Registration Dossier

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

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

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1999
Report date:
1999

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
methyl (2E)-3-methoxy-2-[2-({[6-(trifluoromethyl)pyridin-2-yl]oxy}methyl)phenyl]acrylate
EC Number:
601-478-9
Cas Number:
117428-22-5
Molecular formula:
C18H16F3NO4
IUPAC Name:
methyl (2E)-3-methoxy-2-[2-({[6-(trifluoromethyl)pyridin-2-yl]oxy}methyl)phenyl]acrylate
Test material form:
solid
Details on test material:
- Purity: 93.3-99.8% (see individual test record for specific details)
Specific details on test material used for the study:
93.3% purity

Test animals

Species:
rabbit
Strain:
New Zealand White
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Age at arrival: not reported
- Weight at study initiation: mean of 3679-3824 g
- Housing: The rabbits were housed individually in mobile rabbit units.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: not reported

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17±3°C
- Humidity (%): 30-70%
- Air changes (per hr): at lease 15 per hour
- Photoperiod (hrs dark / hrs light): 12 hour light and 12 hour dark

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples were extracted with water acetonitrile and portions of supernatant solutions were diluted with acetonitrile, as appropriate, to give sample solution concentrations within the range of the calibration standards. These were analyzed by High Performance Liquid Chromatography (HPLC).

Nominally 100 mg of the test substance was accurately weighed into a volumetric flask and diluted with 50 mL water, swirling gently to dissolve. The solution was made to volume with acetonitrile and was sonicated for 5 minutes (nominally 1.0 mg/mL). Further appropriate dilutions were made with acetonitrile in volumetric flasks to give a range of solutions, nominally within 2 μg/mL to 10 μg/mL. The purity of the test substance was not taken into account in the preparation of the standard solutions.

Appropriate portions of samples, 1 g ± 0.5 g, were transferred to volumetric flasks, diluted to volume with 50 mL of water, made to volume with acetonitrile and sonicated for 5 minutes. The samples were further diluted, as required, with acetonitrile to a known nominal concentration within the range of calibration standards.

High Performance Liquid Chromatography Conditions are as follows: Pump: 600 Series (Waters); Mobile phase: Acetonitrile (65% v/v), Water (35% v/v), and Phosphoric acid (0.1% v/v); Flow rate: 0.8 mL/min; Detector: 486 Series (Waters); Detector wavelength: 245 nm; Column: 15 cm x 3.9 mm id Symmetry Cl8 (Waters); Column temperature: Ambient; Sample introduction: 717 plus (Waters); Injection volume: 20 μL; and Data handling: Waters 860 (Waters).

The analysis system was calibrated using a range of standards to determine the linearity of response. An appropriate standard of known concentration was interspersed at intervals throughout the analysis.

The limit of detection was calculated to be approximately 0.1 μg/mL test substance in the analyzed solution, corresponding to a formulation concentration of 0.1 mg/mL.
Details on mating procedure:
Virgin female New Zealand White rabbits were paired (at the suppliers) with males of the same strain. The day when mating was observed was designated day 1 of gestation. The animals were delivered to the test facility on either day 2 or day 3 of gestation (within a period of 2 weeks).
Duration of treatment / exposure:
Days 8-20 of gestation
Frequency of treatment:
Daily on days 8-20 (inclusive) of gestation.
Duration of test:
30 days
Doses / concentrationsopen allclose all
Dose / conc.:
8 mg/kg bw/day (actual dose received)
Dose / conc.:
25 mg/kg bw/day (actual dose received)
Dose / conc.:
100 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
20
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels selected for this study were based on the results of dose range finding studies in the pregnant rabbit conducted at the test facility.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Within 1 hour of being dosed and towards the end of each working day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily and where appropriate, at the same time that the body weights were recorded

BODY WEIGHT: Yes
- Time schedule for examinations: Day 4, immediately prior to dosing on days 8-20 (inclusive) and on days 23, 26, and 30 of gestation

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Time schedule for examinations: Measured over 3 or 4 day intervals and calculated to give a daily value (g food/rabbit/day)

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 30
- Organs examined: External observation and an examination of the thoracic and abdominal viscera. The pregnancy status of each animal was determined.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes / No / No data
Fetal examinations:
- External examinations: Yes
- Soft tissue examinations: Yes
- Skeletal examinations: Yes
- Head examinations: Yes
Statistics:
See the "Any other information on materials and methods" section for information on the statistical methods used in this study.
Indices:
Percentage pre-implantation loss and percentage post-implantation loss were calculated as follows:

% pre-implantation loss number of corpora lutea - number of implantations x 100/number of corpora lutea

% post-implantation loss = number of implantations - number of live fetuses x 100/number of implantations

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
There was a slightly increased incidence of animals in the 25 and 100 mg/kg/day groups producing only few faeces and/or with signs of diarrhoea, in comparison with the control group.

There were no other clinical findings which were considered to be treatment-related.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
There was a statistically significant loss of bodyweight between days 8 and 11 following the onset of treatment with 100 mg/kg/day of the test substance. Bodyweight gain was observed thereafter.

Bodyweights (adjusted for day 8 bodyweight) of the animals in the 25 mg/kg/day were essentially comparable with the control group throughout the study. A small loss of bodyweight was observed on day 9 only. The bodyweights (adjusted for day 8 bodyweight) of the animals in the 8 mg/kg/day group were higher than in the control group throughout the dosing and post-dosing periods.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Following the onset of treatment on day 8 with 100 mg/kg/day of the test substance, a statistically significant reduction in food consumption was observed for days 8-11 and days 14-17. Food consumption was lower than that of the control animals throughout the dosing period although food consumption was slightly greater in the post-dosing period. Food consumption of the animals in the 25 mg/kg/day group was slightly lower than in the control group throughout the dosing period although a statistically significant difference was attained for days 8-11 only. Parity with controls was attained in the post-dosing period. The food consumption of the animals in the 8 mg/kg/day group was comparable with the animals in the control group throughout the dosing and post-dosing periods.
Food efficiency:
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
effects observed, non-treatment-related
Description (incidence and severity):
The number of corpora lutea in the 100 mg/kg/day group was lower than in the other groups. This difference was established prior to treatment with the test substance as was the increase in pre-implantation loss observed in this group.
Total litter losses by resorption:
effects observed, non-treatment-related
Description (incidence and severity):
1 litter was totally resorbed at termination in the low-dose group (1/20). There were no other resorptions.
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
The number of pregnant animals with live fetuses in utero on day 30 of gestation was 17, 16, 18 and 16 in the 0, 8, 25 and 100 mg/kg/day groups, respectively.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
25 mg/kg bw/day
Basis for effect level:
body weight and weight gain
clinical signs
food consumption and compound intake

Maternal abnormalities

Key result
Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
Mean foetal weights were 43.1 g, 42.3 g, 40.7 g, and 44.3 g in the control, 8, 25, and 100 mg/kg/day groups, respectively.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not examined
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
The mean number of live foetuses were 8.35, 9.00, 9.56, and 6.94 in the control, 8, 25, and 100 mg/kg/day groups, respectively.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
The percentages of male foetuses were not different (49.5, 54.2, 51.9, and 51.6) in the control, 8, 25, and 100 mg/kg/day groups, respectively.
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The percentage of foetuses with minor external/visceral defects in the each of the test substance groups was either comparable with or lower than in the control group and provided no evidence for any treatment-related effect. Consideration of the specific defects provided no evidence for any effect of the test substance on external or visceral development of the foetuses.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The percentage of foetuses with minor skeletal defects in the test substance groups showed no relationship with dose and there were no statistically significant differences from control. Consideration of the specific defects revealed only one finding, incomplete ossification of the odontoid, the incidence of which was statistically significantly increased in the 100 mg/kg/day group in comparison with the control group. This increase is considered to be incidental to treatment in the absence of any other indication of reduced ossification of the foetal skeleton.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The percentage of foetuses with minor external/visceral defects in the each of the test substance groups was either comparable with or lower than in the control group and provided no evidence for any treatment-related effect. Consideration of the specific defects provided no evidence for any effect of the test substance on external or visceral development of the foetuses.
Other effects:
effects observed, treatment-related
Description (incidence and severity):
The percentage of foetuses with skeletal variants was statistically significantly higher in the 100 mg/kg/day group in comparison with the control group. For the specific skeletal variants, the incidence of fetuses with 13 thoracolumbar ribs of long length was increased in the 25 and 100 mg/kg/day groups and, statistically significant in comparison with the control group. The incidence of this finding in both groups was within the range seen previously in control rabbits in the testing facility. At 100 mg/kg, the incidence of 13th throacolumbar ribs of long length, although within the range of historical control values, coincided with a slightly increased incidence of 27th pre-pelvic vertebrae and for this reason, an association with treatment at the maternally toxic dose of 100 mg/kg cannot entriely be dismissed . At 25 mg/kg, the statistically significant increase in incidence of 13th thoracolumbar ribs of long length was within historical control range, ws not associated with any other skeletal findings, and is considered not to reflect any effect of treatment. In summary, the increased incidence of foetuses with 27th pre-pelvic vertebrae and 13th thoracolumbar ribs of long length at 100 mg/kg represent small changes in the proportions of common variants, and as such are considered not to be of toxicological significance.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: highest dose tested

Overall developmental toxicity

Key result
Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
100 mg/kg/day of the test substance produced clear evidence of maternal toxicity.
25 mg/kg/day is the dose level at which no maternal toxicologically significant findings were observed.
100 mg//kg/day is the dose level at which no toxicologically significant developmental effects were observed.
Executive summary:

Groups of 20 pregnant rabbits were administered by gavage, 0, 8, 25, or 100 mg/kg/day of the test substance dissolved in 1% carboxymethyl cellulose on days 8-20 of gestation.

 

Administration of 100 mg/kg/day of the test substance was associated with maternal toxicity manifest as a loss of bodyweight and reduced food consumption. Signs of diarrhoea and reduced faecal output were also noted. There were no maternal toxicologically significant findings attributable to 8 or 25 mg/kg/day of the test substance. There was no adverse effect of the test substance on the number, growth or survival of the foetuses in utero and no evidence of teratogenicity. Evaluation of the minor external/visceral foetal defects and variants provided no evidence for an adverse effect of the test substance on development. Evaluation of the foetal skeletal variants revealed only two findings which may be related to treatment, namely an increased incidence of foetuses with 27 pre-pelvic vertebrae and an increased incidence of foetuses with 13 thoracolumbar ribs of long length. However, these findings reflect small changes in the proportions of common variants and as such are considered not to be of toxicological significance.