Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1 June 2012 to 21 July 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)
Version / remarks:
adopted 22 March 1996.
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Specific details on test material used for the study:
Identity : Benzyl methacrylate
Alternative names : VISIOMER® BNMA; Benzylmethacrylat
CAS number : 2495-37-6
EINECS number : 219-674-4
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Sprague Dawley [Crl:CD(SD)BR] rats from Charles River Italia S.p.A., Calco (Lecco), Italy
- Age at study initiation: 7 to 8 weeks old and weighing 211-232 g for males and 135-199 g for females, at receipt and approximately 10 weeks at study initiation.
- Weight at study initiation: (P) Males: 335g-338g; Females: 233g-238 g
- Fasting period before study: no
- Housing: 5 sex/cage
- Diet (e.g. ad libitum): ad libitum except for clinical pathology investigation
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 14 days

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

IN-LIFE DATES: From: To: 24 May 2012 to 21 July 2012
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was administered orally by gavage and the formulations were prepared daily.

DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:

VEHICLE
- Justification for use and choice of vehicle (if other than water): solubility of test item
- Concentration in vehicle: 10, 35 and 100 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg body weight
- Lot/batch no. (if required): ----
- Purity: ----
Details on mating procedure:
Mating was monogamous (one male to one female). A vaginal smear was taken from the day after the start of pairing until positive identification of copulation (sperm identification, vaginal plug in situ or copulation plugs found in the cage tray).
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 (content check and homogeneity). Results of the analyses were within the limits of acceptance.
The stability was found to be 24 hours at room temperature in the concentration range of 10 to 100 mg/mL.
Samples of the formulations prepared on Week 1 and last Week were also analysed to check the concentration and homogeneity. Results of the analyses were within the limits of acceptance.
Chemical analysis was carried out by the Analytical Chemistry Department at RTC according to a validated method (RTC Study No. 90820), in the range from 1 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, post coitum and post partum periods until Day 3 post partum (the day before sacrifice).

Dose volumes were adjusted once per week for each animal according to the last recorded body weight.
During the post coitum 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 a day
Dose / conc.:
50 mg/kg bw/day (nominal)
Dose / conc.:
175 mg/kg bw/day (nominal)
Dose / conc.:
500 mg/kg bw/day (nominal)
No. of animals per sex per dose:
3 groups of 10 males and 10 females received the test item at the dose levels of 50, 175 and 500 mg/kg/day. A similarly constituted group of
animals received the vehicle alone (corn oil) and acted as control.
Control animals:
yes, concurrent vehicle
Details on study design:
The oral route was selected as it is a possible route of exposure of the test item in man.
Dose levels of 50, 175 and 500 mg/kg/day were selected by the Sponsor based on information from previous non GLP compliant studies (RTC Study nos.: 90830EXT and 90840EXT).
Positive control:
no
Parental animals: Observations and examinations:
Mortality

Throughout the study, all animals were checked early in each working day in the morning and 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

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 approximately 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 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); an assessment of grip strength was also performed. For males the tests were performed on Day 29 of the study 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 (at least 5 minutes) by an automated activity recording device. Measurements were performed using a computer generated random order. For males the tests were performed on Day 29 of the study and for females on Day 3 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.

Food consumption

The weight of food consumed by each cage of males and females was recorded weekly, where possible, during the pre-mating period following allocation. 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.

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 (females with viable litters) 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

The measurements performed on blood samples are listed below:

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







Oestrous cyclicity (parental animals):
Vaginal smears

Vaginal smears were taken daily in the morning starting two weeks before pairing until a positive identification of copulation was made. The vaginal smear data were examined to determine the following:

a) anomalies of the oestrous cycle;
b) pre-coital interval (i.e., the number of nights paired prior to the detection of mating).

Sperm parameters (parental animals):
Parameters examined in [P] male parental generation:
[testis weight, epididymis weight, morphological evaluation of the seminiferous epithelium (staging of spermatogenic cycle) ]
Litter observations:
Parturition and gestation length

A parturition check was performed from Day 20 to Day 25 post coitum.
Gestation length was calculated as the time between the day of successful mating (Day 0 post coitum) and the day of commencement of birth (i.e. first detected presence of offspring in the cage). The day that offspring were first detected in the cage was considered Day 0 post partum.

Pups identification, weight and observation

As soon as possible, after parturition was considered complete (Days 0 or 1 post partum), all pups (live and dead) were counted, sexed and live pups were
identified.
Live pups were individually weighed on Days 1 and 4 post partum.
Pups dying during the lactation period were weighed before the despatch to necropsy.
Observation was performed once daily for all litters.
Postmortem examinations (parental animals):
Parental animals were killed by exsanguination under isofluorane anaesthesia. All animals were subjected to necropsy.

Parental males:
Males were sacrificed after completion of the mating period 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 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).
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 groups killed at term.
b) All abnormalities in all groups











Postmortem examinations (offspring):
Pups were sacrificed by intraperitoneal injection of Sodium Thiopental
All pups found dead in the cage were examined for external and internal abnormalities.
All live pups sacrificed at termination were killed and examined for external abnormalities and sex confirmation by gonadal inspection.
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.
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 mean values, standard deviations and statistical analysis were calculated from actual values in the computer without rounding off.

Reproductive indices:
The following reproductive indices were calculated:

Males
Copulatory Index (%) = no. of animals mated/no. of animals paired x 100
Fertility Index (%)= no. of males which induced pregnancy/no. of animals paired x 100

Females
Copulatory Index (%) = no. of animals mated/no. of animals pairedx 100
Fertility Index (%) = no. of pregnant females/no. of females paired x 100

Males and Females

Pre-coital Interval = Mean number of days between pairing and mating

Sex ratios were calculated at birth and on Day 4 post partum and are presented as the percentage of males per litter.
Offspring viability indices:
Females
Pre-birth loss was calculated as a percentage from the formula:

(No. of visible implantations-total litter size at birth)/No. of visible implantations x 100

Pup loss at birth was calculated as a percentage from the formula:

(Total litter size-live litter size)/Total litter size x 100

Cumulative pup loss on Day 4 post partum was calculated as a percentage from the formula:

(Total litter size at birth-live litter size at Day 4 post partum)/Total litter size at birth x 100

Pre- implantation loss was calculated as a percentage from the formula:

(no. of corpora lutea - no. of implantations)/ no. of corpora lutea x 100
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
no effects observed
Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
- CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
No mortality occurred throughout the study.
All females proved to be pregnant.
The number of females with live pups on Day 4 post partum was 10 both in the control and in all treated groups.

Clinical observations (Functional Observation Battery Tests) - (Table 2)
Neurotoxicity assessment (removal of animals from the home cage and open arena)
Observation of animals at removal from the cage and in an open arena (neurotoxicity assessment) did not reveal changes attributable to the test item.

- BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Means of body weight and body weight gain were, in general, comparable between control and treated groups both in males and females throughout the
study.
No relevant differences in food consumption were observed in treated animals of both sexes during the study.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
Oestrus cycle and reproductive parameters (pre-coital intervals, copulatory and fertility indices) were similar in treated and control groups

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Oestrus cycle and reproductive parameters (pre-coital intervals, copulatory and fertility indices) were similar in treated and control groups
Implantation, pre-implantation loss data, pre-birth loss data and gestation length of females: No significant differences were observed in treated and control groups for these parameters. All dams gave birth between Days 21 and 23 post coitum

ORGAN WEIGHTS (PARENTAL ANIMALS)
Terminal body weight was unaffected by treatment in both sexes.
Changes noted in the absolute and relative organ weights were of low magnitude and no histological findings were observed at microscopic examination, therefore they were considered of no toxicological relevance.

GROSS PATHOLOGY (PARENTAL ANIMALS)
No treatment-related changes were noted

HISTOPATHOLOGY (PARENTAL ANIMALS)
No treatment-related changes were noted.
In addition seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and no alterations were noted.

OTHER FINDINGS (PARENTAL ANIMALS)
Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Critical effects observed:
no
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
no effects observed
Gross pathological findings:
no effects observed
Litter data at birth, on Day 1 and on Day 4 post partum of females and sex ratio of pups: Litter data parameters and sex ratios at birth and on
Day 4 post partum were similar between groups.

Clinical signs of pups: Pre-weaning clinical signs were comparable between treated and control groups.

Necropsy findings in decedent pups and in pups sacrificed on Day 4 post partum: Necropsy findings in decedent pups and in pups sacrificed on
Day 4 post partum did not reveal any treatment-related effect.

Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Critical effects observed:
not specified
Reproductive effects observed:
not specified
Conclusions:
On the basis of the results obtained in this study, 500 mg/kg/day could be considered the NOAEL (No Observed Adverse Effect Level) for animals of both sexes.
Executive summary:

SUMMARY

 Study design

The toxic effects on rats of both sexes after repeated oral dosing withBenzyl methacrylate, as well as on effects of the test item on male and female reproductive performance, such as gonadal function, conception, parturition and early lactation of the offspring were investigated. The vehicle was corn oil. All doses were administered at a constant volume of 5 mL/kg body weight.

 

Group

Number

Treatment

(mg/kg/day)

Number of animals

1

2

3

4

0

50

175

500

10M+10F

10M+10F

10M+10F

10M+10F

Males were treated for 2 weeks prior to pairing and during pairing with females until the day before necropsy, for a total of 33 days. Females were treated for 2 weeks prior to pairing, during pairing and throughout the gestation and lactation periods until Day 3post partum.

The following investigations were performed in all groups: body weight, clinical signs (including neurotoxicity assessment, motor activity and sensory reaction to stimuli), food consumption, oestrous cycle, mating performance, clinical pathology investigations (haematology and clinical chemistry), litter data, macroscopic observations, organ weights and histopathological examination.

Clinical signs and macroscopic observations of pups were also performed. The histopathological examination was performed only on control and high dose groups (five animals/sex/group selected randomly). It included identification of the stages of the spermatogenic cycle in five males.

 Mortality and fate of females

 No mortality occurred throughout the study.

All females proved to be pregnant. The number of females with live pups on Day 4post partumwas 10 both in the control and in all treated groups.

 

Clinical signsand clinical observations (Functional ObservationBatteryTests)

 

No relevant clinical signs were observed throughout the study in all treated animals of both sexes.

Neurotoxicity assessment (removal of animals from the home cage and open arena)

Observation of animals at removal from the cage and in an open arena (neurotoxicity assessment) did not reveal changes attributable to the test item.

 

Body weight and body weight gain

No relevant differences in body weights were recorded in animals of both sexes compared to the control group, throughout the study.

 

Food consumption

 Food consumption was, in general, comparable between control and treated groups.

 

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 recorded.

No significant change was recorded in the coagulation test.

 

 Clinical chemistry

 The alteration (decrease in cholesterol level) noted in males receiving 500 mg/kg/day could not be conclusively attributed to treatment since it was observed in only one sex.

 

  Oestrus cycle, reproductive parameters, pairing combination and mating performance

 Oestrus cycle, pre-coital intervals, copulatory index and fertility index did not show intergroup differences.

 

 Implantation, pre-implantation loss data, pre-birth loss data and gestation length of females

  No significant differences were observed in treated and control groups for these parameters.

All dams gave birth between Days 21 and 23post coitum.

 

Litter data at birth, on Day 1 and on Day 4post partumof females and sex ratio of pups

 Litter data parameters and sex ratios at birth and on Day 4post partumwere similar between groups.

 

Clinical signs of pups

  Pre-weaning clinical signs were comparable between treated and control groups.

 

Necropsy findings in decedent pups and in pups sacrificed on Day 4post partum

 Necropsy findings in decedent pups and in pups sacrificed on Day 4post partumdid not reveal any treatment-related effect.

 

Terminal body weight and organ weights

 Terminal body weight was unaffected by treatment in both sexes.

Changes noted in the absolute and relative organ weights were of low magnitude and no histological findings were observed at microscopic examination, therefore they were considered of no toxicological relevance.

 

Macroscopic observations

  No treatment-related changes were noted.

 

Microscopic observations

  No treatment-related changes were noted.

In addition seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and no alterations were noted.

 

Conclusions

 On the basis of the results obtained in this study, 500 mg/kg/day could be considered the NOAEL (No Observed Adverse Effect Level) for animals of both sexes.

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.

 

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
500 mg/kg bw/day
Study duration:
subacute
Species:
rat
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

For BENZYL METHACRYLATE, a reliable (RL=1), relevant and adequate study is available on toxicity to reproduction:

In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test according to OECD Guideline 422 (22 March 1996) BENZYL METHACRYLATE was administered to 10 Sprague Dawley [Crl:CD(SD)BR] rats/sex/dose orally by gavage at dose levels of 0 (control), 50, 175 and 500 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. No clinical signs of toxicological significance were reported. No relevant differences in body weights and food consumption were recorded in animals of both sexes compared to the control group, throughout the study.

No relevant differences were noted in motor activity and sensory reaction to stimuli between control and treated groups. Haematology and urinalysis revealed no changes of toxicological significance.

A decrease in cholesterol levels noted in males receiving 500 mg/kg bw/day could not be conclusively attributed to treatment since it was observed in only one sex.

Measurements of oestrus cycle, pre-coital intervals and copulatory index did not show differences between treated and control groups. 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. All dams gave birth between Days 21 and 23 post coitum.

Litter data parameters and sex ratios at birth and on Day 4 post-partum were similar between groups. Pre-weaning clinical signs were comparable between treated and control groups. Necropsy findings in decedent pups and in pups sacrificed on Day 4 post-partum did not reveal any treatment-related effect.

At necropsy, changes noted in the absolute and relative organ weights were of low magnitude and no histological findings were observed at microscopic examination, therefore they were considered to be of no toxicological relevance.

No treatment-related macroscopic or microscopic changes were noted. In addition, seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and no alterations were noted.

No relevant toxic effects were seen in parental animals up to the highest dose group of 500 mg/kg bw/d. Based on the results obtained in the study, the NOAEL for both, general toxicity and reproduction/developmental toxicity was 500 mg/kg bw/d (males/females). 

 In conclusion, based on studies in experimental animals, there is no evidence for impairment of fertility as a result of exposure to Benzyl methacrylate.

 

Short description of key information:

In an OECD Guideline 422 and GLP study with Benzyl methacrylate, the NOAEL for reproductive toxicity is 500 mg/kg/day for all relevant endpoints, which was the highest dose tested. The derived NOAEL is based on the absence of any effect in dams or pups. Justification for selection of Effect on fertility via oral route: OECD guideline 422 study, no deviations, GLP


Justification for selection of Effect on fertility via oral route:
OECD guideline 422 study, no deviations, GLP

Effects on developmental toxicity

Description of key information

In a reproductive and developmental toxicity study according to OECD guideline 422 (GLP conform) with Benzyl methacrylate in rats, the NOAEL for development of offspring was at the highest test dose of 500 mg/kg/day.

Furthermore, the absence of a teratogenic potential was also demonstrated for the primary metabolites, MAA and Benzyl alcohol.

For MMA as source substance of MAA, there was no evidence for critical effects on developmental toxicity or teratogenicity in animal studies. The developmental toxicity has also been extensively analysed in studies with the other primary metabolite, benzyl alcohol and its metabolites benzoic acid and its salts. Teratogenic effects were reported only when severe effects on maternal toxicity were observed.

In conclusion, based on studies in experimental animals, there is no evidence for toxicity of Benzyl methacrylate to the reproductive system.

Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1 June 2012 to 21 July 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
other: OECD 422
Version / remarks:
adopted 22 March 1996
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Specific details on test material used for the study:
Identity : Benzyl methacrylate
Alternative names : VISIOMER® BNMA; Benzylmethacrylat
CAS number : 2495-37-6
EINECS number : 219-674-4
Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Italy S.p.A., Calco (Lecco), Italy

- Age at study initiation: (P) Males/females: approximately 8 - 9 wks
- Weight at study initiation: (P) Males: 196.5 - 204.7 g; Females: 166.1 - 189.3 g

- Fasting period before study:
- Housing: Pre mating period: no more than 5 per cage in clear polycarbonate cages measuring 59X39X20 cm with a stainless steel
mesh lid and floor (Techniplast - Gazzada S.a.r.l., Buguggiate, Varese). Each cage tray will hold absorbent material which will be
inspected daily and changed at least three times a week.
During mating period: 1 male to one female per cage in clear polycarbonate cages measuring 36X19X24 cm with a stainless steel
mesh lid and floor (Techniplast - Gazzada S.a.r.l., Buguggiate, Varese). Each cage tray will hold absorbent material which will be
inspected daily.
Pregnant females: will be transferred to individual cages after mating: solid bottomed, breeding cages (Techniplast - Gazzada S.a.r.l.,
Buguggiate, Varese), for the gestation period, birth and lactation.
Suitable nesting material will be provided and will be changed as necessary.
- Diet: ad libitum, commercially available laboratory rodent diet (4 RF 21, Mucedola S.r.l., Via G. Galilei, 4,
20019, Settimo Milanese (MI), Italy)
- Water: ad libitum, supplied via water bottles

- Acclimation period: aproximately 14 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ±2 °C
- Humidity (%): 55 ±15 %
- Air changes (per hr): 15 - 20 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hrs dark/ 12 hrs light
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was administered orally by gavage and the formulations were prepared daily.

DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:

VEHICLE
- Justification for use and choice of vehicle (if other than water): solubility of test item
- Concentration in vehicle: 10, 35 and 100 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg body weight
- Lot/batch no. (if required): ----
- Purity: ----
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Once during week 1 and week 6 of treatment, samples of prepared formulations were analysed for verification of concentration.
Details on mating procedure:
Mating was monogamous (one male to one female). A vaginal smear was taken from the
day after the start of pairing until positive identification of copulation (sperm identification,
vaginal plug in situ or copulation plugs found in the cage tray).
The female was paired with the same male until positive identification occurred.
Duration of treatment / exposure:
Males
Animals were dosed once a day, 7 days a week, for a minimum of 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 a minimum of 2 consecutive weeks prior
to pairing and thereafter during pairing, post coitum and post partum periods until Day 3
post partum or the day before sacrifice.
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.:
50 mg/kg bw/day (nominal)
Dose / conc.:
175 mg/kg bw/day (nominal)
Dose / conc.:
500 mg/kg bw/day (nominal)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
The oral route was selected as it is a possible route of exposure of the test item in man.
Dose levels of 50, 175 and 500 mg/kg/day were selected by the Sponsor based on information from previous non GLP compliant studies (RTC
Study nos.: 90830EXT and 90840EXT).
Maternal examinations:
yes
Ovaries and uterine content:
yes
Fetal examinations:
yes
Statistics:
Standard deviations were calculated as appropriate. For continuous variables the
significance of the differences amongst group means were 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 nonparametric
Kolmogorov-Smirnov test if n was more than 5.
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 nonparametric
version of the Williams test. The criteria for statistical significance were p<0.05
and p<0.01.
The mean value, standard deviations and statistical analysis were calculated from actual
values in the computer without rounding off.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Terminal body weight was unaffected by treatment in both sexes. Changes noted in the absolute and relative organ weights were of low magnitude and no histological findings were observed at microscopic examination, therefore they were considered of no toxicological relevance.
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
No changes of toxicological significance were recorded.
No significant change was recorded in the coagulation test.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
The alteration (decrease in cholesterol level) noted in males receiving 500 mg/kg/day could not be conclusively attributed to treatment since it was observed in only one sex.
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
no effects observed
Details on maternal toxic effects:
Maternal toxic effects:no effects
Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (nominal)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
no effects observed
Skeletal malformations:
not examined
Visceral malformations:
not examined
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects
Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Conclusions:
Benzyl methacrylate did not show developmental toxicity via oral gavage to rats at doses as high as 500 mg/kg bw/day in the decribed reproductive/developmental toxicity screening study according to OECD 422.
Executive summary:

Benzyl methacrylate did not show developmental toxicity via oral gavage to rats at doses as high as 500 mg/kg bw/day in the decribed reproductive/developmental toxicity screening study according to OECD 422 (for more details please also see IUCLID chapter 7.5.1 and 7.8.1). There were no signs of neonatal toxicity or external malformations at doses of 500 mg/kg bw/day.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the methacrylic metabolite:

According to RAAF 2019, the analogue read-across approach, Scenario 1: “read-across based on (bio)transformation to common compound(s)” has been identified as appropriate in order to fulfill the data gap.

Thus, the information requirement (according to REACH Regulation Annex IX) is covered with read across to the methacrylic metabolites.

For detailed information and justification please refer to the attached Read across Justification Document: “Read across assessment according to ECHA’s Read Across Assessment Framework (RAAF)”.
Reason / purpose for cross-reference:
read-across source
Dose / conc.:
50 mg/kg bw/day (nominal)
Remarks:
actual dose: 41 mg/kg bw/d
Dose / conc.:
150 mg/kg bw/day (nominal)
Remarks:
actual dose: 132 mg/kg bw/d
Dose / conc.:
450 mg/kg bw/day (nominal)
Remarks:
actual dose: 406 mg/kg bw/d
Clinical signs:
no effects observed
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
There were no test substance-related or spontaneous mortalities in any group.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
The mean body weights of the low-, mid- and high-dose rabbits (50; 150 and 450 mg/kg bw/d) were not significantly different from the concurrent control throughout the course of the study. The average body weight gain of the mid- and high-dose rabbits was statistically significantly reduced by about 27% and 31% during the treatment period. A significant reduction of mean body weight gain was also noted for the the high-dose rabbits on GD 19-21.

Corrected (net) body weight gain: Mean carcass weights and the corrected body weight gain (terminal body weight on GD 29 minus weight of the unopened uterus minus body weight on GD 6) were comparable among all groups.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
The food consumption in the high-dose females (450 mg/kg bw/d) was distinctly and statistically significantly reduced during a significant part of the treatment period (GD 15-23). During the entire treatment period (GD 6-28) the total average food consumption of the high dose rabbits was about 18% below controls. The food consumption of the mid dose females (150 mg/kg bw/d) was similarly affected in terms of magnitude and course of reduction, however the reduction of food consumption reached statistical significance only on GD 22-24. During the treatment period (GD 6-28) the total average food consumption of the mid-dose rabbits was about 13% below controls. Overall, the food consumption of the low-dose does (50 mg/kg bw/d) did not show test substance-related impairments. The reduced food consumption at the 150 and 450 mg/kg bw/d levels is considered to be related to the treatment.
Food efficiency:
not specified
Ophthalmological findings:
not specified
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not specified
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
The mean gravid uterus weights of all test groups did not show staatistically significant differences in comparison to control.
Gross pathological findings:
no effects observed
Description (incidence and severity):
At necropsy, only spontaneous findings were seen in single females of every test group. No test substance-related findings were observed in the doses.
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
The conception rate reached 96% in test groups 1 and 3 (50 and 450 mg/kg bw/d) and 100% in test groups 0 and 2 (0 and 150 mg/kg bw/d). Importantly, a sufficient number of pregnant females was available for the purpose of the study, as 24-25 pregnant rabbits per group had implantation sites in the uterus, at terminal sacrifice. There were no test substance-related and/or biologically relevant differences between the control and all dosed groups in conception rate, in the mean number of corpora lutea and implantation sites or in the values calculated for the pre- and the postimplantation losses, the number of resorptions and viable fetuses. Gestational parameters were within the normal range for animals of this strain and age.
Dose descriptor:
NOAEL
Effect level:
450 mg/kg bw/day (nominal)
Based on:
test mat.
Remarks on result:
other: Read-across to Methyl methacrylate CAS: 80-62-6. no adverse effects observed; actual dose 406 mg/kg/d
Dose descriptor:
NOEL
Remarks:
food consumption
Effect level:
50 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
food consumption and compound intake
Remarks on result:
other: Read-across to Methyl methacrylate CAS: 80-62-6. actual dose: 41 mg/kg/d
Abnormalities:
effects observed, treatment-related
Fetal body weight changes:
no effects observed
Description (incidence and severity):
The mean fetal weights of all treated groups were not influenced by the test substance. Neither female nor male weights showed statistically significant or biologically relevant differences between the test substance-treated groups and the controls.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
The sex distribution of the fetuses in test groups 1-3 (50; 150 and 450 mg/kg bw/d) was comparable to the control fetuses. Observable differences were without biological relevance.
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
One sole external malformation (unilateral microphthalmia) was recorded for two fetuses from 2 litters in the high-dose group (450 mg/kg bw/d). This malformation is present in the historical control data. Thus an association of these individual findings to the treatment is not assumed. The total incidences of external malformations were comparable to the historical control data.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Malformations of the fetal skeletons were noted in fetuses of test groups 0, 2 and 3 (0; 150 and 450 mg/kg bw/d). Neither statistically significant differences between treated groups and the control were calculated nor a dose-response relationship was observed. All individual skeletal malformations were present in the historical control data at a comparable frequency.
- Fetal skeletal variations: For all test groups, variations in different skeletal structures were detected with or without effects on the corresponding cartilages. The observed skeletal variations were related to various parts of the fetal skeletons and were statistically significant higher in the low- and the high-dose groups on a fetus per litter basis. Several specific skeletal variations were statistically significant higher than the concurrent control in the dosed groups (on a fetus per litter basis). These findings are delays or minor disturbances of ossification which are reversible or do not considerably affect the integrity of the underlying structures. Such slight changes of the ossification process occur very frequently in gestation day 29 rabbit fetuses of this strain and all observed incidences were within the historical control data. Thus an association of these findings to the treatment is not assumed.
- Fetal skeletal unclassified cartilage observations: Additionally, isolated cartilage findings without impact on the respective bony structures, which were designated as unclassified cartilage observations, occurred in all groups including the control. The observed unclassified cartilage findings did not show a relation to dosing and were comparable to historical control data and, therefore, regarded to be spontaneous in nature.
Visceral malformations:
no effects observed
Description (incidence and severity):
- Fetal soft tissue malformations: The examination of the soft tissues revealed a variety of malformations in fetuses of all test groups including the controls (0; 50; 150 and 450 mg/kg bw/d). With the exception of a lateral pouch in the tongue of 2 fetuses all individual soft tissue malformations were present in the historical control data at comparable frequencies. No statistically significant differences between the test groups and the control were observed. The total incidences of external malformations were comparable to the historical control data. No malformation pattern was evident. Thus an association of these findings to the treatment is not assumed.
- Fetal soft tissue variations: A number of soft tissue variations, such as absent lung lobe (lobus inferior medialis) and malpositioned carotid branch, was detected in each test group including the controls. Incidences were without a relation to dosing. Neither statistically significant differences between the test groups nor differences to the historical control data were noted.
- Fetal soft tissue unclassified observations: Unclassified soft tissue observations, such as infarct of liver, hemorrhagic thymus or ovary and blood coagulum around urinary bladder, were recorded for some fetuses of test groups 0, 1, 2 and 3 (0; 50; 150 and 450 mg/kg bw/d). A relation to dosing is not present if normal biological variation is taken into account. Therefore, a test substance induced effect is not assumed.
Details on embryotoxic / teratogenic effects:
- Abstract of all classified fetal external, soft tissue and skeletal observations: Various external, soft tissue and skeletal malformations occurred throughout all test groups including the control. All individual malformations are present in the historical control data, with the exception of lateral pouches in the tongue of 2 fetuses. They did neither show a consistent pattern since a number of morphological structures of different ontogenic origin were affected nor a clear dose-response relationship. The overall incidence of malformations was comparable to the historical control data. One external (paw hyperflexion), two soft tissue (absent lobus inferior medialis and malpositioned carotid branch) and a broad range of skeletal variations occurred in all test groups including the controls. All fetal and litter incidences for these variations and the corresponding mean percentages of affected fetuses/litter were not significantly different from the concurrent control and their frequency is comparable to the historical control data. Therefore, they were not considered to be related to the treatment. A spontaneous origin is also assumed for external, soft tissue and unclassified skeletal cartilage observations which were observed in several fetuses of all test groups including controls (0, 50; 150 and 450 mg/kg bw/d). Distribution and type of these findings do not suggest relation to treatment.
Key result
Dose descriptor:
NOAEL
Remarks:
prenatal developmental toxicity
Effect level:
450 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: Read-across to Methyl methacrylate CAS: 80-62-6. no adverse effects observed
Abnormalities:
no effects observed
Developmental effects observed:
no

Table: Occurence of statistically significantly increased fetal skeletal variation (expressed as mean percentage of affected fetuses/litter)

Finding

Group 0

0 mg/kg/d

Group 1

50 mg/kg/d

Group 2

150 mg/kg/d

Group 3

450 mg/kg/d

HCD

(range)

Incomplete ossification of parietal; unchanged cartilage

0.0

0.0

1.9*

2.1*

0.4

(0.0 – 2.6)

Incomplete ossification of hyoid; cartilage present

11.2

11.4

19.1

20.4*

9.8

(0.0 – 21.6)

Splitting of skull bone

0.4

3.3*

3.3

2.3

2.9

(0.0 – 7.7)

Incomplete ossification of cervical centrum; unchanged cartilage

2.5

2.2

3.6

7.3*

2.5

(0.0 – 9.3)

Supemumerary 13th rib; cartilage not present

2.5

9.8

6.1

9.9*

6.6

(0.0 – 17.5)

Total fetal skeletal variations

46.3

63.7*

59.3

71.6**

63.5

(46.3 – 81.9)

HCD = Historical control data; * = p ≤ 0.05, ** = p ≤ 0.01 (Wilcoxon-Test [one-sided])

 

 

Table: Total fetal malformations

 

 

Group 0

0 mg/kg/d

Group 1

100 mg/kg/d

Group 2

300 mg/kg/d

Group 3

1000 mg/kg/d

Litter

Fetuses

N

N

25

171

24

154

25

157

24

158

Fetal incidence

N (%)

4 (2.3%)

2 (1.3%)

6 (3.8%)

9 (5.7%)

Litter incidence

N (%)

4 (16%)

1 (4.2%)

4 (16%)

7 (29%)

Affected fetuses/litter

Mean%

2.3

1.2

3.6

6.2

 

 

Table: Total fetal variations

 

 

Group 0

0 mg/kg/d

Group 1

100 mg/kg/d

Group 2

300 mg/kg/d

Group 3

1000 mg/kg/d

Litter

Fetuses

N

N

25

171

24

154

25

157

24

158

Fetal incidence

N (%)

106 (62%)

106 (69%)

106 (68%)

122 (77%)

Litter incidence

N (%)

21 (84%)

24 (100%)

24 (96%)

23 (96%)

Affected fetuses/litter

Mean%

59.9

69.8

64.3

74.2

 

Conclusions:
Read-across to Methyl methacrylate CAS: 80-62-6:
In conclusion, the no observed adverse effect level (NOAEL) for maternal toxicity is 450 mg/kg bw/d and the no observed effect level (NOEL) for maternal toxicity is 50 mg/kg bw/d based on effects on food consumption. The NOAEL for prenatal developmental toxicity is 450 mg/kg bw/d. No adverse fetal findings of toxicological relevance were evident at any dose.
Executive summary:

Read-across to Methyl methacrylate CAS: 80-62-6:

The study was performed according to OECD TG 414 in compliance with GLP.

Methyl Methacrylate was tested for its prenatal developmental toxicity in Himalayan rabbits. The test substance was administered as an aqueous preparation to 25 inseminated female Himalayan rabbits by stomach tube at doses of 50; 150 and 450 mg/kg body weight/day on gestation days (GD) 6 through GD 28. The control group, consisting of 25 females, was dosed with the vehicle (1% Carboxymethylcellulose CB 30.000 in drinking water and a few drops Cremophor EL and one drop hydrochloric acid [1% CMC]) in parallel. A standard dose volume of 10 mL/kg body weight was used for each test group. At terminal sacrifice on GD 29, 24-25 females per group had implantation sites.

The following test substance-related adverse effects/findings were noted:

Test group 3 (450 mg/kg body weight/day):

-        Reduced food consumption (-18%) and body weight gain (-31%)

-        No test substance-related adverse effects on gestational parameters or fetuses

 

Test group 2 (150 mg/kg body weight/day):

-        Reduced food consumption (-13%) and body weight gain (-27%)

-        No test substance-related adverse effects on gestational parameters or fetuses

 

Test group 1 (50 mg/kg body weight/day):

-        No test substance-related adverse effects on does, gestational parameters or fetuses

In conclusion, the no observed adverse effect level (NOAEL) for maternal toxicity is nominal 450 mg/kg bw/d (actual 406 mg/kg bw/d), the highest dose tested. The no observed effect level (NOEL) for maternal toxicity is nominal 50 mg/kg bw/d (effective 41 mg/kg bw/d) based on effects on food consumption being a consequence of reduced appetite observed at the LOEL (Lowest Observed Effect Level) of 150 mg/kg bw/d (actual 132 mg/kg bw/d).

The no observed adverse effect level (NOAEL) for prenatal developmental toxicity is nominal 450 mg/kg bw/d (actual 406 mg/kg bw/d). No adverse fetal findings of toxicological relevance were evident at any dose.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the alcohol metabolite

According to RAAF 2019, the analogue read-across approach, scenario 1:"read-across based on (bio) transformation to common compound(s)" has been identified as appropriate in order to fulfill the data gap.
Thus, the information requirement (according Regulation 1907/2008, Annex IX) is covered with read-across to the alcohol metabolite.

For detailed information and justification, please refer to the attached read-across justification document: Read across assessement according to ECHA's Read Across Assessment Framework.
Reason / purpose for cross-reference:
read-across source
Dose / conc.:
750 mg/kg bw/day (actual dose received)
Remarks:
Dose was determined in a preliminary dose range finder study
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Cited from the report:
Animals which showed cyanosis, dyspnoea, hypothermia, hypoactivity, appeared thin or had red staining at the perineum all died during the test.
Other signs recorded included body tremors, hunching, subdued behaviour, prostration, ataxia, swelling and/or cyanosis of the abdomen and piloerection.
These signs appeared both in animals which died, and in those which went on to produce litters.
Mortality:
mortality observed, treatment-related
Description (incidence):
18/50 females died during the treatment period and one females died on Gestation Day 15.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
cited from the report: A significant reduction in the mean Day 18 body weight was noted, both in the unadjusted data (P<0.05)
and in the data adjusted for litter size on Day 1 post partum (P<0.001). There was also a marginal reduction in the maternal weight on Day 3 post
partum (P<0.05) in the treated group when compared with control data. As expected, the body weight gain over Days 7-18 of gestation was reduced for
both unadjusted (P<0.05) and adjusted (P<0.001) data.
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
no effects observed
Key result
Dose descriptor:
LOAEL
Effect level:
750 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
clinical signs
mortality
Remarks on result:
other: Read-across to the alcohol metabolite Benzyl alcohol CAS: 100-51-6.
Fetal body weight changes:
effects observed, treatment-related
Description (incidence and severity):
cited from the report:
The following differences from control data were noted: a decreased mean litter mean pup weight on Day 1 post partum (P<0.01); a decreased mean litter mean pup weight on Day 3 post partum (P<0.001); a decreased mean litter weight
change Days 1-3 post partum (P<0.05); a decreased mean litter mean pup weight change Days 1-3 post partum (P<0.001). Percent weight changes were
consequently affected.
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
effects observed, treatment-related
Description (incidence and severity):
cited from the report:
The following differences from control data were noted: a decreased mean litter mean pup
weight on Day 1 post partum (P<0.01); a decreased mean litter mean pup weight on Day 3 post partum (P<0.001); a decreased mean litter weight
change Days 1-3 post partum (P<0.05); a decreased mean litter mean pup weight change Days 1-3 post partum (P<0.001). Percent weight changes were
consequently affected.

Litter size is unaffected by treatment.
Changes in postnatal survival:
no effects observed
External malformations:
not examined
Skeletal malformations:
not examined
Visceral malformations:
not examined
Key result
Dose descriptor:
LOAEL
Effect level:
750 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes
Remarks on result:
other: Read-across to the alcohol metabolite Benzyl alcohol CAS: 100-51-6.
Key result
Abnormalities:
not examined
Key result
Developmental effects observed:
yes
Lowest effective dose / conc.:
750 mg/kg bw/day (actual dose received)
Treatment related:
yes
Relation to maternal toxicity:
developmental effects as a secondary non-specific consequence of maternal toxicity effects
Dose response relationship:
no
Relevant for humans:
not specified
Executive summary:

Read-across to the alcohol metabolite Benzyl alcohol CAS: 100-51-6:

The Reproduction Tests as carried out in this programme indicated the following:

Ethylene diamine, Benzyl alcohol, Nitrofurazone, Diethylene glycol, and Diethylene glycol (dibutyl ether) would be suspected as being potential reproductive hazards. The latter 2 substances showed effects which were less in degree than those showed by the former 3, but a "positive" response was concluded for them. In all these cases, further inyestigation of the effects demonstrated would be indicated.

However, there was some tendency for severity of effects on the pups to correlate with severity of toxicity to the dams. Such a correlation,

if confirmed, would complicate interpretation of results for test substances showing either very high or very low maternal toxicity in the test. An examination of results obtained by other investigators in this programme may show whether this is likely to be a significant complication.

Today, it is well known that the reduced litter and/ or mean pup weight are secondary effects due to the

reduced body weight of the female rats, because the reduced body weight leads to the interruption of the Leptin-Ghrelin-

Kisspeptin-GnRH Pathway (Yuan, 2000)1.

1 Yuan, C., Attele, A., Zhang, L. et al. (2000): Leptin Reduces Body Weight Gain in Neonatal Rats. Pediatr Res. 2000.48: 380–383.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the alcohol metabolite

According to RAAF 2019, the analogue read-across approach, scenario 1:"read-across based on (bio) transformation to common compound(s)" has been identified as appropriate in order to fulfill the data gap.
Thus, the information requirement (according Regulation 1907/2008, Annex IX) is covered with read-across to the alcohol metabolite.

For detailed information and justification, please refer to the attached read-across justification document: Read across assessement according to ECHA's Read Across Assessment Framework.
Reason / purpose for cross-reference:
read-across source
Dose / conc.:
550 mg/kg bw/day (actual dose received)
Ovaries and uterine content:
Any female showing no signs of delivering a litter by Gestation Day 22 was sacrificed and the uterus examined ofr evidence of pregnancy.
Clinical signs:
no effects observed
Description (incidence and severity):
One animal in the treated group died from toxicity. Clincial signs noted before death were languid behaviour, labored respiration, rough haircoat. This animal died on day 5 of treatment.
Mortality:
no mortality observed
Description (incidence):
The mortality rate for benzyl alcohol did not significantly differ from the vehicle control group. 1/50 animal in the treated group died from toxicity. Clincial signs noted before death were languid behaviour, labored respiration, rough haircoat. This animal died on day 5 of treatment.
Body weight and weight changes:
no effects observed
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
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
550 mg/kg bw/day (nominal)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
not examined
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
not examined
Skeletal malformations:
not examined
Visceral malformations:
not examined
Key result
Dose descriptor:
NOAEL
Effect level:
550 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the alcohol metabolite Benzyl alcohol CAS: 100-51-6.
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Executive summary:

Read-across to the alcohol metabolite Benzyl alcohol CAS: 100-51-6:

Cited from the report:

The test compound Benzyl alcohol, administered at the predicted LD10, did not produce evidence in this test system of developmental toxicity.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the methacrylic metabolite:

According to RAAF 2019, the analogue read-across approach, Scenario 1: “read-across based on (bio)transformation to common compound(s)” has been identified as appropriate in order to fulfill the data gap.

Thus, the information requirement (according to REACH Regulation Annex IX) is covered with read across to the methacrylic metabolites.

For detailed information and justification please refer to the attached Read across Justification Document: “Read across assessment according to ECHA’s Read Across Assessment Framework (RAAF)”.
Reason / purpose for cross-reference:
read-across source
Dose / conc.:
99 ppm
Remarks:
corresponding to 412 mg/m3 or 0.41 mg/L
Dose / conc.:
304 ppm
Remarks:
corresponding to1285 mg/m3 or 1.29 mg/L
Dose / conc.:
1 178 ppm
Remarks:
corresponding to 4900 mg/m3 or 4.9 mg/L
Dose / conc.:
2 028 ppm
Remarks:
corresponding to 8436 mg/m3 or 8.44 mg/L
Clinical signs:
no effects observed
Description (incidence and severity):
The only clinical sign noted was a minimal increase in the incidence of scant feces at 2028 ppm.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Treatment-elated decreases on maternal body weight and feed consumption were noted at all exposure levels. The decreases in maternal body weight at 99 and 304 ppm were minimal and transient since they occurred only during the first 2 days of exposure and returned to control values by the next weighing period. The body weight and feed consumption values returned to control values for all groups during the post exposure period (GD 16-20). At 1178 and 2028 ppm, treatment-related effects included losses in maternal body and/or decreased body weight gain throughout the exposure period (GD 6 - 16) and decreased corrected maternal body weight gain.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Treatment related decrease waere noted on feed consumption between days 6-10 G at all exposure levels teasted. Decteases in feed consumption continued between days 6-10 G at 304, 1178, and 2028 ppm. Feed consumption of all treated groups returend to control value during post-treatment period (days 16-20 G)
Gross pathological findings:
no effects observed
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Details on maternal toxic effects:
Details on maternal toxic effects:
No animals died and no treatment-related clinical signs were noted for the dams in the 99, 304 or 1178 ppm groups. Scant feces was noted in the 2028 ppm group throughout the exposure period (GD 6-15). Treatment-related decreases on maternal body weight and feed consumption were noted at all exposure levels. The decreases in maternal body weight at 99 and 304 ppm were minimal and transient since they occurred only during the first 2 days of exposure and returned to control values by the  next weighing period. The body weight and feed consumption values returned to control values for all groups during the post exposure period  (GD 16-20). At 1178 and 2028 ppm, treatment-related effects included losses in maternal body and/or decreased body weight gain throughout the exposure period (GD 6 - 16) and decreased corrected maternal body weight gain. The gross necropsy evaluations did not indicate any treatment-related effects and there were no treatment-related differences between the control and treated groups in any reproductive parameter.  
Dose descriptor:
LOEC
Remarks:
maternal toxicity
Effect level:
ca. 0.41 mg/L air (analytical)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEC
Remarks:
maternal toxicity
Effect level:
8.44 mg/L air (analytical)
Based on:
test mat.
Remarks on result:
other: no adverse effects observed
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment-related increase were detected in the type of external, visceral, or skeletal malformations.
Spontaneous malformations detected in the study included duplicated hypothalamus in a control fetus, hydrocephaly in a fetus at 304 ppm and an omphalocele in one fetus and enlarged adrenal glands in two fetuses at 1178 ppm was not considered to be treatment related as at 2028 ppm no increase was observed.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The significant increase in skeletal developmental variations at 1178 ppm was not detected at 2028 ppm, the highest doese tested. The slightly, not statistically significant number of fetuses with partial or unossified sternebra at 304 ppm was not considered a result of treatment since no dose response was demonstrated.
Visceral malformations:
no effects observed
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
Details on embryotoxic / teratogenic effects:
 Fetal body weight was not affected by exposure to MMA vapors. The fetal external, visceral and skeletal examinations did not show any treatment related effects.
Dose descriptor:
NOAEC
Remarks:
fetotoxicity
Effect level:
8.44 mg/L air (analytical)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: Read-across to Methyl methacrylate CAS: 80-62-6. corresponding to 2028 ppm; no substance related effects observed
Key result
Dose descriptor:
NOAEC
Remarks:
teratogenicity
Effect level:
8.44 mg/L air (analytical)
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: Read-across to Methyl methacrylate CAS: 80-62-6. corresponding to 2028 ppm; no substance related effects observed
Abnormalities:
no effects observed
Developmental effects observed:
no

Mean measured concentrations (± SD) within the chambers for the 0, 100, 300, 1200 and 2000 ppm groups were 98.8 (±3.4), 304.4 (±9.1), 1178.1  (±69.1) and 2028.2 (±107.3) ppm, respectively.

Conclusions:
Read-across to Methyl methacrylate CAS: 80-62-6:
Exposure by inhalation to methyl methacrylate concentrations up to 8.44 mg/L (2028 ppm) resulted in no embryo or fetal toxicity or malformations even at exposure levels that resulted in maternal toxicity.
Executive summary:

Read-across to Methyl methacrylate CAS: 80-62-6:

In a developmental toxicity study (1991) acc. OECD 414 by inhalation on CRl: CD Br rats pregnant rats were exposed to methyl methacrylate at concentrations of 0 (control), 99, 304, 1178 and 2028 ppm on days 6-15 of gestation. A maternal no observed level was not demonstrated since losses in maternal body weight or decreases in maternal body weight gain and decreases in maternal feed consumption were noted at all exposure levels tested. No embryo or fetal toxicity was evodent and no increase in the incidence of malformations or variations was noted at exposure levels up to and including 2028 ppm. Therefore toxicity to the conceptus was not evident even at expsoure levels that resulted in overt maternal toxicity.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the alcohol metabolite

According to RAAF 2019, the analogue read-across approach, scenario 1:"read-across based on (bio) transformation to common compound(s)" has been identified as appropriate in order to fulfill the data gap.
Thus, the information requirement (according Regulation 1907/2008, Annex IX) is covered with read-across to the alcohol metabolite.

For detailed information and justification, please refer to the attached read-across justification document: Read across assessement according to ECHA's Read Across Assessment Framework.
Reason / purpose for cross-reference:
read-across: supporting information
Analytical verification of doses or concentrations:
not specified
Dose / conc.:
1.75 mg/kg bw/day (actual dose received)
Dose / conc.:
8 mg/kg bw/day (actual dose received)
Dose / conc.:
38 mg/kg bw/day (actual dose received)
Dose / conc.:
175 mg/kg bw/day (actual dose received)
Clinical signs:
not specified
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
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
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
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:
not examined
Changes in number of pregnant:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Executive summary:

Read-across to the metabolite Sodium benzoate CAS: 532-32-1:

Cited from the report:

The administration of up to 175 mg/kg (body weight) of the test material to pregnant rats for 10 consecutive days had no clearly discernible effect on nidation or on maternal or fetal survival. The number of abnormalities seen in either soft or skeletal tissues of the test groups did not differ from the number occuring spontaneously in the sham-treated controls.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the alcohol metabolite Sodium benzoate

According to RAAF 2019, the analogue read-across approach, scenario 1:"read-across based on (bio) transformation to common compound(s)" has been identified as appropriate in order to fulfill the data gap.
Thus, the information requirement (according Regulation 1907/2008, Annex IX) is covered with read-across to the alcohol metabolite.

For detailed information and justification, please refer to the attached read-across justification document: Read across assessement according to ECHA's Read Across Assessment Framework.
Reason / purpose for cross-reference:
read-across source
Dose / conc.:
1.75 mg/kg bw/day (actual dose received)
Dose / conc.:
8 mg/kg bw/day (actual dose received)
Dose / conc.:
38 mg/kg bw/day (actual dose received)
Dose / conc.:
175 mg/kg bw/day (actual dose received)
Clinical signs:
not specified
Mortality:
mortality observed, non-treatment-related
Body weight and weight changes:
no effects observed
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
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
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:
not examined
Changes in number of pregnant:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 175 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Developmental effects observed:
no
Executive summary:

Read-across to the metabolite Sodium benzoate CAS: 532-32-1:

cited from report:

The administration of up to 175 mg/kg (body weight) of the test material to pregant mice for 10 consecutive days had no clearly discernible effect on nidation or on maternal or fetal survival. The number of abnormalities seen in either soft or skeletal tissues of the test groups did not differ grom the number occuring spontaneously in the sham-treated contols.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the alcohol metabolite

According to RAAF 2019, the analogue read-across approach, scenario 1:"read-across based on (bio) transformation to common compound(s)" has been identified as appropriate in order to fulfill the data gap.
Thus, the information requirement (according Regulation 1907/2008, Annex IX) is covered with read-across to the alcohol metabolite.

For detailed information and justification, please refer to the attached read-across justification document: Read across assessement according to ECHA's Read Across Assessment Framework.
Reason / purpose for cross-reference:
read-across: supporting information
Dose / conc.:
2.5 mg/kg bw/day (actual dose received)
Dose / conc.:
12 mg/kg bw/day (actual dose received)
Dose / conc.:
54 mg/kg bw/day (actual dose received)
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Clinical signs:
not specified
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
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
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
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 number of pregnant:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Executive summary:

Read-across to the metabolite Sodium benzoate CAS: 532-32-1:

Cited from the report:

The administration of up to 250 mg/kg (body weight) of the test material to pregnant rabbits for 13 consecutive days had no clearly discernible effect on nidation or on maternal or fetal survival. The number of abnormalities seen in either soft or skeletal tissues of test groups did not differ from the number occuring spontaneously in the sham-treated controls.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
Read across to the alcohol metabolite

According to RAAF 2019, the analogue read-across approach, scenario 1:"read-across based on (bio) transformation to common compound(s)" has been identified as appropriate in order to fulfill the data gap.
Thus, the information requirement (according Regulation 1907/2008, Annex IX) is covered with read-across to the alcohol metabolite.

For detailed information and justification, please refer to the attached read-across justification document: Read across assessement according to ECHA's Read Across Assessment Framework.
Reason / purpose for cross-reference:
read-across: supporting information
Dose / conc.:
3 mg/kg bw/day (actual dose received)
Dose / conc.:
14 mg/kg bw/day (actual dose received)
Dose / conc.:
65 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Clinical signs:
not specified
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
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
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
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:
not examined
Changes in number of pregnant:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 300 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
> 300 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
Read-across to the metabolite Sodium benzoate CAS: 532-32-1.
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Executive summary:

Read-across to the metabolite Sodium benzoate CAS: 532-32-1:

Cited from the report:

The administration of up to 300 g/kg (body weight) of the test material to pregnant hamsters for 5 consecutive days had no clearly discernible effect on nidation or on maternal or fetal survival. The number of abnormalities seen in either soft or skeletal tissues of the test groups did not differ from the number occuring spontaneously in the sham-treated controls.

For read across purposes according to ECHA's RAAF (2019), this study is applicable with a high level of confidence.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
500 mg/kg bw/day
Study duration:
subacute
Species:
rat
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

BENZYL METHACRYLATE has been analysed for reproductive and developmental toxicity in a GLP-study according to OECD guideline 422. In a Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test according to OECD Guideline 422 (22 March 1996). BENZYL METHACRYLATE was administered to 10 Sprague Dawley [Crl:CD(SD)BR] rats/sex/dose orally by gavage at dose levels of 0 (control), 50, 175 and 500 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. No clinical signs of toxicological significance were reported. No relevant differences in body weights and food consumption were recorded in animals of both sexes compared to the control group, throughout the study.

No relevant differences were noted in motor activity and sensory reaction to stimuli between control and treated groups. Haematology and urinalysis revealed no changes of toxicological significance.

A decrease in cholesterol levels noted in males receiving 500 mg/kg bw/day could not be conclusively attributed to treatment since it was observed in only one sex.

Measurements of oestrus cycle, pre-coital intervals and copulatory index did not show differences between treated and control groups. 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. All dams gave birth between Days 21 and 23 post coitum.

Litter data parameters and sex ratios at birth and on Day 4 post-partum were similar between groups. Pre-weaning clinical signs were comparable between treated and control groups. Necropsy findings in decedent pups and in pups sacrificed on Day 4 post-partum did not reveal any treatment-related effect.

At necropsy, changes noted in the absolute and relative organ weights were of low magnitude and no histological findings were observed at microscopic examination, therefore they were considered to be of no toxicological relevance.

No treatment-related macroscopic or microscopic changes were noted. In addition, seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and no alterations were noted.

No relevant toxic effects were seen in parental animals up to the highest dose group of 500 mg/kg bw/d. Based on the results obtained in the study, the NOAEL for both, general toxicity and reproduction/developmental toxicity was 500 mg/kg bw/d (males/females). 

This is supported by the fact that BENZYL METHACRYLATE is rapidly metabolised in vivo and the primary metabolites, methacrylic acid as well as benzyl alcohol (and its metabolites Benzoic acid and Sodium benzoate), demonstrate an absence of concern for reproductive and developmental toxicity.

MMA as metabolite donor substance for MAA has been tested in an oral OECD 414 study in rabbits at 0, 50, 150 and 405 mg/kg bw/d. The NOAEL for developmental toxicity was reported as 450 mg/kg bw/d. There were no adverse effects observed in the pups, even in the presence of maternal toxicity manifested in a reduced body weight due to reduced food consumption. The derived maternal NOEL was 50 mg/kg bw/d.

In a supportive inhalation study according to OECD 414, MMA was administered by inhalation exposure to 99, 304, 1178 and 2028 ppm (corresponding 412, 1285, 4900 and 8436 mg/m³). No relevant maternal treatment-related effects were noted at any concentrations tested. Furthermore, no embryo or foetal toxicity was observed. The incidence of malformations or variations was not increased by MMA. The NOAEC for developmental toxicity was 2028 ppm (8436 mg/m³).

Based on the available results on MMA, it was concluded in the OECD SIDS of MAA that ”Data on reproductive toxicity of MAA in animals or humans does not exist. From studies with MMA no concern in relation to reproductive toxicity of MAA has to be assumed”(OECD SIDS, 2002). This statement is supported by the results of an oral OECD 414 in rabbits showing no effects on the pups, even in the presence of maternal toxicity.

In the study performed by NIOSH in mice in 1983, BA was orally administered at a concentration of 750 mg/kg bw/d. The dose of 750 mg/kg bw/d correspond to the calculated maximum tolerated dose (MTD) that lay between 645 and 1300 mg/kg bw/d based on the results of a preliminary dose range finder study. The control group received the vehicle, distilled water, alone.The animals were allowed to deliver their litters and nurse their pups until Day 3 post-partum, at which time necropsies were performed. Maternal body-weight gain and mortality, mating, gestation, numbers of live and dead pups per litter, total litter weight on days 1 and 2 post-partum, litter weight change between days 1 and 3 post-partum, and pup survival on days 1 and 3 post-partum were recorded. During the treatment period, 18 of 50 females died. All deaths were treatment-related. One further female died on Gestation Day 15, one day after the final dosing. Most of the female mice showed severe clinical signs during the treatment period, e.g. hunched posture, tremors, inactivity, prostration, hypothermia, ataxia, dyspnea, swollen or cyanotic abdomen, and piloerection. The maternal body weight from Gestation Day 4 and 7, was comparable to the control, but a statistically significant reduction was reported on Gestation Day 18 and on Day 3 post-partum. Maternal body weight gain from Gestation Day 7 to 18 was significantly lower than the control value. Furthermore, the mating and gestation indices, the gestation length and the number of resorptions is unaffected by the treatment. Taken together, the female mice showed severe signs of toxicity and the LOAEL was 750 mg/kg bw/.

The maternal toxicity negatively influenced the pups body weight, the mean pup weight per litter on Day 1 and 3 post-partum, the mean pup weight change between Day 1 and 3 post-partum and the mean litter weight change from Day 1 and 3 post-partum. The survival of the pups and the number of live pups is unaffected by BA administration to the dams. Taken together, the LOAEL for developmental toxicity was 750 mg/kg bw/d.

In the study performed in 1986, 50 mice were treated with 550 mg/kg bw/d BA from Gestation Day 6-15. The dose of 550 mg/kg bw/d correspond to the LD10 value determined in two preliminary dose range finder studies. The control group received the vehicle, corn oil, alone. The dams were allowed to deliver naturally and rear the pups until Day 3 post-partum. Maternal body-weight gain and mortality, mating, gestation, numbers of live and dead pups per litter, total litter weight on days 1 and 2 post-partum, litter weight change between days 1 and 3 post-partum, and pup survival on days 1 and 3 post-partum were recorded. Mortality was not significantly increased in the dosed animals. One female showing languid behaviour, laboured breathing, and a rough coat died. No further clinical signs were reported in the treated animals. Body weight and body weight until Day 3 post-partum is unaffected by the administration of BA. All other parameters examined for the dams and the pups were comparable to the control group. The NOAEL was 550 mg/kg bw/d for maternal and developmental toxicity.

 

Sodium Benzoate

Additionally, there are studies on developmental toxicity available with sodium benzoate on rat, mouse, hamster and on the non-rodent species rabbit. The mentioned studies belong to one FDA study which consists of 4 parts for each species.

Rats:

A study using pregnant Wistar rats, orally dosed with 1.75, 8, 38 or 175 mg/kg sodium benzoate from Gestation Days 6-15 of gestation showed no effect on nidation, maternal or foetal survival. The number of abnormalities of soft and skeletal tissues did not differ from controls. The NOAEL for maternal and developmental toxicity was 175 mg/kg bw.

 

Mice:

In a gavage-study pregnant mouse were dosed with 1.75, 8, 38 or 175 mg/kg sodium benzoate from Gestation Days 6-15. The mice showed no effect on nidation, maternal or foetal survival. The number of malformations or variations was comparable to the control group. Therefore, the NOAEL for maternal and developmental toxicity was 175 mg/kg bw/d.

 

Rabbits:

In an oral study, 2.5, 12, 54 or 250 mg/kg sodium benzoate were administered to pregnant rabbits from Gestation Day 6-18. The animals showed no effect on nidation, maternal or foetal survival. The substance did not induce any soft tissue or skeletal malformation or variation. Consequently, the NOAEL for maternal and developmental toxicity was 250 mg/kg bw/d.

 

Hamster:

In a gavage-study pregnant hamster were dosed with 3, 14, 65 and 300 mg/kg sodium benzoate from Gestation Days 6-10. The animals showed no effect on nidation, maternal or foetal survival. The number of malformations or variations was comparable to the control group. Therefore, the NOAEL for maternal and developmental toxicity was 300 mg/kg bw/d.

Additionally, Benzoic acid (BAc) was tested four-generation study in rats in feeding study. The animals received 0, 0,5 or 1% (corresponding to 375 or 750 mg/kg bw/d). The first and second generation were treated lifelong, the third until week 16 and the fourth until breeding. The pregnancy rate, the number of pups/ litter, number of reared pups/ litter. There were no effects on any of the mentioned parameters observed. Therefore, the NOAEL was 750 mg/kg bw/d.

Several committees evaluated the available data on BA, BAc and NaB to assess the risk for developmental toxicity, e.g. JECFA (Joint FAO/WHO Expert Committee on Food Additives) concludedthat “The Committee was satisfied that the data reviewed for compounds in this group were sufficient to demonstrate the lack of teratogenic, reproductive or carcinogenic potential. Consequently, the Committee concluded that further studies were not required, and the group ADI of 0-5 mg per kg of body weight as benzoic acid equivalents was maintained”. SCF (Scientific committee for food) concluded that “the metabolism of benzyl alcohol to benzoic acid by man is well established. In special studies, biochemical effects have been investigated. The toxicity has been studied extensively, including acute, short-term and long-term toxicity, carcinogenicity, genotoxicity and developmental toxicity.

Taken together, the available data on developmental toxicity/ teratogenicity in combination with the lack of treatment-related effects on reproductive organs in chronic/ subchronic studies demonstrate that BA does not induce teratogenic effects.

 

BNMA did not show any developmental effects in a reproductive/ developmental screening study (OECD 422). This result is supported by study results obtained in reliable studies with the metabolites MMA (as donor substance for MAA), BA, NaB and BAc (as donor substance for BA) that also showed no developmental toxic effects. Altogether, these studies support the results of the available screening study with BNMA that there is no concern for BNMA being a reproductive toxicant.

 

 

References:

JECFA (1996): Benzyl acetate, Benzyl alcohol, Benzaldehyde, and Benzoic acid and its salts.available online.http://www.inchem.org/documents/jecfa/jecmono/v37je05.htm 

OECD SIDS (2002): Methacrylic acid; available online: https://www.petrochemistry.eu/wp-content/uploads/2018/01/79414.pdf

SCF (Scientific Committee on Food) (2002): Opinion of the Scientific Committee on Food on Benzoic acid and its salts. SCF/CS/ADD/CONS/48 Final, 17 Sept 2002, available online: http://ec.europa.eu/food/fs/sc/scf/out137_en.pdf

Justification for classification or non-classification

Based on the available data, BNMA does not need to be classified for toxicity to reproduction, developmental toxicity and teratogenicityaccording to the criteria given in regulation (EC) 1272/2008. Thus, no labelling is required.

Additional information