N,N'-ethylenebis[N-acetylacetamide]

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study is in accordance to common test guidelines and GLP and therefore rated as reliable without restrictions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report date:

1988

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

adopted 26 May 1983

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

Version / remarks:

adopted 26 May 1983

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

N,N'-ethylenebis[N-acetylacetamide]

EC Number:

234-123-8

EC Name:

N,N'-ethylenebis[N-acetylacetamide]

Cas Number:

10543-57-4

Molecular formula:

C10H16N2O4

IUPAC Name:

N,N'-ethylenebis[N-acetylacetamide]

Details on test material:

- Name of test material (as cited in study report): TAED
- Colour: white
- Aggregate state at rt: solid
- Analytical purity: approx. 99.5%
- Lot/batch No.: E 12 T 24020 dated April, 1988
- Storage condition of test material and stability: room temperature, moisture protected, stable for months

Method

Target gene:

-

Species / strain

Species / strain / cell type:

other: S. typhimurium TA 1535, TA 1537, TA 1538, TA 98, TA 100; E. coli WP2

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

microsomal fraction from Aroclor 1254 induced rat livers

Test concentrations with justification for top dose:

10, 33.3, 100, 333.3 and 500 µg/plate

Vehicle / solvent:

- solvent used: deionized water
- Justification for choice of solvent/vehicle: solubility of test item

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: WITHOUT METABOLIC ACTIVATION sodium azide (TA 1535, TA 100), 4-nitro-o-phenylene-diamine (TA 1537, TA 1538, TA 98), MMS (WP2), WITH METABOLIC ACTIVATION 2-aminoanthracene (all strains)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
Plates were incubated upside down for 72 hours at 37°C .



NUMBER OF REPLICATIONS: 2 independent experiments, 3 plates per concentration

DETERMINATION OF CYTOTOXICITY
- Method: evaluation of number of spontaneous revertants and bacterial background lawn

Evaluation criteria:

A test item is considered as a mutagen if in strain TA 100 the number of reversions is at least twice as high and in strains TA 1535, TA 1537, TA 1538 and TA 98 is at least three times higher as compared to the spontaneous reversion rate.
A dose-dependent increase in the number of revertants is regarded as an indication of possibly existing mutagenic potential of the test item regardless whether the highest dose induced the above described enhancement factors or not.

Results and discussion

Test results

Species / strain:

other: S. typhimurium TA 1535, TA 1537, TA 1538, TA 98, TA 100; E. coli WP2

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:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: can be excluded
- Effects of osmolality: can be excluded
- Evaporation from medium: none
- Water solubility/ Precipitation:
The limit of solubility of the test article in H2O bidest was obtained at a concentration of 500.0 pg/plate. At this concentration still some particles of the test article remained undissolved.
The undissolved particles had no influence on the data recording. It was not possible to test higher concentrations of the test article than 500.0 pg/plate as no data recording could be performed.

- Other confounding effects: none

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

see attachment

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative with and without metabolic activation

Under the experimental conditions reported the test item did not induce point mutations by base pair changes or
frameshifts in the genome of the strains used.

Executive summary:

TAED was tested in the plate incorporation assay at concentrations of 10, 33.3, 100, 333.3 and 500 µg/plate using the Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 and in addition the Escherichia coli strain WP2 with and without S9-mix (microsomal fraction from Aroclor 1254 induced rat livers). Two independent experiments were performed. Each concentration, including negative, solvent and positive controls, was tested in triplicate. Deionized H₂O was chosen as solvent for TAED. 500 µg/plate was selected as the highest test concentration, since the test substance was partially insoluble at this concentration. Alternative solvents like DMSO, DMF, ethanol or acetone showed no better solubility properties. No distinct toxic effects occurred in the test groups. Up to the highest dose, no significant and reproducible increase in revertant colony numbers was obtained in any of the strains used.
TAED did not induce mutations in the genome of the bacterial strains used.
The study is in accordance with common test guidelines and GLP and therefore rated as reliable without restrictions (data quality score 1).