Potassium carbamoylcarbamate

Repeated dose toxicity: oral

Administrative data

Endpoint:

chronic toxicity: oral

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Published U.S. National Cancer Institute screening study for chronic toxicity and carcinogenicity: 12-month exposure with 4-month recovery period; however, limited toxicological investigations.

Justification for type of information:

Data of reference deemed to adequately apply to Potassium carbamoylcarbamate.

Cross-reference

Reason / purpose for cross-reference:

read-across source

Reference

Endpoint:

chronic toxicity: oral

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Published U.S. National Cancer Institute screening study for chronic toxicity and carcinogenicity: 12-month exposure with 4-month recovery period; however, limited toxicological investigations.

Justification for type of information:

Data of reference deemed to adequately apply to Potassium carbamoylcarbamate.

Reason / purpose for cross-reference:

read-across source

Principles of method if other than guideline:

12-month screening carcinogenesis/chronic toxicity study followed by a 4-month recovery period with limited assessment of toxicological endpoints.

GLP compliance:

not specified

Limit test:

no

Species:

mouse

Strain:

other: C57BL/6

Sex:

male/female

Details on test animals or test system and environmental conditions:

Animals were obtained from Charles River and randomly assigned to dose groups. Animals were 6 weeks old when assigned to the study and were group housed (5/sex/cage). Food and water were available ad libitum. The mean ambient air temperature was 23 degrees C and a 12-hour light/dark cycle was maintained. Diets were prepared weekly.

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): once each week
- Mixing appropriate amounts with (Type of food): The test substance was initially formulated/prepared as a concentrated hand-blended premix. The mixture and remainder of the stock diet were then thoroughly blended in a Patterson-Kelley stainless steel Twin Shell V-Blender for 20-30 minutes.
- Storage temperature of food: Test diets were stored at 4 degrees C.

Stability studies on the test substance admixed with feed were performed on day 1 and day 14 using a colorimetric urea nitrogen method. The analytical results indicated the test substance was stable over a period of 14 days. Diet mixes older than two weeks were discarded.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

Animals were exposed to urea for 12 months, followed by a 4-month recovery period.

Frequency of treatment:

Continuous (ad libitum)

Remarks:

Doses / Concentrations:
4500, 9000, 45000 ppm
Basis:
nominal in diet

No. of animals per sex per dose:

50/sex

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: no data
- Rationale for animal assignment (if not random): the random casual method was used
- Rationale for selecting satellite groups: no satellite groups were used
- Post-exposure recovery period in satellite groups: not applicable
- Section schedule rationale (if not random): not applicable

Positive control:

Not required.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: Individual bodyweights were recorded pre-test and at study termination. Cage weights were recorded weekly during the study.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

Five animals/sex/group were sacrificed at the end of the 365-day exposure period and a comprehensive list of tissues were investigated histopathologically; interim deaths were similarly investigated. Brain, lung, trachea, heart, thymus, pituitary, thyroids, parathyroids, adrenals, esophagus, stomach, duodenum, jejunum, ileum, colon, liver, spleen, lymph nodes, bone and bone marrow, skin, salivary and mammary glands were collected and fixed in 10% neutral buffered formalin. Tissues were paraffin embedded, cut at 4-6 um, stained with H&E, and examined microscopically to determine presence of neoplastic/preneoplastic changes or signs or any toxic syndrome.

All remaining animals were sacrificed after the 4-month recovery period and investigated histopathologically.

Statistics:

The purpose of the statistical analysis of tumors was to determine whether animals receiving test substance developed a significantly (P<0.05) different proportion of tumors than control animals. Statistical analyses of the incidences of specific types of tumors were made using the Fisher exact test to compare each control group with each group of treated animals at each dose. If more than one dose level of test substance was used, the Cochran-Armitage test for linear trend in proportions with continuity correction was used.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Female mice had a significant occurrence of hematopoietic tumors, namely, malignant lymphomas.

Details on results:

Urea caused a non-dose related increase in the incidence of malignant lymphomas among the mid-dose females. Since no lymphomas occurred in the low dose or high dose groups, the study authors concluded that the lesion was of questionable biological significance.

There were no other signs of toxicity among the limited study endpoints examined; that is, bodyweights and survival were unaffected by treatment. Necropsy did not reveal any effects of treatment (with the exception of the spurious incidence on malignant lymphomas in the mid-dose females, which had no dose-response relationship and was not observed in any other treatment group).

Dose descriptor:

NOAEL

Effect level:

45 000 ppm

Sex:

male/female

Basis for effect level:

other: No dose-related, biologically significant adverse effects were attributable to the test substance up to the highest dose level (4.5% in the diet)

Critical effects observed:

not specified

Based on the limited toxicological endpoints evaluated in this study, there were no dose-dependent, biologically significant carcinogenic or toxic effects at dose levels of up to 45000 ppm.

Conclusions:

In this study with limited evaluation of toxicological endpoints, chronic administration of urea to the mouse produced no dose-dependent, biologically significant chronic or carcinogenic effects that could be clearly attributable to the test substance.

Executive summary:

In a 12-month carcinogenicity/chronic screening assay, C57BL/6 mice (50/sex/group) were exposed to urea in the diet at concentrations of 4500, 9000 or 45000 ppm for 12 months. Five animals/sex/group were sacrificed at the end of the 365-day exposure period and a comprehensive list of tissues was investigated histopathologically; interim deaths were similarly investigated. All remaining animals were sacrificed after the 4-month recovery period and investigated histopathologically. There were no signs of chronic toxicity based on the limited toxicological assessment; survival and bodyweights were unaffected by treatment. Gross and microscopic pathology did not reveal dose-dependent, biologically significant adverse effects (chronic or carcinogenic). This study concluded that urea is non-carcinogenic and of low chronic toxicity.

Data source

Reference

Reference 1

Reference Type:

other: secondary source

Title:

Toxicological Review of Urea: Studies on kidneys of newborn rats after chronic urea administration to the mother

Author:

United States Environmental Protection Agency (U.S. EPA)

Year:

2011

Bibliographic source:

Integrated Risk Information System (IRIS). Available at: http://www.epa.gov/iris/toxreviews/1022tr.pdf

Reference 2

Reference Type:

publication

Title:

Carcinogenesis bioassay of acetamide, hexanamide, adipamide, urea and P-tolylurea in mice and rats

Author:

Fleischman, R.W. Baker, J.R. Hagopian, M. Wade, G.G. Hayden, D.W. Smith, E.R. Weisburger, J.H. Weisburger, E.K.

Year:

1980

Bibliographic source:

Journal of Environmental Pathology and Toxicology 3(5-6): 149-70

Materials and methods

Principles of method if other than guideline:

12-month screening carcinogenesis/chronic toxicity study followed by a 4-month recovery period with limited assessment of toxicological endpoints.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Urea

EC Number:

200-315-5

EC Name:

Urea

Cas Number:

57-13-6

Molecular formula:

CH4N2O

IUPAC Name:

urea

Test material form:

not specified

Details on test material:

Urea from Wako Pure Chemical Industries, Ltd

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

Animals were obtained from Charles River and randomly assigned to dose groups. Animals were 6 weeks old when assigned to the study and were group housed (5/sex/cage). Food and water were available ad libitum. The mean ambient air temperature was 23 degrees C and a 12-hour light/dark cycle was maintained. Diets were prepared weekly.

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): once each week
- Mixing appropriate amounts with (Type of food): The test substance was initially formulated/prepared as a concentrated hand-blended premix. The mixture and remainder of the stock diet were then thoroughly blended in a Patterson-Kelley stainless steel Twin Shell V-Blender for 20-30 minutes.
- Storage temperature of food: Test diets were stored at 4 degrees C.

Stability studies on the test substance admixed with feed were performed on day 1 and day 14 using a colorimetric urea nitrogen method. The analytical results indicated the test substance was stable over a period of 14 days. Diet mixes older than two weeks were discarded.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

Animals were exposed to urea for 12 months, followed by a 4-month recovery period.

Frequency of treatment:

Continuous (ad libitum)

Doses / concentrations

Remarks:

Doses / Concentrations:
4500, 9000, 45000 ppm
Basis:
nominal in diet

No. of animals per sex per dose:

50/sex

Details on study design:

- Dose selection rationale: no data
- Rationale for animal assignment (if not random): the random casual method was used
- Rationale for selecting satellite groups: no satellite groups were used
- Post-exposure recovery period in satellite groups: not applicable
- Section schedule rationale (if not random): not applicable

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: Individual bodyweights were recorded pre-test and at study termination. Cage weights were recorded weekly during the study.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

Five animals/sex/group were sacrificed at the end of the 365-day exposure period and a comprehensive list of tissues were investigated histopathologically; interim deaths were similarly investigated. Brain, lung, trachea, heart, thymus, pituitary, thyroids, parathyroids, adrenals, esophagus, stomach, duodenum, jejunum, ileum, colon, liver, spleen, lymph nodes, bone and bone marrow, skin, salivary and mammary glands were collected and fixed in 10% neutral buffered formalin. Tissues were paraffin embedded, cut at 4-6 um, stained with H&E, and examined microscopically to determine presence of neoplastic/preneoplastic changes or signs or any toxic syndrome.

All remaining animals were sacrificed after the 4-month recovery period and investigated histopathologically.

Statistics:

The purpose of the statistical analysis of tumors was to determine whether animals receiving test substance developed a significantly (P<0.05) different proportion of tumors than control animals. Statistical analyses of the incidences of specific types of tumors were made using the Fisher exact test to compare each control group with each group of treated animals at each dose. If more than one dose level of test substance was used, the Cochran-Armitage test for linear trend in proportions with continuity correction was used.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

The mid-dose group of male rats showed decreased survival (89%) relative to diet controls (95%).

Mortality:

mortality observed, treatment-related

Description (incidence):

The mid-dose group of male rats showed decreased survival (89%) relative to diet controls (95%).

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Among the high dosed rats, there was a statistically significant increase in incidence of interstitial cell adenomas of the testes.

Details on results:

The lesion of interstitial cell adenomas of the testes found in the high dose group males may occur in 100% of controls (historically); thus, its biological significance is questionable according to the study authors.
There were no bodyweight effects. The decreased survival of the mid-dose group males had no dose-relationship as survival of all other test groups remained unaffected by treatment.

Effect levels

Dose descriptor:

NOAEL

Effect level:

45 000 ppm

Sex:

male/female

Basis for effect level:

other: No dose-related, biologically significant adverse effects were attributable to the test substance up to the highest dose level (4.5% in the diet)

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Based on the limited toxicological endpoints evaluated in this study, there were no dose-dependent, biologically significant carcinogenic or toxic effects at dose levels of up to 45000 ppm.

Applicant's summary and conclusion

Conclusions:

In this study with limited evaluation of toxicological endpoints, chronic administration of urea to the rat produced no dose-dependent, biologically significant chronic or carcinogenic effects that could be clearly attributable to the test substance.

Executive summary:

In a 12-month carcinogenicity/chronic screening assay, Fischer 344 rats (50/sex/group) were exposed to urea in the diet at concentrations of 4500, 9000 or 45000 ppm for 12 months. Five animals/sex/group were sacrificed at the end of the 365-day exposure period and a comprehensive list of tissues was investigated histopathologically; interim deaths were similarly investigated. All remaining animals were sacrificed after the 4-month recovery period and investigated histopathologically. There were no clear dose-dependent signs of chronic toxicity based on the limited toxicological assessment; bodyweights were unaffected by treatment, while only the mid-dose group of males had reduced survival. Gross and microscopic pathology did not reveal dose-dependent, biologically significant adverse effects (chronic or carcinogenic). This study concluded that urea is non-carcinogenic and of low chronic toxicity.