4,5-dihydroxy-1,3-dimethylimidazolidin-2-one

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Effect on fertility: via oral route

Endpoint conclusion:

no study available

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

No data are available for 4,5-Dihydroxy-1,3-dimethylimidazolidin-2-one.

Effects on developmental toxicity

Description of key information

Key, Prenatal Developmental Toxicity Study, rat, RL1

In the key Prenatal Developmental Toxicity Study in pregnant rats according to OECD 414, oral treatment from gestation day 5 up to day 19 with 4,5-dihydroxy-1,3-dimethylimidazolidin-2-one at the dose levels of 1000, 300 and 1000 mg/kg bw/day did not cause death, clinical signs, macroscopic changes in maternal organs and changes in maternal thyroids. Slightly lower food consumption and body weight gain from G.D. 5 to 8 as well as 14 to 17, as well as slightly lower body weight gain between G.D. 5 and 20 might be attributed to the treatment, however based on the slight manner, not considered as adverse. There was no test item effect proven on the FT3, FT4 and TSH hormone values. The treatment with the test item had no impact on the intrauterine parameters. There was no test item-related effects proven on external, visceral and skeletal examination of fetuses. Based on the observations the NOAEL (maternal toxicity) was 1000 mg/kg bw/day and the NOAEL (developmental toxicity, including teratogenicity) was 1000 mg/kg bw/day.

 

Supporting, read-across, Prenatal Developmental Toxicity Study, rat, RL2

In a supporting study with the analogue substance Dimethyloldihydroxyethylene urea (CAS 1854-26-8) according to OECD guideline 414 and GLP, the test substance was neither maternally nor embryotoxic in rats. There were no signs of toxicity in prenatal development of fetuses. The NOAEL was ≥ 640 mg/kg bw/d (for 100% test substance).

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2021-05-25 to 2021-12-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.31 (Prenatal Developmental Toxicity Study)

Version / remarks:

2008-05-30

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Version / remarks:

1998-08

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Version / remarks:

2018-06-25

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

Han: WIST

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: TOXI-COOP ZRT. 1103 Budapest, Cserkesz u. 90.
- Age at study initiation: Approx. 8 weeks (females), 37-38 weeks (males) at the start of mating
- Weight at study initiation: 172-227 g (first day of gestation)
- Fasting period before study: No
- Housing: 1-2 females per cage (before mating), 1 male and 1-2 females / cage (during mating) and 2 sperm positive females per cage, if not possible 1 sperm positive female per cage (during gestation) in type II polypropylene/polycarbonate cages
- Diet: Ad libitum (ssniff® SM R/M-Z+H "Autoclavable complete feed for rats and mice - breeding and maintenance")
- Water: Ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.1-23.1
- Humidity (%): 44-68
- Air changes (per hr): Above 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From day 5 to day 20 of gestation

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Formulations was prepared in the formulation laboratory of the Test Facility beforehand for not longer than 3 days and stored at room temperature (15 – 25°C) until use. The formulations were considered as stable at room temperature for four days at the concentrations of between 20 and 500 mg/mL according to the method validation study.

VEHICLE
- Concentration in vehicle: 0, 47.2, 141.8 and 472.4 mg/L
- Amount of vehicle: 5 mL/kg bw/day
- Lot/Batch No: 202102004

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analytical control of dosing formulations (verification of concentrations and homogeneity) was performed in the analytical laboratory of the test facility two times during the study. Five aliquots of 10 mL of each formulation to be administered to the animals and five aliquots of 10 mL control item (vehicle) were taken from different places (top, middle and bottom). The mean concentration of formulation samples was in the range of 96.3-98.0 % compared to the corresponding nominal concentration. The formulations were considered as homogenous. The suitability of the chosen vehicle (recovery and stability) for the test item at the intended concentrations was analytically verified up front. 4,5-dihydroxy-1,3-dimethylimidazolidin-2-one was proven to be stable in ultrapure water at the intended concentrations at room temperature for up to 4 days. A separate analytical report provided these data.

Details on mating procedure:

- Impregnation procedure: Cohoused
- M/F ratio per cage: 1 male/1-3 females
- Length of cohabitation: 2-4 hours
- Further matings after two unsuccessful attempts: No
- Verification of same strain and source of both sexes: Yes
- Proof of pregnancy: Presence of vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

From day 5 to day 20 of gestation

Frequency of treatment:

Daily

Duration of test:

15 days

Dose / conc.:

1 000 mg/kg bw/day (actual dose received)

Remarks:

Doses and nominal concentrations determined on the basis of 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one and its organic impurities (42.34 %) contained in the product. Dose of the product: 2360 mg/kg bw/day

Dose / conc.:

300 mg/kg bw/day (actual dose received)

Remarks:

Doses and nominal concentrations determined on the basis of 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one and its organic impurities (42.34 %) contained in the product. Dose of the product: 710 mg/kg bw/day

Dose / conc.:

100 mg/kg bw/day (actual dose received)

Remarks:

Doses and nominal concentrations determined on the basis of 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one and its organic impurities (42.34 %) contained in the product. Dose of the product: 236 mg/kg bw/day

Dose / conc.:

0 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

26 female animals per dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose setting with 0, 236, 710 and 2360 mg/kg bw/day was based on the results of a dose range finding study and the results of a 90-day repeated dose toxicity study with 4,5-dihydroxy-1,3-dimethylimidazolidin-2-one in rats (see IUCLID section 7.5). 4,5-dihydroxy-1,3-dimethylimidazolidin-2-one did not cause adverse effects in male or female rat, Han: WIST of Wistar origin rats after the consecutive 14-day or 90-day oral (by gavage) administration at 236, 710 or 2360 mg/kg bw/day. Thus, in this study, the limit dose of 1000 mg/kg bw/day defined in OECD guideline 414 was chosen as the highest dose (corresponding to 2360 mg/kg bw/day product concentration).
- Rationale for animal assignment: The sperm positive females were allocated to each experimental group on each mating day in such a way that the group averages of the body weight were as similar as possible on the first day of gestation. Females paired with the same male were allocated to different groups on the same mating day.
- Fasting period before blood sampling for dam thyroid hormones: No
- Time of day for (rat) dam blood sampling: In the morning on the day of necropsy within a short timeframe (not exceeding two hours)

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Once a day, in the mornings. An inspection for signs of morbidity and mortality was made once on GD 0, after randomization, and twice daily from gestation day 1 to 19 (in the mornings and at the end of the work period of the laboratory personnel) as well as once on GD 20 (in the morning).
- Cage side observations included: Behavior and general condition

BODY WEIGHT: Yes
- Time schedule for examinations: On gestation days 0, 3, 5, 8, 11, 14, 17 and 20

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes (food consumption was measured between gestation days 0 to 3, 3 to 5, 5 to 8, 8 to 11, 11 to 14, 14 to 17 and 17 to 20)
- 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

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: Viscera were subjected to gross pathological examinations. The uterus with cervix and the left ovary and the thyroid glands together with the parathyroid glands were weighted. The number of corpora lutea in each ovary and implantation sites in each uterine horn, live fetuses, early and late embryonic death and fetal death were counted.

OTHER:
- Histological examination was performed on the thyroid glands of all females.

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes (uterus with cervix and the left ovary)
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Blood sampling:

- Plasma: No
- Serum: Yes
- Volume collected: At least 1.0 mL

Fetal examinations:

- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: Yes: [half per litter]
- Anogenital distance of all live rodent pups: The anogenital distance (AGD) of each fetus was determined (accuracy 1 mm).

Statistics:

The statistical evaluation of data was performed with the program package SPSS PC+4.0. The homogeneity of variance between groups was checked by Bartlett’s homogeneity of variance test.
Where no significant heterogeneity is detected a one-way analysis of variance (ANOVA) was carried out. If the obtained result is significant Duncan’s Multiple Range test was used to assess the significance of inter-group differences. If the result of the Bartlett’s test was significant, the Kruskal-Wallis analysis of variance was used and the inter-group comparisons were performed using Mann-Whitney U-test. Dams or litters were excluded from the data evaluation in cases of:
− Non pregnant females (no implantation, no corpora lutea), total exclusion
− Females with total pre-implantation loss (no implantation but corpora lutea), partial exclusion (only the intrauterine parameters were evaluated).

Although these animals were excluded from the data evaluation the Study Report contains
all data. A male/female fetus was considered as retarded in body weight, when its weight was below the average minus twofold standard deviation of the control male/female fetuses.

Indices:

% Pre-implantation loss: Number of corpora lutea - Number of implantations / Number of corpora lutea x 100

% Post-implantation loss: Number of implantations - Number of live foetuses / Number of implantations x 100

% Sex distribution: Number of Male (Female) fetuses / Number of fetuses x 100

% External, visceral and skeletal abnormalities/litter: Number of fetuses with abnormality / Number of fetuses x 100

Historical control data:

Yes, background pregnancy and fetal data from prenatal developmental toxicity studies in HAN: WIST rats between 2017 and 2021 were provided. This comprised 17 oral gavage studies (6x water, 1x olive oil, 7x sunflower oil and 3x PEG).

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

2360 mg/kg bw/day group: The body weight gain was slightly but statistically significantly lower (p<0.05) between G.D. 5 and 8, 14 and 17 as well as 5 and 20. Based on the slight manner, it was not considered as adverse effect. This conclusion is supported by the body weight as well as corrected body weight/gain values where no significant effects were observed.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

2360 mg/kg bw/day group: Slightly lower (p<0.05) food consumption was observed between G.D. 5 and 8, as well as 14 to 17. These findings are in line with the slightly lower body weight gain in these periods. Based on the slight manner, this was not considered as adverse, which was supported by the body weight and corrected body weight/gain parameters which were unaffected.

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

Endocrine findings:

effects observed, treatment-related

Description (incidence and severity):

The mean FT3 level was statistically significantly lower (p<0.05) in the 2360 mg/kg bw/day group, however, values were within the historical control range (at accuracy of 0.0001 ng/mL), hence this effect was not regarded as adverse. There were no significant differences in the FT4 and TSH levels.

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Dilated renal pelvis was found in one animal in the control, 236 and 2360 mg/kg bw/day group respectively and none at 710 mg/kg bw/day considering the equal distribution and low incidence, this was judged as unrelated to the treatment. Two females, one in the 710 and one in the 2360 mg/kg bw/day group had diaphragmatic hernia. This developmental disorder was not attributed to the treatment. This finding may occur in control rats according to the background data.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

no effects observed

Changes in number of pregnant:

no effects observed

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

1000 mg/kg bw/day corresponds to the substance concentration determined on the basis of 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one and its organic impurities (42.34 %) contained in the product. Dose of the product: 2360 mg/kg bw/day

Key result

Abnormalities:

no effects observed

Fetal body weight changes:

no effects observed

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

effects observed, non-treatment-related

Description (incidence and severity):

236 mg/kg bw/day group: Statistically significant difference was indicated in the sex distribution (p<0.05) by Duncan’s multiple range test and (p<0.01) by Chi square test (sex ratio 38.3: 61.7% males: females). As the sex distribution was approx. 50:50 in all other groups, the difference in the low dose group was not considered as treatment related.

Changes in litter size and weights:

no effects observed

Anogenital distance of all rodent fetuses:

no effects observed

Changes in postnatal survival:

not examined

External malformations:

effects observed, non-treatment-related

Description (incidence and severity):

There was one single malformed fetus in the 710 mg/kg bw/day dose group. This fetus was found with hypoplastic mandible. This malformation has been observed also in control animals of the historical control database and, also because of the lack of dose-dependence, was not considered to be treatment-related.

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

There were three fetuses found as malformed in the control group (two with bent scapula, one
with dumb-bell shaped thoracic vertebra), one in the 236 mg/kg bw/day group (bent scapula)
and one in the 710 mg/kg bw/day group (hypoplastic mandible, absence of Meckel's cartilage
and cleft palate; same fetus which was found with a hypoplastic mandible at external examination). Considering the low incidences and the lack of dose response, the observed malformations were not proven to be treatment related.

Skeletal variations such as delayed ossification of the skull bones, hyoid, sternebra, vertebrae, pectoral girdle (unossified os pubis, os ishii), as well as metacarpal and metatarsal were evaluated as variations and were found among the dose groups without statistically significant differences. Other variations, such as slightly open palate (one fetus in the 236 mg/kg bw/day group), bipartite or misaligned sternebra, dumb-bell shaped and bipartite (and/or asymmetric) vertebral centra and slightly bent scapula (one in the 236 mg/kg bw/day group) were observed with low incidences and without statistically significant differences and dose-response relationship. Hence, there were no adverse effects proven of the test item at skeletal examinations.

Visceral malformations:

effects observed, non-treatment-related

Description (incidence and severity):

One fetus was found with microphthalmia in the 236 mg/kg bw/day dose group as a single
case. As there was no dose-dependency and this finding has also been observed in the laboratory's background data, it was considered not treatment-related. Statistically significant occurence of variations in the treatment groups were of low incidence, not dose-dependent or were within the range of the laboratory's historical control data.

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

1000 mg/kg bw/day corresponds to the substance concentration determined on the basis of 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one and its organic impurities (42.34 %) contained in the product. Dose of the product: 2360 mg/kg bw/day

Key result

Abnormalities:

no effects observed

Key result

Developmental effects observed:

no

Conclusions:

In a Prenatal Developmental Toxicity Study in pregnant rats according to OECD 414, oral treatment from gestation day 5 up to day 19 with 4,5-dihydroxy-1,3-dimethylimidazolidin-2-one at the dose levels of 1000, 300 and 1000 mg/kg bw/day did not cause death, clinical signs, macroscopic changes in maternal organs and changes in maternal thyroids. Slightly lower food consumption and body weight gain from G.D. 5 to 8 as well as 14 to 17, as well as slightly lower body weight gain between G.D. 5 and 20 might be attributed to the treatment, however based on the slight manner, not considered as adverse. There was no test item effect proven on the FT3, FT4 and TSH hormone values. The treatment with the test item had no impact on the intrauterine parameters. There was no test item-related effects proven on external, visceral and skeletal examination of fetuses. Based on the observations the NOAEL (maternal toxicity) was 1000 mg/kg bw/day and the NOAEL (developmental toxicity, including teratogenicity) was 1000 mg/kg bw/day.

Executive summary:

In a Prenatal Developmental Toxicity Study in pregnant rats according to OECD 414 and GLP, groups of 26 sperm-positive female Han: WIST rats were treated with the test item in water (Aqua purificata) by oral gavage administration at three dose levels of 100, 300 and 1000 mg/kg bw/day, respectively, from gestation day 5 to 19. The substance concentrations were determined on the basis of 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one and its organic impurities (42.34 %) contained in the product. Applied doses of the product were 236, 710 and 2360 mg/kg bw/day. A concurrent control group of 26 sperm positive females was treated with the vehicle alone. The dose volume was 5 mL/kg/bw. Dosing formulations were analysed for homogeneity and achieved concentrations of the test item two times during the treatment period using a validated HPLC-RID (refractive index detection) method. The determined test item concentrations were within 10% of the nominal concentration and, therefore, the dosing formulations were considered acceptable. A homogenous distribution of the test item in the vehicle was demonstrated for all analyzed dosing formulations. For mating, the females were cohoused in the mornings with males of the same strain and source (1 male :1-2 females) and the day of detection of sperm in the vaginal smear of females was regarded as gestation day 0. During the study animals were checked daily twice for mortality and once for clinical signs. Body weight and food consumption of the dams were recorded repeatedly. Blood collection for determination of thyroid hormone FT3 (free T3) and FT4 (free T4), as well as TSH level was performed on gestation day 20. A Caesarean section and gross pathology were performed on gestation day 20. Organs of the dams were examined macroscopically. Thyroid glands of all sperm positive females were weighed and histological examination was performed on the thyroid glands of all females. The number of implantations, early and late resorptions, live and dead fetuses in each uterine horn and the number of corpora lutea were recorded. Each fetus was weighed and examined for sex and external abnormalities. The ano-genital distance was measured. The placentas were weighed and examined externally. The body of about half of each litter was subjected to visceral examination by means of a dissecting microscope after fixation in Sanomiya mixture. The heads were examined by Wilson's free-hand razor blade method. After double staining, the skeletons of the remaining fetuses (not assigned to visceral examination) were examined by means of a dissecting microscope. All abnormalities found during the fetal examinations were recorded.

 

Evaluated litters

On gestation day 20, there were 22 to 26 litters with live fetuses in the experimental groups.

 

Mortality, clinical signs

There was no mortality in the experimental groups before scheduled necropsy and there were no clinical signs recorded for the maternal animals.

 

Food consumption, body weight of dams

Slightly lower food consumption and body weight gain was observed in the 2360 mg/kg bw/day group between G.D. 5 and 8, as well as 14 to 17 and the body weight gain was slightly lower between G.D. 5 and 20. These slight differences might be attributed to the treatment of the dams with the test item, however based on the slight manner, not considered as adverse effects.

 

Thyroid hormone (FT3 and FT4) and TSH level

Statistically significantly (p<0.05) lower free thyroid hormone level (FT3) at 2360 mg/kg bw/day was at the historical control level. The test item had no effect on the FT4 and TSH value.

 

Thyroid weight, necropsy findings of dams, histopathology of thyroid glands

There were no differences indicated in the thyroid weight and there were no lesions found at histopathology. The necropsy findings with low incidences were unrelated to the treatment.

 

Intrauterine mortality

There was no increase indicated in the pre- and post-implantation loss or mean number of viable fetuses. There was no test item effect proved in the sex distribution of the fetuses. Statistically significant difference in the 236 mg/kg bw/day group was without a dose response.

 

Fetal- and placental weight, ano-genital distance

There was no test item effect indicated.

 

External examination of placentas

There were no test item related findings observed.

 

Fetal examinations

There was no test item effect proved at external, visceral and skeletal examination.

 

Oral treatment of pregnant Han: WIST rats from gestation day 5 up to day 19 (the day before Caesarean section) with 4,5-dihydroxy-1,3-dimethylimidazolidin-2-one at the dose levels of 2360, 710 and 236 mg/kg bw/day did not cause death, clinical signs, macroscopic changes in maternal organs and changes in maternal thyroids which could have been observed at histopathological evaluation. Slightly lower food consumption and body weight gain from G.D. 5 to 8 as well as 14 to 17, as well as slightly lower body weight gain between G.D. 5 and 20 might be attributed to the treatment, however based on the slight manner, not considered as adverse. There was no test item effect proven on the FT3, FT4 and TSH hormone values. The treatment with the test item had no impact on the intrauterine parameters. There was no test item-related effects proven on external, visceral and skeletal examination of fetuses. Based on the observations the NOAEL (maternal toxicity) was 2360 mg/kg bw/day and the NOAEL (developmental toxicity, including teratogenicity) was 2360 mg/kg bw/day, both corresponding to 1000 mg/kg bw/day when corrected for 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one content and its organic impurities.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Reliable without restrictions

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Key, Developmental toxicity, rat, RL1

In a Prenatal Developmental Toxicity Study in pregnant rats according to OECD 414 and GLP, groups of 26 sperm-positive female Han: WIST rats were treated with the test item in water (Aqua purificata) by oral gavage administration at three dose levels of 100, 300 and 1000 mg/kg bw/day, respectively, from gestation day 5 to 19. The substance concentrations were determined on the basis of 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one and its organic impurities (42.34 %) contained in the product. Applied doses of the product were 236, 710 and 2360 mg/kg bw/day. A concurrent control group of 26 sperm positive females was treated with the vehicle alone. The dose volume was 5 mL/kg/bw. Dosing formulations were analysed for homogeneity and achieved concentrations of the test item two times during the treatment period using a validated HPLC-RID (refractive index detection) method. The determined test item concentrations were within 10% of the nominal concentration and, therefore, the dosing formulations were considered acceptable. A homogenous distribution of the test item in the vehicle was demonstrated for all analyzed dosing formulations. For mating, the females were cohoused in the mornings with males of the same strain and source (1 male :1-2 females) and the day of detection of sperm in the vaginal smear of females was regarded as gestation day 0. During the study animals were checked daily twice for mortality and once for clinical signs. Body weight and food consumption of the dams were recorded repeatedly. Blood collection for determination of thyroid hormone FT3 (free T3) and FT4 (free T4), as well as TSH level was performed on gestation day 20. A Caesarean section and gross pathology were performed on gestation day 20. Organs of the dams were examined macroscopically. Thyroid glands of all sperm positive females were weighed and histological examination was performed on the thyroid glands of all females. The number of implantations, early and late resorptions, live and dead fetuses in each uterine horn and the number of corpora lutea were recorded. Each fetus was weighed and examined for sex and external abnormalities. The ano-genital distance was measured. The placentas were weighed and examined externally. The body of about half of each litter was subjected to visceral examination by means of a dissecting microscope after fixation in Sanomiya mixture. The heads were examined by Wilson's free-hand razor blade method. After double staining, the skeletons of the remaining fetuses (not assigned to visceral examination) were examined by means of a dissecting microscope. All abnormalities found during the fetal examinations were recorded.

 

Evaluated litters

On gestation day 20, there were 22 to 26 litters with live fetuses in the experimental groups.

 

Mortality, clinical signs

There was no mortality in the experimental groups before scheduled necropsy and there were no clinical signs recorded for the maternal animals.

 

Food consumption, body weight of dams

Slightly lower food consumption and body weight gain was observed in the 2360 mg/kg bw/day group between G.D. 5 and 8, as well as 14 to 17 and the body weight gain was slightly lower between G.D. 5 and 20. These slight differences might be attributed to the treatment of the dams with the test item, however based on the slight manner, not considered as adverse effects.

 

Thyroid hormone (FT3 and FT4) and TSH level

Statistically significantly (p<0.05) lower free thyroid hormone level (FT3) at 2360 mg/kg bw/day was at the historical control level. The test item had no effect on the FT4 and TSH value.

 

Thyroid weight, necropsy findings of dams, histopathology of thyroid glands

There were no differences indicated in the thyroid weight and there were no lesions found at histopathology. The necropsy findings with low incidences were unrelated to the treatment.

 

Intrauterine mortality

There was no increase indicated in the pre- and post-implantation loss or mean number of viable fetuses. There was no test item effect proved in the sex distribution of the fetuses. Statistically significant difference in the 236 mg/kg bw/day group was without a dose response.

 

Fetal- and placental weight, ano-genital distance

There was no test item effect indicated.

 

External examination of placentas

There were no test item related findings observed.

 

Fetal examinations

There was no test item effect proved at external, visceral and skeletal examination.

 

Oral treatment of pregnant Han: WIST rats from gestation day 5 up to day 19 (the day before Caesarean section) with 4,5-dihydroxy-1,3-dimethylimidazolidin-2-one at the dose levels of 2360, 710 and 236 mg/kg bw/day did not cause death, clinical signs, macroscopic changes in maternal organs and changes in maternal thyroids which could have been observed at histopathological evaluation. Slightly lower food consumption and body weight gain from G.D. 5 to 8 as well as 14 to 17, as well as slightly lower body weight gain between G.D. 5 and 20 might be attributed to the treatment, however based on the slight manner, not considered as adverse. There was no test item effect proven on the FT3, FT4 and TSH hormone values. The treatment with the test item had no impact on the intrauterine parameters. There was no test item-related effects proven on external, visceral and skeletal examination of fetuses. Based on the observations the NOAEL (maternal toxicity) was 2360 mg/kg bw/day and the NOAEL (developmental toxicity, including teratogenicity) was 2360 mg/kg bw/day, both corresponding to 1000 mg/kg bw/day when corrected for 4,5-dihydroxy-1,3 dimethylimidazolidin-2-one content and its organic impurities.

 

Supporting, read-across, Developmental toxicity, rat, RL2

BG Chemie (1998) reported a developmental/teratogenicity study with Dimethyloldihydroxyethylene urea (CAS 1854-26-8). Wistar rats were administered (gavage) concentrations of 250, 500 or 1000 mg/kg from day 7 - 16 of pregnancy. There were no deaths during the study. No clinical signs were observed in any of the animals. Body weights and food consumption were not affected by the administration of the test compound. No compound-related effects were observed at necropsy of the animals. Gravid uterus weights, crown-rump lengths, litter size, sex ratios, foetal and placental weights remained unaffected by the administration of the test compound. There was no increase in the number of early or late conceptuses undergoing resorption. Morphological examination of the fetuses did not reveal any compound-related effect. In conclusion, the oral administration of Dimethyloldihydroxyethylene urea to the rat at doses of 250, 500 or 1000 mg/kg body weight from day 7 - 16 of pregnancy did not cause any maternal toxicity or embryotoxicity.

Justification for classification or non-classification

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data, the test item is not classified for reproductive toxicity according to Regulation (EC) No 1272/2008 (CLP), as amended for the eighteenth time in Regulation (EU) 2022/692.