Manganese, 4-[(5-chloro-4-methyl-2-sulfophenyl)azo]-3-hydroxy-2-naphthalenecarboxylic acid complex

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:

2006 - 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Calcium 3-hydroxy-4-[(4-methyl-2-sulphonatophenyl)azo]-2-naphthoate

EC Number:

226-109-5

EC Name:

Calcium 3-hydroxy-4-[(4-methyl-2-sulphonatophenyl)azo]-2-naphthoate

Cas Number:

5281-04-9

Molecular formula:

C18H14N2O6S.Ca

IUPAC Name:

calcium 3-hydroxy-4-[(4-methyl-2-sulfonatophenyl)diazenyl]-2-naphthoate

Test material form:

solid: nanoform

Details on test material:

- Purity: approximately 98% [ Containing as impurities : NaCl, CaCl2, Ca(OH)2, and water ]
- Lot/Batch No : T-1322-2
- Storage condition of test material : Stored in cold-dark conditions
- Stability in the vehicle: Stable for 4 hours in the vehicle at the concentrations ranging between 0.925 and 50.0 mg/ml, according to the stability test performed by the laboratory.

Test materials used in this dossier are all considered to fall under the definition of nano-materials according to the European Commission Recommendation 2011/696/EU as the synthesis and manufacturing of this pigment always yields particulate material with a fine particle size distribution.

Specific details on test material used for the study:

- Purity: approximately 98% [ Containing as impurities : NaCl, CaCl2, Ca(OH)2, and water ]
- Lot/Batch No : T-1322-2
- Storage condition of test material : Stored in cold-dark conditions
- Molecular weight: 424.45
- Stability in the vehicle : Stable for 4 hours in the vehicle at the concentrations of 3 and 50.0 mg/ml, according to the stability test performed by the laboratory.

Method

Target gene:

histidine operon

Species / strain

Species / strain 1

Species / strain / cell type:

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

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A pKM 101

Metabolic activation:

with and without

Metabolic activation system:

Phenobarbital/beta-Naphthoflavone induced rat liver S9 and S9 liver microsomal fraction prepared from the liver of 7 - 8 weeks old male Syrian golden hamsters

Test concentrations with justification for top dose:

- Pre-Experiment/Experiment I : 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate
- Experiment II: 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate

Vehicle / solvent:

DMSO
The solvent was chosen because of its solubility properties and its relative non-toxicity to the bacteria

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

congo red

methylmethanesulfonate

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

Remarks:

strains TA 100 and TA 1535 without S9

Details on test system and experimental conditions:

In the pre-experiment the concentration range of the test item was 3 - 5000 microgramm/plate. It was the plate-incorporation experiment. The pre-experiment is reported as experiment I since the criteria mentioned above were met and 5000 microgramm/plate were chosen as maximal concentration. Due to minor toxic effects and the precipitation of the test item in the pre-experiment, seven concentrations were tested in experiment II. Experiment II was performed with hamster liver S9-mix in the pre-incubation set-up.

Evaluation criteria:

- A test item is considered as a mutagen if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA 98, TA 100, and WP2 uvrA) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control is observed.
- A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.
- An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
- A dose dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered biologically relevant.

Statistics:

No statistical evaluation of the data was required.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and 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 2

Species / strain:

E. coli WP2 uvr A pKM 101

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

Additional information on results:

Precipitation of the test item was observed from 333 microgramm/plate up to 5000 microgramm/plate in the test tubes. Precipitation was also observed on the agar plates at 1000 and 2500 microgramm/plate without S9 mix and from 1000 to 5000 microgramm/plate with S9 mix in experiment I, and from 1000 microgramm/plate up to 5000 microgramm/plate with and without S9 mix in experiment II. The undissolved particles had no influence on the data recording.
The plates incubated with the test item showed normal background growth up to 5000 microgramms per plate with and without metabolic activation in both experiments. Minor toxic effects, evident as a reduction in the number of revertants (below a factor of 0.5), were observed in some strains at concentrations of 2500 microgramms per plate or higher.