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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
3 January 1983 - 4 February 1983
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The study is performed according to internationally accepted guidelines (FDA & CSM) and similar to the current OECD 414. The study is performed under GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1983
Report Date:
1983

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
other: Food and Drug Administration (1966). Guidelines for Reproduction Studies for Safety Evaluation of Drugs for Human Use.
Deviations:
no
Qualifier:
according to
Guideline:
other: Committee on Safety of Medicines. Guidelines on Reproduction Studies for Guidance of Applicants for Product Licences and Clinical Trial Certificates, June 1974.
Deviations:
no
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
The number of pregnant animals in the high dose groups is too low (13 of 20) and slightly over 10 % of the animals in the high dose group died during the study (3 of 20).
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 2-Ethylhexyl-p-methoxy cinnamate
- Physical state: liquid
- Stability under test conditions: about 1 year (pure); at least 14 days (dissolved)
- Storage condition of test material: refrigerator

Test animals

Species:
rabbit
Strain:
other: Swiss hare
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Institute for Biological and Medical Research, CH-4414 Fiillinsdorf, Switzerland
- Age at study initiation: no data
- Weight at study initiation: 2510 - 3670 g
- Housing: stainless steel cages
- Diet (e.g. ad libitum): Nafag 814 cubes ad libitum, analysed for contaminants
- Water (e.g. ad libitum):ad libitum (periodically analysed)
- Acclimation period: a minimum of 4 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): about 18 °C
- Humidity (%): about 50 %
- Air changes (per hr): fully airconditioned
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: SSV: 0.5 % Carboxymethylcellulose, 0.5 % Benzyl-EtOH, 0.4 % TWEEN 80, 0.9 % NaCl
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test compound was formulated freshly each week in the vehicle. High shear mixing was used during formulation and the suspension was kept homogeneous during dosing by the use of a magneto stirrer.

VEHICLE
- Justification for use and choice of vehicle: solubility
- Amount of vehicle: 2 mL/kg
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
Not relevant
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1/1
- Length of cohabitation: 4 - 6 hours
- the mounted females were mated again with another male and were then randomly allocated to the dosage groups.
- Proof of pregnancy: Day of copulation referred to as day 1 of pregnancy
Duration of treatment / exposure:
Day 7 - Day 20 of gestation.
Frequency of treatment:
Once daily.
Duration of test:
Day 1 - Day 30 (necropsy) of gestation.
No. of animals per sex per dose:
20 mated females
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
In a dose range finding study for maternal toxicity and embryotoxicity of the test substance, doses of 200 and 500 mg/kg were administered to pregnant rabbits. No adverse effects could be observed except a slight impairment of body weight development of dams treated with 500 mg/kg. In respect to these findings we chose doses of 80, 200 and 500 mg/kg for the main study.
- Rationale for animal assignment: After successful mating the females were distributed to the dosage groups by the use of a computer generated randomization table.

Examinations

Maternal examinations:
OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations: all changes in behaviour, general condition, signs of pharmacological action, etc., rabbits which died during the experiment were autopsied.

BODY WEIGHT: Yes
- Time schedule for examinations: on day of gestation 1, 7, 20 and 30.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 30
- Organs examined: Only uterus examined
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of resorptions: Yes
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all per litter
- Skeletal examinations: Yes: all per litter
- Head examinations: Yes: all per litter
All the young were tested for their 24 h viability in an incubator at 34 °C.
Statistics:
1. Descriptive Statistics
For each group, median and interquartilerange are indicated.
2. Significance tests
For the statistical examination of each quantity, the following testalgorithm is applied:
a) Trend test
b) Simultaneous test without ordered alternative
c) Two sample tests for each dosage group against control
3. Experimental unit
The dam is considered the independent experimental unit within all significant tests.
Indices:
4. Relative numbers
The numbers of resorptions and all numbers of pubs reported are transformed for the test calculations. For each dam the relative number (relative number = absolute number/number of implantations) is determined.
Therefore, significant test results refer to the relative number, which can be interpreted as a rate, the number of implantations being 100 %.
Historical control data:
Yes, present at facility

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects: no effects

Details on maternal toxic effects:
No obvious signs of toxicity in treated females could be observed at any dosages except a slight Impairment of the body weight gain and an slightly increased frequency of constipated and anorectic dams in the 500 mg/kg group. All maternal deaths which occured during the study resulted from either broken vertebral column, enteritis or from application accidents. Therefore, the cause of deaths was not considered to be drug related.

Effect levels (maternal animals)

open allclose all
Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (actual dose received)
Basis for effect level:
other: maternal toxicity
Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (actual dose received)
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects: no effects

Details on embryotoxic / teratogenic effects:
The parameters which could not have been influenced by the treatment, i.e. % of pregnant females, average number of corpora lutea and average number of iinplantations sites were within normal limits in all groups. The number of resorptions in % of implantations is lower in all treatment groups (12.5, 32.5 and 18.8 %) than in concurrent control (34.1 %). The control group of the present study represent an extremly high resorption rate when compared with the mean value of the historical control data (17.8%). This is mainly due to 1 dam (No. 127) exhibiting 10 complete resorptions.
The relatively high resorption rate which resulted from the group treated with 200 mg/kg is due to two dams (No 326 and 336) which both resorbed their implantations completely and therefore contributed 19 out of a total of 40 resorptions to this group (200 mg/kg). However, the distribution of resorptions does not follow a dose effect relation and it is therefore considered that there is no drug related effect on intrauterine death.
Two dams (No 433 and 439) of the highest dosage group (500 mg/kg) aborted their fetuses and died prior to term or were moribund at the autopsy day. The abortions are considered to be due to the poor health status of the mother animals and not to an abortive property of the test compound.
Both animals are excluded from statistical evaluation.

In the highest dosage group (500 mg/kg) the median individual body weight of fetuses was decreased significantly, but was comparable between the other dosage groups and the control. Whether this effect is directly due to a drug action or indirectly to the poor bodyweight gain of dams can not be definitely established from the available data.

Examination of fetuses at necropsy or following skeletal processing revealed no drugrelated deviations. All macroscopically observed abnormalities such as spina bifida, missing or rudimentary tail, torsion in limbs or ectopy of intestines are isolated findings, distributed randomly amongst all groups and are known to arise spontaneously in this rabbit strain. The examination of the head using a serial sectioning method revealed no drugrelated abnormalities at any dose groups.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (actual dose received)
Basis for effect level:
changes in postnatal survival
skeletal malformations
visceral malformations

Fetal abnormalities

Key result
Abnormalities:
not specified

Overall developmental toxicity

Key result
Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
It may be concluded that under the conditions of the present experiment 2-Ethylhexyl 4-methoxycinnamate is neither embryotoxic nor teratogenic in rabbits at oral doses up to 500 mg/kg bw/day. A NOAEL of 500 mg/kg bw/day was therefore established for both maternal and developmental toxicity, indicating that the substance does not need to be classified as reprotoxic based on the criteria outlined in Annex I of 1272/2008/EC.
Executive summary:

2-Ethylhexyl 4-methoxycinnamate, a UV-B filter for topical use, was tested for embryotoxic and teratogenic action in rabbits according to the guidelines of the American FDA and the English CSM. Doses of 80, 200 and 500 mg/kg bodyweight were administered by oral gavage from Day 7 to Day 20 inclusive of gestation. A control group received the vehicle only (ssv, 2 mL/kg). All females were sacrificed on Day 30 of gestation. Fetuses were removed by ovariohysterectomy, tested for viability (24 h) and examined for skeletal and visceral abnormalities.

A slight decrease of maternal bodyweight gain during the treatment period and a slightly increased frequency of constipated and anorexic animals was observed in the 500 mg/kg group exclusively. No other drug related clinical sings of toxicity could be observed in treated female rabbits. The reproductive parameters were within the normal range of the rabbit strain, however 30 day old fetuses of the highest dosage group (500mg/kg) were slightly lighter when compared with the concurrent control offspring as well as with historical controls. None of the fetuses showed any drug related skeletal or visceral abnormalities at any dosage group. All adverse findings are isolated, distributed randomly amongst groups and are known to arise spontaneously in the rabbit strain.

The 24h survival rate of fetuses was not impaired by the treatment of mother animals during gestation.

It may be concluded that under the conditions of the present study 2-Ethylhexyl 4-methoxycinnamate is neither embryotoxic nor teratogenic in rabbits at oral doses up to 500 mg/kg . Whether the lighter fetuses observed in the highest dosage group (500 mg/kg) resulted from a direct intrauterine drug action during gestation or are secondarily correlated to the reduced bodyweight gain of mother animals can not be definitely established from the available data. A NOAEL of 500 mg/kg bw/day was therefore established for both maternal and developmental toxicity, indicating that the substance does not need to be classified as reprotoxic based on the criteria outlined in Annex I of 1272/2008/EC.