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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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
publication
Title:
Relative Developmental Toxicities of Acrylates in Rats following Inhalation Exposure.
Author:
Saillenfait AM, Bonnet P, Gallissot F, et al.,
Year:
1999
Bibliographic source:
Toxicol. Sciences 48: 240-254

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Groups of 20-29 bred female rats (17-25 pregnant) were exposed to the compound 6h/day on days 6 through 20 of gestation by inhalation. Control animals were exposed concurrently to filtered room air. The test concentrations of ethylhexyl acrylate were 50, 75, and 100 ppm (corresponding to approx. 0.38, 0.56, and 0.75 mg/L)*.
* Calculation of concentrations (mg/L) based on Derelanko MJ (2000). Toxicologist's Pocket Handbook, CRC Press, conversion table, p. 57
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Test substance: 2-ethylhexyl acrylate
- Chemical name: 2-propenoic acid, 2-ethylhexyl ester
- Analytical Purity: 99.7 % (GC)
- Supplier: Elf Atochem (France)

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: IFFA CREDO Breeding Laboratories (Saint-Germain-sur-l' Arbresle, France)
- Age at study initiation: Young, nulliparous females
- Weight at study initiation: 200-220 g
- Housing: Single in clear polycarbonate cages with stainless-steel wire lids and hardwood shaving as bedding.
- Diet: Food pellets (UAR Alimentation Villemoisson, France), ad libitum
- Water: Filtered tap water, ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2
- Humidity (%): 50 ± 5
- Photoperiod (hrs dark / hrs light): 12 hrs dark/12 hrs light

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
EXPOSURE
Exposures were conducted in 200-L glass/stainless-steel inhalation chambers with dynamic and adjustable laminar air flow (6-20 m3/h). The chamber temperature was set at 23 ± 2°C, and the relative humidity at 50 ± 5 %. The air-flow rate passed through the fritted disk of a heated bubbler containing the test chemical. Concentrations of acrylate ester were monitored continuously with a gas-chromatograph equipped with a flame ionization detector and an automatic gas-sampling valve. In addition, exposure levels were determined once during each 6-h exposure period by collecting atmosphere samples through glass tubes packed with activated charcoal. The charcoal samples were then desorbed with carbon disulfide. The resulting samples were analyzed by gas chromatography using appropriate internal standards. Since 2-EHA has a rather low vapour pressure (0.14 mm Hg at 20 °C), the presence of liquid particles was evaluated at the highest concentration generated (i.e. 100 ppm). Airborn particles were measured with an Aerodynamic Particle Sizer.
No differences in particle counts were observed between the clean filtered air (control) and the vapor-laden air in the exposure chambers.

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Glass/stainless-steel inhalation chambers
- Source and rate of air: Test atmospheres were generated through an additional air-flow  rate passed through the fritted disk of a heated bubbler containing ethylhexyl acrylate.  The vaporized compound was introduced into the main air inlet pipe of the exposure chamber.
- Air flow rate: 6-20 m3/h

TEST ATMOSPHERE
- Brief description of analytical method used: GC/FID
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analytical concentrations (mean ± SD):
51.0 ± 4.3 ppm (nominal: 50 ppm)
75.4 ± 9.2 ppm (nominal: 75 ppm)
102.5 ± 7.9 ppm (nominal: 100 ppm)
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 1/2-3
- Length of cohabitation: Overnight
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
Duration of treatment / exposure:
day 6-20 of gestation
Frequency of treatment:
6h /day
Duration of test:
until gestation day 21
Doses / concentrationsopen allclose all
Dose / conc.:
50 ppm (nominal)
Remarks:
corresponding to approx. 0.38
Dose / conc.:
75 ppm (nominal)
Remarks:
corresponding to approx. 0.56 mg/L
Dose / conc.:
100 ppm (nominal)
Remarks:
corresponding to approx. 0.75 mg/L
No. of animals per sex per dose:
20
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale:
Exposure concentrations were based on preliminary studies. Only minimal maternal toxicity was observed after exposure to 90 ppm ethylhexyl acrylate. Nevertheless, l00 ppm ethylhexyl acrylate was used as the highest concentration for the definitive developmental toxicity study, since preliminary level-setting studies have indicated that 100 ppm was the highest reliable vapor concentration technically possible.

Examinations

Maternal examinations:
BODY WEIGHT: Yes
- Time schedule for examinations: On gestation day (GD) 0, 6, 13 and 21.

FOOD CONSUMPTION: YES
Food consumption was measured for the intervals GD 6-13 and 13-21.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: The uteri were removed and weighed. The number of implantation sites, resorptions, and dead and live fetuses were recorded.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
The number of implantation sites, resorptions, and dead and live fetuses were recorded. Uteri which had no visible implantation sites were stained with ammonium sulfide (10 %) to detect very early resorptions.

Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: No data
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
Live fetuses were weighed, sexed, and examined for external anomalies including those of the oral cavity. Half of the live fetuses from each litter were preserved in Bouin's solution and examined for internal soft tissue changes. The other half were fixed in ethanol (70 %), eviscerated, and then processed for skeletal staining with alizarin red S for subsequent skeletal examination.

- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: Yes: all per litter
Statistics:
Data were presented as mean ± SD. The number of  implantation sites and live fetuses and the various body weights were analyzed by one-way analysis of variance (ANOVA), followed by Dunnett's test if differences were found. The percentages of non-live  implants and  resorptions and the proportions of fetuses with alterations in each litter were evaluated by using the Kruskal-Wallis test, followed by the Dixon-Massey test where appropriate. Rates of pregnancy, fetal sex ratio, and percentage of litters with malformations or external, visceral, or skeletal variations were analyzed by using Fisher's test. Where applicable, least-squares analysis was carried out. For all statistical tests, the level of significance was set a priori at alpha = 0.05.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
No test dams died. Except for a significant decrease in absolute weight gain at 100 ppm, there were no significant changes in maternal weight gain of females exposed to ethylhexyl acrylate, compared of those of controls. Rats from the 100 ppm-group showed a significant decrease in food intake throughout the entire exposure period (8-11 % reduction).

Effect levels (maternal animals)

Dose descriptor:
NOAEC
Effect level:
ca. 0.56 mg/L air (nominal)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity

Results (fetuses)

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

Details on embryotoxic / teratogenic effects:
No adverse effects were observed on the mean number of implantations and live fetuses among litters exposed to ethylhexyl acrylate. The statistically significant reductions in the incidence of non-live implants and resorptions at 50 and 100 ppm were not considered to be of toxicological significance. There was a statistically significant trend toward decreased fetal body weights, but the pairwise comparisons to the concurrent control group were not significantly different. No significant differences were observed between control and treated groups in the incidence of fetal malformations or variations.

Effect levels (fetuses)

Dose descriptor:
NOAEC
Effect level:
ca. 0.75 mg/L air (nominal)
Based on:
test mat.
Basis for effect level:
other: teratogenicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Maternal body weights:

Concentration [ppm/6h/d]

Maternal body weight GD 6 [g]

Absolute weight gain [g]

0

289 ± 17

42 ± 11

50

294 ± 19

35 ± 15

75

299 ± 21

32 ± 18

100

292 ± 18

24 ± 16**

Absolute weight gain: (Day 21 body weight) - (gravid uterus weight) - (Day 6 body weight)

Reproductive parameters:

 

Conc. [ppm/6h/d]

No. of litters

No. of implantation sites/litter

% of non-live implants/litter

% of resorption sites/litter

No. of live fetuses/litter

Average fetal body weight/litter [g]

0

25

15.64 ± 2.87

10.12 ± 10.57

10.12±10.57

14.20 ± 3.57

 5.74 ± 0.33

50

24

15.42 ± 4.23

3.65 ± 5.26*

3.65 ± 5.26*

14.88 ± 4.21

 5.69 ± 0.41

75

23

16.30 ± 3.80

6.44 ± 8.26

6.07 ± 7.97

15.26 ± 3.99

 5.58 ± 0.40

100

23

15.96 ± 2.46

3.93 ± 4.54*

3.93 ± 4.54*

15.30 ± 2.27

 5.53 ± 0.31

Concentration [ppm/6h/d]

0

50

75

100

Mean % of fetuses with:

- any malformations/litter

0.27 ± 1.33

1.89 ± 6.91

0.94 ± 2.54

0

- external variations/litter

0

4.41 ± 20.39

0

0

- visceral variations/litter

2.46 ± 5.98

7.33 ± 14.90

8.27 ± 11.36

4.76 ± 10.54

- skeletal variations/litter

20.62 ± 20.98

22.34 ± 18.63

17.40 ± 21.21

16.30 ± 15.21

- any variations/litter

11.86 ± 10.96

19.09 ± 21.09

12.78 ± 11.49

10.62 ± 10.63

* ,** Significant differences from the control (0 ppm) value, p 0.05, and p 0.01, respectively.

Applicant's summary and conclusion