3A-Flux

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 27 April 2021 to 14 May 2021

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Report date:

2021

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Potassium hydrogen fluoride, reaction product with boric acid and dipotassium salt of boric acid

Cas Number:

2787482-26-0

Molecular formula:

not applicable (UVCB)

IUPAC Name:

Potassium hydrogen fluoride, reaction product with boric acid and dipotassium salt of boric acid

Test material form:

solid - liquid: suspension

Method

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9-mix
Obtained by Trinova Biochem GmbH, Gießen, Batch no. 4344, produced from the livers of male Sprague-Dawley rats which were treated with Phenobarbital/5,6-Benzoflavone.

Test concentrations with justification for top dose:

Experiment 1: 5000, 1500, 500, 150 and 50 μg/plate.
Experiment 2 and 2b: 5000, 2500, 1250, 625, 313, 156, 78 μg/plate.
5000 µg/plate was used as top dose as this is the recommended maximum test concentration according to OECD 471 guideline.

Vehicle / solvent:

Based on these results of the non-GLP pre-test, demin. water was used as solvent in the experiments

Controls

Negative solvent / vehicle controls:

yes

Remarks:

Dimethylsulfoxide (DMSO), CAS No. 67-68-5, for the positive controls Demineralized water for the test item and the positive control Sodium chloride (0.9 % NaCl) for the positive control

Positive controls:

yes

Positive control substance:

sodium azide

benzo(a)pyrene

mitomycin C

other:

Details on test system and experimental conditions:

- Number of cultures per concentration: Triplicate
- Number of independent experiments: 2
- Experiment 1: Plate incorporation method
Experiment 2: Pre-incubation method
- Exposure duration/duration of treatment: 48 h

Evaluation criteria:

A result is considered as positive 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 concentrations occurs in at least one tested strain with or without metabolic activation.

A biologically relevant increase is described as follows:
- if in the bacteria strains S. typhimurium TA98, TA100, TA102 the number of revertants is at least twice as high than the reversion rate of the negative controls (increase factor of at least 2.0)
- if in the bacteria strains S. typhimurium TA1535 and TA1537 the number of revertants is at least three times higher than the reversion rate of the negative controls (increase factor of at least 3.0).

A substance is not mutagenic if it does not meet the criteria above.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

Based on the results of this study it is concluded that 3A-Flux is not mutagenic in the Salmonella typhimurium test strains TA98, TA100, TA102, TA1535 and TA1537 in the absence and presence of metabolic activation under the experimental conditions of the present study.

Executive summary:

The study was performed with the plate incorporation (experiment 1) and pre-incubation method (experiments 2 and 2b) in the absence and presence of a metabolic activation system (S9). Under these conditions the influence of the test item on bacterial test strains was evaluated. The test item 3A-Flux showed no relevant or dose-related increase in the number of revertants in the Salmonella typhimurium test strains TA98, TA100, TA102, TA1535 and TA1537 in all evaluated experiments.