietic tissues was not observed in the 12000-ppm group. Doses selected for the 2-year studies of mice were 5000 and 10000 ppm monuron in feed.

A NOAEL was not defined in the subchronic toxicity studies. The dietary concentration of 900 ppm (72 mg/kg bw) can be considered as Lowest Adverse Effect Level (LOAEL). Severe effects such as lymphoid depletion were only obseved at higher dosage levels (>100 mg/kg bw).

Inhalation and dermal repeated dose toxicity studies were waived as the oral route is the most appropriate administration route (based on low vapour pressure and low dermal penetration of Monuron).

Justification for classification or non-classification

Based on the key results and according to CLP (No. 1272/2008 of 16 December 2008), Monuron has no serious effects < 100 mg/kg bw and therefore has no obligatory labelling requirement for repeated dosel toxicity.