Registration Dossier

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
08 Apr 2015 - 23 Nov 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: solidified melt
Details on test material:
- Name of test material (as cited in study report): Vinyloctadecylether
- Analytical purity: 83.5 %
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 20150128
- Expiration date of the lot/batch:
- Purity: 85.5 area-%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature.
- Stability under test conditions: The stability of the test substance under storage conditions
over the test period was guaranteed by the sponsor, and the sponsor holds this responsibility.
- Solubility and stability of the test substance in the solvent/vehicle: The oily test substance preparations
were prepared at the beginning of the administration period and thereafter at intervals, which took
into account the period of established stability.



Test animals

Species:
rat
Strain:
Wistar
Remarks:
The Crl:WI(Han) strain was selected since extensive historical control data is available from the test facility for Wistar rats. This specific strain has been proven to be sensitive to substances with a teratogenic potential.
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH
- Age at study initiation: 10-12 weeks
- Weight at study initiation: 142.5 - 191.3 g
- Fasting period before study:
- Housing: individual
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%.
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 hours light from 6.00 h to 18.00 h and 12 hours darkness from 18.00 h to 6.00 h

I

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The oily test substance preparations were prepared at the beginning of the administration period and thereafter at intervals,
which took into account the period of established stability. The preparations were kept at room temperature.
For the test substance preparations, the test substance was liquefied at a temperature of about 70°C in a drying chamber.
Thereafter, the specific amount of test substance was weighed, topped up with corn oil in a graduated flask and intensely
mixed with a magnetic stirrer until it is dissolved.
During administration, the preparations were kept homogeneous with a magnetic stirrer.


VEHICLE
- Amount of vehicle (if gavage): volume administered each day was 4 ml/kg body weight.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The results of the analysis of the oily test substance preparations confirmed the correctness of the prepared concentrations.
The analytical values of the samples corresponded to the expected values within the limits of the analytical method,
i.e. were always above 90% and below 110% of the nominal concentrations.
Details on mating procedure:
The animals were paired by the breeder (“time-mated”); the day of evidence of mating (= detection
of vaginal plug/sperm) was referred to as GD 0. The animals arrived on the same day (GD 0) at the
experimental laboratory. The following day was designated as “GD 1”.
The animals were acclimated to the laboratory conditions between start of the study (beginning
of the experimental phase) and first administration (GD 6).
Duration of treatment / exposure:
GD 6 to GD 19
Frequency of treatment:
once a day
Duration of test:
On GD 20, the females were sacrificed in a randomized order and examined macroscopically.
The fetuses were removed from the uterus and investigated
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day
Dose / conc.:
300 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- A cage-side examination was conducted at least once daily for any signs of morbidity, pertinent
behavioral changes and/or signs of overt toxicity. If such signs occurred, the animals were examined
several times daily (GD 0-20).
- Mortality: A check was made twice a day on working days or once a day on Saturdays, Sundays or on
public holidays (GD 0-20).

BODY WEIGHT: Yes
- Time schedule for examinations: All animals were weighed on GD 0, 1, 3, 6, 8, 10, 13, 15, 17, 19 and 20. The body weight
change of the animals was calculated based on the obtained results.
- Corrected (net) body weight gain: Furthermore, the corrected body weight gain was calculated after terminal sacrifice (terminal
body weight on GD 20 minus weight of the unopened uterus minus body weight on GD 6).

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes



POST-MORTEM EXAMINATIONS: Yes / No / No data
- Sacrifice on gestation day 20
- Organs examined: uterue, ovaries

Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes / No / No data
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
Fetal examinations:
- External examinations: Yes:
At necropsy each fetus was weighed, sexed, and external tissues and all orifices were examined
macroscopically. The sex was determined by observing the distance between the anus vand the base of the genitalia.
Furthermore, the viability of the fetuses and the condition of placentae, umbilical cords, fetal membranes, and fluids
were examined. The placentas were weighed and their individual weights were recorded.
Thereafter, the fetuses were sacrificed by a subcutaneous injection of pentobarbital.
After these examinations, approximately one half of the fetuses per dam were eviscerated, skinned and fixed in
ethanol; the other half were placed in Harrison’s fluid for fixation.
- Soft tissue examinations: Yes
The fetuses fixed in Harrison’s fluid were examined for any visceral findings according to the
method of BARROW and TAYLOR. After this examination these fetuses were discarded.
- Skeletal examinations: Yes
The skeletons of the fetuses fixed in ethanol were stained according to a modified method of KIMMEL and TRAMMELL.
Thereafter, the skeletons of these fetuses were examined under a stereomicroscope.
Statistics:
Simultaneous comparison of all dose groups with the control group using the DUNNETT-test (two-sided) for the hypothesis of equal means was used for the following parameters:
Food consumption, body weight, body weight change, corrected body weight gain (net maternal body weight change), carcass weight, weight of unopened uterus, number of corpora lutea, number of implantations, number of resorptions, number of live fetuses, proportions of preimplantation loss, proportions of postimplantation loss, proportions of resorptions, proportion of live fetuses in each litter, litter mean fetal body weight, litter mean placental weight

Pairwise comparison of each dose group with the control group using FISHER'S EXACT test (onesided) for the hypothesis of equal proportions was used for the following parameters:
Female mortality, females pregnant at terminal sacrifice, number of litters with fetal findings

Pairwise comparison of each dose group with the control group using the WILCOXON-test (one-sided) for the hypothesis of equal medians was used for the following parameters:
Proportions of fetuses with malformations, variations and/or unclassified observations in each litter.
Indices:
Conception rate, preimplantation loss, postimplantation loss

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Some females (up to 6 out of 25) of the high-dose group (1000 mg/kg bw/d) showed transient salivation
during the treatment period (GD 16-19). Salivation persisted in the respective animals only for some minutes
after daily gavage dosing (i.e. up to 15 minutes) and was initially observed on GD 16.
It is considered to be treatment-related, likely as a result of the bad taste of the test substance/vehicle
preparation or due to local irritation of the upper digestive tract. It is not considered to be a sign of systemic toxicity

No further clinical signs or changes of general behavior, which may be attributed to the test substance,
were detected in any female at dose levels of 100, 300 or 1000 mg/kg bw/d during the entire study period.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
The mean body weights (BW) and the average body weight gains (BWC) of the low-, midand
high-dose dams (100, 300, 1000 mg/kg bw/d) were in general comparable to the controls
throughout the entire study period. The statistically significantly increased BWC values during
GD 15-17 in test group 2, and for the treatment period (GD 6-19) in test groups 2 and 3 were
assessed as being incidental.
Corrected (net) body weight gain:
Mean carcass weight of test group 2 was statistically significantly increased in comparison to
the control group. However, the increase was small and not dose-related. Therefore, this was
assessed as incidental.
The corrected body weight gain (terminal body weight on GD 20 minus weight of the unopened
uterus minus body weight on GD 6) was statistically significantly higher than the control
in test groups 1-3 (100, 300 and 1000 mg/kg bw/d). As for the carcass weight, the increase
was rather small and had no effect whatsoever on the well-being of the animals. Thus
this apparent effect was not considered to be toxicologically relevant.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
The mean food consumption of the dams in test groups 1, 2 and 3 (100, 300 and 1000 mg/kg
bw/d) was generally comparable to the concurrent control throughout the entire study period.
The statistically significantly increased food consumption values in test group 3 during
GD 13-17 were assessed to be incidental.
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Uterus weight
The mean gravid uterus weights of the animals of test groups 1-3 (100, 300 and 1000 mg/kg
bw/d) were not influenced by the test substance. The differences between these groups and
the concurrent control groups revealed no dose-dependency and were assessed to be
without biological relevance.
Gross pathological findings:
no effects observed

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: no adverse effects observed

Results (fetuses)

Fetal body weight changes:
no effects observed
Changes in sex ratio:
no effects observed
External malformations:
no effects observed
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Skeletal malformations were recorded for one fetus each of test groups 0 and 1 (0 and 100 mg/kg bw/d).
The total incidence of skeletal malformations in treated animals did not differ significantly from the control group
and was comparable to the historical control data.
For all test groups, skeletal variations of different bone structures were observed, with or without effects on corresponding
cartilages. The observed skeletal variations were related to several parts of fetal skeleton and appeared without a relation
to dosing. The overall incidences of skeletal variations were covered by the historical control data.
Fetal skeletal unclassified cartilage observations:
Additionally, some isolated cartilage findings without impact on the respective bone structures, which were
designated as unclassified cartilage observations, occurred in all test groups. The observed unclassified cartilage
findings were related to the skull, the sternum and ribs.
The incidence of ‘bipartite processus xiphoideus’ was statistically significantly increased in test group 2.
As a consequence of this occasional increase, the incidence of total fetal skeletal unclassified cartilage
observations was statistically significantly increased in this test group. Since there is no dose-response
relationship and the finding can be found in the historical control data at a higher frequency, an association to
the treatment and a toxicological relevance is not assumed.
Furthermore, the incidence of ‘notched cartilage between basisphenoid and basioccipital’ was statistically significantly
increased in test group 3 (1000 mg/kg bw/d, mean of affected fetuses/litter: 2.6* %). The mean value was well within
the range of the historical control data (mean 2.4 %, range of 0.0 – 10.0 %) and, therefore, the finding was not
assessed as treatment-related and adverse.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
One fetus each in test groups 0 and 2 (0 and 300 mg/kg bw/d) had soft tissue malformations. In one case,
these malformations were associated with external malformations. The total incidence of soft tissue
malformations did not differ significantly from the concurrent control group. The findings were covered by the
historical control data and, therefore, not assessed as treatment-related.
Three soft tissue variations were detected, i.e. short innominate, dilated renal pelvis and dilated ureter.
These variations were neither significantly different from the concurrent control nor dose-dependently altered.
Therefore, they were not considered biologically relevant.
Details on embryotoxic / teratogenic effects:
Fetal external malformations
Three fetuses of one control litter (0 mg/kg bw/d) had external malformations. In two cases, these external malformations
were associated with either soft tissue or skeletal malformations. There were no external malformations in any of the treated
groups.
Fetal external unclassified observations:
One unclassified external observation, i.e. placentae fused, was recorded in two fetuses of the mid-dose group
Since the finding was without statistical significance and not related to dose, it was not considered to be test substance-related.

Assessment of all fetal external, soft tissue and skeletal observations
There were noted external, soft tissue and skeletal malformations in the test groups 0-2 (0, 100 and 300 mg/kg bw/d).
Three fetuses were multiple malformed. One female control fetus showed mandibular micrognathia and a cleft palate
(comprising a short mandible and a small palatine bone), while another female fetus had multiple external malformations
affecting the head (i.e. mandibular micrognathia, cleft palate, open eye) combined with a small lung. Furthermore,
one female mid-dose fetus had multiple soft tissue malformations, i.e. situs inversus (thoracic cavity),
fused or absent lung lobes, membraneous ventricular septum defect. No ontogenetic pattern is recognizable
for these individual malformations nor was there any cluster of any of these individual malformations seen in the other
offspring of these test groups. Most of them can be found in the historical control data of the rat strain.
An association of these findings to the treatment is not assumed. One malformation, i.e. misshapen tuberositas deltoidea,
of test group 1 was not related to the dose and can be found in the historical control data. An association of this finding
to the treatment is not assumed.
External variations did not occur in any of the fetuses in this study. Some soft tissue variations and a range of
skeletal variations were noted in all test groups including the controls. None of the incidences showed a relation to
dosing. The skeletal variations are equally distributed about the different test groups, if normal biological variation
is taken into account, and can be found in the historical control data at a comparable frequency.
No unclassified soft tissue observations were recorded for any of the fetuses in this study.
One unclassified external observation occurred in two mid-dose fetuses. This finding was not considered
to be substance-related. A spontaneous origin was assumed for the unclassified skeletal cartilage observations
which were observed in several fetuses of all test groups( 0, 100, 300 and 1000 mg/kg bw/d).
The distribution and type of these findings do not suggest any relation to treatment.
Finally, fetal examinations revealed that there is no effect of the compound on the respective morphological structures
up to the highest dose tested (1000 mg/kg bw/d).

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Under the conditions of this prenatal developmental toxicity study, the oral administration of Octadecylvinylether
to pregnant Wistar rats from implantation to one day prior to the expected day of parturition (GD 6-19) at doses
as high as 1000 mg/kg bw/d caused neither evidence of maternal nor developmental toxicity.
In conclusion, the no observed adverse effect level (NOAEL) for maternal and prenatal developmental toxicity is 1000 mg/kg bw/d.