Solvent naphtha (petroleum), heavy arom.

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

migrated information: read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-GLP, non-guideline, animal experimental study, restrictions in design and/or reporting but otherwise adequate for assessment

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

publication

Title:

Mutagenicity of 1,3-butadiene inhalation in somatic and germinal cells of mice.

Author:

Adler I-D, Cao J, Filser JG, Gassner P, Kessler W, Kliesch U, Neuhäuser-Klaus A, Nüsse M.

Year:

1994

Bibliographic source:

Mutat Res. 309; 307-314.

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

not specified

Principles of method if other than guideline:

not applicable

GLP compliance:

not specified

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

1,3-butadiene

IUPAC Name:

1,3-butadiene

Details on test material:

- Name of test material (as cited in study report): 1,3-butadiene
- Purity: 99.5%
- Source: Linde, Unterschleissheim, Germany

Test animals

Species:

mouse

Strain:

other: 102/E1 x C3H/E1)F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: GSF animal colony
- Age at study initiation: 10-12 weeks
- Weight at study initiation: The average weight of groups of 20 animals before exposure was: 26.7 ± 1.0 g(control), 25.6 ± 0.9 g (50 ppm), 26.4 ± 1.0 g (200 ppm), 25.6 ± 0.8 g (500 ppm) and 26.1 ± 1.0 g (1300 ppm).
- Housing: 10 sex/group
- Diet: Altronin ad libitum except for during, and for 1 hour following exposure
- Water: ad libitum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS IN CHAMBERS
- The chambers were supplied with a stream (80 1/min) of charcoal-filtered and climatized air (25°C, 55% moisture)

IN-LIFE DATES: no data

Administration / exposure

Route of administration:

inhalation: gas

Vehicle:

air

Details on exposure:

TYPE OF INHALATION EXPOSURE: whole body

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: glass chambers (240 L) with room for two stainless steel wire cages (65 x 40 X 12 cm).
- System of generating test atmosphere: 1,3-butadiene was metered at a known and constant flow rate into the air stream to achieve the desired exposure concentration.

TEST ATMOSPHERE
- Brief description of analytical method used: Septa for air sampling were positioned 30 cm before the inlet and behind the outlet of the chamber. The atmosphere was analyzed for 1,3-butadiene concentration at 15-min intervals during the first hour of daily exposure and at 30-min intervals thereafter by gas chromatography.
- Samples taken from breathing zone: yes

Duration of treatment / exposure:

6 h per day for 5 consecutive days

Frequency of treatment:

Daily

Post exposure period:

None

Doses / concentrations

Remarks:

Doses / Concentrations:
50, 200, 500 or 1300 ppm
Basis:
nominal conc.

No. of animals per sex per dose:

Bone marrow micronucleus test: 5 per sex
Peripheral blood micronucleus test: 2 per sex

Control animals:

yes, concurrent no treatment

Positive control(s):

none

Examinations

Tissues and cell types examined:

Bone marrow and peripheral blood

Details of tissue and slide preparation:

BONE MARROW MICRONUCLEUS TEST
TREATMENT AND SAMPLING TIMES:
- Bone marrow was sampled 18-24 h after the end of exposure.

DETAILS OF SLIDE PREPARATION:
- Bone marrow preparation, staining and scoring were performed (Adler, 1984).

METHOD OF ANALYSIS:
- Per animal 2000 polychromatic erythrocytes (PCE) were scored microscopically for the presence of micronuclei and the means were expressed as micronucleated PCE (MPE) per 1000 PCE.
- To determine a shift in erythroblast proliferation the number of PCE was counted in the microscope fields that contained 2000 normochromatic erythrocytes (NCE) and the mean values are expressed as % PCE of the total erythrocyte counts. Micronucleated NCE were also recorded.

PERIPHERAL BLOOD MICRONUCLEUS TEST
TREATMENT AND SAMPLING TIMES:
- Blood (30-50 µL) was collected from the orbital vein of mice into heparinized tubes
- The samples were taken 18-24h after the end of the exposure to 0, 50, 200 and 1300 ppm of 1,3-butadiene.

METHOD OF ANALYSIS:
- A sample of 5 µL of blood was used for two measurements per animal. Dual laser flow cytometry and sorting was performed.

SCORING:
- Microscopic scoring: For comparison, the blood samples were also scored microscopically using the acridine orange supravital staining method of Hayashi et al. (1990). For each animal, 1000 type I, II and III PCE (Vander et al 1963) were scored for the presence of micronuclei. Samples from exposure groups, 0, 50, 200 and 1300 ppm were analyzed.

Evaluation criteria:

see below

Statistics:

Significance of differences between exposure groups was determined with the Mann-Whitney test (Sachs, 1974). The dose response was determined by regression analysis.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

positive

Remarks:

Bone marrow micronucleus

Additional information on results:

none

Any other information on results incl. tables

The buta-1,3-diene chamber concentrations did not deviate more than 12% at 50 ppm, 1% at 200 ppm, 0.2% at 500 ppm and 2% at 1300 ppm from the nominal concentration throughout the exposure duration.

The micronucleus frequency in bone marrow PCE increased with exposure concentration. The lowest exposure concentration caused a significant increase over the control level (p < 0.01, Mann-Whitney test). The slope of the dose-response curve did not remain constant but flattened with increasing concentration. In the concentration range between 500 and 1300 ppm only a small rise of the micronucleus frequency of about 17% was found. At 500 and 1300 ppm the response of the males was higher than that of the exposed females (p < 0.05, Mann-Whitney test). 1,3-Butadiene exposure had no effect on erythroblast proliferation.

Results of bone marrow micronucleus test

Concentration (ppm)	Individual animal counts MPE/2000 PCE		Mean MPE (% ±SD)	PCE (%)
	Male	Female
0	1, 1, 3, 4, 6	1, 1, 1, 1, 6	1.3 ± 0.6	49.0
50	8, 9, 10, 12, 17	8, 9, 10, 10, 14	5.4 ± 1.0	49.6
200	19, 21, 24, 26, 27	13, 16, 18, 20, 25	10.5 ± 1.4	49.1
500	35, 35, 36, 42, 44	24, 24, 30, 31, 32	16.7 ± 1.8*	49.7
1300	39, 40, 43, 46, 53	28, 32, 33, 37, 39	19.5 ± 1.6*	49.6

*The male response was significantly higher than the female response (p<0.05)

The micronucleus frequency in PCE of peripheral blood also increased with exposure concentration. When scored manually the same sensitivity difference as in the bone marrow micronucleus test between males and females was obvious at 200 and 1300 ppm. The dose-response curve had approximately the same shape. There was no difference between the micronucleus yields in bone marrow and peripheral blood, the latter scored either manually or by flow cytometry.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): positive
The data demonstrate that Inhalation exposure of mice to 1,3-butadiene (50, 200, 500 or 1300 ppm for 6 h per day for 5 days) induced micronuclei in bone marrow and peripheral blood, with male mice more sensitive than females at the higher exposure concentrations. The dose-response curve is consistent with the metabolism of 1,3-butadiene to epoxides, which may then induce micronuclei.

Executive summary:

Inhalation exposure of mice to 50, 200, 500 or 1300 ppm (110, 442, 1106 or 2876 mg/m3) of 1,3 -butadiene for 6 h per day for 5 consecutive days caused micronuclei in mouse bone marrow and peripheral blood erythrocytes. The dose response was non-linear. The slope of the curve flattened with increasing exposure concentration.