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Developmental toxicity / teratogenicity

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developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Publication from the open literature. No detailed description of the substance tested and on test method. All information on teratogenicity will be considered together as weight of evidence for this endpoint.

Data source

Reference Type:
Cholesterol induced palatal clefts in the rat.
Buresh J.J., Urban T.J.
Bibliographic source:
Arch Oral Biol 12; 1221-1228

Materials and methods

Test guideline
no guideline followed
not applicable
(study was performed in pre-guidance period)
Principles of method if other than guideline:
A total of 20 female rats were used. For mating purposes males were placed with individual females 1 day prior to estrus (determined by means of vaginal smears) and remained 1 day after oestrus. The females were then kept in a community cage. The day of oestrus was counted as the zero day of pregnancy. Of the 20 females (number of rats for each specific dose not indicated), 15 became pregnant (3, 5 and 7 at 0, 5 and 10 mg cholesterol) and delivered 131 pups. All groups received subcutaneous injections during days 8-14 of pregnancy, with controls receiving vegetable oil and two groups receiving 5 mg cholesterol or 10 mg cholesterol, respectively in vegetable oil. The rats were sacrificed by ether on day 18 and the young were removed from the uteri and examined (dissection microscope) for any gross oral anomalies or other teratogenic manifestation. The mandibles and the tongues were removed for examination of the palates. The heads were fixed in alcohol and prepared using standard paraffin histological techniques. 10 µ sections were taken from the heads and stained using haematoxylin and eosin. Blood was taken from 10 rats at the time of sacrifice and serum cholesterol was determined using the method of Zarrow, Yochim and McCarthy, 1964 (spectrophotometer).
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Details on test material:
- Name of test material (as cited in study report): cholesterol

Test animals

other: Hotzmann albino
Details on test animals or test system and environmental conditions:
- Weight at study initiation: females were reported to have an average BW of 219.2 grams
- Housing: individually with male during mating, thereafter females were kept in a community cage.
- Diet: commercially prepared food ad libitum
- Water: ad libitum

Administration / exposure

Route of administration:
vegetable oil
Duration of treatment / exposure:
days 8-14 of pregnancy
Frequency of treatment:
once a day
Duration of test:
Until day 18 of pregnancy
Doses / concentrations
Doses / Concentrations:
0, 5, 10 mg
other: suspended in 2 mL vegetable oil and subcutaneously applied
No. of animals per sex per dose:
Number of pregnant females: 3 in control group, 5 in 5 mg cholesterol group, 7 in 10 mg cholesterol group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
In previous work (Buresh and Urban, see other study summary in this dossier) with cholesterol 25 mg as dosage level resulted in resorption and 15 mg gave 56.8% of palatal anomalies.


Maternal examinations:
Blood was taken from ten rats (4 controls, 2 females from 5 mg cholesterol group and 4 females from 10 mg cholesterol group) at the time of sacrifice and blood serum cholesterol was determined.
Ovaries and uterine content:
no data
Fetal examinations:
- External examinations: Yes, all per litter
- Head examinations: Yes, all per litter
not performed

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: increased blood serum cholesterol

Details on maternal toxic effects:
Average blood serum cholesterol was (not dose-related) increased by about a factor 2 in cholesterol treated animals.

Effect levels (maternal animals)

Dose descriptor:
Effect level:
5 other: mg/day
Based on:
test mat.
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes. Remark: increased number of abnormal palates and increase in resorption

Details on embryotoxic / teratogenic effects:
Increased number of abnormal palates, with 27% and 51.7% abnormalities at 5 mg and 10 mg cholesterol, respectively; in high dose group increased total number of resorptions (11 compared to zero in control). Dose related increase in number of complete clefts (0, 2 and 9, respectively) and in number of abnormal (splitting, microanatomical and complete clefts) palates (0, 10 and 30, respectively). in high dose group an increased number of resorptions was observed.

Effect levels (fetuses)

Dose descriptor:
Effect level:
5 other: mg/day
Based on:
test mat.
Basis for effect level:
other: teratogenicity

Fetal abnormalities

not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

The pups from females reeciving cholesterol were reported to be smaller in size than the controls, which was considered to indicate that the embryonic age was less than the chronological age in these animals.

Applicant's summary and conclusion

Cholesterol injections of 5 and 10 mg during days 8 to 14 of pregnacy resulted in Hotzmann albino rats in dose related increase in total number of abnormal palates in Hotzmann albino rats. In the 10 mg cholesterol group an increase in resorption was observed. Blood serum cholesterol levels of treated pregnant rats were (not dose-related) increased.
Executive summary:

In pregnant albino female rats the effect of 5 and 10 mg cholesterol injections during days 8 -14 of pregnancy was investigated. In both dose groups, a dose-related increased number of pups with abnormal palates was observed (27 and 51% abnormalities in 5 mg and 10 mg, respectively). In the 10 mg cholesterol group the number of resorptions was increased. Blood serum cholesterol levels of treated pregnant rats was increased (about factor 2), without dose-relationship.

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