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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

For this endpoint information from three different assays is available. The data were obtained in the following assays: 1) Bacterial in vitro mutagenicity 2) Mammalian cell mutation assay3) Chromosomal abberation assayAll these data have been obtained acording to the relevant OECD protocols.

Link to relevant study records
Reference
Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Oct 10- 19, 2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
The study was performed in compliance with the Good Laboratory Practice (GLP) regulations (revised in 1997, ENV/MC/CHEM(98)17).The method followed that described in the OECD Guidelines for Testing of Chemicals (Adopted: 4 April 1984) No 471 "Bacterial Reverse Mutation Test".
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
Principles of method if other than guideline:
none
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay
Target gene:
HIS operon (S. thyphimurium)TRY operon (E. coli)
Species / strain / cell type:
S. typhimurium TA 1535
Details on mammalian cell type (if applicable):
his G 46, uvrB, rfa
Additional strain / cell type characteristics:
other: mutations in the histidine operon
Species / strain / cell type:
S. typhimurium TA 1537
Details on mammalian cell type (if applicable):
his C 3076, uvrB, rfa
Additional strain / cell type characteristics:
other: mutations in the histidine operon
Species / strain / cell type:
S. typhimurium TA 98
Details on mammalian cell type (if applicable):
his D 3052, uvrB, rfa + R-factor
Additional strain / cell type characteristics:
other: mutations in the histidine operon
Species / strain / cell type:
S. typhimurium TA 100
Details on mammalian cell type (if applicable):
his G 46, uvrB, rfa + R-factor
Additional strain / cell type characteristics:
other: mutations in the histidine operon
Species / strain / cell type:
S. typhimurium TA 102
Details on mammalian cell type (if applicable):
his G 428, rfa + R-factor
Additional strain / cell type characteristics:
other: mutations in the histidine operon
Species / strain / cell type:
E. coli WP2
Details on mammalian cell type (if applicable):
trp C 3076, uvrB, rfa
Additional strain / cell type characteristics:
other: mutations in the tryptophan operon
Metabolic activation:
with and without
Metabolic activation system:
rat liver homogenate (S9 mix) with standard co-factors with metabolic activation (Aroclor)
Test concentrations with justification for top dose:
The test material concentrations were used selected according to the EC and OECD guidelines for this test system and the requirements of the Labor Ministry of Japan: 5-5000 µg/plate have been used in the 1st and 2nd series respectively.
Vehicle / solvent:
Acetone
Positive controls:
yes
Positive control substance:
cumene hydroperoxide
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
N-ethyl-N-nitro-N-nitrosoguanidine
Positive controls:
yes
Positive control substance:
9-aminoacridine
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
Remarks:
with S9
Positive controls:
yes
Positive control substance:
other: daunomycine
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
with S9
Details on test system and experimental conditions:
The assessment of test material-induced effects is dependent on the number of spontaneous revertants of each bacterial strain (solvent controls) and the increase in the number of revertants at the test material concentration which shows the highest number of colonies. The following criteria, based upon the historical controls of the laboratory and statistical considerations, are established:-----------------------------------------------------------------------------------------Mean Number of Colonies Maximal Mean Number of Colonies over the Actual(Solvent Control) Solvent Control (Test Material)-----------------------------------------------------------------------------------------<=10 <=9 >=30<=30 <=19 >=40<=80 <=29 >=80<=200 <=49 >=120<=500 <=79 >=200Assessment No increase Clear increase-----------------------------------------------------------------------------------------All further results, ranging between "no" and "clear", are assessed as "weak in-creases".Interpretations:A test material is defined as non-mutagenic in this assay if "no" or "weak increases" occur in the first and second series of the main experiment. ("Weak increases" randomly occur due to experimental variation.)A test material is defined as mutagenic in this assay if: - a dose-related (over at least two test material concentrations) increase in the number of revertants is induced, the maximal effect is a "clear increase", and the effects are reproduced at similar concentration levels in the same test system;- "clear increases" occur at least at one test material concentration, higher concentrations show strong precipitation or cytotoxicity, and the effects are reproduced at the same concentration level in the same test system.In all further cases, a third test series with the bacterial strain in question should be performed. If the criteria for a positive test result are not fulfilled in at least two out of the three series, the test material is defined as being non-mutagenic in this test system.
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium 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
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Precipitation occured at concentrations or equal 500 µg/plate. Toxicity to bacteria was not observed.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Conclusions:
Interpretation of results (migrated information):negative with and without metabolic activationWith and without addition of S9 mix as the external metabolizing system, the test item was not mutagenic under the experimental conditions described.
Executive summary:

Purpose

The purpose of this in vitro reverse gene mutation test is to identify agents that cause mutations in bacteria cells thus providing information on possible health hazards for the test material and serve as a rational basis for risk assessment to the genotoxic potential of the test item in man.

Study Design

The investigations for mutagenic potential were performed using Salmonella typhimurium tester strains TA 98, TA 100, TA 102, TA 1535 and TA 1537, and Escherichia coli WP2 uvrA pkM101. The plate incorporation test with and without addition of liver S9 mix from Aroclor 1254-pretreated rats was used. Two independent experimental series were performed for all strains. In the series with S9 mix, 10 % or 30 % S9 in the S9 mix were used inn the 1st and 2nd series, respectively.

Results

The test material was dissolved in acetone and tested at concentrations ranging from 5.00 to 5000 pg/plate. Precipitation of the test material on the agar plates occurred at concentrations larger/equal 500 µg/plate. Toxicity to the bacteria was not observed.

Daunomycin, N-ethyl-N'-nitro-N-nitroso-guanidine, 9-aminoacridine and cumene hydroperoxide served as strain specific positive control compounds in the absence of S9 mix. 2-Aminoanthracene and benzo[a]pyrene were used for testing the bacteria and the activity of the S9 mix. Each treatment with the substances used as positive controls led to a clear increase in revertant colonies, thus showing the expected reversion properties of all strains and good metabolic activity of the S9 mix used.

No increase in revertant colonies was obtained after treatment with the test material up to the highest concentrations investigated, thus showing the absence of mutagenic potential in vitro with and without external metabolizing system.

Conclusions

With and without addition of S9 mix as the external metabolizing system, the test material was not mutagenic under the experimental conditions described.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Additional information from genetic toxicity in vitro:

For this endpoint information from three different assays is available. The data were obtained in the following assays:

1) Bacterial mutagenicity (OECD TG 471)

2) Mammalian cell mutation assay (OECD TG 476), read across from a structural similar liquid crystalline compound

3) Chromosomal abberation assay (OECD TG 473), read across from a structural similar liquid crystalline compound

All these data have been obtained according to the relevant OECD protocols.

The results are negative, not mutagenic and not clastogenic.

Justification for read across

Type of read across:one-to-one

Based on:

(1) chemical similarity

(2) almost identical physicochemical profile

(3) same response in biological systems relevant for the endpoint of mutagenicity as shown below:

 

Assay

Test item

surrogate (CAS 129738-34)

Appearance

solid

solid

Tanimoto Score

1

> 0.85

Water solubility

< 0.02 mg/L

< 0.03 mg/L

logP

> 5.7

> 6.5

Molar mass

234.4 g/mol

262.5 g/mol

Melting point

34.4 °C

51.4 °C

Boiling Point

329 °C

351°C

Vapour Pressure

0.015 Pa               

0.0031 Pa

Bacterial Mutagenicity

OECD TG 471 negative

OECD TG 471 negative



Justification for selection of genetic toxicity endpoint
This study was performed according to GLP and the methods applied are fully compliant with OECD TG 471. Additional supportive data have been taken from read across to chemical similar compounds with analogous physico chemical properties.

Justification for classification or non-classification

Based on the results provided there is no need for classification in this endpoint.