Registration Dossier

Diss Factsheets

Administrative data

Description of key information

Oral:


OECD 422:


Wistar Han rats were treated with Propylidynetrimethanol, ethoxylated, esters with acrylic acid (TMPeoTA) by daily oral gavage at dose levels of 100, 300 and 1000 mg/kg/day in a combined 28-day repeated dose toxicity study with the reproduction/developmental toxicity screening test, in accordance to OECD 422. The rats of the control group received the vehicle, corn oil, alone. Males were treated for 2 weeks prior to mating, during mating, and up to termination (for 29 days). Females that delivered offspring were treated for 2 weeks prior to mating, during mating, during post-coitum, and at least 13-15 days of lactation (for 51-55 days (most females) or 63 days (one Group 3 female)). Females which did not mate or failed to deliver were treated for 52 days or 42 days, respectively. Based on the results, the following No Observed Adverse Effect Levels (NOAEL) for Propylidynetrimethanol, ethoxylated, esters with acrylic acid (TMPeoTA) were established:


Parental (systemic) NOAEL: 1000 mg/kg/day


Parental (local) NOAEL: 100 mg/kg/day


 


OECD 408:


The administration of the read-across substance (GPTA) - Glycerol, propoxylated, esters with acrylic acid (CAS 52408-84-1) by daily oral gavage according to OECD TG 408 at dose levels of 50, 150 and 375 mg/kg bw/day was well tolerated in rats at levels up to 150 mg/kg bw/day (ReachCentrum, 2020). At 375 mg/kg bw/day adverse findings were noted in the forestomach only. These findings are consistent with a response to an irritant material. Based on these results, the following was concluded:


Parental (systemic): 375 mg/kg bw/day


Parental (local): 150 mg/kg bw/day


 


In addition,GPTA was tested in a Combined 28-Day Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test according to OECD TG 422 and in compliance with GLP (ReachCentrum, 2019). Wistar Han rats were treated by daily oral gavage at dose levels of 150, 375 and 750 mg/kg bw/day. Microscopic evaluation showed local alterations of the forestomach starting at 150 mg/kg bw/day in males (non-adverse) and at 375 mg/kg bw/day in females. In addition, ulcers which were degenerative in nature were noted in males at 375 mg/kg bw/day and at 750 mg/kg bw/day (in both males and females). At 750 mg/kg bw/day, two males and two females were sacrificed in extremis and one female was found dead. The systemic NOAEL is 375 mg/kg bw/day, based on treatment related mortality at 750 mg/kg bw/day. The local NOAEL is 150 mg/kg bw/day, based on adverse local forestomach effects at 375 mg/kg bw/day (males) and 750 mg/kg bw/day (both sexes).


Parental (systemic) NOAEL: 375 mg/kg/day


Parental (local) NOAEL: 150 mg/kg/day,


 


Dermal:


No data available.


Supportive data available from a carcinogenicity, mice, dermal, 94 weeks (2 times/week/25µL 10% v/v acetone): not carcinogenic, no systemic effects NOAEL ≥ 19.5 mg/kg bw/d (Cytec, Kettering Laboratory, 1987) but less reliable as performed with an outdated protocol (Klimisch score 3).  


 


Inhalation:


No data available


 


For DNEL derivation, a NOAEL of 375 mg/kg bw/day from the OECD 408 study using GPTA (read-across substance) is used.

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
12 Jun 2019 - 18 Nov 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Please note: The Lead has been transferred recently form BASF Health and Care Products France S.A.S. to BASF SE. Accordingly its BASF SE (actual LEAD), who is submitting in time the requested information (OECD 408), which has been addressed as a Decision on a testing proposal to BASF Health and Care Products France S.A.S (old LEAD) before the LEAD change took place. Please do not hesitate to come back to us if you have open questions on this topic.
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
2018
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
Version / remarks:
1998
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Species:
rat
Strain:
Wistar
Details on species / strain selection:
The rat was chosen as the animal model for this study as it is an accepted rodent species for preclinical toxicity testing by regulatory agencies.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: 6-7 weeks
- Weight at study initiation: males weighed between 146 and 174 g and females weighed between 121 and 156 g
- Housing: in polycarbonate cages (Makrolon type IV)
- Diet: ad libitum, rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany)
- Water: ad libitum, municipal tap water
- Acclimation period: 14 days

DETAILS OF FOOD AND WATER QUALITY: The feed was analyzed by the supplier for nutritional components and environmental contaminants. It was considered that there were no known contaminants in the feed that would interfere with the objectives of the study. Periodic analysis of the water was performed, and it was considered that there were no known contaminants in the water that would interfere with the objectives of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 to 23
- Humidity (%): 47 to 68
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The dosing formulations were prepared weekly, filled out in daily portions and stored in the refrigerator protected from light. The dosing formulations were removed from the refrigerator and stirred for at least 30 minutes before dosing. The dosing formulations were dosed within 6 hours after adding the vehicle to the test item or removal from the refrigerator. Test item dosing formulations were kept at room temperature until dosing. If practically possible, the dosing formulations and vehicle were continuously stirred until and during dosing. Adjustment was made for specific gravity of the vehicle and test item. No correction was made for the purity/composition of the test item.

VEHICLE
- Justification for use and choice of vehicle (if other than water): The test item is soluble in corn-oil.
- Concentration in vehicle: 0, 10, 30, 75 mg/mL
- Amount of vehicle: 5 mL/kg
Analytical verification of doses or concentrations:
yes
Remarks:
Acquity UPLC system
Details on analytical verification of doses or concentrations:
The concentrations analyzed in the formulations of Groups 2, 3 and 4 were in agreement with target concentrations (i.e. mean accuracies between 90% and 110%). At first instance, the mean accuracies for the formulation of Groups 2, 3 and 4, prepared for use in Week 1, were below the target concentration (i.e. 73%, 81% and 72% of target). Differences in peaks from the vehicle were observed and this could result in differences in test item concentration but also in poor homogeneity of the solutions. Therefore it was decided to shake the samples again and re-analyze. The concentrations re-analyzed in the formulations of all groups were in agreement with target concentrations. Therefore it was concluded that formulation were prepared accurately.
No test item was detected in the Group 1 formulations. The formulations of Group 2 and Group 4 were homogeneous (i.e. coefficient of variation ≤ 10%).
Duration of treatment / exposure:
90 days
Frequency of treatment:
daily
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
Group 1
Dose / conc.:
50 mg/kg bw/day (nominal)
Remarks:
Group 2
Dose / conc.:
150 mg/kg bw/day (nominal)
Remarks:
Group 3
Dose / conc.:
375 mg/kg bw/day (nominal)
Remarks:
Group 4
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
The dose levels were selected based on results of a combined 28-day repeated dose toxicity study with reproduction/developmental toxicity screening with oral exposure of the test item. In the previous study, test item related mortality was noted at 750 mg/kg bw/day and adverse effects on the stomach were noted at 750 mg/kg bw/day and 375 mg/kg bw/day (males only), no adverse effects were noted at 375 mg/kg bw/day (females) and 175 mg/kg bw/day. Dose levels for this study were selected in an attempt to produce graded responses to the test item. The high-dose level should produce some toxic effects, but not excessive that would prevent meaningful evaluation. The mid-dose level is expected to produce minimal to moderate toxic effects. The low-dose level should produce no observable indications of toxicity.

- Fasting period before blood sampling for clinical biochemistry: yes
Positive control:
None
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: Animals were weighed individually weekly, starting on Day 1. A fasted weight was recorded on the day of necropsy.
In order to monitor the health status, Animal No. 35 was also weighed on Day 81.

FOOD CONSUMPTION: Yes
Food consumption was quantitatively measured weekly starting on Day 1 and continuing weekly throughout the Dosing Period.

WATER CONSUMPTION:
Subjective appraisal was maintained during the study, but no quantitative investigation introduced as no effect was suspected.

OPHTHALMOSCOPIC EXAMINATION: Yes
The eyes were examined using an ophthalmoscope after application of a mydriatic agent (Tropicol 5 mg/ml solution) during pretreatment in all animals, and at the end of the Dosing Period in Week 13 in all Group 1 and 4 animals.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: end of treatment
- Anaesthetic used for blood collection: Yes, isoflurane
- Animals fasted: Yes
- How many animals: all
- Parameters checked: White blood cells (WBC), Neutrophil (absolute), Lymphocyte (absolute), Monocyte (absolute), Eosinophil (absolute), Basophil (absolute), Large unstained cells (LUC) (absolute), Red blood cells, Reticulocyte (absolute), Red Blood Cell Distribution Width (RDW), Haemoglobin, Haematocrit, Mean corpuscular volume (MCV), Mean corpuscular haemoglobin (MCH), Mean corpuscular haemoglobin concentration (MCHC), Platelet, Prothrombin Time (PT), Activated Partial Thromboplastin Time (APTT)

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: end of treatment
- Animals fasted: Yes
- How many animals: all
- Parameters checked: Alanine aminotransferase (ALAT), Glucose, Aspartate aminotransferase (ASAT), Cholesterol, Alkaline Phosphatase (ALP), HDL and LDL Cholesterol, Total protein, Sodium, Albumin, Potassium, Total Bilirubin, Chloride, Urea, Calcium,
Creatinine, Inorganic Phosphate (Inorg. Phos)

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
Functional tests were performed on the first 5 animals per sex per group during Week 12-13. These tests were performed after dosing and after completion of clinical observations (including arena observation, if applicable).
- Battery of functions tested: Hearing ability (HEARING), Pupillary reflex (PUPIL L/R), Static righting reflex (STATIC R), Fore- and hind-limb grip strength, recorded as the mean of three measurements per animal, Locomotor activity

IMMUNOLOGY: No

OTHER:
Estrous Cycle Determination
On the day of necropsy a vaginal lavage was performed to determine the stage of the estrus for all females. Estrous cycles were evaluated by examining the vaginal cytology of samples obtained by vaginal lavage.
Data were only used for evaluation of extreme thyroid hormone effects in females and are not included in the results section.

Thyroid Hormone
Serum samples at a target volume of 1.0 mL were collected in tubes without anticoagulant. Blood samples were processed for serum, which was analyzed for the parameters Triiodothyronine (T3), Thyroxine (T4) and Thyroid-Stimulating Hormone (TSH)

Sacrifice and pathology:
GROSS PATHOLOGY: Yes
Main study and recovery animals were subjected to a complete necropsy examination, which included evaluation of the carcass and musculoskeletal system; all external surfaces and orifices; cranial cavity and external surfaces of the brain; and thoracic, abdominal, and pelvic cavities with their associated organs and tissues.

HISTOPATHOLOGY: Yes
See tabels
Statistics:
Parametric/Non-Parametric:
Levene’s test was used to assess the homogeneity of group variances.
The groups were compared using an overall one-way ANOVA F-test if Levene’s test was not significant or the Kruskal-Wallis test if it was significant. If the overall F-test or Kruskal-Wallis test was found to be significant, then pairwise comparisons were conducted using Dunnett’s or Dunn’s test, respectively.
Non-Parametric:
The groups were compared using an overall Kruskal-Wallis test. If the overall Kruskal-Wallis test was found to be significant, then the above pairwise comparisons were conducted using Dunn’s test.
ANCOVA:
The data corresponding to a response variable of interest and to a related covariate were submitted to an analysis of covariance (ANCOVA), including only groups with at least three non-missing paired values and if found to be significant, then pairwise comparisons were conducted using Dunnett’s test.
Incidence:
A Fisher’s exact test was used to conduct pairwise group comparisons of interest.

Statistics for data collected/processed in Toxdata:
All statistical tests were conducted at the 5% significance level. All pairwise comparisons were conducted using two sided tests and were reported at the 1% or 5% levels.

Parametric
Datasets with at least 3 groups (the designated control group and 2 other groups) were compared using Dunnett-test (many-to-one-t-test). For the motor activity data set (at least 3 groups) parametric (ANOVA) tests on group means were applied with Bonferroni correction for multiple testing. Mixed modelling techniques, comparing six different covariance structures, were used in order to select the best fitting statistical model.
Non-Parametric
Datasets with at least 3 groups were compared using a Steel-test (many-to-one rank test).
Incidence
An overall Fisher’s exact test was used to compare all groups. The above pairwise
comparisons were conducted using Fisher’s exact test whenever the overall test was significant.

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
No toxicologically relevant clinical signs were noted in surviving animals during daily detailed clinical observations or during weekly arena observations.
Salivation seen after dosing among all animals treated at 150 and 375 mg/kg bw/day from Days 11 and 2 onwards, respectively, was considered not toxicologically relevant, taking into account the nature and minor severity of the effect and its time of occurrence (i.e. after dosing). This sign was considered to be a physiological response related to taste of the test item rather than a sign of systemic toxicity. Other clinical signs noted during the Dosing Period occurred within the range of background
findings to be expected for rats of this age and strain which are housed and treated under the conditions in this study and did not show any apparent dose-related trend. At the incidence observed, these were considered to be unrelated to treatment with the test item.
Mortality:
mortality observed, non-treatment-related
Description (incidence):
There were two premature deaths at 375 mg/kg bw/day, both on Day 81.
Male No. 31 was found dead on Day 81. No relevant signs preceded its death and body weight was normal. Noteworthy microscopic findings included erosion/ulcer, inflammation, and exudate in the caudal ventral nasal cavity and trachea, and erosion/ulcer in the larynx. Collectively, the type and distribution of these lesions is highly suggestive of gastroesophageal reflux and aspiration of gastric content/test item, which was considered to be the cause of death. Male No. 35 was euthanized due to poor condition on Day 81. This animal showed laboured breathing and abnormal breathing sounds, hunched posture and erected fur prior to sacrifice. Additionally, a body weight loss of 7% was recorded over the last 3 days prior to sacrifice.
Noteworthy microscopic findings included erosion/ulcer, inflammation and exudate in the trachea (without concurrent lesions in the nasal cavity, nasopharynx, or larynx), and this was considered to be the major factor contributing clinical signs leading to euthanasia in this animal. Without clear corroborative upper airway lesions, there is less evidence for nasogastric reflux/aspiration in this animal, thus the lesions in the trachea may be related to a gavage error. Similar to terminally euthanized animals at 375 mg/kg/day, test item-related squamous hyperplasia of the non-glandular stomach (forestomach) mucosa was also noted in both preterminal animals, and erosion/ulcer was present in Animal No. 31.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
No toxicologically relevant effect on body weight and body weight gain was noted after treatment up to 375 mg/kg/day.
A reduced body weight gain was noted for males treated at 375 mg/kg/day from Day 78 onwards, resulting in lower mean body weights on Days 85 and 91 (-5%, not statistically significant) compared to controls. Based on the magnitude of the change this was considered not toxicologically relevant.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Food consumption of animals treated up to 375 mg/kg/day was similar to the control level over the study period.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No ophthalmology findings were noted that were considered to be related to treatment with the test item.
The nature and incidence of ophthalmology findings noted during the Pretreatment Period and in Week 13 was similar among the groups, and occurred within the range considered normal for rats of this age and strain. These findings were therefore considered to be unrelated to treatment with the test item.
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
No test item-related changes were noted in hematological parameters after treatment up to 375 mg/kg bw/day.
Basophil count was statistically significantly increased in females treated at 375 mg/kg/day compared to controls. Based on the absence of changes in other white blood cell parameters and on the minimal magnitude of the change, this was considered not toxicologically relevant. Other statistically significant changes in haematology parameters were considered to be unrelated to treatment as these occurred in the absence of a dose-related trend.
No test item-related changes were noted in coagulation parameters after treatment up to 375 mg/kg/day. The statistically significant lower prothrombin time (PT) of males treated at 50 and 375 mg/kg/day and females treated at 50 and 150 mg/kg/day was considered to be of no toxicological relevance as the opposite effect (i.e. an increase) would be expected in case of target organ toxicity.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
No test item-related changes were noted in clinical chemistry parameters after treatment up to 375 mg/kg bw/day.
An increased urea concentration was noted for females treated at 150 and 375 mg/kg bw/day (1.24x and 1.20x of controls, respectively). In absence of a dose related response and as all values remained within the historical control range1, this was considered not toxicologically relevant.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
Hearing ability, pupillary reflex, static righting reflex and grip strength were normal in all examined animals. Motor activity was similar between treated and control groups. All groups showed a similar motor activity habituation profile with a decreasing trend in activity over the duration of the test period.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
There were no test item-related alterations in organ weights.
Any differences, including those that reached statistical significance were considered not to be related to treatment with the test item due to the direction of the change, lack of doserelated pattern, and/or general overlap and variability in individual values.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Test item-related macroscopic changes were observed in the stomach of males and females dosed at 375 mg/kg bw/day group males and females. In the non-glandular mucosa of the stomach (forestomach), irregular mucosal surface was noted in 7/8 males and 6/10 females. The remainder of the recorded macroscopic findings were within the range of background gross observations encountered in rats of this age and strain.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Test item-related microscopic findings after treatment with the test item were noted in the stomach of the 150 and 375 mg/kg bw/day males and females.

Squamous cell hyperplasia of the stomach (forestomach), up to moderate degree, was noted in males and females starting at 150 mg/kg/day, with a dose-related increase in incidence and severity. This was characterized by diffuse thickening of the squamous mucosa, accompanied by hyperkeratosis (predominantly orthokeratotic). When mild or moderate, the epithelium was often thrown into simple folds, without cellular atypia.
Focal or multifocal erosion/ulcer of the non-glandular mucosa (forestomach) was noted, in conjunction with hyperplasia in males and females at 375 mg/kg/day. There were no other test item-related histologic changes. The remainder of the recorded
microscopic findings were within the range of background pathology encountered in rats of this age and strain. There was no test item-related alteration in the prevalence, severity, or histologic character of those incidental tissue alterations. A single (minimal) focal erosion/ulcer at the limiting ridge, specifically at the junction between the squamous and glandular mucosa was noted in one female in each of the control, 50 mg/kg/day, and 150 mg/kg/day group and was interpreted to be a spontaneous change when only observed at that location.
Histopathological findings: neoplastic:
not examined
Key result
Dose descriptor:
NOAEL
Remarks:
local
Effect level:
150 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
histopathology: non-neoplastic
Remarks on result:
other: These findings are consistent with a response to an irritant material.
Key result
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
375 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: highest dose tested
Key result
Critical effects observed:
no
Conclusions:
The administration of the read-across substance (GPTA) - Glycerol, propoxylated, esters with acrylic acid (CAS 52408-84-1) by daily oral gavage according to OECD TG 408 at dose levels of 50, 150 and 375 mg/kg bw/day was well tolerated in rats at levels up to 150 mg/kg bw/day (ReachCentrum, 2020). At 375 mg/kg bw/day adverse findings were noted in the forestomach only. These findings are consistent with a response to an irritant material. Based on these results, the following was concluded:
Parental (systemic): 375 mg/kg bw/day
Parental (local): 150 mg/kg bw/day
Executive summary:

The administration of the read-across substance (GPTA) - Glycerol, propoxylated, esters with acrylic acid (CAS 52408-84-1) by daily oral gavage according to OECD TG 408 at dose levels of 50, 150 and 375 mg/kg bw/day was well tolerated in rats at levels up to 150 mg/kg bw/day (ReachCentrum, 2020). At 375 mg/kg bw/day adverse findings were noted in the forestomach only. These findings are consistent with a response to an irritant material. Based on these results, the following was concluded:

Parental (systemic): 375 mg/kg bw/day

Parental (local): 150 mg/kg bw/day

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
375 mg/kg bw/day
Study duration:
subchronic
Experimental exposure time per week (hours/week):
37
Species:
rat
Quality of whole database:
One OECD 422 study performed in accordance to GLP available for TMPeoTA (CAS 28961-43-5)
One OECD 408 study performed in accordance to GLP available for the read-across substance GPTA (CAS 52408-84-1) - read-across substance
One OECD 422 study performed in accordance to GLP available for the read-across substance GPTA (CAS 52408-84-1) - read-across substance

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

A valid short-term study oral toxicity study (OECD 422) is available to assess the repeated dose toxicity of ethoxylated trimethylolpropane triacrylate (TMPeoTA CAS: 28961-43-5). Further information about the oral toxicity of similar acrylate substances (OECD 408) can be adapted from the read-across substance Glycerol, propoxylated, esters with acrylic acid (GPTA CAS: CAS 52408-84-1).

 

Read across to structural similar substances:

Information about the oral toxicity of similar acrylate substances (OECD 408) can be adapted from the read-across substance Glycerol, propoxylated, esters with acrylic acid (GPTA CAS: CAS 52408-84-1).  

 

Oral:

A study according OECD TG 408 was conducted to determine the potential toxicity of the read-across substance (GPTA) item after repeated exposure. Wistar Han rats were treated with the test item for 13 weeks by daily oral gavage at dose levels of 50, 150 and 375 mg/kg bw/day. The control group received the vehicle corn oil. Chemical analyses of formulations were conducted on Day 1 and in Weeks 6 and 13 to assess accuracy and homogeneity. The following parameters and end points were evaluated in this study: mortality/moribundity, clinical signs, body weights, food consumption, ophthalmology, functional observation tests, clinical pathology parameters (hematology, coagulation and clinical chemistry), gross necropsy findings, organ weights, and histopathologic examinations. Test formulations prepared were considered homogeneous at the concentrations tested and analysis of the accuracy revealed acceptable levels. There were two premature deaths during the course of the study, both on Day 81. One male at 375 mg/kg/day was found dead and the lesions observed in this animal were suggestive of gastro-esophageal reflux and aspiration of gastric content/test item as the cause of death. Another male at 375 mg/kg/day was euthanized in extremis based on labored breathing and abnormal breathing sounds, hunched posture and erected fur. Erosion/ulcer, inflammation and exudate in the trachea was considered to be the cause of the clinical signs leading to euthanasia. In absence of corroborative upper airway lesions, this death was likely related to a gavage-related incident. At necropsy of the surviving animals, a test item-related irregular mucosal surface was noted in the stomach of males and females treated at 375 mg/kg bw/day. This correlated to the microscopic changes in the forestomach (non-glandular stomach, squamous cell epithelium) at 150 (non-adverse) and 375 mg/kg bw/day, consisting of diffuse hyperplasia, which was occasionally accompanied by minimal focal or multifocal erosions/ulcers. These findings are consistent with a response to an irritant material. This was interpreted to represent a local effect rather than a systemic effect as no other test item-related changes were noted in any other examined organs. Due to the moderate severity grade and the presence of erosions/ulcers, the changes at 375 mg/kg bw/day were considered to be adverse. The test item-related squamous cell hyperplasia of the forestomach at 150 mg/kg bw/day was considered non-adverse, in absence of erosions/ulcers. No toxicologically significant changes were noted in any of the remaining parameters investigated in this study (i.e., clinical appearance, body weight, food consumption, functional observations, ophthalmoscopy, clinical laboratory investigations (i.e., hematology, coagulation and clinical chemistry) and organ weights).

In conclusion, administration of the test item daily oral gavage was well tolerated in Wistar Han rats at levels up to 150 mg/kg bw/day. At 375 mg/kg bw/day adverse findings were noted in the stomach. Based on these results, the NOAEL for local effects was considered to be 150 mg/kg bw/day, the NOAEL for systemic toxicity was at 375 mg/kg bw/day (ReachCentrum, 2020).

In addition, the GPTA was tested in a Combined 28-Day Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test according to OECD TG 422 and in compliance with GLP (ReachCentrum, 2019). Wistar Han rats were treated by daily oral gavage at dose levels of 150, 375 and 750 mg/kg bw/day. The rats of the control group received the vehicle, corn oil, alone. Males were treated for 2 weeks prior to mating, during mating, and up to termination (for 29 days). Females that delivered offspring were treated for 2 weeks prior to mating, during mating, during post-coitum, and at least 14-16 days of lactation (for 50-58 days). Females that failed to deliver pups were treated for 39-54 days. Chemical analyses of formulations were conducted once during the study to assess accuracy and homogeneity. The following parameters and end points were evaluated in this study: mortality/ moribundity, clinical signs, functional tests, body weight and food consumption, estrous cycle length and regularity, clinical pathology, serum level of thyroid hormone T4 (F0-males), gross necropsy findings, organ weights and histopathologic examination. In addition, the following reproduction/developmental parameters were determined: mating and fertility indices, precoital time, number of implantation sites, gestation index and duration, parturition, maternal care, sex ratio, live litter size and early postnatal pup development (mortality, clinical signs, body weight, anogenital distance, areola/nipple retention, serum level of thyroid hormone T4 in PND 14-16 pups and macroscopy). Test formulations prepared were considered homogeneous at the concentrations tested and analysis of the accuracy revealed acceptable levels.Microscopic evaluation showed local alterations of the forestomach starting at 150 mg/kg in males (non-adverse at this dose level) and at 375 mg/kg in females (non-adverse at this dose level), which consisted of ulcers and/or lymphogranulocytic inflammation and/or squamous cell hyperplasia and/or hyperkeratosis and/or edema in the submucosa. In addition, ulcers which were degenerative in nature were noted in males at 375 mg/kg and at 750 mg/kg (in both males and females). In a dose-related incidence, an irregular surface of the stomach was noted at necropsy in treated animals. In addition, other corroborative findings included salivation observed in treated animals and increased neutrophil values noted in males at 750 mg/kg. The local forestomach effects were considered to be adverse at 375 mg/kg (males) and 750 mg/kg (both sexes). At 750 mg/kg, two males and two females were sacrificed in extremis and one female was found dead. For one male and one female, test item-related findings in the forestomach were considered to be the main cause of moribundity. For the other male the cause of mortality could not be established. For the two remaining females the observed mortality was considered to be related to the oral gavage procedure. At 375 mg/kg, the death of a single female was to be considered incidental. Other changes noted in treated rats, mostly at 750 mg/kg, were considered non-adverse as discussed below. In males, a treatment-related reduction in body weight gain was noted at 750 mg/kg, corresponding to a decreased food consumption during the first week of treatment. As the reductions in mean body weight (gain) and food consumption remained slight and appeared to recover in subsequent intervals, the differences were considered to be not adverse. Test item related increases in mean kidney and liver weights were observed at 375 (kidneys) and at 750 mg/kg (kidneys and liver). The increase in liver weights was correlated with centrilobular hepatocellular hypertrophy observed at microscopic examination. In absence of any degenerative or inflammatory changes, the slight change in liver weights was considered non-adverse. For the increase in kidney weight, no microscopic correlate was found and was considered to be non-adverse. An increased incidence and severity of extramedullary hematopoiesis was observed in females at 750 mg/kg. At the incidence and severity observed, this was considered to be a non-adverse finding. Exposure to the test item was not associated with obvious toxicity in the remaining parental parameters examined in this study (i.e. functional observations and clinical laboratory investigations (including male T4 thyroid hormone levels)).

In conclusion, based on the results of this combined 28-day repeated dose toxicity study with the reproduction/developmental toxicity screening test, the following no-observed-adverse effect level (NOAEL) of the substance were established:

Parental (systemic) NOAEL: 375 mg/kg bw/day, based on treatment related mortality at 750 mg/kg bw/day.

Parental (local) NOAEL: 150 mg/kg bw/day, based on adverse local forestomach effects at 375 mg/kg bw/day (males) and 750 mg/kg bw/day (both sexes).

 

Assessment repeated dose toxicity:

All available information is taken into account to assess the repeated dose toxicity of TMPeoTA. Key data available is from the OECD 422 oral study performed with TMPeoTA, the systemic NOAEL was 1000 mg/kg bw/day and NOAEL for local effects observed at 100 mg/kg bw/day. Further key data from an 90D oral study (OECD 408) with the read-across substance GPTA showed a systemic NOAEL at 375 mg /kg bw/day and NOAEL for local effects at 150 mg/kg bw/day. For DNEL derivation, the oral NOAEL of 375 mg/kg bw/day from the 90D study using GPTA (read-across substance) is used.

 

Key study assignment:

The OECD 408 study evaluating the repeated oral dose toxicity of GPTA (read-across substance) is considered the key study.

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. The substance is not considered to be classified for repeated dose toxicity according to Regulation (EC) No 1272/2008 (CLP), as amended for the twelfth time in Regulation (EU) 2019/521.

Categories Display