4,11-diamino-2-[3-(2-methoxyethoxy)propyl]-1H-naphth[2,3-f]isoindole-1,3,5,10(2H)-tetrone

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:

1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Principles of method if other than guideline:

None

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

4,11-diamino-2-[3-(2-methoxyethoxy)propyl]-1H-naphth[2,3-f]isoindole-1,3,5,10(2H)-tetrone

EC Number:

265-334-3

EC Name:

4,11-diamino-2-[3-(2-methoxyethoxy)propyl]-1H-naphth[2,3-f]isoindole-1,3,5,10(2H)-tetrone

Cas Number:

65059-45-2

Molecular formula:

C22H21N3O6

IUPAC Name:

4,11-diamino-2-[3-(2-methoxyethoxy)propyl]-1H,2H,3H,5H,10H-naphtho[2,3-f]isoindole-1,3,5,10-tetrone

Test material form:

solid: particulate/powder

Details on test material:

None

Method

Target gene:

The Salmonella typhimurium histidine (his) reversion system measures his- & his+ reversions

Species / strain

Species / strain / cell type:

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

Details on mammalian cell type (if applicable):

None

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

with and without liver microsomal activation. (S9 mix )

Test concentrations with justification for top dose:

Exp. I: 33.3; 100.0; 333.3; 1000.0; 2500.0; and 5000.0 µg/plate
Exp. II: 33.3; 100.0; 333.3; 666.6; 1000.0; and 2500.0 µg/plate
active ingredient

Vehicle / solvent:

No data

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

Salmonella TA 1535, TA 100 ; without metabolic activation

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylene-diamine

Remarks:

TA 1537, TA 98;without metabolic activation

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

TA 102;without metabolic activation

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

Salmonella typhimurium TA 15351 TA 1537, TA 98, TA 1001 TA 102 ; with metabolic activation

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)
Selective Agar
2. 0 % Vogel-Bonner-Glucose-Minimal-Agar was used as selective agar. Each petri dish was filled with 20 ml of this nutrient medium. Sterilisations were performed at 121° C in an autoclave.

Overlay Agar
The overlay agar contains per litre:
6.0 g Merck Agar Agar*
6.0 g NaCl*
10.5 mg L-histidine x HCl X H20*
12.2 mg biotin*
* (MERCK, D-64293 Darmstadt)
Sterilisations were performed at 121° C in an autoclave.

NUMBER OF REPLICATIONS:
Each concentration, including the controls, was tested in triplicate.

Evaluation criteria:

A test article is considered as positive if either a dose related and reproducible increase in the number of revertants or a significant and reproducible increase for at least one test concentration is induced. A test article producing neither a dose related and reproducible increase in the number of revertants nor a significant and reproducible positive response at any one of the test points is considered non-mutagenic in this system. A significant response is described as follows:
A test article is considered as mutagenic if in strain TA 100 and TA 102 the number of reversions is at least twice as high and in strains TA 1535, TA 1537, and TA 98 it is at least three times higher as compared to the spontaneous reversion rate (4). Also, a dose-dependent and reproducible increase in the number of revertants is regarded as an indication of possibly existing mutagenic potential of the test article regardless of whether the highest dose induced the above-described enhancement factors or not.

Statistics:

No appropriate statistical method is available

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

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

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

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

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 3

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

Untreated negative controls validity:

valid

True negative controls validity:

not examined

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

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 5

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

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Test results 6

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

Untreated negative controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

None

Any other information on results incl. tables

None

Applicant's summary and conclusion

Conclusions:

FAT 36152/G induced point mutations by frameshifts in the genome of the strain TA 98 with metabolic activation.

Executive summary:

A study was performed to investigate the potential of FAT 36152/G; Terasil Blau BGE Langamin roh feucht (laborgetrocknet) to induce gene mutations according to the plate incorporation test (experiment I and II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100, and TA 102. 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 article was tested at the following concentrations: 33.3; 100.0; 333.3; 1000.0; 2500.0; and 5000.0 µg/plate (active ingredient). No toxic effects occurred in experiment I and II with and without metabolic activation in all strains used. The plates incubated with the test article showed normal background growth up to 5000.0 µg/plate with and without S9 mix in all strains used. Up to the highest investigated dose a clear reproducible dose dependent increase in revertant colony numbers was observed with metabolic activation in tester strain TA 98. 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 article induced point mutations by frameshifts in the genome of the strain TA 98 with metabolic activation. Therefore, FAT 36152/G; Terasil Blau BGE Langamin roh feucht (laborgetrocknet) is considered to be mutagenic in this Salmonella typhimurium reverse mutation assay.