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Toxicological information

Genetic toxicity: in vitro

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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:
other: test in accordance with recognised standards, with acceptable restricition: not GLP, and no data on potential cytotoxicity at highest concentrations than 1000 µg:plate for 3 of the tested strains. In addition, no positive controls included in this test

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1977

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
The test was performed according to the method of Ames et al. (1975).
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Method

Target gene:
Histidine operon
Species / strain
Species / strain / cell type:
S. typhimurium, other: TA 1535, TA 1537, TA 1598, TA 98, TA 100
Metabolic activation:
with and without
Metabolic activation system:
: Rat S9 mix. Liver S9 homogenate was prepared from rats that have been induced with Arochlor 1254
Test concentrations with justification for top dose:
0, 4, 20, 100, 500 and 1000 µg/plate
Vehicle / solvent:
- Solvent used: DMSO
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
other: not in this test. The reversion properties of each strain are regularly checked
Positive control substance:
other: Methylmethanesulfonate, 4-aminobiphenyl, 9-aminoacridine, N-mehtyl-N-nitro-N-nitrosoguanidine
Details on test system and experimental conditions:


METHOD OF APPLICATION:

The procedures used in this mutagenicity assay are described in detail by Ames et al. (1975). Briefly the procedure was as follows:
to 2.5 ml molten soft agar were added 0.1 ml of a fully grown culture of one of the tester strains, 0.1 ml of an appropriate dilution/suspension of the test compound and the liver microsome system if indicated.
The ingredients were thoroughly mixed and immediately poured onto minimal glucose agar plates. After the top agar had been allowed to harden, the plates were incubated at 370c for three days. Then the colonies (revertants which are histidine­ independent) were counted, and the background lawn of bacterial growth examined.
Based on the results of preliminary toxicity tests, the test materials were examined at levels up to 1000 µg/plate. All determinations were carried out in triplicate and appropriate controls were included in each assay.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium, other: TA 1535, TA 1537, TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Species / strain:
S. typhimurium, other: TA 1538, TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: yes, at the highest concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid

Any other information on results incl. tables

Table 1: Number of revertants per plate (mean of 3 plates)
  TA 1535 TA 1537 TA 1538 TA 98 TA 100
Conc. [unit] - MA +MA - MA + MA - MA + MA - MA + MA - MA + MA
0  25 17  12  15   11 14   16  20 61   54
 4  30 13  20  12   13 8 17  22  47   62
 20  24 23  11  24   17  14  20 21   55 54 
 100  19  14 13   15  10  10  14  26 45  69 
 500  23  17  13 14 * PP   17 24  47  54 
1000   13 13   11  12 11 * PP  19   35  PP PP 

*: background lawn of bacterial growth less dense than in control plates

PP: background lawn of bacterial growth sparse or absent, plates crowded with small his- colonies

Applicant's summary and conclusion

Conclusions:
tert-butyl cumyl peroxide did not show any mutagenic activity in the bacterial reverse mutation test with Salmonella typhimurium.
Executive summary:

The potential of the test item (tert-butyl cumyl peroxide at a 90 -98 % purity) to induce reverse mutation in Salmonella typhimurium (strains: TA 1535, TA 1537, TA 1538, TA 98 and TA 100) was evaluated in an Ames test.

The test item did not induce any noteworthy increase in the number of revertants, both with and without S9 mix, in any of the five strains.

Incorporation of 4 µg up to non-inhibitory amounts of the test material per plate (i.e. 20-1000 µg) did not induce an increase in the number of his+ revert plates in any of the tester strains, either in the presence or in the absence of the S-9 mix.

 

At 1evels of 500 and/or 1000 µg/ plate the test substance had a clearly toxic effect on the strains TA 1538 and TA 100. At 1000 µg/plate, the test substance had no toxic effects on strains TA 1535, TA 1537 and TA98.

Under our experimental conditions, tert-butyl cumyl peroxide did not show any mutagenic activity in the bacterial reverse mutation test with Salmonella typhimurium.