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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
5 May 1994 - 2 February 1996.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
A category approach is proposed for hexyl salicylate using data from other salicylate substances. The members of this category are demonstrated to be metabolised rapidly by esterases to salicylic acid; systemic exposure will therefore be to the metabolites to a much greater extent. The systemic toxicity of the salicylates is comparable and is attributable to the salicylic acid generated by metabolism. The other product of metabolism is the alcohol liberated from the sidechain following esterase metabolism. In the case of hexyl salicylate, this alcohol will be 1-hexanol which is of low toxicity and is rapidly metabolised and excreted and/or incorporated into normal metabolism. Hexanol is of lower toxicity than other alcohols generated from the metabolism of other category members (e.g. methanol); the source data therefore represents a worst case approach.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1996
Report Date:
1996

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: liquid
Details on test material:
- Name of test material (as cited in study report): Cyclohexylsalicylate
- Physical state: colourless liquid
- Lot/batch No.: 111 833 34/2
- Expiration date of the lot/batch: December 1995
- Stability under test conditions: Stable in arachidis oil, DAB 10
- Storage condition of test material: At room temperature

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, Wiga GmbH, D-97633 Sulzfeld.
- Age at study initiation: 8 weeks
- Weight at study initiation: mean approximately 214g
- Fasting period before study: Not documented
- Housing: Housed individually in Makrolon Type M3 cage with standard softwood bedding.
- Diet (e.g. ad libitum): Pelleted Altromin Maintenance Diet 1324 ad libitum.
- Water (e.g. ad libitum): Community tap water from Dusseldorf ad libitum.
- Acclimation period: 5 days under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 25°C
- Humidity (%): 44 - 75%
- Air changes (per hr): 10 - 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light / 12 hours dark at lux units of 25 - 550.

IN-LIFE DATES: From: To: Not documented

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test substance was prepared daily before administration. The formulation was analysed and the concentrations of the test substance in arachis oil, DAB 10, based on the results of the determination of the HPLC method were:

0.8% = 0.4g/50ml (40 mg/kg) = 0.415 g/50ml ± 0.007
2.4% = 1.2g/50ml (120 mg/kg) = 1.260 g/50ml ± 0.014
7.2% = 3.6g/50ml (360 mg/kg) = 3.720 g/50ml ± 0.014

DIET PREPARATION
- Rate of preparation of diet (frequency): Daily
- Mixing appropriate amounts with (Type of food): Not relevant as gavage method of administration used
- Storage temperature of food: Not relevant as gavage method of administration used

VEHICLE
- Justification for use and choice of vehicle (if other than water): Arachidis oil, DAB 10.
- Concentration in vehicle: Not documented
- Amount of vehicle (if gavage): Not documented
- Lot/batch no. (if required): Not documented
- Purity: Not documented
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The test concentrations were analysed using High Performance Liquid Chromatography.
Details on mating procedure:
No information on mating procedure provided as the rats obtained from the source were primiparous time-mated females.
Duration of treatment / exposure:
GD 6-15
Frequency of treatment:
Once daily
Duration of test:
20 days post coitum
No. of animals per sex per dose:
24 female rats per dose group
Group 1: 23 pregnant rats (0mg/kg)
Group 2: 23 pregnant rats (40 mg/kg)
Group 3: 23 pregnant rats (120 mg/kg)
Group 4: 22 pregnant rats (360 mg/kg)
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The rationale for dose levels was based on the results of previous toxicological results.
- Rationale for animal assignment (if not random): Animals were randomly assigned.
- Other: No additional information.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily on working days

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule: Day 0, 6, 16 and 20 post coitum.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No data

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

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: All maternal oragans, with emphasis on the uterus, uterine contents, position of foetuses in the uterus and the number of corpura lutea

OTHER:
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No data
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: No data
- Other:
Fetal examinations:
- External examinations: Yes: all live foetuses
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: Yes: half per litter
Statistics:
If the variables could be assumed to follow a normal distribution, the Dunnett-test based on pooled variance was applied for comparison between the treated and and control groups. The Steel-test was applied when the data could not be assumed to follow a normal distribution. Fisher's exact test for 2 x 2 tables was applied if the variables could be dichotomized without loss of information (Bonferroni-Holm-corrected).
Indices:
No information provided
Historical control data:
No information provided

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
no effects observed
Description (incidence and severity):
No compound related symptoms were observed in any treatment groups
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
No deaths occurred in any of the test groups examined
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weight profiles of the pregnant females were essentially similar in all groups. Mean corrected body weight gain of the treatment groups 3 and 4 were statistically different to group 1 (the vehicle control).
Food consumption and compound intake (if feeding study):
no effects observed
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
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
Description (incidence and severity):
No macroscopic changes were noted in the dams of groups 1 - 4
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed

Maternal developmental toxicity

Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
In test groups 2 - 4, the sum of post-implantation loss was decreased by 5%, as was the sum of total embryonic deaths.The sum of embryonic deaths was also decreased by 1% in group 3. In groups 2 and 4, the sum of total foetuses was increased by 5% and in group 3 by 1%.
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
not examined
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): not examined
Changes in number of pregnant:
not examined
Other effects:
no effects observed
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
No deaths occurred in any of the test groups examined. No compound related symptoms were observed in any treatment groups. Body weight profiles of the pregnant females were essentially similar in all groups. Mean corrected body weight gain of the treatment groups 3 and 4 were statistically different to group 1 (the vehicle control). No compound related differences were noted between the mean reproduction data of the test groups in comparison to the control group. In test groups 2 - 4, the sum of post-implantation loss was decreased by 5%, as was the sum of total embryonic deaths.The sum of embryonic deaths was also decreased by 1% in group 3. In groups 2 and 4, the sum of total foetuses was increased by 5% and in group 3 by 1%. These findings were considered to be incidental and in the normal range. No macroscopic changes were noted in the dams of groups 1 - 4.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
360 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects

Maternal abnormalities

Key result
Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
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:
not examined
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Other effects:
no effects observed
Details on embryotoxic / teratogenic effects:
None of the visceral examinations revealed any treatment related abnormalities.

Group 1 (control) : 140 foetuses examined
9 foetuses displayed hydronephrosis
12 foetuses displayed waved ureters
9 foetuses displayed ureter dilatation
1 runt
1 foetus displayed hydrocephalous internus

Group 2 (40 mg/kg bw/d) : 152 foetuses examined
21 foetuses displayed hydronephrosis
12 foetuses displayed waved ureters
9 foetuses displayed ureter dilatation

Group 3 (120 mg/kg bw/d)
152 foetuses examined
16 foetuses displayed hydronephrosis
16 foetuses displayed waved ureters
8 foetuses displayed ureter dilatation
2 foetuses displayed gut protrusion out of the abdomen (considered an artefact)

Group 4 (360 mg/kg bw/d) : 162 foetuses examined
12 foetuses displayed hydronephrosis
2 foetuses displayed waved ureters
5 foetuses displayed ureter dilatation
1 foetus displayed haematoma periorbital
1 runt

On skeletal examination, statistically significant differences were considered to be incidental. In all treatment groups, there were no relevant findings observed when retardation was examined. Any variations observed concerning ossification and the number of ribs where not considered to be statistically significant and were incidental. Skeletal malformations were observed in one case in group 1, with irregularly placed skull bones, micrognathia, cleaved vertebrae (cervical/ thoracic/ lumbar /sacral) and non-ossification of the coccygeal vertebrae. In treatment group 3, there was one case of micrognathia and one case of partial ossification of the skull bones and non-ossification of the caudal vertebral column.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
360 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects

Fetal abnormalities

Key result
Abnormalities:
no effects observed

Overall developmental toxicity

Key result
Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
There was no evidence of developmental toxicity in this study at dose levels of up to 360 mg/kg bw/d, which were sufficient to result in mild maternal toxicity.
Executive summary:

The developmental toxicity of cyclohexyl salicylate was investigated in the rat. Groups of 23 mated female SD rats were gavaged with the test material (in arachis oil) on Days 6-15 of gestation. Clinical condition and reaction to treatment were recorded at least once daily. B ody weight was recorded on Days 0, 6, 16 and 20 of gestation.  All surviving females were sacrificed on Day 20 of gestation and the foetuses removed by caesarean section. Necropsy was performed and macroscopic examinations of the females conducted. Live foetuses were weighed, sexed and examined for visceral and skeletal abnormalities.  There were no signs of toxicity and no mortalities observed at any dose level. Maternal weight gain was not affected by any treatment and no treatment related abnormalities were noted at necropsy.  No treatment related foetal abnormalities were observed at necropsy and there were no effects on reproductive data. Maternal and developmental NOAELs of 360 mg/kg bw/d are determined for this study in the absence of any effects of toxicological significance at the highest dose level.