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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 based on grouping of substances (category approach)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant, guideline study, published in peer reviewed literature, fully adequate for assessment
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1987

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
EPA OPP 83-3 (Prenatal Developmental Toxicity Study)
Deviations:
not specified
Principles of method if other than guideline:
not applicable
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 1,3-butadiene
- Physical state: gas
- Analytical purity: 99.88% (by gas chromatography)
- Lot/batch No.: F-909
- Storage condition of test material: Constant temperature of 72°F
- Other: To prevent the appearance of high concentrations of dimer in the 1,3-butadiene atmospheres used for the exposures, cylinder usage was limited to 80% of the net contents, and the acceptable dimer concentration in material sampled from the headspace was specified to be <500 ppm .

Test animals

Species:
mouse
Strain:
CD-1
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratory, Kingston, NY, USA
- Age at delivery: 7-8 weeks
- Housing: 10/sex/cage during the acclimatisation period; 2 females:1 male during overnight pairing; females housed individually during gestation and lactation.
- Diet: NIH-07 open formula diet ad libitum except during exposure
- Water: ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature: 72±3°F
- Humidity: 50±15%
- Air changes (per hr): no data
- Photoperiod: no data

IN-LIFE DATES: From: 29 November 1985 To: 22 December 1985

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Stainless-steel chambers. The total volume of the chamber was 2 .3 M3, and the active mixing volume was 1 .7 M3.
- Method of holding animals in test chamber: There were three levels of caging in each chamber; each level was split into two tiers, which were offset from each other and from the chamber walls. Drawer-like stainless steel cage units, consisting of individual animal cages, were suspended in the space above each tier.
- System of generating particulates/aerosols: For generation of chamber atmospheres, 1,3-butadiene was withdrawn directly from the gas cylinder through a solenoid valve and, subsequently, through a check-valve filter-flow-limit switch and a flow meter, which accurately metered gas to a distribution manifold, where it was initially diluted with filtered air. An air-driven vacuum pump delivered the butadiene-air mixture to the exposure chamber inlet for final dilution to the desired concentration .
- Temperature and humidity, in air chamber: Temperature range 72-76°F; Humidity range 29-68%

TEST ATMOSPHERE
- Brief description of analytical method used: Chamber monitoring of 1,3-butadiene concentrations was performed using an HIP 5840 gas chromatograph equipped with an FID, a Valco (1-ml loop) sampling valve and a Valco stream-select valve capable of sampling eight different sites.
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Eight sites were sampled every 16 minutes: four exposure chambers, two "holding" chambers, the exposure room and the GC standard . Data from the monitor were accumulated by the HIP 85B computer and compared with limit values for the specific sampling location. When the value exceeded the control limits, the HP 85B computer transmitted the information to the executive computer, which initiated an appropriate action to correct the concentration.
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 1:2
- Length of cohabitation: 5 successive nights
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: examination for copulation plug, referred to as day 0 of pregnancy
Duration of treatment / exposure:
Daily, for 6 hours/day
Frequency of treatment:
From 6 through 15 day.
Duration of test:
Until gestation day 18
Doses / concentrations
Remarks:
Doses / Concentrations:
40, 200 and 1000 ppm
Basis:
nominal conc.
No. of animals per sex per dose:
32, 33, 31 and 32 plug-positive females per group (0, 40, 200 and 1000 ppm respectively)
Control animals:
yes, concurrent no treatment
Details on study design:
Females were mated with unexposed males.
Three days prior to the initiation of exposure, the animals were housed in the exposure chambers in the exposure room.
From day 16 until sacrifice at day 18, all animals were housed in exposure chambers with filtered-air atmospheres. The 5 days of mating resulted in staged starts and cessations of exposures. Accordingly, "filler" animals (excess males and females) were used to maintain a constant animal load in the exposure chambers.

Examinations

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

BODY WEIGHT: Yes
- Time schedule for examinations: During the week prior to mating, and on days 0, 6, 11, 16 and 18 of gestation

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

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 18 (following euthanasia with CO2)
- Examined for gross abnormalities: yes
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: Placentas were examined and weighed. Apparently nongravid uteri were stained with ammonium sulfite to detect implantation sites.
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all per litter
- Skeletal examinations: Yes: all per litter
- Head examinations: Yes: approximately half per litter
- Other: Examinations for foetal lens opacities were conducted by removing the eyelid and examining the eye in situ. In addition, the eyeballs were removed for observation under the dissecting microscope.
Statistics:
Analysis of variance (ANOVA; Steel and Torrie, 1980) was used to analyze weight data and, if the result of the analysis was significant, a t-test was performed. Response proportions, such as the number of resorptions, implants, live, dead, or affected foetuses/litter, were also analyzed by ANOVA following arcsin transformation of the response proportion. An orthogonal contrast (Winer, 1971) was used to test trends for dose dependency. In the case of maternal weights, which were repeated over time, analyses for differences among growth curves employed a randomization test (Zerbe, 1979). Binary-response variables between groups were compared, using chi-square or Fischer's exact test (Siegel, 1956). These variables included numbers of pregnant females/number inseminated.
Indices:
none
Historical control data:
Not given

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
A statistically significant reduction in maternal bodyweight gain during gestation was seen at 200 ppm (14% reduction) and 1,000 ppm (20% reduction). Therefore the NOAEL for maternal toxicity was 40 ppm.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEC
Effect level:
40 other: ppm (88 mg/m3)
Basis for effect level:
other: developmental toxicity
Dose descriptor:
NOAEC
Effect level:
40 other: ppm (88 mg/m3)
Basis for effect level:
other: maternal toxicity

Results (fetuses)

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

Details on embryotoxic / teratogenic effects:
Retardation in foetal bodyweight were seen at 1000 and 200 ppm (both sexes) and at 40 ppm in males. The frequency of minor skeletal abnormalities was increased at 200 and 1,000 ppm, concentrations which also produced evidence of maternal toxicity.

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Summary of exposure chamber concentrations

Observation

Chamber concentrations (ppm)

 

40

200

1000

Mean chamber concentration ± SD

39.9 ± 0.6

199.8 ± 3.00

1000 ± 13.1

Mean of target concentration (%)

100

100

100

 

Compared to control values, exposure to 1000 ppm resulted in significantly reduced body weight and extragestational weight at sacrifice and a depression in extragestational gain and weight gain from 11 to 16 days. Significant differences in mice exposed to 200 ppm were confined to weight gain during late exposure and extragestational gain, while values for these two parameters and extragestational weight only tended to be lower in animals exposed to 40 ppm. These results suggest that 1,3-butadiene was toxic to mice at atmospheric concentrations of 200 ppm and higher.

 

Summary of changes in maternal bodyweight (g)

Gestation day

Chamber concentrations (ppm)

 

0 (number=18)

40 (number=19)

200 (number=21)

1000 (number=20)

0 – 6

2.7

3.0

2.5

2.3

6 - 11

5.5

5.8

5.6

4.8

11 – 16

13.3

12.7*

11.4*

10.6*

16 - 18

5.5

5.7

4.7

4.8

E.wt gain

7.6

6.9

6.2

5.9

*statistically significant from control (p≤0.05)

 

E.wt gain: extragestational weight minus bodyweight on day 0 of gestation

The overall percentage of pregnant mice was low (60%), but no differences among treatments were detected in the number of pregnancies or implantation sites/dam. The incidence of early resorptions was higher in control mice than in animals exposed to 200 ppm, but no effect was observed on the percentageof total resorptions and live foetuses/litter.

Depressed foetal body weights were seen at 200 and 1000 ppm in both sexes and male foetuses were affected at 40 ppm, a level at which there were no clear signs of maternal toxicity. Significant decreases in body weights of females occurred at the next higher dose level (200 ppm). Differences in the depression of placental weights were also associated with the sex of the foetuses, ie ., placentas of male foetuses were significantly affected at 200 ppm and those of female foetuses at 1000 ppm.

 

Foetal and placental measures for mouse litters

 

1,3-butadiene concentration (ppm)

Observation

0

40

200

1000

No. Litters

18

19

21

20

No. Foetuses

211

237

259

244

Bodyweight (g)

1.34 ± 0.03

1.28 ± 0.01

1.13 ± 0.02*

1.04 ± 0.03*

Females

1.30 ± 0.02

1.25 ± 0.01

1.10 ± 0.02*

1.02 ± 0.03*

Males

1.38 ± 0.03

1.31 ± 0.02*

1.13 ± 0.02*

1.06 ± 0.02*

Sex ratio: % males

51 6 ± 3.91

49.8 ± 3.06

51.5 ± 3.68

51.8 ± 3.29

Placental weight mg)

86.8 ± 2.99

85.4 ± 2.29*

78.6 ± 3.24*

72.6 ± 1.88*

Females

83.1 ± 3.03

80.9 ± 2.46

74.7 ± 3.52*

70.1 ± 2.33*

Males

89.3 ± 3.05

89.5 ± 2.27

80.1 ± 2.35*

74.5 ± 1.81*

 

The incidence of malformations, which included exencephalus, hydrocephalus, cleft palate and thoracogastroschisis, was not significantly different among treatment groups.No lens opacities of the foetal eye were detected during examination. However, incidences of foetal variations (supernumerary ribs and reduced ossification of the sternebrae) were significantly increased in litters from mice exposed to 200 and 1000 ppm.

 

Mean % of reduced ossification, supernumery ribs and abnormal sternebrae per litter

 

buta-1,3-diene concentration (ppm)

Observation

0

40

200

1000

reduced ossification (a)

1.7 ± 1.7

1.2 ± 1.5

2.7 ± 2.7

3.9 ± 2.6*

supernumery ribs

1.7 ± 2.3

1.6 ± 2.1

6.0 ± 3.6 b

9.9 ± 3.0*

abnormal sternebrae

0.6 ± 0.9

0.4 ± 0.7

0.4 ± 0.8

0.8 ± 1.3*

(a) - all sites of ossification combined

b - misaligned, scrambled or cleft sternebrae

* - Significantly greater than control (p<0.05)

 

Applicant's summary and conclusion

Conclusions:
Exposure of mice to 1,3-butadienee (up to 1000 ppm, 2212 mg/m3) from days 6-15 of gestation in a pre-natal developmental toxicity study resulted in some fetotoxicity at maternally toxic doses. There was no evidence of teratogenicity in the absence of maternal toxicity.
Executive summary:

CD1 mice were exposed to 1,3-butadiene from days 6-15 of gestation at concentrations of 40, 200 or 1000 ppm (88, 442 or 2212 mg/m3) 6h/day, in a pre-natal developmental toxicity study. 1,3 -Butadiene produced significant signs of maternal toxicity (reduced body weight gain) at concentrations of 200 and 1000 ppm buta-1,3-diene. The NOAEC for maternal toxicity was 40 ppm (88 mg/m3). Foetal growth retardation, decreased placental weights, and increased incidences of morphologic variations occurred at 200 and 1000 ppm (increased percentages of supernumerary ribs per litter at 200 ppm, and increased percentages of reduced ossification sites and abnormal sternebrae at 1000 ppm) and were attributed to the maternal toxicity. The original authors described fetotoxicity in male mice at 40 ppm (a non-maternally toxic exposure level) and therefore no NOAEC was established for developmental toxicity. Subsequent re-analyses, however, demonstrated that the original analysis of foetal toxicity was incorrect and established a NOAEC for foetal toxicity of 40 ppm and therefore 40 ppm (88 mg/m3) as the overall NOAEC in this study.