Registration Dossier

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 2001-08-14 to 2002-03-07
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP and guideline conform study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
: two listed deviations which according to the author had not an impact on the validity or integrity of the study from a scientific or GLP perspective.
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): EXP 3982 (N-2-hydroxyethalurea)
- Physical state: clear colorless liquid
- Analytical purity: aqueous solution containing 57.58% of hydroxyethyl urea
- Lot/batch No.: 84714
- Expiration date of the lot/batch: 2002-01-04

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc, Portage, Michigan
- Age at study initiation: approximately 84 to 90 days of age
- Weight at study initiation: from 200 to 225 g
- Fasting period before study: no
- Housing: individually in suspended stainless steel cages
- Diet (e.g. ad libitum): PMI Certified Rodent Chow® #5002 (Purina Mills, Inc) ad libitum
- Water (e.g. ad libitum): municipal tap water ad libitum
- Acclimation period: three hours on one day to Elizabethan collars

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 26
- Humidity (%): 30 to 70
- Air changes (per hr): 10 to 15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
dermal
Vehicle:
unchanged (no vehicle)
Details on exposure:
TEST SITE
- Area of exposure: no data
- % coverage: approximately 10%
- Time intervals for shavings or clipplings: for shavings - 18 hours, for clipplings - prior to the first application (gestation day 5) and when necessary thereafter

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

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.15 mL/animal (100 mg/kg/day group), 0.49 mL/animal (330 mg/kg/day group), 1.49 mL/ animal (control and 1000 mg/kg/day groups)
- Constant volume or concentration used: no

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes
Analytical verification of doses or concentrations:
no
Details on mating procedure:
- Impregnation procedure: purchased timed pregnant
- Length of cohabitation: three days
Duration of treatment / exposure:
6 h/day
Frequency of treatment:
daily
Duration of test:
from gestation day 6 through gestation day 19
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 100, 330, 1000 mg/kg/day
Basis:
nominal conc.
based on hydroxyethyl urea contained in test substance at a level of 57.58%
No. of animals per sex per dose:
25
Control animals:
other: yes: reverse osmosis deionized (RODI) tap water
Details on study design:
- Dose selection rationale: dosage levels for the study were selected in an attempt to produce graded responses to the test article. The high-dose level was expected to produce toxic effects, but not excessive lethality. The mid-dose level was expected to produce no or minimal observable effects. The low-dose level was expected to produce no observable effects. Dosage levels were selected on the basis of available data from previous studies, including the dose range-finding leratology study in rats.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: on gestation day 0 (from animal supplier) and a minimum of once daily

BODY WEIGHT: Yes
- Time schedule for examinations: on gestation days 0 (from animal supplier), 5, 6, 9, 12, 15, 19 and 20. Body weight changes were reported for the following gestation intervals: 0-5, 5-6, 6-9, 9-12, 12-15, 15-19, 19-20 and 6-19 (summary only).

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: viscera of thoracic, abdominal and pelvic cavities
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: viable fetuses, fetal sex ratios and mean fetal body weights
Fetal examinations:
- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: Yes
Statistics:
Inferential statistical analyses were performed by the SLI Alpha DS-10 computer system. Maternal survival, the incidence of females with total implant loss, the incidence of females with viable fetuses, and fetal malformation and variation data were compared among the groups by R x C Chi-square test, followed by Fisher's Exact Test for group-wise comparisons to the control group, when appropriate. Maternal body weights, body weight gain, food consumption, gravid uterine weights, corpora lutea counts, number of implantation sites, number of live fetuses, and fetal body weights were analyzed by one-way analysis of variance (ANOVA) followed by Dunnett's test for group-wise comparisons to the control group, when appropriate. In addition to mean and standard deviation (SD) calculations, group mean percentages were calculated for the following cesarean section parameters: pre-implantation loss, fetal sex distributions, post-implantation loss, dead fetuses, and early and late resorptions. The percentage data, the number of dead pups, and the number of early and late resorptions were analyzed by Kruskal-Wallis non-parametric ANOVA. When employed, Kruskal-Wallis was used for both the multiple group comparisons and group-wise comparisons to the control group, as appropriate. All calculations and statistical analyses were based on the dam or litter as the experimental unit. All statistical comparisons were two-tailed and utilized a minimum level of alpha at p < 0.05.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
No mortality or clinical signs of toxicity were noted during the study. Mean food consumption of females in the 1000 mg/kg/day group was statistically lower than in controls during gestation days 12-15 and 15-19 and throughout the overall treatment period (gestation days 6-19). However, there were no statistically significant differences in mean body weights or body weight gain between the control and test article-treated groups. Similarly, there were no statistically significant differences in gravid uterine weights, maternal body weight changes or corrected maternal body weight change between the control and test article-treated groups on gestation day 20. Maternal gross necropsy findings were generally unremarkable. There were no statistically significant differences between the control and test article-treated groups in the cesarean section parameters, including the mean number of corpora lutea, implantation sites, viable foetuses, early and late resorptions, pre- and post-implantation loss, foetal sex ratios and mean foetal body weights.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
act. ingr.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
act. ingr.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
There were no statistically significant or toxicologically meaningful differences in the incidence of foetal malformations or developmental variations between the control and test article-treated groups.

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Based on the results, a dosage level of 1000 mg/kg/day (active ingredient) was considered to be the no-observed-adverse-effect level (NOAEL) for maternal toxicity and a dosage level of 1000 mg/kg/day (active ingredient) was considered to be the no-observed-effect level (NOEL) for developmental toxicity in this study following dermal administration of EXP3982 (N-2-hydroxyethylurea) to pregnant rats during the period of major organogenesis.
Executive summary:

In the GLP and OECD guideline conform study, 25 female Sprague-Dawley rats per group were exposed dermally to 0 (RODI water), 100, 330 and 1000 mg/kg bw/day of hydroxyethyl urea (based on test material EXP 3982 (N-2 -hydroxyethylurea)) for approximately six hours daily from gestation day 6 through gestation day 19. Elizabethan collars were placed around the neck of each animal during the six-hour exposure period, the collars were then removed and the test site was wiped clean with gauze moistened with tap water. The dosing sites were clipped free of hair prior to the first application and when necessary thereafter. A collar check was performed a minimum of once daily during the six-hour exposure period. The animals were observed daily for clinical signs of toxicity. The test site was examined for erythema and oedema prior to dosing on gestation days 6, 9, 12, 15 and 19 and prior to scheduled euthanasia on gestation day 20. Individual body weights were recorded on gestation days 0 (from the supplier), 5, 6, 9, 12, 15, 19 and 20. Individual food consumption was recorded on gestation days 6, 9, 12, 15, 19 and 20. All females were euthanised by carbon dioxide inhalation on gestation day 20 and subjected to cesarean section. Foetuses were individually weighed, sexed and examined for external and visceral or skeletal abnormalities.

No mortality or clinical signs of toxicity were noted during the study. Mean food consumption of females in the 1000 mg/kg/day group was statistically lower than controls during gestation days 12-15 and 15-19 and throughout the overall treatment period (gestation days 6-19). However, there were no statistically significant differences in mean body weights or body weight gain between the control and test article-treated groups. Similarly, there were no statistically significant differences in gravid uterine weights, maternal body weight changes or corrected maternal body weight change between the control and test article-treated groups on gestation day 20. Maternal gross necropsy findings were generally unremarkable. There were no statistically significant differences between the control and test article-treated groups in the cesarean section parameters, including the mean number of corpora lutea, implantation sites, viable foetuses, early and late resorptions, pre- and post-implantation loss, foetal sex ratios and mean foetal body

weights. There were no statistically significant or toxicologically meaningful differences in the incidence of foetal malformations or developmental variations between the control and test article-treated groups.

Based on the results of this study, a dosage level of 1000 mg/kg/day (based on hydroxyethyl urea) was considered to be the no-observed-adverse-effect level (NOAEL) for maternal toxicity and developmental toxicity in this study following dermal administration of EXP3982 (N-2-hydroxyethylurea) to pregnant rats during the period of major organogenesis. Bsed on these results hydroxyethyl urea is not classified for developmental toxicity by CLP/GHS criteria as no effects were seen.