Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

Following daily oral administration (by gavage) of the Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl] methacrylamide and methacrylic acid at dose levels of 66.7, 200 and 600/400 mg/kg bw/day (i.e, 93, 279 and 836/557 mg/kg bw/day in terms of registered substance) to Sprague-Dawley rats (10/gender/dose) for at least 5 weeks of dosing in a GLP OECD 421-compliant study, the NOAEL for reproductive performance and the NOAEL for toxic effects on progeny were both set at 600/400 (836/557) mg/kg/day in the absence of any toxicologically relevant treatment-related effects at this dose-level.

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Remarks:
based on test type
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 27 April to January 2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study with no restrictions.
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories France, l’Arbresle, France.
- Age at study initiation: (P) 10 wks for males, 9 wks for females
- Weight at study initiation: (P) Males: 363-433 g; Females: 189-248 g
- Fasting period before study: no
- Housing: individually housed, except during pairing, in polycarbonate cages (Tecniplast 2154, 940 cm²) with stainless steel lids and containing autoclaved sawdust (SICSA, Alfortville, France). Toward the end of gestation and during lactation with their litter, autoclaved wood shavings (SICSA, Alfortville, France) were provided as nesting material, a few days before delivery and during the lactation period. Each cage contained an object (rat hut) for the enrichment of the environment of the rats. nor applicable
- Diet (e.g. ad libitum): free access to SSNIFF R/M-H pelleted maintenance diet, batch No. 2626975 (SSNIFF Spezialdiäten GmbH, Soest, Germany), which was distributed weekly.
- Water (e.g. ad libitum): free access to bottles containing tap water (filtered with a 0.22 µm filter).
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): approximately 12 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12h/12h

IN-LIFE DATES: From: 30 May 2012 To: 23 July 2012
Route of administration:
oral: gavage
Vehicle:
water
Remarks:
drinking water, treated by reverse osmosis
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was administered as a solution in the vehicle. The test item was mixed with the required quantity of vehicle. The test item dose formulations were prepared on a daily basis and were stored and delivered at room temperature in brown flasks.

DIET PREPARATION
not applicable

VEHICLE
- Justification for use and choice of vehicle (if other than water): not applicable
- Concentration in vehicle: 0; 13.3; 40 and 120/80 mg/mL
- Amount of vehicle (if gavage): constant dosage-volume of 5mL/kg bw/d
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: until mating occurs or 14 days have elapsed
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility: not applicable
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged (how): no data
- Any other deviations from standard protocol: no
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentrations of the test item in the dose formulations have been quantified by a validated analytical method.
The validation of the analytical method was conducted in CiToxLAB France/Study No. 38954 VAA and precise details concerning the checked parameters, acceptance criteria and obtained results are documented in the corresponding validation report. The concentration of the test item in samples of each control and test item dose formulation prepared for use in weeks 1, 3 and 5 was determined.
Duration of treatment / exposure:
in the males:
- 2 weeks before pairing (from study days 1 to 14),
- during the pairing period (3 weeks) (from study day 15 until study days 16 to 28),
- until sacrifice (at least 5 weeks in total) (from study days 17 to 29 until study day 36).
in the females:
- 2 weeks before pairing (from study days 1 to 14),
- during the pairing period (3 weeks) (from study days 15 to 28),
- during gestation (from study days 16 to 29 until study days 36 to 49),
- during lactation until day 4 post-partum inclusive
(from study days 37 to 50 until study days 41 to 54).
Frequency of treatment:
daily
Details on study schedule:
- Age at mating of the mated animals in the study: 12 weeks for males, 11 weeks for females
Remarks:
Doses / Concentrations:
0; 66.7; 200 and 600/400 mg/kg bw/d
Basis:
other: based on N-[2-(2-oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid content.
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: in a preliminary study, the test item was given to rats (3/sex/group), by daily oral administration (gavage) for 1 week at 0, 100, 300 or 1000 mg/kg bw/day. At 1000 mg/kg bw/day, all males had marked to severe clinical signs (ptyalism, piloerection, loud breathing, dyspnea, abdominal breathing, chromorhynorrhea, soiled snout and/or hypoactivity); 1/3 females had ptyalism. There was a minimal decrease in mean body weight (-6.7% vs. controls) at the end of the treatment period and a marked decrease in mean food consumption in males only (-25.7% vs. controls).
At necropsy, 1/3 males (1000 mg/kg bw/day) had a thymus reduced in size and stomach wall with red discoloration. One out of 3 females (300 mg/kg bw/day) had an uterus dilated and with a translucent content.
There were no treatment-related findings at 300 or 100 mg/kg bw/day.
Overall, 1000 mg/kg bw/day were considered to be an excessive dose-level. Therefore, 600 mg/kg bw/day were selected as the high-dose level. The low-dose and mid-dose have been selected using a ratio representing a three-fold interval (i.e. 66.7 and 200 mg/kg bw/day).

On study day 5 of the current study, 1/10 high dose males was euthanized for ethical reason and on study day 6, 3/9 surviving males from the same group had loud breathing, abdominal breathing and/or chromorhinorrhea and 1/10 females had piloerection and loud breathing. Therefore, 600 mg/kg/day was proved to be an excessive dose-level. It was thus decided to lower the dose administered to animals from the high-dose group from study day 6 (400 mg/kg/day instead of 600 mg/kg/day).
Positive control:
not required
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once a day before the treatment period and at least twice a day during the treatment period, including weekends and public holidays.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once a day

BODY WEIGHT: Yes
- Time schedule for examinations: male: on the first day of treatment (day 1), then once a week until sacrifice; female: on the first day of treatment (day 1), then once a week until mated (or until sacrifice) and on days 0, 7, 14 and 20 post-coitum (p.c.) and days 1 and 5 p.p..

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION: No
Oestrous cyclicity (parental animals):
The estrous cycle stage was determined from a fresh vaginal lavage (stained with methylene blue), each morning during the pairing period, until the females are mated.
Sperm parameters (parental animals):
Parameters examined in male parental generation: testis and epididymis weight for all tested males
Litter observations:
STANDARDISATION OF LITTERS
Not applicable

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities

GROSS EXAMINATION OF DEAD PUPS:
yes, for external abnormalities. No tissues were preserved.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals at the end of the pairing period (at least 5 weeks of treatment in total)
- Maternal animals: All surviving animals on day 5 p.p..

GROSS NECROPSY
- Gross necropsy consisted of external and inernal examinations including the external surfaces, all orifices, the cranial cavity, the external surfaces of the brain and spinal cord, the thoracic, abdominal and pelvic cavities with their associated organs and tissues and the neck with its associated organs and tissues. The numbers of corpora lutea and implantation sites were also recorded for females sacrificed as scheduled on day 5 post-partum.

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in Table 7.8.1/1 were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
SACRIFICE
Animals were subjected to postmortem macroscopic examinations.

GROSS NECROPSY
- Gross necropsy consisted of external examinations only. No tissues were preserved.

HISTOPATHOLOGY / ORGAN WEIGTHS
Not applicable as no tissues were preserved.
Statistics:
Data are compared by one-way analysis of variances and Dunnett test (mean values being considered as normally distributed, variances being considered as homogenous) or by Fischer exact probability test (proportions).

PathData software (version 6.2d2) was used to perform the statistical analysis of organ weight data (level of significance of 0.05 or 0.01).
Reproductive indices:
pre-implantation loss:
Number of corpora lutea - Number of implantation sites
_____________________________________________ x 100
Number of corpora lutea

post-implantation loss (manually calculated):
Number of implantation sites - Number of live pups
_____________________________________________ x 100
Number of implantations

mating index:
Number of mated animals
_____________________ x 100
Number of paired animals

fertility index:
Number of pregnant female partners
_______________________________ x 100
Number of mated pairs

gestation index:
Number of females with live born pups
________________________________ x 100
Number of pregnant females
Offspring viability indices:
live birth index:
Number of live born pups
_____________________ x 100
Number of delivered pups

viability index on day 4 post-partum:
Number of surviving pups on day 4 post-partum
_______________________________________ x 100
Number of live born pups


Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Mortality at 600/400 mg/kg bw/d. See more details below
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weight loss between days 1 and 8 for the males in the highest tested group. Lower body weight gain during the period of days 1 to 15 for the females treated with the top dose. Then mean body weight was comparable to the one of the control group.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Body weight loss between days 1 and 8 for the males in the highest tested group. Lower body weight gain during the period of days 1 to 15 for the females treated with the top dose. Then mean body weight was comparable to the one of the control group.
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
see details below
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Test substance intake: Decrease in mean food consumption in boh sexes from the high dose-group during the premating period. During the lactation period, lower mean food consumption than in the control group in the high dose group (but not statistically significant).
Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related
Description (incidence and severity):
See tables 7.8.1/3 and 7.8.1/4 and details below
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS): there were no unscheduled deaths in the control, 66.7 or 200 mg/kg bw/d groups. Deaths were recorded in the 600/400 mg/kg bw/d group. 2/10 males were euthanasied for ethical reason (day 5 and 8 respectively) and 3/10 females were found dead or sacrificed for ethical reason (day 8, 15 and 20 respectively). Before sacrifice or death, animals suffered of ptyalism, piloerection, hypoactivity, chromorhinorrhea, dyspnea, loud and abdominal breathing and presented a round back and emaciated appearance or were cold to the touch. In the surviving animals clinical signs were observed and listed in table 7.8.1/2. Piloerection, round back, emaciated appearance, loud and abdominal breathing were considered to be related to the test item treatment. Ptyalism was considered to be related to the treatment with the test item but of minor toxicological significance. All others clinical signs (areas of hair loss, cutaneous lesions, abnormal growth of teeth and/or chromodacryorrhea) are commonly observed in this species and strain. A treatment-related effect was considered unlikely.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS): In the group treated at 600/400 mg/kg/day, the pre and post-implantation losses were higher than in the control group and resulted in a non-statistically significant lower mean number of pups delivered (12.7 vs. 13.2 in the Historical Control Data). This minor difference was mainly due to one litter (with 57.1% of post-implantation loss) and therefore a test item treatment-related effect was considered unlikely.

GROSS PATHOLOGY (PARENTAL ANIMALS): gas distension of the stomach/cecum was found in 3/5 decedents, associated in one male with red and depressed foci which correlated with microscopic gastric erosions. In another decedent male, the thymus was gelatinous, which correlated with interlobular edema, and there was stress-associated thymic lymphoid atrophy and increased apoptotic cell numbers microscopically. In the surviving animals, few macroscopic findings were noted at the end of the treatment period in males but were of those commonly recorded in the Sprague-Dawley rat and none were considered to be related to the test item administration. There were no macroscopic findings in females.

HISTOPATHOLOGY (PARENTAL ANIMALS): The cause of deaths/morbidity in these rats was not evident at microscopic examination. In the surviving animals, there were no test item-related changes in the testis, epididymis, ovaries and oviducts.
Dose descriptor:
NOAEL
Remarks:
Parental toxicity
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
other: Dose adjusted for N-[2-(2-oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid content.
Sex:
male/female
Basis for effect level:
other: Mortality, clinical signs and effects on body weight and food consumption observed at 600/400 mg/kg bw/d
Dose descriptor:
NOAEL
Remarks:
Parental toxicity
Effect level:
279 mg/kg bw/day (actual dose received)
Based on:
other: Dose of substance as registered (including impurities residual water necessary for stability)
Sex:
male/female
Basis for effect level:
other: Mortality, clinical signs and effects on body weight and food consumption observed at 557/836 mg/kg bw/d
Dose descriptor:
NOAEL
Remarks:
Reproductive performance
Effect level:
400 - 600 mg/kg bw/day (actual dose received)
Based on:
other: Dose adjusted for N-[2-(2-oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid content.
Sex:
male/female
Basis for effect level:
other: Non-statistically significant increase in post-implantation losses in the highest dose group which was mainly due to one litter and therefore considered not to be adverse.
Dose descriptor:
NOAEL
Remarks:
Reproductive performance
Effect level:
557 - 836 mg/kg bw/day (actual dose received)
Based on:
other: Dose of substance as registered (including impurities residual water necessary for stability)
Sex:
male/female
Basis for effect level:
other: Non-statistically significant increase in post-implantation losses in the highest dose group which was mainly due to one litter and therefore considered not to be adverse.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
See details below and in Table 7.8.1/5.
Mortality / viability:
mortality observed, treatment-related
Description (incidence and severity):
See details below.
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
See details below and in Table 7.8.1/5.
Histopathological findings:
not examined
VIABILITY (OFFSPRING): In the groups treated at 200 or 600/400 mg/kg/day, the viability indexes were significantly reduced when compared with controls. However, there was no dose-relationship and the viability index of the high-dose group was similar to the mean viability index of the Historical Control Data (95.5% vs. 94.9%). Therefore, theses findings were considered not to be adverse.

CLINICAL SIGNS AND GROSS PATHOLOGY (OFFSPRING): There were dose-related increases in the percentages of litters with pups having clinical signs from 200 mg/kg/day. These increases were mainly due to litter Y23635 (200 mg/kg/day) and litter Y23648 (600/400 mg/kg/day) and there were no dose-related increases in the percentage of litters with pups having gross external abnormalities. Therefore a test item treatment-related effect was considered unlikely.
Dose descriptor:
NOAEL
Remarks:
toxicity on progeny
Generation:
F1
Effect level:
400 - 600 mg/kg bw/day (actual dose received)
Based on:
other: Dose adjusted for N-[2-(2-oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid content.
Sex:
male/female
Basis for effect level:
other: Viability indexes were slightly reduced in the 200 and 600 mg/kg bw/d groups but still comparable to historical control and therefore considered not to be adverse.
Dose descriptor:
NOAEL
Remarks:
toxicity on progeny
Generation:
F1
Effect level:
557 - 836 mg/kg bw/day (actual dose received)
Based on:
other: Dose of substance as registered (including impurities residual water necessary for stability)
Sex:
male/female
Basis for effect level:
other: Viability indexes were slightly reduced in the 557 and 836 mg/kg bw/d groups but still comparable to historical control and therefore considered not to be adverse.
Reproductive effects observed:
not specified

Table 7.8.1/2 : Clinical signs observed in the surviving animals

Sex

Male

Female

Dose-level (mg/kg/day)

0

66.7

200

600/400

0

66.7

200

600/400

Piloerection

0

1

0

4

0

0

0

0

Round back

0

0

0

2

0

0

0

0

Emaciated appearance

0

0

0

2

0

0

0

0

Aggressive behaviour

1

2

0

1

0

0

0

0

Loud breathing

0

0

0

5

0

0

2 (P/G)

2 (P/G)

3 (L)

Abdominal breathing

0

0

0

2

0

0

0

0

Ptyalism

0

1

0

8

0

0

0

2 (P)

5 (G)

Aera of hair loss on forelimb, abdomen or neck

1

2

1

0

0

1 (P/G/L)

0

0

Cutaneous lesion on neck

1

0

2

1

1 (G)

0

0

0

Abnormal growth of teeth

0

0

1

0

0

0

0

0

Chromodacryorrhea

0

0

0

0

1 (P/G)

0

0

0

P: premating and mating periods, G: gestation period, L: lactation period.

Table 7.8.1/3:Mating and fertility data results

Dose-level (mg/kg/day)

0

66.7

200

600/400

Number of animals paired (M + F)

10 + 10

10 + 10

10 + 10

8 + 8 (a)

Number of males mated

10

10

10

7(b)

Number of females mated

10

10

10

7(b)

Mean number of days taken to mate

3.1

3.8

2.4

3.6

Number of pregnant females

10

10

10

7

Male fertility index(%)

100

100

100

100

Female fertility index(%)

100

100

100

100

(a): one female was sacrificed the first day of pairing period and another one was found dead before the start of the pairing period.

(b):one female was sacrificed after a 5-day pairing period with no evidence of mating.

Table 7.8.1/4:Delivery data results

Dose-level (mg/kg/day)

0

66.7

200

600/400

Number of pregnant females

10

10

10

7

Number of females which delivered

10

10

10

7

Mean duration of gestation (days)

21.2

21.1

21.1

21.3

Mean number ofcorpora lutea

16.4

17.0

15.2

15.9

Mean number of implantations

16.2

16.8

14.8

15.1

Mean pre-implantation loss (%)

1.1

1.3

2.9

5.1

Mean number of pups delivered

14.6

15.0

13.5

12.7

Mean post-implantation loss (%)a

10.8

10.9

10.1

17.5

a: manually calculated, no statistics performed.

Table 7.8.1/5: Clinical signs and gross necropsy results in the offspring 

 

Clinical signs

Gross external abnormalities

Dose-level (mg/kg/day)

0

66.7

200

600/400

0

66.7

200

600/400

Scab on tail

 

Y23621-3

 

 

 

 

 

 

Dehydration

 

Y23621-13

 

 

 

 

 

 

Emaciated appearance

 

Y23621-13

Y23635-6

Y23635-10

Y23635-12

Y23639-14

Y23644-6

 

 

Y23635-6

 

Hind limb necrosis

 

 

Y23631-8

Y23648-7

 

 

Y23631-8

Y23648-7

Generalized pallor

 

 

Y23635-6

Y23635-12

 

 

 

 

 

Cold to the touch

 

 

Y23635-6

Y23635-12

Y23639-14

Y23648-12

 

 

 

 

Hematoma on back/abdomen

 

 

 

Y23643-9

 

 

 

 

Hematoma on hind limb

 

 

 

Y23648-12

 

 

 

 

 

 

 

 

 

 

 

 

 

Number of affected pups

0

2

5

4

0

0

2

1

Number of litter examined

10

10

10

7

10

0

10

7

Number of affected litters

0
(0.0)

1
(10.0)

3
(30.0)

3
(42.9)

0
(0.0)

0
(0.0)

2
(20.0)

1
(14.3)

In brackets: percentage (%) of affected litters.

Conclusions:
Based on the experimental conditions of this study:
- the No Observed Adverse Effect Level (NOAEL) for parental toxicity was considered to be 200 mg/kg/day (based on mortality, clinical signs and effects on body weight and food consumption recorded for both sexes at 600/400 mg/kg/day),
- the NOAEL for reproductive performance (mating, fertility and delivery data) was considered to be 600/400 mg/kg/day,
- the NOAEL for toxic effects on progeny was considered to be 600/400 mg/kg/day.
Executive summary:

The potential effects of the Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl] methacrylamide and methacrylic acid, on reproductive and developmental parameters were assessed in an OECD 421 compliant study following daily oral administration (by gavage) to male and female rats from before mating, through mating and, for females, through gestation until day 4post-partum (p.p.).

 

Three groups of ten male and ten female Sprague-Dawley rats received the test item, Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid (batch No. MWAM12059A), daily, by oral administration (gavage), 2 weeks before mating, during mating and, for the males, until sacrifice, for the females, throughout gestation until day 4p.p., at dose-levels of 66.7, 200 or 600/400 mg/kg/day (i.e, 93, 279 and 836/557 mg/kg bw/day in terms of registered substance). An additional group of ten males and ten females received the vehicle control, drinking water, under the same experimental conditions. The dosing volume was 5 mL/kg/day.

Animals were checked daily for clinical signs and mortality. Body weights and food consumption were recorded weekly until mating and then at designated intervals throughout gestation and lactation. The animals were paired for mating after 2 weeks of treatment and the dams were allowed to litter and rear their progeny until day 5p.p.. The total litter sizes and numbers of pups of each sex were recorded after birth. The pups were observed daily for clinical signs of toxicity and pup body weights were recorded on days 1 and 5p.p..

The males were sacrificed after completion of the mating period. Dams were sacrificed on day 5p.p.. Body weights and selected organs weights were recorded and a complete macroscopicpost-mortemexamination performed, with particular attention paid to the reproductive organs.

Microscopic examination was performed on the reproductive organs from control and high-dose males and females at terminal sacrifice and from premature decedents, and on all macroscopic observations in all groups.

Pups, including those found dead before study termination, were also submitted for a macroscopicpost-mortemexamination.

.

The test item concentrations in the administered dose formulations analyzed in weeks 1, 3 and 5 remained within an acceptable range of -9.0% to -0.4% when compared to the nominal values. Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid was not detected in control samples

 

As death occurred in one male exposed to the dose of 600 mg/kg bw/d on day 5 (animal sacrificed for ethical reason), the high-dose level was lowered to 400 mg/kg bw/d for the rest of the study period. An another male was euthanized for ethical reason on day 8 and 3/10 females were found dead or sacrificed on day 8, 15 and 20 respectively, at the highest dose-level. Before sacrifice or death, animals suffered of ptyalism, piloerection, hypoactivity, chromorhinorrhea, dyspnea, loud and abdominal breathing and presented a round back and emaciated appearance or were cold to the touch. In the surviving animals clinical signs were observed (piloerection, round back, emaciated appearance, loud and abdominal breathing) and were considered to be related to the test item treatment. In the surviving animals, few macroscopic findings were noted at the end of the treatment period in males but were of those commonly recorded in the Sprague-Dawley rat and none were considered to be related to the test item administration.

 

With regards to reproductive and developmental parameters, the following observations were made:

 

Mating and fertility data: there were no effects on the mean number of days taken to mate. All mated females were pregnant.

 

Delivery data: there were no obvious treatment related effects. In the group treated at 600/400 mg/kg/day, the post-implantation loss was higher than in the control group but with no statistical significance. This difference was mainly due to one litter (with 57.1% of post-implantation loss).

 

Pups mortality: there were no obvious treatment-related effects on pup mortality.

 

Pups clinical signs and external abnormalities: there were no test item treatment-related findings.

 

Pup viability: there were no treatment-related effects on live birth and lactation indexes. In the groups treated at 200 or 600/400 mg/kg/day, the viability indexes were statistically significantly reduced (94.8% and 95.5% respectively,vs.controls) but still comparable to Historical Control Data (94.9%) and therefore considered not to be adverse.

 

Pup body weight: there were no effects on mean pup body weights and mean pups body weight changes when compared with controls.

 

Pup sex ratio: there were no treatment-related effects on sex ratio (% of male pups) both on days 1 and 5p.p..

 

Pathology: At terminal sacrifice, the test item administration did not induce any organ weight or macroscopic changes. There were no significant microscopic findings in the testis, epididymis, ovaries and oviducts.

 

In conclusion, based on the experimental conditions of this study:

- the No Observed Adverse Effect Level (NOAEL) for parental toxicity was considered to be 200 mg/kg/day (based on mortality, clinical signs and effects on body weight and food consumption recorded for both sexes at 600/400 mg/kg/day),

- the NOAEL for reproductive performance (mating, fertility and delivery data) was considered to be 600/400 mg/kg/day,

- the NOAEL for toxic effects on progeny was considered to be 600/400 mg/kg/day

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
557 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The study available is a reprotoxic/developmental toxicity screening test with a good reliability (Klimisch score = 1).
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

Only one study available

The potential effects of the Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl] methacrylamide and methacrylic acid, on reproductive and developmental parameters were assessed in an OECD 421 compliant study following daily oral administration (by gavage) to male and female rats from before mating, through mating and, for females, through gestation until day 4 post-partum (p.p.).

 

Three groups of ten male and ten female Sprague-Dawley rats received the test item, Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid (batch No. MWAM12059A), daily, by oral administration (gavage), 2 weeks before mating, during mating and, for the males, until sacrifice, for the females, throughout gestation until day 4p.p., at dose-levels of 66.7, 200 or 600/400 mg/kg/day (i.e, 93, 279 and 836/557 mg/kg bw/day in terms of registered substance). An additional group of ten males and ten females received the vehicle control, drinking water, under the same experimental conditions. The dosing volume was 5 mL/kg/day.

Animals were checked daily for clinical signs and mortality. Body weights and food consumption were recorded weekly until mating and then at designated intervals throughout gestation and lactation. The animals were paired for mating after 2 weeks of treatment and the dams were allowed to litter and rear their progeny until day 5p.p.. The total litter sizes and numbers of pups of each sex were recorded after birth. The pups were observed daily for clinical signs of toxicity and pup body weights were recorded on days 1 and 5p.p..

The males were sacrificed after completion of the mating period. Dams were sacrificed on day 5p.p.. Body weights and selected organs weights were recorded and a complete macroscopicpost-mortemexamination performed, with particular attention paid to the reproductive organs.

Microscopic examination was performed on the reproductive organs from control and high-dose males and females at terminal sacrifice and from premature decedents, and on all macroscopic observations in all groups. Pups, including those found dead before study termination, were also submitted for a macroscopicpost-mortemexamination.

 

With regards to reproductive and developmental parameters, there were no toxicologically relevant treatment-related effects in any of the parameters studied, regarding mating, fertility, delivery and pup growth or development.

 

In conclusion, the No Observed Adverse Effect Level (NOAEL) for reproductive performance and the NOAEL for toxic effects on progeny were both set at 600/400 (836/557) mg/kg/day in the absence of any toxicologically relevant treatment-related effects at this dose-level.

Effects on developmental toxicity

Description of key information

Based on the results in this prenatal developmental toxicity study the maternal and developmental No Observed Adverse Effect Level (NOAEL) for Reaction Mass of n-[2-(2- oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid was established as being at least 500 mg/kg.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 03 june 2016 to 21 april 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
Temporary deviations from the daily mean relative humidity occurred
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
Purity/composition correction factor : Yes, correction factor is 1.39 based on purity
Test item handling: No specific handling conditions required
Stability at higher temperatures: Not available
Chemical name (IUPAC), synonym or trade name: Reaction mass of N-[2-(2-oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid ; SIPOMER WAM II
CAS Number: Not indicated. EC number: 934-058-1, used for registration
Molecular structure: Not indicated
Molecular formula: Not indicated
Molecular weight: 119 (The molecular weight is given as a mean value of the molecular weight of the components and additives of the reaction mass, based on the typical concentrations measured on a sample of the multiconstituant substance)
Irritant or corrosive: Yes, severely irritant to the eyes
pH: 2-5 at concentration of 10%
Specific gravity/density: 1.1086 (relative density at 20°C)
Solubility in vehicle:
• Water: 999 g/L
Stability in vehicle:
• Water: Stability for at least 1 day at room temperature, 8 days in the refrigerator and 3 weeks in the freezer is confirmed over the concentration range 1 to 200 mg/g, Test Facility Study No. 512223
Species:
rat
Strain:
other: Crl:WI (Han)
Details on test animals and environmental conditions:
- Number: F0-generation: 88 females. F1-generation: 886 fetuses.
- Strain and sanitary status: Rat: Crl:WI(Han) (outbred, SPF-Quality).
- Breeder: Charles River Deutschland, Sulzfeld, Germany.
- Age/Weight: Females were approximately 10-14 weeks.
- Housing: Females were individually housed in Macrolon plastic cages (MIII type, height 18 cm). Sterilized sawdust as bedding material (Lignocel S 8-15, JRS - J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) and paper as cageenrichment/ nesting material (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom) were supplied.
- Food and water: Diet : Free access to pelleted rodent diet (SM R/M-Z from SSNIFF®Spezialdiäten GmbH, Soest, Germany). Water: Free access to tap-water.
- Acclimation: At least 5 days prior to treatment.
- Allocation to study:

Group Dose level * (mg/kg) Number of females Animal numbers
1 0 22 1-22
2 50 22 23-44
3 150 22 45-66
4 500 22 67-88

- Identification: By indelible ink.

ENVIRONMENTAL CONDITIONS
- temperature: 18 to 24°C,
- relative humidity: 40 to 70%,
- light/dark cycle: 12h/12h,
- ventilation: at least 10 air changes/hour
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
Dose formulation preparation
Formulations (w/w) were prepared daily within 6 hours prior to dosing and were homogenized to a visually acceptable level. No adjustment was made for specific gravity/density of the vehicle. Adjustment was made for specific gravity of the test item
(1.1086). A correction was made for the purity/composition of the test item. A correction factor of 1.39 was used.
Appearance of formulations: Solution (Groups 2-4).
Storage conditions: At room temperature.
Formulations were placed on a magnetic stirrer during dosing.

VEHICLE
- Lot/batch no. (if required): water (Elix, Millipore S.A.S., Molsheim, France)
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analyses were conducted on a single occasion during the treatment phase (25 July 2016), according to a validated method (Test Facility Study No. 512223). Samples of formulations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations).
The accuracy of preparation was considered acceptable if the mean measured concentrations were 90-110% of the target concentration. Homogeneity was demonstrated if the coefficient of variation was ≤ 10%.
Details on mating procedure:
- Mating: Untreated females were mated at the Supplier, and were at Day 0 or 1 post-coitum on arrival at the Test Facility (Day 0 post-coitum was the day of successful mating; confirmed by vaginal plug).
Duration of treatment / exposure:
From Days 6 to 20 post-coitum, inclusive.
Frequency of treatment:
Once daily for 7 days per week, approximately the same time each day with a maximum of 6 hours of difference between the earliest and latest dose.
Duration of test:
Two months and a half
Dose / conc.:
50 mg/kg bw/day
Dose / conc.:
150 mg/kg bw/day
Dose / conc.:
500 mg/kg bw/day
No. of animals per sex per dose:
22 mated female per group
Control animals:
yes, concurrent vehicle
Details on study design:
Rationale for dose-level selection
Dose levels were selected based on results of the dose range finding study (Test Facility Study No.512216), in which dose levels of 100, 300 and 1000 mg/kg were tested. Three out of six females at 1000 mg/kg were killed in extremis on Day 11 post-coitum, as they showed lethargy, hunched posture and piloerection. Moreover, rales were noted for one female given 1000 mg/kg. Body weight loss (12 to 14%) was noted for these females on Day 9 post-coitum and food consumption was slightly reduced on Days 6-9 post-coitum. At necropsy, dark red foci on the glandular mucosa of the stomach was noted for one female given 1000 mg/kg. The remaining three females at 1000 mg/kg showed rales (2/3), food consumption was slightly reduced on Days 6-15 post-coitum and lower gravid uterus corrected body weight gain were noted. No toxicologically relevant effects were observed at 100 and 300 mg/kg.
Maternal examinations:
Mortality / Viability:
At least twice daily.

Clinical signs:
At least once daily from Day 2 post-coitum onwards up to the day prior to necropsy. The time of onset, grade and duration of any observed signs were recorded. Signs were graded for severity and the maximum grade was predefined at 3 or 4. Grades were coded as slight (grade 1), moderate (grade 2), severe (grade 3) and very severe (grade 4). For certain signs, only its presence (grade 1) or absence (grade 0) was scored. In the data tables, the scored grades were reported, as well as the percentage of animals affected in summary tables.

Body weights:
Days 2, 6, 9, 12, 15, 18 and 21 post-coitum.

Food consumption:
Days 2-6, 6-9, 9-12, 12-15, 15-18 and 18-21 post-coitum.

Water consumption:
Subjective appraisal was maintained during the study, but no quantitative investigation was introduced as no treatmentrelated effect was suspected.


POST-MORTEM EXAMINATIONS:
- All animals were sacrificed on Day 21 post-coitum using an oxygen/carbon dioxide procedure and subsequently subjected to an external, thoracic and abdominal examination, with special attention being paid to the reproductive organs.
- All macroscopic abnormalities were recorded, collected and fixed in 10% buffered formalin (neutral phosphate buffered 4% formaldehyde solution, Klinipath, Duiven, The Netherlands).
- Each ovary and uterine horn of all animals was dissected and examined as quickly as possible.

Ovaries and uterine content:
The ovaries and uterus of the females were examined to determine:
• The number of corpora lutea.
• The weight of the (gravid) uterus.
• The number and distribution of live and dead fetuses.
• The number and distribution of embryo-fetal deaths (early and late resorptions).
• The weight of each fetus.
• The sex of each fetus from the ano-genital distance (during necropsy) and also from gonadal inspections (during further fetal examination).
• Externally visible macroscopic fetal abnormalities.

The following definitions were applicable for implantation data:
• Fetal (late) resorptions were defined as a dead fetus with external degenerative changes and presence of distinguishable features such as head or limbs.
• Embryonic (early) resorptions were defined as evidence of implantation without presence of distinguishable features such as head or limbs.
• Dead fetus was defined as a non-viable fetus without external degenerative changes and presence of distinguishable features such as head or limbs.
• Post-implantation loss included embryonic (early) resorptions, fetal (late) resorptions and dead fetuses.

In case implantations were not macroscopically visible, the uterus was stained using the Salewski technique in order to determine any former implantation sites (Salewski staining prepared at Charles River Den Bosch using Ammoniumsulfide-solution 20% (Merck, Darmstadt, Germany) and Milli-Ro water (Millipore Corporation, Bedford, USA)).

Fetal examinations:
External, visceral, and skeletal findings were recorded as developmental variations (alterations in anatomic structure that are considered to have no significant biological effect on animal health or body conformity and/or represent slight deviations from normal) or malformations (those structural anomalies that alter general body conformity, disrupt or interfere with normal body function, or may be incompatible with life).

External examination
Each viable fetus was examined in detail, weighed and sexed. All live fetuses were euthanized by administration of approximately 0.05 mL (=10mg) of sodium pentobarbital (Euthasol® 20%; AST Farma B.V., Oudewater) into the oral cavity using a small flexible plastic or metal feeding tube. Nonviable fetuses (for which the degree of autolysis was minimal or absent) were examined and weighed. For late resorption A079-009, a gross external examination was performed. As no malformation was observed, this late resorption together with the early resorptions were discarded.

Visceral (Internal) examination
Approximately one-half of the fetuses (live and dead) in each litter (all groups) were examined for visceral anomalies by dissection in the fresh (non-fixed) state. The thoracic and abdominal cavities were opened and dissected using a technique described by Stuckhardt and Poppe. This examination included the heart and major vessels. Fetal kidneys were examined and graded for renal papillae development as described by Woo and Hoar. The sex of all fetuses was confirmed by internal examination.
As prominent visceral malformations were noted for two fetuses (A049-03 and A069-05), selected for skeletal examination, additional visceral examination of these fetuses was performed. The heads were removed from approximately one-half of the fetuses in each litter and placed in Bouin's solution (Klinipath, Duiven, The Netherlands) for soft-tissue examination of all groups using the Wilson sectioning technique. Any remaining tissues (from the fetuses used for fresh visceral examination) were discarded. The carcasses were processed and stained with Alizarin Red S (as described below), but not examined in first instance.

Skeletal examination
From the other one-half of the fetuses (live and dead) in each litter (all groups), the sex was confirmed by internal examination. All fetuses were eviscerated, fixed in 96% aqueous ethanol, macerated in potassium hydroxide (Merck, Darmstadt, Germany) and stained with Alizarin Red S (Klinipath, Duiven, The Netherlands) by a method similar to that described by Dawson. Skeletal examination was done for one-half of the fetuses (i.e. the fetuses with heads). The specimens were archived in glycerin (BRENNTAG Nederland B.V., Dordrecht, The Netherlands) with bronopol (Alfa Aesar, Karlsruhe, Germany) as preservative. A few bones were not available for skeletal examination because they were accidentally damaged or lost during processing. The missing bones were listed in the raw data; evaluation by the fetal pathologist and study director determined that there was no influence on the outcome of the individual or overall skeletal examinations, or on the integrity of the study as a whole.
Statistics:
The following statistical methods will be used to analyze the data:
- If the variables can be assumed to follow a normal distribution, the Dunnett-test (many-to-one t-test) based on a pooled variance estimate will be applied for the comparison of the treated groups and the control groups for each sex.
- The Steel-test (many-to-one rank test) will be applied if the data cannot be assumed to follow a normal distribution.
- The Fisher Exact-test will be applied to frequency data.
- The Mann Whitney test will be used to compare mean litter proportions (percent of litter) of the number of viable and dead fetuses, early and late resorptions, total resorptions, pre- and post-implantation loss, and sex distribution.
- Mean litter proportions (percent per litter) of total fetal malformations and developmental variations (external, visceral and skeletal), and each particular external, visceral and skeletal malformation or variation will be subjected to the Kruskal-Wallis nonparametric ANOVA test to determine intergroup differences. If the ANOVA reveals statistically significant (p<0.05) intergroup variance, Dunn’s test will be used to compare the compound-treated groups to the control group.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Rales were noted for seven females at 500 mg/kg for 1 to 11 days at a slight to moderate degree. Piloerection and hunched posture, in addition to rales, were noted for single females at 500 mg/kg (no. 70 and 76, respectively) on one single day. As these signs were observed in the high dose group only, these were considered to be treatment-related. Incidental findings that were noted included alopecia and diarrhoea. These findings occurred within the range of background findings to be expected for rats of this age and strain which are housed and treated under the conditions in this study. At the incidence observed, these were considered signs of no toxicological relevance.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
No unscheduled mortality occurred.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Mean body weights and body weight gains of animals treated up to 500 mg/kg remained in the same range as controls. One female at 500 mg/kg (no. 76) showed significantly lower body weights and body weight gain over the entire treatment period, when compared to the other females in the 500 mg/kg group. This female had a body weight loss of 11% from Day 6 to 9 post-coitum. The days after, body weight was slightly increased. Over the entire treatment period, i.e. from Day 6 to
20 post-coitum, a body weight loss of 4% was noted for this female. As only one out of twenty-two females of the 500 mg/kg group was affected, this was not considered to be toxicologically relevant.
Weight gain corrected for gravid uterus was slightly lower in the treated groups, compared to controls. The mean corrected weight gain was 28.5, 22.7, 23.6 and 21.7 gram in respectively the control, 50, 150 and 500 mg/kg groups. This decrease reached statistical significance at 500 mg/kg. As the changes in all treatment groups were only slight, within the range considered normal for rats of this age and strain and without a dose-response, they were not considered to be toxicologically relevant.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
No toxicologically relevant changes in mean food consumption before and after correction for body weight were noted up to 500 mg/kg. The trend towards slightly reduced food consumption before and after correction for body weight in the treated groups compared to controls was not considered to be toxicologically relevant. The reduction was only slight, no dose-response relationship was observed and the values remained within the range considered to be normal for rats of this age and strain. Moreover, the statistically significantly lower relative food consumption at 150 mg/kg on Days 12 to 15 post-coitum was not considered to be toxicologically relevant as the change was only slight and without dose-response.
Female no. 76 had a significantly lower food intake over the entire treatment period when compared to the other females of the 500 mg/kg group, which was the cause of the significantly lower body weight gain of this female.
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
Immunological findings:
not examined
Other effects:
no effects observed
Description (incidence and severity):
Macroscopic Examination:
One female at 500 mg/kg (no. 76) was emaciated and showed a thymus reduced in size. Moreover, the jejunum, ileum, caecum and colon of this female were distended with gas. These findings were related to the observations during the in-life phase, i.e. rales, hunched posture and significant lower body weight and food consumption (see previous sections). Alopecia was noted in the control, 50 and 500 mg/kg group for single females, confirming the clinical sign observed during the in-life phase. At the incidence observed, this was not considered to be treatment-related.
Details on results:
Treatment up to 500 mg/kg resulted in significant lower body weight and food consumption of one single female given the high dose. This female showed rales and hunched posture during the in-life phase, and was emaciated and showed a thymus reduced in size at necropsy. Moreover, its jejunum, ileum, caecum and colon were distended with gas. For the remaining females treated at 500 mg/kg, no toxicologically relevant changes in body weight and food consumption were noted and no treatment-related clinical signs or macroscopic findings were observed. As only one out of twenty-two females in the 500 mg/kg group was affected, this was not considered to be toxicologically relevant.
No maternal toxicity was observed in the 50 and 150 mg/kg groups.
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
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Changes in number of pregnant:
no effects observed
Other effects:
no effects observed
Details on maternal toxic effects:
Maternal Pregnancy Data:
Three females in the 500 mg/kg group (nos. 80, 83 and 88) were not pregnant. This was not considered to be treatment-related as treatment started from implantation onwards (i.e. Day 6 post-coitum). All other females were pregnant and had litters with viable fetuses. The lower number of pregnant females in Group 4 did not affect the evaluation as there were sufficient litters available for a proper toxicological evaluation. The numbers of pregnant females, corpora lutea and implantation sites, and pre-implantation loss in the control and test groups were similar and in the range of normal biological variation.
Key result
Dose descriptor:
NOAEL
Remarks:
maternal toxicity
Effect level:
500 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: no relevant effects
Fetal body weight changes:
no effects observed
Description (incidence and severity):
There were no effects on mean fetal body weights (both sexes) noted up to 500 mg/kg. Female no. 76 had 10 fetuses which had significant lower fetal body weights. The mean fetal body weight of this litter was 3.4 gram, compared to a mean value of 5.3 gram for Group 4. As the remaining litters of Group 4 had normal fetal weights, this was considered related to the maternal observations of this single dam during the in-life phase, i.e. rales, hunched posture and significant lower body weight and food consumption. Mean combined (male and female) fetal body weights were 5.2, 5.3, 5.2 and 5.3 gram for the control, 50, 150 and 500 mg/kg groups, respectively.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Description (incidence and severity):
The male:female ratio was unaffected by treatment up to 500 mg/kg. Mean sex ratios (males:females) were 48:52, 52: 48, 50:50 and 58:42 for the control, 50, 150 and 500 mg/kg groups, respectively.
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
There were no effects on litter size of any group. Mean litter sizes were 10.0, 10.8, 10.7 and 10.2 viable fetuses/litter for the control, 50, 150 and 500 mg/kg groups, respectively.
Changes in postnatal survival:
no effects observed
External malformations:
no effects observed
Description (incidence and severity):
There were no treatment-related effects on external morphology following treatment up to 500 mg/kg. The only external malformation that occurred in this study was observed in Group 3 fetus A049-03. This fetus had a small lower jaw and was found to be missing an eye. At subsequent skeletal examination, both malformations were confirmed, but as these occurred singly, they were considered to be chance findings. External variations were not seen in any group.
Skeletal malformations:
no effects observed
Description (incidence and severity):
There were no treatment-related effects on skeletal morphology following treatment up to 500 mg/kg. The only skeletal malformation (a bent scapula) in this study was noted in control fetus A021- 03 and as such was considered a chance finding.
Skeletal variations occurred at an incidence of 76.3%, 89.0%, 76.6% and 74.9% per litter in Groups 1, 2, 3 and 4, respectively. Noteworthy is the variation of bent ribs for which the incidence in Group 4 was statistically significantly lower than the control value. Mean litter incidences of this finding were 17.9%, 15.6%, 11.2% and 3.9% per litter in Groups 1, 2, 3 and 4, respectively. As all values were within the historical control data range (0.8% - 22.3% per litter) and as a decreased incidence of this finding is not an adverse effect, this lower value in Group 4 was not considered to be toxicologically relevant.
Other skeletal variations that were noted in this study occurred at low incidences, in the absence of a dose-related incidence trend and/or at frequencies that were within the range of available historical control data, and were therefore not considered to be toxicologically relevant.
Visceral malformations:
no effects observed
Description (incidence and severity):
There were no treatment-related effects on visceral morphology following treatment up to 500 mg/kg. Visceral malformations were observed in respectively 1, 1, 1 and 2 fetuses in Groups 1, 2, 3 and 4. The two affected fetuses of Group 4 were from the same litter (i.e. fetuses A069-02 and -05) and both had a right-sided aortic arch. Hence, the origin was considered to be likely genetic rather than toxicological. The other affected fetuses were the Group 3 fetus (A049-03) with an absent eye and Group 1 and 2 fetuses A014-03 and A028-05, respectively, of which all organs were laterally transposed. Due to the single occurrence and/or occurrence in a control fetus only, these were considered to be chance findings. The visceral variations that were noted in this study occurred singly or at low incidences in the absence of a dose-related incidence trend and therefore were not considered to be treatment-related.
Other effects:
no effects observed
Details on embryotoxic / teratogenic effects:
The significantly lower fetal body weights of one single litter at 500 mg/kg were considered related to the maternal observations of this single dam during the in-life phase rather than a developmental cause. No developmental toxicity was observed in the 50, 150 and 500 mg/kg groups.
Key result
Dose descriptor:
NOAEL
Remarks:
Development
Effect level:
500 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no relvant effects
Abnormalities:
no effects observed
Developmental effects observed:
no

see attached document

Conclusions:
Based on the results in this prenatal developmental toxicity study the maternal and developmental No Observed Adverse Effect Level (NOAEL) for Reaction Mass of n-[2-(2- oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid was established as being at least 500 mg/kg.
Executive summary:

The objective of this prenatal development toxicity study (2017) was to evaluate the potential toxic effects of the test item, Reaction Mass of n-[2-(2-oxoimidazolidin-1- yl)ethyl]methacrylamide and methacrylic acid, in rats by oral gavage according to the OECD guideline n° 414.

 

Eighty-eight mated female Wistar Han rats were assigned to four dose groups. The test item was administered once daily by oral gavage from Days 6 to 20 post-coitum at doses of 50, 150 and 500 mg/kg (Groups 2, 3 and 4 respectively). The high dose of 500 mg/kg was considered to be the highest feasible dose in this type of study as treatment-related mortality was observed at 1000 mg/kg for half of the females in the dose range finding study. The rats of the control group received the vehicle, water, alone. Females were checked daily for the presence of clinical signs. Food consumption and body weight were determined at periodic intervals. Formulations prepared on one day during treatment were analyzed for accuracy and homogeneity. All animals surviving to Day 21 post-coitum were subjected to an examinationpost-mortemand external, thoracic and abdominal macroscopic findings were recorded. Gross lesions were collected and fixed from all animals at necropsy. A laparohysterectomy was performed on each surviving female of the groups. The uteri, placentae and ovaries were examined, and the numbers of fetuses, early and late resorptions, total implantations and corpora lutea were recorded. Gravid uterine weights were recorded, and corrected body weights (changes) were calculated. The fetuses were weighed, sexed and examined for external, visceral and skeletal malformations and developmental variations. All live fetuses were euthanized.One half of the fetuses were decapitated and the heads were fixed in Bouin’s fixative; these fetuses were dissected and examined for visceral anomalies. The other one-half of the fetuses were processed and stained with Alizarin Red S for skeletal examinations.

 

Maternal findings

Treatment up to 500 mg/kg resulted in significant lower body weight and food consumption of one single female given the high dose. This female showed rales and hunched posture during the in-life phase, and was emaciated and showed a thymus reduced in size at necropsy. Moreover, its jejunum, ileum, caecum and colon were distended with gas. For the remaining females treated at 500 mg/kg, no toxicologically relevant changes in body weight and food consumption were noted and no treatment-related clinical signs or macroscopic findings were observed. As only one out of twenty-two females in the 500 mg/kg group was affected, this was not considered to be toxicologically relevant. No maternal toxicity was observed in the 50 and 150 mg/kg groups.

 

Developmental findings

The significantly lower fetal body weights of one single litter at 500 mg/kg were considered related to the maternal observations of this single dam during the in-life phase rather than a developmental cause. No developmental toxicity was observed in the 50, 150 and 500 mg/kg groups.

 

Based on the results in this prenatal developmental toxicity study the maternal and developmental No Observed Adverse Effect Level (NOAEL) for Reaction Mass of n-[2-(2- oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid was established as being at least 500 mg/kg.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
500 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
The study available is a prenatal developmental toxicity test with a good reliability (Klimisch score = 1).
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

Only one study available

The objective of this prenatal development toxicity study (2017) was to evaluate the potential toxic effects of the test item, Reaction Mass of n-[2-(2-oxoimidazolidin-1- yl)ethyl]methacrylamide and methacrylic acid, in rats by oral gavage according to the OECD guideline n° 414.

 

Eighty-eight mated female Wistar Han rats were assigned to four dose groups. The test item was administered once daily by oral gavage from Days 6 to 20 post-coitum at doses of 50, 150 and 500 mg/kg (Groups 2, 3 and 4 respectively). The high dose of 500 mg/kg was considered to be the highest feasible dose in this type of study as treatment-related mortality was observed at 1000 mg/kg for half of the females in the dose range finding study. The rats of the control group received the vehicle, water, alone. Females were checked daily for the presence of clinical signs. Food consumption and body weight were determined at periodic intervals. Formulations prepared on one day during treatment were analyzed for accuracy and homogeneity. All animals surviving to Day 21 post-coitum were subjected to an examinationpost-mortemand external, thoracic and abdominal macroscopic findings were recorded. Gross lesions were collected and fixed from all animals at necropsy. A laparohysterectomy was performed on each surviving female of the groups. The uteri, placentae and ovaries were examined, and the numbers of fetuses, early and late resorptions, total implantations and corpora lutea were recorded. Gravid uterine weights were recorded, and corrected body weights (changes) were calculated. The fetuses were weighed, sexed and examined for external, visceral and skeletal malformations and developmental variations. All live fetuses were euthanized.One half of the fetuses were decapitated and the heads were fixed in Bouin’s fixative; these fetuses were dissected and examined for visceral anomalies. The other one-half of the fetuses were processed and stained with Alizarin Red S for skeletal examinations.

 

Maternal findings

Treatment up to 500 mg/kg resulted in significant lower body weight and food consumption of one single female given the high dose. This female showed rales and hunched posture during the in-life phase, and was emaciated and showed a thymus reduced in size at necropsy. Moreover, its jejunum, ileum, caecum and colon were distended with gas. For the remaining females treated at 500 mg/kg, no toxicologically relevant changes in body weight and food consumption were noted and no treatment-related clinical signs or macroscopic findings were observed. As only one out of twenty-two females in the 500 mg/kg group was affected, this was not considered to be toxicologically relevant. No maternal toxicity was observed in the 50 and 150 mg/kg groups.

 

Developmental findings

The significantly lower fetal body weights of one single litter at 500 mg/kg were considered related to the maternal observations of this single dam during the in-life phase rather than a developmental cause. No developmental toxicity was observed in the 50, 150 and 500 mg/kg groups.

 

Based on the results in this prenatal developmental toxicity study the maternal and developmental No Observed Adverse Effect Level (NOAEL) for Reaction Mass of n-[2-(2- oxoimidazolidin-1-yl)ethyl]methacrylamide and methacrylic acid was established as being at least 500 mg/kg.

Toxicity to reproduction: other studies

Additional information

not applicable

Justification for classification or non-classification

Harmonized classification:

No harmonized classification is available according to the Regulation (EC) No 1272/2008 including ATP3.

Self-classification:

In the absence of any treatment-related adverse effects on reproductive and developmental parameters in an OECD 421 compliant study, Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl] methacrylamide and methacrylic acid is considered as non reprotoxic according to the Regulation (EC) No 1272/2008 (CLP) and the Directive 67/548/EEC.

In the absence of any treatment-related adverse effects on developmental parameters in an OECD 414 compliant study, Reaction mass of N-[2-(2-Oxoimidazolidin-1-yl)ethyl] methacrylamide and methacrylic acid is considered as non developmental toxic according to the Regulation (EC) No 1272/2008 (CLP) and the Directive 67/548/EEC.