Manganese, 3-hydroxy-4-[(1-sulfo-2-naphthalenyl)azo]-2-naphthalenecarboxylic acid complex

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The test substance itself showed no mutagenic effects in three salmonella strains with and without microsomal activation, in an Ames test comparable to OECD TG 471, non-GLP (1985).

An Ames test with five bacterial strains, including Prival modification was not performed for the test item. Reliable experimental data from two analogue substances are available. These substances were not considered to be mutagenic in S. typhimurium TA 1535, TA 100, TA 1537, TA 98 and E.coli WP2 uvrA in the absence or presence of a metabolic activation system (according to OECD TG 471 and GLP).

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Please, see the attached justification.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

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

True negative controls validity:

not examined

Positive controls validity:

valid

Species / strain:

E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Reference 2

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

10 - 21 january 1985

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Only 3 strains tested, non-GLP study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

3 strains had been tested instead of 5

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

other: S. typhimurium TA 98, TA 100 and TA 1537

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254-induced rat liver S-9 fraction

Test concentrations with justification for top dose:

The test substance was tested in concentrations of 20, 78, 313, 1250 and 5000 µg/ 0.1 mL with and wihout S9-mix

Vehicle / solvent:

DMSO

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene (all strains)

Remarks:

With microsomal activation

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: daunorubicin-HC1 (TA 98), 4-nitroquinoline-N-oxide (TA 100), 9(5)aminoacridine-hydrochloride (TA 1537)

Remarks:

Without microsomal activation

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: The plates were incubated for about 48 hours at 37 ± 1.5 °C in darkness

NUMBER OF REPLICATIONS: Three Petri dishes were prepared per strain and per group.

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

Evaluation criteria:

The test substance is generally considered to be nonmutagenic if the colony count in relation to the negative control is not doubled at any concentration.

Statistics:

When the colonies had been counted, the arithmetic mean was calculated.

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:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

reduction in the colony count at the highest concentration

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

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:

not examined

True negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: At the concentrations of 1250 and 5000 µg/ 0.1 mL the substance precipitated in soft agar.

RANGE-FINDING STUDIES: A preliminary toxicity test was carried out with the concentrations ranging from 0.08 to 5000 µg/0.1 mL.

COMPARISON WITH HISTORICAL CONTROL DATA: The spontaneous reversion in the control was within the historical control range for each strain.

ADDITIONAL INFORMATION ON CYTOTOXICITY: Owing to a growth-inhibition effect of the substance on strain TA 100 a reduction in the colony count was observed at the highest concentration.

Table 1: Results of a salmonella mutagenicity test with three strains

 

Without microsomal activation Number of back-mutant colonies per plate (arithmetic mean)
Strain	TA 98	TA 100	TA 1537
Control	20	136	5
25 µg/ 0.1 mL	18	138	6
75 µg/ 0.1 mL	24	133	7
225 µg/ 0.1 mL	25	140	8
675 µg/ 0.1 mL	21	106	6
2025 µg/ 0.1 mL	15	57	6
Positive controls
Daunorubicin-HC1 (10 µg/ 0.1 mL)	843
4-nitroquinoline-N-oxide(0.25 µg/ 0.1 mL)		1388
9(5)aminoacridine-hydrochloride(100 µg/ 0.1 mL)			971
With microsomal activation Number of back-mutant colonies per plate (aritmethic mean)
Strain	TA 98	TA 100	TA 1537
Control	29	114	9
20 µg/ 0.1 mL	35	126	12
78 µg/ 0.1 mL	29	123	11
313 µg/ 0.1 mL	28	134	10
1250 µg/ 0.1 mL	35	159	12
5000 µg/ 0.1 mL	30	97	7
Positive control of the microsomal activation
2-amincanthracene (250 µg/ 0.1 mL)	1000	770	84

Conclusions:

Under the conditions of this study, no mutagenicity was observed in the screening test with 3 strains of S. typhimurium.

Executive summary:

In a reverse gene mutation assay in bacteria, three strains of S. typhimurium (TA 98, TA 100 and TA 1537) were exposed to five concentrations of the test item in the presence and absence of mammalian metabolic activation. The test substance was dissolved in DMSO and the test was performed with the following concentrations of the trial substance with and without microsomal activation: 20, 78, 313, 1250 and 5000 µg/0.1 mL. DMSO was used for the negative controls and a positve control was tested for each strain. No evidence of the induction of point mutations by the test item or by the metabolites of the substance formed as a result of microsomal activation was detectable in the strains of S. typhimurium used in these experiments. The spontaneous reversion in the control was within the historical control range for each strain and the positive controls induced the appropriate responses in the corresponding strains. Therefore, the test results are considered valid.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

The genotoxic potential of the test substance itself was assessed in a reverse gene mutation assay. The conducted study satisfies OECD TG 471 (Bacterial Reverse Mutation Assay) with one deviation: 3 bacteria strains had been tested instead of 5 (S. typhimurium TA 98, TA100 and TA 1537). The test substance was dissolved in DMSO and the test was performed with the following concentrations of the trial substance with and without microsomal activation: 20, 78, 313, 1250 and 5000 µg/0.1 mL. DMSO was used for the negative controls and a positve control was tested for each strain. No evidence of the induction of point mutations by the test item or by the metabolites of the substance formed as a result of microsomal activation was detectable in the strains of S. typhimurium used in these experiments.The spontaneous reversion in the control was within the historical control range for each strain and the positive controls induce the appropriate responses in the corresponding strains.Therefore, the test results are considered valid.

 

An Ames test with 5 bacterial strains, including Prival modification was not performed with the test substance, but reliable experimental data for an analogue approach are available. Please, see the attached read-across justification in section 7.6.1 or 13.

 

CAS 6417-83-0:

The analogue substance was tested for its mutagenic potential based on the ability to induce point mutations in selected loci of several bacterial strains, i.e. S. typhimurium (TA 1535, TA 100, TA 1537, TA 98) and E.coli (WP2 uvrA), in a reverse mutation assay (Ames standard plate test and Prival preincubation test) at concentrations of 33 μg - 5400 μg/plate with and without metabolic activation. Precipitation of the test substance was found depending on the test conditions from about 100 μg/plate onward. A weak bacteriotoxic effect was occasionally observed depending on the strain and test conditions from about 2700 μg/plate onward. A biologically relevant increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the prival preincubation test either without S9 mix or after the addition of a metabolizing system. Thus, under the experimental conditions of this study, the test substance is not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay in the absence and the presence of metabolic activation.

 

CAS 7023-61-2:

The mutagenicity of the test substance was assessed in an in vitro reverse mutation assay (Ames standard plate test and Prival preincubation test) following OECD TG 471 and the principles of GLP. The test substance was administered to five bacterial strains (S. typhimurium TA 1535, TA 100, TA 1537, TA 98 and E.coli WP2 uvrA pKM 101) at concentrations of 3; 10; 33; 100; 333; 1000; 2500 and 5000 µg/plate with and without metabolic activation. Precipitation of the test item was observed from 333 µg/plate onwards in the test tubes and on the agar plates at different concentrations in both experiments. 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, were observed in some strains at concentrations of 2500 microgramms per plate or higher. No biologically relevant increase in the number of his+ or trp+ revertants was observed in the standard plate test or in the prival preincubation test neither without nor with a metabolizing system.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. No indication of genotoxicity was observed in the Ames test (OECD 471, GLP). As a result, the substance is not considered to be classified for mutagenicity under Regulation (EC) No. 1272/2008, as amended for the fourteenth time in Regulation (EC) No. 2020/217.