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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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well documented study report, comparable to OECD guideline study with acceptable restrictions (non-GLP, no data on analytical purity)

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1988
Report date:
1988
Reference Type:
secondary source
Title:
Diesters Category of the Aliphatic Esters Chemicals (Test Plan and Robust Summaries for Substances in the HPV Test Plan)
Author:
US-EPA (American Chemistry Council's Aliphatic Esters Panel)
Year:
2010
Bibliographic source:
High Production Volume (HPV) Chemical Challenge Program (201-16837A and 201-16837B)

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
(adopted 2001)
Deviations:
yes
Remarks:
15 females/group instead of 20 females/group; only two dose levels tested instead of three as recommended by guideline, no GLP, no data on test item purity)
GLP compliance:
no
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Bis(tridecyl) adipate
EC Number:
241-029-0
EC Name:
Bis(tridecyl) adipate
Cas Number:
16958-92-2
Molecular formula:
C32H62O4
IUPAC Name:
ditridecyl adipate

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, New York
- Age at study initiation: approx. 9 weeks old
- Weight at study initiation: control: 237.2 ± 7.3 g; low dose: 239.8 ± 8.7 g; high dose: 234.9 ± 10.1 g
- Fasting period before study: no
- Housing: individually
- Diet: Purina Certified Rodent Chow #5002 (Meal); ad libitum
- Water: tap water, via an automatic watering system; ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-22
- Humidity (%): 40-60
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:
dermal
Vehicle:
unchanged (no vehicle)
Details on exposure:
TEST SITE
- Area of exposure: clipped, intact dorsal skin
- Type of wrap: open application, no wrap used
- Time intervals for shavings or clipplings: on gestation day 0 and once weekly thereafter

REMOVAL OF TEST SUBSTANCE
- Washing: no washing

TEST MATERIAL
- Constant volume or concentration used: yes
- The test material was measured using a 1.00 mL syringe (calibrated in 0.01 mL) for both groups, and during dispensing was spread evenly on the clipped dorsal skin of the rats using the tip of the syringe. (No needle was used.)

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes
To minimize ingestion of the test material, the rats were fitted with cardboard Elisabethan-style collars. These collars were lined with latex tubing to minimize the development of irritation or lesions.
Analytical verification of doses or concentrations:
no
Details on mating procedure:
- If cohoused: cohoused with male rats from in-house breeder population
- M/F ratio per cage: 1:1
- Duration of mating period: 5 days
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
Duration of treatment / exposure:
gestation days 0-19
Frequency of treatment:
once daily
Duration of test:
20 days
Doses / concentrationsopen allclose all
Dose / conc.:
800 mg/kg bw/day (nominal)
Dose / conc.:
2 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
15
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: The dose levels were chosen based on the results of a 13-week study conducted previously with the same material (ExxonMobil, Project 40552).
- Dermal control: Presumed-pregnant rats were clipped as the animals from the treatment groups. The intact dorsal skin of each rat was stroked with the tip of a 1.00 mL syringe, but no test material was applied.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once daily
- Cage side observations: mortality, clinical signs of ill-health (pathosis), appearance, abortion, premature delivery, behaviour

BODY WEIGHT: Yes
- Time schedule for examinations: on days 0, 3, 6, 10, 13, 16, and 20 of gestation

FOOD CONSUMPTION: Yes
- Food consumption for each animal was calculated for gestation day intervals 0-3, 3-6, 10-13, 13-16, and 16-20.

CLINICAL CHEMISTRY: Yes
- Blood samples were collected at the time of sacrifice from the abdominal aorta of each rat and allowed to clot in an SST Serum Separator Tube (Beckman-Dickinson, Rutherford, NJ).
- Parameters:
- Alanine aminotransferase (ALT),
- albumin,
- albumin/globulin ratio,
- alkaline phosphatase (ALP),
- aspartate aminotransferase (AST),
- total bilirubin,
- calcium,
- chloride,
- cholesterol,
- creatine,
- globulin,
- glucose,
- iron,
- lactate dehydrogenase (LDH),
- inorganic phosphorus,
- potassium,
- sodium,
- sorbitol dehydrogenase (SDH),
- total protein,
- triglycerides,
- urea nitrogen,
- uric acid.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day #20
- Organs examined: The thoracic and abdominal cavities were opened and all organs were examined grossly for evidence of pathological changes.

Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number and location of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes: all per litter
- Parameters: gender, body weight, length, gross external examination

- Soft tissue examinations: Yes: half per litter
- Fetuses were fixed in Bouin's solution, stored in 70% Ethanol and evaluated. Visceral analysis was performed usng a modification of the Wilson technique of free-hand sectioning by razor blade.

- Skeletal examinations: Yes: half per litter
- Fetuses were peeled, eviscerated, fixed in 95% ethanol, macerated in potassium hydroxide, differentially stained for cartilage and bone, cleared in glycerine, and examined for skeletal anomalies.

- Head examinations: Yes: half per litter
Statistics:
Maternal biophase and caesarean section data, and fetal data were evaluated by analysis of variance followed by group comparisons using Fisher's Exact or Dunnet's Test. Fetal skeletal data were evaluated statistically by ANOVA followed by group comparisons using Fisher's Exact Test. Fetal visceral data were evaluated statistically using Fisher's Exact Test. Statistical analyses of clinical chemistry data were performed separately on individual serum components using SAS procedures. First the F-test was employed to do an analysis of variance on the serum data obtained from control and exposed groups. Next, the Student-Newman-Keul's multiple comparison test was employed to identified the specific group subsets within the serum data sets identified as having nonrandom variance. The observed statistically significant differences were reported only when a dose-related effect was found in the treated animals.
In general, for all statistical tests, differences between control and treated groups were considered statistically significant if the probability of the difference being due to chance was less than 5% (p<0.05).
Historical control data:
yes

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: Body weights were statistically significantly decreased in the animals of the high-dose group. Statistically significant differences between the serum data from control and treated rats were observed for a number of parameters.

Details on maternal toxic effects:
- Observations during gestation:
Red nasal exudate, chromodacryorrhea and neck lesions were observed equally in control and treated animals. According to the author of the study, these are common findings in animals that were collared.
Signs of dermal irritation, including erythema and flaking of the skin, were observed in most of the animals from both treatment groups. In general, the irritation was very mild. Scabs were observed at the side of test substance application in two animals exposed to 800 mg/kg bw/day.

- Maternal body weights and food consumption:
In general, the mean body weight of the pregnant rats from all of the groups increased throughout gestation, following what would be regarded as a normal growth curve of weight gain. However, animals from the high-dose group gained significantly less (p<0.01) weight during the first and last interval of gestation, which subsequently contributed to the reduced weight gain throughout the entire period of gestation. Animals from the low-dose group demonstrated a significant reduction in body weight gain only during gestation days 0-3. Overall net body weight change was significantly lower in the 2000 mg/kg bw/day-group when compared to the control. With the exception of the first two intervals of food consuption recording, animals exposed to the substance consumed more food than control animals. It should be noted, however, that the total amount of food consumed during gestation by each of the experimental groups was comparable.

- Observations of dams at caesarean section:
At the time od necropsy, no findings attributable to exposure to the test substance were observed.

- Clinical chemistry analyses:
Statistically significant differences between the serum data from control and treated rats were observed for glucose, ALT, alkaline phosphatase, creatinine, cholesterol, triglycerides, total protein, iron, and globulin. A linear relationship (>99% confidence level) was found between dose and serum level for these components except glucose and ALT. When the historical serum reference values were taken into consideration, a portion of the dose-response curves for alkaline phosphatase, cholesterol, total protein, and globulin fall outside the normal range.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Remarks:
maternal
Effect level:
2 000 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: The decrease in body weight seen at 2000 mg/kg bw/d was not considered to be an adverse effect.

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes. Remark: In the fetuses exposed to the test substance at 2000 mg/kg bw/day a statistically significantly increased incidence of levocardia was observed. In addition, in the high-dose fetuses an increased incidence of hydronephrosis was noted.

Details on embryotoxic / teratogenic effects:
- Fetal body measurements:
Mean fetal body weights and crown-rump lengths were not affected by exposure to the test substance.

- Fetal examinations:
With the exception of one fetus exposed in utero to the test substance at 2000 mg/kg bw/day, no remarkable findings were observed during external examination of the fetuses. The mentioned fetus, in addition to being edematous, had a shortened thoracic-lumbar (trunk) region, malrotated hindlimbs, absence of digits (ectrodactyly) on its forepaws, and no tail (acaudia). Due to their occurrence in only one fetus (fetal incidence = 1/191 = 0.5%), these anomalies were probably not related to test substance exposure.
A variety of fetal skeletal anomalies were observed in all groups at a low incidence. They were probably not related to substance exposure due to their low incidence, their occurrence in control fetuses, and/or lack of a dose-response-relationship.
Fetal visceral anomalies observed included levocardia (malrotation of the heart) and hydronephrosis, as well as several other renal developmental variations. Of the levocardia and the hydronephrosis (both classified as malformations), only the former was observed significantly more often in the 2000 mg/kg bw/day group than in the control group. All other findings were classified as variations.

Effect levels (fetuses)

open allclose all
Key result
Dose descriptor:
LOAEL
Remarks:
developmental
Effect level:
2 000 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: increased incidence of levocardia (malrotation of the heart)
Key result
Dose descriptor:
NOAEL
Remarks:
developmental
Effect level:
800 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: increased incidence of levocardia (malrotation of the heart) at 2000 mg/kg bw/day

Fetal abnormalities

Key result
Abnormalities:
effects observed, treatment-related
Localisation:
visceral/soft tissue: cardiovascular
Description (incidence and severity):
levocardia (malrotation of the heart); significantly increased at 2000 mg/kg bw/day

Overall developmental toxicity

Key result
Developmental effects observed:
yes
Lowest effective dose / conc.:
2 000 mg/kg bw/day (nominal)
Treatment related:
yes
Relation to maternal toxicity:
developmental effects in the absence of maternal toxicity effects
Dose response relationship:
yes
Relevant for humans:
not specified

Any other information on results incl. tables

Table 1: Mean body weights (g)

 

 

 

Dose [mg/kg bw/day]

Day

 

800

2000

0.0

0

Mean (SD)

239.8 (8.7)

234.9 (10.1)

237.2 (7.3)

3

 

245.2 (11.7)

234.0 (11.6)**

252.4 (7.6)

6

 

258.3 (11.4)

250.6 (9.3)**

264.5 (7.0)

10

 

281.9 (11.7)

270.0 (10.8)**

285.2 (11.6)

13

 

295.2 (13.9)

284.5 (14.1)*

298.6 (10.4)

16

 

323.5 (12.5)

311.2 (17.5)

322.2 (10.4)

20

 

385.5 (17.3)

366.1 (21.7)*

386.0 (13.5)

significantly different from control: * p<0.05, ** p<0.01

 

 

Table 2: Serum chemistry data

 

Serum parameters [unit]

Dose [mg/kg bw/day]

 

800

2000

0.0

 

Mean (SD)

%

Mean (SD)

%

Mean (SD)

Glucose [mg/dL]

105.2 (13.8)

 

102.7 (9.3)*

-10

114.0 (12.3)

ALT [IU/L]

46 (8)

 

51 (7)*

+19

43 (7)

Alk. Phosphatise [IU/L]

118 (46)

 

140 (55)*°

+50

93 (31)

Creatinine [mg/dL]

0.63 (0.07)*

-7

0.61 (0.04)*

-10

0.68 )0.05)

Cholesterol [mg/dL]

72.6 (15.1)

 

59.4 (8.2)*°

-25

79.4 (10.5)

Triglycerides [mg/dL]

270.7 (83.9)

 

204.5 (58.8)*

-39

335.3 (123.3)

Total protein [mg/dL]

5.0 (0.4)*

-6

4.8 (0.2)*°

-9

5.3 (0.3)

Globulin [g/dL]

2.3 (0.3)*

-8

2.2 (0.1)*°

-12

2.5 (0.1)

Iron [µg/dL]

106 (35)

 

141 (53)*

+52

93 (26)

significantly different from control: * p<0.05; ° outside of historical control range

 

Table 3: Summary of visceral malformations

 

Group

 

Dose [mg/kg bw/day]

 

 

800

2000

0.0

Number examined

Litters

13

13

15

 

Fetuses

94

93

108

Number normal [%]

Litters

7 (53.8%)

3 (23.1%)*

10 (66.7%)

 

Fetuses

88 (93.6%)

72 (77.4%)*

102 (94.4%)

 

 

 

 

 

Levocardia

Litters

5 (38.5%)

10 (76.9%)*

5 (33.3%)

 

Fetuses

5 (5.3%)

21 (22.6%)*

6 (5.6%)

Hydronephrosis, right

Fetuses

1 (7.7%)

0 (0%)

0 (0%)

 

Litters

1 (1.1%)

0 (0%)

0 (0%)

Hydronephrosis, left

Fetuses

0 (0%)

2 (15.4%)

0 (0%)

 

Litters

0 (0%)

2 (2.2%)

0 (0%)

Hydronephrosis, bilateral

Fetuses

0 (0%)

1 (7.7%)

0 (0%)

 

Litters

0 (0%)

1 (1.1%)

0 (0%)

significantly different from control: * p<0.05

Applicant's summary and conclusion