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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1996
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Conducted according OECD TG 471. GLP.

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1996

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
CAS #3319-31-1; 1,2,4-Benzenetricarboxylic acid, tris(2-ethylhexyl) ester

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
S9
Test concentrations with justification for top dose:
without S9: 0, 313, 625, 1250, 2500, 5000 ug/plate (five strains)
with S9: 0, 313, 625, 1250, 2500, 5000 ug/plate (five strains)
Vehicle / solvent:
Acetone
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Remarks:
Acetone, DMSO
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: Without S9 mix, 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (TA100, WP2, TA98) Sodium azide (TA1535) 9-Aminoacridine (TA 1537); With S9 mix, 2- Aminoanthracene (five strains)
Details on test system and experimental conditions:
Prior to assay initiation, a toxicity pretest was performed using tester strain TA100. Based on these results, the doses for the final assay were determined. Each chemical was tested initially at half-log dose intervals up to a dose that elicited toxicity, or to a dose immediately below one which was toxic in the preliminary toxicity test. Chemicals that were not toxic were tested, with few exceptions, to a maximum dose of 10 mg/plate. In the definitive assay, each of the five strains was dosed with either the test substance; a vehicle control (DMSO); or a nontreated control and a positive control. The test mixture containing the tester strain and test substance with or without S9 was added to the surface of petri dishes containing Vogel-Bonner medium. The S-9 (9,000g supernatant) fractions of Aroclor 1254-induced, male Sprague-Dawley rat and male Syrian hamster livers were prepared as described previously [Haworth et al, 1983]. The S-9 mixes were prepared immediately prior to use and contained either 10% or 30% S-9; occasionally, other levels were used. The histidine-independent colonies that formed on the plates were counted following a two-day incubation at 37°C. Positive controls were as follows: 2 aminoanthracene (all strains with S9); sodium azide (without S9, TA1535, TA100), 4-nitro-ophenylenediamine (without S9, TA98) and 9-aminoacridine (without S9, TA 97, TA1537). There were 3 plates/dose group/strain/treatment. The test results were verified by repeating the assay. If the results were negative, they were repeated first without S9 and then with 30% S9.

Species/Strain: Salmonella typhimurium TA100, TA1535, TA98, TA1537; Escherichia coli WP2 uvrA

Positive controls:
Without S9 mix, 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (TA100, WP2, TA98) Sodium azide (TA1535)
9-Aminoacridine (TA 1537)
With S9 mix, 20 Aminoanthracene (five strains)

S9: Rat liver, induced with phenobarbital and 5,6-benzoflavone
Evaluation criteria:
Chemicals were judged to be mutagenic if the test results produced a dose-related, reproducible increase in histidine revertants over control. It was not a requirement for mutagenic responses to reach two-fold over background.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Remarks:
no genotoxic effects were observed at the highest dose tested (5000 ug/plate)
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Remarks:
Toxicity was not observed up to 5000 ug/plate in five strains with and without metabolic activation (S9 mix).
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Remarks:
no genotoxic effects were observed at the highest dose tested (5000 ug/plate)
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Remarks:
Toxicity was not observed up to 5000 ug/plate in five strains with and without metabolic activation (S9 mix).
Vehicle controls validity:
valid
Untreated negative controls validity:
not applicable
Positive controls validity:
valid
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative with metabolic activation at the highest dose tested (5000 ug/plate)
negative without metabolic activation at the highest dose tested (5000 ug/plate)

No genotoxic effects were observed at the highest dose tested (5000 ug/plate) in all five strains of bacteria, with or without metabolic activation.

Classification as a genotoxic agent is not warranted under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP) or under Directive 67/548/EEC for dangerous substances and Directive 1999/45/EC for preparations.
Executive summary:

The test material did not induce a statistically signifiant increase in revertant numbers in any of the test strains, either in the absence or in the presence of S-9. This study was therefore considered to have provided no indication of any test material mutagenic activity.

Classification as a genotoxic agent is not warranted under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP) or under Directive 67/548/EEC for dangerous substances and Directive 1999/45/EC for preparations.