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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:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: comparable to guideline study
Cross-reference
Reason / purpose:
reference to same study

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Unnamed
Year:
1995
Report Date:
1995
Reference Type:
study report
Title:
Unnamed
Year:
1990
Report Date:
1990
Reference Type:
study report
Title:
Unnamed
Year:
1990
Report Date:
1990

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 2-methyl-1-propanol, MEP
- Analytical purity: 99.8%
- Impurities (identity and concentrations): no data
- Purity test date: no data
- Lot/batch No.: no data
- Stability under test conditions: yes
- Storage condition of test material: no data

Test animals

Species:
rabbit
Strain:
Himalayan
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Thomae GmbH, Biberach/Riss, Germany
- Age at study initiation: 24 - 29 weeks
- Weight at study initiation: 2.5 - 2.7 kg
- Fasting period before study: no data
- Housing: individually in wire-mesh cages, type K 300/8 (EBECO, Becker and Co., Castrop Rauxel, Germany), air conditioned rooms
- Diet (e.g. ad libitum): KLIBA rabbit laboratory diet 24-341-4, 10 mm pellets (Klingentalmühle AG, Kaiseraugst, Switzerland)
- Water (e.g. ad libitum): tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 /12

Administration / exposure

Route of administration:
inhalation
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: horizontal-flow whole-body exposure chamber (glas/steel construction, volume 1.1.m³ (BASF AG, Ludwigshafen, Germany)
- Method of holding animals in test chamber: individually in wire cages
- Source and rate of air: clean air
- Method of conditioning air: no data
- System of generating particulates/aerosols: evaporator maintained at 50-70°C, TS was delivered by a continuously operating pump
- Temperature, humidity, pressure in air chamber: 21-24°C, 49-64%, minimal negative pressure
- Air flow rate: 275 L/minute
- Air change rate: 15 air changes per hour
- Method of particle size determination: no particle size determination
- Treatment of exhaust air: no data

TEST ATMOSPHERE
- Brief description of analytical method used: gas chromatography (Hewlett-Packard GC, Model 5840 A with FID detector)
- Samples taken from breathing zone: no data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of the inhalation atmosphere were analyzed hourly during exposure.
Mean concentrations ± SD measured in inhalation chamber: 0.50 ± 0.010 mg/L, 2.51 ± 0.091 mg/L, 10.00 ± 0.370 mg/L

Details on mating procedure:
- Impregnation procedure: artificial insemination
- Verification of same strain and source of both sexes: no data
- Proof of pregnancy: day of insemination referred to as day 0 of pregnancy, the following day was defined as day 1 of pregnancy or postinsemination (pi) respectively
Duration of treatment / exposure:
day 7 - 19 of pregnancy (pi)
Frequency of treatment:
6 hours / day
Duration of test:
29 days
Doses / concentrations
Remarks:
Doses / Concentrations:
0.5; 2.5; 10 mg/L
Basis:
nominal conc.
No. of animals per sex per dose:
15 female rabbits per group
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: preliminary range-finding study
- Rationale for animal assignment (if not random): random

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule:

BODY WEIGHT: Yes
- Time schedule for examinations: day 0, 3, 7, an from then on at 3-day intervals until day 29

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 29 by intravenous application of pentobarbitone
- Organs examined: uterus and ovaries

OTHER: Yes
- gross pathology for all animals
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 of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all per litter; fixation in Bouin's solution according to the method of Barrow and Taylor (1969)
- Skeletal examinations: Yes: all per litter; fixation in ethyl acohol and staining according to a modified method of Dawson (1926)
- Head examinations: Yes: No data
Statistics:
The Dunnett test (Dunnett, 1955, 1964) was used to statistically compare body weight, body weight changes, the number of corpora lutea, implants, resorptions, live fetuses, and pre- or postimplantation losses. The Fisher's exact test (Dixon, 1981) was used for evaluating the conception rate, maternal mortality, and all fetal findings.
Historical control data:
Historical control data were used to further evaluate findings of uncertain relevance.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
One rabbit of the 0.5 mg/L group was killed on day 21 of gestation due to abortion. One rabbit exposed to 2.5 mg/L was found dead on day 24 of gestation. Both cases were considered not to be related to isobutanol inhalation.

10 mg/L group
Slight retardation of body weight gain of the dams, especially during the first of the exposure periods.
Indication of an irritant eye effect (reddish, lid closure, slight discharge)

2.5 mg/L and 0.5 mg/L group
no substance-related effects were observed.

Gross-pathologiy examination revealed no effects that could be attributed to the exposure with isobutanol.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
2.5 mg/L air
Basis for effect level:
other: maternal toxicity

Results (fetuses)

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

Details on embryotoxic / teratogenic effects:
All treatment groups
The fetuses did not showed any substance-related effects (no indications of any embryo-/fetotoxicity or any teratogenic effect).

The uterine weights of the treated animals were not significantly different from the controls. Compound related effects occured neither for conception rate, mean number of corpora lutea, and implantation sites nor in the values calculated for the pre- and postimplantation loss and the number of resorptions as well as viable fetuses.

Effect levels (fetuses)

open allclose all
Dose descriptor:
NOAEL
Effect level:
10 mg/L air
Basis for effect level:
other: fetotoxicity
Dose descriptor:
NOAEL
Effect level:
10 mg/L air
Basis for effect level:
other: embryotoxicity
Dose descriptor:
NOAEL
Effect level:
10 mg/L air
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

Table 1     Data on Reproduction, Maternal Body Weight Change and Uterine Weight

Dose [mg/L]

0 (sham)

0.5

2.5

10.0

Number of animals

15

15

15

15

Number of dams (pregnant animals)

15

15

14a,b

14a

Corpora lutea/dam

8.4

8.6

7.7

7.6

Implants/dam

7.5

6.7

6.5

7.0

Live fetuses/dam

7.0 (1.20)

6.2 (1.72)

6.0 (2.20)

7.1 (1.32)

Dead implants/dam

0.5

0.5

0.5

0.4

Sex ratio (male / female

48.6 / 51.4

51.7 / 48.3

44.9 / 55.1

50.0 / 50.0

Mean fetal weight [g]

39.0 (3.74)

39.7 (4.01)

41.0 (3.46)

39.4 (1.61)

Mean placental weight [g]

4.3 (0.48)

4.6 (0.56)

4.8* (0.66)

4.4 (0.40)

Mean corrected maternal body weight change [g] **

-71.9 (65.77)

-45.3 (63.91)

-47.1 (41.43)

-67.0 (77.51)

Intact uterine weight [g]

367.0 (52.48)

329.3 (69.99)

330.1 (110.81)

344.3 (111.43)

Numbers in parentheses indicate standard deviations

* significantly different from control, p < 0.05

** corrected maternal body weight change = (body weight on day 20 of pregnancy) - (body weight on day 6 of pregnancy) - (uterine weight)a    one not pregnant animalb    one animal aborted

Table 2               Fetal Anomalies

Dose [mg/L]

0 (sham)

0.5

2.5

10

Number of litters evaluated

15

14

13

13

Number of fetuses evaluated

105

87

78

92

Total fetal external malformations

Fetal incidence

0

0

0

0

Litter incidence

0

0

0

0

Total fetal soft tissue malformation

Fetal incidence

0

0

0

0

Litter incidence

0

0

0

0

Total fetal soft tissue variations

Fetal incidence

21 (20.0)

13 (14.9)

17 (21.8)

30 (32.6)

Litter incidence

11 (73.3)

9 (64.3)

7 (53.8)

12 (92.3)

Total fetal skeletal malformations

Fetal incidence

2 (1.9)

1 (1.1)

0

1 (1.1)

Litter incidence

2 (13.3)

1 (7.1)

0

1 (7.7)

Total fetal skeletal variations

Fetal incidence

9 (8.6)

14 (16.1)

8 (10.3)

16 (17.4)

Litter incidence

5 (33.3)

10 (71.4)

5 (38.5)

8 (61.5)

Numbers in parentheses indicate percentage of fetuses/litters affected

* significantly different from control; p < 0.05

** significantly different from control; p < 0.01

The examination of external changes showed no malformations or variations.

For fetal soft tissue no malformations were observed.

Variations were seen in all groups including controls. An increased incidence of separated origin of carotids in the 10 mg/L group was statistically significant (p < 0.01). But historical control data showed that the incidence of fetuses showing this variation was in the range of biological variation. The significantly increased occurence of this variation at the highest concentration was therefore assessed to be incidental.

The skeletal examination revealed various malformations of the sternebrae and/or the vertrebral column. Variations seen in skull, the vertrebal column, the sternum and the ribs were found in all groups without apparent concentration relationships or statistically significant differences between the groups.

Any observed differences between groups were within the range of biological variation and/or occurred without a clear concentration dependency.

Applicant's summary and conclusion

Conclusions:
A slight maternal toxicity was seen at the highest exposure (10 mg/L, borderline concentration) in this inhalation study. No signs of developmental toxicity or teratogenicity were noted even with the highest exposure concentration.
Executive summary:

The developmental toxicity of isobutanol (purity 99.8%) was tested in an inhalation study in pregnant Himalayan rabbits. 15 female animals were exposed to vapors of isobutanol at concentrations of 0, 0.5, 2.5 and 10 mg/L for 6 hours/day from day 7 through 19 of gestation. All animals were killed on day 29 of gestation. The fetuses were removed and examined for compound-related effects.

In the high concentration exposure group (10 mg/L) retardation in body weight change was observed especially during the first phase of the exposure. This is an indication of maternal toxicity. The exposure to 10 mg/L seems to be a borderline concentration for causing maternal toxicity. Thus the maternal NOAEL is 2.5 mg/L . 

The fetuses did not exhibit any signs of treatment-related embryo-/fetotoxicity or teratogenic effects at all exposure levels. The developmental NOAEL is 10 mg/L.

(Klimisch 1990).

This inhalation developmental toxicity study in the rabbit is classified acceptable and satisfies the guideline requirements for a developmental toxicity study (OECD 414).