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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report date:
2013

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
1996
Deviations:
yes
Principles of method if other than guideline:
Deviations: The liver weights of male no. 15 (recovery group 1) and female no. 100 were not determined
and a few tissues were not available for histopathology. Sufficient data was available for evaluation.
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2,3,3,4,4-pentafluoro-2,5-bis(1,1,1,2,3,3,3-heptafluoropropan-2-yl)-5-methoxyoxolane
EC Number:
812-244-2
Cas Number:
957209-18-6
Molecular formula:
C11H3F19O2
IUPAC Name:
2,3,3,4,4-pentafluoro-2,5-bis(1,1,1,2,3,3,3-heptafluoropropan-2-yl)-5-methoxyoxolane
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 3M Company, Batch: L-21343
- Expiration date of the lot/batch: 27 August, 2014
- Purity test date: 27 August, 2012
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature in the dark.
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The test article was dosed undiluted.

Test animals

Species:
rat
Strain:
Wistar
Details on species / strain selection:
Wistar Han sources from Charles River Deutschland, Sulzfeld, Germany.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Females (if applicable) nulliparous and non-pregnant: Yes
- Age at study initiation: Approximately 11 weeks
- Weight at study initiation: Male mean: 323.5 g, Female mean: 208.5 g
- Fasting period before study: not specified.
- Housing: Pre-mating: animals were housed in groups of 5/sex/cage in Macrolon plastic cages (MIV typ
e, heigh 18 cm). This was also applicable for Recovery animals throughout the complete study eriod.
Mating: Main females were caged together with Main males on a one-to-one basis in Macrolon plast
ic cages (MIII type, height 18 cm). Post-mating: Main males were housed in their home cage (Macrol
on plastic cages, MIV type, height 18 cm) with a maximum of 5 animals per cage. Main females were
individually housed in Macrolon plastic cages (MIII ype, height 18 cm). Lactation: Pups were kept with
the dam until termination in Macrolon plastic cages (MIII type, height 18 cm). During locomotor activity
monitoring of the dams, the pups were kept warm in their home cage using bottles filled with warm water.
- Diet (e.g. ad libitum): Pelleted rodent diet (SM R/M-Z from SSNIFF), ad libitum.
- Water (e.g. ad libitum): Tap water, ad libitum.
- Acclimation period: At least 5 days prior to the start of treatment.
ENVIRONMENTAL CONDITIONS
- Temperature (°C):18-24
- Humidity (%): 40-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12
IN-LIFE DATES: From: 17 October, 2012 To: 28 December, 2012

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test article was dosed unchanged.
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: Maximum 14 days.
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged: Individually
Analytical verification of doses or concentrations:
no
Remarks:
Test article was dosed unchanged so the initial purity verification was sufficient.
Duration of treatment / exposure:
All rats were exposed once daily for 7 days per week; all males were exposed for a total of 29 days, including 2 weeks of exposure prior to mating and during mating for rats in the 4 main groups, and all females were exposed for a total of 41-47 days prior to mating, during mating, during post mating, and lactation periods. Males in the recovery groups were allowed to recover for 14 days following the last exposure.
Frequency of treatment:
All rats were exposed once daily for 7 days per week; all males were exposed for a total of 29 days, including 2 weeks of exposure prior to mating and during mating for rats in the 4 main groups, and all females were exposed for a total of 41-47 days prior to mating, during mating, during post mating, and lactation periods. Males in the recovery groups were allowed to recover for 14 days following the last exposure.
Details on study schedule:
- F1 parental animals not mated as this was a screening study.
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw (total dose)
Remarks:
Control animals were dosed with water.
Dose / conc.:
100 mg/kg bw (total dose)
Dose / conc.:
300 mg/kg bw (total dose)
Dose / conc.:
1 000 mg/kg bw (total dose)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent no treatment
Details on study design:
- Dose selection rationale: Based on a range-finding study.
- Rationale for animal assignment (if not random): Random
- Rationale for selecting satellite groups: Random
- Post-exposure recovery period in satellite groups: 14 Days
Positive control:
None

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily
BODY WEIGHT: Yes
- Time schedule for examinations: First day of exposure and weekly thereafter.
FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averag
es from the consumption and body weight gain data: Yes, weekly.
OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Week 4
- Dose groups that were examined: 5 Main animals/sex/group and all Recovery animals.
HAEMATOLOGY: Yes
- Time schedule for collection of blood: On the day of necropsy
- Anaesthetic used for blood collection: Yes (isofluorane)
- Animals fasted: Yes
- How many animals: 5 Main animals/sex/group and all Recovery animals.
- Parameters checked: white blood cell count, differential leucocyte count, red blood cell count, red blood
cell distribution width, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin
concentration, platelets, prothrombin time, activated partial thromboplastin time
CLINICAL CHEMISTRY: Yes
Time schedule for collection of blood: On the day of necropsy
- Anaesthetic used for blood collection: Yes (isofluorane)
- Animals fasted: Yes
- How many animals: 5 Main animals/sex/group and all Recovery animals.
- Parameters checked: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, total
protein, albumin, total bilirubin, urea, creatinine, glucose, cholesterol, sodium, potassium, chloride, calciu
m, inorganic phosphate, bile acids
URINALYSIS: No
NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Week 4
- Dose groups that were examined: The selected Main males and all Recovery males were tested durin
g Week 4 of treatment and the selected Main females were tested towards the end of the scheduled la
ctation period (all before blood sampling) (from lactation Day 4 onwards) and all Recovery females were
tesed on the first day a Main female was tested. Since no treatment-related findings were noted in any of
the above tests, these tests were not conducted for Recovery animals at the end of the recovery phase.
- Battery of functions tested: sensory activity / grip strength / motor activity / other:
IMMUNOLOGY: No
Sacrifice and pathology
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Histopathology included adrenal glands, brain, colon, duodenum, eyes, female mammary gland area,
femur including joint, heart, ileum, jejunum, kidneys, liver, lung, lymph nodes, peyer's patches, pituitary
gland, rectum, sciatic nerve, skeletal muscle, spinal cord, spleen, sternum with bone marrow, stomach, th
ymus, thyroid, trachea, urinary bladder, cervix, clitoral gland, coagulation gland, epididymides, ovaries, p
reputial gland, prostate gland, seminal vesicles, testes, uterus, and vagina.
Sperm parameters (parental animals):
Parameters examined in 5 Main males in Groups 1 and 4 male parental generations:
testis weight, epididymis weight, staging of spermatogenesis.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: No

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring: number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain.

GROSS EXAMINATION OF DEAD PUPS:
yes, for external abnormalities; possible cause of death was determined for pups born or found dead.

ASSESSMENT OF DEVELOPMENTAL NEUROTOXICITY: No

ASSESSMENT OF DEVELOPMENTAL IMMUNOTOXICITY: No
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: Main males: Following completion of the mating period (a minimum of 28 days of dose administration).
Recovery males: After the recovery period of at least 14 days, which was at least 14 days after the scheduled kill of Main males.
- Maternal animals: Main Females which delivered: Lactation Days 5-7.
Main Females which failed to deliver (nos. 70 and 75): Post-coitum Day 25 (females with evidence of mating).
Recovery Females: After the recovery period of at least 14 days, which was at least 14 days after the first scheduled kill of Main females.

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS
Histopathology included adrenal glands, brain, colon, duodenum, eyes, female mammary gland area, femur including joint, heart, ileum, jejunum, kidneys, liver, lung, lymph nodes, peyer's patches, pituitary gland, rectum, sciatic nerve, skeletal muscle, spinal cord, spleen, sternum with bone marrow, stomach, th ymus, thyroid, trachea, urinary bladder, cervix, clitoral gland, coagulation gland, epididymides, ovaries, preputial gland, prostate gland, seminal vesicles, testes, uterus, and vagina.
Organ weights were noted for: Adrenal glands, brain, epididymides, heart, kidneys, liver, ovaries, spleen, testes, thymus, uterus, prostate, seminal vesicles including coagulating glands, thyroid including parathyroid.
Postmortem examinations (offspring):
SACRIFICE
Pups surviving to planned termination were killed by decapitation on Day 5-7 of lactation.

GROSS NECROPSY
All pups were sexed and descriptions of external abnormalities were recorded. Abnormalities were collected and fixed in 10% buffered formalin. The stomach of pups not surviving to the scheduled necropsy date were examined for the presence of milk, if possible. If possible, defects or cause of death were evaluated.
.
Reproductive indices:
The following were calculated: Mating index, fertility index, conception index, duration of gestation.
Offspring viability indices:
The following were calculated: Gestation index, percentage of live males at first litter check, percentage of live females at first litter chekc, percentage of postnatal loss, viability index.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
no effects observed
Description (incidence and severity):
No clinical signs of toxicity related to the test article were noted in any animal.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
No treatment related mortality occurred during the study period.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No toxicologically relevant changes in body weight and body weight gain were noted.
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
no effects observed
Description (incidence and severity):
Food consumption before or after allowance for body weight was similar between treated and control animals.
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Pupillary reflex was normal in all selected animals tested in the FOB.
Haematological findings:
no effects observed
Description (incidence and severity):
Hematological parameters of treated rats were considered not to have been affected by treatment.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
Clinical biochemistry parameters of treated rats were considered not to have been affected by treatment.
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 selected animals.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Hearing ability, pupillary reflex, static righting reflex and grip strength were normal in all selected animals.
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
There were no treatment-related microscopic findings.
Histopathological findings: neoplastic:
no effects observed
Description (incidence and severity):
There were no treatment-related microscopic findings.

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
no effects observed
Description (incidence and severity):
No sprem parameters were altered by treatment with the test-article.
Reproductive performance:
no effects observed
Description (incidence and severity):
No toxicologically relevant effects on reproductive parameters were noted.

Details on results (P0)

No toxicologically relevant effects on reproductive parameters were noted. No toxicologically relevant effects on gestation index and duration, parturition, maternal care and postnatal pup development were observed.

Effect levels (P0)

Key result
Dose descriptor:
NOAEL
Effect level:
ca. 1 000 mg/kg bw (total dose)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food efficiency
ophthalmological examination
haematology
clinical biochemistry
organ weights and organ / body weight ratios
gross pathology
neuropathology
histopathology: non-neoplastic
histopathology: neoplastic
reproductive function (sperm measures)
reproductive performance

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
No treatment-related findings were observed.
Dermal irritation (if dermal study):
not examined
Mortality / viability:
mortality observed, non-treatment-related
Description (incidence and severity):
No treatment-related findings were observed.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No treatment-related findings were observed.
Food consumption and compound intake (if feeding study):
not examined
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
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
No treatment-related findings were observed.
Histopathological findings:
not examined

Developmental neurotoxicity (F1)

Behaviour (functional findings):
not examined

Developmental immunotoxicity (F1)

Developmental immunotoxicity:
not examined

Effect levels (F1)

Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
ca. 1 000 mg/kg bw (total dose)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
clinical signs
mortality
body weight and weight gain
gross pathology

Overall reproductive toxicity

Key result
Reproductive effects observed:
no

Applicant's summary and conclusion

Conclusions:
Based on the results of the study, the test article has a No Observed Adverse Effect Level of 1000 mg/kg/day.
Executive summary:

The repeated dose oral toxicity potential and the reproductive / developmental toxicity potential of the test article was evaluated in male and female Wistar Han rats via daily oral gavage for 28 days. The study was performed in compliance with OECD GLP (1997). The study design was based on OECD 422 (1996) and US EPA OPPTS 870.3650 (2000), and essentially conforms to OECD 421 (1995), US EPA OPPTS 870.3550 (2000), EC 440/2008 Part B B7. L142 (2008), OECD 407 (2008), and US EPA OPPTS 870.3050 (2000). The test article was dosed undiluted and given at a dose volume of 0.055-0.553 mL/kg to four groups of rats by oral gavage. Rats (10/sex/group) were dosed with 0, 100, 300, and 1000 mg/kg-body weight test article per day. Additional rats (5 males/group) were exposed at 0 mg/kg-day or 1000 mg/kg-day for a recovery study. All rats were dosed once daily for 7 days per week; all males were exposed for a total of 29 days, including 2 weeks of exposure prior to mating and during mating for rats in the 4 main groups, and all females were exposed for a total of 41-47 days prior to mating, during mating, during post mating, and lactation periods. Males in the recovery groups were allowed to recover for 14 days following the last dose. Observational parameters evaluated in all main and recovery groups rats included; mortality (twice daily), clinical signs (daily), body weights (first day of exposure and weekly thereafter), food consumption (weekly), and general reproduction data (during mating and post-mating). Pups born to mating mothers were evaluated for mortality (daily), clinical signs (at least once daily), body weights (Days 1 and 4 of lactation), and sex determination (Days 1 and 4 of lactation). Hematology, clinical biochemistry, necropsy, and histopathology and organ weight collection were performed on selected tissues of rats in the main exposure groups (5/sex/group). Histopathology included adrenal glands, brain, colon, duodenum, eyes, female mammary gland area, femur including joint, heart, ileum, jejunum, kidneys, liver, lung, lymph nodes, peyer's patches, pituitary gland, rectum, sciatic nerve, skeletal muscle, spinal cord, spleen, sternum with bone marrow, stomach, thymus, thyroid, trachea, urinary bladder, cervix, clitoral gland, coagulation gland, epididymides, ovaries, preputial gland, prostate gland, seminal vesicles, testes, uterus, and vagina. No treatment related mortality occurred during the study period. No clinical signs of toxicity related to the test article were noted in any animal. Hearing ability, pupillary reflex, static righting reflex and grip strength were normal in all selected animals tested in a functional observational battery. No toxicologically relevant changes in body weight and body weight gain were noted. Food consumption before or after allowance for body weight was similar between treated and control animals. Hematological and clinical biochemistry parameters of treated rats were considered not to have been affected by treatment. Macroscopic observations at necropsy did not reveal any alterations that were considered to have arisen as a result of treatment. No toxicologically relevant changes were noted in organ weights and organ to body weight ratios. There were no treatment-related microscopic findings. No toxicologically relevant effects on reproductive parameters were noted. No toxicologically relevant effects on gestation index and duration, parturition, maternal care and postnatal pup development were observed. Based on the results of the study, the test article has a No Observed Adverse Effect Level of 1000 mg/kg/day.