Dibenzyl ether

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:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study according OECD TG 471

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

This study was performed to investigate the potential of dibenzyl ether, CAS 103-50-4 to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100, and the Escherichia coli strain WP2 uvrA.
The assay was performed in two independent experiments both with and without liver microsomal activation.

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Dibenzyl ether

EC Number:

203-118-2

EC Name:

Dibenzyl ether

Cas Number:

103-50-4

Molecular formula:

C14H14O

IUPAC Name:

[(benzyloxy)methyl]benzene

Test material form:

other: liquid

Details on test material:

Identification: dibenzyl ether
Empirical Formula: C14H14O
Molecular Mass: 198.3 g/mol
CAS Registry Number: 103-50-4
Purity: 99.0 % (GC-analysis)
Appearance: Liquid, colorless-yellowish
Storage Conditions:
(provided by the Sponsor) Room temperature, light protected, moisture protected, N2
Purpose of Use: Industrial chemical
Stability in Solvent: Stable in DMSO for 4 hours at 20 ± 5˚C

Method

Species / strain

Species / strain 1

Species / strain / cell type:

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

Details on mammalian cell type (if applicable):

The histidine dependent strains are derived from Salmonella typhimurium strain LT2 through mutations in the histi¬dine locus. Additionally due to the "deep rough" (rfa-minus) mutation they possess a faulty lipopolysaccharide envelope which enables substances to penetrate the cell wall more easily. A further mutation causes a reduction in the activity of an excision repair system. The latter alteration includes mutational processes in the nitrate reductase and biotin genes produced in a UV-sensitive area of the gene named "uvrB-mi¬nus". In the strains TA 98 and TA 100 the R-factor plasmid pKM 101 carries the ampicillin resistance marker.

Additional strain / cell type characteristics:

not specified

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Details on mammalian cell type (if applicable):

Strain WP2 and its derivatives all carry the same defect in one of the genes for tryptophan biosynthesis. Tryptophan-independent (Trp+) mutants (revertants) can arise either by a base change at the site of the original alteration or by a base change elsewhere in the chromosome so that the original defect is suppressed. Additionally, the uvrA derivative is deficient in the DNA repair process (excision repair damage).

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

sodium azide

other: 4-nitro-o-phenylene-diamine, methyl methane sulfonate, 2-aminoanthracene

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

The test item dibenzyl ether, CAS 103-50-4 was assessed for its potential to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100, and the Escherichia coli strain WP2 uvrA.

The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration and the controls, were tested in triplicate.

The test item was tested at the following concentrations in both experiments: 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate

The test item precipitated in the overlay agar in the test tubes from 1000 to 5000 µg/plate. Precipitation of the test item in the overlay agar on the incubated agar plates was observed in experiment II in all strains without S9 mix at 5000 µg/plate.

No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with dibenzyl ether, CAS 103-50-4 at any concentration level, neither in the presence nor absence of metabolic activation (S9 mix).

Appropriate reference mutagens were used as positive controls. They showed a distinct increase in induced revertant colonies.

Applicant's summary and conclusion

Conclusions:

Interpretation of results: negative

Executive summary:

This study was performed to investigate the potential of dibenzyl ether, CAS 103-50-4 to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100, and the Escherichia coli strain WP2 uvrA.

The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration, including the controls, was tested in triplicate.

The test item was tested at the following concentrations in both experiments: 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate

The test item precipitated in the overlay agar in the test tubes from 1000 to 5000 µg/plate. Precipitation of the test item in the overlay agar on the incubated agar plates was observed in experiment II in all strains without S9 mix at 5000 µg/plate.

The plates incubated with the test item showed reduced background growth in all strains used.

Toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in nearly all strains used.

No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with dibenzyl ether, CAS 103-50-4 at any dose level, neither in the presence nor absence of metabolic activation (S9 mix).

Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.

In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used.

Therefore, dibenzyl ether, CAS 103-50-4 is negative in this Salmonella typhimurium and Escherichia coli reverse mutation assay.