Naphthalene sulfonic acid, reaction products with isobutanol,sodium salts

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Study period:

2016-10-11 to 2016-10-31

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

See chapter 13 for support for read-across within the category of Alkyl Naphthalene Sulfonates (ANS).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Naphthalenesulfonic acid, bis(1-methylethyl)-, Me derivs., sodium salts

EC Number:

272-715-8

EC Name:

Naphthalenesulfonic acid, bis(1-methylethyl)-, Me derivs., sodium salts

Cas Number:

68909-82-0

Molecular formula:

UVCB substance

IUPAC Name:

Aromatic hydrocarbons, C10-13, reaction products with isopropylalcohol, sulphonated, sodium salts

Test material form:

solid: granular

Details on test material:

Name: Naphthalenesulfonic acid, bis(1-methylethyl)-, methyl derivs., sodium salt
Product: MORWET IP Powder
Chemical Name: Naphthalenesulfonic acid, bis(1-methylethyl)-, methyl derivs., sodium salt
CAS No.: 68909-82-0
Batch No.: 1452486
Physical State: solid, powder
Colour: tan
pH: 7.5 to 10 in 5 % solution
Active Components: UVCB substance with 100% purity
Average molecular weight: 346 g/mol (range from 213-474 g/mol)
Purity: 100 %
Storage Conditions: room temperature
Expiry Date: 07 June 2021
Safety Precautions: The routine hygienic procedures were sufficient to assure personnel health and safety.

Method

Target gene:

His

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

rat S9 liver microsomal fraction

Test concentrations with justification for top dose:

Experiment I:
31.6, 100, 316, 1000, 2500 and 5000 µg/plate
Experiment II:
10.0, 31.6, 100, 316, 1000, 2500 and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: A. dest.

Controls

Untreated negative controls:

yes

Remarks:

A. dest., Eurofins Lot No. 160905, 160915

Negative solvent / vehicle controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

methylmethanesulfonate

other: 4-nitro-o-phenylene-diamine; 2-aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
-For the plate incorporation method the following materials were mixed in a test tube and poured over the surface of a minimal agar plate:
100 µL Test solution at each dose level, solvent control, negative control or reference mutagen solution (positive control),
500 µL S9 mix (for testing with metabolic activation) or S9 mix substitution buffer (for testing without metabolic activation),
100 µL Bacteria suspension (cf. Preparation of Bacteria, pre-culture of the strain),
2000 µL Overlay agar.
After solidification the plates were inverted and incubated at 37 °C for at least 48 h in the dark.

- Preincubation period: 60 min; 100 µL of the test item preparation was pre-incubated with the tester strains (100 µL) and sterile buffer or the metabolic activation system (500 µL) for 60 min at 37 °C prior to adding the overlay agar (2000 µL) and pouring onto the surface of a minimal agar plate.
- Exposure duration: at least 48 h; After solidification the plates were inverted and incubated at 37 °C for at least 48 h in the dark.

NUMBER OF REPLICATIONS: For each strain and dose level, including the controls, three plates were used. (In a one case only two plates were evaluated)

DETERMINATION OF CYTOTOXICITY
- Method: clearing or rather diminution of the background lawn or a reduction in the number of revertants down to a mutation factor of approximately ≤ 0.5 in relation to the solvent control

Evaluation criteria:

The Mutation Factor is calculated by dividing the mean value of the revertant counts through the mean values of the solvent control (the exact and not the rounded values are used for calculation).
A test item is considered as mutagenic if:
- a clear and dose-related increase in the number of revertants occurs and/or
- a biologically relevant positive response for at least one of the dose groups occurs
in at least one tester strain with or without metabolic activation.
A biologically relevant increase is described as follows:
- if in tester strains TA 98, TA 100 and TA 102 the number of reversions is at least twice as high
- if in tester strains TA 1535 and TA 1537 the number of reversions is at least three times higher
than the reversion rate of the solvent control.

Statistics:

According to OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.

Results and discussion

Test results

Key result

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

experiment I and II

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

No precipitation of the test item was observed in any tester strain used in experiment I and II (with and without metabolic activation).

Toxic effects of the test item were noted in most tester strains evaluated in experiment I and II.

In experiment I toxic effects of the test item were observed in tester strains TA 98 and TA 1535 at a concentration of 5000 µg/plate and in tester strains TA 100 and TA 1537 at concentrations of 2500 µg/plate and higher (all without metabolic activation).

In experiment II toxic effects of the test item were noted in tester strains TA 98 and TA 1535 at concentrations of 2500 µg/plate and higher (without metabolic activation) and at a concentration of 5000 µg/plate (with metabolic activation). In tester strains TA 100 and TA 1537 toxic effects of the test item were observed at concentrations of 2500 µg/plate and higher (with and without metabolic activation).

Applicant's summary and conclusion

Conclusions:

The substance did not induce mutant colonies over background levels in the reverse gene mutation assay.

Executive summary:

In order to investigate the potential of Naphthalenesulfonic acid, bis(1-methylethyl)-, methyl derivs., sodium salt for its ability to induce gene mutations the plate incorporation test (experiment I) and the pre-incubation test (experiment II) were performed with the Salmonella typhimurium strains TA 98, TA 100, TA 1535, TA 1537 and TA 102.

In two independent experiments several concentrations of the test item were used. Each assay was conducted with and without metabolic activation. The concentrations, including the controls, were tested in triplicate. The following concentrations of the test item were prepared and used in the experiments:

Experiment I: 31.6, 100, 316, 1000, 2500 and 5000 µg/plate

Experiment II: 10.0, 31.6, 100, 316, 1000, 2500 and 5000 µg/plate

No precipitation of the test item was observed in any tester strain used in experiment I and II (with and without metabolic activation).

Toxic effects of the test item were noted in most tester strains used in experiment I and II:

- at concentrations of 2500 µg/plate and higher (without metabolic activation), depending on the particular tester strain.

- In experiment II toxic effects of the test item were noted at concentrations of 2500 µg/plate and higher (with and without metabolic activation), depending on the particular tester strain.

No biologically relevant increases in revertant colony numbers of any of the five tester strains were observed following treatment with Naphthalenesulfonic acid, bis(1-methylethyl)-, methyl derivs., sodium salt at any concentration level, neither in the presence nor absence of metabolic activation in experiment I and II.

All criteria of validity were met.

In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, Naphthalenesulfonic acid, bis(1-methylethyl)-, methyl derivs., sodium salt did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used.

Therefore, Naphthalenesulfonic acid, bis(1-methylethyl)-, methyl derivs., sodium salt is considered to be non-mutagenic in this bacterial reverse mutation assay.