Aromatic hydrocarbons, C6-8, naphtha-raffinate pyrolyzate-derived

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration

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: Not GLP, but key cytogenetic parameters measured comparable to guideline study.

Data source

Reference

Reference Type:

publication

Title:

Dose-related clastogenic effects induced by benzene in bone marrow cells and in differentiating spermatogonia of Swiss CD1 mice.

Author:

Ciranni R, Barale R and Adler I-D.

Year:

1991

Bibliographic source:

Mutagenesis 6 (5), 417-421

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)

Deviations:

yes

Remarks:

only 3-4 animals/group

Principles of method if other than guideline:

Mice given single oral gavage dose of benzene (1 mL/kg bw) and chromosomal aberrations in bone marrow and spermatogonal cells assessed at time points up to 48 h post-treatment.

GLP compliance:

not specified

Type of assay:

other: bone marrow chromosome aberration assay and mammalian germ cell cytogenetic assay

Test material

Test material information

Constituent 1

Reference substance name:

Benzene

EC Number:

200-753-7

EC Name:

Benzene

Cas Number:

71-43-2

Molecular formula:

C6H6

IUPAC Name:

benzene

Details on test material:

- Name of test material (as cited in study report): benzene
- Source: Farmitalia, Carlo Erba, Italy

Test animals

Species:

mouse

Strain:

CD-1

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Valco Co., Italy
- Age at study initiation: 2 months
- Weight at study initiation: 30-35 g
- no further details

ENVIRONMENTAL CONDITIONS
- no data

IN-LIFE DATES:
- no data

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Olive oil

Duration of treatment / exposure:

Single oral dose

Frequency of treatment:

Single oral dose

Post exposure period:

Up to 48 h

Doses / concentrations

Remarks:

Doses / Concentrations:
1 mL/kg
Basis:
actual ingested

No. of animals per sex per dose:

3-4 male mice

Control animals:

yes, concurrent vehicle

Examinations

Tissues and cell types examined:

Bone marrow and spermatogonal cells

Statistics:

The binomial dispersion test was applied to test homogeneity of results from control animals. Statistical differences between treated and solvent control groups were determined by Fisher's exact test. To compare the sensitivity of the two cell types the doubling doses were calculated. The doubling dose (DD) is defined as the dose that induces as many aberrations as occur spontaneously per cell generation. Based on the linear dose-response, Y = a + b D, where Y is the yield of aberrant cells, a the spontaneous frequency and b the linear regression coefficient, the doubling dose is calculated as the ratio of a to b (DD = a/b).

Results and discussion

Test results

Test results 1

Sex:

male

Genotoxicity:

positive

Remarks:

Chromosomal aberration test; mouse (bone marrow)

Toxicity:

not specified

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

not applicable

Test results 2

Sex:

male

Genotoxicity:

positive

Remarks:

Germ cell chromosome aberration test; mouse (spermatogonia)

Toxicity:

not specified

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

not valid

Any other information on results incl. tables

Bone Marrow:

Benzene showed high clastogenic activity in bone marrow cells at all sampling times with a peak at 24 and 30 hours (approx. 20% aberrant cells (excl. gaps) versus 1% in controls). The dose-response was determined 24 h after treatment with 0.1, 0.5 or 1.0 mL/kg benzene (equivalent to 88, 440 and 880 mg/kg bw). All three doses were clearly positive and a dose-dependency was established.

 

Chromatid aberrations in bone marrow cells of mice treated with 1 mL/kg (880 mg/kg) of benzene: time-response

Time (h)	No. cells scored a	No. of aberrations			Highly damaged cells (n) b	Aberrant cells (%±SE)
		gaps	breaks	exchanges		including gaps	excluding gaps
Control	1000	24	11	-	-	3.2 ± 0.6	1.1 ± 0.1
6	600	47	29	-	-	11.3 ± 0.1	4.3 ± 0.6**
12	600	89	98	7	-	24.6 ± 2.4	16.6 ± 0.4**
18	600	59	112	7	3	22.6 ± 4.5	15.3 ± 3.0**
24	600	53	206	6	13	25.1 ± 2.7	20.8 ± 3.3**
30	600	117	237	15	13	28.7 ± 7.4	19.8 ± 0.5**
36	600	10	24	-	-	4.8 ± 0.4	3.5 ± 0.5**
42	600	26	31	-	-	8.6 ± 1.4	5.3 ± 0.4**
48	600	41	44	-	-	12.0 ±1.7	6.3 ± 1.1**

**P <0.01 (Fisher's exact test).

a  200 cells scored per animal.

b  Cells with more than 10 aberrations.

 

Spermatogonia:

After administration of 1 mL/kg (880 mg/kg bw) the maximum response was obtained 24 h after treatment (6.3% aberrant cells versus 1.2% in negative controls). In the dose-response study, all doses tested (0.25, 0.5 and 1.0 mL/kg bw, equivalent to 220, 440 and 880 mg/kg bw) increased the aberration frequency in a dose-dependent manner; at 880 mg/kg again 6.3% of the spermatogonia were aberrant. Since bone marrow clastogenicity was investigated in parallel, it can be concluded that clastogenicity in bone marrow cells and spermatogonia was induced in the same dose range, although effects were less pronounced in spermatogonia.

 

Chromatid aberrations in differentiating spermatogonia of mice treated with 1 mL/kg (880 mg/kg) of benzene: time-response

Time (h)	No. cells scored a	No. of aberrations			Aberrant cells (%±SE)
		gaps	breaks	exchanges	including gaps	excluding gaps
Control	1000	49	12	-	5.5 ± 0.9	1.12 ± 0.2
6	600	32	6	-	6.3 ± 0.1	1.0 ± 0.3
12	600	63	18	1	12.3 ± 0.3	3.3 ± 0.4**
18	600	67	24	-	13.3 ± 1.2	4.0 ± 0.5**
24	600	54	36	3	14.8 ± 2.6	6.3 ± 1.6**
30	600	33	16	1	7.8 ± 1.6	2.6 ± 0.5*
36	600	31	23	1	8.3 ± 1.0	3.5 ± 0.5**
42	600	21	14	-	5.2 ± 1.3	2.0 ± 0.5
48	600	19	23	-	18.2 ±2.0	3.5 ± 0.7**

**P <0.01 (Fisher's exact test).

a  200 cells scored per animal.

 

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): positive Chromosomal aberration test (mouse bone marrow) and Germ cell chromosome aberration test (mouse spermatogonia)
Benzene was positive in the chromosomal aberration test (mouse bone marrow) and germ cell chromosome aberration test (mouse spermatogonia), following a single oral dose of 1 mL/kg to male mice.

Executive summary:

The ability of benzene to induce chromosome damage in vivo was assessed by determining the frequencies of chromosomal aberrations in bone marrow and spermatogonial cells of male Swiss CDI mice. Initially a single dose of 1 mL benzene/kg (880 mg/kg) was assessed using a wide range of times (6, 12, 18, 24, 30, 36, 42 and 48 hours) to determine the time of maximum response. Benzene showed high clastogenic activity with a peak between 24 and 30 hours in bone marrow cells or 24 hours in differentiating spermatogonia. The effect in bone marrow cells was greater than in spermatogonia. Secondly, the dose response 24 hours after treatment was determined. Additional doses of benzene used were: 0.1 mL/kg (88 mg/kg) and 0.5 mL/kg (440 mg/kg) for bone marrow cells; 0.25 mL/kg (220 mg/kg) and 0.5 mL/kg (440 mg/kg) for differentiating spermatogonia.

Benzene was positive in this test with dose dependent clastogenic effects in both cell types. All dose levels showing a statistically significant increase in the incidence of aberrant cells.

It is concluded that benzene is a clastogen in male germ cells and the bone marrow of mice.