NALCO 73280

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

Study period:

25 April 2001 to 29 May 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was conducted in accordance with International Guidelines and in accordance with the principles of Good Laboratory Practise (GLP).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Details on test material:

- Name of test material (as cited in study report): EH&S 01-114
- Lot/batch No.:XC1B1263A0

Method

Target gene:

Histidine for Salmonella typhimurium
Tryptophan for E.Coli.

Species / strain

Species / strain 1

Species / strain / cell type:

E. coli WP2 uvr A

Details on mammalian cell type (if applicable):

Not applicable

Species / strain 2

Species / strain / cell type:

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

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Aroclor induced rat livers (S9 fraction)

Test concentrations with justification for top dose:

Rangefinding test:
6.7 - 5000 μg/plate

Concentration range in main test:
With metabolic activation: 100, 333, 1000, 3333, 5000 μg/plate
Without metabolic activation: 100, 333, 1000, 3333, 5000 μg/plate

Vehicle / solvent:

Water

Controls

Untreated negative controls:

no

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: See remarks

Remarks:

Used with S9 mix: 2-aminoanthracene. Used without S9 mix: 2-nitrofluorene, sodium azide, 9-aminoacridine, methyl methanesulfonate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium; in agar (plate incorporation);
DURATION
- Preincubation period: overnight
- Exposure duration: 48 - 72 hours
- Fixation time (start of exposure up to fixation or harvest of cells):48 - 72 hours


NUMBER OF REPLICATIONS: 3 plates


DETERMINATION OF CYTOTOXICITY
- Method: Bacterial Background Lawn Evaluation -
The condition of the bacterial background lawn was evaluated for evidence of cytotoxicity (using a dissecting microscope) and test article precipitate (by visual examination without magnification). Evidence of cytotoxicity and degree of precipitation was scored relative to the vehicle control plate.


Counting revertant colonies -
The number of revertant colonies per plate for the vehicle controls and all plates containing test article were counted either entirley by automated colony counter or entirely by hand unless the assay was the preliminary toxicity assay or the plate exhibited toxicity. Plates with sufficient test article precipitate to intefere with automated colony counting were counted manually

Evaluation criteria:

Criteria For A Positive Response:

For the test substance to be evaluated positive, it must cause a dose related increase in the mean revertants per plate of at least one tester strain over a minimum of two increasing concentrations of test substance. Data sets for tester strains TA 1535 and TA1537 were judged positive if the increase in mean revertants at the peak of the dose response is equal to or greater than three times the mean vehicle control value. Data sets for tester strains TA98, TA 100 and WP2 uvrA were judged positive if the increase in mean revertants at the peak of the dose response is equal to or greater than two times the mean vehicle control value

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

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

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

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitate was observed at any dose levels

RANGE-FINDING/SCREENING STUDIES:
Neither appreciable toxicity or preciptae was observed upto the maximum dose tested of 5000μg per plate. Based on these findings of the toxicity assay, the maximum dose plated in the mutagenicity assay was 5000μg per plate.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Please refer to attachment S7.6.1 Genetic Toxicity Ames Results Tables 1 - 28.pdf

Preliminary Toxicity Assay Tables 1 - 5

Initial Mutagenicity Assay

Individual plate counts Tables 6 - 15

Summary of results Table 26

Independent Repeat Assay

Individual plate counts Tables 16 - 25

Summary of results Table 27

Historical Negative and Positive Control Values Table 28

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

The test substance was not mutagenic to bacteria under the conditions of the test.

Executive summary:

The test substance was evaluated in the Bacterial Reverse Mutation Assay with an Independent Repeat Assay using Salmonella typhimurium tester strains TA 98, TA 100, TA 1535 and TA 1537 and Escherichia coli tester strain WP2 uvrA in the presence and absence of Aroclor-induced rat liver S9. The assay was conducted on the test substance along with vehicle control and positive control

and was performed using the plate incorporation method.

The test substance doses levels in the mutagenicity assay, 100, 333, 1000, 3333 and 5000μg per plate were selected based on the results of a preliminary test. No positive responses were observed at any dose levels of the test substance with any of the tester strains.