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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:
weight of evidence
Study period:
05 - 27.07.1994
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study without detailed documentation

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1997
Report date:
1997

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
May 12, 1981
Qualifier:
according to guideline
Guideline:
other: Off. J. Europ. Commun. L 133, May 30, 1988; 87/302/EEC
Qualifier:
according to guideline
Guideline:
other: Off. J. Europ. Commun. L 180, March 01, 1991; 91/325/EEC
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Reference substance name:
Fatty acids, C16-18, esters with ethylene glycol
EC Number:
292-932-1
EC Name:
Fatty acids, C16-18, esters with ethylene glycol
Cas Number:
91031-31-1
Molecular formula:
C18-38H36-74O3-4
IUPAC Name:
Fatty acids, C16-18, esters with ethylene glycol

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River, Wiga GmbH, D-97633 Sulzfeld
- Age at study initiation: 8 weeks
- Weight at study initiation: mean 198 g
- Fasting period before study: no
- Housing: One animal / Makrolon Type M3 cage (Ebeco, 0-44579 Castrop-Rauxel) with standard softwood bedding (ARWI-Center, 0-45307 Essen).
- Diet: Pelleted Altromin Maintenance Diet 1324, lot No. 170994/1340 (Fa. Altromin GmbH, 0-32770 Lage) ad libitum
- Water: Community Tap Water from Düsseldorf ad libitum (analysed monthly for use as drinking water)
- Acclimation period: 5 days under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 24 °C
- Humidity (%): 47 - 82 %
- Air changes (per hr): 10 - 15 air changes per hour
- Photoperiod (hrs dark / hrs light): lux units 40 - 500, 12 hours artificial fluorescent light /12 hours dark

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
0.5 % sodium carboxymethylcellulose + 0.25 % Cremophor in aqua desto (CMCC) was used as vehicle for the test substance. The test article was prepared daily before administration. The concentrations of the test substance in CMCC, based on the results of analysis of the dry weight, are as foliows:
1 % (100 mg/kg) = 1.1 %
3 % (300 mg/kg) = 3.3 %
10 % (900 mg/kg) = 9.6 %
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The formulation of the test article was analysed once for achieved concentration by "Henkel TTA-Analytical Centre". No details are available.
Details on mating procedure:
The females were mated at the supplier with an accurate day of mating. They were received at the testing facility on day 0 of gestation.
Duration of treatment / exposure:
The test substance was administered once daily in the morning from day 6 p.c. to day 15 p.c. (10 applications).
Frequency of treatment:
Once daily.
Duration of test:
20 days
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
group 2
Dose / conc.:
300 mg/kg bw/day (nominal)
Remarks:
group 3
Dose / conc.:
900 mg/kg bw/day (nominal)
Remarks:
group 4
No. of animals per sex per dose:
at least 24 females
Control animals:
yes, concurrent vehicle

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice daily (working days)

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: at least twice daily (working days)

BODY WEIGHT: Yes
- Time schedule for examinations: on day 0, 6, 16 and 20

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Details: Gross macroscopic examination of all maternal organs with emphasis on the uterus, uterine contents, position of fetuses in the uterus and number of corpora lutea was performed and the data recorded. Number and distribution of intrauterine implantations were classified as live or dead fetuses, late intrauterine deaths (resorptions), early intrauterine (resorption sites).
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Details: Gross macroscopic examination of all maternal organs with emphasis on the uterus, uterine contents, position of fetuses in the uterus and number of corpora lutea was performed and the data recorded. Number and distribution of intrauterine implantations were classified as live or dead fetuses, late intrauterine deaths (resorptions), early intrauterine (resorption sites).
Fetal examinations:
The fetuses were removed from the uterus. Intrauterine deaths were classified on the basis of the presence (late) or absence (early) of fetal or decidual tissue in addition to placental tissue. The live fetuses were sexed, weighed individually including placentae, examined for gross external abnormalities and allocated to one of the following procedures:
1) Half of the fetuses from each litter was non-individually fixed in Bouin's solution in order to examine viscera and brain by Wilson's slicing technique. After examination the sections were not preserved. All abnormalities were recorded.
2) The remaining fetuses from each litter were fixed non-individually in ethanol for the following staining with alizarin red (Shandon Varistain 24-T). The skeletons were examined and preserved in pure glycerine in plastic containers. All abnormalities were recorded.
The uteri (including content) of all females were weighed at necropsy on day 20 p.c. to enable the calculation of the corrected body weight gain.
Statistics:
The following statistical methods were used: If the variables could be assumed to follow a normal distribution, the Dunnett-Test, based on a pooled variance, was applied for the comparison between the treated groups and the control group.
The Steel-Test was applied when the data could not be assumed to follow a normal distribution. Fisher's Exact test for 2x2 tables was applied if the variables could be dichotomized without loss of information (Bonferroni-Holm-corrected).
Historical control data:
Not available.

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 the treatment groups in comparison to the control group.
Mortality:
no mortality observed
Description (incidence):
No death occurred in the dams of the vehicle control group and in the test groups.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Body weight profiles of the pregnant females were similar in all groups. Mean corrected body weight gain (corrected for uterus weight) of the treatment groups compared favourably with the control values.
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:
no effects observed
Description (incidence and severity):
At scheduled necropsy no macroscopic changes were noted in the dams of the groups.
Gross pathological findings:
not examined
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined

Maternal developmental toxicity

Number of abortions:
no effects observed
Description (incidence and severity):
Abortions of test groups in range of control group.
group 1: 15, group 2: 18, group 3: 12, group 4: 19
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
Pre- and post implantation loss of test groups in range of control group.
Pre-implantation loss (% of corp. lutea): group 1: 8.8, group 2: 9.9, group 3: 10.9, group 4: 10.4
Post-implantation loss (% of impl. sites): group 1: 4.3, group 2: 5.1, group 3: 3.6, group 4: 6.1
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
Total litter losses of test groups in range of control group.
Embryonic death: group 1: 15, group 2: 18, group 3: 12, group 4: 19
Early or late resorptions:
no effects observed
Description (incidence and severity):
Resorptions of test groups in range of control group.
Embryonic resorption: group 1: 14, group 2: 17, group 3: 12, group 4: 16
Fetal resorption: group 1: 1, group 2: 1, group 3: 0, group 4: 3
Dead fetuses:
no effects observed
Description (incidence and severity):
Number of dead fetuses of test groups in range of control group.
Dead fetuses: group 1: 0, group 2: 0, group 3: 1, group 4: 1
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
not examined

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEC
Effect level:
900 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: no adverse effects observed

Maternal abnormalities

Key result
Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
The mean weight of live male and female fetuses of the group 3 were significantly increased. The weights of live fetuses of the group 2 and 4 exibited no significant differences on a litter and individual basis e. g. mean weight in comparison to the control group.
Reduction in number of live offspring:
not examined
Changes in sex ratio:
no effects observed
Description (incidence and severity):
Sex ratio of test groups in range of control group.
Males: group 1: 159, group 2: 174, group 3: 174, group 4: 160
Females: group 1: 177, group 2: 163, group 3: 171, group 4: 146
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
Litter size and weights of test groups in range of control group.
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
No macroscopical findings were noted at external examination of fetuses which were considered to be an effect of the treatment with the test article. The following abnormal spontaneous findings were ascertained during external examination:
One fetus with a missing tail (see skeletal examination) (female 23, group 1).
One dead fetus approx. on day 13 (no signs of malformation) (female 52, group 3).
One dead fetus approx. on day 16 with hydrops/hydrocephalus (female 75, group 4).
One fetus with beginning hydrops fetalis (see visceral examination: runt, cleft palate) (female 81, group 4)
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Following abnormal spontaneous findings were noted:
One fetus with acaudia (coccygeal vertebrae non ossified, sacral vertebrae 2 - 5 non ossified) (female no. 23, group 1).
One fetus with incompletely ossified palate (female no. 25, group 2).
Two fetuses with thickened and nodular ribs, shortened intervertebral spaces in the thoracic region and curved clavicles (female no. 59, group 3).
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The findings were as follows:
Group I, 160 examined fetuses: 10 (6.3 %) hydronephrosis; 8 (5.0 %) ureter dilatation; 5 (3.1 %) ureter waved
Group 2, 164 examined fetuses: 19 (11.6 %) hydronephrosis; 10 (6.1 %) ureter dilatation; 7 (4.3 %) ureter waved
Group 3, 165 examined fetuses: 15 (9.1 %) hydronephrosis; 8 (4.8 %) ureter dilatation; 1 (0.6 %) ureter waved; 1 ( 0.6 %) esophagus dilatation
Group 4, 146 examined fetuses: 28 (19.2 %) hydronephrosis; 13 (8.9 %) ureter dilatation; 7 (4.8 %) ureter waved; 1 (0.7 %) hematoma submandibular region; 1 (0.7 %) runt, cleft palate
The visceral examination of the preserved fetuses did not reveal any treatment related abnormalities.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
A generally retarded ossification was found in:
One fetus (female no. 17, group 1).
Two fetuses (female no. 82, group 4).

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
900 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed

Fetal abnormalities

Key result
Abnormalities:
no effects observed

Overall developmental toxicity

Key result
Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
The test item does not reveal any embryotoxic or teratogenic potential at dose levels up to 900 mg/kg body weight/day in an OECD 414 test.
Executive summary:

The effect of the test item on embryonic and fetal development in pregnant CD-rats was assessed in a test according to OECD guideline 414. The test item was tested at dose levels of 0 (group 1), 100 (group 2), 300 (group 3) and 900 (group 4) mg/kg bw/day. 24 female rats per group at the study start (pregnant rats: Group 1: 23; group 2: 23; group 3: 24; group 4: 22). The test item was administered orally by gavage once daily from day 6 to day 15 of gestation (10 applications). A standard dose volume of 10 mL/kg body weight was used adjusted to the body weight of day 6 post coitum (p.c.). Control animals were dosed with the vehicle alone (0.5 % sodium carboxymethylcellulose + 0.25 % Cremophor in aqua dest. - CMCC) over the period described. Clinical condition and reaction to treatment were recorded at least once daily. Body weights were reported for days 0, 6, 16 and 20 of gestation. All surviving females were sacrificed on day 20 of gestation and removed by caesarean section. At necropsy the females and live fetuses were weighed, sexed and examined for skeletal abnormalities. The dams tolerated the applied dose levels up to 900 mg/kg bw/day test item without lethality. No substance-related symptoms were observed in the treatment groups 2 - 4 in comparison to the control group 1. Maternal body weight gain was not affected by treatment. No treatment-related abnormalities were found at necropsy of the females. All females had viable fetuses. Pre-implantation loss, post-implantation loss, mean numbers of resorptions, embryonic deaths and total fetuses were not affected by treatment. Mean fetal placental and uterus weights were not affected by treatment. Fetal sex ratio was comparable in all groups. No treatment-related fetal abnormalities were found at necropsy. There were no treatment-related effects in the reproduction data. The examined fetuses showed no treatment-related malformations. The figures of visceral variations in the test groups were considered to be similar to the control group. The figures of skeletal ossifications and variations showed no treatment-related deviations. The results of this study showed that repeated oral administration (day 6 p.c. to day 15 p.c.) of the test item to pregnant rats caused no symptoms of cumulative toxicity up to 900 mg/kg body weight/day. According to the study described, the test item does not reveal any embryotoxic or teratogenic potential at dose levels up to 900 mg/kg body weight/day.

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