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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP status unknown, guideline study, published in peer reviewed literature, no restrictions, fully adequate for assessment.

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2003

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- CAS number: 100-41-4
- Purity: > 99.5%
- Source: Fluka chemie AG (Buchs, Switzerland)

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)
- Body weight at study initiation: 180 - 200 g
- Housing: Individually in clear polycarbonate cages with stainless-steel wire lids and corn cob granules as bedding
- Diet: Food pellets (UAR Alimentation Villemoisson, France) ad libitum except during exposures
- Water: filtered tap water ad libitum except during exposures
- Acclimation period: 2 weeks

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

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
other: air
Details on exposure:
- Exposure apparatus: 200 L glass/stainless steel inhalation chambers with dynamic and adjustable laminar air flow (6-8 m3/h), maintained at a negative pressure of ≤3 mm water.
- System of generation: a constant rate of liquid chemical was delivered (with a HPLC pump for 2000 ppm or with an infusion pump for the other concentrations). This liquid chemical was delivered at the top of a heated glass column filled with glass beads. Compressed air heated by a glass heater was introduced at the bottom of the glass column in a counter-current fashion to the liquid flow. Under both conditions, the vaporized compounds were introduced into the main air-inlet pipe of the exposure chambers.
- Temperature, humidity, pressure in air chamber: 23±2°C, 50±5%

TEST ATMOSPHERE
- Brief description of analytical method used: gas chromatography with a flame ionization detector
- Samples taken from breathing zone: no data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The actual concentrations were determined by gas chromatography with a flame ionization detector. The column temperature was maintained at 80°C. The concentrations determined by analyses were essentially the same as the target concentrations.
Details on mating procedure:
Nulliparous females were housed overnight with adult males (one male to two or three females) from the same strain and supplier. The day that vaginal smears were found to be sperm-positive was considered day 0 of gestation (GD).
Duration of treatment / exposure:
6 hr/day
Frequency of treatment:
Daily, from day 6 through 20 of gestation.
Duration of test:
21 days
No. of animals per sex per dose:
23 - 26 mated females/group; 20 - 26 pregnant females/group
Control animals:
yes, concurrent vehicle
Details on study design:
Control animals were exposed concurrently to filtered room air in an adjacent chamber identical to those of the treatment groups.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- recorded on GD 0, 6, 13 and 21
- Body weight changes were calculated for the following gestation intervals: 0-6, 6-13 and 13-21.
- The corrected weight gain was the body weight gain between GD 6-21 subtracted from gravid uterus weight.

FOOD CONSUMPTION: Yes
- Measured for the intervals GD 6-13 and 13-21

POST-MORTEM EXAMINATIONS: Yes - killed on gestation day 21
- Organs examined: Uterus
Ovaries and uterine content:
The uterus was removed and weighed. The number of corpora lutea, implantation sites, resorptions, and dead and live foetuses were recorded. Uteri with no visible implantation sites were stained with ammonium sulphide (10%) to detect very early resorptions.
Fetal examinations:
Live foetuses were weighed, sexed, and examined for external anomalies including those of the oral cavity. Half of the live foetuses 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.
Statistics:
Where appropriate, the data were presented as mean ± SD. One-way analysis of variance was used to analyse the number of corpora lutea, implantation sites and live foetuses, maternal food consumption and body weights and was followed by Dunnett's test where differences were found. The Kruskal-Wallis test was used to evaluate the percentages of non-live implants, resorptions, and males, and the proportions of foetuses with alterations in each litter and was followed by the Mann-Whitney test where appropriate. Pregnancy rates and percentages of litters with any malformations or with external, visceral or skeletal variations were analysed using Fisher's test. Least-squares analysis was carried out where applicable. The level of statistical significance reported was P<0.05. The litter was the unit of analysis for foetal variables.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
- Mortality: No maternal deaths.
- Bodyweight: Exposure to 1000 or 2000 ppm resulted in a significant decrease between GD 6 and GD 13, GD 13 and 21 and GD 6 and 21. Corrected weight gain was lower than controls by 49% and 97% respectively)
- Food consumption: Exposure to 1000 or 2000 ppm resulted in a significant decrease between GD 6 and GD 13, GD 13 and 21 and GD 6 and 21.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEC
Remarks:
ethyl benzene
Effect level:
500 ppm
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEC
Remarks:
ethyl benzene
Effect level:
500 ppm
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
- No effects on mean number of implantation sites, live foetuses, incidence of non-live implants and resorptions although 3 females exposed to 2000 ppm had 100% total litter loss.
- Foetal body weights were significantly lower than controls by 7% at 1000 ppm and 18% at 2000 ppm.
- The occurrences of foetuses with external or visceral variations did not differ between the control and the treatment groups. Visceral malformations occurred sporadically in one or two foetuses and were distributed across the groups.
- Increased incidence of foetuses with skeletal variations in the 1000 and 2000 ppm groups.

Effect levels (fetuses)

Dose descriptor:
NOAEC
Remarks:
ethyl benzene
Effect level:
2 000 ppm
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

Intergroup comparison of the incidence of skeletal variations

(Table based on Saillenfait AM et al, 2003, Food and Chemical Toxicology 41 415-429,Table 6)

Parameter

Exposure (ppm)

0

100

500

1000

2000

Total no foetuses (litters)

177 (24)

174 (24)

175 (24)

175 (25)

119 (18)

No (%) foetuses with skeletal variations

28 (15.8)

35 (20.1)

30 (17.1)

46 (26.3)#

40 (33.6)##

Mean % foetuses with skeletal variations per litter

16.1±17.3

20.7±26.5

16.7±18.1

26.1±21.0

35.6±26.0**

# ## denotes significant difference from control P<0.05 and P<0.01 respectively, Fischer's test

** denotes significant difference from control P<0.01, Mann-Whitney test

Applicant's summary and conclusion

Conclusions:
The NOAEC for maternal toxicity and foetal effects was 500 ppm (2171 mg/m3).
Executive summary:

Inhalation exposure of Sprague-Dawley rats from gestation days 6-20 to ethylbenzene resulted in maternal toxicity at 1000 and 2000 ppm. Foetal toxicity (reduced foetal weight and an increased incidence of skeletal variations) was seen at 1000 and 2000 ppm, in the presence of maternal toxic effects. Ethylbenzene was not teratogenic up to 2000 ppm. The NOAEC for maternal and developmental toxicity was 500 ppm.