Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Toxicological information

Endpoint summary

Currently viewing:

Administrative data

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Effects on fertility

Effect on fertility: via oral route
Endpoint conclusion:
no study available
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

The extended one-generation reproductive toxicity study does not need to be conducted because there are no results from available repeated dose toxicity studies that indicate adverse effects on reproductive organs or tissues, or reveal other concerns in relation with reproductive toxicity.  

Effects on developmental toxicity

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
September 15, 2022 - January 10, 2023
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
June 25, 2018
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Species:
rat
Strain:
Wistar
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Animal Breeding Facility, Jai Research Foundation
- Age at study initiation: approximately 13 weeks at start of acclimatisation
- Weight at study initiation: 219.3 ± 11.0 g (198.3 – 239.7 g) at start of acclimatisation
- Housing: individually except for the mating period
- Diet (e.g. ad libitum): standard rodent pelleted diet (Certified Teklad Global 14% Protein Rodent Diet, Batch No 2014SC-010922MA, procured from Envigo, Inc., USA), ad libitum
- Water (e.g. ad libitum): Reverse Osmosis (RO) water, ad libitum
- Acclimation period: 6 days before cohabitation

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-23
- Humidity (%): 65-67
- Air changes (per hr): 16
- Photoperiod (hrs dark / hrs light): 12/12 (156 - 167 lux)
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The required amount of the test item was received for each dose level separately from the Test Item Control Office (TICO). The test item was mixed with vehicle to prepare dose formulations of required concentrations. The prepared dose formulations were thoroughly mixed using a magnetic stirrer before dosing and with cannula intermittently during dosing. The dose formulations were prepared on each day of dosing.

VEHICLE
- Justification for use and choice of vehicle (if other than water): based on solubility
- dose volume: 4 mL/kg bw
- Lot/batch no.: MKCP9878
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The stability of test item in the vehicle was established in JRF Study N° 228-2-13-30087. The test item was found stable in the vehicle (corn oil) up to 8 days from the time of preparation. Active ingredient (a.i.) concentration and homogeneity of dose formulations were analysed by the Department of Chemistry, using a validated analytical method. The active ingredient (A.I.) concentration and homogeneity of the test item in dose formulations were analyzed once before initiation of treatment and once during the treatment period. Duplicate samples of control (one layer) and the dose formulations (upper, middle, and lower layers) of each treatment group were taken to determine the homogeneity and concentration of the test item. A set of samples were sent for analysis and a second set were stored at frozen condition (-20 ± 5°C).

The mean concentration was determined and compared to the nominal value, and the coefficient of variation was calculated. The acceptance criteria was ± 10% deviation from the nominal value and %CV < 10. Samples were analysed at JRF using a validated analytical method.
Samples were analyzed by the following instrumental parameters:
Instrument : GC-FID (Agilent Technologies, 8890 with OpenLab Software)
Detector : FID
Column : DB Wax (30 m X 250 µm x 0.25 µm) or equivalent
Carrier Gas : N2
Injection Volume (µL) : 2.0 µL
Injector Temperature : 240 °C
Detector Temperature : 240 °C
Flow Rate (mL/minute) : 1.0
Split Ratio : Splitless
H2 Flow (mL/minute) : 35.0
Air Flow (mL/minute) : 350
Oven Temperature : 60 °C (hold 2 min) to 180 °C @ 20 °C/min (hold 5 min) to 220 °C @ 10 °C/min (hold 5 min).
Retention Time : 7.2 (approximately)
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:1
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
Duration of treatment / exposure:
GD 3 through 19
Frequency of treatment:
daily for 17 days (GD 3-19)
Duration of test:
20 d
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
750 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose levels were selected based on the results of the dose range finding study “Prenatal Developmental Oral Toxicity Study of Methyl benzoate in Wistar Rats (Dose Range Finding)’’ (JRF Study N°: RES-1-04-30089). Treatment related morality was observed in the 1000 mg/kg b. wt./day dose group.
Maternal examinations:
MORTALITY AND MORBIDITY: Yes
- Time schedule: twice daily

CLINICAL OBSERVATIONS: Yes
- Time schedule: visible clinical signs, such as changes in the skin, fur, eye, mucous membranes, and the behavioral pattern of the mated female rats, were recorded twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: individually on days 0, 3, 5, 8, 11, 14, 17, and 20 of gestation
- body weight gain was determined for the following intervals: Days 0–3, 3–5, 5–8, 8–11, 11–14, 14–17, 17-20, and 5-20

FOOD CONSUMPTION: Yes
- determined by differentiating the weight of feed input and leftover on gestation day 0, 3, 5, 8, 11, 14, 17 and 20
- food consumption calculated for female rats for the following intervals: days 0 - 3, 3 - 5, 5 - 8, 8 - 11, 11 - 14, 14 - 17, 17 – 20, and 5-20

WATER CONSUMPTION: No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day #20
- Ovaries and gravid uteri, including the cervix
- thyroid glands (post-fixation) and brain were collected and weighed from all female rats; thyroid glands were preserved for histopathology
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: Number of live fetuses , Number of dead fetuses
Blood sampling:
- Plasma: No
- Serum: Yes
- Volume collected: approximately 1.0 mL
Fetal examinations:
- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: Yes:
- Anogenital distance of all live rodent pups: yes
Statistics:
Parametric:
- Body weight, body weight change/gain, food consumption, corrected body weight/carcass weight, absolute and relative gravid uterine weights, organ weight, organ weight ratio, male sex ratio/male percent, pre-and postnatal loss (%), live fetus (%), and resorptions (%), ano-genital distance (normalized), fetal observations (percent fetal)
- 5% and 1% levels between control and treated groups
Non-parametric:
- Reproductive performance/Pregnancy rate, fetal observations (percent litter), fetal count, number of corpora lutea, number of implants, number of live fetus, number of dead fetus, number of pre-and postnatal losses, number of resorptions
- 5% level between control and treated groups
Historical control data:
refer to attached material
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Clinical sign such as salivation was observed in pregnant rats of the groups treated with 500 and 750 mg/kg b. wt./day. Salivation was not observed during the morning clinical observation. It is well known that salivation is often observed in gavage studies and may be a reaction to the taste or irritation of the test article rather than an indication of toxicity (Wook-Joon Yu et al., 2011). Therefore, salivation observed in the study was considered a physiological response to the dose formulation and toxicologically non-adverse in nature.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
The mean food consumption of the pregnant female rats was comparable between the control and the test item treated group up to the dose level of 500 mg/kg bw/day.
Statistically, a significant increase in mean food consumption during gestation days 11-14, 14-17, and 5-20 was observed in pregnant female rats in the group treated with 750 mg/kg bw/day when compared with that of the control group. However, this increase in food consumption did not cause an increase in the body weight and body weight change/gain. These opposite effects in food consumption and body weight were due to decreased feed efficiency and considered as an adverse effect of the test item.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Endocrine findings:
no effects observed
Description (incidence and severity):
The serum level of thyroid hormones (T3, T4, and TSH) were comparable between control and the test item treated group up to the dose level of 750 mg/kg bw/day .
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Description (incidence and severity):
The mean absolute and relative weights of brain and thyroid gland of pregnant rats were comparable between the control and treatment groups up to the dose level of 750 mg/kg b. wt./day.
Gross pathological findings:
no effects observed
Description (incidence and severity):
External and internal (gross) examination of terminally sacrificed female rats did not reveal any macroscopic lesion up to the dose level of 750 mg/kg bw/day.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
Congenital lesions, namely ectopic thymus tissue (Rat N° 99) was observed in the group treated with 750 mg/kg b. wt./day, which was considered as unrelated to the test item treatment due to being present in only 1 out of 25 rats in the respective group. Ultimobranchial cyst {Rat N° 1, 15 (G1) and 32 (G2)} was observed in the control and the group treated with 250 mg/kg bw/day, which were considered as unrelated to the test item due to being a congenital lesion.
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
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Statistically, a significant decrease in the mean absolute and relative (relative to both terminal body weight and brain weight) gravid uterus weight of pregnant female rats in the group treated with 750 mg/kg bw/day was observed when compared with that of the control group. This effect was considered as a treatment-related adverse effect of the test item.
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
food consumption and compound intake
other: gravid uterine weight
Abnormalities:
no effects observed
Fetal body weight changes:
effects observed, treatment-related
Description (incidence and severity):
The mean litter weight, mean weight of male, female, and total fetuses (male + female) was comparable between the control and the test item treated groups up to the dose level of 500 mg/kg bw/day.
Statistically, a significant decrease in the mean litter weight, mean fetus weight of male, female and for composite of both sexes were observed in pregnant female rats in the group treated with 750 mg/kg bw/day when compared to the control group. These effects were considered as an adverse effect of test item treatment.
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):
Statistically, a significant decrease in the mean litter weight, mean fetus weight of male, female and for composite of both sexes were observed in pregnant female rats in the group treated with 750 mg/kg bw/day when compared to the control group. These effects were considered as an adverse effect of test item treatment.
Anogenital distance of all rodent fetuses:
no effects observed
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment related external anomalies were observed in fetuses of the control and the treatment groups up to the dose level of 750 mg/kg b. wt./day. However, fetuses with hemorrhage and runt fetuses were observed in the control and the treatment groups as specified in the attached information.
Skeletal malformations:
effects observed, treatment-related
Description (incidence and severity):
Statistically, a significant increase in the percent litter with 14th rib: extra ossification center (left) was observed in the group treated with 250 mg/kg bw/day. This finding was considered as incidental due to the lack of dose dependency.
Statistically, a significant increase in the percent litter affected with xiphisternum: bipartite ossification was observed in the group treated with 500 mg/kg bw/day. This finding was considered as incidental due to the lack of dose dependency.
Statistically, a significant increase in the percent of fetuses with various anomalies was observed in the group treated with 750 mg/kg bw/day as specified in the attached information.

A significant increase in the percent litter affected with xiphisternum: unossified was considered as non-adverse effect of the test item caused by a mild; non-statistically significant decrease of the fetal mean body weight in the group treated with 500 mg/kg bw/day.
The increased incidences of percent fetus and percent litter affected with various anomalies were well correlated with the decreased fetal mean body weight in the group treated with 750 mg/kg bw/day.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment related visceral anomalies were observed in fetuses of the control and the treatment groups up to the dose level of 750 mg/kg bw/day. However, fetuses with hemorrhagic adrenals, dilated renal pelvis, hemorrhagic kidneys, malposition liver, hemorrhagic lungs, elongated thymus, and convoluted ureter were observed in the control and the treatment groups as specified in the attached information.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
Head Razor Observations:
No treatment related anomalies were observed in fetuses of the control and treatment groups up to the dose level of 750 mg/kg bw/day. However, fetuses with dilated 3rd and lateral ventricle were observed in the control and the treatment groups as specified in the attached information.
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes
other: skeletal variations
Abnormalities:
effects observed, treatment-related
Localisation:
skeletal: vertebra
Description (incidence and severity):
The increased incidences of percent fetus and percent litter affected with unossified or incompletely ossified sternebrae in the group treated with 750 mg/kg bw/d were correlated with decreased maternal body weight gain and uterus weight (absolute and relative) as well as a reduction in mean fetal weight.

The increased incidences of percent fetus and percent litter affected with absent 1st, 2nd and 3rd caudal vertebrae in the group treated with 750 mg/kg bw/d were considered as treatment-related adverse effect.

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 occurring together with maternal toxicity effects, but not as a secondary non-specific consequence of maternal toxicity effects
Dose response relationship:
yes
Relevant for humans:
not specified
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:
yes
Relevant for humans:
not specified

Please refer to the attached information.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
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

This study was performed to evaluate the prenatal developmental and maternal toxicity potential of Methyl benzoate when administered through oral gavage in pregnant Wistar rats (25/dose level) at dose levels of 0 (control), 250, 500, and 750 mg/kg bw/d in corn oil (dose volume: 4 mL/kg bw/d) from day 3 of gestation to day 19 of gestation in accordance with OECD Guideline 414 (June 25, 2018).


 


Maternal toxicity (effect on mortality, clinical sign, body weight, body weight gain, food consumption, uterine weight, thyroid weight etc.) and developmental toxicity (effect on fetal weight, ano-genital distance, fetal count, fetal gross external examination, fetal visceral examination, fetal head razor examination, fetal skeletal examination etc.) were evaluated.


 


No mortality or morbidity was observed in control and in the groups treated with 250, 500 and 750 mg/kg bw/d.


Clinical signs of toxicity such as salivation was observed in pregnant rats of the groups treated with 500 and 750 mg/kg bw/d. Salivation is often observed in gavage studies and may be a reaction to the taste or irritation of the test article rather than an indication of toxicity This was considered a physiological response to the dose formulation and toxicologically non-adverse in nature (Wook-Joon Yu et al., 2011).


The reproductive performance/pregnancy rate was comparable between the control and the test item treated groups.


No treatment-related adverse effects were observed in the groups treated with 250 and 500 mg/kg bw/d for any of the evaluated maternal and fetal parameters.


Treatment-related adverse effects were observed in the group treated with 750 mg/kg bw/d dose group as follows:



  • A decrease in the mean body weight gain of the pregnant female rats was observed during the gestation period of GD 8-11, 11-14, 17-20, and 5-20.

  • A statistically significant decrease in the mean body weight gain of the pregnant female rats was observed during the gestation period of GD 14-17.

  • A statistically significant increase in the mean food consumption of the pregnant female rats was observed during the gestation period of GD 11-14, 14-17, and 5-20.

  • A statistically significant decrease in the mean absolute and mean relative (relative to both terminal body weight and brain weight) gravid uterine weight of the pregnant female rats was observed.

  • A statistically significant decrease in the mean fetal body weight of male, female, mean total fetal body weight (male + female) and mean litter weight was observed.

  • A statistically significant increase was observed in the percent fetus and percent litter with 1st sternebrae: unossified, 2nd sternebrae: incomplete ossification, 3rd sternebrae: unossified, 4th sternebrae: unossified, xiphisternum: unossified, and 1st, 2nd, and 3rd caudal vertebrae: absent for fetal skeletal evaluation.

  • A statistically significant increase was observed in the percent fetus with pubis: unossified (bilateral), 2nd sternebrae: unossified, and 5th sternebrae: unossified for fetal skeletal evaluation.


 


The results of the present study suggest that a dose of 750 mg/kg bw/d produced reduction in maternal body weight gain and uterus weight (absolute and relative). Reduction in mean fetal weight as well as increase in fetal skeletal anomalies were also observed at the dose level of 750 mg/kg bw/d.


 


The increased incidences of percent fetus and percent litter affected with unossified or incompletely ossified sternebrae in the group treated with 750 mg/kg bw/d were correlated with decreased maternal body weight gain and uterus weight (absolute and relative) as well as a reduction in mean fetal weight and can therefore be considered as growth retardation secondary to maternal toxicity.


 


The increased incidences of percent fetus and percent litter affected with absent 1st, 2nd and 3rd caudal vertebrae in the group treated with 750 mg/kg bw/d and were considered as treatment-related adverse effect.


 


Considering that similar observations were not observed at the dose level of 500 mg/kg bw/d, it is concluded that the “No Observed Adverse Effect Level (NOAEL)” of methyl benzoate for maternal and developmental toxicity is 500 mg/kg bw/d.

Justification for classification or non-classification

The report of the PNDT study is currently under review and not finalised, yet. Therefore, no final conclusion can be reached at this stage.


The results will be thoroughly assessed, and the study report will be finalised within the next 3 months. The dossier will then be updated accordingly.

Additional information