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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study performed to GLP and guideline.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
EPA OPP 82-1 (90-Day Oral Toxicity)
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Dimethylhydantoin
-Physical state: white crystals
-Analytical purity: >99.5%
-Lot/batch No.: F9431299
-Storage condition of test material: room temperature

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
Test animals
Source: Charles River Breeding Laboratories, Kingston, New York
Age at study initiation: 7 weeks old
Weight at study initiation: Male: 204.5 to 270.3 g - Female: 161.7 to 214.1 g
Housing: individually housed during experiment. Group housed (2-3 per cage) during acclimation period.
Diet: ad libitum
Water: ad libitum
Acclimation period: 14 days

Environmental conditions:
Temperature (°C): 68-76 °C.
Humidity (%): 40-70 %
Photoperiod (hrs dark / hrs light): 12 hours dark and 12 hours light

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The samples were diluted and analysed by HPLC. Analyses of samples at the 1% and 10% nominal concentration levels demonstrated homogeneity. Concentration verififcation analyses were within 9 % of the target value.

HPLC parameters:
Column: Alltech Absorbosphere C8, 25cm x 4.6 mm, 10 micron particle size.
Mobile phase: 70% water / 30% acetonitrile
Detector: waters Associates UV 450 Variable Wavelength/LDC SM4000 Multi-wavelength UV
Pump: Waters Corporation 510
Integrator: Hewlett-Packard HP3396A/Waters Associates Baseline 810 Computer system
Autosampler: waters corporation WISP 710B
Injection size: 5 microliters / 10 microliters
Chart speed 0.5 / 1.0 cm/min
Flow rate: 0.8 ml/min isocratic
Attenuation: 2^10
Duration of treatment / exposure:
90 days (13 weeks)
Frequency of treatment:
5 days per week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
100 mg/kg
Basis:
nominal in water
Remarks:
Doses / Concentrations:
300 mg/kg
Basis:
nominal in water
Remarks:
Doses / Concentrations:
1000 mg/kg
Basis:
nominal in water
No. of animals per sex per dose:
15/sex/dose
Control animals:
yes, concurrent vehicle

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
Time schedule: once daily

DETAILED CLINICAL OBSERVATIONS: Yes
Time schedule: weekly

BODY WEIGHT: Yes
Time schedule for examinations: weekly

WATER CONSUMPTION: No

FOOD CONSUMPTION: Yes
Time schedule for examinations: weekly

HAEMATOLOGY: Yes
Time schedule for collection of blood: end of study
How many animals: first ten surviving animals/sex/group examined at necropsy
Parameters checked: Haematocrit, haemoglobin concentration, erythrocyte count, total and differential leukocyte count, platelet count

CLINICAL CHEMISTRY: Yes
Time schedule for collection of blood: end of study
How many animals: first ten surviving animals/sex/group examined at necropsy
Parameters checked: sodium, potassium, chloride, glucose, calcium, globulin (calculated), blood urea nitrogen, total bilirubin, creatinine, total protein, albumin, alanine aminotransferase, aspartate aminotransferase, phosphorous, alkaline phosphatase, gamma glutamyl transferase, A/G ratio (calculated)

URINALYSIS: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Prior to study initiation and during final week of study

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY:
Performed on all animals. The necropsy included: brain, spinal cord, pituitary, thyroid, parathyroid, thymus, oesophagus, salivary glands, stomach, sternum with marrow, small and large intestines, liver, pancreas, kidneys, adrenals, spleen, heart, trachea, lungs, aorta, uterus, mammary glands, prostate, urinary bladder, lymph nodes, skin, eye.

HISTOPATHOLOGY:
Histopathology performed on first 10 surviving animals/sex/dose level.
Statistics:
Bartletts test was used to determine if dose groups had equal variance.
Parametric procedures used one way ANOVA using F distribution If significant differences were seen Dunnett’s test was used to determine which treatment groups differed significantly from the control.
For non parametric procedures test of equality of means was performed using Kruskal-Wallis test. If significant differences were seen Dunn’s Summed Rank test was used to determine which treatment groups differed significantly from the control.
In addition Jonckheere’s test for monotonic trend in the dose response was performed.

Results and discussion

Results of examinations

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):
effects observed, treatment-related
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:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS
Sporadic evidence of alopecia, missing/broken/maloccluded incisors and red/clear ocular discharge.

MORTALITY
One male in the control group was euthanized on day 41 because of a broken snout. One female in the 300 mg/kg dose group was found dead on day 74 due to miss-dosing.

BODY WEIGHT GAIN
Decreased body weight gain in male rats in the high dose group.
Increased body weight in females in all treatment groups.

FOOD CONSUMPTION AND COMPOUND INTAKE
Reduction in food consumption in the high dose male rats.

OPHTHALMOSCOPIC EXAMINATION
No effects

HAEMATOLOGY
No effects

CLINICAL CHEMISTRY
No effects

URINALYSIS
Not conducted

NEUROBEHAVIOUR
Not examined

ORGAN WEIGHTS
The mean absolute and relative liver weights for male rats in the high dose group were decreased. The mean absolute and relative liver weigh values for females were increased in the high dose group. The mean absolute kidney weights were also increased in the high dose females but was considered to be a reflection of the increased body weight.

GROSS PATHOLOGY AND HISTOPATHOLOGY (NON-NEOPLASTIC)
Abnormalities noted at very low incidences included entire GI tract distended with gas, emaciation, smaller than normal seminal vesicles, clear yellow liquid in stomach, distended uterus, larger than normal ovaries, liver mass, liver thickening, lung discoloration and distended cecum. None of these appeared to be related to the treatment.

Effect levels

Dose descriptor:
NOAEL
Effect level:
> 1 000 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: Based upon the observation of no-effects related to treatment on any studied parameters.

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
The results of this thirteen week oral gavage study indicate that dimethylhydantoin has little or no potential to produce toxic effects when administered to rats at dosages as high as 1000 mg/kg/day. The NOAEL is therefore determined to be > 1000 mg/kg.