Estradiol

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:

February 2003

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:

2004

Report Date:

2004

Materials and methods

Test guideline

Qualifier:

according to

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Reference

Name:

Unnamed

Type:

Constituent

Test material form:

solid

Specific details on test material used for the study:

- Solubility and stability of the test substance in the solvent/vehicle: The solution and further dilutions were prepared immediately before addition to the test bacteria. Thus, no remarkable instability is expected which could influence the outcome of
the study.

Method

Target gene:

Histidine gene locus

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium, other: TA 98, TA 100, TA 1535, TA 1537 and TA 1538

Additional strain / cell type characteristics:

not applicable

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 induced male Sprague-Dawley rat liver S9 mix

Test concentrations with justification for top dose:

0.05 - 5.0 mg/plate (with/without S9 mix; plate incorporation and the experiment with preincubation)

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

sodium azide

N-dimethylnitrosamine

benzo(a)pyrene

cyclophosphamide

ethylmethanesulphonate

other: 4-nitro-o-phenylenediamine, 2-aminoanthracene , N-Methyl-N´-nitro-N-nitrosoguanidine

Evaluation criteria:

A positive response was considered if the number of revertants of the compound groups compared to the number of revertants of the negative group was reproducibly higher than 2-fold. A dose-dependent increase in the number of
revertants was also considered to indicate a mutagenic effect.

Statistics:

The arithmetic means of the number of mutant colonies of the 3 parallel plates in the negative control groups were compared with those of the compound groups. However, in the tables the given mean values are rounded to the nearest integer.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium, other: TA 98, TA 100, TA 1535, TA 1537 and TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A

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:

not examined

Positive controls validity:

valid

Additional information on results:

Precipitates in the agar were found starting from 0.5 mg/plate onwards.
Generally, growth inhibition of the background lawn was observed at 2.5 and 5.0 mg/plate.
The preincubation experiment with and without S9 mix on strain TA 1537, however, was repeated in order to check the results of the positive control anthracene-2-amine.

Any other information on results incl. tables

None of the six tester strains S. typhimurium TA1535, TA100, TA1537, TA1538, TA98 and E.coli WP2uvrA showed increased reversion to prototrophy in assays with the test item at the doses tested between 0.05 and 5.0 mg/plate, either in the absence or presence of metabolic activation. Precipitates in the agar were found starting from 0.5 mg/plate onwards. Growth inhibition of the background lawn was observed in all strains at 2.5 and 5.0 mg/plate without and with metabolic activation.The counts recorded on appropriate negative control plates confirmed the characteristically spontaneous reversion rates of the tester strains. Furthermore, appropriate positive controls with known mutagens produced the expected distinct increase in the number of revertant colonies.

Applicant's summary and conclusion

Executive summary:

The mutagenic potential of the test substance was evaluated in a Salmonella/microsome test with the S. typhimurium strains TA 98, TA 100, TA 1535, TA 1537, TA 1538 and E. coli WP2uvrA in the presence and absence of S9 mix according to OECD TG 471. Evidence of mutagenic activity was not seen up to the maximum recommended dose level of 5.0 µl/plate. No substantial increases in revertant colony numbers of any of the six tester strains were observed at any dose level in the presence and absence of metabolic activation. Therefore, the test substance was considered to be non-mutagenic in the Salmonella typhimurium reverse mutation assay.