Registration Dossier

Administrative data

Description of key information

Subacute oral toxicity was tested in an OECD 422 using 1,4 butanediol dimethacrylate. The NOAEL is 300 mg/kg/d due to histopathological findings in the forestomach (males and females) (focal hyperplasia of the squamous epthelium of the forestomach associated with hyperkeratosis) and a reduced body weight and food consumption at 1000 mg/kg bw/d (see expert statement, Chapter 13.2). Further effects (e.g. reduced thymus weight, changes in clinical biochemistry) are considered to be stress-related due to the emerging Methacrylic acid leading to a reduced food consumption and consequently a reduced body weight. The increased kidney weight is considered as not adverse since there were microscopic changes or functional restrictions observed. The minimal degree of multifocal perilobular hepatocytic vacuolation observed in the liver of 3/10 high dose females is considered as normal background observation in animal studies and therefore not adverse.  No effects on the nervous system were observed.


1,4-Butanedioldimethacrylate will rapidly be hydrolysed by unspecific carboxylesterases in the liver into methacrylic acid and 1,4-butanediol (for details, see Chapter 7.1.1, toxicokinetics).


Since ECHA has recently rejected the read across approach, it was decided to conduct the OECD 408 with the registered substance. The dossier will be updated as soon as the results will become available to the Lead Registrant.

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
sub-chronic toxicity: oral
Data waiving:
other justification
Justification for data waiving:
other:
Justification for type of information:
The performance of the OECD 408/ Subchronic toxicity oral is in delay.
The toxicological endpoint requested by ECHA in the decision of October 2017 was initially addressed by read across to the metabolites, i.e. Methacrylic acid/ Methyl methacrylate and 1,4-Butanediol/ gamma Butyrolactone.
In the draft decision of May 2020, ECHA requested the performance of the study. In our comment, we argued to strengthen the read-across.
Meanwhile, we decided to perform the study since inconsistencies in the toxicological profiles of the parental ester and the alcohol metabolites (1,4-Butanediol and gamma butyrolactone) have been identified. Although the toxicological profile of the parental ester and the methacrylic metabolite is identical (see OECD 422 incl. Expert statement and OECD 416 with Methyl methacrylate), the differences in the toxicological profile of the parental ester and the alcohol metabolite cannot be explained since no data is available to support our hypothesis on different toxicokinetics.

The study will be commissioned as soon as possible.

The REACH dossier will be updated without undue delay once the study finalised.
For this issue, please also refer to the "Letter to ECHA" as attached in IUCLID Section 13.2..



Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 20 April to 25 June 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Identity: 1,4-Butanediol dimethacrylate
Alternative names: VISIOMER® 1.4 BDDMA
CAS number : 2082-81-7
EINECS number : 218-218-1
Appearance : Yellowish clear liquid
Appearance at first us : Colourless liquid
Storage conditions: Room temperature, protected from light
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Italy s.r.l., San Pietro al Natisone (UD), Italy. Animals were bred by Harlan Laboratories Netherland - Kreuzelweg 53, 5961 NM Horst P.O. Box 6174 NL 5960 AD Horst (Netherlands).
- Age at study initiation: 9 to 10 weeks (day of allocation)
- Weight at study initiation: Males 279-280g ; Females 214-216g (day of allocation)
- Housing: 5 sex/cage
- Diet (e.g. ad libitum): ad libitum excepted for clinical pathology
- Water (e.g. ad libitum): ad libitum excepted for clinical pathology
- Acclimation period: circa 3 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C +/-2°C
- Humidity (%): 55% +/- 15%
- Air changes (per hr): 15 to 20 air changes per hour
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: from 26 April (day of allocation) to 25 June 2012
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
The test item was administered orally by gavage at a dose volume of 5 mL/kg body weight. Control animals received the vehicle alone at the same dose volume.
The required amount of 1,4-Butanediol dimethacrylate was suspended in the vehicle (corn oil) at the final concentrations of 20, 60 and 200 mg/mL. The formulations were prepared daily and the concentrations were calculated and expressed in terms of test item as supplied.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Prior to commencement of treatment, analysis was performed to confirm that the proposed formulation procedure was acceptable (check of concentration and homogeneity).
The stability was found to be 24 hours at room temperature in the concentration range of 20 to 200 mg/mL.
Samples of the formulations prepared on Weeks 1 and 6 (when all females were present) were also analysed to check the concentration and homogeneity.
The overall results of the analyses were within the limits of acceptance stated in RTC’s SOPs for concentration (90-110%) and homogeneity (CV <10%).
Chemical analysis was carried out by the Analytical Chemistry Department at RTC according to a validated method (RTC Study No. 90770), in the range from 1.0 to 200 mg/mL. The software used for this activity was the Empower® Pro build No. 2154.
Duration of treatment / exposure:
Males
Animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing and thereafter through the day before necropsy.
Dose volumes were adjusted once per week for each animal according to the last recorded body weight.

Females
Animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing and thereafter during pairing (up to Day 40 of pairing for female no. 90760079 which did not mate), post coitum (up to Day 26 for non pregnant females) and post partum periods until Day 3 post partum. Dose volumes were adjusted once per week for each animal according to the last recorded body weight.
During the gestation period, dose volumes were calculated according to individual body weight on Days 0, 7, 14 and 20 post coitum and on Day 1 post partum. Thereafter individual dose volumes remained constant.
Frequency of treatment:
Once daily
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10 male and 10 female rats
Control animals:
yes
Details on study design:
This study design was in agreement with the procedures described in OECD Guideline no. 422 adopted on 22 March 1996 and subsequent revisions.
The test item was administered orally by gavage to parental animals at 5 mL/kg body weight.
The dosages, selected in consultation with the Sponsor, were 100, 300 and 1000 mg/kg/day.
The oral route was selected as it is a possible route of exposure of the test item in man.
Observations and examinations performed and frequency:
Mortality
Throughout the study, all animals were checked early in each working day and again in the afternoon. At weekends and Public Holidays a similar procedure was followed except that the final check was carried out at approximately mid-day.

Clinical signs
All clinical signs were recorded for individual animals.
Once before commencement of treatment and at least once daily during the study, each animal was observed and any clinical signs were recorded.
Observations were performed at the same time interval each day, the interval was selected taking into consideration the presence of post-dose reactions.

Clinical observations (Functional Observation Battery Tests)
Once before commencement of treatment and at least once a week thereafter, each animal was given a detailed clinical examination. Each animal was
removed from the home cage and observed in an open arena. The tests included observation of changes in gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypies or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, pupil size, unusual respiratory pattern).
All observations were recorded for individual animals.

Grip strength and sensory reactivity to stimuli
Once during the study, towards the end of treatment, 5 males and 5 females were randomly selected from each group for evaluation of sensory reactivity to stimuli of different modalities (e.g. auditory, visual and proprioceptive stimuli) and for assessment of grip strength. Measurements were performed using a computer generated random order. For males the tests were performed 4 days before necropsy and for females on Day 3 post partum.

Motor activity assessment (MA)
Once during the study, towards the end of treatment, 5 males and 5 females were randomly selected from each group and the motor activity was measured (for approximately 5 minutes) by an automated activity recording device. For males the tests were performed 4 days before necropsy and for females on Day 3 post partum.

Food consumption
The weight of food consumed by each cage of males and females was recorded weekly during the pre-mating period following allocation. The interval between allocation and treatment initiation was less than a full week. Individual food consumption for the females was measured on Days 7, 14 and 20 post coitum starting from Day 0 post coitum and on Day 4 post partum starting from Day 1 post partum.

Body weight
Males were weighed on the day of allocation to treatment groups, on the day of treatment commencement, weekly thereafter and at termination. Females were weighed on the day of allocation to treatment groups, on the day of treatment commencement, weekly thereafter until pairing and on Days 0, 7, 14 and 20 post coitum. Dams were also weighed on Days 1 and 4 post partum.

Clinical pathology investigations
As a part of the sacrificial procedure, samples of blood were withdrawn under isofluorane anaesthesia from the abdominal vena cava from 5 males and 5 females (all females with viable litters with the exception of one not pregnant female of Group 4) randomly selected from each group, under condition of food deprivation.
The blood samples collected were divided into tubes as follows:
EDTA anticoagulant for haematological investigations
Heparin anticoagulant for biochemical tests
Citrate anticoagulant for coagulation tests

Haematology
Haematocrit
Haemoglobin
Red blood cell count
Reticulocyte count
Mean red blood cell volume
Mean corpuscular haemoglobin
Mean corpuscular haemoglobin concentration
White blood cell count
Differential leucocyte count
- Neutrophils
- Lymphocytes
- Eosinophils
- Basophils
- Monocytes
- Large unstained cells
Platelets
Coagulation tests
Prothrombin time

Clinical chemistry
Alkaline phosphatase
Alanine aminotransferase
Aspartate aminotransferase
Gamma-glutamyltransferase
Urea
Creatinine
Glucose
Triglycerides
Bile acids
Phosphorus
Total bilirubin
Total cholesterol
Total protein
Albumin
Globulin
A/G Ratio
Sodium
Potassium
Calcium
Chloride

Urinalysis (Only males)
At the same time interval as the clinical pathology investigations, individual overnight urine samples were also collected from the same animals under the same conditions. Before starting urine collection, water bottles were removed from each cage and each animal received approximately 10 mL/kg of drinking water by gavage, in order to obtain urine samples suitable for analysis.

Appearance
Volume
Specific gravity
pH
Protein
Glucose
Ketones
Bilirubin
Urobilinogen
Blood

The sediment, obtained from centrifugation at approximately 3000 rpm for 10 minutes, was examined microscopically for:

Epithelial cells
Leucocytes
Erythrocytes
Crystals
Spermatozoa and precursors
Other abnormal components
Sacrifice and pathology:
Parental males:
The males were killed after the mating of all females (after 33 days of treatment).

Parental females:
The females with live pups were killed on Day 4 post partum.
The clinical history of the males and females of the parental generation was studied and a detailed post mortem examination was conducted (including examination of the external surface and orifices).
Changes were noted, the requisite organs weighed and the required tissue samples preserved in fixative and processed for histopathological examination.

Females:
All females were examined also for the following:
a) external and internal abnormalities;
b) number of visible implantation sites (pregnant animals);
c) number of corpora lutea (pregnant animals).
Organ weights

From all animals completing the scheduled test period, the organs were dissected free of fat and weighed.
The ratios of organ weight to body weight were calculated for each animal.

Tissues fixed and preserved

Samples of all the tissues listed in Annex 1 were fixed and preserved in 10% neutral buffered formalin (except testes and epididymides which were fixed in modified Davidson's fluid and preserved in 70% ethyl alcohol).

Histopathological examination

After dehydration and embedding in paraffin wax, sections of the tissues were cut at 5 micrometer thickness and stained with haematoxylin and eosin.
In addition, the testes and epididymides were cut at 2-3 micrometer thickness and stained with Periodic Acid Schiff (PAS). The morphological evaluationof the seminiferous epithelium (staging of spermatogenic cycle) was performed.

In the first instance the examination was restricted as detailed below:

a) Tissues from 5 males and 5 females randomly selected (animals evaluated for clinical pathology) in the control and high dose group.
b) All abnormalities in all groups.
The examination of stomach (both sexes) and liver (only females) was extended to include the remaining 5 males and 5 females (animals not evaluated for clinical pathology) of the control and high dose group.
In addition, the examination of stomach (both sexes) and liver (only females) was extended to animals of the other dose groups since possible treatment-related changes were observed in the high dose group.



Statistics:
Standard deviations were calculated as appropriate. For continuous variables the significance of the differences amongst group means was assessed by Dunnett’s test or a modified t test, depending on the homogeneity of data.
Statistical analysis of histopathological findings was carried out by means of the non-parametric Kolmogorov-Smirnov test (for stomach and liver).
The non-parametric Kruskal-Wallis analysis of variance was used for the other parameters. Intergroup differences between the control and treated groups were assessed by the non-parametric version of the Williams test.
The criterion for statistical significance was p<0.05 or p<0.01.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weight and body weight gain were lower in the high dose group compared to controls throughout the study.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Food consumption was reduced in the high dose group compared to controls.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
The main relevant change was an increased value of bile acids in treated groups compared to controls with a clear dose-relation in males.
Urinalysis findings:
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Treatment-related findings were limited to the high dosed animals and were seen in the stomach of both sexes and in the liver of the females only.
Histopathological findings: neoplastic:
no effects observed
Other effects:
not examined
Details on results:
Mortality and fate of females
No mortality occurred in the study.
A total of 8 females were found not pregnant at necropsy: one each in the control and low dose groups and 6 in the high dose group.
The number of females with live pups on Day 4 post-partum were: 9 in the control group, 9 in the low dose group, 10 in the mid-dose group and 4 in the high dose group.

Clinical signs and neurotoxicity assessment (Functional Observation Battery Tests)
No clinical signs of toxicological significance were reported.

Body weight and body weight gain
Body weight and body weight gain were lower in the high dose group compared to controls throughout the study.

Food consumption
Food consumption was reduced in the high dose group compared to controls.

Motor activity and sensory reaction to stimuli
No relevant differences were noted in all parameters investigated between control and treated groups.

Haematology
No changes of toxicological significance were found.

Clinical chemistry
The main relevant change was an increased value of bile acids in treated groups compared to controls with a clear dose-relation in males.

Urinalysis
No changes were recorded.

Oestrus cycle, reproductive parameters, pairing combination and mating performance
Measurements of oestrus cycle, pre-coital intervals and copulatory index did not show differences between treated and control groups. On the
contrary, fertility index was markedly reduced in the high dose group (40% compared to 90% of the control group).

Implantation, pre-birth loss data and gestation length of females
No significant differences were observed in the number of implantation, corpora lutea, total litter size, pre-implantation loss, pre-birth loss and
gestation length between control and treated groups.

Litter data and sex ratios
Reduced litter and mean pup weights were found in the high dose group compared to controls. The percentage of cumulative pup loss on Day 4 post partum starting from the total litter size at birth, was increased in the high dose group.
No differences were found in sex ratio between the groups.

Clinical signs of pups
Small pups were generally observed in all groups including the control group. Cold to touch and apparently no food intake were the signs noted in pups of the treated groups only.

Necropsy findings in decedent pups and in pups sacrificed on Day 4 post partum
No relevant differences were recorded in decedent pups between the groups.
No abnormalities were observed in pups sacrificed at term.

Terminal body weight and organ weights
Terminal body weight was lower in the high dose group compared to controls and this difference was statistically significant in females.
Statistically significant higher kidneys weight was observed in high dose males and females compared to controls. In addition, thymus weight was significantly decreased in high dose males.

Macroscopic observations
No treatment-related changes were noted at macroscopic examination.

Microscopic observations
Treatment-related findings were limited to the high dosed animals and were seen in the stomach of both sexes and in the liver of the females only.
Stomach (non-glandular): The treatment-related change seen in the high dosed animals (1/10 and 5/10, respectively in males and females), consisted of mild diffused hyperplasia of the squamous epithelium in the non-glandural stomach, which was associated with mild thickening (i.e., hyperkerathosis) of the keratin layer. This change was not associated with any indication of inflammation and/or ulceration
Liver: In 3/10 high dose animals (females), minimal degree of multifocal perilobular hepatocytic vacuolaiton, which is suggested to be consistent with fatty change, was noted. The vacuoles were of mixed type (i.e., micro- and macrovesicular) and no presence of inflammation and/or necrosis was noted.
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
food consumption and compound intake
gross pathology
Critical effects observed:
yes
Lowest effective dose / conc.:
1 000 mg/kg bw/day (nominal)
System:
gastrointestinal tract
Organ:
stomach
Treatment related:
yes
Dose response relationship:
no
Relevant for humans:
no
Conclusions:
On the basis of the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) could be considered 300 mg/kg/day for males and females.
Executive summary:

In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test according to OECD Guideline 422 (22 March 1996) 1,4-Butanediol dimethacrylate was administered to 10 Hsd: Sprague Dawley SD rats/sex/dose orally by gavage at dose levels of 0 (control), 100, 300 and 1000 mg/kg bw/d. The treatment schedule included 2 weeks before pairing, during pairing, post coitum and post partum periods up to day 3 post partum. Animals were administered for approximately 5 and 8 weeks for males and females, respectively.


 


No mortality occurred in the study. A total of 8 females were found not pregnant at necropsy: one each in the control and low dose groups and 6 in the high dose group.
The number of females with live pups on Day 4 post-partum were: 9 in the control group, 9 in the low dose group, 10 in the mid-dose group and 4 in the high dose group.
No clinical signs of toxicological significance were reported.


Body weight and body weight gain were lower in the high dose group compared to controls throughout the study. 
Food consumption was reduced in the high dose group compared to controls. No relevant differences were noted in all parameters investigated between control and treated groups. In accordance with the expert statement, the reduced food consumption and the body weight loss is a direct consequence of the emerging Methacrylic acid (see Chapter 7.1.1. Toxicokinetics and expert statement, Chapter 13.2). 
The main relevant change was an increased value of bile acids in treated groups compared to controls with a clear dose-relation in males. 
No changes were recorded in urinalysis.
Measurements of oestrus cycle, pre-coital intervals and copulatory index did not show differences between treated and control groups. On the contrary, fertility index was markedly reduced in the high dose group (40% compared to 90% of the control group). In accordance with the expert statement, the reduced fertility index is the direct of consequence of maternal toxicity due to the markedly reduced body weight/ food consumption. The markedly reduced body weight leads to the interruption of the Leptin-Ghrelin-Kisspetin-GnRH hormon axis (see expert statement, Chapter 13.2). 


No significant differences were observed in the number of implantations, corpora lutea, total litter size, pre-implantation loss, pre-birth loss and gestation length between control and treated groups.
Reduced litter and mean pup weights were found in the high dose group compared to controls. The percentage of cumulative pup loss on Day 4 post partum starting from the total litter size at birth, was increased in the high dose group. The reduced litter and mean pup weight is also a secondary effect of the reduced maternal body weight and pup's survial is also directly linked with their general condition after birth. No differences were found in sex ratio between the groups.
Small pups were generally observed in all groups including the control group. Cold to touch and apparently no food intake were the signs noted in pups of the treated groups only.
No relevant differences were recorded in decedent pups between the groups.
No abnormalities were observed in pups sacrificed at term.
Terminal body weight was lower in the high dose group compared to controls and this difference was statistically significant in females. The significantly reduced body weight in females leads to the interruption of the Leptin-Ghrelin-Kisspetin-GnRH hormon axis. This is well-known and widely described in the literature.  
Statistically significant higher kidneys weight was observed in high dose males and females compared to controls. In addition, thymus weight was significantly decreased in high dose males. The higher kidney weights are considered to the consequence of the high volume intake due to the gavage administration (10 ml/kg bw) and not adverse since they are not accompanied by microscopic changes. The reduced thymus weight (thymus atrophy) is considered to be stress-related due to the hyperplasia in the forestomach and consequently reduced food consumption and body weight (see expert statement, Chapter 13.2).
No treatment-related changes were noted at macroscopic examination.
Treatment-related findings at microscopic observations were limited to the high dosed animals and were seen in the stomach of both sexes and in the liver of the females only. The treatment-related change seen in the high dosed animals (1/10 and 5/10, respectively in males and females), consisted of focal hyperplasia of the squamous epithelium in the non-glandular stomach, which was associated with thickening (i.e., hyperkeratosis) of the keratin layer. This change was not associated with any indication of inflammation and/or ulceration.


The findings in the forestomach of the animals are considered as main result of the study, treatment-related and adverse. The reason for the adverse alteration in the forestomach is the emerging Methacrylic acid due to the well-known, well-described and widely accepted metabolism of Methacrylates. (see Chapter 7.1.1., Toxicokinetics, see expert statement, Chapter 13.2, see CLH proposal of Finnish MSCA, Chapter 13.2).


 


In 3/10 high dose animals (females), minimal degree of multifocal perilobular hepatocytic vacuolation, which is suggested to be consistent with fatty change, was noted. The vacuoles were of mixed type (i.e., micro- and macrovesicular) and no presence of inflammation and/or necrosis was noted. The minimal degree of multifocal perilobular hepatocytic vacuolation in the liver of females which is not accompanied with necrosis or functional restrictions is considered to be not adverse since it is part of the normal background changes in the liver observed in animal studies (see expert statement, Chapter 13.2). 




Evaluation of the spermatogenic cycle did not show differences between the groups. Regular layering in the germinal epithelium was noted.


 


Conclusions


On the basis of the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) for both general toxicity could be considered 300 mg/kg bw/d for both sexes. The NOAEL on reproduction/developmental toxicity could be considered 1000 mg/kg/day.


 


NOTE: Any of data in this dataset are disseminated by the European Union on a right-to-know basis and this is not a publication in the same sense as a book or an article in a journal. The right of ownership in any part of this information is reserved by the data owner(s). The use of this information for any other, e.g. commercial purpose is strictly reserved to the data owners and those persons or legal entities having paid the respective access fee for the intended purpose. 

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
300 mg/kg bw/day
Study duration:
subacute
Species:
rat
Organ:
stomach

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

For 1,4-BDDMA, a reliable (RL=1), relevant and adequate study is available on repeated dose toxicity:


 In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test according to OECD Guideline 422 (22 March 1996) 1,4-Butanediol dimethacrylate was administered to 10 Hsd: Sprague Dawley SD rats/sex/dose orally by gavage at dose levels of 0 (control), 100, 300 and 1000 mg/kg bw/d. The treatment schedule included 2 weeks before pairing, during pairing, post coitum and post partum periods up to day 3 post partum. Animals were administered for approximately 5 and 8 weeks for males and females, respectively.


 


No mortality occurred in the study. A total of 8 females were found not pregnant at necropsy: one each in the control and low dose groups and 6 in the high dose group.
The number of females with live pups on Day 4 post-partum were: 9 in the control group, 9 in the low dose group, 10 in the mid-dose group and 4 in the high dose group.
No clinical signs of toxicological significance were reported.


Body weight and body weight gain were lower in the high dose group compared to controls throughout the study. 
Food consumption was reduced in the high dose group compared to controls. No relevant differences were noted in all parameters investigated between control and treated groups. In accordance with the expert statement, the reduced food consumption and the body weight loss is a direct consequence of the emerging Methacrylic acid (see Chapter 7.1.1. Toxicokinetics and expert statement, Chapter 13.2). 
The main relevant change was an increased value of bile acids in treated groups compared to controls with a clear dose-relation in males. 
No changes were recorded in urinalysis.
Measurements of oestrus cycle, pre-coital intervals and copulatory index did not show differences between treated and control groups. On the contrary, fertility index was markedly reduced in the high dose group (40% compared to 90% of the control group). In accordance with the expert statement, the reduced fertility index is the direct of consequence of maternal toxicity due to the markedly reduced body weight/ food consumption. The markedly reduced body weight leads to the interruption of the Leptin-Ghrelin-Kisspetin-GnRH hormon axis (see expert statement, Chapter 13.2). 


No significant differences were observed in the number of implantation, corpora lutea, total litter size, pre-implantation loss, pre-birth loss and gestation length between control and treated groups.
Reduced litter and mean pup weights were found in the high dose group compared to controls. The percentage of cumulative pup loss on Day 4 post partum starting from the total litter size at birth, was increased in the high dose group. The reduced litter and mean pup weight is also a secondary effect of the reduced maternal body weight and pup's survial is also directly linked with their general condition after birth. No differences were found in sex ratio between the groups.
Small pups were generally observed in all groups including the control group. Cold to touch and apparently no food intake were the signs noted in pups of the treated groups only.
No relevant differences were recorded in decedent pups between the groups.
No abnormalities were observed in pups sacrificed at term.
Terminal body weight was lower in the high dose group compared to controls and this difference was statistically significant in females. The significantly reduced body weight in females leads to the interruption of the Leptin-Ghrelin-Kisspetin-GnRH hormon axis. This is well-known and widely described in the literature.  
Statistically significant higher kidneys weight was observed in high dose males and females compared to controls. In addition, thymus weight was significantly decreased in high dose males. The higher kidney weights are considered to the consequence of the high volume intake due to the gavage administration (10 ml/kg bw) and not adverse since they are not accompanied by microscopic changes. The reduced thymus weight (thymus atrophy) is considered to be stress-related due to the hyperplasia in the forestomach and consequently reduced food consumption and body weight (see expert statement, Chapter 13.2).
No treatment-related changes were noted at macroscopic examination.
Treatment-related findings at microscopic observations were limited to the high dosed animals and were seen in the stomach of both sexes and in the liver of the females only. The treatment-related change seen in the high dosed animals (1/10 and 5/10, respectively in males and females), consisted of focal hyperplasia of the squamous epithelium in the non-glandular stomach, which was associated with thickening (i.e., hyperkeratosis) of the keratin layer. This change was not associated with any indication of inflammation and/or ulceration.


The findings in the forestomach of the animals are considered as main result of the study, treatment-related and adverse. The reason for the adverse alteration in the forestomach is the emerging Methacrylic acid due to the well-known, well-described and widely accepted metabolism of Methacrylates. (see Chapter 7.1.1., Toxicokinetics, see expert statement, Chapter 13.2, see CLH proposal of Finnish MSCA, Chapter 13.2).


 


In 3/10 high dose animals (females), minimal degree of multifocal perilobular hepatocytic vacuolation, which is suggested to be consistent with fatty change, was noted. The vacuoles were of mixed type (i.e., micro- and macrovesicular) and no presence of inflammation and/or necrosis was noted. The minimal degree of multifocal perilobular hepatocytic vacuolation in the liver of females which is not accompanied with necrosis or functional restrictions is considered to be not adverse since it is part of the normal background changes in the liver observed in animal studies (see expert statement, Chapter 13.2). 




Evaluation of the spermatogenic cycle did not show differences between the groups. Regular layering in the germinal epithelium was noted.


 


Conclusions


On the basis of the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) for both general toxicity could be considered 300 mg/kg bw/d for both sexes. The NOAEL on reproduction/developmental toxicity could be considered 1000 mg/kg/day.


 


 


 


The 90d-study is in progress and the dossier will be update accordingly as soon as the results will become available to the lead registrant.


 


There are no data gaps for the endpoint repeated dose toxicity. No human data are available. However, there is no reason to believe that these results from rat would not be applicable to humans.



Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
OECD guideline 422 study, no deviations, GLP


 


Compliance to REACh requirements


The requirements are covered with an OECD 422 oral rat study with the substance itself, and subchronic/ chronic study is in progress. The mentioned study is reliable (Reliability 1).

Justification for classification or non-classification

Based on the available data, 1,4-BDDMA does not need to be classified for repeated dose toxicity according to the criteria given in regulation (EC) 1272/2008 or the former European directive on classification and labelling 67/548/EEC. Thus, no labelling is required.