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Effects on fertility

Description of key information

Fertility:

- 90-day oral repeated dose toxicity (OECD TG 408): No fertility effects in the male and female reproductive organs were observed at the maximum dose of 500 mg/kg bw

- 90 -day dermal /oral repeated dose toxicity studies in rat and mice effects cannot be used for deriving systemic and thus fertility effects.

- 28-day repeated dose (OECD TG 407): No fertility effects in the male and female reproductive organs were observed at the maximum dose of 1000 mg/kg bw.

- An extended one-generation study is imposed in a final decision by ECHA

Developmental toxicity:

- Developmental toxicity in rat (OECD TG 414): Maternal NOAEL =240 mg/kg bw. Developmental NOAEL =>480 mg/kg bw

- Developmental toxicity in rabbit (OECD TG 4149: Maternal NOAEL is 200 mg/kg bw. Developmental NOAEL is >=500 mg/kg bw

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
Two oral gavage studies of 28- and 90-day no fertility effects are seen at the highest dose tested of 500 and 1000 mg/kg bw.
An extended one-generation reproduction toxicity study will be performed in the near future.
Effect on fertility: via dermal route
Quality of whole database:
The dermal studies are insufficient for deriving information on systemic toxicity including fertility a summary of all repeated dose studies is presented below.
Additional information

Reproductive toxicity in the repeated dose toxicity 90 days in rat - oral

The safety of the test substance OTNE was examined in a subchronic (13 week) oral toxicity study in Wistar rats, according to OECD TG 408. OTNE was administered by daily oral gavage as a dilution in corn oil at levels of 0 (vehicle control), 30, 120 and 500 mg/kg body weight/day to groups of 10 rats/sex during 13 weeks. No relevant findings were seen on fertility and there were no histopathological lesions in reproductive organs. The increase of the relative testes weight at the mid dose only is not toxicologically relevant (absolute is more important than relative weight for this organ. Some dilatation of the uterus was reported, however as it this was also observed in the control group it was not considered an effect related to exposure to OTNE. Overall no effects on fertility were observed in the animals exposed to OTNE. The NOAEL for fertility is >=500 mg/kg bw,

Reproductive toxicity in the repeated dose toxicity 90 days in mouse and rat - dermal

The study was performed to assess the dermal repeated dose systemic toxicity as a result of dermal exposure to OTNE in the mouse and rat. The study was performed comparable to OECD TG 411, with some deviations (Dermal application without occlusion (not justified); partial oral exposure through grooming is expected, the top dose tested in mice was above the recommended dose)). Solutions of OTNE in ethanol were applied to the skin of mice and rats for 5 days per week for 3 months. There were 10 rodents in each dose group. Doses for which reproductive effects were evaluated were 25% and 50% OTNE in ethanol and 100% OTNE. These doses correspond approximately to 500 to 2000 mg/kg bw/day in mice and 125 to 500 mg/kg bw/day in rat. During the course of this study, samples were collected for estrous cycle characterization. At the end of the study, samples were collected for reproductive tissue evaluations. OTNE exposure via dermal application showed some minor but significant fewer sperm and lower sperm motility in the 2000 mg/kg bw/day male group. In females at 2000 mg/kg bw the estrus cycle was significantly extended at this dose with one day. None of these effects were observed in rats. The effects in mice are all observed at the high dose of 2000 mg/kg bw where very increased of relative liver weight was seen (up to 90%). Up to 1000 mg/kg bw no adverse effects on fertility are seen in male and female mice. The NOAEL for fertility is set at 1000 mg/kg bw.

Reproductive toxicity in repeated dose toxicity 28 days in rat - oral

This study was performed to assess the systemic toxicity of OTNE to the rat according to OECD TG 407. OTNE was administered by oral gavage, once daily, to three groups of rats for a minimum of twenty eight consecutive days, at dosage levels of 15, 150 or 1000 mg/kg/day. The test material was prepared as suspensions in corn oil at concentrations of 0.3, 3.0 or 20% w/v and was administered at a dosage volume of 5 ml/kg bw/day. After the 28-day exposure period, no effects on fertility were observed in the animals exposed to OTNE, therefore the NOAEL is set at >= 1000 mg/kg bw.

Effects on developmental toxicity

Description of key information

Rat: Prenatal development toxicity study (OECD TG 414): maternal NOAEL 240 mg/kg bw/day, developmental NOAEL 480 mg/kg bw/day

Rabbit: Prenatal developmental toxicity study (OECD TG 414): maternal toxicity NOAEL is 200 mg/kg bw, developmental NOAEL >=500 mg/kg bw.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The quality of the database is high because in two different tests fertility has been investigated: the developmental toxicity study and a study in which the transfer across the placenta and into milk of rats during and after pregnancy following repeated oral administration was studied. These studies adequately fulfill the REACH requirements.
Additional information

Rat developmental toxicity study

A developmental toxicity study was performed in accordance with the US FDA Guideline on detection of toxicity to reproduction for medicinal products, which is equivalent to OECD TG 414. The study was performed under GLP conditions. Groups of pregnant rats (25/dose group) were gavaged with 0 (control), 96, 240, or 480 mg/kg bw/day on gestation days 7-17 at dosage volumes of 0.5, 0.1, 0.25, or 0.5 ml/kg bw, respectively. Viability, abnormal clinical signs, body weights, abortions, premature deliveries, and feed consumption were recorded. Mating performance was assessed. On gestation day 21, surviving rats were euthanized by carbon dioxide asphyxiation and Caesarean sections were performed to remove the foetuses. Rats underwent gross necropsy. Foetuses were assessed for litter parameters and gross external, soft tissue, or skeletal malformations. Maternal animals showed reduced body weights and persistent clinical signs (e.g., increased salivation and urine-stained abdominal fur) at the highest dose. There were no treatment-related effects on developmental parameters at any dose, although reduction (not statistically significant) in foetal body weight was noted at the highest dose. Based on these results, the maternal NOAEL was 240 mg/kg bw/day and the developmental NOAEL was 480 mg/kg bw/day.

Rabbit developmental toxicity study

In this rabbit developmental toxicity test according to OECD 414 (in compliance with GLP) the effects of the test substance was tested on pregnant New Zealand White rabbits and the development of the embryo and fetus following daily oral administration by gavage during gestation days (GD) 6-28.

Method: The dose levels were set at 0, 75, 200 and 500 mg/kg bw based on the absence of maternal toxicity in a dose range finding study in the same species. Eighty-eight time-mated rabbits arrived at the test facility and were allocated to one control group and three groups receiving the test substance by oral gavage. A dose volume of 2 mL/kg body weight was applied in all groups and corn oil was used as vehicle. Control animals received the vehicle only. In-life parameters included signs of morbidity and mortality, body weight and food consumption. On gestation day 29 the dams were sacrificed and examined macroscopically. Fetuses, placentas, reproductive organs and kidneys and livers were weighed. The fetuses were macroscopically examined and processed for visceral and skeletal examinations.

Results, Maternal toxicity:

Two animals died before the scheduled necropsy on gestation day 29, these deaths were not treatment-related. Indications of maternal toxicity were noted in the high dose group. Mean body weight of these animals was lower throughout the study compared to the control and other treatment groups, reaching a difference of ~5% on day 29. This is mainly ascribed to a significantly lower food consumption (~32% compared to the control) and the resulting body weight loss during the first three days of dosing (day 6 to 9). This growth retardations reflects the lower body weights throughout the study. The overall food consumption in the high dose group was reduced with ~17% compared to the control group.

Results, Developmental toxicity:

No effects of treatment were observed on pre-implantation and post-implantation loss. The number of live fetuses per dam was not affected by the treatment. The ossification status of the vertebrae and sternebrae did not indicate a treatment-related delayed development of the fetuses. Incomplete ossification of the forelimb and hindlimb was observed in all groups, as is expected in normal fetal development. Skeletal malformations of the vertebra and fused ribs occurred incidental and were randomly distributed in all groups, indicating no treatment related effect. This is supported by the absence of a treatment-related effect on variations in the shape vertebra. Malformation and variations in the shape of sternebrae were more common than skeletal malformation of the vertebra and ribs, but lacked a treatment-related trend. This results in a NOAEL of >=500 mg/kg bw for developmental toxicity.

Conclusions

The significant increase in the relative liver weight in the high dose group may be adaptive (induced to metabolise the substance) but is conservatively used to help setting the NOAEL. The body weight effects and reduced food consumption and the increased relative liver weights are used to derive the NOAEL of 200 mg/kg bw for maternal toxicity. Based on the absence of effects on implantation sites , i.e. resorptions, live/dead fetuses, and fetal development, i.e. malformations, variations, teratomas, the NOAEL for developmental toxicity in rabbits is ≥ 500 mg/kg bw/day.

Justification for classification or non-classification

Based on the results, which indicate no reproduction and developmental toxicity effects of OTNE, the substance does not need to be classified as toxic to reproduction according to EU CLP Regulation (EC 1272/2008 and its amendments).