Registration Dossier

Diss Factsheets

Administrative data

Endpoint:
one-generation reproductive toxicity
Remarks:
based on generations indicated in Effect levels (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 19 May 2014 to 12 Dec 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study performed according to OECD Guideline and GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report date:
2014

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
10/01/2013
Limit test:
no

Test material

Constituent 1
Test material form:
solid: particulate/powder

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Number: 82 rats (41 males and 41 females) were received at CiToxLAB France on 03 June 2014.
- Strain and sanitary status: Sprague-Dawley, RjHan: SD (Rats CD®).
- Breeder: Janvier, Le Genest-Saint-Isle, France.
- Age/Weight: At the beginning of the treatment period, the males were 10 weeks old and had a mean body weight of 454 g (range: 406 g to 475 g). The females were 9 weeks old and had a mean body weight of 213 g (range: 196 g to 242 g). The animals were sexually mature and were not siblings. The females were virgin.

- Housing: The animals were individually housed, except during mating and lactation, in polycarbonate cages (Tecniplast 2154, 940 cm2) with stainless steel lids and containing autoclaved sawdust (SICSA, Alfortville, France).
Individual housing was chosen since it is preferable for pregnant animals and to avoid any aggressive behavior around mating.
Toward the end of gestation and during lactation, autoclaved wood shavings (SICSA, Alfortville, France) were provided to females and their litters as nesting material.
Each cage contained an object for the environmental enrichment of the animals (rat hut).

- Diet (e.g. ad libitum): All animals had free access to SSNIFF R/M-H pelleted maintenance diet, batch No. 6730736 (SSNIFF Spezialdiäten GmbH, Soest, Germany), which was distributed weekly.
- Water (e.g. ad libitum): The animals had free access to bottles containing tap water (filtered with a 0.22 µm filter).
- Acclimation period: The animals were acclimated to the study conditions for a period of 7 days before the beginning of the treatment period. One supplementary animal per sex than necessary were allocated to the study and acclimated, in order to permit the selection and/or replacement of individuals.

ENVIRONMENTAL CONDITIONS
- temperature: 22 ± 2°C,
- relative humidity: 50 ± 20%,
- light/dark cycle: 12h/12h,
- ventilation: About 12 cycles/hour of filtered, non-recycled air.


IN-LIFE DATES: From: 03 June 2014 To: 01 August 2014

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test item was administered as a solution in the vehicle. The weighing of the test item was performed using de humidified bag system (glove box under nitrogen/argon), then the test item was mixed with the required quantity of vehicle.
The test item dose formulations were prepared with a frequency (8 days) depending on their stability according to an homogeneity and stability study performed at CiToxLAB France (CiToxLAB France/Study No. 41182 AHS), stored at +2-8°C protected from light prior to use and delivered to the study room at room temperature and protected from light.

VEHICLE
The vehicle was prepared by CiToxLAB France and was used for preparation of test item dose formulations and administered to control group (control dose formulation).
The vehicle was drinking water treated by reverse osmosis using ELIX 5 (Millipore SA).
Details on mating procedure:
The females were paired with males from the same dose-level group. One female was placed with one male, in the latter's cage, during the night.
Confirmation of mating was made in the morning by checking for the presence of a vaginal plug or for sperm in a vaginal lavage.
The day of confirmed mating was designated Day 0 p.c..
Each female was placed with the same male until mating occurs or 7 days had elapsed. Pairs with no evidence of mating after 7 days were separated and the female was placed for a further 7 days with a different male from the same dose-level group who had already mated.
The pre-coital time was calculated for each female.
Group 3 female C27975, with no evidence of mating after 7 days with the second male was sacrificed 25 days after the end of the mating period.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Diluted samples of dose formulations were analyzed in single by Ion Chromatography (IC) with conductivity detection.
The test item peak area was determined for each sample and the corresponding concentration was calculated using the equation obtained from the calibration data.
All the results are expressed as mg/mL of LiTFSI PURE.
The test item concentrations in the administered dose formulations analyzed in Weeks 01, 03 and 05 remained within an acceptable range of variation (+0.9% to +4.8%) when compared to the nominal values (± 10% of the nominal concentrations).
LiTFSI PURE was not detected in control samples.
Duration of treatment / exposure:
In the males:
- 2 weeks before mating,
- during the mating period (up to 2 weeks),
- until sacrifice (at least 4 weeks in total).

In the females:
- 2 weeks before mating,
- during the mating period (up to 2 weeks),
- during gestation,
- during lactation until Day 4 p.p. inclusive,
- until sacrifice for females with no evidence of mating or no delivery.

Day 1 corresponds to the first day of the treatment period.
Frequency of treatment:
Daily
Doses / concentrations
Remarks:
Doses / Concentrations:
15; 30 and 60 mg/kg/day
Basis:
actual ingested
No. of animals per sex per dose:
10
Control animals:
yes, concurrent no treatment
Details on study design:
- Dose selection rationale:
The dose-levels were selected, following the results of a previous 4-week toxicity study by oral route (gavage) in rats followed by a 2-week treatment-free period. In this previous study, the test item was administered at 15, 45 and 90 mg/kg/day.
The No Observable Effect Level (NOEL) was established at 15 mg/kg/day and the Lowest Observable Adverse Effect Level (LOAEL) was established at 45 mg/kg/day. Therefore, 60 mg/kg/day was selected as the high dose-level. The low-dose and mid-dose were selected using a ratio representing a 2-fold interval (i.e. 15 and 30 mg/kg/day).

Examinations

Parental animals: Observations and examinations:
Morbidity and mortality
Each animal was checked for mortality and morbidity once a day before the treatment period and at least twice a day during the treatment period, including weekends and public holidays.
Animal showing signs of poor condition, especially when death appeared imminent, was humanely sacrificed. Animals found dead or prematurely sacrificed were subjected to a macroscopic post-mortem examination.

Clinical signs
From arrival, the animals were observed once a day as part of routine examinations.
From the start of the treatment period, each animal was observed once a day, at approximately the same time, for the recording of clinical signs.

Body weight
The body weight of each male was recorded on the first day of treatment (Day 1), then once a week until sacrifice.
The body weight of each female was recorded on the first day of treatment (Day 1), then once a week until mated (or until sacrifice for females with no evidence of mating), on Days 0, 7, 14 and 20 post-coitum and on Days 1 and 5 p.p.

Food consumption
The quantity of food consumed by each male was measured once a week, over a 7-day period, from the first day of treatment until the start of the mating period.
The quantity of food consumed by each female was measured once a week, over a 7 day period, from the first day of treatment until the start of the mating period, during gestation for the intervals Days 0-7, 7 14 and 14-20 p.c. and during lactation for the interval Days 1-5 p.p..
During the mating period, food consumption was not measured for males or females.
Food intake per animal and per day was calculated by noting the difference between the food given and that in the food-hopper the next time.
Oestrous cyclicity (parental animals):
The estrous cycle stage was determined from a fresh vaginal lavage (stained with methylene blue), each morning during the mating period, until the females were mated.
Litter observations:
Females were allowed to litter normally and rear their progeny until Day 5 p.p..
The morning when parturition is completed was designated Day 1 p.p.. The length of gestation was calculated.

Each live pup was identified individually on Day 1 p.p., by subcutaneous injection of Indian ink.

Litter size
The total litter size and sex of each pup was recorded as soon as possible after birth.
The litters were observed daily in order to note the number of live, dead and cannibalized pups.

Clinical signs
The pups were observed daily for clinical signs, abnormal behavior and external abnormalities.

Body weight
The body weight of each pup was recorded on Days 1 and 5 p.p..
Postmortem examinations (parental animals):
SACRIFICE
On completion of the treatment period, all surviving animals were deeply anesthetized by an intraperitoneal injection of sodium pentobarbital and sacrificed by exsanguination:
- males: after the end of the mating period (at least 4 weeks of treatment in total),
- females: on Day 5 p.p.,
- females which do not deliver: on Day 25 p.c. (after a body weight recording to check for a possible un noticed delivery; sacrifice by inhalation of carbon dioxide gas followed by cervical dislocation were used if gestation was suspected),
- female with no evidence of mating: 25 days after the end of the mating period as no delivery occurred (sacrifice by inhalation of carbon dioxide gas followed by cervical dislocation were used if gestation was suspected),
- females with total litter loss: as appropriate.

MACROSCOPIC POST MORTEM EXAMINATION
A macroscopic post-mortem examination of the principal thoracic and abdominal organs was performed on all F0 animals including the two males found dead or sacrificed prematurely. Special attention was paid to the reproductive organs.
The numbers of corpora lutea and implantation sites were recorded for female sacrificed as scheduled on Day 5 p.p. and for female sacrificed 25 days after the end of the mating period with no evidence of mating and for females sacrificed on Day 25 p.c. due to no delivery.
For apparently non-pregnant females the presence of implantation scars on the uterus was checked using the ammonium sulphide staining technique (Salewski, 1964).

HISTOPATHOLGY
Epididymides, Mammary gland area, ovaries, prostate, seminal vesicles, testes, uterus, vagina

ORGAN WEIGHT
Epididymides, testes
Postmortem examinations (offspring):
SACRIFICE
- pups whose mother died: as soon as possible,
- surviving pups: on Day 5 p.p..

MACROSCOPIC POST MORTEM EXAMINATION
A detailed soft tissue examination of thoracic (including a detailed examination of the heart and great vessels) and abdominal organs was performed on all pups sacrificed on Day 5 p.p..
Macroscopic lesions were preserved without any further investigations in the first instance.
The findings were described according to the glossary of the International Federation of Teratology Societies (IFTS) and classified as malformations or variations (Chahoud et al., 1999, Makris et al., 2009):
- malformation refers to a permanent structural change that is likely to adversely affect the survival or health,
- variation refers to a change that occurs within the normal population under investigation and is unlikely to adversely affect survival or health (this might include a delay in growth or morphogenesis that has otherwise followed a normal pattern of development).

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive performance:
effects observed, treatment-related

Details on results (P0)

MORTALITY (PARENTAL ANIMALS)
MALES
In controls and at 15 mg/kg/day:
There were no unscheduled deaths.

At 30 mg/kg/day:
One male was found dead on Day 27 without any clinical signs prior to death. At macroscopic post-mortem examination, the ventricles of the heart, the liver and the kidneys were enlarged. Ureters showed translucent or brown content and were dilated. Dilated pelvis was noted in one kidney whereas black discoloration was seen in the other one.

At 60 mg/kg/day:
One male was sacrificed prematurely on Day 2. Prior to sacrifice, signs of poor clinical condition were observed (ptyalism, half-closed eyes, round back, tremors, clonic contraction, convulsions, hypoactivity, prostration, decrease in grasping reflex, locomotory difficulties and lateral decubitus). At macroscopic post-mortem examination, the prostate was gelatinous in consistency and the urinary bladder was dilated. Enlarged kidneys along with dilated ureters and dilated pelvis confirmed at microscopic evaluation were observed. These changes that could be noted occasionally in untreated animals contributed to the death of the animal.

FEMALES
In controls and at 15 mg/kg/day:
There were no unscheduled deaths.

At 30 mg/kg/day:
- One female was sacrificed for no delivery. This female was not pregnant,
- Three females were sacrificed for dead litter.
Prior to sacrifice, piloerection was observed in one female (C27979) sacrificed for dead litter. No necropsy findings were found at macroscopic post-mortem examinations.

At 60 mg/kg/day:
- Two females were sacrificed for no delivery. These females were not pregnant,
- Two females were sacrificed for humane reasons (moribunds) and one female was found dead,
- Five females were sacrificed for dead litter.

CLINICAL SIGNS (PARENTAL ANIMALS)
At 60 mg/kg/day, marked to severe clinical signs were observed both in males and females. They were mostly neurological disorders (i.e. tremors, clonic contraction, convulsions, tonic contraction, hypoactivity, prostration, decrease in grasping reflex, locomotory difficulties and/or hypersensitivity to the touch)
At 30 mg/kg/day, marked clinical signs were observed in one male on study Day 26 and were consistent with the neurological disorders described at the high dose-level (i.e. tremors, convulsions, and tonic contraction).
At 15 mg/kg/day, there were no test item treatment-related findings.

BODY WEIGHT (PARENTAL ANIMALS)
MALES
At 15 or 30 mg/kg/day, and when compared with controls, there were no effects.
At 60 mg/kg/day, and when compared with controls, there was a minimal lower mean body weight on Day 8 (-4.6%, p < 0.01) resulting in a lower mean body weight gain over the first week of treatment (+24 g vs. +38 g, p < 0.05).

FEMALES
Pre-mating and gestation periods: There were no effects.

Lactation period: When compared with controls, there were dose-related decreases in mean body weight and mean body weight change at 15 and 30 mg/kg/day. The difference was only statistically significant for mean body weight change at 15 mg/kg/day (+16 g vs. +26 g in controls, p < 0.05). The absence of statistically significant differences at the high dose-level was considered to be due to large variation between the females.

FOOD CONSUMPTION (PARENTAL ANIMALS)
MALES
From 15 mg/kg/day, and when compared with controls, there were minimal reductions in mean food consumption during the first week of the treatment period (down to 32 g/animal/day at 60 mg/kg/day vs. 36 g/animal/day). Thereafter, mean food consumption returned towards control values.

FEMALES
Pre-mating and gestation periods: There were no effects.
Lactation period: From 15 mg/kg/day, and when compared with controls, there was a lower mean food consumption (down to 35 g/animal/day at 30 mg/kg/day vs. 48 g/animal/day, p < 0.01).

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
A slightly higher mean pre-coital time (mean number of days taken to mate) was observed in females given 60 mg/kg/day, this was particularly due to one female which mated after 13 days (the female was blocked in diestrous for several days).
Lower fertility and gestation indexes were noted in females given 60 mg/kg/day.
All females were pregnant, except one and two females at 30 and 60 mg/kg/day, respectively.

At 60 mg/kg/day, the lower fertility and gestation indexes were considered to be related to the treatment with the test item.

ORGAN WEIGHTS (PARENTAL ANIMALS)
No changes that could be related to treatment were noted in testis and epididymis weights.

GROSS PATHOLOGY (PARENTAL ANIMALS)
All the necropsy findings observed in unscheduled deaths or at terminal sacrifice were recognized as those that are commonly noted in the rats of this strain and age, and none were considered to be of toxicological relevance.

HISTOPATHOLOGY (PARENTAL ANIMALS)
No microscopic changes that could be related to treatment were noted in the examined organs of males and females from group 4 (60 mg/kg)
Increased incidence and severity of follicles development were seen in the ovaries from unscheduled dead or surviving females in group 4. In 2/10 females, ovocytes were noted in the lumen of oviducts indicating that estrus just occurred. Recent basophilic corpora lutea were also observed in 3 of these females. All these changes are related to the estrus cycle and suggested that females from group 4 were possibly at a different stage than controls. An evaluation of these females to determine a more precise cycle stage was impossible in the absence of uterus and vagina examination. A relationship to treatment of this slight change in group 4 remained unclear.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
< 15 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on effect on body weight at this dose-level and, mortality and clinical signs from 30 mg/kg/day.
Dose descriptor:
NOAEL
Remarks:
reproductive performance and embryo/fetal development
Effect level:
30 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on lower fertility and gestation indexes at 60 mg/kg/day.

Results: F1 generation

General toxicity (F1)

Clinical signs:
effects observed, treatment-related
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Sexual maturation:
not examined

Details on results (F1)

VIABILITY (OFFSPRING)
At 15, 30 and 60 mg/kg/day, and when compared with controls, there was a dose-related decrease in viability index on Day 4 p.p. (79.6%, 47.5% and 0.0% vs. 95.7%, respectively). The decreases in viability indexes were the consequence of increased incidences of found dead and cannibalized pups on Days 1 and 2 p.p.

At 60 mg/kg/day, and when compared with controls, there was a high incidence of post-implantation loss (30.1% vs. 11.9) and, only a few females delivered with all pups found dead during the period of Days 1 4 p.p.
The mean number of pups per litter (14.1, 15.2 and 13.3 for the control, low-dose, mid-dose groups, respectively) was comparable in all groups. The number of male and female litter mates was comparable in all groups. No effects were observed on the live born index or on the number of stillborn pups.

All these findings were considered to be test item treatment-related.

MORTALITY (OFFSPRING)
There were dose-related increased percentages of found dead and cannibalized pups from 15 mg/kg/day.
Found dead and cannibalized pups were observed mostly on Days 1 and 2 p.p..
These findings were considered to be test item treatment-related and observed in a context of severe to excessive maternal toxicity.

CLINICAL SIGNS (OFFSPRING)
When compared with controls, there were a dose-related increased numbers of litters with pups with absence of milk in the stomach from 15 mg/kg/day.

BODY WEIGHT (OFFSPRING)
When compared with controls, on Days 1 and 5 p.p., there were lower mean body weights and mean body weight changes from 15 mg/kg/day.

GROSS PATHOLOGY (OFFSPRING)
In found dead pups (non autolyzed ones), there was absence of milk in the stomach. In surviving pups, there were no test item treatment-related findings at necropsy and in particular no findings at heart and cardiovascular examinations.

Effect levels (F1)

Dose descriptor:
NOEL
Remarks:
Toxic effect on progeny
Generation:
F1
Effect level:
< 15 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on lower survival index on Day 4 p.p. at this dose-level.

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

After 2 or 27 days of treatment at 60 or 30 mg/kg/day respectively, excessive toxicity resulted into a panel of clinical signs, premature euthanasia and/or deaths, both in males and/or females.

In term of body weight and food consumption, males were mostly affected at 60 mg/kg/day while toxicologically significant reductions were observed in females from 15 mg/kg/day (mainly during the lactation period).

In this context of marked to severe parental toxicity, there were no effects on mating/fertility data up to 30 mg/kg/day (lower fertility and gestation indexes at 60 mg/kg/day), no effects on pregnancy parameter up to 30 mg/kg/day (increased post-implantation at 60 mg/kg/day), and decreased pups viability from 15 mg/kg/day.

The decreases in viability indexes were the consequence of increased incidences of found dead and cannibalized pups on Days 1 and 2 p.p.. At necropsy, found dead pups (non autolyzed ones) had no milk in the stomach.In this study, the marked to severe toxicity may have dramatically modified the maternal behavior leading to decreased pup viability from 15 mg/kg/day.

 

Despite a context of severe to excessive maternal toxicity, there was no indication of any teratogenic potential (no test item treatment-related findings at necropsy of found dead or surviving pups).

Applicant's summary and conclusion

Conclusions:
Based on the experimental conditions of this study:
- the No Observed Adverse Effect Level (NOAEL) for parental (male and female) toxicity was considered to be lower than 15 mg/kg/day based on effect on body weight at this dose-level and, mortality and clinical signs from 30 mg/kg/day,
- the NOAEL for reproductive performance and embryo/fetal development was considered to be 30 mg/kg/day based on lower fertility and gestation indexes at 60 mg/kg/day,
- the No Observed Effect Level (NOEL) for toxic effects on progeny was considered to be lower than 15 mg/kg/day based on lower survival index on Day 4 p.p. at this dose-level.
Despite a context of severe to excessive maternal toxicity, there was no indication of any teratogenic potential.
Executive summary:

The objective of this study was to evaluate the potential toxic effects of the test item, LiTFSI PURE, following daily oral administration (gavage) to male and female rats from before mating, through mating and, for females, through gestation until Day 4 post-partum (p.p.), according to the OECD guideline n°421 and under GLP conditions. This study provided initial information on male and female reproductive performance, such as gonadal function, mating behavior, conception, development of the conceptus and parturition.

Three groups of ten male and ten female Sprague-Dawley rats received the test item,LiTFSI PURE (batch No. TFSILi-PU-1305001), daily, by oral administration (gavage), before mating, during mating and, for the females, throughout gestation until Day 5 p.p., at dose-levels of 15, 30 or 60 mg/kg/day for at least 4 weeks in males and up to 6 weeks in females.An additional group of ten males and ten females received the vehicle control, drinking water treated by reverse osmosis, under the same experimental conditions. The dosing volume was 5 mL/kg/day.

Animals were checked daily for clinical signs, mortality, and detailed clinical observations were conducted weekly. Body weights and food consumption were recorded weekly until mating and then at designated intervals throughout gestation and lactation.

The animals were paired for mating after 2 weeks of treatment and the dams were allowed to litter and rear their progeny until Day 5 p.p.. The total litter sizes and numbers of pups of each sex were recorded after birth. The pups were observed daily for clinical signs of toxicity and pup body weights were recorded on Days 1 and 5 p.p..

The males were sacrificed after completion of the mating period. Dams were sacrificed on Day 5 p.p.. Body weights and selected organs weights were recorded and a complete macroscopic post-mortem examination performed, with particular attention paid to the reproductive organs. A microscopic examination was also conducted on selected organs from the animals in the control groups and the high‑dose groups. Microscopic examination was conducted on all macroscopic lesions from all groups.

Pups, including those found dead before study termination, were also submitted for a macroscopic post‑mortem examination.

The LiTFSI PURE concentrations in the administered dosage forms were within acceptable range of variations (± 10% of the nominal concentrations).LiTFSI PURE was not detected in control samples.

 

Mortality: Fifteen animals died during the study, nine females and one male in the high-dose group and four females and one male in the mid-dose group. In males and females, all deaths recorded at 30 and 60 mg/kg/day were considered to be related to the test item treatment. In females, these deaths were considered to represent excessive maternal toxicity at 30 and 60 mg/kg/day

Clinical signs: there were no test item treatment-related findings at 15 mg/kg/day. There were signs of neurological disorders (i.e. tremors, convulsions, and tonic contraction) from 30 mg/kg/day, in males and at 60 mg/kg/day in females.

 

Body weight and body weight change:

In males, at 60 mg/kg/day, and when compared with controls, there was a minimal lower mean body weight on Day 8 (-4.6%, p < 0.01) resulting in a lower mean body weight gain over the first week of treatment (+24 gvs. +38 g, p < 0.05).

In females, there were dose-related decreases in mean body weight and mean body weight change from 15 mg/kg/day.

Food consumption:

In males, from 15 mg/kg/day, and when compared with controls, there were minimal reductions in mean food consumption during the first week of the treatment period (down to 32 g/animal/day at 60 mg/kg/day vs. 36 g/animal/day).

In females during the lactation period, from 15 mg/kg/day, and when compared with controls, there were lower mean food consumptions (down to 35 g/animal/day at 30 mg/kg/dayvs. 48 g/animal/day, p < 0.01).

 

Mating and fertility data: at 60 mg/kg/day, there were toxicologically significant lower fertility and gestation indexes.

 

Delivery data:

.            at 15, 30 and 60 mg/kg/day, and when compared with controls, there was a dose-related decrease in viability index on Day 4 p.p.(79.6%, 47.5% and 0.0%vs.95.7%, respectively),

.            at 60 mg/kg/day, and when compared with controls, there was a high incidence of post-implantation loss (30.1%vs. 11.9) and, only a few females delivered with all pups found dead during the period of Days 1-4 p.p..

 

Pup mortality: there were dose-related increased percentages of found dead and cannibalized pups from 15 mg/kg/day.

 

Pup clinical signs: when compared with controls, there was a dose-related increased number of litters with pups with absence of milk in the stomach from 15 mg/kg/day.

 

Pup body weight and body weight gain: on Days 1 and 5 p.p., and when compared with controls, there were lower mean body weights and mean body weight changes from 15 mg/kg/day.

 

Pups necropsy: at heart and cardiovascular examinations of the pups, there were no findings.

 

Pathology: there were no organs weight changes and macroscopic or microscopic findings considered to be test item treatment-related.

Based on the experimental conditions of this study:

.            the No Observed Adverse Effect Level (NOAEL) for parental (male and female) toxicity was considered to be lower than 15 mg/kg/day based on effect on body weight at this dose-level and, mortality and clinical signs from 30 mg/kg/day,

.            the NOAEL for reproductive performance and embryo/fetal development was considered to be 30 mg/kg/day based on lower fertility and gestation indexes at 60 mg/kg/day,

.            the No Observed Effect Level (NOEL) for toxic effects on progeny was considered to be lower than 15 mg/kg/day based on lower survival index on Day 4 p.p. at this dose-level.

 

Despite a context of severe to excessive maternal toxicity, there was no indication of any teratogenic potential.