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Description of key information

No treatment-related effects indicating local or systemic toxicity were observed in male or female animals at any of the dose levels investigated.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Study duration:
subacute
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

A Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity screening test was conducted with the structural analogue in Sprague Dawley SD rats up to Day 4 post partum. The effects of the test item systemic or local toxicity as well as any possible effects on male and female reproductive performance, such as gonadal function, mating behaviour, conceptuses, parturition and early lactation of the offspring, were investigated at the dose levels of 62.5, 250 and 1000 mg/kg body weight/day. The reversibility of toxic effects after repeated dosing up to 4 consecutive weeks followed by a 2 week treatment-free period were also investigated, in order to assess recovery from any adverse effects observed during the dosing phase. The animals were assigned to four treatment main groups (Groups 1 to 4) of 10 animals/sex and two treatment recovery groups (Groups 5 and 6) of five animals/sex. The animals were administered orally by gavage at a constant volume of 5 mL/kg body weight. The test item was formulated in sesame oil at the final concentrations of 12.5, 50 and 200 mg/mL for treatment of Groups 2, 3 and 4/6 respectively. Control animals (Groups 1/5) received the vehicle alone at the same dose volume. Furthermore, genotoxicity assessment was also evaluated in males of the main groups, in order to assess the ability of the test item to induce cytogenetic damage in rat bone marrow, as measured by the induction of micronuclei in polychromatic erythrocytes.

Main groups: According to the study design, males were treated for 2 weeks prior to pairing and during pairing with females until the day before necropsy, up to 45 days. Females were treated for 2 weeks prior to pairing, during pairing and throughout the gestation and lactation periods until Day 3 post partum, for approximately 41-54 days, depending on the female’s performance. The following investigations were performed in all groups: mortality check, clinical signs (including neurotoxicity assessment, motor activity, grip strength and sensory reaction to stimuli), body weight, food consumption, oestrous cycle, mating performance, clinical pathology investigations (haematology and clinical chemistry), litter data, macroscopic observations and organ weights. Evaluation of body weight, clinical signs and macroscopic observations of pups were also performed. Routine histopathological examination was performed on control and high dose groups (five animals/sex/group randomly selected). The morphological evaluation of the seminiferous epithelium (staging of spermatogenic cycle) was also included.

Recovery groups: Animals of the recovery groups were treated for a total of 4 consecutive weeks and sacrificed after 2 weeks of recovery (Groups 5, 6). The following parameters were evaluated in these animals: mortality, clinical signs (including neurotoxicity assessment, motor activity, grip strength and sensory reaction to stimuli), body weight, food consumption, clinical pathology investigations (clinical chemistry and haematology with the exception of coagulation), macroscopic observations and organ weights. As no adverse effects were noted in main group animals, histopathology was not performed in recovery animals.

Mortality and fate of females: No mortality occurred throughout the study. Non-pregnant females were found in the mid- and in the high dose groups. One mid-dose female had unilateral implantation. The number of females with live pups on Day 4 post partum was: 10 in each of the control and low dose groups, 7 in the mid-dose group and 9 in the high dose group.

Clinical signs and observation of cage tray: Green staining of tail was the main clinical sign recorded during treatment and recovery period in the high dose animals. At observation of the cage tray, green staining was also recorded in the high dose group during the treatment period. Moreover, treated animals of the recovery group (Group 6) showed green staining in the cage tray also during the recovery period. This discolouration was due to the colour of the test item and its properties as textile dye.

Neurotoxicity assessment (removal of animals from the home cage and in an open arena) did not reveal changes attributable to the test item. No relevant differences in motor activity, grip strength and sensory reactivity to stimuli were noted between control and treated groups.

Body weight: Body weight and body weight gain were unaffected by treatment both in males and females throughout the main or recovery phases.

Food consumption: No changes in food consumption were recorded in male and female animals throughout the main or recovery phases.

Clinical pathology: No changes of toxicological relevance were recorded in clinical pathology investigation at the end of dosing or recovery periods.

Reproductive performance: Oestrous cycle, pre-coital intervals, copulatory and fertility indices did not show intergroup differences.

Implantation, pre-birth loss data and gestation length of females: No significant differences were observed in the number of implantations, corpora lutea, total litter size, pre-implantation loss and pre-birth loss between control and treated groups. The majority of dams gave birth on Day 22 post coitum.

Litter data: Reproductive outcomes of dams which included the number and body weight of pups and the percentage of live pups on Days 1 and 4 post partum did not differ between groups. Sex ratio of pups was also comparable between groups.

Clinical signs of pups: There were no treatment-related effects at clinical observation of pups.

Necropsy findings in deceased pups and in pups sacrificed on Day 4 post partum: No treatment-related findings were recorded at necropsy of deceased pups or those sacrificed at term.

Terminal body weight and organ weights: No relevant differences of toxicological significance were seen in terminal body weight or organ weights between controls and treated animals, both in main and recovery groups.

Macroscopic observations: Animals killed at termination did not show relevant macroscopic changes that could be treatment-related. No relevant changes were noted at post mortem examination in recovery animals, when compared with controls.

Microscopic observations: No microscopic alterations were noted in treated animals that could be considered treatment related.

Spermatogenic cycle: Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.

 

Conclusion: No treatment-related effects indicating local or systemic toxicity were observed in male or female animals at any of the dose levels investigated. No effects on sexual function and fertility or in developmental parameters and lactation were observed at any of dose level investigated.

Based on the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) for both general toxicity and reproduction/developmental toxicity was considered to be above 1000 mg/kg body weight/day for males and females of the parental and F1 generation.

Justification for classification or non-classification

No treatment-related effects indicating local or systemic toxicity were observed in male or female animals at any of the dose levels investigated