sodium 4,4-dimethyl-2,5-dioxoimidazolidin-1-ide

Endpoint summary

Description of key information

Repeat oral dose studies, via gavage (90 days) with rats and chronic dietary studies in rats and dogs (104 weeks), on DMH produced no adverse effects in the rat at the highest dose level tested (NOAEL = 1000 mg/kg bw/day). In a 1-year dietary repeat-dose study in the dog, signs of toxicity, decreasing bodyweight, increase in adrenal weight and mild hypertrophy in the adrenal cortex, were observed at the highest dose level tested, however these were not considered to be of an adverse nature (NOAEL = 40,000 ppm, ie, 1000 mg/kg/d, NOEL = 12,000 ppm, ie, 300 mg/kg/d). 
A repeat dose (90 day) dermal study was performed in the rat at dose concentration of 39, 130 and 390 mg/kg/day. The highest dose tested was limited by solubility. No systemic toxicity or dermal irritation was observed. 

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

repeated dose toxicity: oral

Remarks:

combined repeated dose and carcinogenicity

Type of information:

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

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed to GLP and guideline.

Reason / purpose for cross-reference:

reference to same study

Qualifier:

equivalent or similar to guideline

Guideline:

other: EPA 83-5

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Testing lab
- Age at study initiation: 8 weeks at start of dosing
- Weight at study initiation: Males: 224-282 g - Females: 147-198 g
- Housing: stainless steel cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 3 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 66-77
- Humidity (%): 40-70
- Air changes (per hr): 10 room air changes per hour.
- Photoperiod: 12 hours per day with fluorescent lightning.

Route of administration:

oral: feed

Vehicle:

other: Basal diet (Ground Purina Certified rodent Chow)

Details on oral exposure:

DIET PREPARATION
- Rate of preparation of diet: Fresh diet was prepared and offered to the animals each week.
- Mixing appropriate amounts with (Type of food): A concentrated premix was prepared by direct addition of DMH to ground rodent feed, milled in a ball mill for 1 hour and then mixed for 1 hour in a Hobart mixer to ensure homogenous distribution of the test substance in the diet. The test diets were prepared by appropriate dilutions of the concentrated premix or higher diet concentration and were mixed in a Hobart mixer for 15 minutes. Dietary concnetrations were not adjusted for percent active ingredeint of the test substance, as received was >99%.
- Storage temperature of food: Prepared diets were stored in closed polyethylene containers at room temperature until fed to the animals.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

DMH was analyzed in the rodent diet using HPLC.
The dosages obtained in terms of test substance consumed for the 100, 300, and 1000 mg/kg/day groups ranged from 88.9 to 112.2, 265.3 to 342.5, and 896.9 to 1132.4 mg/kg/day for the males and 90.9 to 117.6, 269.0 to 347.7, and 908.8 to 1147.9 mg/kg/day for the females respectively.

Duration of treatment / exposure:

104 weeks

Frequency of treatment:

7 days per week

Remarks:

Doses / Concentrations:
100 mg/kg/day
Basis:
nominal in diet

Remarks:

Doses / Concentrations:
300 mg/kg/day
Basis:
nominal in diet

Remarks:

Doses / Concentrations:
1000 mg/kg/day
Basis:
nominal in diet

No. of animals per sex per dose:

60/sex/dose level

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: Prior to initiation of the chronic study, the potential for DMH to produce toxicity in rats was evaluated in a 14-day dietary dose range-finding study. In the range finding study rats were exposed to DMH in the diet at concentrations of 0, 7000, 14000 or 20000 ppm for 14 days. These dietary concentrations corresponded to approximate mean intake levels of 571, 1157 and 1663 mg/kg/day for males and 596, 1160, and 1717 mg/kg/day for females respectively. Measured parameters included food consumption, body weights and body weight gains and gross pathology. There were no treatmnet related effects observed and DMH was considered to be palatable and not toxic to rats when administered in the diet at concentrations as high as 20000 ppm. Based on this information, the limit dose of 1000 mg/kg was selected as the target dosage level for the high dose group for the chronic study.

Observations and examinations performed and frequency:

CLINICAL OBSERVATIONS: Yes
- Time schedule: clinical signs were checked twice daily, detailed clinical observations were performed once each week.

MORTALITY: Yes
- Time schedule: Twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly for the first 14 weeks of the study and every other week thereafter.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: weekly for the first 14 weeks of the study and every other week thereafter.

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: prior to first exposure and following study period of 104 weeks.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at 6, 12, 18 and 24 months
- How many animals: 15/sex/group
- Parameters examined: Haematocrit, haemoglobin concentration, erythrocyte count, total and differential leukocyte count, platelet count, reticulocyte count, mean corpuscular volume, mean corpuscular haemogoblin, mean corpuscular haemoglobin concentration.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at 6, 12, 18 and 24 months
- How many animals: 15/sex/group
- Parameters examined: sodium, potassium, chloride, calcium, phosphorus, glucose (fasting), total cholesterol, urea nitrogen, total bilirubin, direct bilirubin, indirect bilirubin, creatinine, total protein, albumin, globulin, A/G ratio, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, gamma glutamyl transpeptidase, creatine kinase.

URINALYSIS: Yes
- Time schedule for collection of urine: at 6, 12, 18 and 24 months
- How many animals: 15/sex/group
- Parameters examined: colour and appearance, volume, specific gravity, pH, protein, glucose, ketones, bilirubin, blood, urobilinogen, microscopic elements.

Sacrifice and pathology:

GROSS PATHOLOGY AND HISTOPATHOLOGY: Yes
- On all surviving animals
- Performed on high dose group and control group
Organs: brain, spinal cord, pituitary, thyroid, parathyroid, thymus, oesophagus, salivary glands, stomach, small and large intestines, liver, pancreas, kidneys, adrenals, spleen, heart, trachea, lungs, aorta, uterus, vagina, mammary gland, prostate, testes, epididymis, seminal vesicles, urinary bladder, lymph nodes, sciatic nerve, sternum, femur, skin, skeletal muscle (thigh), eyes.
- Performed on low and intermediate dose groups: Kidneys, pituitary gland and all gross lesions and mammary glands of female rats.

Other examinations:

Organ weights: Yes
from all surviving animals
Liver, kidneys, adrenals, testes, ovaries, spleen, brain, heart.

Statistics:

Levene’s test for equality of variances, analysis of variance (ANOVA) and tests (when F value from ANOVA was significant).
Non-parametric data were evaluated using Kruskal-Wallis test followed by the Mann-Whitney U test when appropriate.
Two untreated controls were included in the study in order to collect data that would provide information regarding the range of normal or control values for the parameters evaluated in the study. These data were used as an aid in the identification of false positive statistical citations as well as confirmation of true treatment-related effects. Based on the independent manner in which the animals were handled and the data collected, it was not considered appropriate to combine the data from the two control groups for the purposes of comparing the combined control data to those from the treated groups.

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

effects observed, treatment-related

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

no effects observed

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS
The number of animals with urine stains tended to be increased in the high dose males and all dose groups of females. The number of females observed as unkept was increased in all dose group female rats which generally paralleled the number of animals that die. Therefore the higher incidence of these findings was not considered to indicate an effect of the test substance on the overall well being of the animals.

MORTALITY
The mean survival time for rats from the 1000mg/kg/day group (626 days for males and 647 days for females) was lower than the control groups (668 to 659 days for males and 669 to 703 days for females). This difference was only statistically significant for males.

BODY WEIGHT GAIN
Although generally not statistically significant slight decreases (~5 to 10%) in mean absolute body weight of animals in the high dose group of both sexes were observed during the last 2 to 3 months of the study.

FOOD CONSUMPTION AND COMPOUND INTAKE
No effects.

OPHTHALMOSCOPIC EXAMINATION
The incidence of several findings increased with age but were not considered to be effects of the test substance.

HAEMATOLOGY
No effects

CLINICAL CHEMISTRY
No effects

URINALYSIS
No effects
GROSS PATHOLOGY No effects
ORGAN WEIGHTS
No effects

HISTOPATHOLOGY: NEOPLASTIC
The incidences of mammary gland tumors (adenomas, fibroadenomas and carcinomas) was slightly increased in the high dose group of female rats although these were not statistically significant therefore the slight increase was attributed to random biologic variation.

Dose descriptor:

NOAEL

Effect level:

300 mg/kg diet

Sex:

male/female

Basis for effect level:

other: Based upon the equivocal decreases in body weight and survival for animals in the high dose group.

Critical effects observed:

not specified

Conclusions:

Based upon the equivocal decreases in body weight and survival for animals in the high dose group, the No-Observable-Effect Level (NOEL) for systemic toxicity is considered to be at least 300 mg/kg/day but may be as high as 1000 mg/kg/day. The NOAEL is therefore 300 mg/kg/day.

Executive summary:

This study has been performed on DMH (Dimethyl Hydantoin) and has been used for read-across purposes.

Based upon the equivocal decreases in body weight and survival for animals in the high dose group, the No-Observable-Effect Level (NOEL) for systemic toxicity is considered to be at least 300 mg/kg/day but may be as high as 1000 mg/kg/day. The NOAEL is therefore 300 mg/kg/day.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

300 mg/kg bw/day

Study duration:

chronic

Species:

rat

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Reference

Endpoint:

repeated dose toxicity: inhalation

Data waiving:

other justification

Justification for data waiving:

other:

Critical effects observed:

not specified

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

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: dermal

Type of information:

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

Adequacy of study:

key study

Study period:

in year 1994

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed to GLP and guideline

Qualifier:

according to guideline

Guideline:

EPA OPP 82-3 (Subchronic Dermal Toxicity 90 Days)

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

other: CD

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Testing lab
- Age at study initiation: 32 days old
- Weight at study initiation: Males: 236-313 g - Females: 159-215 g
- Housing: stainless steel cages (1 animal per cage during study)
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 3 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 66-77 °F
- Humidity (%): 40-70%
- Air changes (per hr): 10 room air changes per hour.
- Photoperiod: 12 hours per day of fluorescent light.

Type of coverage:

occlusive

Vehicle:

water

Details on exposure:

TEST SITE
- Area of exposure: Dosing solution was applied directly to the back
- Type of wrap if used: Wrapped with VETRAP® Bandaging Tape (3M Personal Care Products) and secured with Elastikon®
- Time intervals for shavings or clipplings: Ten days prior to initiation of study


REMOVAL OF TEST SUBSTANCE
- Washing (if done): Rinsed with water
- Time after start of exposure: After 6 hours


TEST MATERIAL
- Amount applied: 0, 39,130 and 390 mg/kg/day corresponding to a dose volume 3.0, 0.3, 1.0 and 3.0 mL/kg/day.
- Constant volume or concentration used: yes - a constant concentration of 13% (w/w) DMH was used.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Concentration verification analyses on solutions used for dosing showed analytical values ranging from 99.2% to 106.3% of nominal for the 6 periods of analysis.

Duration of treatment / exposure:

90 days

Frequency of treatment:

6 hours/day; 5 days per week

Remarks:

Doses / Concentrations:
39
Basis:
nominal per unit body weight

Remarks:

Doses / Concentrations:
130
Basis:
nominal per unit body weight

Remarks:

Doses / Concentrations:
390
Basis:
nominal per unit body weight

No. of animals per sex per dose:

15/sex/group

Control animals:

yes

Details on study design:

- Dose selection rationale: Previously a 5-day repeated dermal toxicity study was conducted with DMH to generate information to aid in the selection of dose levels for the 90 day study. Male rats were treated dermally 6 hours/day (occluded) for 5 days with 2 mL/kg of 5 or 10% aqueous solutions of DMH or 25 or 50% aqueous mixtures of DMH. These treatments corresponded to dose levels ranging from 100 to 1000 mg/kg/day. No toxicity or skin irritation was observed. Results of the dose solution preparation indicated that even mixtures of DMH in water could not be prepared above saturating DMH concentrations (~13%).

DMH dose levels for the 90 day study were selected based on the results of the preliminary 5 day study, the maximum aqueous solubility of DMH (13%) and the maximum dose volume which could be adminsitered in the test system (3.0 mL/kg/day).

Observations and examinations performed and frequency:

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice daily

DERMAL IRRITATION (if dermal study): Yes

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION: Yes
- Time schedule for examinations: weekly

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: prior to first exposure and following sixty-third treatment

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at end of study
- How many animals: all animals
- Parameters: Haematocrit, haemoglobin, erythrocyte count, total and differential leukocyte count, platelet count, reticulcyte count, mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC).

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at end of study
- How many animals: all animals
- Parameters examined: sodium, potassium, chloride, calcium, phosphorus, glucose, cholesterol, urea nitrogen, total bilirubin, direct bilirubin, indirect bilirubin, creatinine, total protein, albumin, globulin, A/G ratio, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, gamma glutamyl transpeptidase.

URINALYSIS: No

Sacrifice and pathology:

GROSS PATHOLOGY AND HISTOPATHOLOGY: Yes
High dose group and controls
organs: brain, spinal cord, pituitary, thyroid, parathyroid, thymic region, oesophagus, salivary glands, stomach, small and large intestines, liver, pancreas, kidneys, adrenals, spleen, heart, trachea, lungs, aorta, uterus, vagina, female mammary gland, prostate, testes, seminal vesicles, urinary bladder, lymph nodes sciatic nerve, femur, sternum (including marrow), eyes, exorbital lacrimal gland, thigh musculature, skin (treated and untreated)
* Lungs, liver, kidneys, treated and untreated skin were examined from the low and mid-dose groups. In addition, submandibular lymph nodes were examined from all low and mid dose male rats

Other examinations:

Not specified

Statistics:

Data for quantitative continuous variables were compared for the 3 treatment groups and control group by use of Levene’s test for equality of variances, analysis of variance (ANOVA), and t-tests. The t-tests were used when the F value from the ANOVA was significant. When Levene’s test indicated homogenous variances, and the ANOVA was significant, a pooled t-test was used for pairwise comparisons. When Levene’s test indicated heterogenous variances all groups were compared by an ANOVA for unequal variances followed, when necessary, by a separate variance t-test for pairwise comparisons. Nonparametric data were statistically evaluated using the Kruskal-Wallis test followed by the Mann-Whitney U test when appropriate. Incidence data were compared using Fishers exact test. For all statistical tests, the probability value of <0.05 (two-tailed) was used as the critical level of significance.

Clinical signs:

no effects observed

Dermal irritation:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

not examined

Details on results:

CLINICAL SIGNS
No treatment effects.

MORTALITY One male from the control group was found dead on day 17 and one male from the low dose group was sacrificed moribund on day 39.

BODY WEIGHT GAIN
No treatment related effects.

FOOD CONSUMPTION
No treatment related effects.

OPHTHALMOSCOPIC EXAMINATION
No adverse effects.

HAEMATOLOGY
No treatment related effects.

CLINICAL CHEMISTRY
No treatment related effects

ORGAN WEIGHTS
No treatment related effects

GROSS PATHOLOGY
No treatment related effects.

HISTOPATHOLOGY: NON-NEOPLASTIC
No treatment related effects.

Dose descriptor:

NOEL

Effect level:

390 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: Based upon the observation of no clinical signs of toxicity or effects on any study parameters.

Critical effects observed:

not specified

Conclusions:

No clinical signs of toxicity or effects on any of these parameters were found. Based on the results of this study, repeated dermal exposure to saturated aqueous solutions of DMH does not result in systemic toxicity or skin irritation in the rat. NOEL = 390 mg/kg bw/d.

Executive summary:

This study has been performed on DMH (Dimethyl Hydantoin) and has been used for read-across purposes.

No clinical signs of toxicity or effects on any of these parameters were found. Based on the results of this study, repeated dermal exposure to saturated aqueous solutions of DMH does not result in systemic toxicity or skin irritation in the rat. NOEL = 390 mg/kg bw/d.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

390 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Oral repeated dose toxicity studies carried out on DMH

 

Federici (1991) – reliability 1

90 day oral toxicity study in rats with dimethylhydantoin

Type of study: subchronic

Goldenthal (1995) – reliability 1

One year chronic dietary toxicity study in dogs

Type of study: chronic

 

Hermansky & Benson (1994) – reliability 1

Chronic dietary toxicity/oncogenicity study with 5,5 -dimethylhydantoin (DMH) in rats

Type of study: combined repeated dose and carcinogenicity

 

Salinas (1986) – reliability 1

90 day repeated dose oral toxicity study on EMH and DMH in rats

Type of study: subchronic

Dermal repeated dose toxicity studies carried out on DMH

 

Chun & Loughran (1994) - reliability 1

90 day dermal toxicity study with 5,5 -dimethylhydantoin (DMH) in CD rats

Type if study: subchronic

Conclusion:

The studies carried out on the DMH demonstrated that dimethylhydantoin did not produce any toxic effects.

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Reliable study without restiction. Study performed to GLP and guideline.

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:
Reliable study without restiction. Study performed to GLP and guideline.

Justification for classification or non-classification

Considering that oral and dermal exposure to DMH does not produce any toxic effects no classification is required.

As previously stated these studies have been performed on DMH (Dimethyl Hydantoin) and have been used for read-across purposes. A summary of data for read-across of NaDMH with DMH is given in Section 13.