Benzene, C10-13-alkyl derivs.

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1992

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5265 (The Salmonella typhimurium Bacterial Reverse Mutation Test)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Alkylate 215

IUPAC Name:

Alkylate 215

Details on test material:

- Composition of test material, percentage of components: <1% C9, 22% C10, 43% C11, 35% C12, 1% C13, <1% C14; average C11.26
- Analytical purity: 98.5%

Method

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Metabolic activation system:

S9 from Aroclor induced rat and mouse livers

Test concentrations with justification for top dose:

10, 40, 200, 1000, 3000, and 10000 μg/plate

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-acetylaminofluorene

9-aminoacridine

2-nitrofluorene

benzo(a)pyrene

other: sodium nitrite

Details on test system and experimental conditions:

METHOD OF APPLICATION:
- in agar (plate incorporation)

Evaluation criteria:

A response was considered positive if three or more treatments were significantly greater than controls, and if there was a significant dose response.

Statistics:

Significant differences analyzed using Bartlett's test. Comparison to controls analyzed using Dunnett's t test. Dose response analyzed with regression analysis for a log-log straight line and t-test.

Results and discussion

Test results

Key result

Species / strain:

S. typhimurium, other: TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

Negative with and without activation. A significant difference was seen in strain TA 98 with S9 at all concentrations. There was also a significant response in strain TA 1535 without S9 at a single concentration. Retesting with strain TA 98 showed no significant increase in revertants. A retest of strain TA 1535 was also negative. Due to lack of a dose-response relationship and lack of reproducibility of the positive results, the test substance is considered negative for mutagenicity.

Any other information on results incl. tables

Table 1: Mean Revertants/plate - without S9

Concentration (μg or nl/plate)	TA 1535	TA 100	TA 1537	TA 98
First run
10	28.3 ± 3.51	307 ± 51.5	8.0 ± 1.0	25.7 ± 5.13
40	23.0 ± 2.65	344 ± 33.5	9.0 ± 3.6	19.3 ± 5.51
200	29.0 ± 8.00	310 ± 17.6	10.7 ± 0.58	17.3 ± 3.79
1000	23.3 ± 3.21	318 ± 33.3	10.0 ± 1.0	18.7 ± 1.15
3000	34.7 ± 5.03	296 ± 57.4	9.0 ± 1.7	21.7 ± 2.08
10,000	27.3 ± 4.62	297 ± 59.2	10.3 ± 1.5	27.3 ± 6.03
Solvent control	21.7 ± 0.58	395 ± 30.8	7.3 ± 3.5	23.7 ± 0.58
NaNO2	691
9-aminoacridine			1620
2-nitrofluorene		627		354
Second run
10		321 ± 17.7
40		297 ± 25.1
200		292 ± 39.0
1000		307 ± 20.3
1500	16.0 ± 1.0
3000	14.3 ± 0.58	322 ± 58.2
4000	12.0 ± 4.0
10,000		322 ± 33.2
Solvent control	13.0 ± 3.46	258 ± 25.5
NaNO2	564
2-nitrofluorene		814
Third run
10		245 ± 11.5
40		236 ± 30.7
200		166 ± 57.6
1000		203 ± 49.2
3000		230 ± 11.8
10,000		179 ± 22.3
Solvent control		233 ± 27.0
2-nitrofluorene		506

Table 2: Mean Revertants/plate - with S9

Concentration (μg or nl/plate)	TA 1535	TA 100	TA 1537	TA 98
First run
10	25.7 ± 5.8	309 ± 44.9	11.3 ± 1.5	72.7 ± 5.8
40	21.3 ± 5.8	294 ± 31.0	10.3 ± 2.3	79.3 ± 5.5
200	20.3 ± 1.2	283 ± 12.7	7.3 ± 2.5	63.7 ± 13.9
1000	24.3 ± 2.1	339 ± 38.4	6.7 ± 1.5	71.7 ± 16.0
3000	22.7 ± 3.8	303 ± 27.4	13.7 ± 2.5	71.7 ± 4.2
10,000	28.7 ± 4.9	367 ± 62.4	12.3 ± 3.2	70.7 ± 0.53
Solvent control	24.3 ± 1.5	314 ± 20.3	9.7 ± 2.1	29.0 ± 9.5
B(a)P		938		459
2-AA	661		155
Second run
10	15.7 ± 3.2	280 ± 46.6		27.3 ± 2.5
40	11.0 ± 3.6	299 ± 15.0		33.7 ± 4.9
200	15.3 ± 2.1	294 ± 24.4		39.0 ± 1.0
1000	21.3 ± 3.1	297 ± 26.8		30.0 ± 7.2
3000	21.0 ± 11.5	301 ± 39.9		32.0 ± 7.2
10,000	13.7 ± 2.5	296 ± 6.8		36.0 ± 4.6
Solvent control	13.7 ± 5.7	304 ± 29.0		27.0 ± 1.7
B(a)P		1611
2-AA	511			896
Third run
10		191 ± 43.0
40		210 ± 3.06
200		227 ± 15.0		39.0 ± 6.2
250				35.3 ± 9.3
1000		229 ± 12.9		32.0 ± 5.3
3000		238 ± 29.7
10,000		204 ± 12.2
Solvent control		219 ± 39.8		34.7 ± 3.2
B(a)P		1649		810

Applicant's summary and conclusion

Conclusions:

Under the conditions described in this study Alkylate 215 did not show mutagenic properties.

Executive summary:

This EPA OPPTS 870.5265 (The Salmonella typhimurium Bacterial Reverse Mutation Test) and GLP compliant study was performed to investigate the mutagenicity of the test substance alkylate 215. Salmonella typhimurium strains TA 1535, TA 100, TA 1537, and TA 98 were exposed to concentrations of 10, 40, 200, 1,000, 3,000, and 10,000 μg/plate of test substance. DMSO was used as a solvent. 9-aminoacridine, sodium nitrite, benzo(a)pyrene, 2 -acetylaminofluorene, and 2 -nitrofluorene were used as positive controls substances. No reproducible, dose-response related positive results were seen in the treatment groups. A significant increase in revertants per plate was seen in the positive controls. The test substance is not mutagenic.